Home

User's Guide Neo-Sensitabs 18th ed 10-2005-2006

1. sessi nennen 95 14 5 Enterococci and VRE Quality Control essere nre trennen ene 96 15 Susceptibility Testing of Fastidious or Problem Organisms seeeeeeeeeeeee nennen eene 98 15 1 Susceptibility Testing of Haemophilus influenzae and parainfluenzae esse 99 15 1 1 Quality Control Limits for Haemophilus influenzae ATCC 49247 sess 102 15 2 Susceptibility Testing of Gonococci ete Den Dee RO Te CERERI E ERR ERE II De e DER FIRE 104 15 2 1 Quality Control Limits for N gonorrhoeae ATCC 49226 essen 105 15 3 Susceptibility Testing of Meningococci sess 107 15 4 Susceptibility Testing of Moraxella catarrhalis eene enne 109 15 5 Susceptibility Testing of Pneumococci rnarnnnnnvnvnnnvnnrnvrrnvrrnerevrvennverarrnrnrernensnensnnenvennvesvsvsvaverasevassresvresnen 110 15 5 1 Quality Control Limits for 5 pneumoniae ATCC 49619 113 15 6 Susceptibility Testing of Beta haemolytic and Viridans Streptococci esrnarnnnnnnnnvnnnvrrnrnrnrveraverarvrervreenen 115 15 7 Susceptibility Testing of 1 en 119 15 8 Susceptibility Testing of Vibrio cholerae 121 15 9 Susceptibility Testing of Helicobacter pylori essent nen
2. Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 17 Page 148 of 170 17 Detection of Other Resistance Mechanisms 17 1 Screening of 16S rRNA Methylases HLR to Aminoglycosides Unlike aminoglycoside modifying enzymes that vary in their substrate profile the acquired 16S rRNA methylases confer high level resistance HLR to almost all clinically important aminoglycosides They have been identified in several nosocomial pathogens including P aeruginosa Serratia marcescens E coli P mirabilis pneumoniae and Acinetobacter spp Enterobacter cloacae Citrobacter freundii 8 9 These enzymes RmtA RmtB RmtC ArmA are capable of conferring very high levels of resistance MIC gt 512 ug ml against amikacin gentamicin isepamicin netilmicn and tobramycin while apramycin neomycin and streptomycin are not affected The responsible genes ArmA RmtA RmtB and RmtC are located in self tramsmissible plasmids 7 Screening method A high level amikacin resistance MIC gt 512 ug ml corresponding to no zone of inhibition around Amikacin 40 ug Neo Sensitabs may be used as a marker for screening the 168 rRNA methylase producing strains The diffusion test is performed on MH agar using a 0 5 McF inoculum and incubation at 35 37 C overnight Strains of Enterobacteriaceae and non fermenters P aeruginosa and Acinetobacter spp showing no zone of inhibition around Amikacin 40 ug Neo Sensitabs should be suspected o
3. The emergence of 16S rRNA methylases in Enterobactericaceae and non fermenters P aeruginosa Acinetobacter Spp in strains that already are ESBL positive may result in the spread of multidrug resistant isolates producing both ESBLs and 16S rRNA methylases becoming an important clinical problem References 1 Galimand M et al Plasmid mediated high level resistance to aminoglycosides in Enterobacteriaceae due to 16S rRNA methylation Antimicr Ag Chemother 47 2565 2571 2003 2 Yohei Doi et al Plasmid mediated 16S rRNA methylase in Serratia marcescens conferring HLR to aminoglycosides Antimicr Ag Chemother 48 491 6 2004 3 Jing Jou Yan et al Plasmid mediated 16S rRNA methylases conferring HL aminoglycoside resistance coli and pneumoniae isolates from 2 Taiwanese hospitals J Antimicr Chemother 54 1007 1012 2004 4 Kumkazu Yamane et al Global spread of multiple aminoglycoside resistance genes Emerg Infect Dis 11 951 953 2005 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 17 Page 149 of 170 5 Galimand M et al Worldwide disseminated armA aminoglycoside resistance methylase gene is borne by composite transposon Tn1548 Antimicr Ag Chemother 49 2949 53 2005 6 Gonz lez Zorn B et al Genetic basis of dessemination of armA J Antimicr Chemother 56 583 5 2005 7 Wachino Jun idri et al Novel plasmid mediated 165 rRNA methylase found in a P mirabilis i
4. 12 mm Nalidixan is a good screening for the detection of decreased fluoroquinolone susceptibility in Salmonella spp Hakanen A et al J Clin Microbiol 37 3572 77 1999 Strains resistant to Nalidixan Neo S zone 28 mm show a decreased susceptibility to quinolones ciprofloxacin MIC gt 0 125 ug ml When testing Pseudomonas use S gt 30 mm I 29 23 mm R lt 23 mm Carbapenem testing on Iso Sensitest agar may give false susceptibility results for isolates that may harbour metallo beta lactamases testing on Mueller Hinton agar may be preferred BSAC 2001 Strains of Klebsiella Salmonella and E coli that produce ESBL may be clinically resistant to therapy with penicillins cephalosporins or aztreonam despite apparent in vitro susceptibility See chapter 16 1 for ESBL screening and confirmatory tests For the detection of staphylococci with reduced susceptibility to vancomycin teicoplanin VISA GISA hVISA see the chapter in User s Guide on detection of strains with decreased susceptibility to vancomycin page 86 O Copyright Rosco Diagnostica A S NEO SENSITABS II b Haemophilus spp S pneumoniae N gonorrhoeae N meningitidis and H pylori Interpretation according to the MIC break points recommended by the Dutch CRG 2000 Inoculum McFarland 0 5 or 3 H pylori Zone diameter 09 2007 2008 Page 35 of 170 Chapter 10 Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Penicil
5. Chiu C H et al Increasing ceftriaxone resistance and multiple alterations of PBPs among penicillin resistant S pneumoniae isolates in Taiwan Antimicr Ag Chemother 51 3404 06 2007 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 115 of 170 15 6 Susceptibility Testing of Beta haemolytic and Viridans Streptococci The recommended medium is Mueller Hinton agar supplemented with 596 defibrinated sheep blood Procedure 1 Growth from an overnight 18 20 h sheep blood agar plate is suspended in Mueller Hinton broth or 0 9 saline to a density equivalent to the 0 5 McFarland standard Plates must be inoculated within 15 minutes The current tablet diffusion procedure described for Neo Sensitabs should be followed but no more than 9 Neo Sensitabs should be placed on a large 150 mm plate or 4 Neo Sensitabs on a 90 100 mm plate Incubate the plate at 35 2 degrees in an atmosphere of 5 7 CO for 20 24 hours before reading the inhibition zones Zone diameter interpretative standards are described in the table below Special interpretation for beta haemolytic streptococci with penicillins and cephalosporins CLSI Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Amoxycillin 30 ug AMOXY gt 28 lt 27 lt 0 25 Ampicillin 33 ug AMP33 228 lt 27 lt 0 25 Cefepime 30 ug CFEPM gt 26 lt 0 5 Cefotaxime 30 ug CFTAX gt 26 lt 0 5 g
6. Three cases of invasive infections caused by Salmonella enterica serotype cholerasuis found in Taiwan 5 The strains were resistant to ciprofloxacin mutations gyrA and ParC and to ceftriaxone presence of plasmid mediated CMY 2 beta lactamase Recent studies 4 show that the outcome of cephalosporin treatment in serious infections due to AmpC beta lactamase producing K pneumonia isolates was poor A standard test for detection of plasmid mediated AmpC beta lactamases is needed Emergence of cefepime hydrolyzing CMY 19 in Japan 8 Detection of Plasmid mediated A mpC beta lactamases Strains suspicious of possessing plasmid mediated AmpC beta lactamases are cefoxitin resistant except AAC 1 and have reduced susceptibility to ceftazidime while currently they are susceptible to cefepime and the carbapenems Procedure Apply Cefotaxime Clavulanate and Ceftazidime Clavulanate Neo Sensitabs At a distance of 10 mm edge to edge of each apply Boronic acid Diatabs Apply Ceftazidime and Cefoxitin Neo Sensitabs At a distance of 5 to 10 mm edge to edge apply Cloxacillin 500 ug Diatabs Interpretation A keyhole or ghost zone synergism between Boronoic acid and any of Cefotaxime Clavulanate or Ceftazidime Clavulanate indicates the presence of a AmpC beta lactamase A keyhole or ghost zone synergism between Cloxacillin 500 ug and any of Ceftazidime or Cefoxitin indicates the presence of an AmpC beta lactamase Inducible AmpC be
7. lt 12 lt 0 06 pen MIC test Oxacillin 5 ug OXA 5 8 S pneumoniae gt 24 lt 24 lt 0 06 pen test d Streptococcus spp gt 20 lt 20 lt 0 12 pen test l gonorrhoeae gt lt 18 lt 0 06 pen test 9 N meningitidis gt 18 5 18 lt 0 06 pen test Amoxycillin 30 ug AMOXY Haemophilus spp lt 30 gt 1 Anaerobes gram pos gt 24 23 18 lt 18 lt 4 gt 16 Anaerobes gram neg gt 32 31 28 lt 28 lt 0 5 gt 1 Amoxycillin Clav 30 15 ug AM CL Haemophilus spp gt 28 lt 4 2 Campylobacter spp 228 27 20 20 lt 4 2 gt 16 2 Anaerobes gt 28 27 20 lt 20 lt 4 2 gt 16 2 Ticarcillin Clavulanate 75 15 ug TI CL Anaerobes gt 24 23 18 lt 18 lt 16 2 gt 64 2 Piperacillin Tazobactam 100 10ug PI TZ Anaerobes gt 28 27 20 lt 20 lt 8 4 gt 64 4 Cephalothin 66 ug CLOTN e Haemophilus spp 26 gt 8 Amp R Ceftizoxime 30 ug CEZOX b pneumoniae gt 32 lt 32 lt 0 5 gt 0 5 valid for 3rd gen cephalosporins Cefotaxime 30 ug CFTAX Ceftriaxone 30 ug CETRX b S pneumoniae use Ceftizoxime N gonorrhoeae 240 lt 0 25 N meningitidis 238 lt 0 25 Campylobacter spp 224 23 18 18 lt 4 232 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 70 of 170 Zone diameter Concentrations in mm critiques NEO SENSITABS POTENCY CODE S R S R Cefoxitin 60 ug CFOXT Anaerobes 22 2
8. 1 Growth method a Select at least 3 to 5 isolated colonies Touch the top of each colony with a wire loop and transfer to a tube containing 4 to 5 ml of a broth medium such as tryptic soy broth b Allow the broth culture to incubate at 35 C until it achieves or exceeds the turbidity of the 0 5 McFarland standard It usually takes 2 to 6 hours c Adjust the turbidity of the broth culture with sterile saline or broth to obtain a turbidity visually comparable to the standard This results in a suspension containing approx 1 to 2 x 10 CFU ml for E coli ATCC 25922 d Within 15 minutes dip a sterile cotton swab into the adjusted suspension and remove inoculum from the swab by exerting firm pressure on the inside of the tube e Inoculate the dried surface of the Mueller Hinton agar plate by streaking the swab over the entire surface 2 Direct colony suspension method It is also possible to set up the tests immediately rather than wait for a broth sub culture 3 This can be done by suspending several morphologically similar colonies from an 18 24 h agar plate non selective into 4 5 ml 0 9 NaCl solution and then immediately adjusting the turbidity to match that of the BaSO4 standard 0 5 McFarland Within 15 minutes swabs are used to inoculate the test plates This method was found to be equivalent to the standard CLSI method and requires less time This method is used in most laboratories for all types of bacteria This approach
9. 55 1055 6 2005 Lesho E et al Fatal Acinetobacter baumannii infection with discordant carbapenem susceptibility CID 41 758 9 2005 Jones R N et al Carbapenem susceptibility discords among Acinetobacter isolates CID 42 158 2005 Thamlikitkul V et al In vitro activity of Tigecycline against B pseudowallei and B thailandensis Antimicr Agents Chemother 50 1555 7 2006 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 126 of 170 16 Detection of Beta Lactamases 16 1 Extended Spectrum Beta Lactamases ESBL Screening and Confirmatory Tests for Extended Spectrum Beta Lactamases ESBL Transferable plasmid mediated beta lactamases that produce resistance towards third generation cephalosporins and monobactams e g aztreonam have been described in strains of Klebsiella pneumoniae K oxytoca E coli and other Enterobacteriaceae These enzymes are classified as extended spectrum beta lactamases ESBL and they have been implicated in clinical resistance to monobactams and broad spectrum cephalosporins such as ceftazidime CEZDI cefotaxime CFTAX and ceftriaxone CETRX Some ESBLs confer high level resistance to these beta lactams and are easily detected as resistant or intermediate by disk tablet diffusion testing But the ESBL may provide low levels of resistance MIC 1 2 ug ml to monobactams and third generation cephalosporins that can be easily overlooked by routine susceptibility
10. CEFTOBIPROLE 30 ug CFBIP 30 36 26 34 23 31 CARBENICILLIN 115 ug CARBE 26 32 21 27 CEFACLOR 30 ug CCLOR 22 27 25 33 CEFAZOLIN 60 ug CFZOL 26 32 28 36 CEFEPIME 30 ug CFEPM 31 38 26 33 CEFPIROME 30 ug CFPIR 31 38 26 32 18 22 CEFOTAXIME 30 ug CFTAX 30 36 28 34 18 25 CEFOTETAN 30 ug CFTTN 30 36 15 20 x CEFOXITIN 60 ug CFOXT 29 35 26 34 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 12 Zone diameter in mm Page 81 of 170 E coli S aureus Ps aeruginosa Ent faecalis NEO SENSITABS CODE ATCC 25922 ATCC 25923 ATCC 27853 ATCC 29212 CEFSULODIN 30 ug CFSUL 27 33 CEFTAZIDIME 30 ug CEZDI 27 33 18 23 22 30 CEFTRIAXONE 30 ug CETRX 29 35 21 28 17 24 CEFUROXIME 60 ug CEFUR 24 30 29 36 CEPHALEXIN 30 ug CFLEX 16 21 CEPHALOTHIN 66 ug CLOTN 21 27 32 40 18 23 Cephalosporins CHLORAMPHENICOL 60 ug CLR60 24 32 23 30 21 27 clear zone CINOXACIN 30 ug CINOX 20 26 CIPROFLOXACIN 10 ug CIP10 30 40 21 29 24 32 17 24 CIPROFLOXACIN 0 5 ug CIP L 24 34 CLARITHROMYCIN 30 ug CLARI 24 31 CLINDAMYCIN 25 ug CLIND 30 38 COLISTIN 2 18 h predif 10 ug Co 10 22 28 17 23 DAPTOMYCIN 30 ug DAPCa 22 28 2 18 h prediffusion DOXYCYCLINE 80 ug DOXYC 25 31 29 35 18 23 DORIPENEM 10 ug DORIP 28 35 33 42 29 35 ERTAPENEM 10 ug ERTAP 29 36 24 31 13 21 ERYTHROMYCIN 78 ug ERYTR 26 33 19 24 FAROPENEM 10 ug FAROP 20 26 27 24 FOSFOMYCIN 70 40
11. NEO SENSITABS Vb Danish blood agar Interpretation valid for Denmark Aflaesningsskema for Neo Sensitabs hurtigvoksende bakterier DIREKTE METODE Resistensplade DBA Semikonfluerende vaekst 09 2007 2008 Chapter 10 Page 53 of 170 Zone diameter i mm MIC pg ml NEO SENSITABS STYRKE KODE 3 2 1 0 3 2 1 0 Amoxycillin 30 ug gt 28 2027 15 19 lt 14 lt 2 3 6 8 32 gt 32 b Amoxycillin 30 15 ug AM CL gt 28 20 27 15 19 lt 14 lt 2 3 6 8 32 gt 32 Clavulanate Amikacin 40 ug AMIKA gt 26 23 25 15 22 lt 14 lt 4 6 8 12 64 gt 64 a Ampicillin 33 ug AMP33 gt 28 2327 15 22 lt 14 lt 2 3 6 8 32 gt 32 Azithromycin 30 ug AZITR 222 19 21 13 18 lt 12 lt 1 5 2 4 6 16 gt 16 c Aztreonam 30 ug AZTRM gt 28 2027 15 19 lt 14 lt 2 3 8 16 32 gt 32 j Chloramphenicol 60 ug CLR60 gt 28 23 27 15 22 14 lt 6 8 16 32 64 gt 64 b c Cefepime 30 ug CFEPM gt 28 20 27 15 19 14 lt 2 3 8 16 32 gt 32 Cefepime Clav 30 10ug CP CL P visning af ESBL b c Cefotaxime 30 ug CFTAX gt 28 20 27 15 19 lt 14 lt 2 3 8 16 31 gt 32 b k Cefoxitin 60 ug CFOXT gt 28 2327 15 22 lt 14 lt 3 4 6 8 12 64 gt 64 S aureus og S lugdunensis gt 30 lt 27 Mec A pos Coag neg staph 234 lt 31 OxaS Mec A pos b k Cefoxitin 10 ug CFO10 S aureus og S lugdunensis gt 22 22 OxaS Mec A pos Coag neg staph 227 22 26 lt 21 OxaS Mec A pos b c Cefpirome
12. P aeruginosa St maltophilia samt Proteus spp b r rapporteres R mod Tetracyclines Neo Sensitabs uanset zonest rrelse P aeruginosa b r ogs rapporteres mod Chlormaphenicol og Streptomycin O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 56 of 170 Cefoxitin 60 ug og 10 ug er bedst til p visning af methicillin resistente staphylococcer Med Cefoxitin 60 ug S aureus med zoner lt 28 mm er mecA positive For coag neg staph anvendes 32 mm Med Cefoxitin 10 ug S aureus med zoner lt 22 mm mec A pos Coag neg staph zoner lt 27 mm mec A pos I Klebsiella og Enterobacter rapporteres altid R mod ampicillin m Staphylokokker resistente overfor gentamicin bgr rapporteres R mod netilmicin og tobramycin n Nalidixan er en god screening til p virkning af nedsat f lsomhed overfor kinoloner hos Salmonella spp Stammer resistente overfor Nalidixan Neo S zone lt 26 mm viser nedsat f lsomhed ovefor kinoloner CIPRO MIC gt 0 125 ug ml 1 0 coli Klebsiella Salmonella stammer der udviser zoner lt 23 mm overfor Cefpodoxime Neo Sensitabs producerer formodentlig ESBL enzymer Til bekreftelse test Ceftazidime Clavulanate og Cefepime Clavulanate mod Ceftazidime Cefepime Neo Sensitabs se side 13 9 1 p Mecillinam Neo Sensitabs kan bruges som screening for beta lactamase produktion hos staphylococcer Mecillinam resistente staphylococcer er ogsa penicillin resistente beta l
13. User s Guide TM NEO SENSITABS SUSCEPTIBILITY TESTING 19th Ed 2007 2008 y gJ 5 um _ me ROSCO DIAGNOSTICA NEO SENSITABS 09 2007 2008 Page 2 of 170 USER s GUIDE NEO SENSITABS SUSCEPTIBILITY TESTING 19th Ed 2007 2008 Contents T Susceptibiaty Testing in General nope DIR SE e IRR nU 6 2 Characteristics of NEO SENSITABS sese enne eene nene trente entren reete bens en EEN ER tnnt tenete nennen trennen nen 7 2 1 Storage of NEO SENSITABS saae are a nen nret nennen teen teen nennen ten tenet nennen nete nnne 7 3 NEO SENSITABS Range Al sort he Cp e RO SOE E EEE EE ER GERD ER CORRUPTI ERR EL DRE PRSE 9 34 jAntb cterials rer EE OR e ecd Rip pe ett s ei e ptt eee 9 32 JAn fungals uer RUE OR E EEEE I quieta t tue e qti 13 4 Susceptibility Test Media diee e e ed nae eee ette 14 5 Primary vs Pure Culture Susceptibility Testing Early Reading of Antibiogramme sse 15 6 Inoculum Standardization and Prediffusion Method eese enne nennen nen 17 6 1 Inoculum Standardization ICS and CLSI Kirby Bauer eese nennen nennen 17 6 1 1 Inoculum according to ICS sonnerie d den enne 17 6 1 2 Inoculum according to CLSI Kirby Bauer essere rene 17 6 2 Pr diffusion Method eee tete PR DERE ERROR EU DOE reiner 18 7 Dispensers and Templates Schabelons reerrrarnrernvnvvnenvnrn
14. dextrose glucose and 0 5 ug ml methylene blue with the undiluted 0 5 McFarland Standard using a sterile cotton wab dipped into the suspension removing excess fluid by pressing aginst the tube Inoculate the agar by streaking while rotating the plate swabbing over the entire agar surface 2 Incubation time Reading of the test should be made as soon as possible i e for yeasts after not more than 18 24 hours A longer incubation time may result in false resistance against imidazoles azoles Plates should always be examined after overnight incubation and if inhibition zones are visible they must be measured If growth is not yet visible with particular strains the plates may be reincubated for up to 24 hours more For Cryptococcus spp incubate at 30 for 42 48 hours 3 Criteria for measuring inhibition zones For the polyenes Amphotericin B Nystatin the clear zone with no visible growth is measured Colonies inside the zone of inhibition must be considered resistant mutants For azoles imidazoles Caspofungin and Terbinafine Candida measure the zones must be measured up to colonies of normal size There is often a zone of growth of partially inhibited colonies smaller at the edge of the real zone They are not resistant mutants Fluorocytosine cannot be tested with MH Glucose Methylene Blue agar because of the presence of antagonists in the agar It may be testing using Shadomy agar or similar agars free of antagonists In
15. pneumoniae 226 25 21 lt 21 lt 1 gt 2 Moraxella catarrhalis gt 30 29 26 lt 26 lt 0 5 gt 0 5 Trimethoprim Sulfa 5 2 240 ug TR SU Haemophilus spp gt 32 31 26 lt 26 lt 16 gt 32 S pneumoniae strep gt 32 31 26 lt 26 lt 16 gt 32 Moraxella catarrhalis gt 32 31 26 lt 26 lt 16 232 Remarks a Beta lactamase production in M catarrhalis can be detected by measuring the zones of inhibition around Amoxycillin and Amoxycillin Clavulanate Neo Sensitabs If the zone around Amoxycillint Clavulanate is gt 5 mm larger than around Amocyxillin alone the strain produces beta lactamase BRO 1 BRO 2 Beta lactamase positive isolates are resistant to penicillin amoxycillin ampicillin piperacillin and ticarcillin b I ug is used for the detection of reduced sensitivity to penicillin in pneumococci Penicillin resistant isolates from the meninges must be considered resistant to ampicillin amoxycillin amoxycillin clavulanate first and second generation cephalosporins c Oxacillin 1 ug may be used for the detection of viridans streptococci with reduced sensitivity to penicillin Strains with zones 16 mm should be tested by an MIC method d Beta lactamase negative ampicillin resistant strains BLNAR are best detected using Ampicillin 2 5 ug Neo Sensitabs BLNAR isolates must be considered resistant to amoxycillin amoxycillin clavulanate as well as first and second generation cephalosporins no matter the size of th
16. 13 12 11 lt 10 lt 8 gt 32 Doxycycline 80 ug DOXYC gt 28 27 24 lt 23 lt 2 28 Gatifloxacin 5ug GATIF 218 lt 1 b Imipenem 15 ug IMIPM 222 4 b d Levofloxacin 5ug LEVOF gt 20 lt 2 b Meropenem 10 ug MEROP 226 z lt 0 5 b d Moxifloxacin 5ug MOXIF gt 18 lt 1 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 100 of 170 Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R d Nalidixan 130ug NALID 25 reduced susceptibility to quinolones b d Ofloxacin 10 ug OFLOX gt 22 2 Penicillin Low 5ug PEN L 220 19 17 lt 16 lt 1 gt 4 Rifampicin 30 ug RIFAM gt 28 27 24 lt 23 lt 1 gt 4 Telithromycin 18 15 ug TEL15 gt 15 14 12 lt 11 lt 4 gt 16 Tetracyclines 10 ug TET10 220 19 17 16 lt 2 gt 8 Tetracyclines 80 ug TET80 gt 30 29 27 lt 26 lt 2 gt 8 Trimethoprim Sulfa 5 2 240 ug TR SU gt 28 27 24 lt 23 lt 0 5 9 5 24 76 Trimethoprim 5 2 ug TRIME gt 23 22 20 lt 19 lt 1 24 Special breakpoints see 4 page 101 MIC breakpoints have not been established by the CLSI a Beta lactamase negative ampicillin resistant BLNAR strains are best detected using Ampicillin 2 5 ug These strains should be considered resistant also to amoxycillin amoxycillint clavulanate ampicillin sulbactam cefaclor cefonicid and cefuroxime despite apparent in vitro susceptibility of some
17. 26 I gt 1 Metronidazole 16 ug MTR16 gt 26 25 23 lt 22 lt 8 gt 16 Tetracycline 10 ug TET10 gt 23 22 20 lt 19 lt 2 gt 8 MIC breakpoints not yet established by the CLSI References 1 Huaxiang X et al Standardisation of Disk Diffusion Test and clinical significance for susceptibility testing of Metronidazole against Helicobacter pylori Antimicr Ag Chemother 38 2357 2361 1994 2 Hachan C Y et al Antimicrobial susceptibility testing of Helicobacter pylory Diagn Microbiol Infect Dis 24 37 41 1996 3 S rberg M et al Risk of development of in vitro resistance to Amoxycillin Clarithromycin and Metronidazole in Helicobacter pylori Antimicro Ag Chemother 42 1222 1228 1998 4 Megraud F et al Antimicrobial susceptibility testing of Helicobacter pylori in a large multicenter trial the MACH 2 study Antimicro Ag Chemother 43 2747 2752 1999 5 Kwon D H et al Isolation and characterization of tetracycline resistant clinical isolates of Helicobacter pylori Antimicro Ag Chemother 44 3203 3205 2000 6 Kalach N et al High levels of resistance to Metronidazole and Clarithromycin in Helicobacter pylori strains in children J Clin Microbiol 39 394 397 2001 7 Glupczynski Y et al European multicenter survey of in vitro antimicrobial resistance in Helicobacter pylori EJCMID 20 820 823 2001 8 McNulty C et al Helicobacter pylori susceptibility testing by disc diffusion
18. Clin Microbio 45 2798 2801 2007 O Copyright Rosco Diagnostica A S NEO SENSITABS 16 7 Detection of multiple beta lactamases in one strain 09 2007 2008 Page 145 of 170 Chapter 16 Diagnostic problems posed by coexistance of different classes of beta lactamases in a single bacterial isolate could be solved by the combined use of various phenotypic detection methods See below example with multiresistant K pneumoniae from Taiwan and USA Neo Sensitabs K pneumoniae Cefoxitin Cefepime Ceftazidime D P A Boronic 250 ug producing Clavulanate Ceftazidime Cefoxitin Clav or or Cefepime Clav Cefepime Imipenem or Clavulanate EDTA Cloxacillin 500 ug synergy synergy Cefoxitin Ceftazidime synergy AmpC R S negative negative POSITIVE ESBL S V I R POSITIVE negative negative AmpC ESBL R I R POSITIVE negative POSITIVE AmpC metallo B R I R negative POSITIVE POSITIVE lactamases AmpC ESBL R I R POSITIVE POSITIVE POSITIVE metallo B lactamases References 1 Jing Jou Yan et al Complexity of Klebsiella pneumoniae isolates resistant to both cephamycins and extended spectrum cephalosporins at a teaching hospital in Taiwan J Clin Microbiol 42 5337 40 2004 2 Smith Moland E at al Klebsiella pneumoniae isolate producing at least 8 different beta lactamases including AmpC and KPC beta lactamases Antimicr Ag Chemother 51 800 801 2007 O Copyright Rosco Diagnostica A S
19. DHA Induc plasmid AmpC DHA ESBL 33 Amp C ESBL DHA ESBL ESBL Metallo beta lactamases 24 28 ESBL 16S rRNA methylases 26 Synergism Ceftazidime Clavulanate and or Cefepime Clavulanate Cefoxitin S Synergism Ceftazidime Clavulanate and or Cefepime Clavulanate Synergism Imipenem and Ceftazidime and or Cefoxitin and Ceftazidime No synergism Ceftazidime Clavulanate and Cefepime Clavulanate Cefoxitin R Ceftazidime R Synergism Cefoxitin and Cloxacillin 500 ug and or Ceftazidime and Cloxacillin 500 ug Antagonism Clavulanate AMC and 3rd generation Synergism Cefoxitin Ceftazidime and Cloxacillin 500 ug Antagonism Clavulanate AMC and 3rd gen cephalosporins DHA Synergism Tazobactam Piperacillin Tazobactam and Ceftazidime Cefepime Synergism Cefepime Clavulanate ESBL 31 Cefoxitin R Ceftazidime R Synergism Cefoxitin or Ceftazidime and Cloxacillin 500 ug AmpC Synergism Ceftazidime Clavulanate and or Cefepime Clavulanate ESBL Antagonism Clavulanate AMC and 3rd gen cephalosporins DHA ACT 1 Synergism Aztreonam Clavulanate Amoxicillin Clavulanate ESBL Synergism Imipenem EDTA metallo beta lactamases Synergism Ceftazidime DPA metallo beta lactamases Synergism Ceftazidime Clavulanate and or Cefepime Clavulanate Cefoxitin S No zone with Amikacin Gentamicin Tobramycin Neo Sensitabs 2 High level K 1 Klebsiella oxytoca no ESBL K 1 ESBL 23 No synergism
20. For susceptibility testing of CNS except S lugdunensis we recommend e Direct inoculum suspension of colonies from an overnight culture in 0 9 96 saline McFarland 0 5 e Diffusion test with Cefoxitin Neo Sensitabs and Oxacillin 1 ug Neo Sensitabs incubated at 33 35 C on Mueller Hinton agar without NaCl added e Incubation for 24 hours with reincubation for a total of 48 hours in doubtful cases e Interpretation For Cefoxitin 60 ug use 5 gt 28 mm oxacillin S and R lt 27 mm mecA positive oxacillin R S gt 18 mm MIC lt 0 25 ug ml and R x 17 mm MIC gt 0 5 ug ml for Oxacillin 1 ug MecA negative isolates that show MICs gt 4 ug ml towards oxacillin should be reported as oxacillin resistant NCCLS 2003 Testing of oxacillin against urine isolates of S saprophyticus is not recommended because mecA negative strains of S saprophyticus often appear resistant by interpretative criteria used for coagulase negative staphylococci M100 S11 NCCLS 2001 In general routine testing of urine isolates of S saprophyticus is not advised because infections respond to antimicrobial agents commonly used to treat acute uncomplicated urinary tract infections 2 Cefoxitin Neo Sensitabs can be used to detect mec A positive strains of S saprophyticus For S lugdunensis see recommendations for S aureus Chapter 13 1 Methicillin resistant CNS are even more resistant to multiple antibiotics than S aureus The observatio
21. HLR gt 2000 LINEZOLID 30 ug LINEZ 23 29 S PENICILLIN LOW 5ug PEN L 12 15 RIFAMPICIN 30 ug RIFAM 21 26 STREPTOMYCIN 500 ug zone HLR gt 2000 TEICOPLANIN 60 ug TEICO 17 23 S TETRACYCLINES 80 ug TET80 24 30 VANCOMYCIN 5ug VAN 5 12 14 I R 16 32 MH agar inoculum McF 0 5 incubation 35 16 18 hours 24 hours for vancomycin References 1 Jenkins R D et al False susceptibility of enterococci to aminoglycosides with blood enriched Mueller Hinton Agar for disk susceptibility testing J Clin Microbiol 22 369 374 1985 2 Sahm D F et al Medium dependent zone size discrepancies associated with susceptibility testing of group D streptococci against various cephalosporins J Clin Microbiol 18 858 865 1983 3 Eliopoulos G M et al Effect of blood product medium supplements on the activity of cefotaxime and other cephalosporins against Enterococcus faecalis Diagn Microbiol Infect Dis 12 149 156 1989 4 CLSI Performance Standards for Antimicrobial Susceptibility Testing 17th Inf Suppl M100 S17 2007 5 Navarro F Mecanismos de resistencia a los glicopeptidos Enferm Infecc Microbiol Clin 14 317 323 1996 Spanish 6 Torres C et al Detection of Aminoglycoside penicillin synergy against Enterococcus faecium using high content aminoglycoside disks Eur J Clin Microbiol Infect Dis 14 878 882 1995 7 Centinkaya Y et al Vancomycin resistant enterococci Clin Microbiol Reviews 1
22. Imipenem EDTA 15 750 ug IMIED Detection of metallo B lactamases lt 4 gt 8 y Ertapenem 10 pg ETP10 Enterobacteriaceae gt 30 lt 30 lt 0 25 gt 2 Glycopeptides Vancomycin 5 ug VAN 5 216 15 14 14 lt 4 gt 8 i 2 2 18 h prediffusion staph lt 22 VISA GISA 2 18 h prediffusion enterococci lt 16 VRE Teicoplanin 30 ug TPN30 gt 16 15 14 lt 14 lt 4 gt 8 i x z 2418 h prediffusion lt 20 VISA GISA y Daptomycin 30 ug DAPCa 2 2 18 h prediffusion Staphylococcus spp 222 lt 1 Enterococcus spp gt 12 lt 4 Macrolides j Erythromycin 78 ug ERYTR u Staphylococcus spp gt 28 27 24 lt 24 lt 1 22 Enterococcus spp Clarithromycin 30 ug CLARI Staphylococcus spp gt 22 21 18 lt 18 lt 1 gt 2 Enterococcus spp Azithromycin 30 ug AZITR Staphylococcus spp 222 21 18 18 1 22 Enterococcus spp Lincosamides Clindamycin 25 ug CLIND Staphylococcus spp gt 26 25 22 lt 22 lt 1 22 Aminoglycosides Gentamicin 40 ug GEN40 Enterobacteriaceae gt 26 25 22 lt 22 lt 2 gt 4 k Staphylococcus spp gt 30 29 26 lt 26 lt 1 gt 1 O Copyright Rosco Diagnostica A S NEO SENSITABS Zone diameter 09 2007 2008 Chapter 10 Page 46 of 170 Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S l R S R Netilmicin 40 ug NETIL Enterobacteriaceae gt 26 25 22 lt 22 lt 2 gt 4 l Staphylococcus spp gt 30 29 26 lt 26 lt 1 gt 1 T
23. NEO SENSITABS 09 2007 2008 Chapter 15 Page 99 of 170 15 1 Susceptibility Testing of Haemophilus influenzae and parainfluenzae The medium recommended for diffusion testing of Haemophilus spp is Haemophilus Test Medium HTM It consist of the following ingredients Mueller Hinton agar 15 ug ml NAD 15 ug ml bovine hematin 5 mg ml yeast extract pH adjusted to 7 2 7 4 To make HTM 50 mg bovine hematin is dissolved in 100 ml 0 01 N NaOH with heat and stirring 30 ml of this solution are added to 1 I MHA with 5 g yeast extract Autoclave cool to 45 50 C and add aseptically 3 ml of NAD sol 50 mg NAD dissolved in 10 ml H 0 and filter sterilized pH adjusted to 7 2 7 4 Procedure 1 Colonies are taken directly from an overnight 20 24 h chocolate agar culture and a suspension prepared in broth or 0 9 saline The suspension is adjusted to a 0 5 McFarland standard using a photometer Within 15 min after adjusting the turbidity of the suspension it should be used for plate inoculation Notice that too high inoculum may lead to false resistant results with some beta lactam antibiotics Apply in general no more than 9 Neo Sensitabs to the surface of a 140 150 mm plate and no more than 4 Neo Sensitabs on 90 100 mm plate Incubate at 35 C in an atmosphere of 5 7 for 16 18 h before measuring the zones of inhibition The zone margin should be considered as the area showing no obvious growth visible with the unaided
24. NEO SENSITABS POTENCY CODE S R Page 112 of 170 Zone diameter Break points in mm MIC pg ml S R f non meningeal criteria 9 f Amoxycillin 30 ug AMOXY gt 26 25 21 lt 20 lt 2 gt 8 f Amoxycillin Clav 30 15 ug AM CL gt 26 25 21 lt 20 lt 2 1 gt 8 4 f Cefepime 30 ug CFEPM gt 28 27 24 lt 23 lt 1 gt 4 f Cefotaxime 30 ug CFTAX gt 28 27 24 lt 23 lt 1 gt 4 f Cefpodoxime 30 ug CFPOX 232 31 27 lt 26 lt 0 5 gt 2 f Ceftriaxone 30 ug CETRX 228 27 24 lt 23 lt 1 gt 4 f Cefuroxime oral 60 ug CEFUR 230 29 27 lt 26 lt 1 gt 4 Note Penicillin cefotaxime or ceftriaxone and meropenem should be tested by a reliable MIC method and reported routinely with CSF isolates of S pneumoniae Comment For cefotaxime use of interpretation criteria for non meningitis requires doses appropriate for serious pneumococcal infections e g at least 1 g adults or 50 mg kg children every 8 hours or more frequently 2 a b d e g h Tentative 17 Isolates with Oxacillin I ug zone gt 20 mm are susceptible MIC x 0 06 ug ml to penicillin A penicillin MIC as well as a cefotaxime ceftriaxone MIC should be determined on isolates of S pneumoniae with Oxacillin 1 ug zone sizes 19 mm penicillin I R Penicillin resistant strains from the CSF should be considered resistant to ampicillin amoxycillin amoxycillin clavulanate and first second generation cephalosporins Str
25. aureus 700788 see chapter 13 5 Store working control cultures on Tryptic soy agar at 4 8 C and subculture weekly Replace working cultures once a month from frozen liophylized or commercial cultures Before testing the strains should be streaked onto agar plates to obtain single colonies Colonies should then be picked and suspended in 0 9 saline for testing according to the recommended susceptibility testing procedures Each laboratory should record the result of consecutive separate analysis using the control cultures and each antimicrobial to be controlled If an unexplained result suggests a change in the organism s susceptibility a fresh culture of the control strain should be obtained Zone Diameters Quality Control Limits Quality control should be performed at least once a week and every time a new lot of Mueller Hinton is introduced The control limits using Mueller Hinton Agar and inoculum according to CLSI Kirby Bauer have been established as follows Control limits on Mueller Hinton agar Inoculum according to CLSI Kirby Bauer Zone diameter in mm E coli S aureus Ps aeruginosa Ent faecalis NEO SENSITABS CODE ATCC 25922 ATCC 25923 ATCC 27853 ATCC 29212 AMIKACIN 40 ug AMIKA 24 30 22 29 25 31 AMOXYCILLIN 30 ug AMOXY 21 26 26 32 AMOXYCILLIN 30 15 ug 22 27 28 36 CLAVULANATE AMPICILLIN 33 ug AMP33 22 27 25 31 AZITHROMYCIN 30 ug AZITR 21 27 AZTREONAM 30 ug AZTRM 29 36 24 30
26. but when properly used the diffusion method is also able to detect CAT producing strains The diffusion method using a break point of 2 ug ml clearly separates chloramphenicol susceptible from resistant strains C Fluoroquinolones In Spain 4 ciprofloxacin resistant H influenzae have been isolated from patients with cystic fibrosis Vigilance for quinolone resistant H influenzae should be maintained in patients with chronic infections treated with quinolones Ciprofloxacin resistant H influenzae have also been isolated in the UK 11 South Africa 12 and Spain 14 The use of Nalidixan 25 mm and Ciprofloxacin 0 5 ug Neo Sensitabs with interpretations S gt 18 mm MIC 0 12 ug ml I 17 15 mm lt 14 mm MIC gt 1 ug ml is recommended in such cases Current CLSI MIC breakpoints will not always detect resistance In a long term care facility in New York 36 levofloxacin resistance was found among influenzae isolates in 2001 9 For the detection of decreased susceptibility to fluoroquinolones due to changes in gyrA and ParC the following breakpoints should be used 14 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE R S R Nalidixan 130ug NALID 25 gt 16 Ciprofloxacin 0 5 ug CIP L gt 18 17 15 lt 14 lt 0 12 gt 1 Ciprofloxacin 10ug CIP10 gt 32 lt 0 12 Levofloxacin 5ug LEVOF 230 lt 0 25 Gatifloxacin 5ug GATIF 230 gt lt 0 25 Moxifloxa
27. colony growth inside the inhibition zone Report as resistant to telithromycin 12 Interpretative criteria for streptococci are based on population distributions of various species pharmacokinetics of the antimicrobials previously published literature and the clinical experience of certain members of the CLSI Subcommittee 2 Oxacillin 1 ug Neo Sensitabs is useful for screening for penicillin susceptibility in streptococci Strains with inhibition zones gt 14 mm are susceptible to penicillin while strains showing zones lt 13 mm I R should be tested for penicillin susceptibility by an MIC method Oxacillin 5 ug Neo Sensitabs is also useful SFM 2002 Since 1983 there has been several reports disclosing high rates of penicillin resistant viridans streptococci isolated from clinical significant infections Penicillin resistance is due to alterations in the PBP s A study of 410 consecutive viridans streptococcal isolates from blood to 22 beta lactams was performed in Barcelona 3 33 6 were found resistant to penicillin MIC 0 5 8 ug ml The most active drug against the resistant strains was imipenem followed by cefpirome cefotaxime ceftriaxone and cefepime Rodriguez Avial et al 6 found 49 erythromycin resistant and 46 penicillin resistant viridans strains Decreased activity of erythromycin against S pyogenes and other beta haemolytic streptococci has been reported from several countries 4 5 7 8 Horizontal gene tra
28. gt 38 37 27 lt 26 lt 0 12 gt 2 f S pneumoniae Test ceftizoxime som surrogat lt 0 12 gt 1 Haemophilus gt 38 37 29 lt 28 lt 0 12 gt 1 Aztreonam 30 ug AZTRM Haemophilus spp 240 39 35 lt 34 lt 0 5 gt 1 Imipenem 15 ug IMIPM Streptococcus spp gt 36 35 24 lt 23 lt 0 06 gt 2 pneumoniae gt 36 35 29 lt 28 lt 0 06 gt 0 5 Haemophilus gt 34 33 29 lt 28 lt 1 gt 2 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 63 of 170 Zone diameter Break points imm MIC pg ml NEO SENSITABS STYRKE KODE S l R S Meropenem 10 ug MEROP Streptococcus spp gt 36 35 24 lt 23 lt 0 06 gt 2 pneumoniae gt 36 35 27 amp 26 lt 0 06 gt 0 5 Haemophilus gt 34 33 29 lt 28 lt 1 gt 2 Ertapenem 10 ug ETP10 Streptococcus spp gt 30 29 26 lt 26 lt 0 5 gt 1 S pneumoniae gt 30 29 26 lt 26 lt 0 5 gt 1 Haemophilus spp Moraxella catarrhalis gt 30 29 26 lt 26 lt 0 5 gt 1 Anaerobes gt 32 31 28 lt 28 1 21 Erythromycin 78 ug ERYTR d 9 X Streptococcus spp S pneumoniae 228 lt 27 lt 0 5 gt 0 5 Moraxella catarrhalis gt 32 31 22 lt 21 lt 0 5 gt 4 Corynebacterium spp Listeria spp 232 31 22 lt 21 lt 0 5 gt 4 Clindamycin 25 ug CLIND d Streptococcus spp S pneumoniae 232 31 27 lt 26 lt 0 5 gt 2 Chloramphenicol 60 ug CLR60 pneumoniae 226 25 23 lt 22 lt 8 gt 16 Haemophilus spp gt 34 33 31 lt 30 lt 2 gt 4 Te
29. gt 2 Tetracyclines 10 ug TET10 gt 20 19 17 lt 17 lt 2 gt 2 Doxycycline 80 ug DOXYC Haemophilus spp HTM gt 28 27 24 lt 24 lt 2 gt 2 Haemophilus spp MH PDM gt 25 24 20 lt 20 lt 4 gt 4 pneumoniae gt 26 25 23 lt 23 lt 1 22 Moraxella catarrhalis Quinolones Nalidixan 130ug NALID Reduced susceptibility Haemophilus spp 30 to quinolones Ciprofloxacin 10ug CIP10 Haemophilus spp gt 30 29 26 lt 26 lt 0 5 gt 0 5 i S pneumoniae 236 35 22 22 lt 0 125 22 Moraxella catarrhalis gt 32 31 28 lt 28 lt 0 125 gt 0 5 Norfloxacin 10 ug NORFX Reduced susceptibility pneumoniae 16 to quinolones O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 51 of 170 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Others Trimethoprim Sulfa 5 2 240 ug TR SU gt 32 31 26 lt 26 lt 0 5 9 5 gt 2 38 Moraxella catarrhalis gt 30 29 26 lt 26 lt 1 19 gt 2 38 Linezolid 30 ug LINEZ pneumoniae gt 28 27 23 lt 23 lt 4 gt 4 Quinupristin Dalfopristin 15 ug SYNI5 S pneumoniae streptococci gt 18 lt 18 lt 2 gt 2 BL pos Beta lactamase producing strains BL neg Non beta lactamase producing strains Haemophilus spp Haemophilus spp and S pneumoniae Remarks a b c d e 8 h Beta lactamase production in M catarrhalis can be detected by measuring the zones of inhibition around Amoxyc
30. k N gonorrhoeae 32 to quinolones i N meningitidis lt 28 Campylobacter spp gt 28 27 24 lt 24 lt 8 gt 16 Trimethoprim Sulfa 5 2 240 ug TR SU Haemophilus spp gt 36 lt 0 5 9 5 gt 1 19 S pneumoniae Streptococcus spp 230 29 26 26 gt 2 38 gt 8 152 Gentamicin 40 ug GEN40 Haemophilus spp gt 26 25 23 lt 23 lt 2 gt 4 Campylobacter spp gt 28 27 23 lt 23 lt 2 gt 4 Tobramycin 40 ug TOBRA Campylobacter spp 228 27 23 23 lt 2 gt 4 Spectinomycin N gonorrhoeae gt 22 lt 22 64 gt 64 Teicoplanin 30 ug TPN30 pneumoniae Streptococcus spp anaerobes gt 18 18 lt 4 gt 8 Vancomycin 5 ug VAN 5 S pneumoniae Streptococcus spp gt 16 16 lt 4 gt 8 Anaerobes gt 18 lt 18 lt 4 gt 8 m Metronidazole 16 ug MTR16 Anaerobes lt 21 lt 4 gt 16 Clostridium difficile 1 gt 32 31 30 lt 30 lt 4 gt 16 Remarks Pneumococci a Oxacillin 1 ug or 5 ug are used for the detection of reduced sensitivity to penicillin in pneumococci Penicillin resistant isolates from the meninges must be considered resistant to ampicillin amoxycillin amox clav and first and second generation cephalosporins b Cefotaxime and ceftriaxone must not be tested against pneumococci by the diffusion method A surrogate test is used instead Ceftizoxime ceftizoxime detects reduced sensitivity to third generation cephalosporins Strains sensitive to ceftizoxime show currently MIC 0 5
31. lt 22 lt 2 28 Swine gt 30 29 27 lt 26 lt 0 5 22 Other organisms 223 22 20 lt 19 lt 4 gt 16 Tiamulin 30 ug TIAMU Spirochaetae gt 28 27 24 lt 23 lt 1 gt 4 Actinobacillus gt 11 zone lt 16 gt 32 Tiamulin Tetra 30 15 ug gt 16 15 14 lt 13 lt 8 4 gt 16 8 Ticarcillin 75 ug TICAR Ps aeruginosa gt 16 lt 15 lt 64 gt 128 Gram neg ent 220 19 17 lt 16 lt 16 2128 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 22 Page 170 of 170 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Ticarcillin Clavulanate 75 15 ug TI CL Ps aeruginosa gt 16 lt 15 lt 64 2 2 128 2 Gram neg enteric organisms gt 20 19 17 lt 16 lt 16 2 2 28 2 Tilmicosin 80 ug TILMI Bovine RD gt 18 17 15 lt 14 lt 8 gt 32 Swine RD gt 15 lt 14 lt 16 gt 32 Trimethoprim 5 2 ug TRIME 220 19 17 lt 16 lt 4 gt 16 9 Trimethoprim Sulfa 5 2 240 ug TR SU S pneumoniae gt 32 31 27 lt 26 lt 0 5 9 5 24 76 Systemic infection gt 32 31 27 lt 26 lt 0 5 9 5 24 76 Urine gt 28 27 24 lt 23 lt 2 38 gt 4 76 c Tylosin 150ug gt 26 25 23 lt 22 lt 4 gt 16 Vancomycin 5 ug VAN 5 216 15 14 lt 13 lt 4 gt 32 Remarks a Results with Ceftriaxone are valid for Cefoperazone b Metronidazole 16 ug is the representative of the nitroimidazole group including ronidazole ornidazole ipronidazole and moxnidazole Results obta
32. lt 24 lt 0 25 gt 0 5 Haemophilus spp gt 28 lt 28 lt 0 25 gt 0 25 Daptomycin 30 ug DAPCa Prediffusion 2418 h Staphylococci 222 21 20 20 lt 1 gt 1 Streptococcus spp non pneumoniae 222 21 20 20 lt 1 gt 1 Enterococci gt 12 lt 12 lt 4 gt 4 nal Use Nalidixan Neo Sensitabs as surrogate test Strains resistant to Nalidixan show reduced susceptibility to fluoroquinolones 5 6 7 Zones tentative for 1 year T Technique described on page 18 Prediffusion technique for detecting GISA VISA strains as well as VRE described on page 18 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 11 Page 79 of 170 11 Interpretation of Results Susceptible S The infection due to the strain tested may be expected to respond to a normal dosage of this antimicrobial Intermediate 1 The intermediate category implies clinical applicability in body sites where the drugs are concentrated e g urine or when high dosage of an antimicrobial can be used e g betalactams The intermediate category also comprises a buffer zone which should prevent small uncontrolled technical factors from causing major discrepancies in interpretations thus when a zone falls within the intermediate range the results may be considered equivocal and if alternative drugs are not available MIC testing may be indicated Resistant R The antimicrobial cannot be recommended for treatment in
33. lt 2 Fluorocytosine 1 pg FLU 1 gt 26 lt 1 Voriconazole 1 ug VOR 1 gt 22 lt 0 25 E Tentative 21 2 2 Candida spp Quality Control modified Shadomy Agar Quality control limits using ATCC strains on modified Shadomy agar are included in the table below C albicans C albicans C parapsilosis C krusei NEOSENSITABS ATCC 64548 ATCC 64550 ATCC 22019 ATCC 6528 Amphotericin B 18 23 19 24 20 26 15 21 Caspofungin 15 22 13 23 15 22 Fluconazole 36 42 11 17 30 36 10 16 Fluorocytosine 1 ug 34 40 26 33 35 43 9 Itraconazole 25 31 9 28 35 16 22 Ketoconazole 36 44 21 29 41 49 20 26 Posaconazole 28 39 25 36 23 31 Voriconazole 31 42 28 37 16 25 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 21 Page 166 of 170 References 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 Galgiani J N Susceptibility of Candida albicans and other yeasts to fluconazole Relation between in vitro and in vivo studies Rev Infect Dis 12 Suppl 3 S272 S275 1990 Barry A L et al Fluconazole disk diffusion procedure for determining susceptibility of Candida species J Clin Microbiol 34 2154 2157 1996 Quind s G et al Multicenter survey of in vitro antifungal resistance in yeasts of medical importance isolated from Spanis
34. 0 12 wk Sh putrefaciens Not integron OXA 55 S S S 1 4 0 25 no wk Sh algae 9 Chromosomal OXA 58 256 4 128 gt 32 2 32 2564 no no A baumanii Plasmid OXA 60 S S R 0 5 2 no R pickettii Chromosomal OXA 62 SoR S gt R SoR 22564 645128 no no Pandorea 10 Chromosomal pnomenusa OXA 23 8532 85232 Ac baumannii Plasmid 27 49 only 23 subgroup 1 D OXA 24 Ac baumannii Chromosomal 8 25 26 40 subgroup 2 OXA 51 gt 1 gt 1 Ac baumannii Chromosomal OXA 64 66 68 71 78 82 OXA 51 like subgroup 3 OXA 58 Ac baumannii Plasmid subgroup 4 only 58 Bold involved in outbreaks O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 140 of 170 Procedure for detection of class A and D carbapenemases 14 Strains with reduced susceptibility to imipenem meropenem MIC gt 1 ug ml I R to ertapenem disk and highly resistant to ceftazidime KPC and GES enzymes should be suspected of possessing carbapenemases particularly KPC It is important to recognize small resistant colonies growing inside the ertapenem disk zone 13 24 Besides Class D enzymes such as oxacillinases OXA 25 OXA 34 OXA 58 OXA 64 66 are isolated mainly from Acinetobacter baumannii outbreaks Strains producing Class A enzymes are isolated more frequently particularly KPC enzymes 1 Synergy test between Imipenem and Amox Clav Neo Sensitabs distance 10 mm from edge to edge The addition of 5
35. 0 25 20 5 Ofloxacin 10 ug OFLOX gt 24 lt 22 lt 1 a Penicillin Low 5 ug PEN L gt 32 31 23 lt 22 lt 0 06 gt 2 Rifampicin 30 ug RIFAM gt 28 27 24 23 24 Spiramycin 200ug SPIRA 226 25 21 lt 20 lt 2 gt 8 Tetracyclines 80 ug TET80 gt 27 26 23 lt 22 lt 2 gt 8 Tetracyclines 10 ug TET10 gt 20 19 17 lt 16 49 gt 8 Trimethoprim Sulfa 5 24240 ug TR SU 232 31 27 26 lt 0 5 9 5 gt 2 38 a Beta lactamase positive strains should be reported resistant to penicillin ampicillin amoxycillin ticarcillin and piperacillin These strains may be detected using Amoxycillin Clavulanate Neo Sensitabs compared to Amoxycillin Neo Sensitabs or Ampicillin 33 ug Neo Sensitabs Synergism 255 mm larger zone with Amoxycillin Clavulanate will be seen in the presence of a beta lactamase BRO 1 BRO 2 The CLSI has not yet defined MIC breakpoints for Moraxella catarrhalis Interpretative criteria are based upon population distributions of MIC s of various agents published data on pharmacokinetics and clinical experience in combination with knowledge from different national guidelines References 1 Westley Catlin B Branhamella catarrhalis an organism gaining respect as a pathogen Clinical Microbiology Reviews Vol 3 No 4 293 320 1990 2 Chaibi E B et al Beta lactamases de Branhamella catarrhalis et leurs implications phenotypiques Res Microbiol 146 761 TT1 1995 3 Verduin C M et al Moraxella catarrha
36. 10 ug CIP10 gt 26 lt 25 lt 0 5 gt 1 Doxycycline 80 ug DOXYC 220 19 17 lt 16 lt 4 gt 16 b Erythromycin 78 ug ERYTR lt 18 gt 16 b d Nalidixan 130ug NALID lt 25 gt 8 Sulfonamides 240ug gt 23 22 20 lt 19 lt 100 2350 c Tetracyclines 80 ug TET80 223 22 20 lt 19 lt 4 gt 16 c Tetracyclines 10 ug TET10 gt 16 15 14 lt 13 lt 4 gt 16 Trimethoprim Sulfa 5 2 240 ug TR SU gt 28 27 24 lt 23 lt 2 38 gt 8 152 a Results for ampicillin are used to predict susceptibility to amoxycillin b Not yet established by the CLSI c Results for tetracycline can be used to predict susceptibility to doxycycline d Strains resistant to nalidixan show a decreased susceptibility to quinolones Ciprofloxacin MIC gt 0 12 mg l References 1 CLSI Performance Standards for Antimicrobial Susceptibility Testing 17th Inf Suppl M100 S17 2007 2 Mukhopadhyay A K et al Emergence af Fluoroquinolone resistance in strains of V cholerae isolated from hospitalized patients with acute diarrhoea in Calcutta India Antim Ag Chemother 42 206 207 1998 3 Garg et al Emergence of fluoroquinolone resistant strains of Vibrio cholerae 01 Biotype El Tor among hospitalized patients with cholera in Calcutta India Antim Ag Chemother 45 1605 1606 2001 4 Lai King Ng et al Can E test be used to determine Vibrio cholerae susceptibility to erythromycin Antimicr Ag Chemother 47 1479
37. 2005 Chu Y W et al EDTA susceptibility leading to false detection of metallo beta lactamase in P aeruginosa by E test and an Imipenem EDTA disk method Intl J Antimicr Agents 26 340 341 2005 Lincopan N et al First isolation of metallo B lactamase producing multiresistant pneumoniae from a patient in Brazil J Clin Microbiol 43 516 9 2005 Pitout J D D et al Detection of P aeruginosa producing metallo B lactamases in a large centralized laboratory J Clin Microbiol 43 3129 35 2005 Marqu S et al Regional occurrence of plasmid mediated carbapenem hydrolyzing oxacillinase OXA 58 in Acinetobacter spp in Europe J Clin Microbiol 43 4885 8 2005 Choi Y S et al Evaluation of Imipenem Disk Hodge Test and double disk synergy tests to detect SIM 1 type metallo B lactamase producing Acinetobacter isolates Presentation D 1705 45th ICAAC 2005 Lee Kyungwon et al Novel acquired metallo B lactamase gene blaSIM 1 in class I integron from Ac baumannii clinical isolates from Korea Antimicr Ag Chemother 49 4485 91 2005 Peleg A Y Dissemination of the metallo lactamase gene blal MP 4 among gram negative pathogens in a clinical setting in Australia CID 41 1549 56 2005 Pournaras et al VIM 12 a novel plasmid mediated metallo B lactamase from pneumoniae that resembles a VIM 1 VIM 2 hybrid Antimicr Ag Chemother 49 5153 6 2005 Berges L et al Prospective evaluation of Imipenem EDTA combined disk
38. 26 32 4 4 References 1 NCCLS Methods for Antimicrobial Susceptibility Testing of Anaerobic Bacteria 7th Ed Approved Standard M11 A7 2007 2 Edwards R et al Mechanism responsible for reduced susceptilility to imipenem in Bacteroides fragilis J Antimicrob Chemother 38 941 951 1996 3 M G et al False resistance to metronidazole by E test among anaerobic bacteria Diagn Microbiol Infect Dis 24 117 119 1996 4 Fang H et al Detection of imipenem resistant and metronidazole resistant Bact fragilis group strains in fecal samples Clin Microbiol Infect 5 753 8 1999 5 SFM Comite de l Antibiogramme de la SFM Communiqu 2006 6 Aldrige K E Detection of carbapenemase production in the B fragilis group using the MBL E test strip correlation with broth microdilution testing Clin Microbio Inf 11 Suppl 2 625 2005 7 Kuijper EJ et al Clostridium difficile ribotype 027 toxinotype The Netherlands Emerg Infect Dis 12 827 830 2006 8 Bogaerts P et al Phenotypic and genotypic analysis of carbapenem resistant B fragilis strains isolated in Belgium Presentation d 246 ICAAC 2007 Chicago USA O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 21 Page 161 of 170 21 Susceptibility Testing of Yeasts The CLSI M27 A2 reference broth microdilution method requires 48 hours incubation and can be cumbersome to perform for most laboratories Therefore there is
39. 26 lt 26 lt 0 25 gt 1 Aminoglycosides b Amikacin 40 ug AMIKA Enterobacteriaceae Acinetobacter spp 224 23 20 20 8 gt 16 Pseudomonas spp 226 25 22 22 8 gt 16 S aureus gt 24 23 20 lt 20 8 gt 16 neg staph gt 30 29 26 lt 26 lt 8 gt 16 b Gentamicin 40 ug GEN40 Enterobacteriaceae Acinetobacter spp gt 26 25 22 lt 22 lt 2 gt 4 Pseudomonas spp gt 28 27 24 lt 24 lt 4 gt 4 k S aureus gt 30 29 26 lt 26 lt 1 gt 1 k Coag neg staph gt 34 lt 34 lt 1 gt 1 b Netilmicin 40 ug NETIL Enterobacteriaceae Acinetobacter spp 226 25 22 22 lt 2 gt 4 Pseudomonas spp gt 28 27 24 lt 24 lt 4 gt 4 S aureus gt 30 29 26 lt 26 lt 1 gt 1 Coag neg staph gt 34 lt 34 lt 1 gt 1 b Tobramycin 40 ug TOBRA Enterobacteriaceae Acinetobacter spp gt 26 25 22 lt 22 lt 2 gt 4 Pseudomonas spp gt 28 27 24 lt 24 lt 4 gt 4 S aureus gt 30 29 26 lt 26 lt 1 gt 1 Coag neg staph gt 34 34 lt 1 gt 1 Others Erythromycin staph 78 ug ERYTR 230 29 26 lt 26 lt 0 5 gt 0 5 Azitromycin staph 30 ug AZITR gt 24 23 20 20 lt 0 5 gt 0 5 Claritromycin staph 30 ug CLARI gt 24 23 20 lt 20 lt 0 5 gt 0 5 Clindamycin staph 25 ug CLIND gt 30 29 26 lt 26 lt 0 5 gt 2 Telithromycin staph 15 ug TELI15 Use Erythromycin Fucidin 100ug FUCID 232 32 lt 0 5 gt 0 5 Doxycycline 80 ug DOXYC 228 27 23 23 lt 2 gt 2 Tetracyclines 80
40. 26 23 22 lt 4 gt 4 Enterococcus spp gt 26 25 23 lt 22 lt 4 gt 4 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 59 of 170 Zone diameter Break points imm MIC pg ml NEO SENSITABS STYRKE KODE S l R S R Vancomycin 5ug VAN 5 216 lt 16 lt 2 gt 4 h2 2 18 t prediffusion BHI blod r Staphylococcus spp 21 VISA GISA t Enterococcus spp MH 16 VRE Teicoplanin 30 ug TPN30 h2 24 18 t pr diffusion BHI blod VISA GISA r Staphylococcus spp lt 20 VISA GISA t Enterococcus spp MH lt 16 VRE Daptomycin 30 ug DAPCa h2 2 18 t prediffusion Staphylococcus spp gt 22 lt 1 Enterococcus spp gt 12 lt 4 Colistin 10 ug CO 10 h2 2 18 t pr diffusion gt 15 lt 15 lt 2 gt 2 Ciprofloxacin 10 ug CIP10 d Enterobacteriaceae 234 33 23 lt 22 lt 0 06 gt 1 Pseudomonas spp Acinetobacter spp gt 28 27 23 lt 22 lt 1 gt 1 Staphylococcus spp gt 24 23 18 lt 17 lt 1 gt 4 Norfloxacin 10 ug NORFX Nedsat f lsomhed Staphylococcus spp lt 20 for kinoloner Moxifloxacin 5ug MOXIF Staphylococcus spp gt 29 28 26 lt 25 lt 0 25 gt 0 5 Nitrofurantoin U 260ug NITRO gt 26 lt 26 lt 32 gt 32 Trimethoprim 5 2 ug TRIME 222 21 19 lt 18 lt 2 gt 4 Trimethoprim Sulfa 5 2 240 TR SU gt 28 27 24 lt 23 lt 16 gt 32 Sulphonamides U 240ug SULFA 222 lt 22 lt 512 gt 512 Nalidixan U 130ug NALID 228 27 24 lt 2
41. 27 23 lt 23 lt 2 gt 8 Doxycycline 80 ug DOXYC 226 25 22 22 lt 4 gt 16 Ertapenem 10 ug ERTAP gt 23 22 19 lt 19 lt 4 gt 16 Fucidin Clostridia 100ug FUCID gt 30 lt 29 lt 2 gt c Imipenem 15 ug IMIPM gt 24 23 20 lt 20 lt 4 gt 16 Imipenem EDTA 15 750 ug IM ED Detection of metallo beta lactamases Linezolid 30 ug LINEZ 221 lt 4 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 20 Page 159 of 170 Zone diameter Break points in mm MIC pg ml NEO SENSITABS S b R S R Meropenem 10 ug MEROP gt 22 21 18 lt 18 lt 4 gt 16 d Metronidazole 16 ug MTR16 gt 22 21 18 18 8 232 Moxifloxacin 5ug MOXIF 223 22 19 19 lt 2 gt 8 a Penicillin Low 5 ug PEN L gt 24 23 18 lt 18 lt 0 5 gt 2 Piperacillin 100ug PIPRA gt 24 23 20 lt 20 lt 32 gt 128 Piperacillin Tazobactam 100 10ug PI TZ gt 24 23 20 lt 20 lt 32 4 gt 128 4 Tetracyclines 80 ug TET80 gt 28 27 23 lt 23 lt 4 gt 16 Ticarcillin 75 ug TICAR gt 24 23 20 lt 20 lt 32 gt 128 Ticarcillin Clavulanate 75 15 ug TI CL gt 24 23 20 lt 20 lt 32 2 gt 128 2 Tigecycline 15 ug TIGEC gt 18 17 15 lt 15 lt 4 gt 16 Vancomycin 5 ug VAN 5 218 lt 4 Break points have not been established by the CLSI a Members of the Bact fragilis group are resistant Other gram negative anaerobes should be screened for beta lactamase with nitrocefin and if positive reported as penicillin a
42. 28 27 23 lt 23 lt 2 gt 4 Quinupristin Dalfopristin 15 ug SYNI5 pneumoniae gt 20 19 17 17 lt 1 gt 2 Vancomycin 5 ug VAN 5 S pneumoniae strep 17 16 16 lt 4 gt 4 C jeikeium urealyticum gt 17 16 16 lt 4 gt 4 Teicoplanin 60 ug TEICO S pneumoniae strep 18 17 16 16 lt 4 gt 4 C jeikeium urealyticum gt 18 17 16 16 lt 4 gt 4 Fucidin 100ug FUCID C jeikeium urealyticum 228 271 24 24 lt 0 5 gt 0 5 Norfloxacin 10 ug NORFX Decreased susceptibility Pneumococcus spp lt 16 to quinolones Nalidixan 130 ug NALID Decreased susceptibility Haemophilus spp lt 30 to quinolones Ciprofloxacin 10 ug CIP 10 i Haemophilus spp gt 30 29 26 lt 26 lt 0 5 gt 0 5 9 S pneumoniae gt 36 35 23 lt 23 lt 0 12 22 Moraxella catarrhalis 230 29 26 lt 26 lt 0 5 gt 0 5 Ofloxacin 10 ug OFLOX Haemophilus spp gt 30 29 26 lt 26 lt 0 5 gt 0 5 9 S pneumoniae 236 35 20 20 lt 0 25 gt 2 Moraxella catarrhalis gt 30 29 26 lt 26 lt 0 5 gt 0 5 Levofloxacin 5 ug LEVOF Haemophilus spp gt 30 29 26 lt 26 lt 0 5 gt 0 5 S pneumoniae 222 21 19 19 lt 2 gt 2 Moraxella catarrhalis gt 30 29 26 lt 26 lt 0 25 gt 0 5 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 43 of 170 Zone diameter Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S l R S R Moxifloxacin 5ug MOXIF Haemophilus spp 230 29 26 lt 26 lt 0 5 gt 0 5
43. 30 ug CFEPM gt 26 25 19 lt 18 lt 4 gt 32 b Cefotetan 30 ug CFTTN gt 26 25 19 lt 18 lt 4 gt 32 b Aztreonam 30 ug AZTRM gt 28 27 19 lt 18 lt 2 gt 32 1 b Meropenem 10 ug MEROP gt 26 25 19 18 2 28 Ps aeruginosa 230 29 24 lt 23 lt 2 gt 8 1 b Imipenem 15 ug IMIPM gt 26 25 21 lt 20 lt 2 gt 8 Ps aeruginosa gt 30 29 24 lt 23 lt 2 gt 8 j Tetracyclines 80 ug TET80 gt 28 27 21 lt 20 lt 1 gt 8 Doxycycline 80 ug DOXYC gt 28 27 21 lt 20 lt 1 gt 8 Chloramphenicol 60 ug CLR60 gt 26 lt 25 lt gt 16 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 74 of 170 Zone diameter Break points inmm MIC ug ml NEO SENSITABS POTENCY CODE S R S R Gentamicin 40 ug GEN40 gt 26 25 21 lt 20 lt 1 gt 8 e Gentamicin 250ug GN250 gt 15 I4 HLR gt 500 Tobramycin 40 ug TOBRA 226 25 21 20 lt 1 gt 8 Netilmicin 40 ug NETIL gt 26 25 21 lt 20 lt 1 gt 8 Amikacin 40 ug AMIKA gt 24 23 19 lt 18 lt 4 gt 32 e Streptomycin 500ug 8 500 gt 15 I4 HLR 1000 Kanamycin 100ug KANAM m Staphylococcus spp gt 23 22 19 lt 18 lt 16 gt 64 f k Vancomycin 5 ug VAN 5 gt 16 lt 15 lt 4 16 k Teicoplanin 30 ug TPN30 gt 16 lt 15 lt 4 gt 16 Fucidin 100 ug FUCID gt 30 lt 29 lt 2 gt 4 Erythromycin 78 ug ERYTR gt 28 27 24 lt 23 lt 1 gt 8 Azithromycin 30 ug AZITR gt 20 19 15 lt 14 lt 2 28 Clindamycin 25 ug
44. 32 lt 0 03 gt 0 12 Meropenem 10ug MEROP gt 30 lt 29 lt 0 25 c Minocycline 80 ug MINOC gt 28 2 lt 27 lt 2 x b Nalidixan 130ug NALID 28 screen quino Ofloxacin 10 ug OFLOX 236 35 32 lt 31 lt 0 03 gt 0 12 a Oxacillin 1 ug OXA 1 gt 10 zone lt 0 06 Oxacillin 5ug OXA 5 216 lt 15 lt 0 06 pen MIC a screen pen Penicillin Low 5 ug PEN L 226 25 23 lt 22 lt 0 06 20 5 c Rifampicin 30 ug RIFAM gt 30 lt 29 lt 0 5 gt 2 c Sulphonamides 240ug SULFA gt 28 lt 27 lt 16 gt 32 c Trimethoprim Sulfa 5 2 240 ug TR SU gt 36 35 32 lt 31 lt 0 12 2 3 2 0 5 9 5 Breakpoints have not been established by CLSI NCCLS a Oxacillin 1 ug and Oxacillin 5 ug Neo Sensitabs are useful to screen for beta lactamase negative meningococci with decreased susceptibility to penicillin chromosomal resistance If the zone is 10 mm Oxacillin 1 ug or lt 15 mm Oxacillin 5 ug perform an MIC test for penicillin b Nalidixan is useful to screen for strains with reduced susceptibility to quinolones If the zone is 28 mm control MIC s of OFLOX etc Strains showing decreased susceptibility to ciprofloxacin have been reported in Spain 7 c Used for prophylaxis only not treatment O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 108 of 170 References 1 Campos J et al Detection of relatively pe
45. 32 32 64 64 128 gt 128 i Norfloxacin U 10 ug NORKX gt 26 1825 13 17 lt 12 lt 0 5 1 4 6 16 gt 16 S pneumoniae lt 16 Nedsat f lsomhed for Kinoloner Novobiocin 5 ug NOVOS gt 16 S75 662 5 gt 2 c Ofloxacin 10 ug OFLOX gt 30 21 29 lt 20 lt 0 25 0 5 2 gt 2 Oxacillin lug OXA 1 9 S aureus gt 16 lt 15 lt 1 21 9 Koag neg staph gt 20 lt 19 lt 0 25 gt 0 5 f pneumoniae gt 24 lt 23 lt 0 06 pen gt 20 12 f N gonorrhoeae screening gt 12 lt 9 lt 0 06 pen d Penicillin Low 5ug PEN L gt 28 2327 1522 lt 14 x025 0 5 1 2 22 Staphylococcus spp 228 lt 27 lt 0 25 BL pos a b Piperacillin 100ug gt 28 21 27 10 20 lt 9 lt 16 32 128 256 gt 256 b Piperacillin 100 10ug PI TZ 228 21 27 10 20 lt 9 lt 16 32 128 256 gt 256 Tazobactam Polymyxins 150ug COI50 gt 22 19 21 13 18 lt 12 lt 4 8 16 32 128 gt 128 Quinupristin 15ug SYNI5 220 19 17 lt 16 lt 1 2 4 gt 4 Dalfopristin Rifampicin 30 ug RIFAM gt 28 23 27 15 22 lt 14 lt 0 5 1 2 4 16 gt 16 Spiramycin 200 ug SPIRA 230 27 29 lt 26 lt 2 4 8 28 j Streptomycin 100 ug ST100 gt 28 23 27 15 22 lt 14 lt 8 12 32 64 128 gt 128 d Enterococcus spp 20 gt 1000 HLR Sulphonamides U 240 ug SULFA 228 23 27 15 22 lt 14 x64 128 256 512 gt 512 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 55 of 1
46. 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 Page 167 of 170 Rementeria A et al Utility of Neo S tablets of Fluconazole and Voriconazole for in vitro susceptibility testing of Candida spp with the NCCLS M44 P method of diffusion on agar VII Congress Mycology Salamanca July 2004 Spanish Pfaller M et al Evaluation of the E test method using MH GMB agar for determining Amphothericin B MIC s for 4936 clinical isolates of Candida spp J Clin Microbiol 42 4977 9 2004 Espinel Ingroff A et al Correlation between Neo S tablets on 3 media NCCLS reference disk diffusion and broth microdilution methods for testing Candida spp and Cryptococcus neoformans with fluconazole and voriconazole Abstract ECCMID 2005 Carrillo Mufioz A J et al In vitro antifungal activity of Sertaconazol compared with 9 other drugs against 250 clinical isolates of dermatophites and S brevicaulis Chemotherapy 50 308 313 2004 Cuenca Estrella M et al Correlation between the procedure for antifungal susceptibility testing for Candida spp of the European Committee on Antibiotic Susceptibility Testing EUCAST and four commercial techniques Clin Microbiol Infect 11 6 486 92 2005 Jun Wanger A Comparison of E test and Yeast one to broth microdilution for MIC testing of 7 antifungal agents using challenge Candida strains Abstract C 236 ASM meeting June 2005 Espinel Ingroff A et al Interna
47. 4 With Trimethoprim Sulphonamides and Trimetroprim Sulfa organisms may grow for several generations before being inhibited resulting in edges of zones of inhibition containing a large number of small colonies In this case zones of inhibition are measured up to colonies of normal size disregard slight growth and measure the more obvious margin If the test organism is Staphylococcus or Enterococcus spp 24 hours of incubation is required and transmitted light plate held up to light is used to examine the oxacillin linezolid and vancomycin zones for light growth minute colonies of methicillin linezolid or vancomycin resistant colonies respectively within apparent zones of inhibition Any discernible growth within the zone of inhibition is indicative of methicillin linezolid or vancomycin resistance Other agents can be read after 16 18 hours incubation 1 With staphylococci and linezolid read with transmitted light and measure only the clear zone smallest zone Serratia marcescens may show a large zone of inhibition around Polymyxins 150 ug colistin and within this zone a band of colonies close to the tablet disk cocarde These colonies are polymyxin resistant and consequently S marcescens is to be considered resistant to polymyxins 2 Brumfitt et al 3 explain the phenomenon of resistance to Amoxycillin Clavulanate and sensitivity to ampicillin found with a few strains by the fact that ampicillin in general is more active
48. 5 gt 1 Nalidixan identification 130 ug NALID gt 18 lt 16 ID gt 128 quinolones 5 gt lt 27 p decreased suscept to quinolones Neomycin 120 ug NEOMY gt 30 29 24 lt 23 lt 8 gt 32 Nitrofurantoin 260 ug NITRO gt 30 29 27 lt 26 lt 8 gt 16 Norfloxacin 10 ug NORFX gt 26 25 23 lt 22 lt 0 5 gt 1 Ofloxacin 10 ug OFLOX gt 26 25 23 22 lt 0 5 gt 1 Tetracyclines 80 ug TET80 gt 28 27 24 lt 23 lt 4 gt 16 Trimethoprim Sulfa 5 2 240 ug TR SU gt 28 27 24 lt 23 lt 2 38 gt 8 152 ID ID purposes Please note Strains resistant to Nalidixan show a decreased sensitivity to the quinolones CIPRO NORFX OFLOX etc O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 120 of 170 References 1 Vanhoof et al Disk sensitivity testing for Campylobacter jejuni Eur J Clin Microbiol 3 160 162 1984 2 Piddock L J V Quinolone resistance and Campylobacter Clin Microbiol Infect 5 239 243 1999 3 Engberg J et al Comparison of 2 agar dilution methods and 3 agar diffusion methods including E test for antibiotic susceptibility testing of thermophilic Campylobacter species Clin Microbiol Infect 5 580 584 1999 4 Saenz Y et al Antibiotic resistance in Campylobacter strains isolated from animals foods and humans in Spain in 1997 1998 Antimicr Ag Chemother 44 267 271 2000 5 Gaudreau Ch et al Antimicrobial resistance of Camp
49. 5 ug ml haemin 5 defibrinated sheep blood and I ug ml vitamin K1 haemin and vitamin K1 may be added before sterilisation Direct suspension of colonies in broth to achieve a turbidity equivalent to a 1 0 McFarland standard 3x10 CFU ml Streak the surface of the agar with a cotton swab e Allow the inoculated plate to remain at room temperature 5 10 min until the surface of the agar looks dry For some fastidious isolates that do not grow on control plates pre reduction of plates in an anaerobic environment may be necessary Apply Neo Sensitabs tablets e nvert the inoculated plate and incubate at 35 C in an anaerobic jar or alternative anaerobic environment for 24 48 hours Zone diameter interpretative standards are correlated to the MIC break points recommended by the CLSI for anaerobic bacteria 1 Anaerobes Supplemented Brucella blood agar Inoculum McFarland 1 0 Incubation in anaerobic environment MIC break points according to CLSI M11 A7 Zone diameter Break points in mm MIC pg ml NEO SENSITABS S I b R S R a Ampicillin 33 ug AMP33 gt 32 31 26 lt 26 lt 0 5 22 Ampicillin Sulbactam 30 30 ug AM SU gt 26 25 22 lt 22 lt 8 4 gt 32 16 Amoxycillin Clav 30 15 ug AM CL gt 28 27 23 lt 23 lt 4 2 2 16 8 Cefotetan 30 ug CFTTN gt 20 19 16 lt 16 lt 16 gt 64 Cefoxitin 60 ug CFOXT gt 24 23 18 lt 18 lt 16 gt 64 Chloramphenicol 60 ug CLR60 226 25 22 22 8 232 Clindamycin 25 ug CLIND gt 28
50. 80 2003 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 122 of 170 15 9 Susceptibility Testing of Helicobacter pylori The recommended agar medium is Mueller Hinton Agar 10 blood Use colonies taken directly from a blood agar culture incubated for 48 hours at 35 37 C under microaerophilic conditions Control microscopically with gram staining that there are only few cocoid cells in the culture Prepare a suspension in bouillon or 0 9 saline to an opacity equivalent to McFarland 3 or 4 Not more than 9 Neo Sensitabs should be placed on a 150 mm plate or 4 Neo Sensitabs into a 90 100 mm plate The plates are incubated at 35 C in microaerophilic atmosphere for 72 hours Helicobacter pylori Mueller Hinton 10 Blood Inoculum 3 4 McFarland Incubation microaerophilic Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S R Amoxycillin 30 ug AMOXY 232 31 27 lt 26 lt 0 5 gt 2 Ampicillin 33 ug AMP33 232 31 27 lt 26 lt 0 5 gt 2 Ampicillin 2 5 ug AMP L 223 22 20 lt 19 lt 0 5 gt 2 Azithromycin 30 ug AZITR gt 30 29 24 lt 23 lt 0 25 gt 1 Ciprofloxacin 0 5 ug CIP L 220 19 17 lt 16 lt 0 5 gt Clarithromycin 30 ug CLARI gt 30 29 24 lt 23 lt 0 25 gt 1 Doxycycline 80 ug DOXYC gt 32 31 29 28 2 28 Erythromycin 78 ug ERYTR 230 20 24 lt 23 lt 1 gt 4 Furazolidone 50 ug FURAZ gt 32 31 29 28 lt 1 gt 4 Levofloxacin 5 ug LEVOF gt 26 lt
51. Antibiogramme de la Societ Francaise de Microbiologie Jan 2006 Zone diameter Concentrations inmm critiques NEO SENSITABS POTENCY CODE S R S R Amikacin 40 ug AMIKA gt 20 19 17 lt 16 lt 8 gt 16 Amoxycillin 30 ug AMOXY 223 22 20 lt 19 lt 4 gt 16 Amoxycillin Clav 30 15 ug AM CL gt 23 22 20 lt 19 lt 4 2 gt 16 2 a Ampicillin 33 ug AMP33 Enterobacteriaceae 223 22 20 lt 19 lt 4 gt 16 Ampicillin Sulbactam 30 30 ug AM SU gt 23 22 20 lt 19 lt 4 8 gt 16 8 Azithromycin 30 ug AZITR 223 27 17 lt 16 lt 0 5 24 0 g Aztreonam 30 ug AZTRM 225 24 21 lt 20 lt 4 gt 32 Bacitracin 40U BACIT gt 20 lt 19 lt 2 gt 2 Cefaclor 30 ug CCLOR gt 25 24 21 lt 2 lt 2 gt 8 Cefadroxil 30 ug CFDRO 220 19 17 lt 16 lt 8 gt 32 0 g Cefepime 30 ug CFEPM gt 23 22 20 lt 19 lt 4 232 Cefepime Clavulanate 30 10ug CP CL Detection of ESBL 9 Cefixime 30 ug CFFIX gt 28 27 24 lt 23 lt 1 gt 2 9 Cefotaxime 30 ug CFTAX gt 23 22 20 lt 19 lt 4 gt 32 Cefotetan 30 ug CFTTN gt 23 22 20 lt 19 lt 4 gt 32 Cefoxitin 60 ug CFOXT gt 23 22 20 lt 19 lt 8 gt 32 S aureus 226 lt 25 Oxa S Mec A pos Coag neg staph 230 lt 29 S pos 9 Cefpirome 30 ug CFPIR 223 22 20 lt 19 lt 4 gt 32 g m Cefpodoxime 30 ug CFPOX 224 lt 23 lt 4 gt 4 Scr ESBL Cefsulodin 30 ug CFSUL Pseudomonas spp 223 22 20 lt 19 lt 8 gt 32 9 Ceftazidime 30 ug CEZDI 223 22 20 lt 19 lt 4
52. Breakpoints in Europe a future perspective 42nd ICAAC Presentation 1022 2002 2 et al European harmonization of MIC breakpoints for antimicrobial susceptibility testing of bacteria J Antimicr Chemother 52 145 148 2003 3 http www srga org eucastwt MICTA B index html O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter I Page 6 of 170 1 Susceptibility Testing in General Susceptibility testing is of great importance to the hospital physician and the practitioner in the treatment of infectious diseases Knowledge of susceptibility to different antibiotics of an infectious agent facilitates the choice of the most effective antibacterial agents In most clinical laboratories an agar diffusion disk tablet method is used routinely for testing the susceptibility of common rapidly growing bacterial pathogens It is generally accepted that reliable results can be obtained by the diffusion test when a standardized methodology is used and the diameter of the inhibition zones has been correlated with minimal inhibitory concentrations MIC The interpretation of zone diameters by the diffusion method should be based on regression lines A regression line for a certain antibiotic contained in a disk tablet is a graphic expression of the correlation existing between MIC s and the diameters of inhibition zones obtained with a number of bacterial strains These lines represent a control of the
53. GENTAMICIN 40 ug GEN40 40 ug GENTAMICIN 250 ug HLR GN250 250 ug NETILMICIN NETIL 40 ug TOBRAMYCIN TOBRA 40 ug SPECTINOMYCIN SPECT 200 ug APRAMYCIN Vet APRAM 40 ug ISEPAMICIN 30 ug ISP30 30 ug G Tetracyclines TETRACYCLINES 80 ug TET80 80 ug Oxytetracycline TETRACYCLINES 10 ug TET10 10 ug DOXYCYCLINE DOXYC 80 ug MINOCYCLINE MINOC 80 ug TIGECYCLINE investigational drug TIG15 15 ug H Chloramphenicol and derivatives CHLORAMPHENICOL 60 ug CLR60 60 ug CHLORAMPHENICOL 10 ug CLR10 10 ug FLORFENICOL Vet FFC30 30 ug I Macrolides Lincosamides Streptogramines Ketolides and Oxazolidinones ERYTHROMYCIN ERYTR 78 ug AZITHROMYCIN AZITR 30 ug CLARITHROMYCIN CLARI 30 ug LINCOMYCIN LINCO 19 ug O Copyright Rosco Diagnostica A S NEO SENSITABS Identification Diffusible code amount of Neo Sensitabs Neo Sensitabs Antimicrobial CLINDAMYCIN 25 ug CLIND 25 ug SPIRAMYCIN SPIRA 200 ug PRISTINAMYCIN PRIST 30 ug VIRGINIAMYCIN VIRGI 30 ug QUINUPRISTIN DALFOPRISTIN SYNI5 15 ug TELITHROMYCIN TEL15 15 ug LINEZOLID LINEZ 30 ug LINCO SPECTIN Vet LI SP 15 200 ug TYLOSIN Vet TYLOS 150 ug TILMICOSIN Vet TILMI 80 ug PIRLIMYCIN Vet PIRLI 10 ug J 1 Glycopeptides VANCOMYCIN 5 ug VAN 5 5ug J 2 Lipoglycopeptides TEICOPLANIN TPN30 30 ug DALBAV ANCIN investigational drug DALBA 30ug TELAVANCIN investigational drug TELAV 30ug J 3 Cyclic lipopeptides Gram positive spectrum DAPTOMYCIN Ca DAPCa 30 ug B
54. Glycopeptide dependent strains Glycopeptide dependent strains require vancomycin teicoplanin for growth 21 They are currently resistant to vancomycin or to both vancomycin and teicoplanin and appear as colonies growing up to the edge of the vancomycin tablet disk When testing Van D isolates comparing semiconfluent growth and McF 0 5 inoculum respectively a large difference in zone size will be obtained 10 mm or more due to the high inocolum effect 11 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 14 Page 94 of 170 Characteristics 5 7 21 A Acquired resistance High Level Variable level Phenotype Van A Van B Van D 11 Van E 13 Van G 15 MIC ug ml Vancomycin gt 64 4 16 1024 16 64 128 6 16 32 16 Teicoplanin gt 16 0 5 1 0 25 4 64 0 5 0 5 Transfer by conjugation 0 0 Resistance induced by Vancomycin 0 Teicoplanin 0 0 0 0 Location Plasmid Plasmid Chromosome Chromosome Chromosome chromosome chromosome Microorganisms E faecium E faecium E faecium E faecalis E faecalis detected E faecalis E faecalis E faecalis E durans E gallinarum high E gallinarum 14 inoculum MIC Vanco 32 128 pg ml effect E casseliflavus E gallinarum E mundtii E raffinosus E avium E hirae E raffinosus Characteristics 5 7 B Intrinsic resistance Low level High level Phenotype Van MIC ug ml Vancomycin 8 16 4 32 gt 1000 Teicoplanin 0 5 0 5 1 2250 T
55. J Antimicr Chemother 49 601 609 2002 9 Tankovic J et al Single and double mutations in gyrA but not in gyrB are associated with low and high level fluoroquinolone resistance in H pylori Antimicr Ag Chemother 47 3942 44 2003 10 Jung Mogg Kim et al Distribution of antibiotic MICs for pylori strains over a 16 year period in patients from Seoul South Korea Antimicr Ag Chemother 48 4843 7 2004 11 Raymond J et al Heterogeneous susceptibility to metronidazole and clarithromycin of H pylori isolates from a single biopsy in adults is confirmed in children Intl J Antimicrobial Agents 26 272 8 2005 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 123 of 170 15 10 Susceptibility Testing of S maltophilia B cepacia and Acinetobacter spp The medium recommended is plain Mueller Hinton Agar Use colonies taken directly from an overnight culture and make a suspension in 0 9 saline equivalent to the 0 5 McFarland standard Use the suspension for plate inoculation within 15 min The plates are incubated in air at 30 C for 20 24 hours Acinetobacter spp at 35 C The CLSI Working Group recommends incubation at 35 C 20 24 hours for S maltophilia and B cepacia Incubation at higher temperatures 35 37 C may result in false susceptibility results particularly with aminoglycosides and polymyxins 7 9 14 S maltophilia show medium dependent susceptibility False susceptibili
56. M et al Activities of 12 quinolones by 3 susceptibility testing methods against a collection of influenzae isolates with different levels of susceptibility to ciprofloxacin evidence of cross resistance J Antimicr Chemother 51 147 151 2002 8 Dabernat et al Characterization of beta lactam resistance in Haemophilus influenzae in France 42nd ICAAC Presentation C2 1889 2002 9 Nazir J et al Levofloxacin resistant Haemophilus influenzae in a long term care facility 42nd ICAAC Presentation C2 645 2002 10 Pitout M J et al Characterization of ESBL activity in Haemophilus parainfluenzae 42 ICAAC Presentation C2 645 2002 11 Bronwald N P et al Detection of ciprofloxacin resistance in influenzae using nalidixic acid and BSAC methodology JAC 52 1311 12 2003 12 Elliott E et al Fluoroquinolone resistance in H influenzae JAC 52 734 5 2003 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 13 14 15 16 17 18 19 Page 103 of 170 Matic V et al Contribution of B lactamase and PBP aminoacid substitutions to amoxicillin clavulanate resistance in B lactamase positive amoxicillin clavulanate resistant H influenzae JAC 52 1018 1021 2003 Perez V zquez M et al Laboratory detection of influeanzae with decreased susceptibility to nalidixic acid ciprofloxacin levofloxacin and moxifloxacin due to gyrA and ParC mutations J Clin Microbi
57. Mueller Hinton agar For the detection of Erythromycin resistance phenotypes use the double tablet induction test see chapter 13 3 page 89 Cefuroxime Neo Sensitabs is used to test both cefuroxime sodium injectable 23 19 and cefuroxime axetil oral 25 20 The different zone sizes correspond to the recommended MIC break points O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 m n 0 q 5 Page 31 of 170 Fosfomycin Indicated for use against E coli and Ent faecalis only M100 S15 2005 Morganella morganii should not be tested against Cefixime because false susceptible results may occur All staphylococci with zone diameters lt 17 mm with Teicoplanin Neo S should be tested using the 2 18 hours prediffusion test with Vancomycin 5 ug and Teicoplanin 30 ug page 86 and page 18 Strains of Klebsiella spp Salmonella and E coli that produce ESBL may be clinically resistant to therapy with penicillins cephalosporins or aztreonam despite apparent in vitro susceptibility M100 S17 2007 See chapter 16 1 for ESBL screening and confirmatory tests Enterobacteriaceae susceptible to nalidixic acid NALI are susceptible to quinolones MIC ciprofloxacin lt 0 06 ug ml Strains resistant to NALI zone 25 mm show a decreased susceptibility to quinolones MIC CIPRO gt 0 125 ug ml Therefore Nalidixic acid is a good screening for the detection of decreased
58. R S k Penicillin Low 5ug PEN L Streptococcus spp 226 25 21 lt 20 lt 0 25 gt 1 Haemophilus gt 20 19 18 lt 18 lt 1 gt 1 Moraxella catarrhalis Test beta laktamase Penase Corynebacterium spp Listeria spp gt 26 25 21 lt 20 lt 0 25 gt 1 Oxacillin lug OXA 1 c Streptococcus spp penicillin gt 17 lt 17 lt 17 lt 0 12 S pneumoniae penicillin 224 lt 23 lt 23 lt 0 06 MIC Amoxycillin Clav 30 15 ug AM CL Haemophilus spp gt 30 29 17 lt 26 lt 2 gt 4 Ampicillin 2 5 ug AMP L Streptococcus spp 224 23 19 18 lt 0 25 gt 1 Haemophilus gt 22 21 19 lt 18 lt 0 5 gt 0 5 m Moraxella catarrhalis Test beta laktamase Penase Ampicillin 33 ug AMP33 Streptococcus spp gt 36 35 29 lt 28 lt 0 25 1 Haemophilus spp test Amp 2 5 gt 34 33 31 lt 30 lt 0 5 gt 0 5 Corynebacterium spp gt 36 35 29 lt 28 lt 0 25 gt 1 Listeria spp gt 34 33 31 lt 30 lt 0 5 gt 1 Cefuroxime 60 ug CEFUR Haemophilus spp gt 34 33 29 lt 28 lt 2 gt 2 Moraxella catarrhalis gt 34 33 29 lt 28 lt 2 gt 2 Corynebacterium spp gt 40 39 31 lt 30 lt 0 12 gt 2 Cefotaxime 30 ug CFTAX Streptococcus spp 238 37 27 lt 26 lt 0 12 gt 2 f S pneumoniae Test ceftizoxime som surrogat lt 0 12 gt 1 Haemophilus spp gt 38 37 29 lt 28 lt 0 12 gt 1 Ceftizoxime 30 ug CEZOX f S pneumoniae gt 32 lt 32 lt 0 5 MIC 3 gen cepha Ceftriaxone 30 ug CETRX Streptococcus Spp
59. R S R Penicillin Low 5 ug PEN L m Staphylococcus spp gt 31 30 29 lt 28 lt 0 12 Penicillinase Enterococcus spp 226 25 11 lt 10 lt 0 25 gt 4 Oxacillin lug OXA 1 S aureus og S lugdunensis gt 16 lt 16 lt 1 gt 1 Coag neg staph 220 20 lt 0 25 gt 0 5 9 Cloxacillin 500ug X CL500 P visning af plasmid AmpC m Ampicillin 33 ug AMP33 8 Enterobacteriaceae 232 31 19 lt 18 lt 1 gt 8 Enterococcus spp gt 26 25 19 lt 18 lt gt 8 Mecillinam 33 ug MECIL Enterobacteriaceae 230 29 21 lt 20 lt 1 gt 8 1 n p Staphylococci penicillin gt 23 lt 22 P visning af penicilinresistens m Piperacillin 100ug PIPRA Enterobacteriaceae gt 28 27 24 lt 23 lt 16 gt 16 Pseudomonas spp Acinetobacter spp gt 28 27 24 lt 23 16 gt 16 i Aztreonam 30 ug AZTRM Enterobacteriaceae gt 34 33 29 lt 28 lt 0 5 gt 1 n Cefuroxime 60 ug CEFUR Enterobacteriaceae gt 26 25 23 lt 22 8 gt 8 i n Cefotaxime 30 ug CFTAX Enterobacteriaceae 234 33 29 lt 28 lt 0 5 gt 1 i n Ceftriaxone 30 ug CETRX Enterobacteriaceae 234 33 29 lt 28 lt 0 5 gt 1 n Ceftazidime 30 ug CEZDI Enterobacteriaceae 228 27 24 lt 23 lt 2 gt 4 Pseudomonas spp Acinetobacter spp gt 23 22 21 lt 20 lt 8 gt 8 f Ceftazidime Clav 30 10 ug CZ CL P visning af ESBL i Cefepime 30 ug CFEPM 234 33 29 28 lt 0 5 gt 1 e f Cefpodoxime 30 ug CFPOX 23 Screen ESBL Cefpodoxime 10ug CFD10 lt 20 Screen ESBL f Cefepime Clavulan
60. Teicoplanin 30 ug Neo Sensitabs have been removed Note the hazy edge around Vancomycin VAN 5 and no clear zone for Teicoplanin TEI30 After prediffusion inoculation and one Vancomycin 5 ug has been placed for comparison of the zone Incubated at 33 35 overnight Sandberg et al 31 ina screening study for hVISA VISA in MRSA isolates from Denmark 1 1 2003 to 31 7 2004 reported the presence of VISA hVISA in Denmark The clinical significance of hVISA strains is still controversial It is difficult to evaluate because conventional susceptibility tests including MIC determination cannot detect hVISA 28 42 Clinicians should bear in mind that an inadequate response to vancomycin in the treatment of MRSA infections could be due to heteroresistance to vancomycin An interesting property of the VISA isolates is the gradually decreasing growth rate of the bacteria which seemed to parallel the increase in the vancomycin MIC 29 Sakoulas et al 38 showed that GISA VISA strains have a decreased hemolysin expression and lack the ability to hemolyze blood on sheep blood agar plates The MIC of daptomycin is higher for GISA than for susceptible strains despite a different mechanism of action or resistance between vancomycin and daptomycin It is attributed to the fact that daptomycin being a large molecule has difficulty in passing through the thickened cell wall 41 Vancomycin resistant S aureus VRSA The first documented case
61. These strains are not clinically resistant Testing with Cefoxitin Neo Sensitabs zone S 2 25 mm will show that the strains are mecA negative cefoxitin susceptible O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 13 Page 86 of 170 b Detection of strains with decreased susceptibility to Vancomycin VISA GISA hVISA Strains of CNS showing intermediate and resistant MIC s towards vancomycin and teicoplanin have been described 5 Strains of S aureus with decreased susceptibility to vancomycin MIC 4 to 8 ug ml were reported from Japan 6 in 1997 and shortly thereafter from USA and France To date all S aureus strains appear to have developed from MRSA Automated commercial tests poorly recognize VISA hVISA and VRSA isolates which necessitates the use of espensive supplemental screening tests 39 According to Liu 23 classification of strains with reduced susceptibility to vancomycin may be as follows 1 hVISA Vancomycin heteroresistant containing a susceptible population and subpopulations of VISA Stage that precedes the development of VISA Shows also reduced susceptibility to teicoplanin Ex Mu 3 strains of Hiramatsu 2 VISA Shows MICs to vancomycin of 8 to 16 ug ml and reduced susceptibility to teicoplanin Ex Mu 50 from Hiramatsu 3 VRSA Shows MICs to vancomycin of gt 32 ug ml We should be aware of the following e VISA may not appear on the primary culture plate until day 2 of in
62. Voriconazole lug VOR 1 gt 22 21 16 lt 15 lt 0 25 gt 2 All MIC breakpoints from EUCAST are tentative 41 krusei should always be reported as resistant to fluconazole 21 1 2 Interpretation table fo Local treatment In local treatment of fungal infections a high concentration of antifungal is placed at site of the infection Consequently other MIC breakpoints and zone interpretations should be used in those cases Local Treatment MH Glucose Methylene Blue Agar or Shadomy McFarland 0 5 inoculum Susceptible gt 20 mm gt 15 mm gt 10 mm Intermediate 12 19 mm 10 14 mm Resistant lt 11 mm no zone no zone Ciclopirox Natamycin Griseofulvin Clotrimazole Nystatin Econazole Itraconazole Fluconazole Isoconazole Ketoconazole Miconazole Tioconazole Terbinafine O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 21 Page 164 of 170 21 1 3 Candida spp Quality Control MH GMB Quality control limits using Candida strains on modified Shadomy agar or MH GMB agar are included in the table below Mueller Hinton Glucose Methylene Blue Inoculum McFarland 0 5 undiluted Incubation 18 24 hours Zone diameters mm C albicans C parapsilosis C krusei NEOSENSITABS ATCC 90028 ATCC 22019 ATCC 6528 Amphotericin B 18 23 20 26 15 21 Caspofungin 15 22 13 23 16 22 Fluconazole 28 39 22 33 Itraconazole 21 30 19 26 16 22 Ketoconazole 31 42 26 35 22 29 Pos
63. Wheat P F et al Effect of temperature on antimicrobial susceptibilities of Ps maltophilia J Clin Pathol 38 1055 58 1985 8 Bonfiglio Livermore D M Effect of media composition on the susceptibility of X maltophilia to beta lactam antibiotics J Antimicrob Chemother 28 837 842 1991 9 Rahmati Bahram A et al Effect of temperature on aminoglycoside binding sites in Stenot maltophilia J Antimicrob Chemother 39 19 24 1997 10 Hsueh Po Ren et al Pandrug resistant Acinetobacter baumanii causing nosocomial infections in a University Hospital Taiwan Emerg Infect Dis 8 August 2002 11 Levin A S Severe nosocomial infections with imipenem resistant Acinetobacter baumanii treated with ampicillin sulbactam Intl J Antimicrob Ag 21 58 62 2003 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 12 13 14 15 16 17 18 Page 125 of 170 Garnacho Montero J et al Treatment of multidrug resistant A baumannii V AP with intravenous colistin a comparison with Imipenem susceptible VAP Clin Infect Dis 36 1111 1118 2003 CLSI Performance Standards for Antimicrobial Susceptibility Testing 15th Inf Suppl M100 S15 2005 King A Susceptibility testing of S maltophilia Effect of temperature and medium on results Clin Microbiol Infect 40 Suppl 3 335 2004 Pournaras S et al Heteroresistance to carbapenems in Acinetobacter baumannii J Antimicr Chemother
64. amp Infect 11 Suppl 2 229 2005 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 14 17 18 19 20 21 22 23 24 25 26 27 Page 97 of 170 del Campo R et al New aac 6 I genes in Enterococcus hivae and E durans effect on B lactam aminoglycoside synergy J Antimicr Chemoth 55 1053 5 2005 Willems R J L et al Global spread of vancomycin resistant E faecium from distinct nosocomial genetic complex Emerg Infect Dis 11 821 8 2005 Ono S et al Mechanisms of resistance to imipenem and ampicillin in Enterococcus faecalis Antimicr Ag Chemother 49 2954 8 2005 Lemming L et al Characterization of atypical Enterococcus faecalis resistant to penicillin but susceptible to ampicillin from bacteremic episodes 45th ICAAC presentation D1651 2005 Courvalin P Vancomycin resistance in gram positive cocci CID 42 Suppl 1 525 534 2006 Metzidie E et al Spread of an unusual penicillin and imipenem resistant but ampicillin susceptible phenotype among E faecalis clinical isolates JAC 57 158 9 2006 Lester C H et al In vivo transfer of the van A resistance gene from an E faecium isolate of animal origin to an E faecium isolate of human origin in the intestines of human volunteers Antimicr Ag Chemother 50 596 99 2006 Leavis H L et al High level ciprofloxacin resistance from point mutations in gyrA and parC confined to global hospital adapted clonal
65. and break points recommended by the Norwegian AFA Group January 2006 Version 1 9 Inoculum according to ICS Media Mueller Hinton Iso Sensitest Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Penicillins Ampicillin 33 ug AMP33 a Enterobacteriaceae gt 34 33 20 lt 20 lt 0 5 gt 8 b Enterococcus spp gt 26 25 18 lt 18 lt 2 gt 8 Amoxicillin 30 ug AMOXY 8 Enterobacteriaceae gt 34 33 20 lt 20 lt 0 5 gt 8 Enterococcus spp gt 28 27 20 lt 20 lt 2 gt 8 Amoxycillin Clav 30 15 ug AM CL Enterobacteriaceae gt 34 33 20 lt 20 lt 0 5 gt 8 Penicillin Low 5 ug PEN L Staphylococcus spp gt 32 31 28 lt 28 lt 0 06 gt 0 125 BL pos Enterococcus spp gt 12 zone lt 4 gt 8 Oxacillin lug OXA 1 c d S aureus gt 16 lt 16 lt 2 gt 2 d Coag neg staph gt 20 lt 20 lt 0 25 gt 0 25 Mecillinam U 33 ug MECIL Enterobacteriaceae 226 25 20 20 lt 2 gt 8 Enterobacteriaceae M H agar gt 22 21 17 lt 17 lt 2 gt 8 Piperacillin Tazobactam 100 10ug PI TZ Enterobacteriaceae gt 30 29 24 lt 24 lt 8 gt 16 Cloxacillin 500 ug CL500 Detection of plasmid mediated Amp C Cephalosporins and cephamycins Cefazolin 66 ug CFZOL Enterobacteriaceae gt 34 33 20 lt 20 lt 1 gt 8 Cephalexin U 30 ug CFLEX Enterobacteriaceae gt 30 29 18 18 lt 1 gt 8 t Cefuroxime 60 ug CEFUR Enterobacteriaceae gt 36 35 23 lt 23 lt 0 5 gt 8 Cefepime 30 ug CFEPM i Cef
66. catarrhalis BL neg BL pos Test beta lact Penicillin Low V 5ug PEN L h Haemophilus spp gt 30 29 14 lt 14 lt 0 5 gt 4 Oxacillin lug OXA 1 b S pneumoniae 24 lt 23 23 lt 0 06 pen MIC c Streptococcus spp gt 17 17 lt 0 12 pen MIC Ampicillin 2 5 ug AMP L d Haemophilus spp gt 22 21 19 lt 18 lt 0 5 gt 0 5 Streptococcus spp gt 24 23 18 lt 18 lt 0 25 gt 1 a Moraxella catarrhalis BL neg BL pos Test beta lact Ampicillin 33 ug AMP33 d Haemophilus spp gt 34 33 30 lt 30 lt 0 5 gt 0 5 Streptococcus spp gt 36 35 28 lt 28 lt 0 25 gt 1 a Moraxella catarrhalis BL neg BL pos Test beta lact Amoxycillin Clav 30 15 ug AM CL Haemophilus spp gt 30 29 26 lt 26 lt 2 gt 4 Cefuroxime 60 ug CEFUR Haemophilus spp gt 34 33 28 lt 28 lt 2 gt 2 Moraxella catarrhalis gt 34 33 28 lt 28 lt 2 22 Cefotaxime 30 ug CFTAX Haemophilus spp 238 37 28 28 lt 0 12 gt 1 S pneumoniae use ceftizoxime Ceftriaxone 30 ug CETRX Haemophilus spp gt 38 37 28 lt 28 lt 0 12 gt 1 e S pneumoniae use ceftizoxime Ceftizoxime 30 ug CEZOX e S pneumoniae gt 32 lt 32 lt 0 5 MIC 3rd gen cephalosporins Cftax Cetrx Aztreonam 30 ug AZTRM Haemophilus spp gt 40 39 34 lt 34 0 5 gt 1 Imipenem 15 ug IMIPM Haemophilus spp gt 34 33 28 lt 28 lt 1 gt 2 S pneumoniae gt 36 35 28 lt 28 lt 0 06 gt 0 5 Meropenem 10ug MEROP Haemophilus spp gt 34 33 28 lt 28 lt 1 gt 2
67. easily detected a with a rapid beta lactamase test e g Beta lactamase acido Diagnostic Tablets ROSCO code 455 21 using several colonies because producers and non producers may coexist in the sample b using Amoxycillin Clavulanate Neo Sensitabs compared to Amoxycillin alone Synergism larger zone with Amoxycillin Clavulanate will be seen in the presence of a beta lactamase Chromosomal resistance to ampicillin due to alteration of PBP s and or reduction in permeability Zerva et al 3 showed in a study of 300 H influenzae strains that testing with a 2 ug ampicillin disk gave a superior interpretative accuracy than the 10 ug ampicillin disk The latter miscategorised as susceptible or intermediate 81 396 of the BLNAR strains tested K rp noja P et al 15 found similar results Testing of Ampicillin 2 5 ug Neo Sensitabs serves better to detect the rare BLNAR strains than testing of Ampicillin 33 ug or of Amoxycillin Clavulanate by the diffusion method Beta lactamase negative Haemophilus resistant to ampicillin amoxycillin should be reported as resistant to combinations with beta lactamase inhibitors Amoxycillin Clavulanate Ampicillin Sulbactam irrespective of zone size because beta lactamase inhibitors have no effect on beta lactamase negative strains The prevalence of BLNAR strains is increasing in Europe 8 17 beta lactamase positive isolates that were resistant to amoxycillin clavulanate were detected in a US National S
68. for high level resistance HLR with enterococci Neomycin Framycetin and Paromomycin have a close chemical affinity and only one of them should be tested Neomycin Neo Sensitabs Nitroimidazoles Cross resistance between Metronidazole Ornidazole and Tinidazole against anaerobes Only one representative Metronidazole 16 ug Neo Sensitabs needs to be tested routinely Polymyxin B and Colistin Very closely related and only one needs to be tested routinely Colistin 10 ug and Polymyxins 150 ug Neo Sensitabs colistin Quinolones Broad spectrum antimicrobials including Ciprofloxacin Gatifloxacin Levofloxacin Moxifloxacin Norfloxacin Ofloxacin and Pefloxacin Small differences in spectrum may require separate testing of the individual agents Sulphonamides The Working Group of the W H O expressed the opinion that only one representative sulphonamide needs to be used in sensitivity tests against drugs of the sulphonamide group Sulphonamides Neo Sensitabs 3 Tetracyclines Most members of this group oxytetracycline tetracycline chlortetracycline are closely related and for routine work it is enough to test one tetracycline Tetracyclines 80 ug Neo Sensitabs 4 Doxycycline and Minocycline should be tested separately Glycopeptides Close chemical affinity but with different activity against some species Vancomycin and Teicoplanin Neo Sensitabs should be tested individually O Copyright Rosco Diagnostic
69. gt 32 Ceftazidime Clav 30 10ug CZ CL Detection of ESBL 9 Ceftizoxime 30 ug CEZOX 223 22 20 lt 19 lt 4 gt 32 9 Ceftriaxone 30ug CETRX 223 22 20 lt 19 lt 4 gt 32 Cefuroxime parenteral 60 ug CEFUR 223 22 20 lt 19 lt 8 232 Cefuroxime oral 60 ug CEFUR 230 29 26 lt 25 lt 1 gt 4 Cephalexin 30 ug CFLEX gt 20 19 17 lt 16 lt 8 232 Cephalothin 66 ug CLOTN 223 22 20 lt 19 lt 8 gt 32 Cephadrine 60 ug CFRAD gt 23 22 20 lt 19 lt 8 gt 32 Chloramphenicol 60 ug CLR60 225 24 21 lt 20 lt 8 gt 16 Ciprofloxacin 10 ug CIP10 gt 25 24 22 lt 21 lt 0 5 gt 1 P aeruginosa Acinetobacter spp gt 23 22 20 lt 19 lt 1 gt 2 Clarithromycin 30 ug CLARI 220 19 17 lt 16 lt 1 gt 4 Clindamycin 25 ug CLIND gt 24 lt 24 lt 2 gt 2 Cloxacillin 500ug 500 Detection of plasmid med Amp C t Colistin 10 ug CO 10 n 24 18 hours prediffusion gt 15 14 11 lt 10 lt 2 gt 2 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 66 of 170 Zone diameter Concentrations in mm critiques NEO SENSITABS POTENCY CODE S R S R Daptomycin 30ug DAPCa 2 18 hours prediffusion Staphylococcus spp 222 lt 20 lt 1 2 Enterococcus spp gt 12 lt 11 lt 4 gt 8 Doxycycline 80 ug DOXYC 223 22 20 lt 19 lt 4 gt 8 Ertapenem 10 ug ERTAP gt 23 22 21 lt 20 lt 2 gt 4 Erythromycin 78 ug ERYTR gt 25 24 21 lt 20 lt 1 gt 4 r Fosfomycin 70 40 ug FOSFO gt 16 lt 16 lt 32
70. gt 32 Fucidin 100ug gt 28 27 21 lt 20 lt 2 gt 16 Gatifloxacin 5 ug GATIF 221 20 18 lt 17 lt 1 22 n Gentamicin 40 ug GEN40 gt 24 23 21 lt 20 lt 2 gt 4 Staphylococcus spp gt 26 lt 26 lt 1 gt 1 P aeruginosa Acinetobacter spp 223 22 20 lt 19 lt 4 gt 8 h Gentamicin HNR 250ug 250 gt 14 lt 14 lt 250 gt 500 Imipenem 15 ug IMIPM gt 23 22 20 lt 19 lt 4 gt 8 Pseudomonas spp gt 28 27 24 lt 23 lt 4 gt 8 Imipenem EDTA 15 750ug IM ED Detection of metallo B lactamases Isepamicin 30 ug 220 19 17 lt 16 lt 8 gt 16 q Kanamycin 100ug KANAM gt 23 22 20 lt 19 lt 8 gt 16 h Kanamycin HNR 500 ug 500 gt 14 lt 14 lt 250 gt 500 Levofloxacin 5ug LEVOF 222 21 19 lt 18 lt 1 gt 2 Enterococcus spp gt 21 20 17 lt 16 lt 1 gt 4 Lincomycin 19 ug LINCO gt 26 25 23 lt 22 lt 2 gt 8 Linezolid 30 ug LINEZ gt 28 27 26 lt 25 lt 2 gt 4 Staphyloccus spp Enterococcus spp 224 24 lt 4 gt 4 Moxifloxacin 5ug MOXIF gt 24 23 21 lt 20 lt 0 5 gt 1 Enterococcus spp 221 20 18 17 lt 1 gt 2 Mecillinam U 33 ug MECIL gt 23 22 20 lt 19 lt 2 gt 8 Meropenem 10 ug MEROP gt 20 19 17 lt 16 lt 4 gt 8 Pseudomonas spp gt 26 25 23 lt 22 lt 4 gt 8 Minocycline 80 ug MINOC 223 22 20 lt 19 lt 4 gt 8 Mupirocin 10 ug MUPIR gt 18 lt 2 p Nalidixan U 130ug NALID 225 24 21 lt 20 lt 8 gt 16 Enterobacteriaceae lt 25 Reduced susceptibility to quinolo
71. gt 4 aeruginosa 223 22 18 17 lt 4 gt 16 q Sparfloxacin 10 ug gt 20 19 17 lt 16 lt 1 gt 4 Spectinomycin 200 ug SPECT gt 20 19 17 lt 16 lt 16 gt 64 not gonococci Spiramycin 200ug SPIRA 226 25 23 22 2 gt 8 Streptomycin 100ug 8 100 226 25 23 lt 22 lt 6 225 f Streptomycin 500 ug 8 500 lt 14 gt 1000 Enterococcus spp HLR Sulphonamides U 240 ug SULFA 223 22 20 lt 19 lt 100 gt 350 Teicoplanin 30 ug TPN30 gt 14 13 11 lt 10 lt 8 gt 32 v 2 18 h prediffusion staph lt 20 GISA TRCNS Telithromycin 15 ug TEL15 222 21 19 18 lt 1 gt 4 Temocillin 30ug TEMOC gt 18 17 15 lt 14 lt 16 gt 32 Tetracyclines 80 ug TET80 222 21 19 lt 18 lt 4 gt 16 b Ticarcillin 75 ug TICAR gt 20 19 17 lt 16 lt 16 gt 128 h Pseudomonas spp gt 16 lt 64 2 128 d Ticarcillin Clavulanate 75 15 ug TI CL gt 23 22 20 lt 19 lt 16 2 gt 128 2 h Pseudomonas spp gt 16 lt 64 2 gt 128 2 Tigecycline 15 ug TIG15 Enterobacteriaceae gt 19 18 15 lt 14 lt 2 gt 8 Staphylococcus spp gt 19 z lt 0 5 Enterococcus spp gt 19 lt 0 25 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 30 of 170 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Tobramycin 40 ug TOBRA gt 23 22 20 lt 19 lt 4 gt 8 Trimethoprim U 5 2 ug TRIME 220 19 17 lt 16 lt 4 gt 16 Trimethoprim Sulfa 5 2 240
72. interest in the validation of the agar based methods of disk diffusion for the antifungal susceptibility testing of yeasts Approximately 10 years ago Rosco Diagnostica developed the first disk diffusion method using modified Shadomy agar with Amphothericin B Fluconazole Fluorocytosine Itraconazole and Ketoconazole Neo Sensitabs Studies with fluconazole and ketoconazole have however shown a good correlation between in vitro and in vivo results using animal models 1 In vitro resistant strains isolated from clinical treatment failure were clearly more difficult to treat in the animal models than typical susceptible strains Troillet et al 1993 demonstrated a good correlation between MICs and zone diameters using a fluconazole disk on Yeast Nitrogen agar Barry et al 2 defined a simple disk diffusion test for rapid determination of the susceptibility of Candida spp to fluconazole using RPMI 1640 glucose agar Quind s et al 3 in a multicenter survey of antifungal resistance using Neo Sensitabs antifungals found a good correlation between the 12 Spanish laboratories involved in the survey Other papers about the use of antifungal Neo Sensitabs have been presented or published elsewhere 4 5 6 7 14 15 18 19 20 21 24 27 33 34 36 37 39 40 44 Subsequent studies modified the procedure to use Mueller Hinton Agar supplemented with 2 glucose because this medium is more readily available in the microbiological laboratory Further studies sho
73. is common in E faecium The MIC distribution of E faecalis without resistance mechanisms normal distribution against ampicillin is 0 125 to 2 ug ml c Staphylococci should be tested beta lactams against penicillin and oxacillin only Oxacillin resistant staphylococci should be considered resistant to all available beta lactam antibiotics Iso sensitest agar should not be used for testing staphylococci against oxacillin because of false sensitivity results BSAC 2002 d Cefoxitin may be used for the detection of mecA positive staphylococci oxacillin S aureus resistant to cefoxitin 60 ug zone 27 mm or less are mecA positive e Valid for S epidermidis S hominis and S haemolyticus Other coagulase negative staphylococci S saprophyticus S lugdunensis S xylosus should be tested using the zones recommended for S aureus 16 14 and or using cefoxitin f Isolates being I R MIC gt 1 ug ml should be tested for ESBL production See Neo Sensitabs User s Guide chapter 16 1 g The MIC breakpoints used are tentative The AFA group has not yet established MIC breakpoints for pseudomonas h Isolates with MIC gt 0 5 ug ml might produce metallo beta lactamases See User s Guide 2003 Imipenem EDTA or other broad spectrum beta lactamases Carbapenem testing on Iso sensitest agar may give false sensitive results with isolates able to produce metallo beta lactamases Testing on Mueller Hinton agar should be preferred BSAC 2
74. lineage CC 17 of Enterococcus faecium J Clin Microbiol 44 1059 64 2006 Bonora M G et al Emergence of linezolid resistance in the vancomycin resistant E faecium multilocus sequence typing C 1 epidemic lineage J Clin Microbiol 44 1153 55 2006 Lee W G et al Van B phenotype Van A genotype E faecium with heterogeneous expression of glycopeptide resistance in a Korean Hospital 46th ICAAC Abstract C2 0210 2006 Jae Hoon Song et al High frequency of UR E faecium isolates with vanB phenotype and van A genotype in Korean Hospitals Diagn Microbiol Infect Dis 56 401 406 2006 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 98 of 170 15 Susceptibility Testing of Fastidious or Problem Organisms The CLSI Kirby Bauer method and other diffusion tests have been standardized for rapidly growing pathogens Larger zones of inhibition will result if the test is performed with organisms that have a slow rate of growth resulting in erroneous results in the susceptibility test Consequently it is important to give optimal growth conditions to the strains being tested This may be achieved by using lower incubation temperature It is well known that methicillin resistant strains of staphylococci sometimes are reported as susceptible to methicillin by disk testing at 37 C but as resistant at 30 35 or with NaCl added to the medium This phenomenon is attributed to the heterogeneit
75. methods and current interpretative criteria 1 These latter isolates may not reach current CLSI breakpoints for resistance yet can be clinically resistant to beta lactam therapy 2 Since some ESBLs are more active on CEZDI while others are more active on CFT AX the choice of cephalosporins tested can also affect the ability of laboratories to detect resistant strains 3 Most ESBLs are inhibited by clavulanic acid tazobactam or sulbactam and can be readily detected by the double disk tablet synergy test 4 Double disk tablet synergy test Inoculate a Mueller Hinton plate as for susceptibility testing and apply Ceftriaxone CETRX Neo Sensitabs Cefotaxime CFTAX Neo Sensitabs Ceftazidime CEZDI Neo Sensitabs Cefepime CFEPM Neo Sensitabs and Aztreonam AZTRM Neo Sensitabs at approximately 20 mm 30 mm from tablet center to tablet center from a tablet containing Amoxycillin Clavulanate Neo Sensitabs AM CL using a dispenser Incubate overnight at 35 Extension of the zone of inhibition synergism towards the tablet containing AM CL indicates the presence of an extended spectrum beta lactamase ESBL Plate 16 1 a Klebsiella pneumoniae ATCC 700603 producing extended spectrum beta lactamases ESBL Note the synergy between cefotaxime Neo Sensitabs CFTAX ceftazidim Neo Sensitabs CEZDI and Amoxyeillin Clavulanate Neo Sensitabs AM CL Another possibility of screening for ESBL is the use of lower MI
76. mm MIC lt 0 5 ug ml and Cefepime Neo Sensitabs S gt 26 mm MIC lt 0 5 ug ml Strains showing zones less than 26 mm with any of the 3 cefalosporins should be sent to a Reference Laboratory 1 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 116 of 170 Streptococci except pneumoniae Mueller Hinton 5 blood Inoculum McFarland 0 5 Incubation in 5 7 CO Break points according to CLSI M100 515 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S R a Amoxycillin viridans 30 AMOXY gt 30 29 21 lt 20 lt 0 25 gt 8 a Ampicillin viridans 33 ug AMP33 gt 30 29 21 lt 20 lt 0 25 gt 8 Azithromycin 30 ug AZITR gt 22 21 19 lt 18 lt 0 5 gt 2 a Cefepime viridans 30 ug CFEPM gt 26 25 23 lt 22 lt 1 gt 4 Cefotaxime viridans 30 ug CFTAX gt 30 29 27 lt 26 lt 1 gt 4 S Cefpirome 30 ug CFPIR 230 29 27 lt 26 lt 1 gt 4 a Ceftriaxone viridans 30 ug CETRX gt 28 27 24 lt 23 lt 1 gt 4 Chloramphenicol 60 ug CLR60 gt 28 27 21 lt 20 lt 4 gt 16 Chloramphenicol 10 ug CLR10 gt 16 15 11 lt 10 lt 4 gt 16 Clarithromycin 30 ug CLARI gt 24 23 21 lt 20 lt 0 25 gt 1 e Clindamycin 25 ug CLIND gt 28 27 24 lt 23 lt 0 25 gt Daptomycin 30 ug DAPCa 222 lt 1 2 18 h prediffusion Doxycycline 80 ug DOXYC 226 25 23 22 lt 2 gt 8 e Erythromycin 78 ug ERYTR gt 28 27 24 lt 23 lt 0 25 gt 1 Ga
77. resistant S aureus that are multidrug resistant should be suspected of being MRSA and zones of inhibition around Oxacillin 1 ug Neo Sensitabs should be examined for the presence of a light film of growth within the zone of inhibition if found report R Cefoxitin is easier to interpret Community acquired MRSA CA MRSA infections appear to be an emerging phenomenon worldwide 11 21 They are susceptible to numerous drugs gentamicin chloramphenicol trimethoprim sulfa fosfomycin rifampicin etc They are mecA positive and can be detected using Cefoxitin Neo Sensitabs R The Vitek 2 system failed to detect MRSA strains having the community acquired MRSA phenotype Such errors may remain undetected in laboratories exclusively using broth based methods vitek 2 Microscan to test the susceptibility of S aureus isolates Disk diffusion and E test detected methicillin resistance 26 Warning Methicillin Oxacillin resistant S aureus and coagulase negative staphylococci CNS should be reported as resistant to all beta lactams including beta lactamase inhibitor combinations and imipenem regardless of in vitro test results with those agents 2 Do not use cephalosporins or carbapenem disks for susceptibility testing of staphylococci Use Cefoxitin and Oxacillin 1 ug Neo Sensitabs BORSA Borderline oxacillin resistant S aureus Some strains of S aureus with high B lactamase activity and mecA negative may show resistance to oxacillin
78. resistant S aureus isolate from a patient in Pennsylvania Antimicrob Ag Chemother 48 275 280 2004 MMWR Weekly April 2004 Brief report Vancomycin resistant Staph aureus New York 2004 Swaber M J et al Failure of broth based tests to detect methicillin resistant S aureus in a clinical specimen Eur J Clin Microbiol Infect Dis 23 348 51 2004 Lewis II J S et al Inducible clindamycin resistance in staphylococci should clinicians and microbiologists be concerned CID 40 280 285 2005 Jae Hoon Song et al Emergence in asian countries of S aureus with reduced susceptibility to vancomycin Antimicr Ag Chemother 48 4926 8 2004 Sieradzki K et al Evolution of VISA strain in vivo multiple changes in the antibiotic resistance phenotypes of a single lineage of MRSA under the impact of antibiotics administered for chemotherapy J Clin Microbiol 41 1687 93 2003 Howden B P et al Identification of a new agar dilution screening method for the accurate detection of hVISA 44th Annual ICAAC presentation D 59 2004 Sandberg A et al First report of heterogeneous vancomycin intermediate S aureus RVISA VISA in Denmark 44th Annual ICAAC presentation AR 07 2004 Nielsen S V Casals J B Detection of decreased susceptibility to glycopeptides in S aureus using tablet disc prediffusion 15th Eur Cong Clin Microbiol Infect Dis ECCMID April 2005 Palazzo I C V et al First report of vancomycin resistant staphy
79. s of 4 16 ug ml 15 Lesho et al 16 and Jones et al 17 documented a greater potency and higher susceptibility rates for imipenem compared to meropenem when Acinetobacter spp are tested Reversed for P aeruginosa where in isolates from the U S meropenem showed a 2 to 4 fold greater potency than imipenem and a percentage susceptibility advantage of 4 5 96 Because of possible discords when testing carbapenems against non fermenters the carbapenem selected for therapy should also be tested by the clinical laboratory References 1 Arpi M et al In vitro susceptibility of 124 Xanthomonas maltophilia Stenotrophomonas maltophilia isolates APMIS 104 108 114 1996 2 Denton M et al Microbiological and clinical aspects of infection associated with Stenotrophomonas maltophilia Clin Microbiol Reviews 14 57 80 1998 3 Valdezate S et al Antimicrobial susceptibilities of unique Stenotrophomonas maltophilia clinical isolates Antimicr Ag Chemother 45 1581 1584 2001 4 H iby N Prevention and treatment of infections in cystic fibrosis Intl J Antimicr Agents 1 229 238 1992 5 Pitt T L et al Type characterisation and antibiotic susceptibility of Burkholderia cepacia isolates from patients with cystic fibrosis in the U K and the Republic of Ireland J Med Microbiol 44 203 210 1996 6 Gowan J R W et al Burkholderia cepacia medical taxonomic and ecological issues J Med Microbiol 45 395 407 1996 7
80. should be reported resistant to all combinations with beta lactamase inhibitors Enterococci For treatment of serious infections S refers to high dose combination therapy When testing enterococci use Gentamicin 250 ug and Streptomycin 500 ug to detect high level resistance HLR Kanamycin 500 ug can be used to detect high level resistance to amikacin Strains that are HLR to gentamicin are HLR to all aminoglycosides including amikacin except streptomycin When testing enterococci and staphylococci plates should be incubated for full 24 hours and examined carefully for the presence of a haze or other growth within the zone indicates resistance see chapter 14 2 CLSI recommend a reference MIC method for staphylococcal isolates showing inhibition zones smaller than the limit for susceptible chapter 13 Pseudomonas aeruginosa Serious infections should be treated with maximum doses of antipseudomonal penicillin or ceftazidine in combination with an aminoglycoside According to CLSI The susceptibility of Ps aeruginosa isolated from patients with cystic fibrosis can be reliably determined by the diffusion method but may require extended incubation up to 24 hours CLSI M100 S17 2007 When blood is added to the medium the interpretation is Novobiocin 5 ug gt 13 mm I 12 11 mm R 10 mm and Fucidin S 26 mm I 25 23 mm R 22 mm Addition of blood should only be used when the strain does not grow well on unsupplemented
81. supplement with thiamin and or menadione for normal growth Some of the supplemental substances may interfere with the activity of the antibiotics e g CO affects the activity of aminoglycosides macrolides and tetracyclines in which case a modification of the zone size interpretation may be necessary Strains that require thymidine must be tested on media supplemented with thymidine because the effect of thymidine on trimethoprim can be disregarded since such organisms are resistant to trimethoprim in vitro and in vivo Species specific breakpoints When testing the susceptibility of slow growing organisms and or species with special requirements Haemophilus spp Neisseria spp pneumococci streptococci the methods must be modified to fit each organism and special interpretation tables are required 3 Special methodologies may be required to detect problem organisms such as methicillin resistant staphylococci resistant enterococci and extended spectrum beta lactamase producing gram negative bacilli Methodologies including interpretation tables are described in the following pages References 1 Manual of Clinical Microbiology Ed by Murray Baron Pfaller Tenover amp Yolken 6th Ed 1995 2 Antibiotics in Laboratory Medicine Ed by V Lorian Williams amp Williams Baltimore 48 50 1980 3 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 2003 O Copyright Rosco Diagnostica A S
82. testing of yeast against Ketoconazole Itraconazole and Fluconazole by an agar diffusion method Scand Meet Bact Gothenburg May 1989 Casals J B Pringler N Improved antifungigramme using Ketoconazole Itraconazole and Fluconazole Neo Sensitabs Joint Meet Brit Scand Soc Mycopath and Swed Soc Clin Mycology Gothenburg Sept 1989 Martinez Suarez J V Rodriguez Tudela J L Patterns of in vitro activity of Itraconazole and Imidazole antifungal agents against C albicans with decreased susceptibility to Fluconazole in Spain Antimicr Agents Chemother 39 1512 1516 1995 Sandven P Detection of Fluconazole resistant Candida strains by a disc diffusion screening test J Clin Microbiol 37 3856 9 1999 Carrillo Mufioz A J Multicenter evaluation of Neo Sensitabs a standardized diffusion method for yeast susceptibility testing Rev Iberoam Micol 16 92 96 1999 Ceballos A et al In vitro antifungal resistance in Candida albicans from HIV patients with and without oral candidosis Rev Iberoam Micol 16 194 7 1999 Carrillo Mufioz AJ et al In vitro resistance to fluconazole and itraconazole in clinical isolates of Candida spp and Cryptococcus neoformans Rev Iberoam Micol 14 50 54 1997 Spanish Carrillo Mufioz AJ Multicenter evaluation of the reproducibility of Neo Sensitabs antifungal sensitivity testing Rev Iberoam Micol 11 56 1994 Spanish Carrillo Mufioz AJ et al Comparison of two methods for sensitivi
83. they show resistance to both VANCO and TEICO The Van B phenotype can also be transferred by conjugation These strains show with the 24 18 hours prediffusion method zone lt 16 mm and hazy edge Van C Strains characterized by a constitutive low level resistance to vancomycin but being susceptible to teicoplanin Strains of phenotype Van C are less frequently encountered species of motile enterococci They can be detected by performing a motility test Motile enterococci show either intrinsic low level resistance to vancomycin zone around Vancomycin 5 ug Neo Sensitabs or have acquired high level resistance no zone around Vancomycin 5 Neo Sensitabs Van A phenotype These strains show with the 24 18 hours prediffusion method zone lt 16 mm and sharp edge E casseliflavus E flavescens and E gallinarum with vancomycin MIC s of 8 16 ug ml intermediate differ from vancomycin resistant enterococci for infection control purposes NCCLS 2000 Heterogeneous phenotype Recently new resistant phenotypes Van D Van E have been described 7 Rabiul Alam M et al 8 describes heteroresistance to Vancomycin in E faecium The diffusion test with Vancomycin 5 ug Neo S will show growth af subcolonies inside the zone of inhibition Van B phenotype Van A genotype E faecium with heterogeneous expression of glycopeptide resistance have been isolated from a Korean hospital Vanco MIC 64 128 ug ml and Teico MIC 4 12 ug ml 26 27
84. ug FOSFO 25 33 28 35 FUCIDIN 100 ug FUCID 28 38 GATIFLOXACIN 5ug GATIF 30 37 27 33 20 28 GENTAMICIN 40 ug GEN40 25 31 25 32 25 31 GENTAMICIN 250 ug GN250 17 23 IMIPENEM 15 ug IMIPM 29 34 23 31 26 31 KANAMYCIN 100 ug KANAM 26 32 25 31 LEVOFLOXACIN 5ug LEVOF 30 38 26 31 21 28 LINEZOLID 30 ug LINEZ 25 31 21 25 MECILLINAM 33 ug MECIL 271 35 MEROPENEM 10 ug MEROP 31 38 32 42 31 38 18 24 MINOCYCLINE 80 ug MINOC 24 30 27 33 MOXIFLOXACIN 5 ug MOXIF 28 35 28 35 17 25 MUPIROCIN 10ug MUPIR 21 26 NALIDIXAN 130 ug NALID 27 33 NEOMYCIN 120 ug NEOMY 23 28 23 30 NETILMICIN 40 ug NETIL 24 30 25 32 22 27 NITROFURANTOIN 260 ug NITRO 24 30 24 30 24 29 NORFLOXACIN 10 ug NORFX 28 35 17 26 19 26 12 17 NOVOBIOCIN 5 ug NOV 5 18 25 OFLOXACIN 10 ug OFLOX 29 38 23 29 22 26 16 22 OXACILLIN 1 pg 1 18 24 OXACILLIN 5 ug 5 22 28 OXOLINIC ACID 10ug OXOLI 30 36 18 24 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 12 Page 82 of 170 Zone diameter in mm E coli S aureus Ps aeruginosa Ent faecalis NEO SENSITABS CODE ATCC 25922 ATCC 25923 ATCC 27853 ATCC 29212 PENICILLIN LOW 5 ug PEN L 26 36 15 21 PIPEMIDIC ACID 30 ug PIPEM 26 32 PIPERACILLIN 100 ug PIPRA 26 31 25 33 25 30 PIPERACILLIN 100 10 ug PI TZ 26 31 25 33 TAZOBACTAM POLYMYXINS 150 ug CO150 19 24 20 25 QUINUPRISTIN 15 ug SYNI5 21 28 DALFOPRISTIN R
85. 0 12 gt 2 a Moraxella catarrhalis BL neg BL pos Test betalactamase prod Penicillin Low V 5 ug PEN L Haemophilus spp gt 30 29 14 lt 14 lt 0 5 gt 4 Oxacillin lug OXA 1 b S pneumoniae gt 24 2 lt 24 lt 0 06 pe j Streptococcus spp gt 17 lt 17 lt 0 12 pen Ampicillin 2 5 ug AMP L c Haemophilus spp gt 20 19 16 lt 16 lt 1 gt 4 Ampicillin 33 ug AMP33 Haemophilus spp gt 32 31 26 lt 26 lt 1 gt 4 d S pneumoniae non meningeal gt 36 35 29 lt 29 lt 0 5 gt 2 Beta haem streptococcus gt 30 lt 0 25 Viridans streptococcus gt 36 35 24 lt 24 lt 0 25 gt 4 a Moraxella catarrhalis BL neg BL pos Test betalactamase prod Amoxycillin Clav 30 15 ug AM CL Haemophilus spp gt 30 29 26 lt 26 lt 2 gt 4 Moraxella catarrhalis gt 34 33 26 lt 26 lt 1 gt 4 Cephalosporins Cefuroxime 60 ug CEFUR Haemophilus spp gt 34 33 28 lt 28 2 gt 4 Moraxella catarrhalis gt 34 lt 34 lt 2 gt 2 d S pneumoniae non meningeal 238 37 32 32 0 5 gt 1 Cefotaxime 30 ug CFTAX e Haemophilus spp gt 38 37 28 28 lt 0 125 gt 1 Moraxella catarrhalis gt 33 32 29 lt 29 lt 2 gt 4 Ceftizoxime 30 ug CEZOX f pneumoniae gt 32 lt 32 lt 0 5 gen cephalosporins CFTAX CETRX Ceftriaxone 30 ug CETRX e Haemophilus spp 238 37 28 lt 28 lt 0 125 gt 1 Monobactams Aztreonam 30 ug AZTRM Haemophilus spp gt 40 39 32 lt 32 lt 0 5 gt 2 catarrhalis gt 28
86. 0 19 17 lt 16 lt 1 1 4 gt 2 2 16 Neomycin 120ug NEOMY 223 22 20 lt 19 lt 6 gt 25 c Nitrofurantoin 260ug NITRO gt 23 22 20 lt 19 lt 32 gt 128 nitrofurans c Novobiocin 5 5 Blood 213 12 11 lt 10 lt 2 gt 4 Plain agar gt 16 15 14 13 lt 2 gt 4 f Oxacillin 1 ug 1 neg staph gt 18 lt 17 lt 0 25 20 5 S aureus gt 13 12 11 lt 10 lt 2 gt 4 S pneumoniae penicillin 220 lt 19 lt 0 06 gt 0 12 c Oxolinic acid 10ug OXOLI 220 19 17 lt 16 lt 4 gt 8 Penicillin Novo 10U 30 ug PEN N Mastitis gt 18 17 15 lt 14 lt 1 2 gt 4 8 Other organisms 217 lt 16 lt 1 2 gt 4 8 Penicillin Low 5 ug PEN L Staphylococcus spp 226 lt 25 lt 0 12 gt 0 25 Streptococcus viridans gt 26 25 13 lt 12 lt 0 12 gt 4 Beta haemolytic gt 26 lt 0 12 Other organisms gt 18 17 11 lt 10 lt 1 gt 4 Pirlimycin 10ug PIRLI gt 18 lt 17 lt 2 gt 4 Rifampicin 30 ug RIFAM S pneumoniae gt 28 27 24 lt 23 lt 1 gt 4 Enterococci gt 26 25 23 lt 22 lt 1 24 Spectinomycin 200ug gt 18 17 15 lt 14 lt 32 gt 128 Pasteur Haemoph Spiramycin 200ug SPIRA 226 25 23 lt 22 lt 2 gt 8 c Streptomycins 100ug 100 Pseudomonas spp gt 32 lt 31 lt 6 gt 25 Other organisms gt 26 25 23 lt 22 lt 6 gt 25 Sulphonamides U 240ug SULFA 223 22 20 lt 19 lt 256 gt 512 Tetracyclines 80 ug TET80 Streptococcus spp Cattle gt 26 25 23
87. 002 1 Concerning detection of VISA GISA strains and the detection of vancomycin resistant enterococci VRE see Neo Sensitabs User s Guide chapter 14 2 jp Erythromycin is the group representative for the macrolides and results are valid for clarithromycin and azithromycin k Staphylococci and enterococci resistant to gentamicin should be reported as resistant to netilmicin and tobramycin 1 Gentamicin is recommended as test drug for staphylococci because netilmicin might give false sensitive results with coagulase negative staphylococci Isolates sensitive to gentamicin will currently be sensitive to netilmicin m Tobramycin sensitive strains will currently be sensitive to gentamicin and netilmicin O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 n 9 r s u x y 7 Page 48 of 170 Gentamicin HLR in enterococci indicates resistance to all aminoglycosides except streptomycin Gentamicin sensitive E faecalis but not E faecium are also sensitive to netilmicin E faecium shows intrinsic resistance towards kanamycin tobramycin and netilmicin due to the production of the enzyme AAC 6 Isolates with MICs gt 0 125 ug ml cipro or MIC gt 0 5 ug ml oflox show reduced sensitivity to ciprofloxacin ofloxacin There is a risk of resistance development during treatment Nalidixan resistance can be used to detect strains with reduced susceptibility to the fluoquinolon
88. 01 15 Kamile Rasheed J et al Characterization of ESBL Reference Strain Kl pneumoniae ATCC 700603 which produces the novel enzyme SHV 18 Antimicr Ag Chemother 44 2382 2388 2000 16 Florijn A et al Comparison of E test and double disk diffusion test for detection of extended spectrum Beta lactamases Eur J Clin Microbiol Infect Dis 21 241 243 2002 17 Tzelepi E et al Detection of ESBL in clinical isolates of Enterobacter cloacae and E aerogenes 1 Clin Microbiol 38 542 6 2000 18 Oliver A et al Mechanisms of decreased susceptibility to Cefpodoxime in E coli Antimicr Ag Chemoter 46 3829 3836 2002 19 Bolmstr m A Cefepime Clavulanic acid in a E test configuration for investigating non determinable ESBL results for NCCLS criteria 42nd ICAAC Presentation D 527 2002 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 Page 131 of 170 Rodriguez Villalobos H et al Evaluation of the combined disk method for the detection of ESBL in Enterobacteriaceae ICAAC 2003 presentation D 202 Fluit A et al Comparison of phenotypic tests for the detection of ESBLs and AmpC beta lactamases ICAAC 2003 presentation D 199 Pitout J D D et al Modification of the doulbe disk test for detection of Enterobacteriaceae producing ESBL and AmpC beta lactamases J Clin Microbiol 44 3933 5 20
89. 03 Decr D et al Outbreak of multiresistant K oxytoca involving strains with ESBLs and strains with extended spectrum activity of the chromosomal B lactamase J Antimicr Chemother 54 881 8 2004 Lartigue M F et al First detection of a carbapenem hydrolyzing metalloenzyme in an Enterobacteriaceae isolate in France Antimicr Ag Chemother 48 4929 30 2004 Kim J Y et al Nosocomial outbreak by P mirabilis producing extended spectrum B lactamase VEB 1 in a Korean University Hospital J Antimicr Chemother 54 1144 7 2004 Yan J J et al Plasmid mediated 16S rRNA methylases conferring high level aminoglycoside resistance in E coli and pneumoniae isolates from two Taiwanese hospitals Antimicr Chemother 54 1007 12 2004 Tristram S G et al Disc diffusion based screening tests for ESBL s in influenzae J Antimicr Chemother 55 570 3 2005 Lincopan N et al First isolation of metallo B lactamase producing multiresistant pneumoniae from a patient in Brazil J Clin Microbiol 43 516 9 2005 Pitout J D D et al Emergence of Enterobacteriaceae producing ESBLs in the community J Antimicr Chemother 56 52 59 2005 Lee Seungok et al Prevalence of Ambler class A and D beta lactamase among clinical isolates of P aeruginosa in Korea J Antimicr Chermother 56 122 27 2005 Song Wonkeun et al Failure of cefepime therapy in treatment of K pneumoniae bacteremia J Clin Microbiol 43 4891 94 2005
90. 07 2008 Chapter 10 Page 32 of 170 10 2 Il Interpretations according to MIC Breakpoints of the Dutch CRG Il a Susceptibility testing of rapidly growing bacteria using Neo Sensitabs and MIC break points as recommended by the Dutch CRG September 2000 Medium Iso Sensitest Inoculum according to ICS Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S Ampicillin 33 ug AMP33 gt 28 27 18 lt 18 lt 2 gt 16 Amoxycillin 30 ug AMOXY 228 27 18 18 lt 2 gt 16 a Amoxycillin Clav 30 15 ug AM CL gt 28 27 18 lt 18 lt 2 0 5 gt 16 4 b Methicillin 29 ug METHI lt 4 gt 4 b Oxacillin MH McF 0 5 lug OXA 1 S aureus 213 13 lt 2 gt 2 Staphylococcus spp gt 18 18 lt 0 25 gt 0 5 c Penicillin Low 5 ug PEN L Staphylococcus spp gt 28 z lt 28 lt 0 25 Others 228 27 10 10 lt 0 25 24 Piperacillin 100ug PIPRA 226 25 20 20 lt 16 gt 64 Piperacillin Tazobactam 100 10ug PI TZ gt 26 25 22 lt 22 lt 1642 gt 32 4 Temocillin 30 ug TEMOC 220 19 15 15 lt 8 gt 32 Ticarcillin 75 ug TICAR gt 26 25 20 lt 20 lt 16 gt 64 Ticarcillin Clavulanate 75 15 ug TI CL gt 26 25 20 lt 20 lt 16 4 gt 6444 d Cefaclor 75415 ug TI CL 226 25 20 20 lt 4 gt 16 Cefazolin 60 ug CFZOL gt 28 27 23 lt 23 lt 4 gt 16 e m Cefotaxime 30 ug CFTAX 226 25 20 20 lt 4 gt 16 Cefotetan 30 ug CFTTN gt 22 21 18 18 lt 4 gt 16 b2 Cefoxitin 60 ug CF
91. 08 Chapter 16 Page 144 of 170 References 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 Walsh T R et al Evaluation of new E test for detecting metallo beta lactamases in routine clinical testing J Clin Microbiol 40 2755 59 2002 Jong D et al Imipenem EDTA disk method for differentiation of metallo beta lactamase producing clinical isolates of Pseudomonas spp and Acinetobacter spp J Clin Microbiol 40 3798 3801 2002 Larrosa M N et al E coli multi resistente productora de una metalo beta lactamasa Spanish SEIMC Congress Bilbao 16 19th May 2004 Lartigue et al First detection of carbapenem hydrolyzing metalloenzyme in an Enterobacteriaceae isolate in France Antimicr Ag Chemother 48 4929 30 2004 Castanheira M et al Molecular characterization of a beta lactamase gene blaGIM 1 a new subclass of metallo beta lactamases Antimicr Ag Chemother 48 4654 61 2004 Jing Jou Yan et al Comparison of the double disk combined disk and E test methods for detecting metallo beta lactamases in gram negative bacilli Diagn Microbiol Infect Dis 49 5 11 2004 Walsh T R et al Metallo beta lactamases The quite before the storm Clin Microbiol Reviews 18 306 325 2005 Tortola M T et al First detection of a carbapenem hydrolyzing metalloenzyme in two Enterobacteriaceae isolates in Spain Antimicr Ag Chemother 49 3492 4
92. 1 1 2 Interpretation table fo Local treatment sss eene nennen 163 21 1 3 Candida spp Quality Control MH GMB sse nennen nennen 164 21 2 Procedure using Shadomy modified agar srvrrvvronovenoverrvvravernnvrarnrernenvvnenvennvnnnvernevevrvevaverssevarraevrneveesvessee 164 21 2 1 Interpretation Tables for Yeasts modified Shadomy agar sese 164 21 2 2 Candida spp Quality Control modified Shadomy Agar sse 165 22 Veterinary Practice CEST iac eu ERREUR UR FRED EUER RUE whee ae 168 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Page 5 of 170 PREFACE The 19th Ed 2007 2008 of the NEO SENSITABS User s Guide contains updated text tables and references all informative information when using Neo Sensitabs tablets for susceptibility testing A table giving the changes from the 19th Ed 2007 2008 is available at our website The different interpretation tables following the CLSI formerly NCCLS recommendations have been updated according to the latest information of the CLSI described in Performance Standards for Antimicrobial Susceptibility Testing 15th Informational Suppl M100 S17 2007 Furthermore the User s Guide includes updated Zone Diameter Interpretative Standards according to CRG SRGA AFA Denmark SFM DIN and BSAC Information and recommendations given by susceptibility testing standardization groups or or
93. 15 15 lt 2 gt 2 p Daptomycin 30 ug DAPCa 2 18 h prediffusion Staphyloccus spp 222 lt 1 gt 1 Enterococcus spp gt 12 lt 4 d Sulphonamides U 240ug SULFA 222 22 lt 256 gt 256 Trimethoprim 5 2 ug TRIME 220 19 16 16 lt 2 gt 4 Trimethoprim Sulfa 5 2 240 ug TR SU gt 28 27 23 lt 23 lt 16 120 gt 32 d Novobiocin U 5ug NOVOS5 gt 16 lt 16 d Mupirocin 10 ug MUPIR Staphylococcus spp gt 20 lt 20 lt 2 gt 2 Tigecycline 15 ug TIG15 Enterobacteriaceae enterococci gt 23 lt 23 lt 1 gt 2 Staphylococcus gt 25 lt 25 lt 0 5 gt 0 5 Remarks Staphylococci do not need to be tested against Ampicillin Amoxycillin and Piperacillin use Penicillin Low Staphylococci resistant to Penicillin Low beta lactamase producers should be reported as resistant to the above mentioned penicillins a b c d e f 8 h Valid for amoxycillin Including azidocillin with enterococci Klebsiella and Enterobacter are always reported R to ampicillin For aminoglycosides the intermediate category is only valid in case of UTI For other tests the intermediate results should be considered as resistant Mecillinam when using Mueller Hinton use S gt 24 mm I 23 16 mm R 16 mm When testing against P mirabilis use S gt 20 mm I 19 16 mm R lt 16 mm Break points not yet established by the SRGA If the media contain blood use S gt 28 mm R l
94. 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 Page 91 of 170 MMWR Staph aureus resistant to vancomycin United States 2002 MMWR Weekly July 5 2002 51 26 565 567 2002 CDR Weekly Staph aureus with reduced susceptibility to vancymycin 17 May 2002 Johnson A P et al Glycopeptide resistant Staph aureus J Antimicrob Chemoter 50 621 3 2002 Cui L et al Cell wall thickening is a commen feature of vancomycin resistance in Staph aureus J Clin Microbiol 41 5 14 2003 Sievert D M et al Investigation of a van A positive Vancomycin resistant Staph aureus infection 42nd ICAAC Presentation LB 6 2002 Chavez Bueno S et al Inducible resistance to clindamycin evidenced by D test in community acquired MRSA in children from 1999 to 2002 The Dallas experience ICAAC 2003 presentation D 246 Vandenesch F et al Commonuty acquired MRSA carrying Panton Valentine Leucocidin genes worldwide emergence Emerg Infect Dis 9 978 984 2003 Siberry G K et al Failure of clindamycin treatment of MRSA expressing inducible clindamycin resistance in vitro Clin Infect Dis 37 1257 60 2003 Liu C et al S aureus with heterogeneous resistance to vancomycin epidemiology clinical significance and critical assessment of diagnostic methods Antimicr Ag Chemother 47 3040 5 2003 Tenover F C et al Vancomycin
95. 17 16 8 232 c Cefazolin 60 ug CFZOL gt 23 22 20 lt 19 8 232 c Cefepime 30 ug CFEPM gt 20 19 17 lt 16 8 gt 32 Cefepime Clav 30 10 ug CP CL Detection of ESBL c m Cefixime 30 ug CFFIX gt 26 25 23 lt 22 lt 1 gt 4 c Cefonicid 30 ug CFCID gt 20 19 17 lt 16 lt 8 gt 32 c Cefotaxime 30 ug CFTAX gt 23 22 20 lt 19 lt 8 gt 32 Cefotetan 30 ug CFTTN 218 17 15 lt 14 lt 16 gt 64 Cefoxitin 60 ug CFOXT 223 22 20 lt 19 lt 8 gt 32 t S aureus and S lugdunensis 225 lt 24 OxaS MecA pos t Coag neg staph gt 28 lt 27 OxaS Mec A pos c Cefpirome 30 ug CFPIR gt 20 19 17 lt 16 lt 8 gt 32 0 Cefpodoxime 30 ug CFPOX 225 24 21 lt 20 lt 2 28 ESBL screening lt 20 gt 8 Cefsulodin 30 ug CFSUL gt 23 22 20 lt 19 lt 8 gt 32 c Ceftazidime 30 ug CEZDI 220 19 17 lt 16 lt 8 gt 32 0 ESBL screening lt 24 gt 2 Ceftazidime Clav 30 10 ug CZ CL Detection of ESBL c Ceftizoxime 30 ug CEZOX gt 23 22 20 lt 19 8 232 S aureus gt 23 lt 20 Oxa S Mec A pos Oxa R c Ceftriaxone 30 ug CETRX gt 20 19 17 lt 16 lt 8 232 0 ESBL screening 24 22 k Cefuroxime parenteral 60 ug CEFUR 223 22 20 lt 19 lt 8 gt 32 k Cefuroxime oral 60 ug CEFUR 225 24 21 lt 20 lt 4 gt 32 c Cephalexin 30 ug CFLEX gt 20 19 17 lt 16 8 232 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 28 of 170 Zone diameter Break
96. 2 13 2003 2 CLSI Performance Standards for Antimicrobial Susceptibility Testing 15th Inf Suppl M100 S15 2005 3 Hoff Gerdi Microbiol Dept Herning Hospital DK Personal communication 1990 4 Gransden W R et al Decreased susceptibility of Neisseria gonorrhoeae to ciprofloxacin The Lancet 335 51 1990 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 5 6 7 8 Page 106 of 170 Knapp J et al Proposed criteria for interpretation of susceptibilities of strains of N gonorrhoeae to Ciprofloxacin Ofloxacin Enoxacin Lomefloxacin and Norfloxacin Antimicrob Agents Chemother 39 2442 2445 1995 Anon Dramatic increase in ciprofloxacin resistant gonorrhoea in England and Wales CDR Weekly 10th April 2003 Regunathan P L et al Nalidixic acid susceptible ciprofloxacin resistant N gonorrhoeae strain in the UK JAC 56 437 2005 Tanaka M et al Analysis of mutations within multiple genes associated with resistance in a clinical isolate of N gonorrhoeae with reduced ceftriaxone susceptibility that shows a multidrug resistant phenotype Intl J Antimicr Ag 27 20 26 2006 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 107 of 170 15 3 Susceptibility Testing of Meningococci The recommended agar medium is Mueller Hinton Agar with 5 Blood added Use colonies taken directly from an overnight culture and make a suspension in broth or 0 9 saline eq
97. 29 24 lt 24 lt 1 gt 2 pneumoniae Test ceftizoxime lt 0 5 gt 2 Streptococcus spp gt 32 31 28 28 lt 0 5 gt 0 5 influenzae Moraxella catarrhalis gt 38 37 34 lt 34 lt 0 12 gt 0 12 N meningitidis gt 38 37 34 lt 34 lt 0 12 gt 0 12 N gonorrhoeae gt 38 37 34 lt 34 lt 0 12 gt 0 12 Ceftazidime 30 ug CEZDI lt 4 gt 8 Enterobacteriaceae gt 30 29 24 lt 24 lt 1 gt 8 Pseudomonas spp gt 28 lt 28 lt 8 gt 8 Ceftriaxone 30 ug CETRX lt 1 gt 2 Enterobacteriaceae gt 30 29 24 lt 24 lt 1 22 S pneumoniae Test ceftizoxime lt 0 5 gt 2 Streptococcus spp gt 32 31 28 lt 28 lt 0 5 gt 0 5 influenzae Moraxella catarrhalis gt 36 35 32 lt 32 0 12 gt 0 12 N meningitidis gt 36 35 32 lt 32 lt 0 12 gt 0 12 N gonorrhoeae gt 36 35 32 lt 32 lt 0 12 gt 0 12 Cefuroxime 60 ug CEFUR lt 4 gt 8 Enterobacteriaceae gt 26 d 26 8 28 S pneumoniae non meningeal gt 36 35 32 lt 32 lt 0 5 gt 1 Streptococcus spp gt 36 35 34 lt 34 lt 0 5 gt 0 5 influenzae Moraxella catarrhalis 236 35 30 lt 30 lt 1 gt 2 Ceftizoxime 30 ug CEZOX S pneumoniae 3 gen cephalosporins 232 31 28 28 lt 0 5 MIC O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 78 of 170 Zone diameter Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S R S R 6 Carbapenems Ertapenem 10 ug ETP10 lt 0 5 gt 1 Enterobacteriaceae gt
98. 3 686 707 2000 8 Rabiul Alam et al Heteroresistance to Vancomycin in Enterococcus faecium J Clin Microbiol 39 3379 81 2001 9 Weinstein M P Comparative evaluation of penicillin ampicillin and imipenem MICs and susceptibility breakpoints for vancomycin susceptible and vancomycin resistant faecalis and E faecium J Clin Microbiol 49 2729 2731 2001 10 Amin N El et al Ampicillin sensitive Imipenem resistant strains of E faecium J Clin Microbiol 40 738 2002 11 Lefort A et al Influence of Van D type resistance on activities of glycopeptides in vitro and in experimental endocarditis due to Enterococcus faecium Antimicr Ag Chemother 47 3515 8 2003 12 Yu Hong Min et al Heterogeneity of macrolide lincosamide streptogramin B resistance phenotypes in enterococci Antimicr ag Chemother 47 3415 20 2003 13 Abad a Patifio L et al Van E type vancomycin resistant E faecalis clinical isolates from Australia Antimicr Ag Chemother 48 4882 5 2004 14 Corso A et al First report of Van A E gallinarum dissemination within an intensive care unit in Argentina Intern J Antimicr Ag 25 51 6 2005 15 Reynolds P E et al Vancomycin resistance in enterococci due to synthesis of precursors terminating in D alanyl D serine Antimicr Ag Chemother 49 21 25 2005 16 Metzidie E et al An unusual resistance phenotype in nosocomial strains of Enterococcus spp in a Greek hospital Clin Microbiol
99. 3 lt 8 gt 32 c Enterobacteriaceae lt 26 Nedsat f lsomhed for kinoloner Staphylococcus spp ingen zone Muligvis nedsat f l undtagen saprophyticus somhed for kinoloner Novobiocin U 5ug NOVOS 216 16 lt 2 gt 2 Tentative Bem rkninger til afl sningsskemaet a b c d e 8 Enterobacteriaceae Klebsiella og Enterobacter rapporteres altid R mod ampicillin Enterobacter og Proteus spp b r rapporteres R mod Nitrofurantoin uanset zonest rrelse Nalidixan er en god screening til p visning af nedsat f lsomhed overfor kinoloner hos Salmonella spp Stammer resistente overfor Nalidixan Neo S zone lt 26 mm viser nedsat f lsomhed ovefor kinoloner CIPRO MIC gt 0 125 ug ml 1 Bem rk nye MIC breakpoints for Kinoloner skandinavisk model Tidligere MIC breakpoints var fra CLSI Bem rk at EUCAST anbefaler en f lsom breakpoint for ciprofloxacin med Enterobacteriaceae p 0 5 ug ml svarende til en zone p 28 mm E coli Klebsiella Salmonella stammer der udviser zoner lt 23 mm overfor Cefpodoxime Neo Sensitabs producerer formodentlig ESBL enzymer Til bekr ftelse test Ceftazidime Clavulanate og Cefepime Clavulanate mod Ceftazidime Cefepime Neo Sensitabs se side 13 9 1 Til p visning af ESBL se kapitel 16 1 Extended spectrum beta lactamases ESBL Til p visning af plasmid mediated Amp C beta lactamases se kapitel 16 4 Plasmid mediated AmpC beta lactamases
100. 30 29 26 lt 26 lt 0 5 gt 0 5 Norfloxacin 10ug NORFX Enterobacteriaceae nal gt 28 27 24 lt 24 lt 0 5 gt 1 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 77 of 170 Zone diameter Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S R S R Ofloxacin 10 ug OFLOX Enterobacteriaceae nal gt 28 27 24 lt 24 lt 0 5 gt 1 Staphylococci gt 26 25 22 lt 22 lt 1 gt 1 S pneumoniae gt 36 35 20 lt 20 lt 0 12 gt 4 H influenzae Moraxella spp nal gt 30 29 26 lt 26 lt 0 5 gt 0 5 Gonococci nal gt 34 lt 34 lt 0 12 gt 0 25 3 Glycopeptides Teicoplanin 30 ug TPN30 Staphylococci Enterococci gt 16 15 14 lt 14 lt 4 gt 8 Streptococci S pneumoniae gt 17 16 lt 16 lt 4 gt 4 Vancomycin 5ug VAN 5 n Staphylococci Enterococci 216 15 14 14 lt 4 gt 8 Streptococci S pneumoniae gt 18 17 16 16 lt 4 gt 4 4 Oxazolidinones Linezolid 30 ug LINEZ Staphylococci Enterococci 226 25 22 22 lt 4 gt 4 Streptococci S pneumoniae gt 28 27 23 lt 23 lt 2 gt 4 5 Cephalosporins Cefepime 30 ug CFEPM lt 4 gt 8 Enterobacteriaceae gt 30 29 24 lt 24 lt 1 gt 8 Pseudomonas spp gt 28 lt 28 lt 8 gt 8 pneumoniae Test ceftizoxime lt 1 gt 2 Streptococcus spp gt 32 31 28 lt 28 lt 0 5 gt 0 5 H influenzae Moraxella catarrhalis gt 35 34 31 lt 31 lt 0 25 gt 0 25 Cefotaxime 30 ug CFTAX lt 1 gt 2 Enterobacteriaceae gt 30
101. 30 29 26 lt 26 lt 0 5 gt 1 Streptococci S pneumoniae gt 30 29 26 lt 26 lt 0 5 gt 0 5 H influenzae Moraxella catarrhalis gt 30 29 26 lt 26 lt 0 5 gt 0 5 Anaerobes gt 32 31 28 lt 28 lt 1 gt 1 Imipenem 15 ug IMIPM lt 2 gt 8 Enterobacteriaceae gt 26 25 20 lt 20 lt 2 gt 8 Pseudomonas spp gt 28 27 24 lt 24 lt 4 gt 8 Acinetobacter gt 26 23 18 lt 18 lt 2 gt 8 Streptococci S pneumoniae gt 28 27 25 lt 25 lt 2 gt 2 Enterococcus spp gt 24 23 20 lt 20 lt 4 gt 8 H influenzae Moraxella catarrhalis gt 32 31 28 lt 28 lt 2 22 Anaerobes 228 271 20 20 lt 2 gt 8 Meropenem 10 ug MEROP Enterobacteriaceae gt 26 25 20 lt 20 x gt 8 Pseudomonas spp gt 30 29 24 lt 24 lt 2 gt 8 Acinetobacter gt 26 25 20 lt 20 lt 2 gt 8 Streptococci S pneumoniae gt 28 27 25 lt 25 lt 2 gt 8 H influenzae Moraxella catarrhalis gt 28 27 24 lt 24 lt 2 gt 2 N meningitidis gt 32 31 28 lt 28 lt 0 25 gt 0 25 Anaerobes 232 31 26 26 lt 2 gt 8 7 Others Aztreonam 30 ug AZTRM lt 4 gt 8 Enterobacteriaceae gt 30 29 24 lt 24 lt 1 gt 8 Pseudomonas spp gt 32 31 20 lt 20 lt 1 gt 16 Tigecycline 15 ug TIG15 lt 0 25 gt 0 5 Enterobacteriaceae gt 24 23 20 lt 20 lt 1 gt 2 Staphylococci gt 26 lt 26 lt 0 5 gt 0 5 Streptococcus gt 25 24 20 lt 20 lt 0 25 gt 0 5 Enterococcus spp 221 21 lt 0 25 gt 0 5 pnemoniae gt 24
102. 30 ug CFPIR 228 2027 15 19 lt 14 lt 2 3 8 16 32 gt 32 0 Cefpodoxime 30 ug CFPOX gt 24 lt 23 lt 4 28 Cefpodoxime 10 ug CPD10 lt 20 Screen ESBL Cefsulodin 30 ug CFSUL gt 28 20 27 15 19 lt 14 lt 2 3 8 12 32 gt 32 b c Ceftazidime 30 ug CEZDI gt 28 20 27 15 19 lt 14 lt 2 3 8 12 32 gt 32 Ceftazidime Clav 30 10ug CZ CL P visning af ESBL b c Ceftriaxone 30 ug CETRX gt 26 18 25 13 17 lt 12 lt 2 3 8 12 32 gt 32 b Cefuroxime 60 ug CEFUR gt 28 2327 15 22 lt 14 x34 6 8 12 64 gt 64 b Cephalothin 60 ug CLOTN gt 28 2327 15 22 lt 14 lt 3 4 6 8 12 64 gt 64 i Ciprofloxacin 10 ug CIP10 gt 30 21 29 lt 20 lt 0 25 0 5 2 gt 2 Clarithromycin 30 ug CLARI 222 19 21 13 18 lt 12 lt 1 5 2 4 6 16 gt 16 Clindamycin 25ug CLIND 228 2327 1522 lt 14 lt 1 1 5 2 4 8 28 Cloxacillin 500 ug CL500 P visning af plasmid Amp C Colistin 10 ug CO 10 s 24 18 t pr diffusion 215 lt 15 lt 2 gt 2 Daptomycin 30 ug DAPCa 5 2 18 t pr diffusion Staphylococcus spp 222 1 Enterococcus spp gt 12 lt 4 Erythromycin 78 ug ERYTR gt 28 23 27 15 22 lt 14 1 2 4 6 32 gt 32 r Ertapenem 10 ug ETP10 230 29 26 lt 26 lt 0 5 gt 1 Fosfomycin 70 40ug FOSFO 218 16 17 13 15 lt 12 Fucidin 3 100 ug FUCID gt 24 23 lt 22 lt 0 5 1 22 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 54 of 170 Zone diameter
103. 32 Imipenem 15 ug IMIPM S pneumoniae gt 32 31 28 lt 28 lt 0 5 gt 2 Campylobacter spp gt 24 23 20 lt 20 lt 4 gt 8 Anaerobes gt 24 23 20 lt 20 lt 4 gt 8 Meropenem 10 ug MEROP S pneumoniae gt 32 31 28 lt 28 lt 0 5 gt 2 Ertapenem 10 ug ERTAP S pneumoniae gt 28 27 24 lt 24 lt 1 gt 2 Haemophilus spp gt 28 0 5 Anaerobes 220 19 18 lt 17 lt 4 gt 8 Erythromycin 78 ug ERYTR c S pneumoniae Streptococcus spp gt 28 27 24 lt 24 lt 1 gt 4 Campylobacter spp gt 28 27 24 lt 24 lt 1 gt 4 n Helicobacter pylori gt 28 27 24 lt 24 lt gt 4 Clindamycin 25 ug CLIND S pneumoniae Streptococcus spp 226 25 lt 2 gt 2 Anaerobes gt 26 lt 25 lt 2 gt 2 Linezolid 30 ug LINEZ S pneumoniae Streptococcus spp 228 27 25 lt 24 lt 2 gt 4 Telithromycin 15 ug TEL15 S pneumoniae I Streptococcus spp gt 21 20 17 lt 17 lt 0 5 22 Tetracyclines 80 ug TET80 Haemophilus spp gt 30 29 26 lt 26 lt 2 gt 4 S pneumoniae Streptococcus spp 228 27 24 24 24 28 N gonorrhoeae 26 1 24 Campylobacter spp 228 27 24 24 lt 4 gt 8 Chloramphenicol 60 ug CLR60 Haemophilus spp gt 34 33 30 lt 30 lt 2 gt 4 S pneumoniae Streptococcus spp 230 29 26 26 8 gt 16 N gonorrhoeae gt 32 31 28 lt 28 lt 4 gt 16 N meningitidis gt 34 lt 2 gt 4 Campylobacter spp gt 30 29 26 lt 26 lt 8 gt 16 Anaerobes gt 30 29 26 lt 26 lt 8 gt 16 Rifampicin 30 ug RIFAM Haemophilus
104. 34 33 32 lt 32 OxaS Mec pos Cefoxitin 10 ug CFO10 S aureus and S lugdunensis 222 22 OxaS Mec A pos Coag neg staph gt 27 26 22 lt 21 OxaS Mec Apos i Cefpodoxime 30 ug CFPOX 228 28 lt 1 Screen ESBL Cefotaxime 30 ug CFTAX Enterobacteriaceae gt 34 32 28 lt 28 lt 0 5 gt 1 Ceftriaxone 30 ug CETRX Enterobacteriaceae gt 34 33 28 27 lt 0 5 gt 1 Ceftazidime 30 ug CEZDI Enterobacteriaceae gt 28 27 24 lt 24 lt 2 gt 4 Pseudomonas spp Acinetobacter spp gt 26 lt 26 8 28 Ceftazidime Clav 30 10 ug CZ CL Detection of ESBL Cefepime 30 ug CFEPM Enterobacteriaceae gt 34 33 28 lt 28 lt 0 5 gt 1 Pseudomonas spp Acinetobacter spp gt 28 22 20 lt 28 lt 8 gt 8 Cefepime Clav 30 10 ug CP CL Detection of ESBL Aztreonam 30 ug AZTRM Enterobacteriaceae gt 34 33 28 lt 28 lt 0 5 gt 1 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 38 of 170 Zone diameter Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S R S R Imipenem 15 ug IMIPM 9 Enterobacteriaceae Acinetobacter spp 228 277 18 18 lt 1 gt 8 9 Pseudomonas spp gt 26 25 20 lt 20 lt 4 gt 8 Imipenem EDTA 15 750ug IM ED Detection of metallo B lactamases Meropenem 10 ug MEROP Enterobacteriaceae gt 34 33 16 16 lt 0 12 gt 8 9 Pseudomonas spp 230 29 20 20 lt 2 gt 8 Acinetobacter spp gt 30 29 20 lt 20 lt 1 gt 8 Ertapenem 10ug ETP10 Enterobacteriaceae gt 32 31
105. 4 appears highly mobile this outbreak involved 5 different gram negative genera Diagnostic laboratories in Australia and other countries must be now in high alert because early detection may limit the wide dispersal of MBL genes Acquired Metallo beta lactamases NON FERMENTERS MBL 3rd gen AZTRM IMIPM MEROP Microorganisms Genetic location Cepha MIC MIC pg ml MIC pg ml MIC pg ml IMP 1 11 gt 128 lt 8 16 gt 8 gt 8 Pseudomonas spp Chromosomal Alcaligenes spp lasmid Acinetobacter baumannii dnas IMP 12 gt 128 32 32 128 Pseudomonas putida gr IMP 13 16 gt 256 4 128 gt 64 gt 64 Pseudomonas aeruginosa Integron VIM 1 3 R SoR 2 128 1 128 Achromobacter xylosoxidans Chromosomal Pseudomonas aeruginosa 1 id Pseudomonas putida VIM 2 and 4 e integron Acinetobacter baumannii VIM 4 11 gt 256 S gt R 32 256 32 256 Pseudomonas aeruginosa SPM 1 gt 256 4 R R Pseudomonas aeruginosa Plasmid not integron GIM 1 16532 8 16 gt 8 gt 8 Pseudomonas aeruginosa Integron SIM 1 2256 128 8 16 16 Acinetobacter baumannii Integron IND 5 1 32 32 32 16 Chryseobact indologenes Chromosomal 23 MBL are not inhibited by clavulanate but are inhibited by EDTA or 2 MPA O Copyright Rosco Diagnostica A S 09 2007 2008 Chapter 16 Page 142 of 170 NEO SENSITABS Acquired Metallo beta lactamases ENTEROBACTERIACEAE MBL 3rd gen AZTRM IMIPM MEROP Microorganisms
106. 46 16 8 1 Detection of B lactam Resistance Phenotypes in Enterobacteriaceae 146 16 8 2 Detection of B lactam Resistance Phenotypes in Non fermenters esseeeee 147 17 Detection of Other Resistance Mechanisms seesesseeseseseeseee eene nennen 148 17 1 Screening of 16S rRNA Methylases to Aminoglycosides eee 148 17 2 Screening for Plasmid mediated Quinolone Resistance QnrA QnrB QnrS in Enterobacteriaceae iui d aars genser 149 17 3 Detection of Resistance Mechanisms General sss eene enne enne rennen enne 151 17 4 Intrinsic Natural Resistance 153 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Page 4 of 170 18 Sources of Error in the Diffusion Test r r r tei oerte bete te tere e e DOR tert eR RR 156 19 Limitations of Diffusion Methods Warning seeeeeeeeeeeeeeeeee eene nennen eene enne 157 20 Susceptibility Testing of Anaetob es i iere erret be eicere ape tvi ede eee 158 2 Susceptibility Testing of Yeasts ettet Re rre ht Ee gu e ree Sese euet che Re etate eR deg 161 21 1 Procedure according to CEST eee Ere Eee de REIR HR ettet Re 162 21 1 1 Interpretation Tables for Yeasts MH GMB sesseeseeeeeeeeeeneerennne ener nnne 162 2
107. 6 1000 HLR f g Kanamycin 100ug KANAM gt 28 27 23 lt 23 4 gt 16 f Kanamycin 500 ug 500 lt 16 gt 1000 HLR f Netilmicin 40 ug NETIL 228 27 23 lt 23 lt 2 gt 8 f 9 Streptomycin 100ug ST100 230 29 23 23 lt 4 gt 16 f Streptomycin 500ug 5 500 16 1000 HLR f Tobramycin 40 ug TOBRA 230 29 23 23 lt 1 gt 4 Neomycin 120ug NEOMY 226 25 20 20 lt 4 gt 8 h Ciprofloxacin 10 ug CIP10 gt 24 23 20 lt 20 lt 1 gt 2 h Norfloxacin U 10 ug NORFX gt 24 23 20 lt 20 lt 1 gt 2 h Ofloxacin 10 ug OFLOX gt 24 23 18 lt 18 lt 1 gt 4 h Levofloxacin 5 ug LEVOF gt 26 25 18 18 lt 0 5 24 h Pefloxacin 10 ug PEFLX gt 24 23 18 18 1 24 Pipemidic acid U 30 ug PIPEM 220 20 8 28 j Nalidixan U 130ug NALID 228 27 23 lt 23 lt 8 gt 8 Cinoxacin U 30 ug CINOX 222 22 8 28 Bacitracin 40U BACIT gt 22 21 18 lt 18 Clarithromycin 30 ug CLARI gt 24 23 18 lt 18 lt 0 5 22 Erythromycin 78 ug ERYTR 228 27 23 lt 23 lt 1 gt 2 Linezolid 30 ug LINEZ gt 28 27 23 lt 23 lt 2 gt 8 Fosfomycin 70 40 ug FOSFO gt 28 27 23 lt 23 lt 16 232 Chloramphenicol 60 ug CLR60 228 27 23 lt 23 lt 4 gt 8 Clindamycin 25 ug CLIND gt 26 25 20 lt 20 lt 1 gt 4 Quinupristin Dalfopristin 15 ug 15 220 19 16 16 1 24 Fucidin 100ug FUCID 232 32 lt 1 gt 1 Nitrofurantoin U 260ug NITRO 224 24 lt 32 gt 32 Polymyxins 150ug 150 gt 22 21 18 lt 18 lt 4 gt
108. 6 Other organisms 225 24 21 lt 20 lt 8 gt 32 i Clindamycin 25 ug CLIND Staphylococcus spp gt 26 25 23 lt 22 lt 0 5 gt 4 c Doxycycline 80 ug DOXYC 223 22 20 lt 19 lt 4 gt 16 Enrofloxacin 10ug ENROF 223 22 17 lt 16 0 5 24 i Erythromycin 78 ug ERYTR Streptococcus spp 228 27 24 23 lt 0 25 gt 1 Other organisms gt 26 25 23 lt 22 lt 0 5 gt Florfenicol 30 ug FFC30 Cattle gt 20 19 17 lt 16 lt 2 gt 8 Swine gt 22 21 19 lt 18 lt 2 gt 8 c Flumequine 30 ug FLUME gt 20 19 17 lt 16 lt 2 gt 4 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 22 Page 169 of 170 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S R Fucidin 100ug FUCID Plain agar 228 27 24 lt 23 lt 1 gt 4 Blood agar gt 26 25 23 lt 22 lt 1 gt 4 c Furazolidone 50 ug FURAZ gt 23 22 20 lt 19 lt 4 gt 8 Gentamicin 40 ug 40 225 24 21 lt 20 lt 2 28 Kanamycin 100ug KANAM 223 22 20 lt 19 lt 16 gt 64 c i Lincomycin 19 ug LINCO gt 26 25 23 lt 22 lt 2 gt 8 c i Lincomycin Neomycin 15460 ug LIN N gt 20 19 17 lt 16 lt 2 4 gt 5 16 Albiotic Forte i Linco spectin 15 200ug LI SP gt 20 19 17 lt 16 lt 4 32 gt 16 64 Imipenem 15 ug IMIPM 220 19 17 lt 16 lt 4 gt 16 c Marbofloxacin 5 ug 5 220 19 15 lt 14 lt 1 gt 4 b c Metronidazole 16 ug MTR16 gt 28 27 24 lt 23 lt 4 gt 8 anaerobes c Naf Pen Strep 5 2 20 ug N P S gt 2
109. 7 2006 11 Jacoby g A et al QnrB another plasmid mediated gene for quinolone resistance Antimicr Ag Chemother 50 1178 82 2006 12 Lavignes P et al qnrA in CTX M producing E coli isolates from France Antimicr Agents Chemother 50 4224 28 2006 13 Poirel L et al Prevalence and genetic analysis of plasmid mediated quinolone resistance determining 5 QnrA and QnrS in Enterobacteriaceae from a French University Hospital Antimicr Ag Chemother 50 3992 97 2006 14 Hyundoo Pai et al Association of QnrB determinants and production of ESBL s or plasmid mediated AmpC beta lactamases in clinical isolates of K pneumoniae Antimicr Ag Chemother 51 366 68 2007 O Copyright Rosco Diagnostica A S NEO SENSITABS 17 3 Detection of Resistance Mechanisms General 09 2007 2008 Page 151 of 170 Chapter 17 Mechanisms of resistance include production of inactivating enzymes alteration of drug targets and altered drug uptake or efflux Find enclosed in the table below the antibiotics recommended to detect certain resistance mechanisms 1 Antibiotic Mechanism of Neo Sensitabs Phenotype resistance Bacteria 1 Beta lactams Penicillin pH indicator Penicillin resistance Penicillinase Staphylococci Beta lactamase D T Haemophilus Gonococci Oxacillin 1 ug Resistance to all beta lactams Additional PBP Staphylococci Cefoxitin 60 ug Resistance to all beta lactams mecA Staphylococc
110. 70 Zone diameter i mm MIC pg ml NEO SENSITABS STYRKE KODE 3 2 1 0 3 2 1 0 Teicoplanin 30ug TPN30 220 19 17 16 lt 2 4 8 gt 16 5 2 18 t pr diffusion 20 GISA VISA Telithromycin 15ug TEL15 gt 22 21 17 lt 16 lt 1 2 gt 2 j Tetracyclines 80 ug TET80 gt 28 23 27 15 22 lt 14 lt 4 4 12 16 64 gt 64 a b Ticarcillin 75 ug TICAR gt 28 21 27 10 20 lt 9 lt 16 32 128 256 gt 256 b Ticarcillin 75 15ug TI CL gt 28 21 27 10 20 lt 9 lt 16 32 128 256 gt 256 Clavulanate Tigecycline 15 ug TIG15 gt 25 lt 24 lt 1 gt 2 m Tobramycin 40 ug TOBRA gt 26 23 25 1522 lt 14 lt 4 6 8 12 64 gt 64 Trimethoprim U 5 2 ug gt 22 19 21 13 18 lt 12 lt 4 6 8 12 16 gt 16 Trimethoprim 5 2 240ug TR SU gt 28 23 27 15 22 lt 14 x16 1 6 3 2 3 2 12 8 gt 12 8 Sulfa e Vancomycin 5ug VAN 5 gt 16 lt 15 lt 2 gt 4 Enterococcus spp Mueller Hinton gt 15 14 13 lt 12 lt 4 6 16 gt 16 5 2 18 t pr diffusion lt 22 VISA GISA Staphylococcus spp Enterococcus spp M H 16 VRE Bem rkninger til aflzesningsskemaet a b d e 8 svares 0 for penicillin resistente stafylokokker svares 0 for methicillin resistente stafylokokker Ved resistensbestemmelse af Pseudomonas samt St maltophilia og Burkholderia cepacia anvendes 3 230mm 2 20 29 mm 0 lt 19 mm Streptokokker enterokokker udviser intrinsisk resistens ove
111. 8 Rifampicin 30 ug RIFAM gt 28 2 lt 28 lt 1 gt 1 9 Sulphonamides 240ug SULFA 230 29 23 23 32 gt 64 Teicoplanin 30 ug TPN30 216 15 13 13 lt 2 gt 4 Tetracyclines 80 ug TET80 gt 28 27 23 lt 23 lt 1 gt 4 Doxycycline 80 ug DOXYC gt 28 27 23 lt 23 lt 1 gt 4 Trimethoprim U 5 2 ug TRIME 222 21 18 18 lt 1 gt 2 g Trimethoprim Sulfa 5 2 240 ug TR SU gt 28 27 23 lt 23 lt 1 19 gt 2 38 i n Vancomycin 5ug VAN 5 gt 15 14 13 lt 13 lt 4 gt 8 Enterococcus spp 217 16 14 14 lt 4 gt 8 i Enterococcus spp 0 5 gt 15 14 13 lt 13 lt 4 gt 4 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 34 of 170 Remarks a b b2 c d e 8 h m Amoxycillin Clavulanate Tentative zone limits until further testing has been completed For staphylococci test Oxacillin 1 ug see b Methicillin The MIC breakpoint is valid only for staphylococci and refers to MICs performed on Mueller Hinton agar at 30 Methicillin resistance in staphylococci is best detected using Mueller Hinton agar inoculum equivalent to McFarland 0 5 10 CFU ml Oxacillin 1ug and incubation at 30 35 C for full 24 hours for S aureus S gt 13 mm R 13 mm and 48 hours for coagulase negative staphylococci S 2 18 mm R 18 mm Staphylococci that are resistant to Oxacillin 1 ug methicillin should also be considered as resistant to a
112. 8 34 Cephalothin 66 ug CLOTN 24 28 Chloramphenicol 60 ug CLR60 31 35 28 34 26 31 Ciprofloxacin 10 ug CIP10 36 44 33 39 24 29 Clindamycin 25 ug CLIND 34 39 Erythromycin 78 ug ERYTR 31 37 Fucidin 100 ug FUCID 26 32 Gentamicin 40 ug GEN40 28 34 31 36 30 35 18 24 Imipenem 15 ug IMIPM 35 41 30 36 Mecillinam 33 ug MECIL 34 40 Methicillin 29 ug METHI 30 37 Nalidixan 130 ug NALID 28 33 Netilmicin 40 ug NETIL 29 34 33 37 28 33 Nitrofurantoin 260 ug NITRO 26 32 26 31 33 38 Ofloxacin 10 ug OFLOX 33 38 27 33 25 29 Oxacillin lug OXA 1 21 27 Penicillin Low 5ug PEN L 34 40 15 19 Piperacillin 100 ug PIPRA 30 36 34 40 31 36 Sulphonamides 240 ug SULFA 25 32 Teicoplanin 60 ug TEICO 20 24 Tetracyclines 80 ug TET80 33 38 34 39 22 27 Ticarcillin 75 ug TICAR 28 34 28 33 Tobramycin 40 ug TOBRA 27 31 31 36 31 35 Trimethoprim 5 2ug TRIME 26 33 22 26 39 43 Trimethoprim Sulfa 5 2 240 ug TR SU 38 43 35 41 42 47 Vancomycin 5 ug VAN 5 17 22 16 20 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 13 Page 85 of 170 13 Detection of Resistant Staphylococci Against Methicillin and Vancomycin 13 1 Staphylococcus aureus a Methicillin resistant Staphylococcus aureus MRSA and S lugdunensis Methicillin resistant staphylococci are one of the leading causes of nosocomial infections world wide Most methicillin resistance is mediated by 1
113. 8 hours incubation 9 Midtvedt amp Midtvedt 6 found a good correlation between results obtained after 8 hours and 18 24 hours incubation respectively In most cases the zones of inhibition were found to be somewhat larger at 18 24 hours than after 8 hours None of the strains observed to be susceptible after 8 hours incubation was found to be resistant after 18 24 hours Saha et al 9 using rapid but low cost susceptibility testing methods disk diffusion obtained antibiogram results 8 hours incubation identical to those obtained by conventional methods 2 Blood cultures Previous studies have shown an agreement of 94 6 or higher when comparing direct primary susceptibility testing by the diffusion method utilizing positive blood cultures as inoculum and standard diffusion susceptibility test methods 2 3 4 Coyle et al 7 used a combination of direct inoculation from blood cultures and early readings 6 hours of disk diffusion 91 of the tests with gram positive organisms and 86 gram negatives were in agreement with standard results Early reading results must be confirmed by re reading after 18 24 hours incubation or by standard susceptibility testing In clinical emergencies when the gram strain suggests that the infection is due to a single species direct susceptibility tests may be performed and the results reported as preliminary 8 Once isolated colonies become available from the initial culture a follow up standardize
114. ACITRACIN BACIT 40 units b Gram negative spectrum COLISTIN 10 ug CO 10 10 ug POLYMYXINS 150 ug Colistin CO150 150 ug K Sulphonamides and similars SULPHONAMIDES SULFA 240 ug TRIMETHOPRIM TRIME 5 2 ug TRIMETHOPRIM SULFA TR SU 5 2 240 ug L Nitrofurans NITROFURANTOIN NITRO 260 ug FURAZOLIDONE FURAZ 50 ug M Quinolone derivatives NALIDIXAN NALID 130 ug CINOXACIN CINOX 30 ug FLUMEQUINE Vet FLUME 30 ug OXOLINIC ACID OXOLI 10 ug PIPEMIDIC ACID PIPEM 30 ug CIPROFLOXACIN 10 ug CIP10 10 ug CIPROFLOXACIN 0 5 ug CIP L 0 5 ug MOXIFLOXACIN MOXIF 5ug GATIFLOXACIN GATIF 5ug ENROFLOXACIN Vet ENROF 10 ug LEVOFLOXACIN LEVOF 5ug MARBOFLOXACIN Vet MARBO 5ug NORFLOXACIN NORFX 10 ug OFLOXACIN OFLOX 10 ug PEFLOXACIN PEFLX 10 ug O Copyright Rosco Diagnostica A S NEO SENSITABS Neo Sensitabs N Others Identification code Neo Sensitabs 09 2007 2008 Chapter 3 Page 12 of 170 Diffusible amount of Antimicrobial FOSFOMYCIN G6 P FUCIDIN METRONIDAZOLE 16 ug MUPIROCIN NOVOBIOCIN 5 ug RIFAMPICIN TIAMULIN Vet PENICILLIN NOVO Vet FOSFO FUCID MTR16 MUPIR NOV05 RIFAM TIAMU PEN N O Copyright Rosco Diagnostica A S 70 40 ug 10 U 30 ug NEO SENSITABS 3 2 Antifungals Neo Sensitabs AMPHOTERICIN B CICLOPIROX CLOTRIMAZOLE ECONAZOLE FLUCONAZOLE 5 FLUOROCYTOSINE 10 ug 5 FLUOROCYTOSINE 1 ug GRISEOFULVIN ISOCONAZOLE ITRACONAZOLE KETOCONAZOLE MICONAZOLE NATAMYCIN NYSTATIN TERBI
115. BL lactamases Synergism Imipenem EDTA and or Ceftazidime DPA Metallo beta lactamases 5 Acinetobacter spp ESBL Synergism Ceftazidime Clavulanate and or Cefepime Clavulanate Synergism Ceftazidime Cefepime and Ticarcillin Clavulanate VEB 1 ESBL Synergism Cefepime and Ticarcillin Clavulanate distance 15 mm Best at 30 in the presence of Cloxacillin 500 ug 6 Achromobacter xylosoxidans VEB 1 ESBL Synergy between Ceftazidime and Clavulanate 7 Haemophilus influenzae ESBL 27 Compare Cefpodoxime and Cefpodoxime Clavulanate Zones 2 5 mm larger with the combination References 1 Sader H S et al Prevalence of important pathogens and the antimicrobial activity of parenteral drugs at numerous Medical Centers in the U S Diagn Microbiol Infect Dis 20 203 208 1994 2 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 6th Ed M2 A6 section 6 6 page 14 15 1997 3 Rasheed J K et al Evolution of extended spectrum beta lactams resistance SHV 8 in a strain of E coli during multiple episodes of bacteremia Antimicr Agents Chemother 41 647 653 1997 4 Casals J B Pringler N Detection in the routine laboratory of new plasmid mediated broadspectrum beta lactamases in Enterobacteriaceae 7th Mediterr Congr Chemother Barcelona 1990 5 Livermore D M et al Antibiotic resistance and production of extended spectrum betalactamases amongst Klebsiella spp from intensive care unit
116. BLNAR strains to these agents Cefaclor and Cephalothin may also be used to detect BLNAR strains Beta lactamase negative strains resistant to cefaclor 19 mm and or cephalothin 23 mm are BLNAR b The CLSI has not yet defined other categories than S due to the current absence of resistant strains c Clinical indications and relevant pathogens include bacterial meningitis and concurrent bacteremia in association with meningitis caused by H influenzae 2 d Fluoroquinolone resistant H influenzae are rare Non susceptible strains should be sent to a Reference Laboratory 2 Strains resistant to nalidixan should be suspected of having reduced susceptibility to quinolones Strains with decreased susceptibility to ciprofloxacin have decreased susceptibility to all quinolones 7 The current CLSI MIC breakpoints for quinolones will not always detect resistance 16 Note Only results of testing with ampicillin one of the third gen cephalosporins chloramphenicol and meropenem should be reported routinely with all blood and CSF isolates of H influenzae recovered from patients with life threating infections e g meningitis bacteremia epiglottitis and facial cellulitis 2 A Beta lactams When testing the susceptibility of Haemophilus spp it should be possible to detect 2 different types of resistance towards ampicillin 1 2 Beta lactamase producing strains plasmidic TEM or ROB 1 Beta lactamase producing strains are
117. C beta lactamase High level expression of Amp C may prevent the recognition of ESBL Cefepime is practically not affected by Amp C and consequently Cefepime Neo Sensitabs should be included as an ESBL screening agent when testing Enterobacter Serratia ect Synergism between Amox Clav and Cefepime will indicate ESBL production 11 12 13 17 19 Strains with Cefepime zones 24 mm should be suspected of ESBL production Recently Schwaber et al 32 found that the Vitek 2 Advanced Expert System identified the ESBL phenotype in only 62 5 96 isolates of Enterobacter spp and erroneously reported cephalosporin susceptibility in 28 96 Cefepime Clavulanate and cefepime Neo Sensitabs should be used in the confirmatory tests for ESBL because they are effective in detecting ESBL in strains of Klebsiella E coli etc that may produce Amp C or are hyperproducers of beta lactamase 31 Pitout et al 22 recommend the use of cefepime and piperacillin tazobactam when testing against strains with high level expression of AmpC beta lactamases E cloacae E aerogenes C freundii S marcescens Synergism between cefepime and tazobactam indicates presence of an ESBL Synergism between Ticarcillin Clavulanate and aztreonam ceftazidime cefepime permit the detection of ESBL producing strains of Ps aeruginosa SFM 2001 These strains show currently no zone around Ceftazidime Neo Sensitabs 14 ESBLs can be obscured by the chromosomal AmpC cephalosporin
118. C break points for ceftazidime and aztreonam Livermore et al 5 showed that most ESBL producers were resistant to CEZDI at 2 ug ml and AZTRM at 1 ug ml The corresponding zones with Neo Sensitabs using McFarland 0 5 inoculum are 24 mm CEZDI and CETRX and 26 mm AZTRM As a consequence Klebsiella spp E coli and Salmonella spp showing zones lt 24 mm with Ceftazidime Cefepime and or Ceftriaxone Neo Sensitabs and or lt 26 mm with Aztreonam and or Cefotaxime Neo Sensitabs should be suspected of ESBL production The CLSI has adopted practically all the same MIC breakpoints Cefpodoxime may also be used in the screening of ESBL Zones lt 20 mm should be suspected of strains with ESBL production 18 Recently the CLSI changed their Cefpodoxime screening breakpoints for ESBL from 2 2 to gt 8 ug ml 18 In a study comparing several ESBL screening methods Vercauteren et al 10 found that the double tablet synergy test using Neo Sensitabs detected 96 9 of ESBL producers while the E test ESBL Screen detected 81 2 96 De Gheldre et al showed that synergism between ceftazidime and cefepime with clavulanate Neo Sensitabs was very useful to detect ESBL in Enterobacter aerogenes from Belgian hospitals 13 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 127 of 170 Confirmatory Tests for ESBL The CLSI 9 recommends the use of Ceftazidime in combination with Clavulanic acid Ceftazidime Clavulan
119. CL zone gt 5 mm larger than AMOXY resistance to penicillin amoxycillin and ampicillin susceptible to amoxycillin clavulanate c Beta lactam resistance except cephamycins and carbapenems d Except Proteus penneri and P vulgaris Note The mentioned zone sizes are valid for McFarland 0 5 inoculum Copyright Rosco Diagnostica A S NEO SENSITABS References 09 2007 2008 Page 153 of 170 1 Clinical Microbiology and Infection Vol 2 Suppl 1 December 1996 2 Hakonen A et al Detection of decreased fluoroquinolone susceptibility in Salmonellas and validation of Nalidixic acid screening test J Clin Microbiol 37 3572 77 1999 17 4 Intrinsic Natural Resistance Antimicrobial resistance can be classified as either intrinsic or acquired Intrinsic resistance may be related to inherent or natural characteristics in a bacteria and may be used for recognition of a bacterial species and results of in vitro susceptibility testing is not relevant to report as treatment options The most relevant drug related natural resistance in a group or species is listed below BACTERIA Enterobacteriaceae NATURAL RESISTANCE Enterobacteriaceae in general Penicillinase stable penicillins Macrolides Fucidin Rifampicin Glycopeptides Enterobacteria group 2 K pneumoniae K oxytoca C diversus Esch hermannii Aminopenicillins Carboxypenicillins Enterobacteria group 3 E cloacae E aerogene
120. CLIND gt 28 27 24 lt 23 lt 1 28 Telithromycin 15 ug TEL15 221 20 17 lt 16 lt 0 5 gt 2 Linezolid 15 ug LINEZ gt 28 27 24 lt 23 lt 2 gt 8 Quinupristin Dalfopristin 15 ug SYNI5 gt 22 21 18 lt 17 lt 1 gt 4 Ofloxacin 10 ug OFLOX gt 24 23 19 18 1 gt 4 Ciprofloxacin 10 ug CIP10 gt 24 23 19 lt 18 lt 1 gt 4 Norfloxacin 10 ug NORFX gt 24 23 19 lt 18 lt 1 gt 4 Sparfloxacin 10 ug SPARF gt 24 23 19 18 lt gt 4 Gatifloxacin Sug GATIF gt 26 25 23 lt 22 lt 1 gt 4 Levofloxacin 5 ug LEVOF gt 26 25 23 lt 22 lt 1 gt 4 Moxifloxacin 5 ug MOXIF gt 26 25 23 lt 22 lt 1 gt 4 d Nalidixan 130ug NALID 228 27 24 lt 23 lt gt 32 Trimethoprim 5 2 ug TRIME gt 20 19 17 lt 16 lt 2 gt 8 Trim Sulfa 5 2 240 ug TR SU gt 28 27 24 lt 23 lt 16 gt 128 Polymyxins colistin 150ug C0150 222 gt lt 21 lt 0 5 gt 4 Pipemidic acid 30 ug PIPEM gt 22 lt 21 lt 4 gt 8 Nitrofurantoin 260 ug NITRO gt 24 lt 23 lt 64 2512 Fosfomycin 70 40 ug FOSFO 224 23 21 lt 20 lt 32 gt 64 Mupirocin 10 ug MUPIR gt 18 lt 17 lt 4 gt 8 Rifampicin 30 ug RIFAM 226 25 23 22 1 24 Remarks Note This table is valid for rapidly growing bacteria Fastidious strains and problem organisms have special interpretation tables haemophilus neisseria pneumococci streptococci etc or need special methodologies See chapter 15 in the User s Guide a Staphylococci resistant to Penicillin Low beta lactamase produc
121. CMY 19 gt 128 gt 128 16 4 0 25 lt 0 06 pneumoniae 8 21 gt 64 64 32 0 5 0 25 0 06 E coli O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 136 of 170 Beta lactamases Cefoxitin Ceftazidime Aztreonam Cefepime Imipenem Meropenem Microorganisms CFE 1 R 64 8 0 25 0 25 E coli inducible DHA 1 128 512 8 64 1 16 lt 0 125 2 lt 0 125 E coli K pneumoniae Salmonella 0 5 P mirabilis inducible DHA 2 16 8 2 0 03 0 25 inducible 1 128 8 1 1 0 25 lt 0 03 E coli pneumoniae FOX 2 256 32 2 0 13 0 5 0 03 E coli FOX 3 64 16 1 lt 0 06 0 12 E coli oxytoca FOX 4 gt 512 gt 128 64 2 0 5 0 12 coli FOX 5 512 128 8 16 0 5 0 5 pneumoniae E coli 7 E coli pneumoniae cloacae LAT 1 64 256 gt 128 64 1 0 25 2 0 06 K pneumoniae LAT 2 CMY 2 256 gt 256 64 256 E coli K pneumoniae E aerogenes LAT 3 CMY 6 256 128 64 0 5 0 25 0 06 E coli LAT 4 CMY 1 64 256 8 256 8 128 0 125 1 0 25 0 125 E coli MIR 1 gt 256 128 128 1 1 0 125 E cloacae pneumoniae MOX 1 R 165 16 0 5 pneumoniae MOX 2 gt 128 4 256 0 25 4 lt 0 125 K pneumoniae Gupta et al 3 describes isolation of multiresistant Salmonella with plasmid mediated AmpC beta lactamase from cattle and humans in the USA
122. Ceftazidime Clavulanate Ceftazidime S Synergy is currently observed with third gen cephalosporins and Clavulanate as well as with Cefepime Clavulanate Synergism Ceftazidime Clavulanate Ceftazidime I R 3a Enterobacter spp Serratia spp Providencia rettgeri Citrobacter freundii ESBL ESBL 16S rRNA methylases ESBL Metallo beta lactamases 3b Morganella morganii ESBL Synergism Cefepime and Clavulanate Amoxycyllin Clavulanate and or Ceftazidime and Clavulanate Synergism Cefepime and Clavulanate and or Ceftazidime Clavulanate No zone with Amicacin Gentamicin Tobramycin Neo Sensitabs Synergism Aztreonam or Cefepime Clavulanate ESBL Synergism Imipenem EDTA and or Ceftazidime DPA Metallo beta lactamases Synergism Cefepime and Tazobactam Piperazillin Tazobactam Synergism Sulbactam Ampicillin Sulbactam and Ceftazidime or Cefotaxime Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 130 of 170 4 Pseudomonas aeruginosa ESBL Ceftazidime no zone of inhibition Ticarcillin resistant Synergism Ceftazidime Clavulanate and or Cefepime Clavulanate Synergism Aztreonam Ceftazidime or Cefepime with Ticarcillin Clavulanate VEB 1 ESBL Synergism between Imipenem and Ceftazidime or Cefepime in the presence of Cloxacillin 500 ug Synergism Ceftazidime Clavulanate and or Cefepime Clavulanate ESBL Metallo beta Synergism Aztreonam or Cefepime Clavulanate ES
123. Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 9 Page 25 of 170 9 Measuring the Inhibition Zones After incubation at 35 overnight the diameter of the zones of inhibition tablets included are measured in millimetres using sliding calipers a ruler or a template prepared for this purpose held on the back of the petri dish illuminated with reflected light If blood has been added to the agar base the zones should be measured from the surface illuminated with the cover removed 1 The diameters of the zones of complete inhibition as judged by the unaided eye are measured including the diameter of the tablet disk 1 Therefore no zone will be registered as 9 mm Faint growth of tiny colonies which can be detected only with a magnifying lens at the edge of the zone of inhibited growth is ignored However discrete colonies growing within a clear zone of inhibition should be subcultured reidentified and retested resistant mutants or mixed culture When the zone limits are clear and sharply outlined ideal readings can be made 2 Strains of Proteus mirabilis and Proteus vulgaris may swarm into areas of inhibited growth The veil of swarming growth is ignored 3 With Chloramphenicol Erythromycin and Tetracyclines bacteriostatic agents the zones of inhibition will contain a gradient of growth Zone diameters should be read halfway between the start of inhibition and complete inhibition CLSI 2006
124. ELI15 27 33 0 008 0 016 Tetracyclines 10 ug TET10 20 26 0 25 Tetracyclines 80 ug TET80 28 35 0 25 Tigecycline 15 ug TIG15 23 29 0 03 0 06 Trimethoprim Sulfa 5 2 240 ug TR SU 32 38 0 25 4 75 Vancomycin 5ug VAN 5 19 26 0 25 Deterioration in Oxacillin paper disks content may result in false resistance when testing pneumococci Therefore the recommends 8 testing Oxacillin lug paper disks against both S pneumoniae ATCC 49619 penicillin intermediate and Staph aureus ATCC 25923 penicillin sensitive acceptable zone diameter 18 24 mm This problem is nonexistent when using Oxacillin lug Neo Sensitabs due to its high stability References 1 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 Section 6 3 page 12 13 2003 2 Barry A L Fuchs P Surrogate disks for predicting Cefotaxime and Ceftriaxone susceptibility of 5 pneumoniae J Clin Microbiol 34 2609 2612 1996 3 Doern et al Antimicrobial resistance of S pneumoniae recovered from outpatients in the U S during the winter months of 1994 to 1995 result of a 30 Center National Surveillance Study Antimicrob Agents Chemother 40 1208 1213 1996 4 Paris M M et al Management of meningitis caused by penicillin resistant pneumoniae Antimicrob Agents Chemother 39 2171 2175 1995 5 Barry A L et al In vitro activity of Cefotaxime Ceftriaxone Ceftazidime Cefpirome and Penicillin against S pneumoniae iso
125. Forsythe S B Selection of a reference lot of Mueller Hinton Agar J Clin Microbiol 24 1 6 1986 3 Hindler J A Inderlied C B Effect of source of Mueller Hinton Agar and resistence frequency on the detection of methicillin resistant Staph aureus J Clin Microbiol 21 205 210 1985 4 Casals J B Effects of medium on the results of antimicrobial susceptibility testing Chemotherapy 2 41 46 Proc 9th International Congress Chemotherapy London 1975 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 5 Page 15 of 170 5 Primary vs Pure Culture Susceptibility Testing Early Reading of the Antibiogramme Direct or primary susceptibility tests in which tablets or disks are applied to plates inoculated from the specimen are widely used in Denmark and the U K but their reliability is often questioned by microbiologists in other countries The advantages of these tests are a speed as results may be available on the following day b identification of small numbers of resistant organisms in a predominantly susceptible population and c their value as selective media in mixed cultures 1 1 Urine Primary susceptibility tests of urine specimen are particularly successful 5 In urgent clinical situations physicians may request more rapid information to guide their selection of antimicrobial agents Therefore it is common practice in some laboratories to report presumptive susceptibilities after 6
126. Genetic location Cepha MIC MIC pg ml MIC g ml MIC pg ml IMP 1 232 0 5 2 0 5 E coli IMP 1 232 0 5 OR 4 128 4 128 S marcescens K pneumoniae K oxytoca E cloacae Integron E aerogenes Cit freundii P rettgeri M morganii IMP 3 64 0 5 1 Shigella flexneri IMP 4 256 3 6 Citrobacter youngae Plasmid IMP 6 128 0 25 2 8 64 E coli IMP 6 gt 128 128 32 gt 128 Serratia marcescens IMP 8 R SoR 0 5 8 0 25 4 Enterobacter cloacae Klebsiella pneumoniae VIM 1 R 8 8 2 E coli P mirabilis integron Plasmid C koseri VIM 1 16 128 5 gt 1 64 1 32 Klebsiella pneumoniae E cloacae Plasmid integron VIM 2 232 SoR gt 1 0 5 gt gt 2 Citrobacter freundii E cloacae Plasmid VIM 2 gt 128 32 16 64 8 64 Serratia marcescens P rettgerii Integron VIM 2 8 16 4 0 1 S Klebsiella oxytoca Plasmid integron VIM 4 gt 32 45R 2 4 0 5 1 K pneumoniae E cloacae Plasmid VIM 12 gt 128 16 8 4 K pneumoniae Plasmid 16 VIM 12 gt 32 1 1 0 25 E coli Plasmid 22 VIM 2 E coli Integron GES7 MBL are not inhibited by clavulanate but are inhibited by EDTA or 2 MPA O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 143 of 170 Procedure for MBL detection Some resistance profiles may suggest MBL production for example a Pseudomonas aeruginosa Pseudomonas spp and Acinetobacter spp All isolates non susceptible to carbapenems and resistant to eith
127. Gonococci Nali zone 28 mm and 4 Vibrio cholerae Nali zone 25 mm With staphylococci the interpretation is valid for amikacin and isepamicin Do not take into account the presence of colonies inside the zone of inhibition Resistant strains show homogeneous resistance For detection of VISA GISA hVISA strains see chapter in User s Guide on detection of staphylococci with decreased susceptibility to vancomycin page 86 Zones tentative for 1 year O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 69 of 170 VI b Haemophilus spp S pneumoniae Streptococcus spp N gonorrhoeae N meningitidis Campylobacter spp and anaerobes Interpretation according to the MIC break points recommended by the Comit de l Antibiogramme de la Societ Francaise de Microbiologie Jan 2006 Inoculum media and incubation conditions acc to SFM 2006 Zone diameter Concentrations in mm critiques NEO SENSITABS POTENCY CODE S R S R Ampicillin 33 ug AMP33 e Haemophilus spp 32 21 Campylobacter spp 226 25 22 22 lt 4 gt 16 e Ampicillin 2 5 ug AMP L Haemophilus spp lt 22 gt 1 Penicillin Low 5ug PEN L N gonorrhoeae 234 33 23 23 lt 0 06 21 Oxacillin 1 ug OXA 1 pneumoniae gt 20 lt 20 lt 0 06 pen MIC test d Streptococcus spp gt 15 lt 15 lt 0 12 pen test l N gonorrhoeae gt 14 lt 14 lt 0 06 pen MIC test 9 N meningitidis gt 12
128. IFAMPICIN 30 ug RIFAM 32 40 SULPHONAMIDES 240 ug SULFA 18 25 23 33 TEICOPLANIN 30 ug TPN30 15 21 TELAVANCIN 30 ug TELAV 16 20 TELITHROMYCIN 15 ug TEL15 24 30 TEMOCILLIN 30 ug TEMOC 18 24 TETRACYCLINES 80 ug TET80 26 32 26 34 15 21 TICARCILLIN 75 ug TICAR 24 30 22 28 TICARCILLIN 75 15ug TICL 25 31 24 30 CLAVULANATE TIGECYCLINE 15 ug TIG15 20 27 20 25 9 13 TOBRAMYCIN 40 ug TOBRA 25 30 26 32 28 34 TRIMETHOPRIM 5 2 ug TRIME 22 29 19 25 21 27 TRIMETHOPRIM 5 2 240 ug TR SU 30 38 29 38 26 32 SULFA VANCOMYCIN 5ug VAN 5 16 21 15 18 Zone diameter in mm E coli NEO SENSITABS CODE ATCC 35218 AMOXYCILLIN CLAVULANATE 30 15 ug AM CL 21 26 AMPICILLIN SULBACTAM 30 30 ug AM SU 21 26 PIPERACILLIN TAZOBACTAM 100 10 ug PI TZ 26 31 TICARCILLIN CLAVULANATE 75 15 ug TI CL 25 31 TEMOCILLIN 30 ug TEMOC 20 26 Frequency of Q C Testing according to CLSI 1 Daily Testing When testing is performed daily for each antimicrobial organism combination 1 out of 20 consecutive results may be out of the acceptable range More than 1 out of control result in 20 consecutive tests requires corrective action 2 Weekly Testing Satisfactory performance should be demonstrated for conversion from daily to weekly Q C testing Test all control strains for 20 consecutive days No more than 3 out of the 30 zone diameters for each antimicrobial organism combination may be out of the acceptable range f any of t
129. LOX gt 32 31 25 lt 24 lt 0 25 22 Oxacillin 1 ug 1 gt 12 lt 0 06 screening penicillin Oxacillin 5 pg 5 gt 18 5 lt 0 06 screening penicillin c Penicillin Low 5 ug PEN L gt 44 43 26 lt 25 lt 0 06 232 Spectinomycin 200ug SPECT 223 22 20 lt 19 lt 32 gt 128 Tetracyclines 80 ug TET80 gt 42 41 34 lt 33 lt 0 25 gt 2 e Tetracyclines 10ug TET10 gt 32 31 26 25 lt 0 25 22 Trimethoprim Sulfa 5 2 240 ug TR SU gt 32 31 27 lt 26 lt 0 5 9 5 22 38 a CLSI has not yet defined other categories than S due to current absence of resistant strains b Ciprofloxacin resistant gonococci should be presumed to be resistant to other quinolones ofloxacin moxifloxacin pefloxacin norfloxacin and gatifloxacin 2 C A positive beta lactamase test predicts resistance to penicillin ampicillin amoxycillin ticarcillin and piperacillin O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 105 of 170 d Nalidixan is useful to detect strains with reduced susceptibility to quinolones If the zone is lt 28 mm control MIC s of CIPRO OFLOX etc Two strains of gonococci NALI susceptible and CIPRO resistant were recently detected in the UK 7 Test both NALI and CIPRO e Gonococci with zonediameters lt 26 mm Tetracyclines 80 ug Neo S or lt 18 mm Tetracyclines 10 ug Neo S usually indicate a plasmid mediated tetracycl
130. Livermore DM et al Interpretative reading recognizing the unusual and inferring resistance mechanisms from resistance phenotypes J Antimicrob Chemother 2001 48 Suppl 1 87 102 2 Comminique January 2005 Societ Francaise de Microbiologie CA SFM O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 18 Page 156 of 170 18 Sources of Error in the Diffusion Test The following common sources of error should be considered when a zone diameter is outside the indicated control limits with ATCC Control Strains e Variability in the agar test medium lot variation e pH value of medium too high or too low If the pH value is too low Zones of inhibition will be smaller with aminoglycosides macrolides and quinolones If the pH value is too high Zones of inhibition will be too small for penicillins and tetracyclines Ca and Mg content of media content too low Zone too small with daptomycin Zone too large with P aeruginosa and aminoglycosides content too high Zone too small with P aeruginosa and aminoglycosides Zone too small with Tetracyclines e Inoculum used too heavy or too light Inoculum for the test prepared from a plate incubated for too long time more than 24 hours old ncorrect incubation temperature and or atmosphere used e Contamination of the control strain e Deteriorated McFarland standard or failure in the vortex procedure of the BaS0 turbidity standard e Loss of disk pot
131. NAFINE VORICONAZOLE CASPOFUNGIN investigational drug POSACONAZOLE investigational drug Identification code Neo Sensitabs AMPHO CICLO CLOTR ECONZ FLUCZ FLU10 FLU 1 GRISE ISOCN ITRAC KETOC MICON NATAM NYSTA TERBI VOR 1 CASP5 POSAC O Copyright Rosco Diagnostica A S Diffusible amount of Antimicrobial 10 ug 50 ug 10 ug 10 ug 25 ug 09 2007 2008 Chapter 3 Page 13 of 170 NEO SENSITABS 09 2007 2008 Chapter 4 Page 14 of 170 4 Susceptibility Test Media In order to achieve satisfactory results a proper medium for susceptibility testing must be used It should fulfil at least two requirements The composition of the medium should give sufficiently good growth conditions for the strains to be tested The medium must not contain material interfering with the test itself or with any antimicrobial contained in Neo Sensitabs The ideal medium for susceptibility testing has not been found yet and this is one of the reasons why different media are used in different countries Mueller Hinton agar is recommended by the FDA and the CLSI in the U S 1 and is routinely used in several European countries Variations in Mueller Hinton agar from different manufacturers were first recognized with tetracyclines and amino glycosides Variation in Mg and Ca will affect results of aminoglycoside and tetracycline tests with Ps aeruginosa Excess zinc ions may reduce zone sizes of carbapenems Excessive cation con
132. NEO SENSITABS 09 2007 2008 Chapter 16 Page 146 of 170 16 8 Detection of B lactam Resistance Phenotypes 16 8 1 Detection of B lactam Resistance Phenotypes in Enterobacteriaceae AMC CLOT CAZ FEP FOX CAZ CLAV IMI IMI CLOXA Comments FEP CLAV EDTA 500 CAZ DPA Boronic 1 E coli P mirabilis Salmonella spp Shigella spp Klebsiella spp C diversus Penicillinase low IR SI S S S S S E coli Penicillinase high R R R S S S S Cephalosporinase low SI SA YR S S S S AmpC high plasmid R R R I R S I R S synergy FOX antagonism CAZ with CAZ or FEP CLAV indicates inducible AmpC ESBL R R R R SR R Synergy S synergy CAZ CLAV IRT R R SI S gt CAZ S zone Oxacillinase RIR SI 8 SCAZ S zone Penicillinase R R R SA S UR S Cephalosporinase Chromosomal K 1 high R UIR R S SIT S false synergy S Klebsiella oxytoca FEP CLAV ESBL R V R VR V R S R synergy S Metallo B lactamase R R R IRIIR R S R Synergy AZT SF Carbapenemase class A R SR R R UR syn CLAV 2 Enterobacter spp C freundii Penicillinase R R R S S R S AmpC derepressed R R R R S R S syn FOX syn CAZ CLAV AmpC ESBL R R R R SR R Synergy S syn FOX syn CAZ CLAV ESBL for E coli etc Metallo B lactamase and Carbapenemases 3 morganii Providencia spp Serratia spp Penicillinase R R R S S R S A
133. NaCl to the medium provide larger inhibition zones making the test more sensitive 2 Synergy test between Imipenem and Imipenem EDTA Neo Sensitabs gt 6 mm larger zone indicates synergy 3 Induction test between Cefoxitin Imipenem Neo Sensitabs and Ceftazidime Dipicolinic acid and Ceftazidime distance 15 mm from edge to edge 4 Carbapenem inactivation assay carbapenemase test page 140 Results Carbapenemases type A will show in most cases a synergism between Imipenem and Clavulanate KPC GES Besides no synergism between Imipenem and EDTA and or Ceftazidime Dipicolinic acid metallo B lactamase negative Carbapenemase test is positive Inducible carbapenemases Sme 1 to Sme 3 NmcA IMI 1 and IMI 2 and OXA 60 are induced by cefoxitin and imipenem and will show antagonism between Cefoxitin Imipenem and Ceftazidime Carbapenem inactivation assay Carbapenemase test 2 In order to determine whether resistance to carbapenems is caused by a carbapenemase a tablet disk diffusion bioassay may be performed A M H agar plate is inoculated with the susceptible strain E coli ATCC 25922 as for disk diffusion Mc Farland 0 5 inoculum Then Imipenem and or Meropenem Neo Sensitabs are applied onto the plate A suspension of the microorganism to be tested for the presence of carbapenemase is adjusted to McF 0 5 Standard and a loop is used to make a 15 20 mm streak on each side of the Imipenem and or Meropenem Neo Sensitabs star
134. O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 h Page 60 of 170 Til p visning af metallo beta lactamaser og andre carbapenemaser se kapitel 16 6 Carbapenemases h2 Pr diffusionsmetode Er beskrevet i detaljer pa side 18 Kan med fordel anvendes til p visning af Colistin resistente P aeruginosa A baumannii Daptomycinresistente staphylococci enterococci samt pavisning af VISA GISA stammer Non fermenters i 12 k m n 0 q 5 Ved resistensbestemmelse af Pseudomonas samt St maltophilia og Burkholderia cepacia anvendes 230mm I 29 20 mm R lt 19mm P aeruginosa St maltophilia samt Proteus spp b r rapporteres R mod Tetracyclines Neo Sensitabs uanset zonest rrelse Ertapenem m ikke testes over for P aeruginosa p Dansk Blod Agar idet det kan resultere i falsk f lsomme resultater Til resistensbestemmelse af Acinetobacter spp St maltophilia og B cepacia se kapitel 15 10 Staphylococcer Staphylococcer og beta lactam testning Til p visning af beta lactamase anvendes Penicillin Low Neo Sensitabs I tvivlstilf lde kan resultaterne kontrolleres med Mecillinam Neo Sensitabs Til p visning af methicillin oxacillin resistens anvendes surrogattestning med Cefoxitin 60 ug eller 10 ug samt Oxacillin 1 ug Neo Sensitabs Med Cefoxitin 60 ug g lder f lgende S aureus og S lugdunensis 230mm I 29 28 og R lt 28 mm Koagulase neg
135. O SENSITABS 09 2007 2008 Chapter 15 Page 111 of 170 P rez Trallero 11 mentions that using fluoroquinolones to treat a strain that had an exising but unapparent first step mutation in the gyr gene probably favoured the development of high level resistance to quinolones observed in the later isolates Current CLSI breakpoints for fluoroquinolones on pneumoniae define many isolates as susceptible even though they harbour QRDR mutations The proposed microbiological resistance breakpoints 13 is the MIC zone in which gt 50 of the isolates carry QRDR mutations 14 19 20 21 Microbiological resistance break points for fluoroquinolones 13 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S Levofloxacin 5ug LEVOF 2 lt 20 gt 1 Moxifloxacin 5ug MOXIF lt 25 gt 0 12 Gatifloxacin 5ug GATIF lt 24 gt 0 25 Norfloxacin 10 ug NORFL lt 12 gt 8 Streptococcus pneumoniae Mueller Hinton 5 blood Inoculum McFarland 0 5 Incubation in 5 7 CO Break points according to CLSI M2 A8 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Azithromycin 30 ug AZITR 222 21 19 lt 18 lt 0 5 gt 2 c Ceftizoxime 30 ug CEZOX gt 30 29 26 lt 25 lt 0 5 cefotaxime ceftriaxone CFTAX etc cefepime cefpirome Chloramphenicol 60 ug CLR60 gt 28 lt 27 lt 4 gt 8 Chloramphenicol 10 ug CLR10 gt 16 lt 15 lt 4 gt 8 Clarith
136. OXT gt 28 27 23 lt 23 lt 4 gt 16 S aureus and S lugdunensis gt 30 29 28 lt 28 OxaS A pos Coag neg staph gt 34 33 32 lt 32 OxaS mec A pos b2 Cefoxitin 10 ug CFO10 S aureus and S lugdunensis 222 22 OxaS A pos Coag neg staph gt 27 26 22 lt 22 OxaS pos Cefsulodin 30ug CFSUL 226 25 20 20 lt 4 gt 16 m Ceftazidime 30 ug CEZDI 226 25 20 20 lt 4 gt 16 e m Ceftriaxone 30 ug CETRX gt 26 25 20 lt 20 lt 4 gt 16 Cefuroxime parenteral 60 ug CEFUR 228 27 23 23 lt 4 gt 16 d Cefuroxime oral 60 ug CEFUR 234 33 23 23 lt 1 gt 16 d Cephalexin 30 ug CFLEX 226 25 20 20 lt 4 gt 16 Cephalotin 66 ug CLOTN gt 28 27 23 lt 23 lt 4 gt 16 d Cephadrine 60 ug CFRAD 226 25 20 20 lt 4 gt 16 Cefixime 30 ug CFFIX gt 30 29 26 lt 26 lt 1 gt 2 Cefepime 30 ug CFEPM gt 26 25 20 lt 20 lt 4 gt 16 Cefpodoxime 30 ug CFPOX gt 32 31 30 lt 30 0 25 gt 0 5 e m Aztreonam 30 ug AZTRM 226 25 22 22 lt 4 gt 8 Imipenem 15 ug IMIPM gt 28 27 20 lt 20 lt 2 gt 8 10 ug MEROP 226 25 20 20 lt 2 gt 8 O Copyright Rosco Diagnostica A S NEO SENSITABS Zone diameter 09 2007 2008 Page 33 of 170 Chapter 10 Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE R S R f Amikacin 40 ug AMIKA 226 25 22 22 lt 4 gt 16 f Gentamicin 40 ug GEN40 gt 30 29 23 lt 23 lt 1 gt 4 f Gentamicin 250ug 250 1
137. S 1 the appropriate inoculum should yield a dense but not completely confluent growth It is prepared by diluting an overnight broth culture of the organism in broth Suitable dilutions of fully grown broth cultures of Enterobacteriaceae and most gram negative rods are usually about 107 1 10 000 while for staphylococci and enterococci a dilution of 10 1 1 000 is usually appropriate 2 The inoculum can also be prepared from isolated colonies on agar Suspend 2 to 3 colonies into 10 ml physiological saline and thereafter dilute 0 1 ml 10 ml for Enterobacteriaceae and other gram negative rods I ml 10 ml for staphylococci and enterococci Take 1 2 drops 9 cm plate or 3 to 4 drops 14 cm plate of the final suspension and apply onto the agar surface and distribute with a bent glassrod or Drigalski spatula The open plate is then dried at 35 37 C for 10 15 min before the Neo Sensitabs are placed onto the agar surface When testing slower growing microorganisms the dilution must be individualized to give semi confluent growth The method of inoculation may vary but it must be such that it gives uniform seeding of the plate and the semi confluent growth desired 6 1 2 Inoculum according to CLSI Kirby Bauer When using the technique of Kirby Bauer the inoculum is standardized according to the method described by the CLSI 2 which normally results in more dense growth and therefore smaller inhibition zones than for semiconfluent growth
138. S pneumoniae by using genetics instead of pharmacokinetics pharmacodynamics Antimicr Ag Chemother 48 3630 5 2004 Lim S et al Antimicrobial susceptibility breakpoints and first step parC mutations in S pneumoniae redifining fluoroquinolone resistance Emerg Infect Dis 9 833 7 2003 Wolter N et al Novel mechanism of resistance to oxazolidinones macrolides and chloramphenicol in ribosomal protein L4 of the pneumococcus Antimicr Ag Chemother 49 3554 7 2005 Faccone D et al Emergence of S pneumoniae clinical isolate highly resistant to telithromycin and fluoroquinoles J Clin Microbiol 43 5800 03 2005 Ribes S et al Evaluation of fosfomycin alone and in combination with ceftriaxone or vancomycin in an experimental model of meningitis caused by 2 strains of cephalosporin resistant S pneumonaie JAC 57 931 6 2006 Rantala M et al S pneumoniae isolates resistant to telithromycin Antimicrob Ag Chemother 50 1855 8 2006 Pletz M W R et al Prevalence of first step mutants among levofloxacin susceptible invasive isolates of S pneumoniae in the U S Antimicr Ag Chemother 50 1561 3 2006 Schurek K W et al Call for the international adoption of microbiological break points for fluoroquinolones and S pneumoniae Intl J Antimicr Ag 28 266 69 2006 Varon E et al Non molecular test for detection of low level resistance to fluoroquinolones in S pneumoniae Antimicr Ag Chermother 50 572 579 2006
139. SBL producing coli K pneumoniae P mirabilis except Temocillin Routine reporting of results from strains isolated from the CSF could be dangerously misleading for patient care in the following cases e Agents administered orally only e Ist and 2nd generation cephalosporins except Cefuroxime sodium e Clindamycin e Macrolides e Tetracyclines e Fluoroquinolones Some antimicrobials are associated with the emergence of resistance during prolonged therapy As a consequence isolates initially susceptible may become resistant within a few days after initiation of treatment This occurs most frequently in e Enterobacter Citrobacter and Serratia spp with third generation cephalosporins and aztreonam e Pseudomonas aeruginosa with most antimicrobials e Staphylococci with quinolones Repeat testing in 3 5 days should be considered Essentially all isolates of Enterobacter aerogenes E cloacae Citrobacter freundii Providencia spp Proteus spp except P mirabilis Serratia marcescens Pseudomonas aeruginosa possess the genes for Group I beta lactamase production Therefore there is no useful information from demonstrating in vitro induction of the enzyme Laboratories should focus on repeat testing every 3 4 days or earlier depending on the patient s condition of isolates repeatedly recovered from infected patients during therapy to detect selection of clones that constitutively produce Group I beta lactamase 1 R
140. Schwaben J J et al Utility of the Vitek 2 Advanced Expert System for identification of ESBL producton in Enterobacter spp J Clin Microbio 44 241 3 2006 Muratani T et al Emergence and prevalence of beta lactamase producing K pneumoniae resistant to cephems in Japan Intl J Antimicr Ag 27 491 9 2006 Wonkeun Song et al Detection of ESBLs by using Boronic acid as an AmpC f lactamase inhibitor in clinical isolates of pneumoniae and E coli J Clin Microbiol 45 1180 84 2007 Wonkeun Song et al Clonal spread of both oxyimino cephalosprin and cefoxitin resistant K pneumoniae isolates coproducing SHV 2a and DHA 1 B lactamase at a bruns intensive unit Int J Antimicrob Agents 28 520 24 2006 Naiemi N et al Widely destributed and predominant CTX M ESBLs in Amsterdam The Netherlands J Clin Microbiol 44 3012 3014 2006 16 2 ESBL Quality Control Klebsiella pneumoniae ATCC 700603 Zone diameter NEO SENSITABS POTENCY CODE in mm MIC ug ml 15 Aztreonam 30ug AZTRM 11 19 64 Cefepime 30ug CFEPM 21 26 1 Cefepime Clavulanate 30 10ug CP CL 28 33 lt 0 12 Cefotaxime 30 ug CFTAX 20 26 8 Cefpodoxime 30 ug CFPOX 14 21 16 Ceftazidime 30 ug CEZDI 11 19 32 Ceftazidime Clavulanate 30 10ug CZ CL 24 31 1 Ceftriaxone 30 ug CETRX 16 24 16 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 132 of 170 16 3 Inducible Cephalosporinases or AmpC Beta lactamases Inducible cephalosp
141. T Hafnia alvei R R R R R R T R R T T R R Enterobacteriaceae with ESBL no inducible R TP R R R R R R R T T TP T B lactamases Enterobacteriaceae with inducible B lactamases R R R R R R R R R T T R R and ESBL Aeromonas with A2 R R T T T T T T T R R R R most sobria Aeromonas with A 1 and A 2 Ps aeruginosa Burkholderia spp S maltophilia R R R R R T R R R R R T R A baumannii R T R R R T T R R T T T T AMG CL Amoxyeillin Clavulanate AMP Ampicillin AZTRM Aztreonam CEFUR Cefuroxime CETRX Ceftriaxone CEZDI Ceftazidime CFEPM Cefepime CFOXT Cefoxitin CFTAX Cefotaxime IMIPM Imipenem MEROP Meropenem PI TZ Piperacillin Tazobactam TI CL Ticarcillin Clavulanate Al Inducible cephalosporinase the enzyme is usually found in A hydrophila and A caviae These species are considered resistant to cephalosporins and cephamycins A2 Penicillinase carbapenemase that hydrolyses imipenem and meropenem The expression may be heterogeneous a R the microorganism is resistant and may possess a resistance mechanism not always detected by the diffusion method T these antimicrobials may be used for testing b Test isolates from urine only Isolates from other sites are considered resistant c Test also for Cefazolin A caviae does not posses a carbapenemase A2 and can be tested against imipenem and meropenem d Use Ampicillin Sulbactam e K oxytoca producing a K 1 enzyme are s
142. a A S NEO SENSITABS 09 2007 2008 Chapter 8 Page 24 of 170 Use of the combination of antibiotics in disks was condemned for the reason stated in the W H O report Diffusion tests employing either a single disk containing two antibiotics at two disks superimposed are useless since it has been shown that the effect obtained is only that produced by the antibiotic giving the wider zone of inhibition when acting alone The combination of trimethoprim and sulphonamides Trimethoprim Sulfa Neo Sensitabs as well as Amoxycillin Clavulanate Ampicillin Sulbactam Ticarcillin Clavulanate and Piperacillin Tazobactam are exceptions because true potentiation or synergism between the two antimicrobials is to be expected in several cases Antifungals Neo Sensitabs representing the polyenes azoles imidazoles and caspofungin fluorcytosine for susceptibility testing of yeast are described in this booklet References 1 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 January 2003 2 Barry A L et al Reassessment of the Class concept of disk susceptibility testing Am J Clin Path 70 909 913 1978 3 W H O Technical report series 210 1961 Standardisation of methods for conducting microbic sensitivity tests Second report of the Expert Committee on Antibiotics 4 Barry A L et al Aerobic and anaerobic susceptibility tests with three tetracyclines Am J Clin Path 70 821 825 1978 O
143. aconazole 24 34 25 36 23 31 Voriconazole 31 42 25 36 23 31 21 2 Procedure using Shadomy modified agar This technique developed by Rosco approx 10 years ago 8 uses the following procedure 1 Media The modified Shadomy Agar contains Yeast Nitrogen Base glucose and asparagine and is buffered with phosphate to pH 7 0 Chloramphenicol has been added to avoid bacterial contamination 2 Inoculum and inoculation technique Prepare a McFarland 0 5 suspension and ilute further 1 1 with saline For C krusei use a McFarland 0 5 suspension diluted 1 10 with saline For Cryptococcus spp use an inoculum equivalent to McFarland 1 0 undiluted 0 5 ml 9 cm plate or 1 ml 14 cm plate of the above suspensionis poured onto the agar surface flooding and excess liquid is removed immediately with a pipette Thereafter the open plate is dried at 35 C for 10 minutes before applying the Neo Sensitabs disks on the agar surface 3 Incubation time Reading of the test should be made as soon as possible i e for yeasts after not more than 18 24 hours A longer incubation time may result in false resistance against imidazoles azoles Plates should always be examined after overnight incubation and if inhibition zones are visible they must be measured If growth is not yet visible with particular strains the plates may be reincubated for up to 24 hours more For Cryptococcus spp incubate at 30 C for 42 48 hours 21 2 1 Interpretation Table
144. actamase produktion Mecillinam zoner gt 22 mm indicerer penicillin f lsomhed hos S aureus 2 q Til p visning af hVISA VISA stammer se afsnittet i User s Guide Detection of strains with decreased susceptibility to Vancomycin side 86 r Ertapenem m ikke testes over for P aeruginosa p Dansk Blod Agar idet det kan resultere i falsk f lsomme resultater s 2 18 timers pr diffusionsteknikken er beskrevet p side 18 Referencer 1 Dragsted U B et al Relapse of multiresistant Salmonella Tiphi after combined therapy with ciprofloxacin and ceftriaxone Eur Soc Clin Microbiol Infect Dis 6 167 8 2000 2 BruunB Gahrn Hansen B Mecillinam susceptibility as an indicator of betalactamase production in Staphylococcus aureus Clin Microbiol amp Infect 8 122 124 2002 3 Skov R et al Tentative interpretative zone diameters for fusidic acid Neo Sensitabs on Mueller Hinton Agar and three blood containing media Int J Antimicrob Ag 22 502 7 2003 O Copyright Rosco Diagnostica A S 09 2007 2008 Page 57 of 170 Chapter 10 NEO SENSITABS 10 5 1 Interpretations According to MIC Breakpoints of the Danish Reference Group for Susceptibility Testing Vc Afl sningsskema for hurtigvoksende bakterier Breakpoints ifolge DSKM Referencegruppe for antibiotika resistens DK Substrat DBA Inoculum Semikonfluerende vaekst Zone diameter Break points imm MIC pg ml NEO SENSITABS STYRKE KODE S l
145. ains show inhibition zones gt 5 mm larger with Imipenem EDTA compared to Imipenem and or synergism between Meropenem and EDTA 8 Since most anaerobic infections involve mixed aerobic anaerobic flora with several different strains it may be difficult to develop reliable predictive values for patient management When new problems are recognised or improvements in these criteria are developed changes will be incorporated into future editions of this booklet and also distributed as informational supplements O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 20 Page 160 of 170 Quality control limits for anaerobes Suppl Brucella Blood Agar Inoc McFarland 1 0 Anaerobic incubation Zone diameter in mm Bact fragilis ug ml thetaiotaomicron MIC pg ml NEO SENSITABS CODE ATCC 25285 ATCC 29741 Amoxycillin Clavulanate 30 15 ug AM CL 32 38 0 5 35 41 0 5 Cefoxitin 60 ug CFOXT 27 33 8 22 27 16 Chloramphenicol 60 ug CLR60 29 34 8 32 37 16 Clindamycin 25 ug CLIND 30 37 1 25 30 4 Imipenem 15 ug IMIPM 33 40 0 06 34 41 0 12 Linezolid 30 ug LINEZ 4 4 Meropenem 10 ug MEROP 29 35 0 12 31 36 0 12 0 25 Metronidazole 16 ug MTR16 29 36 0 5 30 36 1 Tetracyclines 80 ug TET80 33 39 0 25 22 26 16 Penicillin Low 5 ug PEN L 9 14 16 9 14 16 Piperacillin 100 ug PIPRA 22 27 4 8 22 28 8 16 Piperacillin Tazobactam 100 10ug PIHTZ 26 32 0 25 4
146. ains susceptible to Oxacillin 1 ug penicillin can be considered susceptible to ampicillin amoxycillin amoxycillin clavulanate cefaclor cefepime cefixime cefotaxime cefpirome ceftibuten ceftriaxone cefuroxime cefpodoxime ceftizoxime and imipenem for approved indications and these agents need not be tested The CLSI has not yet defined other categories than S due to the absence of vancomycin resistant strains If strains showing smaller zones are observed they should be submitted to a Reference Laboratory According to the CLSI for CSF isolates cefotaxime and ceftriaxone should not be tested by the diffusion method Ceftizoxime Neo Sensitabs is used to predict the susceptibility of S pneumoniae to cefotaxime CFTAX and ceftriaxone CETRX Strains with zones 2 30 mm are predictably susceptible to CFTAX and CETRX Strains with zone lt 29 mm should be tested by an MIC method 7 S lt 0 5 ug ml 2 ug ml Break points not yet established by the CLSI If reported laboratories should indicate that rifampicin should not be used alone for therapy For use against S pneumoniae in acute otitis media acute sinusitis and comunity acquired pneumonia non meningeal interpretative criteria Breakpoints have been based primarily on pharmacokinetic and pharmaco dynamic considerations NCCLS 1998 2001 Amoxycillin results are valid for Penicillin G and ampicillin Norfloxacin is used to screen for fluoroquinolone resistance Isola
147. amicin 40 ug GEN40 223 22 20 lt 19 lt 4 gt 8 f Gentamicin 250ug 250 lt 14 gt 500 Enterococci HLR all amino glycosides c Imipenem 15 ug IMIPM gt 20 19 17 lt 16 lt 4 216 Imipenem EDTA 15 750 ug IM ED Detection of metallo B lactamases Isepamicin 30 ug 220 19 17 lt 16 lt 8 gt 32 Kanamycin 100ug KANAM gt 25 24 21 lt 20 lt 6 gt 25 f Kanamycin 500 ug 500 lt 14 gt 1000 Enterococci HLR amikacin q Levofloxacin 5ug LEVOF 216 15 14 13 2 gt 8 Staphylococcus spp gt 19 18 16 lt 15 lt 1 gt 4 Lincomycin 19 ug LINCO gt 26 25 23 lt 22 lt 2 28 Linezolid 30 ug LINEZ Enterococcus spp 223 22 21 20 lt 2 gt 8 Staphylococcus spp gt 21 lt 20 lt 4 a Mecillinam U 33 ug MECIL gt 18 17 15 lt 14 lt 8 gt 32 c Meropenem 10 ug MEROP 218 17 15 lt 14 lt 4 gt 16 X Pseudomonas spp gt 26 25 21 lt 20 lt 4 gt 16 c Methicillin 29 ug METHI gt 20 19 17 lt 16 lt 8 gt 16 Minocycline 80 ug MINOC gt 22 21 19 lt 18 lt 4 gt 16 q Moxifloxacin 5ug MOXIF gt 19 18 16 lt 15 lt 2 gt 8 Staphylococcus spp gt 24 23 21 lt 20 lt 0 5 22 Mupirocin 10 ug MUPIR gt 14 lt 13 lt 4 gt 8 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 29 of 170 Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R a q Nalidixan U 130ug NALID 225 24 21 lt 20 lt 8 gt 32 Enterobacteriaceae gt lt 25 Reduc
148. an udf res ved at anvende Norfloxacin Neo S Hvis h mningszonen er lt 16 mm eller MIC gt 16 ug ml er der stor sandsynlighed for udvikling af resistente mutanter in vivo Gonococcer Ciprofloxacinresistente gonococcer anvend Nalidixan som surrogat test m betragtes som resistente overfor alle kinoloner Oxacillin 1 ug kan anvendes til p visning af beta lactamase negative gonococcer med nedsat f lsomhed over for penicillin kromosomal resistens Se speciel afl sning Meningococcer Oxacillin 1 ug kan anvendes til screening af meningococcer med nedsat f lsomhed over for penicillin kromosomal resistens Se speciel afl sning Nalidixan er anvendelig til screening af stammer med nedsat f lsomhed over for kinoloner Moraxella catharralis Beta lactamaseproduktion hos M catharralis kan p vises ved at teste b de Amoxycillin og Amoxycillin Clav Neo S Beta lactamase positive stammer vil udvise zoner gt 5 mm med Amox Clav i forhold til Amoxycillin alene Listeria spp Listeria er intrinsisk resistent overfor cephalosporiner aztreonam clindamycin fosfomycin colistin og nalidixan og m derfor ikke testes med ovenn vnte antibakterielle midler Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 65 of 170 10 6 VI Interpretation according to MIC Breakpoints of SFM France Vi a Rapidly growing bacteria Interpretation according to the MIC break points recommended by the Comite de l
149. anate may be used for screening purposes Klebsiella oxytoca hyperproducing K 1 beta lactamase may show a false positive result potentiation of cofotaxime and or cefepime Only when the strain is resistant to ceftazidime and shows synergism between ceftazidime and clavulanate should it be reported as ESBL positive ESBL AmpC beta lactamases In strains possessing both chromosomal plasmidic AmpC beta lactamases and ESBLs Boronic acid can be used as inhibitor of the AmpC beta lactamase 34 35 For ex 3 Boronic acid Diatabs are placed on the agar plate and eliminated after 1 hour at room temperature prediffusion The plate is now inoculated and the corresponding Neo Sensitabs Cefotaxime Ceftazidime and Cefepime are placed where the Boronic acid disks were previously When using the combination disks the same procedure with Boronic acid Diatabs is followed An increase in the zones of inhibition of 3 mm compared to the single drugs prediffused with boronic acid indicates the presensce of an ESBL In most cases this procedure is not necessary because with the use of Cefepime Cefepime Clavulanate almost not affected by AmpC beta lactamases and Ceftazidime Ceftazidime Clavulanate it is possible to detect ESBLs in the presence of AmpC beta lactamases NON FERMENTERS Here are particularly P aeruginosa and A baumannii that may possess several types of beta lactamases Non fermenters showing reduced susceptibility to ceftazidime and
150. ance to ampicillin MIC gt 64 ug ml zone lt 12 mm may not be susceptible to the synergistic effect with aminoglycosides Ampicillin susceptibility is used to predict the susceptibility of amoxycillin acylampicillins ampicillin sulbactam amoxycillint clavulanate piperacillin and piperacillin tazobactam for non beta lactamase producing enterococci cross resistance An unusual resistance phenotype 16 was recently detected in 33 nosocomial strains of E faecalis in a Greek hospital The strains were resistant to penicillin MIC gt 32 ug ml and susceptible to ampicillin and amoxycillin clavulanate MIC 0 4 1 5 ug ml The same resistance phenotype has been observed in Denmark 20 Weinstein 9 recommends the use of ampicillin surrogate disk as representative for imipenem when testing against E faecalis and E faecium Recently some ampicillin sensitive imipenem resistant strains have been isolated in Europe 10 Consequently sensitivity testing with imipenem should be done when enterococcal infections with this drug is considered for treatment Ampicillin susceptible MIC lt 2 ug ml but penicillin MIC gt 16 ug ml and imipenem MIC gt 8 ug ml resistant strains of E faecalis were widely disseminated 31 4 96 of the E faecalis with this phenotype in a Greek hospital 22 Ono et al 19 reported ampicillin MIC 8 16 ug ml and imipenem MIC 4 32 ug ml resistant isolates of Van A E faecalis Resistance is due to poin
151. and E test for detection of MBL producing P aeruginosa Clin Microbiol Infect 12 Suppl 4 P1451 2006 Gornaglia et al Metallo Blactamases as emerging resistance determinants in Gram negative pathogens open issues Int J Antimicr Ag 29 380 88 2007 Franklin C et al Phenotypic detection of carbapenem susceptible metallo B lactamase producing Gram negative bacilli in the clinical laboratory J Clin Microbiol 44 3139 3144 2006 Moland E S et al Prevalence of newer B lactamases in gram negative cllinical isolates collected in the U S from 2001 to 2002 J Clin Microbiol 44 3318 3324 2006 Deshpande L M et al Emergence of serine carbapenemases KPC and SME among clinical strains of Enterobacteriaceae in the U S Medical Centers Report from the MYSTIC Program 1999 2005 Iconomidis A et al First occurrence of an E coli clinical isolate producing theVIM1 VIM2 hybrid metallo B lactamase VIM12 Antimicr Ag Chemother 51 3038 39 2007 Perilli M et al Identification and characterisation of a new metallo B lactamase from a clinical isolate of Chryseobacterium indologenes Antimicr Ag Chemother 51 2988 2990 2007 Anderson K F et al Evaluation of methods to identify the K pneumoniae carbapenemase in Enterobacteriaceae J Clin Microbiol 45 2723 25 2007 Kim Soo Young et al Convenient test using a combination of chelating agents for the detection of metallo beta lactamases in the clinical laboratory J
152. ant to Cefoxitin 13 5 VISA GISA Quality Control Using the 20 hours 2 18 h prediffusion technique recommended for detection of VISA GISA hVISA strains on Brain Heart Infusion 5 horse blood and inoculum 0 5 McFarland Q C results should be as follows Strains S aureus ATCC 700788 VISA S aureus ATCC 43300 VSSA Teicoplanin 30 ug Vancomycin 5 ug 2 18 h prediffusion 2 18 h prediffusion Zone Diameter MIC ug ml Zone Diameter MIC pg ml S aureus ATCC 70078 9 12 16 15 20 4 8 S aureus ATCC 43300 23 27 0 5 23 27 1 References 1 Baker C N et al Optimizing testing of methicillin resistant Staphylococcus species Diagn Microbiol Infect Dis 19 167 170 1994 2 Performance Standards for Antimicrobial Susceptibility Testing 17th Inf Suppl M100 S17 2007 3 Mackenzie A M R et al Evidence that the NCCLS disk test is less sensitive than the screen plate for detection of low expression class methicillin resistant S aureus J Clin Microbiol 33 1909 1911 1995 4 Coombs G W et al Problems in detecting low expression class methicillin resistance in S aureus with batches of Oxoid Mueller Hinton agar J Antimicr Chemother 38 551 553 1996 5 Schalbe R S et al Selection for vancomycin resistance in clinical isolates of S haemolyticus J Infect Dis 161 4551 1990 6 Hiramatsu K et al Letter J Antimicrob Chemother 1997 in press 7 Climo M W et al Combinations of vancomyci
153. apenem during therapy for Shewanella alga bacteremia J C M 44 1172 4 2006 10 Schneider I et al Novel carbapenem hydrolyzing oxacillinase OXA 62 from Pandoraea pnomenusa Antimicr Ag Chemother 50 1330 35 2006 11 Villegas M V et al First identification of P aeruginosa isolates producing a KPC type carbapenem hydrolysing B bactamase Antimicr Ag Chemother 51 1553 55 2007 12 Tenover F C et al Carbapenem resistance in K pneumoniae not detected by automated susceptibility testing Emerg Inf Dis 12 1209 1213 2006 13 Sundin D R Rapid screening for KPCs Presentation D 1555 ICAAC september 2007 Chigago USA 14 Queenan A M et al Carbapenemases the versatile B lactamases Clin Microbiol Reviews 20 440 458 2007 O Copyright Rosco Diagnostica A S 09 2007 2008 Page 141 of 170 Chapter 16 NEO SENSITABS 16 6 2 Detection of acquired Metallo beta lactamases MBL The worldwide spread of acquired metallo beta lactamases MBL in gram negative aerobes is of great concern MBL production in clinical isolates of key gram negatives P aeruginosa E cloacae S marcescens and K pneumoniae should be carefully monitored 5 MLBs are classified into 5 major types IMP VIM SPM and GIM and SIM type enzymes In Enterobacteriaceae only IMP and VIM enzymes have been found as yet MBLs hydrolyze most beta lactams carbapenems and large expectrum cephalosporins except aztreonam This phenotype of m
154. ase in P aeruginosa 30 Cloxacillin 500 ug or Boronic acid Diatabs can be used to inhibit AmpC for example by prediffusing 1 hour one of these compounds on the agar before inoculation and before adding the antibiotic tablets Neo Sensitabs placed on the same spots With Klebsiella oxytoca synergism between Amoxycillin Clavulanate AM CL and Aztreonam or Ceftriaxone but not with ceftazidime indicates the presence of hyperproduction of K 1 chromosomal beta lactamase but negative for ESBL Strains producing ESBL show synergism between AM CL and ceftazidime Use Ceftazidime Clavulanate The use of cefotaxime ceftriaxone cefepime aztreonam with AM CL may result in false positive results for ESBL in Klebsiella oxytoca Vitek The emergence of ESBL in Salmonellae merits concern They cause frequently neonatal meningitis in many developing countries and are often already resistant to ampicillin and chloramphenicol 7 Karas et al 8 reports clinical failure due to ESBL in spite of the organism being susceptible with disk diffusion and MIC test CFTAX MIC 0 75 ug ml The double disk diffusion test indicated the presence of an ESBL but the test was first performed when therapy with cefotaxime was stopped due to treatment failure The laboratory report should indicate that ESBL producing strains may be resistant clinically to all penicillins cephalosporins and aztreonam 9 For serious systemic infections even if the isolate appears susce
155. ate 30 10 ug CP CL Pavisning af ESBL Imipenem 15 ug IMIPM Enterobacteriaceae gt 28 27 19 lt 18 lt 1 gt 8 Pseudomonas spp gt 26 25 21 lt 20 lt 4 gt 8 Acinetobacter spp gt 28 27 19 lt 18 lt 1 gt 8 h Imipenem EDTA 15 750ug IM ED P visning af Metallo beta lactamases Meropenem 10ug MEROP Enterobacteriaceae gt 34 33 17 lt 16 lt 0 12 gt 8 Pseudomonas spp gt 30 29 21 lt 20 lt 2 gt 8 Acinetobacter spp gt 30 29 21 lt 20 lt 1 gt 8 O Copyright Rosco Diagnostica A S NEO SENSITABS Zone diameter 09 2007 2008 Chapter 10 Page 58 of 170 Break points imm MIC ug ml NEO SENSITABS STYRKE KODE S l R S R j2 Ertapenem 10 ug ETP10 Enterobacteriaceae gt 30 29 26 lt 26 lt 0 5 gt 1 Cefoxitin 60 ug CFOXT aureus og lugdunensis gt 30 29 28 lt 28 OxaS Mec A pos with MH agar gt 28 27 26 lt 26 OxaS Mec A pos l Coag neg staph gt 34 33 32 lt 32 OxaS A pos with MH agar 232 31 30 lt 30 OxaS Mec A pos Cefoxitin 10 pg CFO10 l S aureus og S lugdunensis gt 22 lt 22 OxaS Mec A pos l Coag neg staph gt 27 26 22 lt 21 OxaS Mec A pos Aminoglycosider Amikacin 40 ug AMIKA Enterobacteriaceae gt 26 25 23 lt 22 4 gt 8 Pseudomonas spp gt 26 25 23 lt 22 lt 8 gt 8 Acinetobacter spp gt 26 25 23 lt 22 lt 4 gt 4 Staphylococcus spp Test Kanamycin lt 4 gt 8 s Gentamicin 40 ug GEN40 Enterobacteriaceae gt 28 27 24 lt 23 lt 2 gt 2 Acinetobacter spp Pseu
156. ate Neo Sensitabs as a phenotypic confirmatory test for the presence of ESBL Perform the antibiogram using Mueller Hinton Agar and McFarland 0 5 inoculum Test both Ceftazidime Clavulanate Cefepime Clavulanate and Ceftazidime Cefepime Neo Sensitabs An increase in zone diameter of gt 5 mm for the combination Ceftazidime Clavulanate or Cefepime Clavulanate compared to Ceftazidime Cefepime alone is confirmatory of the presence of an ESBL Rodriguez Villalobos et al 20 and Fluit et al 21 showed that the double disk Neo Sensitabs synergy test has a higher sensitivity for the detection of ESBL than all combination disks Oxoid E test Steward et al 12 showed that synergism between cefepime and clavulanate Cefepime Clavulanate Neo Sensitabs is very useful to detect ESBL in Klebsiella pneumoniae differentiating strains producing ESBL synergy between cefepime and clavulanate from strains producing Amp C or hyperproducers of beta lactamase Florijn et al 16 conclude that the use of ceftazidime ceftriaxone and amoxycillin clavulanate as Neo Sensitabs is a cheap and reliable method for detection of E coli Klebsiella spp and P mirabilis isolates suspected of carrying ESBL It performs better in a routine setting than the E test which often yields a result that cannot be interpreted Enterobacter Serratia Morganella morganii Providencia Citrobacter freundii and Pseudomonas aeruginosa produce chromosomally encoded inducible Amp
157. ated from animals approved standard 2002 and M31 S1 Informational Supplement 2004 3 Jones N et al Tiamulin activity against fastidious and non fastidious veterinary and human bacterial isolates initial development of in vitro susceptibility test methods J Clin Microbiol 40 461 5 2002 4 Petersen A et al Harmonization of antimicrobial susceptibility testing among veterinary diagnostic laboratories in the five nordic countries Microbial Drug Resistance 9 381 388 2003 5 Gray J T et al Antibiotic susceptibility testing of bacteria isolated from animals Clin Microbiol Newsletter 27 131 5 2005 6 Rich M et al Clindamycin resistance in MRSA isolated from animals Veterinary Microbiology 111 237 40 2005 7 Xian Zhi Li Beta lactam resistance and beta lactamases in bacteria of animal origin Vet Microbiol 121 197 214 2007 8 L thje P et al Molecular basis of resistance to macrolides and lincosamines among staphylococci and streptococci from various animal sources collected in the resistance monitoring program B of T Germ Vet Int J Antimicr Agents 29 528 535 2007 Copyright Rosco Diagnostica A S
158. ation the bottom of each tube is labelled with a short name 5 digit alpha numeric code also marked on each tablet of the antimicrobial contained in the cartridge The dispensers ensure uniform tablet location by a pre determined pattern The bottom plate holes are provided with chutes ensuring that the tablets are correctly placed onto the agar surface The dispensers are equipped with excentrical legs which are easily turned to make a perfect fit for the petri dishes The dispensers may be cleaned with 70 alcohol particularly the holes in the top plate in order to make a good fitting for the cartridges If one cartridge is found too small try cleaning of the dispenser top holes to eliminate tablet dust A mechanized technique for antibiograms has been described using ROSCO dispensers 2 2 Dispenser 101 and Dispenser 104 for Neo Sensitabs These dispensers are to be used with Rosco Neo Sensitabs cartridges with a spring where the potency of the tablets are according to recommendations of Clinical and Laboratory Standards Institute formerly NCCLS 1 Models available from Rosco 1 Adaptable to 8 10 cm petri dishes delivering up to 7 Neo Sensitabs at a time 2 Adaptable for square petri dishes 12 x 12 cm delivering up to 16 Neo Sensitabs at a time The Dispenser 101 and Dispenser 104 are made of hard plastic and are operated by pushing the handle down and the Neo Sensitabs will be transferred to the agar surface When usin
159. ative staph S gt 34 mm I 33 32 og R lt 32 mm Med Cefoxitin 10 ug g lder S aureus og S lugdunensis S gt 22 mm R lt 22 mm Koagulase negative staph S gt 27 mm I 26 22 og R lt 21 mm Staphylococcer resistente overfor cefoxitin m betragtes som resistente overfor alle beta lactam antibiotika uanset Zone st rrelse svares 0 for penicillin resistente stafylokokker svares 0 for methicillin resistente stafylokokker Til p visning af clindamycin resistens hos erythromycin resistente staphylokokker se afsnittet Double tablet induction test D zone test side 89 Mecillinam Neo Sensitabs kan bruges som screening for beta lactamase produktion hos staphylococcer Mecillinam resistente staphylococcer er ogs penicillin resistente beta lactamase produktion Mecillinam zoner gt 22 mm indicerer penicillin f lsomhed hos S aureus 2 Staphylokokker resistente overfor gentamicin b r rapporteres R mod netilmicin og tobramycin Til p visning af hVISA VISA stammer se afsnittet Detection of strains with decreased susceptibility to vancomycin VISA GISA hVISA side 86 Enterococcer Streptokokker enterokokker udviser intrinsisk resistens overfor aminoglykosider og b r kun testes for HLR high level resistens overfor Gentamicin 40 ug og Streptomycins 100 ug Neo Sensitabs Gentamicin 40 ug zone lt 20 mm HLR MIC gt 500 ug ml Streptomycins 100 ug zone lt 20 mm HLR MIC gt 1000 ug ml For enterokokker anvendes Va
160. bs BORON 250 ug CEFTAZIDIME DIPICOLINIC ACID CAZ D 30 250 ug DIPICOLINIC ACID Diatabs D P A 250 ug 1 Cephalosporins Cephems CEPHALOTHIN CLOTN 66 ug CEFACLOR CCLOR 30 ug CEFADROXIL CFDRO 30 ug CEFAZOLIN CFZOL 60 ug CEPHALEXIN CFLEX 30 ug CEPHRADINE CFRAD 60 ug CEFUROXIME CEFUR 60 ug CEFONICID CFCID 30 ug CEFIXIME CFFIX 30 ug CEFPODOXIME CFPOX 30 ug CEFOTAXIME CFTAX 30 ug CEFTAZIDIME CEZDI 30 ug CEFTRIAXONE CETRX 30 ug CEFTIZOXIME CEZOX 30 ug CEFTIOFUR Vet CFTIO 30 ug CEFEPIME CFEPM 30 ug O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 3 Page 10 of 170 Identification Diffusible code amount of Neo Sensitabs Neo Sensitabs Antimicrobial CEFPIROME CFPIR 30 ug CEFSULODIN CFSUL 30 ug CEFQUINOME Vet CFQUI 30 ug CEFTOBIPROLE BAL 9141 CFBIP 30 ug C 2 Cephamycins and Oxacephems CEFOXITIN CFOXT 60 ug C 3 Cephalosporins active against MRSA CEFTOBIPROLE investigational drug CFBIP 30 ug CEFTAROLINE investigational drug CFARL 30 ug D Penems and Carbapenems IMIPENEM IMIPM 15 ug MEROPENEM MEROP 10 ug ERTAPENEM ERTAP 10 ug DORIPENEM investigational drug DORIP 10 ug FAROPENEM investigational drug ree mem E Monobactams AZTREONAM AZTRM 30 ug F Aminoglycosides STREPTOMYCINS 100 ug ST100 100 ug STREPTOMYCIN 500 ug HLR ST500 500 ug KANAMYCIN 100 ug KANAM 100 ug KANAMYCIN 500 ug KA500 500 ug NEOMYCIN Framycetin NEOMY 120 ug AMIKACIN AMIKA 40 ug
161. bs should be suspected of ESBL h Klebsiella Enterobacter and Proteus should be reported R to nitrofurantoin i Klebsiella end Enterobacter should be reported R to ampicillin amoxycillin j Ps aeruginosa St maltophilia and Proteus spp should be reported R to tetracyclines k Concerning detection of VISA GISA strains see User s Guide chapters 13 1 and 13 5 m Interpretation valid for amikacin and isepamicin with staphylococci References 1 Methoden zur Empfindlichkeitspr fung von mikrobiellen Krankheitserregern gegen Chemotherapeutika DIN 58940 4 1 Jan 2000 2 GENARS German Network for Antimicrobial Resistance Surveillance November 2002 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 76 of 170 10 8 VIII Interpretation according to the MIC break points recommended by EUCAST VIII Interpretation according to the MIC break points recommended by the EUCAST Media Iso sensitest Inoculum Semiconfluent growth ICS Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S R 1 Aminoglycosides Amikacin 40 ug AMIKA gt 24 23 20 lt 20 lt 8 gt 16 Coag neg staph gt 30 29 26 lt 26 Gentamicin 40 ug GEN40 Enterobacteriaceae gt 26 25 22 lt 22 lt 2 gt 4 Pseudomonas spp Acinetobacter spp gt 28 27 24 lt 24 lt 4 gt 4 S aureus gt 30 29 26 lt 26 lt 1 gt 1 Coag neg staph gt 34 lt 34 lt 1 gt 1 Netilmicin 40 ug NETIL Ent
162. carbapenemase when imipenem is used for treatment of serious infections caused by S algae 9 Acquired carbapenemases Ambler class A and D MICs ug ml IMIPENEM Ambler 3rd gen CLAV Genetic class Enzymes PIPER cepha AZTRM IMIPM MEROP synergy Inducible Organisms Location A NmcA S S 0 25 2 4 216 2 8 wk E cloacae Chromosomal Sme 1 S 0 25 0 5 4 64 226 0 25 8 wk S marcescens Chromosomal to Sme 3 IMI 1 gt 256 1 2 8 gt 64 4 E cloacae Chromosomal IMI 2 165128 0 1 2 4 8 64 4 32 E asbariae Plasmid KPC 1 gt 128 gt 32 gt 64 16 16 no pneumoniae Plasmid KPC 2 gt 64 gt 8 gt 16 8 16 216 t no K pneumoniae Plasmid oxytoca Salmonella Enterobacter 11 KPC 2 2256 gt 256 256 256 no P aeruginosa Plasmid Chrom KPC 3 256 256 gt 256 gt 4 gt 4 no K pneumoniae Plasmid Enterobacter E coli KPC 4 gt 16 gt 16 Enterobacter GES 2 128 gt 32 16 4516 4 16 t no E cloacae Plasmid P aeruginosa integron GES 3 128 645256 64 0 25 0 5 t no K pneumoniae Plasmid CEFOX R GES 4 128 R R 8 8 no pneumoniae CEFOX R GES 5 R R 8 8 P aeruginosa Integron D OXA 23 gt 256 gt 256 gt 256 4 64 4 128 wk baumanni Chromosomal to OXA 27 integron OXA 40 R 45128 45128 232 232 wk no Ac haemolyticus Plasmid K pneumoniae P aeruginosa OXA 48 8 gt R S gt R S gt R 2364 0 25564 wk no K pneumoniae Plasmid OXA 54 32 S S 1
163. ccording to Hoff 3 and confirmed by our own investigations Oxacillin I ug Neo Sensitabs is useful to detect beta lactamase negative gonococci with decreased susceptibility to penicillin chromosomal resistance as the case is with pneumococci S 2 12 mm R no zone Oxacillin 5 ug Neo Sensitabs may be used for the same purpose Strains of gonococci with decreased susceptibility towards ciprofloxacin have been isolated from patients who did not respond to ciprofloxacin treatment 4 6 CDC has proposed criteria for the interpretation of susceptibilities of gonococci to the quinolones 5 The use of a disk with lower content of antimicrobial is recommended for detecting these strains We recommend the use of Ciprofloxacin 0 5 ug Neo Sensitabs see table 15 2 1 Quality Control Limits for N gonorrhoeae ATCC 49226 G C Agar Base with Supplements Inoculum McFarland 0 5 Incubation at 35 in 5 7 CO NEO SENSITABS POTENCY CODE Zone diameter in mm MIC pg ml Azithromycin 30 ug AZITR 29 35 Ceftriaxone 30 ug CETRX 41 53 0 004 0 016 Cefuroxime 60 ug CEFUR 32 38 0 25 1 Ciprofloxacin 0 5 ug CIP L 34 42 0 001 0 008 Gatifloxacin 5 ug GATIF 45 56 0 004 0 008 Penicillin Low 5ug PEN L 23 31 0 25 1 Spectinomycin 200 ug SPECT 27 33 8 32 Tetracyclines 10 ug TET10 22 28 0 25 1 Tigecycline 15 ug 15 30 40 References 1 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 section 6 2 page 1
164. cephalosporins except cefpirome and cefepime cephamycins and monobactams irrespective of the size of the inhibition zone Plate 16 3 a Plate 16 3 b Demonstration of the presence of inducible beta lactamases in Enterobacter cloacae ATCC 13047 Note the flattened edges of Cefotaxime Neo Sensitabs CFTAX and Ceftazidime Neo Sensitabs CEZDI zones adjacent to Cefoxitin Neo Sensitabs CFOXT Plate 16 3 a and Imipenem Neo Sensitabs IMIPM Plate 16 3 b respectively References 1 Dunne W M et al Use of several inducer and substrate antibiotic combinations in a disk approximation assay format to screen for AmpC induction in patient isolates of P aeruginosa Enterobacter spp Citrobacter spp and Serratia spp J C M 43 5945 9 2005 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 133 of 170 16 3 1 Testing Reporting of Susceptibility to Beta lactams against Enterobacteriaceae and Non fermenters Use the table below for testing reporting of susceptibility to beta lactams against Enterobacteriaceae and non fermenters causing serious infections when inducible beta lactamases are present AMP AM CL CFTAX CETRX CEZDI CFEPM CFOXT AZTRM IMIPM MEROP TIHCL PIHTZ E aerogenes cloacae C freundii R R R R R R T R R T T R R S marcescens Prov stuartii rettgeri Morg morganii P vulgaris penneri R T R R R T T T R T T T T Klebsiella oxytoca R T R R R T T T R T T T
165. cin 5ug MOXIF 230 lt 0 25 gt Ofloxacin 10ug OFLOX 230 lt 0 25 3 D Telithromycin According to Bogdanovich et al 18 H influenzae strains with telithromycin MIC s gt 0 5 ug ml zone x 25 mm have efflux present resistance mechanism Antimicrobial therapy may be ineffective O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Page 102 of 170 Chapter 15 15 1 1 Quality Control Limits for Haemophilus influenzae ATCC 49247 HTM agar Inoculum McFarland 0 5 Incubation at 35 in 5 7 CO NEO SENSITABS POTENCY CODE Zone diameter in mm MIC pg ml Amoxycillin Clavulanate 30415 ug AM CL 20 26 4 2 8 4 Ampicillin 2 5 ug AMP L 10 14 2 8 Ampicillin 33 ug AMP33 22 27 2 8 Azithromycin 30 ug AZITR 15 22 1 4 Ceftobiprole BAL 9141 30ug 28 36 0 25 0 5 Cefaclor 30 ug CCLOR 13 18 128 Cefotaxime 30 ug CFTAX 34 42 0 12 0 5 Ceftazidime 30 ug CEZDI 29 36 0 12 1 Ceftriaxone 30 ug CETRX 32 39 0 06 0 25 Cefuroxime 60 ug CEFUR 20 26 16 Cephalothin 66 ug CLOTN 20 25 32 Chloramphenicol 10 ug CLR10 27 34 0 25 1 Chloramphenicol 60 ug CLR60 36 43 0 25 1 Ciprofloxacin 10 ug CIP10 34 42 0 004 0 03 Clarithromycin 30 ug CLARI 12 18 4 16 Doripenem 10 ug DORIP 21 31 0 12 Erythromycin 78 ug ERYTR 21 27 8 Faropenem 15 22 Gatifloxacin 5 GATIF 33 41 0 008 0 016 Iclaprim 24 33 Levofloxacin 5 ug LEVOF 32 40 0 016 Moxifloxacin 5 ug MOXIF 31 39 0 008 0 03 Nalidixan 130 ug NALID 36 44 1 Quinupris
166. cubation 48 h e VISA grows more slowly than typical vancomycin susceptible S aureus e VISA isolates may demonstrate variable colony morphologies e Some colonies might be present inside the inhibition zone of the glycopeptides e Vancomycin and teicoplanin inhibition zones show a sharp edge if staphylococci are susceptible With VISA GISA strains the edge of the zone of inhibition around Vancomycin 5 and Teicoplanin Neo Sensitabs is currently hazy e Most VISA lack the ability to hemolyze sheep blood 38 Howden et al 30 showed that the addition of 5 horse blood or 20 96 horse serum to Brain Heart Infusion and incubation for 24 48 hours allowed highly sensitive dectection of clinically important hVISA The current diffusion method with vancomycin and teicoplanin disks or tablets is not useful for detecting VISA GISA hVISA strains Consequently a modification of the method is necessary Using horse blood added to the medium as recommended by Howden and a 20 hours prediffusion technique Nielsen amp Casals 32 obtained a clear separation between VISA GISA hVISA strains and VSSA The technique is described below Prediffusion procedure for detecting VISA hVISA a Screening of suspicious strains MRSA strains showing zones of inhibition 15 mm around Cefoxitin 60 ug Neo Sensitabs indicating high level resistance to oxacillin in the current diffusion test should be suspected of possibly being VISA hVISA b Confirmat
167. d test should be performed Navon Venecia et al 11 inoculated 0 2 ml from incubated aerobic bottles blood culture into Mueller Hinton agar plates and tested for ESBL production by the disk method High concordance between direct testing and the standard protocol was achieved Weinbren et al 13 reported ESBL results after 5 6 hours incubation with coli and Klebsiella spp 3 Faeces For detecting the faecal carriage of antimicrobial resistant E coli a faecal swab was directly plated onto a McConkey plate and antimicrobial disks applied onto the seeded plate The same swab was tested with a conventional method The rapid screening method showed a good specificity and good concordance with the conventional method Paradisi et al 12 Kronwall et al 15 Bartolini et al 16 using a direct plating method of fecal samples on McConkey agar studied the status on antimicrobial resistance in commensal E coli in preschool children from low resource countries This inexpensive sensitive and simple method detected high resistance rates of E coli to ampicillin 95 Trim Sulfa 94 Tetracycline 93 and Chloramphenicol 70 35 of the strains were resistant to Nalidixic acid 4 Sputum Cercenado et al 10 in patients with ventilator associated pneumonia evaluated the Gram strain and direct susceptibility test by the diffusion method E test on respiratory samples Results of the difusion test correlated with the microdilu
168. dividual large colonies inside the zone are usually resistant mutants 21 1 1 Interpretation Tables for Yeasts MH GMB 21 1 1 1 Interpretation according to CLSI breakpoints When using the procedure recommended by the CLSI for the diffusion test M H agar 2 glucose 0 5 ug ml methylene blue and McF 0 5 inoculum the interpretation will be as follows 25 26 29 30 44 47 48 49 50 MH Glucose Methylene Blue Agar Inoculum McFarland 0 5 undiluted MIC breakpoints according to CLSI M44 A Zone diameter Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S l R S R Amphotericin B 10 ug AMPHO gt 15 14 10 lt 10 1 22 Caspofungin 5ug CASP5 gt 15 14 12 lt 11 lt 2 gt 2 Fluconazole 25 ug FLUCZ gt 19 18 15 DD lt 14 8 gt 64 Itraconazole 50 8 ug ITRAC 223 22 14 DD lt 13 0 12 2 Ketoconazole 15 ug KETOC 228 27 21 lt 20 lt 0 12 gt 0 5 Posaconazole 52 5ug POSAC gt 17 16 14 lt 13 lt 1 gt 4 Voriconazole l ug VOR 1 gt 17 16 14 lt 13 lt 1 gt 4 DD Dosis Dependent C krusei should always be reported as resistant to fluconazole no matter the zone size Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 21 Page 163 of 170 Azole resistance Development of azole resistance may be a progressive phenomenon that may be related to the accumulation of stepwise mutations with MIC of fluconazole being the first to increase after the selective presure made by any a
169. domonas spp gt 28 27 24 lt 23 lt 4 gt 4 q Staphylococcus aureus gt 30 29 27 lt 26 lt 1 gt 1 q Coag neg staph 234 lt 33 lt gt 1 Netilmicin 40 ug NETIL Enterobacteriaceae gt 28 27 24 lt 23 lt 2 gt 2 Acinetobacter spp Pseudomonas spp gt 28 27 24 lt 23 lt 4 gt 4 q Staphylococcus aureus gt 30 29 27 lt 26 lt 1 gt 1 q Coag neg staph 234 lt 33 lt 1 gt 1 Tobramycin 40 ug TOBRA Enterobacteriaceae 228 27 24 lt 23 lt 2 gt 2 Acinetobacter spp Pseudomonas spp 228 27 24 23 lt 4 gt 4 q Staphylococcus aureus gt 30 29 27 lt 26 lt 1 gt 1 q Coag neg staph 234 33 lt 1 gt 1 Kanamycin 100ug KANAM Staphylococcus spp gt 26 25 24 lt 23 lt 4 gt 16 5 Streptomycin 100ug 100 Staphylococcus spp gt 28 lt 27 lt 8 gt 16 Erythromycin 78 ug ERYTR Staphylococcus spp gt 30 lt 29 lt 0 5 gt 0 5 Clindamycin 25 ug CLIND 0 Staphylococcus spp gt 32 31 28 lt 27 lt 0 5 gt 2 Fucidin 100ug FUCID Staphylococcus spp gt 25 lt 24 lt 0 5 gt 0 5 Chloramphenicol 60 ug CLR60 Enterobacteriaceae gt 26 lt 26 lt 8 gt 16 Staphylococcus spp gt 26 lt 26 lt 8 gt 16 j Tetracyclines 80 ug TET80 Staphylococcus spp 232 31 29 28 lt 2 gt 2 Tigecycline 15 pg TIG15 gt 25 lt 24 lt 1 gt 2 Rifampicin 30 ug RIFAM Staphylococcus spp gt 35 34 31 lt 30 gt 2 Enterococcus spp gt 28 lt 28 lt gt 2 Linezolid 30 ug LINEZ Staphylococcus spp 227
170. e cloxacilline et d acide boronique Oct 2006 Internal study 12 Rupp E et al First detection of the Ampler Class C1 AmpC beta lactamase in Citrobacter freundii by a new simple double disk synergy test J Clin Microbiol 44 4204 4207 2006 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 138 of 170 16 5 Inhibitor Resistant TEM Beta lactamases IRT Strains with this phenotype give patterns of antibiotic resistance similar to TEM 1 or 2 or SHV 1 beta lactamases but they are resistant to amoxicillin clavulanate IRT are found mainly in E coli and Klebsiella pneumoniae Are resistant to amoxicillin clavulanate Zone diameter for Amoxicillin Clavulanate Neo Sensitabs 19 mm Are S generally susceptible to cephalosporins cephalothin cefoxitin cefotaxime References 1 Reguera J A et al Factors determining resistance to beta lactams combined with beta lactamase inhibitiors J Antimicr Chemother 27 569 75 1991 2 Lemozy J et al First characterization of inhibitor resistant TEM IRT beta lactamases in Klebsiella pneumoniae strains Antimicr Ag Chemother 33 2580 2 1995 16 6 Carbapenemases Carbapenemases are beta lactamases that significantly hydrolyze at least imipenem and or meropenem Carbapenemases involved in acquired resistance are of Ambler classes A B and D They may be plasmid or chromosomally encoded Because several of these carbapenemases confer only
171. e 4 control strains must fall within the specified limits Ability to detect methicillin resistance in staphylococci Quality control with S aureus ATCC 43300 Ability to produce clear inhibition zones within the control limits around Trimethoprim and Trimethoprim Sulfa with enterococci Quality control with E faecalis ATCC 29212 Only Mueller Hinton medium formulations that have been tested according to and that meet the acceptance limits described in the CLSI document M6 should be used 1 Neo Sensitabs have been standardized with the following susceptibility test media Mueller Hinton Agar Iso sensitest Agar and Danish Blood Agar The pH value of each batch of agar medium should be checked when the medium is prepared The agar medium should have a pH of 7 2 7 4 at room temperature after gelling If the pH is too low certain antimicrobials will appear as having lower potency e g aminoglycosides macrolides and the strains will appear as more resistant while other antimicrobials may appear as having excessive activity e g penicilins and the corresponding strains will appear as more susceptible If the pH is too high the opposite effects can be expected 1 The thickness of the agar must be 4 mm 1 mm References 1 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests M2 A8 8th Ed January 2003 2 Pollock H M Barry A L Gavan T L Fuchs P C Hansen S Thornsberry C L Frankel H
172. e detected in the same plate 7 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 135 of 170 16 4 1 Differentiation of AmpC beta lactamases in E coli Mirelis et al 9 found a simple phenotypic method for the differentiation between plasmid mediated and chromosomal AmpC B lactamases in E coli using Cloxacillin 500 ug Neo Sensitabs and by visual examination of the antibiogram plates The presence of scattered colonies located near the edge of the zone of inhibiton of cefoxitin cefotaxime ceftazidime and aztreonam indicated the presence of plasmid mediated AmpC beta lactamases Cloxacillin 500 ug alone do not distinguish between chromosomal or plasmidic AmpC beta lactamases E coli AMC I R Cefotaxime I R Ceftazidime I R Cefoxitin most I R Plasmid AmpC Inducible Chromosomal Not plasmid AmpC AmpC hyperprod AmpC Cloxacillin 500 ug Synergy with Synergy with Synergy with No synergy ceftazidime or cefoxitin ceftazidime or cefoxtin ceftazidime or cefoxitin Boronic acid Synergy with Synergy with Synergy with Ceftazidime Clav Ceftazidime Clav Ceftazidime Clav and or Cefotaxime Clav and or Cefotaxime Clav and or Cefotaxime Clav Cefoxitin No antagonism with Antagonism with 3rd Antagonism or not Imipenem 3rd gen cephalosporins gen cephalosporins depending on the degree of derepression Antibiogram Scattered colonies Scattered colonies Well defined edge resistant mutants of z
173. e inhibition zone Cephalothin or cefaclor may be used as surrogate tablets for screening BLNAR e Cefotaxime and ceftriaxone must not be tested against penumococci by the diffusion method false results A surrogate test ceftizoxime is used instead Ceftizoxime detects reduced sensitivity to third generation cephalosporins Strains sensitive to ceftizoxime show currently MIC 0 5 ug ml towards cefotaxime ceftriaxone susceptible while isolates resistant to ceftizoxime should be tested by an MIC method f Erythromycin is the representative of the macrolide group The normal population of Haemophilus spp is categorized as intermediate to erythromycin g The break points are chosen in such a way that the normal population of pneumococci is categorized as intermediate to ciprofloxacin and ofloxacin h The break points are chosen in such a way that the normal population of Haemophilus spp is categorized as intermediate to penicillin V oral i Strains with reduced sensitivity to ciprofloxacin MIC gt 0 125 ug ml show decreased sensitivity to all quinolones jp Erythromycin Interpretation valid for azithromycin and clarithromycin Break points not yet established by the SRGA O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 44 of 170 10 4 IV Interpretations according to MIC Breakpoints of the Norwegian AFA Group IV a Susceptibility testing of rapidly growing bacteria using Neo Sensitabs
174. ed as resistant to all beta lactams even if they appear susceptible in vitro Strains of Klebsiella spp and E coli may be clinically resistant to cephalosporins and aztreonam therapy by producing ESBL extended spectrum beta lactamase Read in chapter 16 1 of User s Guide how to detect ESBL These high content Neo Sensitabs are used to detect high level resistance to the aminoglycosides Read in chapter 14 3 how to detect HLR HNR The interpretation of results is valid for other combinations of Trimethoprim Sulphonamide Strains showing inhibition zones smaller than the limit for susceptible should be tested by an MIC method MIC breakpoints concentrations critiques not yet established by the Comit de l Antibiogramme SFM Strains showing zone lt 23 mm with Cefpodoxime Neo Sensitabs should be suspected of producing ESBL E coli Klebsiella Salmonella For ESBL confirmatory tests see chapter 16 1 Results with gentamicin and staphylococci are also valid for netilmicin Staphylococci that are resistant to gentamicin should be reported as resistant against both netilmicin and tobramycin enzymes APH 2 AAC 6 Synergism between TTC TI CL and Aztreonam Ceftazidime Cefepime permits the detection of ESBL producing strains in Ps aeruginosa Nalidixan is useful to detect strains with reduced susceptibility to quinolones in 1 Enterobacteriaceae Nali zone lt 25 mm 2 Haemophilus influenzae Nali zone 25 mm 3
175. ed susceptibility to quinolones Neomycin 120ug NEOMY 225 24 21 lt 20 lt 6 gt 25 Netilmicin 40 ug NETIL gt 20 19 17 lt 16 lt 12 gt 32 a Nitrofurantoin U 260 ug NITRO 223 22 20 19 32 gt 128 q Norfloxacin U 10 ug NORFX gt 16 15 14 lt 13 lt 4 gt 16 i Novobiocin 5 ug 5 gt 16 15 14 lt 13 lt 2 q Ofloxacin 10 ug OFLOX gt 18 17 15 lt 14 lt 2 gt 8 Staphylococcus spp 220 19 17 lt 16 lt 1 gt 4 c Oxacillin 1 pg OXA 1 S aureus 213 12 11 lt 10 lt 2 gt 4 Coag neg staph gt 18 lt 17 lt 0 25 20 5 c Oxacillin 5 ug 5 S aureus gt 16 15 14 lt 13 lt 9 a Oxolinic acid U 10 ug OXOLI gt 16 15 14 lt 13 lt 4 gt 8 q Pefloxacin 10 ug PEFLX gt 18 17 15 lt 14 lt 2 gt 8 Penicillin Low 5ug PEN L 226 25 23 lt 22 b Staphylococcus spp gt 26 lt 26 lt 0 1 Beta Lactamase e Enterococcus spp gt 10 gt 16 Pipemidic acid U 30 ug PIPEM gt 20 19 17 lt 16 lt 4 gt 16 Piperacillin 100ug PIPRA gt 23 22 20 lt 19 lt 16 gt 128 Pseudomonas spp gt 18 lt 64 gt 128 d Piperacillin Tazobactam 100 10ug PI TZ 223 22 20 lt 19 lt 16 4 gt 128 4 h Pseudomonas spp gt 18 lt 64 4 gt 128 4 5 Polymyxins colistin 150ug C0150 gt 20 19 17 lt 16 lt 2 24 Pristinamycin 30ug PRIST 223 22 20 lt 19 lt 2 28 Quinupristin Dalfopristin 15 ug SYNIS5 gt 19 18 16 lt 15 lt 1 gt 4 Rifampicin 30 ug RIFAM 226 25 23 lt 22 lt 1
176. eferences 1 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 2003 2 Performance Standard for Antimicrobial Susceptibility Testing 17th Inf Suppl M100 S17 2007 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 20 Page 158 of 170 20 Susceptibility Testing of Anaerobes Increasing resistance to clindamycin and some beta lactam agents among the Bacteroides group has been found at some hospitals Organisms recognized as virulent or commonly resistant should be considered for testing These include the Bacteroides fragilis group Prevotella and Porphyromonas spp Clostridium perfringens Cl ramosum CI septicum and CI difficile 7 A hypervirulent strain of CI difficile ribotype 027 reported as cause of outbreaks in Holland Canada USA and the UK The strain has a characteristic pattern since it is resistant to CIPRO and ERY but susceptible to Clindamycin and Metronidazole 7 Cl ramosum shows intrinsically low level resistance to vancomycin and linezolid Cl innocuum shows intrinsically low level resistance to vancomycin and daptomycin CI clostridioforme shows intrinsically low level resistance to teicoplanin The recommended procedure for susceptibility testing of anaerobes by the diffusion method is the following e Use supplemented Brucella blood agar it supports good growth for essentially all anaerobes Brucella agar base is supplemented with
177. en cepha nate 7 3rd gen cepha 3rd gen cepha 3rd gen cepha Ceftazidime SOR R zone 20 mm Ceftazidime R Cefepime S S Cloxacillin 500 ug Synergism Synergism Synergism Synergism Boronic acid Cloxacillin cefoxitin Cloxacillin cefoxitin Cloxacillin cefoxitin Cloxacillin ceftazidime Cloxacillin ceftazidime Cloxacillin ceftazidime Boronic acid ceftazidime Boronic acid ceftazidime Boronic acid ceftazidime Boronic acid ceftazidime Enterobacter spp C freundii M morganii Hafnia alvei Providencia spp Proteus indole positive and Serratia marcescens all produce an inducible chromosomal AmpC beta lactamase which is not inhibited by clavulanate There may be seen an antagonism between amoxycillin and clavulanate smaller zone with the combination that with amoxycillin alone due to the presence of the inducible beta lactamase All these strains should be reported as resistant to ampicillin amoxycillin and to amoxycillin clavulanate except P vulgaris Using an Amoxycillin Clavulanate disc Neo Sensitabs better performance is obtained due to the dual action of clavulanic acid 1 induces expression of inducible plasmid mediated AmpC beta lactamases antagonism with 3rd gen cephalosporins and 2 permits the detection of an ESBL by enlarging inhibition zones of 3rd gen cephalosporins synergism 7 In the presence of an ESBL an inducible plasmid mediated AmpC both antagonism and synergism can b
178. ency which is unlikely with Neo Sensitabs e Error in reading the inhibition zones e Error in transcribing the data Some errors can be readily resolved by examining the test plates If no obvious deviations are found daily control tests should be performed for at least 5 consecutive test days or until the problem is resolved 1 References 1 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 2003 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 19 Page 157 of 170 19 Limitations of Diffusion Methods Warning Studies are not yet adequate to develop reproducible standards to interpret diffusion tests with other microorganisms not listed in the tables of this booklet Dangerously misleading results can occur when certain antimicrobials are tested against specific microorganisms 1 2 These combinations include the following e All beta lactam antibiotics except oxacillin methicillin and cefoxitin against methicillin resistant staphylococci e Cephalosporins aminoglycosides except testing for high level resistance clindamycin and trimethoprim sulfa against enterococci e First and second generation cephalosporins cephamycins and aminoglycosides against Salmonella and Shigella species e Cephalosporins against Listeria spp e Glycopeptides against S aureus with reduced susceptibility to vancomycin e Penicillins Cephalosporins and aztreonam against E
179. epime Clavulanate 30 10ug CP CL Detection of ESBL Cefotaxime 30 ug CFTAX f Enterobacteriaceae gt 30 29 22 lt 22 lt 1 gt 4 Ceftriaxone 30 ug CETRX f Enterobacteriaceae 230 29 22 lt 22 lt 1 gt 4 Ceftazidime 30 ug CEZDI f Enterobacteriaceae gt 30 29 18 lt 18 lt 1 gt 8 9 Pseudomonas gt 30 29 20 lt 20 lt 2 gt 8 Ceftazidime Clav 30 10ug CZ CL Detection of ESBL Cefpodoxime 30 ug CFPOX 2 lt 28 lt 1 Screen ESBL O Copyright Rosco Diagnostica A S NEO SENSITABS Zone diameter 09 2007 2008 Page 45 of 170 Chapter 10 Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S R S R Cefpirome 30 ug CFPIR Enterobacteriaceae gt 30 29 18 lt 18 lt 1 gt 8 9 Pseudomonas 230 29 20 20 lt 2 gt 8 d Cefoxitin 60 ug CFOXT Staphylococcus aureus gt 30 29 28 lt 28 Oxa S mec A pos Coag neg staph gt 34 33 32 lt 32 Oxa S mec A pos Cefoxitin 10 ug 10 aureus Iso Sensitest 222 lt 22 Oxa S mec A pos S aureus Mueller Hinton gt 18 lt 18 Oxa S mec A pos Monobactams Aztreonam 30 ug AZTRM f Enterobacteriaceae gt 30 29 18 lt 18 lt 1 gt 8 9 Pseudomonas 230 29 20 20 lt gt 8 Carbapenems Meropenem 10ug MEROP h Enterobacteriaceae gt 30 29 23 lt 23 lt 0 5 gt 2 9 Pseudomonas gt 28 27 20 lt 20 lt 4 gt 8 Imipenem 15 ug IMIPM Enterobacteriaceae gt 32 31 24 lt 24 lt 0 5 gt 2 9 Pseudomonas Enterococcus faecalis 226 25 20 lt 20 lt 4 gt 8
180. er M phenotype resulting in a lower level of resistance It is important to be able to detect such strains Besides the high level macrolides lincosamides streptogamin B MLS resistance which may be constitutive or inducible clindamycin zone distorted in the vicinity of erythromycin Shortridge et al 6 detected a novel low level macrolide resistance with clindamycin susceptibility in 4146 of the erythromycin resistant S pneumoniae examined These data suggest that macrolide resistant pneumococci and streptococci should not be assumed to be inducible resistant to clindamycin without performing an induction test 10 Concerning detection of inducible clindamycin resistance see page 117 double tablet induction test for streptococci Faccone et al 16 describe the emergence of an S pneumoniae clinical isolate with high level telithromycin resistance MIC 256 ug ml and simultaneous resistance to fluoroquinolones MIC levofloxacin 64 ug ml Rantala et al 18 observed in 26 of 210 erythromycin resistant erm B positive isolates showing heterogeneous resistance to telithromycin manifested by the presence of colonies inside the inhibition zone When cultured and tested these cells showed stabler homogeneous and high level resistance to telithromycin 4 Fluoroquinolones As consequence of the increasing use of fluoroquinolones resistance has now emerged to this group of compounds 11 O Copyright Rosco Diagnostica A S NE
181. er is recommended NCCLS 2001 Cephalosporins aminoglycosides clindamycin and trimethoprim sulfa should not be tested and or reported against enterococci Reporting of these results can be dangerously misleading except for screening for high level aminoglycoside resistance 4 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 14 Page 96 of 170 Double tablet induction test D zone test 12 For the detection of macrolide resistance phenotypes in enterococci use the double tablet induction test described for streptococci in page 117 Instead of Clindamycin enterococci are resistant use Tylosin 16 membered ring and Erythromycin Neo Sensitabs at a distance of 10 mm between the tablets Absence of significant zone of inhibition around the two Neo Sensitabs indicates constitutive 5 resistance cMLSg Blunting of the Tylosin or of the Erythromycin zones indicates inducible MLSs resistance 1MLSg The iMLSg phenotype with blunting of erythromycin zone near the Tylosin tablet indicates reversibly inducible MLS resistance riMLSg 14 5 Enterococci HLR and VRE Quality Control E faecalis ATCC 51299 van B Zone diameter in mm E faecalis NEO SENSITABS POTENCY CODE ATCC 51299 MIC pg ml AMOXYCILLIN 30 ug AMOXY 25 30 AMPICILLIN 33 ug AMP33 25 31 CHLORAMPHENICOL 60 ug CLR60 9 14 R DOXYCYCLINE 80 ug DOXYC 26 32 ERYTHROMYCIN 78 ug ERYTR zone R GENTAMICIN 250 ug zone
182. er ticarcillim ticarcillin clavulanate or ceftazidime should be tested for MBL production b Enterobacteriaceae For E coli Klebsiella spp P mirabilis Salmonella spp and Shigella spp All carbapenem S I R isolates that are resistant to cefoxitin and amoxicillin clavulanate and are non susceptible to ceftazidime inhibition zone lt 18 mm should be tested for MBL production In all other cases all isolates are non susceptible to carbepenems 18 Procedure Apply an EDTA Diatabs on the plate At 10 mm distance apply Imipenem Neo Sensitabs at one side and Meropenem at the other side 10 mm distance In case the strains shows no zone around Imipenem Meropenem the distance to EDTA should be reduced to 5 mm Apply Ceftazidime Neo Sensitabs and Dipicolinic acid DPA Diatabs at a distance of 5 mm from each other Apply Imipenem EDTA and Ceftazidime DPA separate from the others Interpretation The use of 2 chelating agents EDTA and DPA will enhance the detection of metallo beta lactamases in the clinical laboratory 25 key hole or ghost zone synergism between EDTA and Imipenem and or Meropenem indicates the presence of MBL 19 key hole or ghost zone between Ceftazidime and Dipicolinic acid indicates the presence of MBL difference of gt 5 mm larger zone with Imipenem EDTA compared to Imipenem and EDTA alone indicates the presence of a MBL False MBL positive results 9 15 may be obtained if EDTA alone is not used fo
183. erator should be used within 6 months With Neo Sensitabs stored at room temperature labelled store below 25 C cartridges that have been opened or are placed in a dispenser should be stored at room temperature and used within 12 months of opening date The stability of antimicrobials in paper disks is decreased compared to Neo Sensitabs The CLSI 1 recommends frozen storage of paper disks containing beta lactam antibiotics In case of Imipenem Cefaclor and Clavulanic acid combinations paper disks should be stored frozen until the day of use In a comparative stability study between Neo Sensitabs and Oxoid paper disks 2 it was observed that disks containing Ticarcillin 75 ug lost activity after 15 days at 4 6 while Ampicillin 10 ug and Amoxycillin Clavulanate 204 10 ug disks lost activity after one month at 4 6 C The corresponding Neo Sensitabs were stable at room temperature and at 4 6 during the study period of six months Steward et al 3 noticed overdetection of imipenem meropenem resistance in the project ICARE most probably due to inactivation of the reagents used Vitek disk diffusion etc and recommended the use of a second independent antimicrobial susceptibility testing method to validate carbapenem intermediate and resistant strains O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 2 Page 8 of 170 References 1 NCCLS Performance Standards for Antimicrobial Disk Su
184. erobacteriaceae 226 25 22 22 lt 2 gt 4 Pseudomonas spp Acinetobacter spp gt 28 27 24 lt 24 lt 4 gt 4 aureus gt 30 29 26 lt 26 lt 1 gt 1 Coag neg staph gt 34 34 lt 1 gt 1 Tobramycin 40 ug TOBRA Enterobacteriaceae 226 25 22 22 lt 2 gt 4 Pseudomonas spp Acinetobacter spp 228 27 24 24 lt 4 gt 4 aureus gt 30 29 26 lt 26 lt 1 gt 1 Coag neg staph gt 34 lt 34 lt 1 gt 1 2 Quinolones Nalidixan 130ug NALID Enterobacteriaceae 28 Reduced susceptibility Haemophilus spp lt 30 lp quinolones Ciprofloxacin 10 ug CIP10 Enterobacteriaceae nal gt 28 27 24 lt 24 lt 0 5 gt 1 Pseudomonas spp gt 30 29 27 lt 27 lt 0 5 gt 1 Acinetobacter spp Staphylococci gt 26 25 22 lt 22 1 gt 1 S pneumoniae gt 36 35 22 lt 22 lt 0 12 gt 2 influenzae Moraxella nal gt 30 29 26 lt 26 lt 0 5 gt 0 5 Gonococci Meningococci nal gt 36 lt 36 lt 0 03 gt 0 06 Levofloxacin 5 ug LEVOF Enterobacteriaceae gt 26 25 23 23 lt 1 22 Pseudomonas spp 228 27 24 24 lt 1 gt 2 Streptococcus spp gt 22 21 19 lt 19 lt 1 gt 2 S pneumoniae gt 22 21 19 lt 19 lt 2 gt 2 H influenzae Moraxella spp nal gt 28 27 24 lt 24 lt 1 gt 1 Moxifloxacin 5ug MOXIF Enterobacteriaceae nal gt 28 27 24 lt 24 lt 0 5 gt 1 Staphylococci gt 28 27 24 lt 24 lt 0 5 gt 1 pneumoniae gt 24 23 22 lt 22 lt 0 5 gt 0 5 influenzae Moraxella spp nal 2
185. ers should be reported as resistant to penicillin amoxycillin ampicillin azlocillin piperacillin and ticarcillin b Oxacillin resistant staphylococci should be reported as resistant to all other beta lactams penicillins beta lactamase inhibitor combinations cephalosporins and carbapenems c Staphylococci resistant to gentamicin should be reported as resistant to all aminoglycosides except streptomycin d E coli Klebsiella and Salmonella strains resistant to Nalidixan Neo Sensitabs zone 28 mm show a decreased susceptibility to quinolones CIPRO MIC gt 0 125 ug ml It may result in treatment failure with quinolones O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 75 of 170 e When testing enterococci use Gentamicin 250 ug and Streptomycin 500 ug to detect high level resistance HLR Strains that are HLR to gentamicin should be reported HLR to all aminoglycosides except streptomycin f When testing enterococci use Vancomycin 5 ug Neo Sensitabs Plates should be incubated for full 24 hours and examined carefully for the presence of a haze or other growth within the zone resistance g Strains of E coli and Klebsiella spp that produce ESBL may be clinically resistant to therapy with penicillins cephalosporins or aztreonam despite apparent in vitro susceptibility See chapter 16 1 for ESBL screening and confirmatory tests Strains showing zone lt 23 mm with Cefpodoxime Neo Sensita
186. es The suggested breakpoint classified the normal population as intermediate Staphylococci may develop resistance during quinolone therapy There is cross resistance between quinolones against staphylococci Valid for systemic therapy E faecalis is resistant to Quinupristin Dalfopristin With these MIC breakpoints the normal population of E coli is categorized as intermediate to cefuroxime For detection of inducible MLS resistance we refer to Neo Sensitabs User s Guide For detection of VISA GISA h VISA strains see chapter in User s Guide on detection of staphylococci with decreased susceptibility to vancomycin page 86 MIC breakpoints taken from SRGA or EUCAST Prediffusion technique 24 18 hours is described on page 18 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 49 of 170 IV b Susceptibility testing of Haemophilus spp Streptococcus pneumoniae Streptococcus spp and Moraxella catarrhalis using Neo Sensitabs and breakpoints recommended by the Norwegian AFA Group January 2006 Version 1 9 Inoculum according to ICS Media HTM Besides PDM MH and ISO all 5 blood and incubation 5 7 CO Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Penicillins Penicillin Low G 5ug PEN L Haemophilus spp 220 19 14 14 lt 1 gt 4 j Beta haem streptococcus 225 lt 0 12 j Viridans streptococcus gt 28 27 17 lt 17 lt
187. es have been found most frequently in species naturally negative for AmpC such as K pneumoniae E coli K oxytoca Salmonella and P mirabilis Recently they were also found in Enterobacter spp 2 The strains with plasmid mediated AmpC show resistance to cefoxitin MIC gt 16 ug ml and ceftazidime MIC gt 16 ug ml corresponding to zones of inhibition lt 20 mm McF 0 5 Strains with plasmid mediated AmpC do not show antagonism between cefoxitin and 3rd generation cephalosporins are not inducible while inducible plasmid mediated AmpC ACT 1 DHA 1 DHA 2 CMY 13 show antagonism between cefoxitin or imipenem and third generation cephalosporins Isolated that coproduce an ESBL and a plasmid mediated AmpC beta lactamase may yield a positive confirmatory test for ESBL using cefepime and cefepime clavulanate synergism Characteristics of AmpC beta lactamases Chromosonally Plasmid Inducible plasmid AmpC mediated AmpC mediated AmpC mediated AmpC overexpression partially derepressed derepressed AmpC ACT 1 DHA 1 P aeruginosa AmpC mutants mutants DHA 2 CFE 1 CMY 13 Cefepime Clavulanate No synergism No synergism No synergism No synergism and or except MOX 1 Ceftazidime Clavula MOX 2 nate Cefoxitin Imipenem Cefoxitin Cefoxitin Cefoxitin No antagonism R zone lt 20 mm R zone lt 20 mm R zone lt 20 mm Imipenem or Amoxycillin Clavula Antagonism with No antagonism with Antagonism with 3rd g
188. etative zones are Tentative for 1 year Breakpoints have not yet been established by the CLSI Proposed MIC breakboints by the CLSI Working Group on Stenotrophomonas and Burkholderia August 2003 The Subcommittee for Antimicrobial Susceptibility Testing has established the Working Group for Susceptibility Testing of S maltophilia and B cepacia CLSI recommends only minocycline levofloxacin and trim sulfa to be reported for S maltophilia isolates and only ceftazidime meropenem and minocycline to be reported for B cepacia isolates 13 a Dueto the high molecular weight of colistin its diffusion on agar is very slow resulting in small differences in inhibition zones between susceptible and resistant strains Zones are tentative for one year Rosco Diagnostica has developed a prediffusion technique for colistin permitting a clear differentiation between susceptible and resistant strains page 18 Place Colistin 10 ug Neo Sensitabs on a non inoculated MH plate and incubate at room temperature for 2 hours Thereafter eliminate the tablet by knocking the place against the table and leave the plate at room temperature for further 18 hours overnight Inoculate the plate and incubate overnight Read the inhibition zones b Resistant subpopulations colonies were grown within the zone of inhibition around imipenem and meropenem discs with A baumannii strains in Greece heteroresistance showing MIC s of 8 32 ug ml imipenem and meropenem MIC
189. eye Faint growth or tiny colonies that may appear to fade from the more obvious zone should be ignored in the measurement 1 Zone diameter interpretative criteria when testing Haemophilus spp are listed in the table below Haemophilus spp HTM agar Inoculum McFarland 0 5 Incubation in 5 7 CO Break points according to CLSI M2 A8 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S R 8 Amoxycillint Clav 30 15 ug AM CL gt 24 24 lt 442 gt 8 4 a Ampicillin 2 5 ug AMP L gt 20 19 17 lt 16 lt 1 24 8 Ampicillin 33 ug AMP33 228 27 24 lt 23 lt 1 gt 4 a Ampicillin Sulbactam 30 30 ug AM SU gt 26 lt 26 lt 2 1 gt 4 2 b Azithromycin 30 ug AZITR gt 16 lt 4 b Aztreonam 30 ug AZTRM gt 28 lt 2 a Cefaclor 30 ug CCLOR 223 22 20 lt 19 lt 8 gt 32 Cefepime 30 ug CFEPM gt 28 lt 2 b Cefixime 30 ug CFFIX gt 32 lt 1 b Cefotaxime 30 ug CFTAX gt 28 lt 2 Cefpirome 30 ug CFPIR 228 lt 2 b Cefpodoxime 30 ug CFPOX 228 5 lt 2 i b Ceftazidime 30 ug CEZDI gt 28 lt 2 b Ceftriaxone 30 ug CETRX gt 28 lt 2 a Cefuroxime 60 ug CEFUR gt 28 27 24 lt 23 lt 4 gt 16 Cephalothin 66 ug CLOTN gt 28 lt 23 lt 8 amp R Chloramphenicol 10 ug CLR10 220 19 17 lt 16 lt 2 gt 8 Chloramphenicol 60 ug CLR60 gt 32 31 27 lt 26 lt 2 gt 8 Ciprofloxacin 10 ug CIP10 gt 26 lt 1 Clarithromycin 30 ug CLARI gt
190. f possessing 16S rRNA methylases a Enterobacteriaceae 16S rRNA methylase positive strains will show Amikacin 40 ug No zone of hibition Gentamicin 40 ug No zone of hibition Netilmicin 40 ug No zone of hibition Tobramycin 40 ug No zone of hibition Neomycin 120 ug Zone of inhibition lt 20 mm or no zone Apramycin 40 ug Zone of inhibition 2 20 mm 6 S Non fermenters 16S rRNA methylase positive strains will show Amikacin 40 ug No zone of hibition Gentamicin 40 ug No zone of hibition Netilmicin 40 ug No zone of hibition Tobramycin 40 ug No zone of hibition Neomycin 120 ug No zone or small zone Streptomycin 100 ug Small zone in most cases Galimand et al 5 found in 12 clinical isolates of Enterobacteriaceae the armA gene associated with ESBL beta lactamase CTXM 3 cefotaxime zone ceftazidime zone on a conjugative plasmid Bogaerts et al 9 investigated the presence of 16 SrRNA methylase mediated high level resistance to aminoglycosides in clinical isolates of Enterobacteriaceae from 2 University Hospitals in Belgium They screened for HLR to gentamicin tobramycin and amikacin resistance and deleted by PCR ArmA genes in 18 K pneumoniae E coli E aerogenes E cloacae and C amaloneticus whereas RmtB was detected in a single E coli isolate These strains were susceptible to Apramycin and Neomycin Neo Sensitabs except 2 strains All 16 SrRNA methylase positive strains produced ESBL s predominantly type CTX M3
191. fferent species in many countries Phenotypically this resistance has been divided into several phenotypes See table page 94 The results are interpreted as follows Compare the test strain with a Reference sensitive strain E faecalis ATCC 29212 1 An inhibition zone smaller than the Reference strain with a sharp edge is typically E gallinarum E casseliflavus 2 The test strain show a zone of inhibition similar to that of the Reference strain and a sharp edge It is sensitive to vancomycin 3 The test strain has a zone of inhibition with a hazy edge i e fine growth visible at the edge of the zone The isolate is VRE with van B type of resistance Van A Strains characterized by an inducible high level resistance to both vancomycin V ANCO and teicoplanin TEICO Both vancomycin and teicoplanin are resistance inducers although vancomycin has a higher inducer activity These strains show no zone of inhibition around Vancomycin 5 ug Neo Sensitabs current diffusion method The Van A resistance is generally plasmid mediated and may be transferred by conjugation to susceptible strains It should be expected that the Van A resistance will spread from enterococci to other microorganisms 5 Van B Strains characterized by variable levels of resistance to vancomycin with MICs 16 1024 ug ml The strains are susceptible to teicoplanin with MICs 0 5 1 ug ml Resistance is only induced by vancomycin but once the strains have been induced
192. fluoroquinolone susceptibility in Salmonella spp Hakanen A et al J Clin Microbiol 37 3572 77 1999 that may be associated with clinical failure CLSI 2003 According to Hakanen et al JCM 43 577 8 2005 Salmonella enterica isolates from Southeast Asia may show a new quinolone resistance pattern NALI susceptible and CIPRO reduced susceptibility MIC 0 12 0 25 ug ml Therefore test both NALI and Ciprofloxacin 0 5 ug Neo Sensitabs Staphylococci that are resistant to gentamicin should be reported as resistant against both netilmicin and tobramycin enzymes APH 2 AAC 6 Rosco Diagnostica has developed a 2 18 hours prediffusion technique for colistin permitting a clear differentiation between susceptible and resistant strains see page 18 Colistin 10 ug disk testing without prediffusion can be used as screening test for high level resistance with P aeruginosa MIC gt 128 ug ml corresponds to no zone of inhibition Cefoxitin should be used to detect MRSA heterogeneous resistance Cefoxitin Neo Sensitabs S gt 25 mm methicillin susceptible and R lt 24 mm methicillin resistant mecA positive For coag neg staph except S lugdunensis use S 28 mm and R lt 27 mm For S aureus incubate for 18 24 h For coagulase negative staphylococci incubate for 24 h Results may be reported at 18 h if resistant Tentative FDA breakpoints September 2005 A 2 18 hours prediffusion technique has been developed by Rosco Diag
193. fusion enterococci lt 16 VRE Virginiamycin 30 ug VIRGI gt 25 24 21 lt 20 lt 1 gt 2 Enterobacteriaceae Pseudomonas spp Acinetobacter spp Staphylococcus spp Enterococcus spp Special technique for testing high molecular weight antimicrobials colistin daptomycin teicoplanin vancomycin 2 18 hours prediffusion method permitting a good separation between susceptible and resistant strains Description of the procedure on page 18 Remarks a Klebsiella spp produces a natural low level beta lactamase that inactivates amino carboxy and ureido penicillins They may appear susceptible in vitro but they should be reported as Intermediate to carboxy and ureido penicillins Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 b d e 8 h m wm n 9 r s Page 68 of 170 Critical concentrations of ampicillin with a fixed concentration of sulbactam 8 ug ml Critical concentrations of amoxycillin with a fixed concentration of clavulanic acid 2 ug ml Critical concentrations of ticarcillin with a fixed concentration of clavulanic acid 2 ug ml Critical concentrations of piperacillin with a fixed concentration of tazobactam 4 ug ml For detecting methicillin oxacillin resistance in staphylococci follow the instructions in User s Guide chapter 13 1 Test Cefoxitin Neo Sensitabs Strains resistant to oxacillin should be report
194. g several dispensers the colour code on the top of the handles can be used for differentiation The holes in the bottom plate ensure that the tablets are placed onto the agar surface in a pre determined pattern The dispensers are easy to disassemble for inside cleaning They must be cleaned occasionally wipe with ethanol and hot water to remove dust from the Neo Sensitabs tablets The tablets come packed in cartridges tubes matching the dispenser top plate holes Each cartridge accommodates 50 Neo Sensitabs and a red block that prevents dispensing when one cartridge is empty Insert the cartridges gently and carefully one by one through the top plates holes For easy identification the bottom of each tube is labelled with a short name 5 digit alpha numeric code also marked on each tablet of the antimicrobial contained in the cartridge Further information and Instructions for use is available on www rosco dk References 1 CLSI Performance Standards for Antimicrobial Susceptibility Testing 15th Inf Suppl M100 S15 2005 2 Guin e P A et al Mechanized technique for phage typing and determination of antibiograms Zbl Bakt Hyg I Abt Orig A246 276 284 1980 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 7 Page 21 of 170 Templates Schabelons In order to simplify the interpretation of the zone sizes a reading chart template for Neo Sensitabs has been devised The template consists of a plate of
195. ganizations in different European countries provide supplementary knowledge of specific considerations and decisions made in each country In the future the use of common European MIC breakpoints would permit the different countries and laboratories to use their separate techniques including inoculum media tablets or disks automatic methods etc and obtain comparable results because the techniques used would be standardized according to common European MIC breakpoints 1 The European Committee on Antimicrobial Susceptibility Testing EUCAST has been reorganized in 2001 2002 giving national breakpoint committees at present France Germany Norway Sweden the Netherlands and the UK a greater role One of the major tasks of the EUCAST is to harmonize MIC breakpoints across Europe and the process is in progress 2 At the moment MIC breakpoints for aminoglycosides cephalosporins carbapenems monobactams fluoroquinolones glycopeptides oxazolidinones tigecycline and daptomycin have been harmonised though EUCAST If included in national recommendations they are included in the interpretation tables I VIII 3 The User s Guide is available at our website www rosco dk and updated information is continuously included Rosco Diagnostica A S is welcoming any feedback and questions on susceptibility testing from users directly info rosco dk or through our representatives ROSCO DIAGNOSTICA A S August 2007 References 1 Kahlmeter G
196. ght Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 118 of 170 References 1 CLSI Performance Standards for Antimicrobial Susceptibility Testing 17th Inf Suppl M100 S17 2007 2 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 Section 6 3 page 12 13 2003 3 Alcaide F et al In vitro activities of 22 beta lactam antibiotics against penicillin resistant and penicillin susceptible viridans group streptococci isolated from blood Antimicr Agents Chemother 39 2243 2247 1995 4 Hsueh et al Decreased activity of erythromycin against Streptococcus pyogenes in Taiwan Antimicr Agents Chemother 39 2239 2242 1995 5 Jiunn Jong Wu et al High incidence of Erythromycin resistant streptococci in Taiwan Antimicr Agents Chemother 41 844 846 1997 6 Rodriguez Avial et al Susceptibility to penicillin and 13 antimicrobial agents in erythromycin resistant viridans streptococci isolated from blood spanish Rev Esp Quimioter 14 sept 2001 7 Descheemaeker P et al Macrolide resistance and erythromycin resistance determinants among Belgian S pyogenes and S pneumoniae isolates J Antimicr Chemother 45 167 173 2000 8 Sepp l H et al Three different phenotypes of erythromycin resistant Streptococus pyogenes in Finland J Antimicr Chemother 32 885 891 1993 9 Malbruny B et al A new phenotype of resistance to lincosamide and streptogrami
197. gonorrhoeae gt 20 19 15 lt 15 lt 0 06 gt 1 b Ciprofloxacin 10 ug CIP10 f N gonorrhoeae gt 36 35 30 lt 30 lt 0 06 gt 1 Tetracyclines 80 ug TET80 N gonorrhoeae 232 31 26 26 lt 1 gt 4 H pylori gt 30 30 4 24 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 36 of 170 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Trimethoprim Sulfa 5 2 240 ug TR SU S pneumoniae gt 32 31 28 lt 28 lt 0 5 9 5 gt 2 38 Vancomycin 5 ug VAN 5 S pneumoniae gt 16 lt 1 Metronidazole 16 ug MTR16 H pylori gt 26 lt 26 lt 8 gt 8 Remarks Haemophilus a Beta lactamase negative ampicillin resistant strains BLNAR are best detected using Ampicillin 2 5 ug Neo Sensitabs BLNAR isolates must be considered resistant to amoxycillin amoxycillin clavulanate as well as first and second generation cephalosporins no matter the size of the inhibition zone b Strains with reduced sensitivity to ciprofloxacin MIC gt 0 125 ug ml show decreased sensitivity to all quinolones S pneumoniae c Oxacillin I ug is used for the detection of strains with reduced sensitivity to penicillin in pneumococci Penicillin resistant isolates from the meninges must be considered resistant to ampicillin amoxycillin amoxycillin clavulanate and first and second generation cephalosporins d Cefotaxime and ceftriaxone must not be tested against pneumococci by t
198. gt 16 mm sharp edge VanB zone 16 mm hazy edge VanC zone 16 mm sharp edge The vanA genotype will show no zone of inhibiton in the current diffusion test with Vancomycin 5 ug Neo Sensitabs O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 6 b Page 19 of 170 Daptomycin testing medium used Mueller Hinton plain inoculum McF 0 5 Staphylococci Daptomycin 30 ug Neo Sensitabs Susceptible zone gt 22 mm corresponding to an MIC of lt I ug ml Enterococci Daptomycin 30 ug Neo Sensitabs Susceptible zone gt 12 mm corresponding to an MIC of lt 4 ug ml III Colistin testing medium used Mueller Hinton plain inoculum McF 0 5 Colistin 10 ug Neo Sensitabs Susceptible zone gt 15 mm corresponding to an MIC of lt 2 ug ml I 14 10 mm R no zone MIC 2 8 g ml References 1 Nielsen S V Casals J B Detection of decreased susceptibility to glycopeptides in S aureus using tablet disc prediffusion 2 3 4 15th Eur Cong Clin Microbiol Inf Dis ECCMID April 2005 Ferreira Nunes AP et al Heterogeneous resistance to vancomycin in S epidermidis S haemolyticus and S warneri clinical strains characterisation of glycopeptide susceptibility profiles and cell wall thickening Intl J Antimicr Ag 27 307 315 2006 Katz B D et al new pre diffusion method for the detection of Daptomycin DAP non susceptible strains using Neo Sensitabs Presen
199. h patients Rev Iberoam Micol 16 97 100 1999 Carrillo Munoz A J et al Abstract P 02 72 In vitro antifungal susceptibility testing standardization Contribution of the Committee of the Spanish Society for Mycology XII congress of the Intl Soc for Human and Animal Mycology Adelaide Australia 1994 Quindos et al Poster I 217 In vitro antifungal susceptibility of Candida isolates from HIV infected patients with oral candidiasis treated with Fluconazole and or Nystatin ICAAC Orlando USA 1994 Carrillo Munoz A J et al Evaluation of an agar diffusion method for in vitro antifungal susceptibility testing Rev Esp Quimioterap 8 221 228 1995 Spanish Carrillo Munoz A J et al In vitro antifungal activity of sertaconazole bifonazole ketoconazole and miconazole against yeast of the Candida genus J Antimicrob Chemother 37 815 819 1996 Casals J B Tablet sensitivity testing of pathogenic fungi J Clin Path 32 719 722 1979 Van Eldere J et al Fluconazole and Amphotericin B antifungal susceptibility testing by NCCLS broth microdilution method compared with E test and semiautomated broth microdilution test J Clin Microbiol 34 842 847 1996 Wanger A et al Comparison of E test and NCCLS broth microdilution method for antifungal susceptibility testing enhanced ability to detect Amphotericin B resistant Candida isolates Antimicr Agents Chemother 39 2520 2522 1995 Casals J B Pringler N Sensitivity
200. he diffusion method A surrogate test is used instead Ceftizoxime Ceftizoxime detects reduced sensitivity to third generation cephalosporins Strains sensitive to ceftizoxime show currently MIC 0 5 ug ml towards cefotaxime ceftriaxone susceptible while isolates resistant to ceftizoxime should be tested by an MIC method e Erythromycin Interpretation valid for azithromycin and clarithromycin Gonococci f Ciprofloxacin resistant gonococci should presumably be considered resistant to all quinolones g A positive beta lactamase test predicts resistance to penicillin amoxycillin ampicillin piperacillin and ticarcillin h Oxacillin 1 ug Neo Sensitabs are useful to detect beta lactamase negative gonococci with decreased sensitivity to penicillin due to chromosomal resistance Meningococci i Oxacillin I ug is used routinely for the detection of reduced sensitivity to penicillins in meningococci chromosomal resistance Helicobacter pylori For species showing slow growth it may be difficult to estabish good correlation between MIC s and zone sizes Use an MIC method References 1 Interpretation for susceptibility tests and susceptibility criteria for antibacterials in the Netherlands CRG Ned Tijdschr Med Microbiol 8 79 81 2000 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 37 of 170 10 3 Ill Interpretation according to MIC Breakpoints of SRGA in Sweden III a Rapidly growing bac
201. he weekly quality control results is out of the acceptable range corrective action is required O Copyright Rosco Diagnostica A S 09 2007 2008 Chapter 12 Page 83 of 170 NEO SENSITABS 12 1 Quality Control Flow Chart Daily testing Max 1 result gt I result of 20 test out of range of 20 tests out of range Test for 30 consecutive days If no more than 3 of 30 results are out of range go to weekly testing Weekly testing Any result to corrective action PA N Reason obvious Reason not obvious Immediate Retest same day corrective action Result in range Em Result out of range return to normal testing Test for 5 days All in range Any result out of range Return to Additional weekly testing corrective action Investigate sources of error O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 12 Page 84 of 170 12 2 Quality Control Zones on Danish Blood Agar Kontrolgr nser SSI substrat Inokulum Semi konfluerende v kst Zone diameter i mm E coli S aureus Ps aeruginosa Ent faecalis NEO SENSITABS CODE ATCC 25922 ATCC 25923 ATCC 27853 ATCC 29212 Amoxycillin Clavulanate 30 15 ug AM CL 25 30 30 37 Ampicillin 33 ug AMP33 26 32 30 38 Cefotaxime 30 ug CFTAX 38 43 Cefoxitin 10 ug CFO10 22 26 Cefoxitin 60 ug 30 36 Ceftazidime 30 ug CEZDI 35 40 32 38 Ceftriaxone 30 ug CETRX 37 42 Cefuroxime 60 ug CEFUR 2
202. hey have been associated with clinical failures As a consequence clinical laboratories should consider screening selected isolates for susceptibility to CFTAX and CETRX as well as to penicillin That is particularly important for blood and CSF isolates Studies have shown that a surrogate disk tablet containing ceftizoxime CEZOX can be used to predict CFTAX and CETRX susceptibility of S pneumoniae 7 This surrogate tablet Ceftizoxime Neo Sensitabs can be used together with Oxacillin 1 ug Neo Sensitabs another surrogate that is already in use to test for susceptibility to penicillin Antimicrobial resistance has clearly emerged as a very serious problem in the United States 3 and other parts of the world The emergence of pneumococci resistant to broad spectrum cephalosporins has limited the choice of antibiotics for the treatment of pneumococcal meningitis 4 It appears that the altered PBP s that reduce the susceptibility of S pneumoniae to penicillin also adversely influence the potencies of CFTAX CETRX and cefpirome in vitro higher MIC values than the susceptible strains Ceftazidime has poor activity 5 Chiu et al 22 observed an increasing ceftriaxone resistance in S pneumoniae from Taiwan 3 Macrolides Telithromycin Resistance to macrolides among S pneumoniae has reached high levels in many countries There are 2 main types of resistance mechanisms one MLS phenotype leading to high level resistance and the oth
203. i are considered part of the viridans group The viridans group also includes S mitis S oralis S sanguis S salivarius S intermedius S constellatus S mutans and S bovis O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 117 of 170 a Strains susceptible to Penicillin Low can be considered susceptible to ampicillin amoxycillin amoxycillin clavulanate ampicillin sulbactam cefaclor cefepime cefotaxime ceftriaxone cefuroxime cefpodoxime ceftizoxime and imipenem for approved indications and need not be tested against these agents Penicillin resistant strains from the CSF should be considered resistant to ampicillin amoxycillin amoxycillin clavulanate and first second generations cephalosporins The CLSI recommends only testing of beta haemolytic streptococci b The CLSI recommends only testing of beta haemolytic streptococci against Ofloxacin Levofloxacin c The CLSI has not yet defined other categories than S due to the absence of vancomycin resistant strains Strains yielding results suggestive of a non susceptible category should be submitted to a Reference Laboratory for further testing d Rifampicin should not be used alone for chemotherapy e Iferythromycin resistant B hemolytic streptococci use D Zone test to detect resistance phenotype f Tentative FDA breakpoints September 2005 g Macrolide resistant isolates erm B may show heterogeneous resistance to telithromycin
204. i mm MIC pg ml NEO SENSITABS STYRKE KODE 3 2 1 0 3 2 1 0 i Gatifloxacin 5ug GATIF 230 21 29 lt 20 lt 0 25 0 5 2 gt 2 m Gentamicin 40 ug GEN40 gt 26 23 25 15 22 lt 14 x4 6 8 12 64 gt 64 d Enterococcus spp lt 20 gt 500 HLR b Imipenem 15 ug IMIPM gt 26 1825 13 17 lt 12 lt 2 3 8 12 16 gt 16 Pseudomonas spp 230 24 29 lt 23 lt 2 4 8 gt 8 Imipenem EDTA 15 750ug IM ED P visning af metallo B lactamase Kanamycin 100 ug gt 28 23 27 15 22 lt 14 lt 4 6 16 32 64 gt 64 i Levofloxacin 5 ug LEVOF gt 30 21 29 lt 20 lt 0 25 0 5 2 gt 2 Linezolid 30 ug LINEZ gt 28 23 27 lt 22 lt 2 4 gt 8 b p Mecilinam U 33 ug MECIL 228 2027 15 19 lt 14 lt 2 3 16 32 64 gt 64 9 Methicillin 29 ug 228 2327 1522 lt 14 lt 1 5 2 6 8 32 gt 32 b Meropenem 10 ug MEROP gt 20 17 19 i lt 16 lt 4 8 32 gt 32 Pseudomonas spp gt 26 21 25 lt 20 lt 2 4 8 gt 8 i Moxifloxacin 5 ug MOXIF gt 30 21 29 lt 20 lt 0 25 0 5 2 22 Mupirocin 10 ug MUPIR gt 20 z lt 19 x1 gt 1 Nalidixan U 130ug NALID gt 28 23 27 15 22 lt 14 lt 4 6 16 32 64 gt 64 Enterobacteriaceae lt 26 Nedsat f lsomhed for Haemophilus spp lt 28 Kinoloner Gonococcer lt 32 Neomycin 120 ug NEOMY gt 28 23 27 1522 lt 14 2 5 m Netilmicin 40 ug NETIL 226 23 25 1522 lt 14 lt 4 6 8 12 64 gt 64 h Nitrofurantoin U 260 ug NITRO 228 23 27 15 22 14
205. iary substances by a dry process using crystalline antimicrobials This procedure results in very uniform tablets homogenous in their content of active ingredients and with an extraordinary stability usually not less than 4 years shelf life at room temperature 2 Neo Sensitabs are standardized according to the MIC breakpoints recommended by Susceptibility Testing Standardization Groups in several countries e g Holland Norway Sweden France Germany UK and Denmark including the CLSI 3 All antimicrobials have received new letter codes see chapter 3 because in order to achieve optimal recognition and zone measurements with automatic instruments a 5 digit code for each Neo Sensitabs type has been chosen References 1 World Health Organization Expert Committee on Biological Standardization Requirements for antibiotic susceptibility tests Requirements for Biological Substances No 26 Geneva September 1981 2 Gylling Pedersen O Standardizing manufacture and control of Neo Sensitabs Acta Clin Belg 28 139 149 1973 3 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Testing 8th Ed M2 A8 January 2003 4 Stes P et al Cefepime activity against Ps aeruginosa evaluation of Etest and two disc diffusion methods J Antimicro Chemother 38 707 711 1996 5 Engberg J et al Comparison of two agar dilution methods and three agar diffusion methods including the Etest for antibiotic susceptibility tes
206. iffucult to assess by the current diffusion method Consequently Rosco Diagnostica is developing a prediffusion method with Amphothericin B Neo Sensitabs that we expect will detect all strains with reduced susceptibility to amphothericin B According to Pfaller et al 31 the fluconazole disk diffusion method using MHGMB agar performed acceptably for testing C neoformans when compared to the reference microdilution method Recently Carillo et al 51 tested Caspofungin and Voriconazole Neo Sensitabs against 184 clinical isolates of Candida spp and other medically important yeasts finding the expected results Diekema et al 52 evaluated Etest and disk diffusion against the broth microdilution reference method using 2171 isolates of Candida spp from 60 medical centres worldwide Posaconazole results with disk diffusion correlated well with the CLSI broth microdilution method demonstrating categorial agreements of 98 interpretations S gt 17 mm and R lt 13 mm O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 21 Page 162 of 170 21 1 Procedure according to CLSI Colonies may be taken from CHROM agar Candida medium or from non differential media as potato dextrose agar 1 Inoculum density and inoculation It is important to standardize the inoculum Therefore use a McFarland 0 5 inoculum Plates may be dried for 20 minutes at 35 C before inoculation Inoculate Mueller Hinton agar plates supplemented with 2
207. illin V 2 stable to penicillinase Methicillin Oxacillin 3 broad spectrum penicillins Ampicillin Amoxycillin 4 penicillins active against pseudomonas Ticarcillin 5 amdino penicillins Mecillinam 6 ureido penicillins Piperacillin 7 alpha methoxypenicillins Temocillin 1 Penicillin Low Neo Sensitabs is the representative of the penicillinase labile penicillins azidocillin penicillin V pheniticillin propicillin penicillin G For pneumococci Oxacillin 1 ug Neo S is used to detect decreased susceptibility to penicillin 2 Methicillin and Oxacillin Neo Sensitabs are the representatives of the penicillinase resistant penicillins cloxacillin dicloxacillin flucloxacillin nafcillin and when testing against staphylococci they are also the representatives of all other beta lactam antibiotics 3 Ampicillin 33 ug and Amoxycillin Neo Sensitabs are the representatives of the group including bacampicillin epicillin hetacillin pivampicillin and talampicillin 4 Ticarcillin Neo Sensitabs is the representative of the group including carindacillin and carfecillin for UTI only 5 Mecillinam Neo Sensitabs is the representative of the group including pivmecillinam 6 Piperacillin Neo Sensitabs 7 Temocillin shows a good activity against Enterobacteriaceae and high stability against beta lactamases Beta lactam Beta lactamase inhibitor Combinations Amoxycillin Clavulanate Ampicillin Sulbactam Ticarcillin Clavula
208. illin and Amoxycillin Clavulanate Neo Sensitabs If the zone around Amoxycillin Clavulanate is gt 5 mm larger than around Amoxycillin alone the strain produces beta lactamase BRO 1 BRO 2 Beta lactamase positive isolates are resistant to penicillin amoxycillin ampicillin piperacillin and ticarcillin Oxacillin 1 ug is used for the detection of reduced sensitivity to penicillin in pneumococci Penicillin resistant isolates from the meninges must be considered resistant to ampicillin amoxycillin amoxycillin clavulanate first and second generation cephalosporins Beta lactamase negative ampicillin resistant strains BLNAR are best detected using Ampicillin 2 5 ug Neo Sensitabs BLNAR isolates must be considered resistant to amoxycillin amoxycillin clavulanate as well as first and second generation cephalosporins no matter the size of the inhibition zone To be used with non meningeal isolates only Breakpoint chosen for therapy of meningitis Cefotaxime and ceftriaxone must not be tested against pneumococci by the diffusion method false results A surrogate test ceftizoxime is used instead Ceftizoxime detects reduced sensitivity to third generation cephalosporins Strains sensitive to ceftizoxime show currently MIC lt 0 5 ug ml towards cefotaxime ceftriaxone susceptible while isolates resistant to ceftizoxime should be tested by an MIC method Breakpoint chosen for therapy of meningitis Erythromycin is the representan
209. illin resistente BLNAR p vises bedst med Ampi 2 5 ug BLNAR isolater m betragtes som resistente over for amox amox clav samt 1 og 2 generations cefalosporiner uanset zonest rrelse b Nalidixanresistente stammer udviser nedsat f lsomhed for ciprofloxacin MIC gt 0 125 ug ml og m betragtes som nedsat f lsomme over for alle kinoloner Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 c d i y k D m n Page 64 of 170 Streptococci Oxacillin 1 ug kan anvendes til p visning af viridans streptococcer med nedsat f lsomhed over for penicillin Se speciel afl sning Til p visning af inducerbar clindamycinresistens hos streptococcer se afsnittet Double tablet induction test D Zone test side 117 S pneumoniae Oxacillin 1 ug anvendes til p visning af stammer med nedsat f lsomhed over for penicillin Penicillinresistente isolater fra spinalv ske m betragtes som resistente over for ampicillin amoxycillin amoxycillin clav samt 1 og 2 generations cefalosporiner Er v ksten semikonfluerende anvendes S gt 24 mm og I R lt 23 mm Er v ksten konfluerende anvendes S gt 20 mm og I R lt 19 mm Cefotaxime og Ceftriaxone m ikke testes over for pneumococcer ved diffusionsmetoder Anvend surrogat testen Ceftizoxime Erythromycin Afl sningsresultatet g lder ogs for Azithromycin og Clarithromycin Screening af pneumococcer med nedsat f lsomhed overfor kinoloner k
210. ine resistant gonococcus TRNG When testing the susceptibility of gonococci towards penicillin it should be possible to detect 2 types of resistance 1 Beta lactamase producing strains plasmid resistance by a constitutive beta lactamase TEM 1 Beta lactamase producing strains of gonococci may be detected a with a rapid beta lactamase test e g Beta lactamase acido Diagnostic Tablets ROSCO code 45521 or b using Amoxycillin Clavulanate Neo Sensitabs compared to Amoxycillin alone Synergism larger zone with Amox Clav will be seen in the presence of a beta lactamase 2 Chromosomal resistance alterations of PBP 1 and PBP 2 or permeability reduction Chromosomally mediated resistance in N gonorrhoeae may represent a continuum in the evolution of the resistance of the gonococcus to various antimicrobial agents In Denmark and Sweden approx 25 of the gonococci show MIC s of 0 25 1 ug ml towards penicillin and 5 are totally resistant to penicillin with MIC s gt 1 ug ml The strains showing decreased sensitivity to penicillin show MIC s 0 1 ug ml and are detected using the current diffusion technique with Penicillin Low Neo Sensitabs Emergence in Japan of beta lactamase negative strains resistant to penicillin MIC 2 8 ug ml and with decreased susceptibility to cefixime MIC gt 0 5 ug ml and ceftriaxone MIC gt 0 125 ug ml These strains had a mosaic PBP2 composed of fragments of PBP2 from N cinerea and N perflava 7 8 A
211. ined with metronidazole are applicable to the others c MIC break points have not yet been given by the CLSI d From August 2005 the FDA no longer allow the use of enrofloxacin for treating infections in poultry to avoid development of resistance in Campylobacter spp e Results of cephalothin susceptibility tests are used to predict susceptibility to the first generation cephalosporins such as cephadroxil and cephalexin f Results of Cefoxitin and Oxacillin with staphylococci are used to predict susceptibility to cloxacillin Cefoxitin resistant staphylococci should be reported as resistant to all beta lactams In case of discordant results between cefoxitin and oxacillin report the strains as resistant R g The results of Trimethoprim Sulfa can be used to predict the susceptibility of other potentiated sulphomanides with thrimethoprim h For detection of ESBL CTX M and AmpC beta lactamases CMY in Salmonella spp see Chapter 16 on Detection of Beta Lactamases 7 i Routine screening of clindamycin inducible resistance in staphyloccci streptococci should be performed double disk induction test page 89 6 Results are also valid for lincomycin References 1 M37 A2 Development of in vitro susceptibility testing criteria and quality control parameters for veterinary antimicrobial agents approved guideline 2002 2 M31 A2 Performance standards for antimicrobial disk and dilution susceptibility tests for bacteria isol
212. inolone resistance associated with the gnr gene in pneumoniae clinicial isolates in the U S Antimicr Ag Chemother 48 1295 9 2004 2 Rodriguez Martinez J M Mechanisms of plasmid mediated resistance to quinolones Enferm Infecc Microbiol Clin 23 25 31 2005 3 Hedi Mammeri et al Emergence of plasmid mediated quinolone resistance in E coli in Europe Antimicr Ag Chemother 49 11 6 2005 4 Xian Zhi Li Quinolone resistance in bacteria emphasis on plasmid mediated mechanisms Intl J Antimicrob Ag 25 453 63 2005 5 Jin Yong Jeong et al Detection of qnr in clinical isolates of E coli from Korea Antimicr Ag Chemother 49 2522 4 2005 6 Poirel L et al Association of plasmid mediated quinolone resistance with ESBL VEB 1 Antimicr Ag Chemother 49 3091 4 2005 7 Robicsek A et al Broda distribution of plasmid mediated quinolone resistance in the U S Antimicr Ag Chemother 49 3001 3 2005 8 Nordmann P Poirel L Emergence of plasmid mediated resistance to quinolones in Enterobacteriaceae J Antimicr Chemother 56 463 9 9 Corkill J E et al High prevalence of the plasmid mediated quinolone resistance determinant anrA in multidrug resistant Enterobacter from blood cultures in Liverpool UK JAC 56 1115 7 2005 10 Poirel L et al In vivo selection of fluoroquinolone resistant E coli isolates expressing plasmid mediated quinolone resistance and ESBL Antimicr Ag Chemother 50 1525 2
213. ion One tablet each of Vancomycin 5 ug and Teicoplanin 30 ug Neo Sensitabs are placed on an uninoculated plate containing Brain Heart Infusion Agar 5 horse blood After 2 hours at room temperature the tablets are removed by knocking the plate against the table and the plate is maintained at room temperature for further 18 hours overnight The plate is now inoculated with the suspicious strain using a 0 5 McFarland inoculum and the plate is incubated at 33 35 overnight Reincubation for further 24 hours is seldom necessary The zones of inhibition are measured and compared to the following c Interpretation VISA GISA hVISA strains will show the following tentative zones of inhibition Teicoplanin 30 ug Neo Sensitabs Inhibition zone 20 mm and or Vancomycin 5 ug Neo Sensitabs Inhibition zone 22 mm Practically all VISA hVISA GISA strains show higher MIC s and consequently higher resistance to teicoplanin than to vancomycin As a consequence the prediffusion test with Teicoplanin 30 ug Neo Sensitabs has the highest sensitivity and accuracy identifying these strains Confirmation of MIC by a reference laboratory is recommended Recently MSSA GISA strains have been isolated in France 36 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 13 Page 87 of 170 Plate 13 1 a Staphylococcus aureus ATCC 700698 VISA Demonstration of pr diffusion 2 18 hours see procedure Vancomycin 5 ug and
214. is sharp Staphylococcus spp may develop resistance during prolonged therapy with quinolones Susceptible isolates may become resistant within 3 4 days after initiation of treatment Testing of repeat isolates may be warranted 11 Rifampicin and fusidate should not be used alone for therapy development of resistance 44 MRSA treatment failures occurred in children when clindamycin was used to treat inducible MLSp resistance Chavez Bueno 20 Siberry et al 22 For the detection of Erythromycin resistance phenotypes use the double tablet induction test also described for streptococci in page 117 inducible 5 constructive MLSg efflux Kanamycin Amikacin Strains possessing the enzyme AAC 3 may show only a minor increase in the MIC of amikacin but the bactericidal and synergistic activities of amikacin with beta lactams or glycopeptides disappear Consequently routine testing of kanamycin not amikacin against all staphylococci should be performed and the results reported as amikacin 41 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 13 Page 89 of 170 Double tablet induction test D Zone test Inducible clindamycin resistance Clindamycin and Erythromycin Neo S are placed approx 20 mm apart from edge to edge Following incubation organisms showing flattening of the clindamycin zone near the Erythromycin tablet indicate inducible clindamycin resistance Such isolates sh
215. is the recommended for testing the fastidious organisms Haemophilus spp gonococci Moraxella catarrhalis pneumococci streptococci and for testing staphylococci for potential methicillin or oxacillin resistance 2 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 6 Page 18 of 170 References 1 Ericsson H M Sherris J C Antibiotic susceptibility testing Report of an International Collaborative Study Acta Path Microbiol Scand Sec B Suppl No 217 1971 2 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests M2 A8 8th Ed January 2003 3 D Amato R F Hochstein L Evaluation of a rapid inoculum preparation method for agar disk diffusion susceptibility testing J Clin Microbiol 15 282 285 1982 6 2 Prediffusion Method Prediffusion Method 2 18 hours for Antimicrobials Diffusing Poorly on Agar High molecular weight antimicrobials vancomycin teicoplanin daptomycin colistin diffuse poorly on agar media resulting in difficult to interpret results when using the disc diffusion method Rosco Diagnostica has developed a 2 18 hours prediffusion technique permitting an easier differentiation between susceptible and resistant strains when testing against these antimicrobials Procedure One Neo Sensitabs of the antimicrobial to be tested is placed on an uninoculated plate containing the susceptibility test medium Mueller Hinton plain or BHI Agar 5 blood After 2 h
216. ity of the agar used It is therefore important to control the quality of the agar used in the diffusion test and this may be achieved by controlling zone sizes with control ATCC strains Separate interpretation tables have been prepared for slow growing and bacteria with special requirements Haemophilus Streptococci Moraxella Neisseria Pneumococci and Anaerobes This booklet describes methods quality control and interpretative criteria recommended in different countries for diffusion susceptibility tests with Neo Sensitabs When new problems are recognized or improvements are developed changes will be incorporated in future editions of the booklet and also distributed as informational supplements References 1 Casals J B Gylling Pedersen O Tablet sensitivity testing a comparison of different methods Acta Path Microbiol Scand sect B 80B 806 816 1972 2 Schumacher et al A procedure for evaluation and documentation of susceptibility test methods using the susceptibility of Klebsiella pneumonae to ciprofloxacin as model J Antimicr Chemother 48 493 500 2001 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 2 Page 7 of 170 2 Characteristics of NEO SENSITABS Neo Sensitabs are produced according to the guidelines of WHO 1 The tablets are 9 mm in diameter and each Neo Sensitabs is printcoded for safe identification The tablets are manufactured with the aid of microbially inert auxil
217. laced on a 150 mm agar plate or 4 Neo Sensitabs onto a 90 100 mm plate The plates are incubated at 35 C with 5 7 CO2 for 20 to 24 hours and the inhibition zones measured to the nearest millimeter The susceptibility of the strains tested is determined according to the interpretation table below Neisseria gonorrhoeae GC Agar Base with Supplements Inoculum McFarland 0 5 Incubation in 5 7 CO Break points according to CLSI M2 A8 Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE S R S R Azithromycin 30 ug AZITR gt 26 lt 25 lt 1 gt 2 Cefepime 30 ug CFEPM gt 32 lt 0 5 Cefixime 30 ug CFFIX gt 32 lt 0 25 Cefotaxime 30 ug CFTAX gt 32 lt 0 5 5 Cefotetan 30 ug CFTTN 226 25 21 gt 20 lt 2 gt 8 Cefoxitin 60 ug CFOXT 232 31 27 26 lt 2 gt 8 a Cefpodoxime 30 ug CFPOX gt 32 lt 0 5 2 Ceftazidime 30 ug CEZDI gt 32 lt 0 5 a Ceftriaxone 30 ug CETRX gt 32 s lt 0 25 Cefuroxime 60 ug CEFUR 232 31 27 26 1 gt A Chloramphenicol 60 ug CLR60 gt 30 29 27 lt 26 lt 4 gt 8 b Ciprofloxacin 10 ug CIP10 gt 41 40 28 lt 27 lt 0 06 gt 1 b Ciprofloxacin 0 5 ug CIP L gt 28 27 15 lt 14 lt 0 06 gt 1 Doxycycline 80 ug DOXYC gt 36 35 27 lt 26 lt 0 25 gt 2 Erythromycin 78 ug ERYTR gt 30 lt 29 lt 1 gt 2 Levofloxacin 5 ug LEVOF gt 32 31 27 lt 26 lt 0 25 gt 1 d Nalidixan quino 130ug NALID 28 Ofloxacin 10 ug OF
218. lanate and AM CL Serratia Citrobacter freundii Cefepime Clavulanate CP CL zone gt 5 mm ESBL confirmatory Enterobacteriaceae than CFEPM alone Cefoxitin Cephalosporins Antagonism indicates Inducible Enterobacteriaceae Imipenem Cephalosporins cephalosporin resistance cephalosporinase AmpC Cefepime Clavulanate No synergy Plasmid mediated Enterobacteriaceae Cefoxitin 3rd gen cepha No antagonism AmpC Cefoxitin R Ceftazidime R Amoxycillint Clavulanate Amoxycillin Clavulanate R Inhibitor resistant E coli Zone lt 19 mm Cefazolin S TEM 8 lactamase Klebsiella Cefazolin Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Page 152 of 170 Chapter 17 Antibiotic Mechanism of Neo Sensitabs Phenotype resistance Bacteria Cefoxitin Antibiotic resistance Porin alteration E coli Klebsiella Aztreonam Ceftazidime Cefepime and Ticarcillin Clavulanate Synergy between 1 and AZTRM CFEPM CEZDI ESBL Ps aeruginosa Imipenem EDTA Synergy between Imipenem Metallo B lactamase Ps aeruginosa and EDTA Acinetobacter E coli Cloxacillin 500 ug Synergy between cefoxitin AmpC beta lactamase Enterobacteriaceae ceftazidime and cloxacillin 500 ug Dipicolinic acid Synergism with Ceftazidime Metallo B lactamase Enterobacteriaceae and or Imipenem Non fermenters Boronic acid Synergism with Cefotaxime4Clav AmpC beta lactamase Enterobacteriaceae and o
219. lates Antimicrob Agents Chemother 39 2193 2196 1995 6 Shortridge V D et al Novel mechanism of macrolide resistance in S pneumoniae Diagn Microbiol Infect Dis 26 73 78 1996 7 Williams Bouyer N et al Predicting susceptibility of Streptococcus pneumoniae to ceftriaxone and cefotaxime by cefuroxime and ceftizoxime disk diffusion testing J Clin Microbiol 37 3707 3710 1999 8 Performance Standards for Antimicrobial Susceptibility Testing 15th Inf Suppl M100 S15 2005 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 9 10 11 12 13 14 15 16 17 18 19 20 21 22 Page 114 of 170 Sahm D F et al In vitro activities of broad spectrum cephalosporins against nonmeningeal isolates of t pneumoniae MIC interpretation using NCCLs M100 S12 recommendations J Clin Microbiol 40 669 674 2002 Descheemaeker P et al Macrolide resistance and erythromycin resistance determinants among Belgian St pyogenes and S pneumoniae isolates J Antimicr Chemother 45 167 173 2000 P rez Trallero E et al Fluoroquinolone and macrolide treatment failure in pneumococcal pneumonia and selection of multidrug resistant isolates Emerg Infect Dis 9 1159 1161 2003 Schrag S J et al Emergence of S pneumoniae with very high level resistance to penicillin Antimicr Ag Chemother 48 3016 23 2004 Smith H J et al Designing fluoroquinolone breakpoints for
220. lin Low 5ug PEN L S pneumoniae Use Oxacillin 1 ug 9 gonorrhoeae gt 34 33 22 lt 22 lt 0 06 gt 1 N meningitidis gt 26 25 22 lt 22 lt 0 06 gt 1 Ampicillin 2 5 ug AMP L a Haemophilus spp 399 21 18 lt 18 lt 0 5 gt 2 Amoxycillin 30 ug AMOXY Haemophilus spp gt 28 27 24 lt 24 lt 1 gt 2 Amoxycillin Clav 30 15 ug AM CL a Haemophilus spp gt 28 27 24 lt 24 1 0 25 gt 2 0 5 Oxacillin lug OXA 1 c S pneumoniae gt 20 lt 20 lt 20 lt 0 06 pen MIC h N gonorrhoeae gt 12 pen screening lt 0 06 pen i N meningitidis gt 10 pen screening lt 0 06 pen MIC Cefuroxime 60 ug CEFUR a Haemophilus spp gt 30 29 26 lt 26 lt 1 gt 2 N gonorrhoeae gt 36 n 36 lt 0 25 Cefotaxime 30 ug CFTAX d pneumoniae Use Ceftizoxime N gonorrhoeae 232 lt 0 25 N meningitidis gt 32 31 28 lt 28 lt 0 5 gt 1 Ceftriaxone 30 ug CETRX d S pneumoniae Use Ceftizoxime N gonorrhoeae gt 32 lt 0 25 N meningitidis gt 32 31 28 lt 28 lt 0 5 gt 1 Ceftizoxime 30 ug CEZOX d pneumoniae gt 30 lt 30 lt 0 5 3rd gen cepha Imipenem 15 ug IMIPM pneumoniae gt 34 33 30 lt 30 lt 0 12 gt 0 5 Azithromycin 30 ug AZITR Campylobacter spp gt 23 22 19 lt 19 lt 1 gt 2 Erythromycin 78 ug ERYTR e S pneumoniae gt 28 27 25 lt 25 lt 0 25 gt 0 5 Clarithromycin 30 ug CLARI H pylori gt 28 27 24 lt 24 lt 0 25 gt 1 b Ciprofloxacin 0 5 ug CIP L f N
221. linical impact of low level resistance in S aureus J Antimicrob Chemother 59 1 4 2007 Tenover F C et al The rationale for revising the CLSI Vancomycin MIC interpretative criteria for S aureus Clin Infect Dis 44 1205 1215 2007 Swenson J M et al The cefoxitin disk test what a clinical microbiologist needs to know Clin Microbiol Newsletter 29 33 40 2007 Yazdankhah S P et al Fusidic acid resistance mediated by fusB in bovine coagulase negative staphylococci J Antimicr Chemother 58 1254 1256 2006 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 14 Page 92 of 170 14 Detection of Resistant Enterococci Enterococci should always be tested on plain agar without blood Use of blood containing media in susceptibility testing of enterococci may result in false susceptibility results with aminoglycosides cefotaxime and other cephalosporins 1 2 3 14 1 Penicillin Ampicillin Resistance Enterococci may be resistant to penicillin ampicillin because of production of low affinity PBPs or less commonly due to the production of beta lactamase sharp edge of the zone of inhibition The Ampicillin 33 ug Neo Sensitabs diffusion test accurately detects isolates with altered PBPs The rare betalactamase producing strains are best detected using a direct nitrocefin test 4 The majority of faecium are resistant to ampicillin low affinity PBPs Enterococci E faecium with high level resist
222. lis from emerging to established pathogen Clin Microbiol Rev 15 125 144 2002 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 110 of 170 15 5 Susceptibility Testing of Pheumococci The recommended medium is Mueller Hinton agar supplemented with 5 defibrinated sheep blood 1 Growth from an overnight 16 20 h sheep blood agar plate is suspended in Mueller Hinton broth or 0 946 saline to a density equivalent to the 0 5 McFarland standard The suspension is used for plate inoculation within 15 min The plates are incubated at 35 in an atmosphere of 5 7 CO for 20 to 24 hours the inhibition zones measured to the nearest millimeter and interpreted according to the table below 1 Penicillin Results with Oxacillin I ug must be reported as susceptibility S or resistance I R to penicillin and not to oxacillin as such Penicillin susceptible strains show MIC lt 0 06 ug ml intermediate or relatively resistant MIC 0 12 1 ug ml and penicillin resistant strains MIC gt 2 ug ml Schrag et al 12 document the emergence of pneumococci with very high level of resistance to penicillin MIC gt 8 ug ml in the USA It is associated with high MIC gt 8 ug ml amoxicillin and cefotaxime MIC s 2 Cephalosporins Many strains of S pneumoniae are only intermediate in susceptibility to penicillin and unfortunately some pneumococci are now resistant to cefotaxime CFTAX and ceftriaxone CETRX and t
223. ll other beta lactams penicillins beta lactamase inhibitor combinations cephalosporins and carbapenems Cefoxitin is now the best test to detect mecA positive staphylococci Staphylococci resistant to Penicillin Low should also be considered as resistant to amoxycillin ampicillin piperacillin and ticarcillin The relatively high MIC breakpoint for I R for the oral cephalosporins is valid only for urinary tract infections When testing Pseudomonas with Cefotaxime Ceftazidime Ceftriaxone and Aztreonam use S 30 mm I 29 23 mm R 23 mm When testing enterococci use Gentamicin 250 ug and Streptomycin 500 ug to detect high level resistance HLR 16 mm Kanamycin 500 ug can be used to detect highlevel resistance to amikacin HLR 16 mm Strains that are HLR to gentamicin are HLR to all aminoglycosides including amikacin except streptomycin When testing Pseudomonas with Kanamycin 100 ug Streptomycins 100 ug Sulphonamides and Trimethoprim Sulfa use S 32 mm I 31 28 mm R 28 mm When testing Pseudomonas with Ciprofloxacin 10 ug Norfloxacin Levofloxacin Ofloxacin and Pefloxacin use S gt 28 mm I 27 23 mm lt 23 mm When testing enterococci use Vancomycin 5 ug Plates should be incubated full 24 hours and examined carefully for the presence of a haze or other growth within the zone indicates resistance It is preferable to use a heavier inoculum equivalent to McFarland 0 5 S 15 mm I 14 13 mm R
224. lococci isolated from healty carriers in Brazil J Clin Microbiol 43 179 185 2005 Steward C D et al Testing for induction of Clindamycin resistance in Erythromycin resistant isolates of S aureus J Clin Microbiol 43 1716 1721 2005 Novotna G et al Prevalence of resistance mechanism against macrolides and lincosamines in methicilin resistant coagulase negative staphylococci in the Czech Republic and occurrence of an undefined mechanism of resistance to lincosamides Antimicr Ag Chemother 49 3586 9 2005 G F W et al Prevalence of GISA among S aureus clinical isolates including methicillin susceptible strains MSSA in a parisian hospital 45th ICAAC presentation D1736 2005 Wootton M et al Evidence for reduction in breakpoints used to determine vancomycin susceptibility to S aureus Antimicr Ag Chemother 49 3982 3 2005 Sakoulas G et al Adaptation of methicillin resistant S aureus in the face of vancomycin therapy CID 42 Suppl 1 540 550 2006 Jones R N Microbiological features of vancomycin in the 21st century MIC creep bactericidal static activity and applied breakpoints to predict clinical outcomes or detect resistant strains CID 42 Suppl 1 513 524 2006 O Sullivan M V N Influence of disk separation distance on accuracy of the disk approximation test for detection of inducible clindamycin resistance in Staphylococcus spp J Clin Microbiol 44 4072 76 2006 Goldstein F The potential c
225. loodstream infections caused by AmpC type beta lactamase producing K pneumoniae Antimicr Ag Chemogher 48 3720 28 2004 5 Wen Chien Ko et al A new therapeutic challenge for old pathogens community acquired invasive infections caused by ceftriaxone resistant Salmonella enterica serotype Cholerasuis CID 40 315 8 2005 6 Nakano R et al CFE 1 a novel plasmid encoded beta lactamse with an ampR gene originating from C freundii Antimicr Ag Chemother 48 1151 8 2004 7 Kyungwon Lee et al Evaluation of phenotypic screening methods for detecting plasmid mediated AmpC beta lactamases producing isolates of E coli and K pneumoniae Diagn Microbiol Infect Dis 53 319 323 2005 8 Wachino Jun ichi et al Horizontal transfer of blaCMY bearing plasmids among clinical E coli and K pneumonia isolates and emergence of cefepime hydrolyzing CMY 19 AAC 50 534 541 2006 9 Mirelis B et al A simple phenotypic method for the differentiation between acquired and chromosomal AmpC beta lactamases in E coli Enferm Infecc Microbiol Clin 24 370 2 2006 10 Wonkeun Song et al Use of boronic acid methods to detect the combined expression of plasmid mediated AmpC beta lactamases and ESBLs in clinical isolates of Klebsiella spp Salmonella spp and P mirabilis Diagn Microbiol Infect Dis 57 315 18 2007 11 Prof P Nordmann Evaluation de tests phenotypiques de detection de cephalospinases integrant l utilisation des disques d
226. lt 28 lt 4 gt 4 O Copyright Rosco Diagnostica A S NEO SENSITABS Zone diameter 09 2007 2008 Page 50 of 170 Chapter 10 Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Carbapenems Imipenem 15 ug IMIPM Haemophilus spp gt 34 lt 1 S pneumoniae streptococci 226 25 24 24 lt 2 gt 2 Anaerobes gt 26 25 20 lt 20 lt 2 gt 8 Meropenem 10 ug MEROP g Haemophilus spp gt 36 lt 0 5 S pneumoniae streptococci 230 29 24 24 lt 0 5 22 S pneumoniae meningit 232 31 24 lt 24 lt 0 25 gt 2 Anaerobes gt 26 25 20 lt 20 lt 2 gt 8 Glycopeptides Vancomycin 5 ug VAN 5 S pneumoniae streptococci gt 18 17 16 lt 16 lt 4 gt 4 Teicoplanin 30 ug TPN30 pneumoniae streptococci gt 18 17 14 lt 14 lt 4 gt 4 Macrolides h Erythromycin 78 ug ERYTR Haemophilus spp gt 34 33 19 lt 19 lt 1 gt 8 j S pneumoniae streptococci gt 28 s 28 lt 0 5 gt 0 5 Moraxella catarrhalis 232 32 lt 1 gt 1 Lincosamides Clindamycin 25 ug CLIND j S pneumoniae streptococci gt 36 35 26 lt 26 lt 0 25 gt 2 Chloramphenicol Chloramphenicol 60 ug CLR60 Haemophilus spp gt 34 33 30 lt 30 lt 2 gt 4 Moraxella catarrhalis S pneumoniae streptococci gt 28 27 23 lt 23 lt 4 gt 4 Chloramphenicol 10 ug CLR10 Haemophilus spp gt 23 22 20 lt 20 lt 2 gt 4 Moraxella catarrhalis Tetracyclines Tetracyclines 80 ug TET80 gt 28 27 24 lt 24 lt 2
227. m Se NE b exti sates eese crspe getestet eemper e 79 12 Quality Control Procedures ete ade et ettet IE Go ce a eee vei 80 12 1 Quality Control Flow Chart 2 iret rette eer reet eet teer dec bee e sees bee bes lee 83 12 2 Quality Control Zones on Danish Blood Agar 84 13 Detection of Resistant Staphylococci Against Methicillin and Vancomyocin esee 85 13 1 StAPNVlOCOC CUS GUTOUS soe to eie i p o e a Pere le t ie eie er et is 85 13 2 Coagulase Negative Staphylococci W W u u u sssesersersereersene renere senere eene ene eene trennen trennen 88 13 3 Comments Concerning Other Antimicrobials eese nennen nennen enne 88 13 4 MRSA Quality Control n nee to E Here REESE DER 90 13 35 VISA GISA Quality Control eec t tee e ete ee pe ti dees xD IN 90 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Page 3 of 170 T4 Detection of Resistant Enterococci tdt eet bed re CPU TETUER rr ee etre 92 14 3 Pemeciln Ampicillin Resistdhce eee eie este eee esent e eerie eicere 92 14 2 Glycopeptide Resistance VRE w uut tere eee rendere pe eee eec beider ete 92 14 3 High level Aminoglycoside Resistance HLR errervrnvrornnnrnnnvnnnnvnnenrverarernevarernrerarvrnevrernvesversvvrsvsrversvessnseer 95 14 4 Comments concerning Other Antibacterials
228. mp C derepressed R R R R S UR S syn FOX syn CAZ CLAV ESBL As for E coli etc Metallo B lactamase and Carbapenemases 4 P vulgaris P penneri Penicillinase R S R S S S S Chromosomal B lactamase R S R R S S S derepressed AMP Ampicillin AMC Amoxicillin clavulanic acid CLOT Cephalothin CAZ Ceftazidime FEP Cefepime FOX Cefoxitin CAZ CLAV Ceftazidime clavulanic acid FEP CLAV Cefepime clavulanic acid IMI Imipenem IMI EDTA Cloxa 500 Cloxacillin 500 ug High AZT Aztreonam Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 147 of 170 16 8 2 Detection of B lactam Resistance Phenotypes in Non fermenters TIC TCC PIP CAZ FEP IMI MERO CAZ CLAV IMI EDTA Remarks FEP CLAV CAZ DPA 1 Pseudomonas aeruginosa Penicillinase R SI S S S S TEM 1 2 PSE 1 4 Amp C partial derep S I any I I S I S S Amp C derepressed IR R IR S S CAZ lt AZT TIC gt CTAX Oxacillinase IR I R S S R S S FEP lt CAZ lt CAZ TIC gt CTAX OXA 31 OXA 1 4 Y R YR any S Y R S S Mex XY OprM Efflux S Y R S S CAZ gt FEP Y R 1 PER 1 2 VEB 1 ESBL R S SA R R S PER 1 S PER 1 Synergy Phenotype PIP S VEB 1 I R VEB 1 CAZ R indicates PER 1 enz ESBL IR any R IR R S S Sy
229. mpicillin resistant b The intermediate range is established because of the difficulty of reading zone endpoints with some bacteria antibiotic combinations and because of clustering of MICs at breakpoint concentrations c Imipenem shows typically MIC s of x 0 12 ug ml against Bact fragilis Imipenem Neo S zone gt 30 mm Strains showing higher imipenem MIC s 2 4 ug ml may represent up to 7 of Bact fragilis isolates 2 and although susceptible to the breakpoint of 4 ug ml they may sometimes conduct to therapeutic failure d Resistance among strict anaerobes to metronidazole in most cases represent laboratory error 3 Strict anaerobiosis is critical for accurate metronidazole susceptibility testing and this environmental failure is the most likely source of false resistant results The SFM recommends Metronidazole Neo Sensitabs for sensitivity testing of anaerobes 5 Beta lactamase testing has limited utility in detecting resistance to certain beta lactam agents among certain anaerobes A positive nitrocefin test can predict resistance to penicillin and ampicillin Other types of tests are not suitable for anaerobes 1 Imipenem resistant and metronidazole resistant Bact fragilis group strains have been detected in fecal samples using the diffusion method 4 Detection of carbapenemase metallo B lactamase production in the B fragilis group is performed using Imipenem and Imipenem EDTA as well as Meropenem EDTA Positive str
230. n 3 220mm Ampicillin 33 Ceftazidime b 2 17 19 mm Cefoxitin 60 ug b Mecillinam b p 0 zio Cefuroxime b Cephalosporins b 3 gt 26 mm Teicoplanin 30 ug Chloramphenicol 60 ug 2 23 25 inm Meropenem b Clindamycin 25 ug 1 15 22 mm Quinupristin Dalfopristin Erythromycin 0 lt 14 mm Teicoplanin 30 ug Kanamycin 100 ug E 2418 t pr diff 20 19 s Lincomycin Amikacin Linezolid Gentamicin 40 ug d m S gt 20 mm Methicillin 9 Netilmicin m R lt 19 mm Nalidixan n Tobramycin m TE i Mupirocin Nr Cefpodoxime 10 ug lt 20 ESBL Nitrofurantoin h 3 gt 26 mm P HE Penicillin Low i 2 18 25 mm Ri 2 3 gt 18 mm ifampicin 1 13 17 mm gt 16 17 Streptomycin 100 ug d 0 lt 12 mm en 1 13 15 Sulphonamides i 2 Tetracyclines 80 ug i Ceftriaxone b c 0 lt 12 mm Trimethoprim Sulfa Imipenem b Fosfomycin Norfloxacin i 3 gt 28 mm Meropenem Pseud 26 20 gt 16 mm 2 21 27 mm ETTE R lt 15 mm 1 10 20 mm 3 gt 24 mm 0 lt 9 mm 2 18 23 mm Novobiocin 5 ug 1 13 17 mm Oxacillin 1 ug f 9 Piperacillin a b 0 lt 12 mm Vancomycin 5 ug q e Piperacillin Tazo b 2418 t pr diff enterococci Ticarcillin a b Tigecycline 25 24 Colistin 10 ug Ticarcillin Clav b Cefpodoxime 30 ug 9 2418 t pr diff 15 14 5 Cefotetan b D aptomycin h j dosering 1 2 g time Fucidin 24 22 24 18 t pr diffusion s enterococci 12 11 O Copyright Rosco Diagnostica A S
231. n A type antibiotics in S agalactiae in New Zealand J Antimicrob Chemother 54 1040 4 2004 10 Pletz M W R et al Fluoroquinolone resistance in invasive S pyogenes isolates due to spontaneous mutation and horizontal gene transfer Antimicr Ag Chemother 50 943 8 2006 11 Jasir A et al Emergence of novel clindamycin resistance phenotype among invasive S pyogenes in Sweden Clin Microbiol Infect 12 Suppl 4 P1594 2006 12 Rantala M et al Telithromycin resistance in pneumococci Clin Microbiol Infect 12 Suppl 4 P1277 2006 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 119 of 170 15 7 Susceptibility Testing of Campylobacter The recommended medium is Mueller Hinton Agar with 596 blood added Use colonies taken directly from an overnight culture and make a suspension in broth or 0 9 saline equivalent to the 1 0 McFarland standard Use the suspension for plate inoculation within 15 min Not more than 9 Neo Sensitabs should be placed on a 150 mm agar plate or 4 Neo Sensitabs onto a 90 100 mm plate The plates are incubated at 36 in Campylobacter atmosphere catalysator for 42 to 48 hours or at 42 for 20 24 hours same conditions and the inhibition zones measured to the nearest mm The disk diffusion and the Etest have been demonstrated to be reliable and convenient methods 3 5 With the increased resistance it becomes important that susceptibility testing is
232. n and beta lactams are synergistic against staphylococci with reduced susceptibilities to vancomycin Ant Agents Chemother 43 1747 53 1999 8 Howe R A et al Interactions between methicillin and vancomycin in MRSA strains displaying different phenotypes of vancomycin susceptibility J Clin Microbiol 37 3068 71 1999 9 Hubert S K et al GISA evaluation of a novel screening method and results of a survey of selected US hospitals J Clin Microbiol 37 3590 3 1999 Hussain Z et al Correlation of oxacillin MIC with MecA gene carriage in coagulasenegative staphylococci J Clin Microbiol 38 752 754 2000 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Testing 8th ed M2 A8 2003 Patel R et al Frequency of isolation of Staph lugdunensis among staphylococcal isolates causing endocarditis a 20 year experience J Clin Microbiol 38 4262 4263 2000 Mougeot C et al Staph aureus nouvelle detection de la resistance intrinsique a la methicilline par la methode de diffusion Pathol Biol 49 199 204 2001 Felten A et al Evaluation of 3 techniques for detection of low level methicillin resistant S aureus MRSA a disk diffusion method with Cefoxitin and Moxalactam the Vitek 2 system and the MRSA screen latex agglutination test J Clin Microbiol 40 2766 2771 2002 10 11 12 13 14 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 13 15
233. n of multiple resistance should be a clue to the possibility of methicillin resistance NaCl should not be added in the diffusion test while MIC tests broth and agar dilution give optimal results with Mueller Hinton added of 2 NaCl and incubation at 33 35 C for a full 24 hours Mackenzie et al found discrepancies in the Oxacillin 1 ug disk test results given by Mueller Hinton agars from three manufacturers when testing against S aureus with low expression class methicillin resistance They recommend to use a low expression class methicillin resistant S aureus as a control for the medium used 3 Coombs et al 4 had problems detecting low expression class methicillin resistance in S aureus with several batches of Oxoid Mueller Hinton agar Vancomycin resistant staphylococci S haemolyticus S epidermidis and S capitis have been isolated from healthy carriers in Brazil 33 For detection use the 20 hours prediffusion technique recommended for VISA hVISA 13 3 Comments Concerning Other Antimicrobials Penicillin susceptible staphylococci are also susceptible to other penicillins cephems and carbapenems Thus susceptibility or resistance to a wide range of beta lactams may be deduced from testing only Penicilin Low Cefoxitin and Oxacillin 1 ug Neo Sensitabs Some rare strains of S aureus produce low levels of B lactamase They may be identified as penicillin resistant because the edge of the zone around Penicillin Low Neo Sensitabs
234. nate and Piperacillin Tazobactam Neo Sensitabs are representatives of combinations of penicillins and betalactamase inhibitors Ceftazidime Clavulanate and Cefepime Clavulanate are useful for detecting ESBLs Imipenem EDTA is useful to detect metallo beta lactamases MBL Cephalosporins 1 First generation of cephalosporins Cross resistance between Cephalothin Cephalexin Cefaclor Cephradine and Cefadroxil Cephalothin Neo Sensitabs is the representative of the group 2 Cefazolin must be tested separately 2 Second generation of cephalosporins Cefuroxime and Cefonicid have a similar spectrum of activity being active in vitro against some strains resistant to cephalothin Cephamycins cefoxitin has a spectrum of activity including anaerobe strains unlike other cephalosporins 3 Third generation of cephalosporins They can be divided into cephalosporins for oral use including cefixime broad spectrum and cephalosporins for injectable use that can be subdivided into two groups A Narrow spectrum good activity against pseudomonas Cefsulodin non stock B Broad spectrum which may be divided into two subgroups a Cephamycins oxa cephems with good activity against anaerobes and low activity against pseudomonas Cefotetan b Moderate activity against anaerobes and b1 good activity against pseudomonas Ceftazidime or b2 moderate activity against pseudomonas Cefotaxime Ceftriaxone Ceftizoxime and Cefodizime 4 Fourth gene
235. ncomycin 5 ug og Mueller Hinton Agar uden blod med kraftig inokulum McFarland 0 5 Afl sning S gt 15 mm I 14 13 mm R lt 12 mm Man kan ikke p vise VanB resistente enterococcer p DBA anvend derfor MH agar O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 61 of 170 Referencer 1 Dragsted U B et al Relapse of multiresistant Salmonella Typhi after combined therapy with ciprofloxacin and ceftriaxone Eur Soc Clin Microbiol Infect Dis 6 167 8 2000 2 Bruun B Gahrn Hansen B Mecillinam susceptibility as an indicator of betalactamase production in Staphylococcus aureus Clin Microbiol amp Infect 8 122 124 2002 3 Skov R et al Tentative interpretative zone diameters for fusidic acid Neo Sensitabs on Mueller Hinton Agar and three blood containing media Int J Antimicrob Ag 22 502 7 2003 4 Klaringsrapport Dansk Selskab for Klinisk Mikrobiologisk Referencegruppe vedr rende Antibiotika Resistensbestemmelse 20 04 2004 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 62 of 170 Vd Streptococcus spp S pneumoniae Haemophilus spp Moraxella catarrhalis Corynebacterium spp Listeria spp Breakpoints ifolge DSKM Referencegruppe for antibiotikaresistens DK Substrat DBA 5 CO Haemophilus spp Chokolade Agar Inoculum Semikonfluerende v kst Zone diameter Break points imm MIC pg ml NEO SENSITABS STYRKE KODE S l
236. ne 18 mm and resistant to Kanamycin 500 ug and Colistin 10ug Fusobacterium is sensitive to Kanamycin 500 ug and Colistin 10 and resistant to Vancomycin 5 ug For species showing slow growth it may be difficult to establish a correlation between MIC s and zone sizes Use an MIC method m Metronidazole Certain strains may show false resistance to metronidazole if anaerobiosis is not correct Helicobacter pylori n Interpretation valid for clarithromycin References 1 Barbut F et al Antimicrobial susceptibilities and serogroups of clinical strains of Clostridium difficile isolated in France in 1991 and 1997 Antimicr Ag Chemother 43 2607 11 1999 2 Communique 2004 de la Societe Francaise de Microbiologie CA SFM 3 Communique Janvier 2006 de la Societe Francaise de Microbiologie CA SFM O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 73 of 170 10 7 VII Interpretation according to MIC Breakpoints of DIN 58940 4 Germany VII Rapidly growing bacteria Interpretation according to the MIC break points recommended by the German DIN 58940 4 January 2000 and GENARS November 2002 Semiconfluent growth ICS Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Penicillin Low 5 ug PEN L a Staphylococcus spp gt 28 E27 lt 0 125 gt 0 25 beta lactamase other gt 28 27 17 lt 16 lt 0 125 gt 2 b Oxacillin lug OXA 1 S au
237. nen enne 122 15 10 Susceptibility Testing of S maltophilia B cepacia and Acinetobacter spp sese 123 16 Detection of Beta Lactamases eee e e EE EH EET irt E e RC rb REESE EUER tee PERIERE 126 16 1 Extended Spectrum Beta Lactamases eene ene nnne enne nene nnne 126 16 2 ESBL Quality Control sp r nest eere RP e ERE ep SH ER E ae EET 131 16 3 Inducible Cephalosporinases or AmpC Beta lactamases esee nennen nennen 132 16 3 1 Testing Reporting of Susceptibility to Beta lactams against Enterobacteriaceae and Non fermentetrs oe ee reka eee 133 16 4 Plasmid mediated AmpC 134 16 4 1 Differentiation of AmpC beta lactamases in E coli see 135 16 5 Inhibitor Resistant TEM Beta lactamases esses eene enne enne enne 138 16 6 Carbapenemases rcs ppt inte ni eret eb e i a i eu te eoe der tete ees 138 16 6 1 Detection of acquired carbapenemases Ambler classes and D 138 16 6 2 Detection of acquired Metallo beta lactamases MBL seen 141 16 7 Detection of multiple beta lactamases in one strain sorsnvrnnorenrverrvernvvrnnvrnrnnernnnvnnenvernvernrrrvrverarerarereevresnee 145 16 8 Detection of B lactam Resistance Phenotypes 1
238. nergy Metallo B lactamase RIR R R R I R I R Synergy AZT SK Carbapenemase Class A R SA R Y R I R I R I R AZT sk Syn IMI CLAV Increased efflux IR VR I R UR IR sk I CAZ gt AZT Loss of Opr porin S S S S S IR S I 2 Stenotrophomonas maltophilia Beta lactamase L 1 RIR R R R R R Synergy AZT S Beta lactamase L 2 S I R R S AZT R Beta lactamase L 1 L 2 RIR R R R R R Synergy AZT R 3 Acinetobacter baumannii TIC PIP PTZ CAZ FEP IMI MERO Penicillinase RIR S S S S S TEM 1 2 Oxacillinase RIR S S S S S lt CAZ 21 37 lt 7 Amp C partial I I SM I S I S S Amp C derepressed R I R S S PER 1 VEB 1 ESBL R any S R R S S Synergy Syn TIC CLAV Syn PIP TAZO ESBL R R R S S Synergy Metallo B lactamase RIR R R R I R I R Synergy AZT SK Carbapenemase Class D R R I R UR I R S I R S U R AZT sk Oxa 23 27 40 51 58 Loss of porins S S S S S I R S I TIC Ticarcillin Ticarcillin clavulanic acid PIP Piperacillin PTZ Piperacillin tazobactam CAZ Ceftazidime FEP Cefepime IMI Imipenem MERO Meropenem CAZ CLAV Ceftazidime clavulanic acid FEP CLAV Cefepime clavulanic acid IMI EDTA Imipenem EDTA AZT Aztreonam References 1 Hocquet D et al Involvement of the MexXY OprM Efflux system in emergence of Cefepime resistance in clinical strains of Ps aeruginosa Antimicr Ag Chemother 50 1347 51 2006
239. nes Neomycin 120ug NEOMY 225 24 21 lt 20 lt 8 gt 16 Netilmicin 40 ug NETIL gt 24 23 21 lt 20 lt 2 gt 4 P aeruginosa Acinetobacter spp gt 23 22 20 lt 19 lt 4 gt 8 Nitrofurantoin U 260ug NITRO 223 22 20 lt 19 lt 32 gt 128 Norfloxacin U 10 ug NORFX gt 25 24 23 lt 22 lt 0 5 gt 1 Ofloxacin 10 ug OFLOX 225 24 22 21 lt 0 5 gt 1 Oxacillin 1 ug lug OXA 1 f Staph aureus 216 16 lt 2 gt 2 Coag neg staph gt 20 lt 20 lt 0 25 gt 0 5 Oxacillin 5 ug 5ug OXA 5 f Staph aureus 220 lt 20 lt 2 gt 2 Oxolinic acid U 10 ug OXOLI gt 18 17 15 lt 14 lt 2 gt 4 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 67 of 170 Zone diameter Concentrations in mm critiques NEO SENSITABS POTENCY CODE S R S R Pefloxacin 10 ug PEFLX 223 22 17 lt 16 lt 1 gt 4 Penicillin Low 5ug PEN L Staphylococcus spp gt 26 26 lt 0 25 beta lactamase Pipemidic acid U 30 ug PIPEM 220 19 17 lt 18 lt 8 gt 16 Piperacillin 100ug PIPRA a Enterobacteriaceae gt 23 22 20 lt 19 lt 8 gt 64 Pseudomonas spp Acinetobacter spp gt 20 19 17 lt 16 lt 16 gt 64 Piperacillin Tazobactam 100 10ug PI TZ e Enterobacteriaceae gt 23 22 20 lt 19 lt 8 4 gt 64 4 Pseudomonas spp Acinetobacter spp gt 20 19 17 lt 18 lt 16 4 gt 64 4 Polymyxins colistin 150ug 150 gt 20 E lt 19 lt 2 gt 2 Pristinamycin 30 ug PRIST gt 25 24 21 l
240. nicillin G resistant N meningitidis by disk susceptibility testing Antimicrob Ag Chemother 31 1478 1482 1987 2 Struillou L et al Rapid emergence of meningococci with reduced susceptibility to penicillin in France the need for vigilance in meningitis treatment Clin Microbiol Infect 4 661 2 1998 3 Campos J Disc testing of meningococci J Clin Microbiol 37 879 880 1999 4 Shultz T R et al An invasive isolate of N meningitidis showing decreased susceptibility to quinolones Antimicro Ag Chemother 44 1116 2000 5 Richter S S et al Neisseria meningitidis with decreased susceptibility to penicillin report from the SENTRY antimicrobial surveillance program North America 1998 99 Diagn Microbiol Infect Dis 41 83 88 2001 6 L et al Bacterial resistance to penicillin by decreased affinity to penicillin binding proteins a mathematical model Emerg Inf Dis 9 411 6 2003 7 Acala B et al N meningitidis showing decreased susceptibility to ciprofloxacin first report in Spain J Antimicr Chemoter 53 409 2004 8 NCCLS CLSI Performance Standards for Antimicrobial Disk Susceptibility Testing 15th Inf Suppl M100 S15 2005 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 109 of 170 15 4 Susceptibility Testing of Moraxella catarrhalis Zone diameter interpretative standards are as indicated in the table below Moraxella catarrhalis Muelle
241. noglycosides except gentamicin and streptomycin Recently 17 new aac 6 Ii like genes have been characterized in hirae and E durans which precludes the synergy between tobramycin or kanamycin amikacin and beta lactams even if the strains are not HLR These results indicate that except for E faecalis association of B lactams with tobramycin and kanamycin amikacin should not be recommended in the therapy of enterococcal infections Penicillin gentamicin synergy may be obtained even in faecium strains exhibiting penicillin ampicillin resistance provided that appropriate penicillin concentrations are used in most cases half of the penicillin MICs 6 14 4 Comments concerning Other Antibacterials High level resistance to ciprofloxacin MIC gt 64 ug ml appears to be associated with ampicillin resistance in genotypically related E faecium isolates and favours the hospital adaptation of lineage CC17 of E faecium 24 New antimicrobials linezolid quinupristin dalfopristin E faecium only show good activity against enterococci A relatively high rate 40 of VR Enterococcus faecium not susceptible MIC gt 8 ug ml to linezolid was observed recently in Italy by Bonosa et al 25 in intensive care unit patients Because of limited alternatives chloramphenicol erythromycin doxycycline minocycline and rifampicin may be tested for vancomycin resistant Enterococci VRE and consultation with an infectious disease practition
242. nostica Zone breakpoints are tentative for 1 year Description of the technique on page 18 P aeruginosa low level resistance to meropenem MIC 8 ug ml cannot be detected by the interpretation recommended by the CLSI Therefore we recommend using Rosco s own interpretation S 2 22 R 18 mm Note For fecal isolates of Salmonella and Shigella spp only ampicillin a quinolone nalidixic acid and trimethoprim sulfa should be tested and reported routinely In addition chloramphenicol and a third generation cephalosporin should be tested and reported for extraintestinal isolates of Salmonella spp Aminoglycosides as well as first and second gen cephalosporins are not effective clinically may appear active in vitro Second and third generation cephalosporins are associated with emergence of resistance during prolonged therapy for Enterobacter Citrobacter and Serratia Susceptible isolates may become resistant within a few days after initiation of therapy Pseudomonas aeruginosa may develop resistance during prolonged therapy with all antimicrobials testing of repeat isolates The susceptibility of Ps aeruginosa isolated from patients with cystic fibrosis can be reliably determined by the diffusion method but may require incubation up to 24 hours CLSI 2001 For Enterobacteriacea isolated from the CSF test cefotaxime or ceftriaxone instead of cephalothin or cefazolin O Copyright Rosco Diagnostica A S NEO SENSITABS 09 20
243. nsfer of fluoroquinolone resistance from S dysgalactiae to S pyogenes is desribed 10 S dysgalactiae may serve as a resistance gene pool for S pyogenes Group B streptococci are susceptible to penicillin ampicillin and cefazolin but may be resistant to clindamycin and or erythromycin When a group B streptococcus is isolated from a pregnant woman with severe penicillin allergy clindamycin and erythromycin should be tested and reported 1 Double tablet induction test D Zone test The test is performed to detect the erythromycin resistance phenotypes Erythromycin resistance is classified on the basis of the double tablet test with Erythromycin and Clindamycin Neo Sensitabs Sepp l 8 The tablets are placed approx 20 mm apart on Mueller Hinton Agar supplemented with 5 blood a Resistance to both erythromycin and clindamycin indicates constitutive MLSs cross resistance ErmB Report resistance to erythromycin and clindamycin b Blunting of the clindamycin zone proximal to Erythromycin Neo Sensitabs indicates an inducible type of MLS ErmA The strain is reported as resistant to both erythromycin and clindamycin c Susceptibility to clindamycin with no blunting of the zone indicates M phenotype efflux mechanism mefA Report erythromycin resistant and clindamycin susceptible d Erythomycin susceptibility S and Clindamycin I R indicate the presence of LSA phenotype lincosamide streptogramin A 9 11 O Copyri
244. obramycin 40 ug TOBRA Enterobacteriaceae 226 25 22 22 lt gt 4 m Staphylococcus spp gt 30 29 26 lt 26 lt 1 gt 1 Daptomycin 30 ug DAPCa 2 2 18 h prediffusion Staphylococcus spp gt 22 lt 1 Enterococcus spp gt 12 lt 4 Streptomycin 500ug X ST500 Enterococcus spp HLR gt 18 lt 18 lt 256 gt 256 Gentamicin 250ug GN500 n Enterococcus spp HLR gt 18 lt 18 lt 128 gt 128 Chloramfenicol Chloramphenicol 60 ug CLR60 gt 26 lt 26 lt 8 gt 8 Enterococcus spp gt 28 lt 28 lt 4 gt 4 Tetracyclines Doxycycline 80 ug DOXYC Enterobacteriaceae 230 20 18 18 28 Staphylococcus spp 228 27 24 24 lt gt 2 Enterococcus spp Tetracyclines 80 ug TET80 Enterobacteriaceae gt 24 23 18 lt 18 lt 4 gt 8 Staphylococcus spp gt 28 27 24 lt 24 lt 1 gt 2 Enterococcus Spp Quinolones 0 Nalidixan U 130ug NALID 224 lt 24 lt 16 gt 16 Enterobacteriaceae lt 28 Reduced susceptibility to quinolones Norfloxacin U 10ug NORFX Enterobacteriaceae gt 28 27 23 lt 23 lt 0 5 gt 1 Ciprofloxacin 10ug CIP10 0 Enterobacteriaceae gt 28 27 23 lt 23 lt 0 5 gt 1 p Staphylococcus spp gt 26 25 22 lt 22 lt 1 gt 1 Enterococcus spp gt 32 31 18 lt 18 lt 0 125 gt 2 9 Pseudomonas 228 27 23 lt 23 lt 1 gt 1 Ofloxacin 10ug OFLOX 0 Enterobacteriaceae gt 28 27 23 lt 23 lt 0 5 gt 1 Enterococcus gt 30 29 16 lt 16 lt 0 25 gt 4 q Staphylococcus s
245. of infection caused by vancomycin resistant S aureus VRSA with vancomycin MIC gt 32 ug ml in a patient in USA has been described 15 17 19 The strain could be detected using the disk diffusion procedure van A gene Tenover et al 24 describe the second VRSA strain in the USA vancomycin MIC 32 ug ml The strain was detected using disk diffusion and vancomycin agar screen while automatic systems Vitek2 and Microscan were unable to detect vancomycin resistance The third VRSA 25 with a van A gene was detected in New York vancomycin MIC gt 64 ug ml Commonly used automatic methods Microscan Vitek failed to detect vancomycin resistance in this VRSA strain while agar based methods detected vancomycin resistance The 4th US case of VRSA Michigan has been confirmed by the CDC March 2005 The strains showed an MIC gt 256 ug ml by broth microdilution and E test Vitek 2 failed to detect resistance with a reported MIC of lt 1 ug ml 28 Consequently additional VRSA infections might have occurred but were undetected by laboratories using only automated methods O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 13 Page 88 of 170 13 2 Coagulase Negative Staphylococci Concerning methicillin resistance in CNS the proportion of resistant bacteria in the heterogenous population is lower than for S aureus making the detection problems more difficult to solve special interpretation required 10
246. ohnson J R et al Direct antimicrobial susceptiblility testing for acute urinary tract infection in women J Clin Microbiol 33 2316 2323 1995 6 Midtved K Midtved T Rapid determination of antibiotic susceptibility by a disc diffusion test for urgent clinical situations Scand J Infect Dis 17 131 132 1985 7 Coyle M B McGonagle L A Plorde J J Clausen C R Schoenknecht F D Rapid antimicrobial susceptibility testing of isolates from blood cultures by direct inoculation and early reading of disk diffusion tests J Clin Microbiol 20 473 477 1984 8 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 2003 9 Saha S K et al Rapid identification and antibiotic susceptibility testing of Salmonella enterica serovar tiphi isolated from blood implications for therapy J Clin Microbiol 39 3583 3585 2001 10 Cercenado E et al Rapid antimicrobial susceptibility testing in patients with ventilator associated pneumonia direct E test on respiratory samples 42nd ICAAC Presentation D 51 2002 11 Navon Venecia S et al Direct testing of ESBL producing E coli and pneumoniae from blood cultures ICAAC 2003 presentation D 204 12 Paradisi F et al Evalutation of a rapid screening method for detection of antimicrobial resistance in the commensal E coli microbiota ICAAC 2003 presentation D 248 13 Weinbren M J et al Rapid detection of ESBL producing organisms in blood cultu
247. ol 42 1185 91 2004 K rp noja P et al Disc diffusion susceptibility testing of Haemophilus influenazae by NCCLS methodology using low strength ampicillin and co amoxyclav discs J Antimicr Chemother 53 660 3 2004 Ho P L et al Invasive H influenzae isolates with decreased levofloxacin susceptibility in Hong Kong JAC 57 366 2006 Bozdogan B et al Combination of altered PBPs and expression of cloned ESBLs confers cefotaxime resistance in H influenzae JAC 57 747 9 2006 Bogdanovich T et al Effect of efflux on telithromycin and macrolide susceptibility in H influenaza Antimicr Ag Chemother 50 893 8 2006 Cerquetti M et al First characterisation of heterogeneous resistance to imipenem in invasive nontypable H influenzae isolates Antimicr Agents Chemother 51 3155 61 2007 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 104 of 170 15 2 Susceptibility Testing of Gonococci The recommended agar medium for testing gonococci is Gonococcus Agar Base to which a 1 of a defined growth supplement is added after autoclaving The use of a cysteine free growth supplement is not required for diffusion testing Procedure 1 Use colonies taken directly from an overnight culture chocolate agar and make a suspension in broth or 0 9 saline equivalent to the 0 5 McFarland standard Use the suspension for plate inoculation within 15 min Not more than 9 Neo Sensitabs should be p
248. ome times 2 zones of growth appear around the CLINDA tablet An inner zone of light growth and an outer zone of confluent growth The inner zone is blunted near to the Ery tablet Report CLINDA resistant Constitutive MLS erm A R R R No hazy zone Growth up to both the ERY and erm A C CLINDA tablets MS M mef negative S R Clear susceptible zone around the CLINDA tablet msr A Report CLINDA susceptible Constitutive L lin A negative Y R S Clear susceptible zone around ERY No zone around CLINDA or reduced zone size 20 mm Report ERY susceptible and CLINDA resistant No resistance none S S S Clear susceptible zone diameter Copyright Rosco Diagnostica A S 09 2007 2008 Page 90 of 170 Chapter 13 NEO SENSITABS 13 4 MRSA Quality Control S aureus ATCC 43300 NEO SENSITABS POTENCY CODE Zone diameter in mm Amoxycillin Clavulanate 30415 ug AM CL 22 25 R b Cefoxitin 60 ug CFOXT 16 20 R Ciprofloxacin 10 ug CIP10 23 24 5 Erythromycin 78 ug ERYTR 9 zone Gentamicin 40 ug 40 16 22 I R Linezolid 30 ug LINEZ 26 32 S 8 Oxacillin lug OXA 1 9 nozone Tetracyclines 80 ug TET80 32 39 5 Trimethoprim 5 2 ug TRIME 24 31 S Vancomycin 5 ug VAN 5 18 23 S Vancomycin 70 ug VAN70 22 27 S a There is a zone around Oxacillin 1 ug but it is overgrown of thin colonies The zone should be read as 9 resistant b The strain is also resist
249. omycin Nitrofurantoin Achromobacter xylosoxidans Aminopenicillins 1 and 2 gen Cephalosporins Aminoglycosides Aztreonam Alc denitrificans Cefotaxime Burkholderia cepacia Aminopenicillins Ureidopenicillins Carboxypenicillins Amoxicillin Clavulanate 1 and 2 gen Cephalosporins Quinolones Aminoglycosides Polymyxins Nitrofurantoin Fosfomycin Chloramphenicol Chryseobacterium meningosepticum Aminoglycosides Carboxypenicillins 1 2 and 3 gen Cephalosporins Polymyxins Tetracyclines Chloramfenicol Ticarcillin Clavulanate Quinolones Ochrobactrum anthropi Ureidopenicillins Carboxypenicillins Ticarcillin Clavulanate 3 gen Cephalosporins Aztreonam Pseudomonas aeruginosa Aminopenicillins Amoxicillin Clavulanate 1 and 2 gen Cephalosporins Cefotaxime Ceftriaxone Chloramfenicol Nalidixic acid Trim Sulfa Tetracyclines Nitrofurantoin Stenotrophomonas maltophilia Listeria Neisseria Branhamella Ureidopenicillins Carboxypenicillins 1 and ord gen Cephalosporins Imipenem Cefotaxime Aztreonam Aminoglycosides Tetracyclines except Minocycline Fosfomycin Oxacillin Cephalosporins Aztreonam Polymyxins Nalidixic acid Clindamycin Fosfomycin Branhamella catarrhalis Lincomycin Clindamycin Trimethoprim Gonococci meningococci Campylobacter Helicobacter Lincomycin Clindamycin Polymyxins Trimethoprim Vancomycin Campylobacter s
250. one near the edge of the zone of cefoxitin cefotaxime ceftazidime and aztreonam The same procedure will be appropriate for K pneumoniae and P mirabilis strains Plasmid mediated AmpC beta lactamases MIC pg ml Beta lactamases Cefoxitin Ceftazidime Aztreonam Cefepime Imipenem Meropenem Microorganisms AAC 1 4 8 232 1 0 25 0 125 0 8 0 03 E coli pneumoniae mirabilis Salmonella C freundii ACT 1 gt 256 4 gt 128 4 gt 128 lt 0 06 8 1 E coli K pneumoniae E cloacae inducible BIL 1 CMY 2 R 16 4 16 1 0 5 0 06 E coli CMY 1 256 4 128 32 0 25 4 lt 0 5 0 06 K pneumoniae CMY 2 32 256 32 128 16 64 0 5 4 lt 0 5 0 06 E coli Salmonella K pneumoniae CMY 3 128 64 32 256 1 0 25 0 03 P mirabilis CMY 4 8 gt 256 8 256 0 5 32 0 06 4 0 25 0 125 E coli Salmonella P mirabilis CMY 5 R 256 64 0 5 1 K oxytoca CMY 6 256 256 64 0 5 0 25 0 06 E coli CMY 7 R gt 32 I 0 25 lt 2 E coli Salmonella CMY 8 gt 256 32 64 0 25 0 5 K pneumoniae CMY 9 gt 128 128 8 0 25 0 5 0 06 E coli CMY 10 128 8 64 4 128 0 12 0 5 0 25 0 5 lt 0 125 E coli E aerogenes pneumoniae CMY 11 gt 256 256 128 E coli CMY 12 256 128 8 32 16 0 25 4 0 5 P mirabilis CMY 13 512 256 64 1 0 25 lt 0 03 E coli inducible CMY 14 128 128 16 32 0 5 32 0 25 2 0 06 P mirabilis CMY 15 512 128 8 32 0 25 8 0 25 16 4 P mirabilis CMY 16 232 232 1 2 2 0 05 P mirabilis Synergy TAZO FEP
251. or cefepime and or aztreonam should be tested for the presence of ESBLs Procedure Apply Ceftazidime Cefepime and Aztreonam Neo Sensitabs At a distance of approx 15 mm edge to edge apply Ticarcillin Clavulanate Neo Sensitabs Separately apply Ceftazidime Clavulanate and Cefepime Clavulanate Neo Sensitabs Interpretation A key hole zone or ghost zone between Ticarcillin Clavulanate and any of Ceftazidime Cefepime or Aztreonam Neo Sensitabs indicates the presence of an ESBL With the combination disks a 5 mm larger zone for Ceftazidime Clavulanate and or Cefepime Clavulanate compared to the single antimicrobials indicates the presence of an ESBL The prediffusion procedure with Boronic acid may also be used 30 when ESBLs can be obscured by the chromosomal AmpC cephalosporinase in P aeruginosa Detection of ESBLs in different strains The presence of ESBLs may be masked by the overexpression of AmpC beta lactamases or by the induction of AmpC beta lactamase by clavulanate used in synergy tests ESBLs may be confused with enzymes such as K oxytoca chromosomal B lactamase K1 Laboratory staff must be aware of the increasing array of different resistance mechanisms and phenotypes O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Page 129 of 170 Chapter 16 1 E coli Klebsiella spp Salmonella spp Proteus mirabilis Shigella sonnei ESBL VEB 1 ESBL 25 AmpC plasmid no ESBL
252. orinases or AmpC beta lactamases are produced by Enterobacter cloacae E aerogenes Serratia marcescens Citrobacter freundii Hafnia alvei Providencia stuartii and Morganelli morganii and they are inhibited by aztreonam but not by clavulanic acid sulbactam or tazobactam Resistant mutants with high beta lactamase activity are present at a high frequency As a result therapy with cephalosporins except fourth generation agents and monobactams may fail because of selection of such mutants The tablet approximation test is useful to demonstrate the presence of inducible cephalosporinases during routine antibiogram testing Neo Sensitabs containing an inducer e g cefoxitin or imipenem and indicators such as piperacillin tazobactam cefotaxime or ceftazidime are placed approx 20 25 mm apart center to center A wider spacing 30 mm may be preferable for e g M morganii and Providencia spp Following overnight incubation at 35 C in air the presence of an inducible beta lactamase is indicated by the blunting of the zone of inhibition around the indicator drug piperacillin tazobactam cefotaxime ceftazidime adjacent to the inducer cefoxitin imipenem Dunne et al 1 have shown that the combination Imipenem and Piperacillin Tazobactam has the highest sensitivity 97 1 96 followed by Imipenem and Ceftazidime 94 2 96 The result should be reported as R resistant for penicillins except temocillin penicillin inhibitor combinations
253. orins Aminoglycosides in vivo Yersinia enterocolitica Gram positive cocci in general Aminopenicillins Carboxypenicillins Amoxicillin Clavulanate 1 and 2 gen Cephalosporins Cefoxitin Aztreonam Nalidixic acid Polymyxins Staphylococci Staphylococcus spp in generel Nalidixic acid Polymyxins S saprophyticus Novobiocin Fosfomycin Penicillin resistant staphylococci Oxa S Penicillin Aminopenicillins Ureidopenicillins Carboxypenicillins Methicillin resistent staphylococci All beta lactams Micrococcus spp Nitrofurantoin Mupirocin O Copyright Rosco Diagnostica A S Chapter 17 NEO SENSITABS BACTERIA Streptococci enterococci 09 2007 2008 Chapter 17 Page 154 of 170 NATURAL RESISTANCE Streptococcus spp Polymyxins Nalidixic acid Aminoglycosides low level Enterococcus faecalis Cephalosporins Clindamycin Mupirocin Aminoglycosides low level HLR test Novobiocin Trim Sulfa in vivo E faecium Cephalosporins Aminoglycosides low level HLR test Nitrofurantoin Trim Sulfa in vivo E gallinarum casseliflavus Vancomycin MIC 4 16 ug ml Arcanobacterium spp Bacitracin Mupirocin Optochin Pediococcus Leuconostoc Lactobacillus Erysipelothrix Non fermenters Glycopeptides Acinetobacter baumanii calcoaceticus Aminopenicillins 1 and Dm gen Cephalosporins Chloramphenicol Trimethoprim Fosf
254. ould be reported as clindamycin resistant NCCLS 2004 Inducible clindamycin resistance is not detected by current susceptibility test methods including automatic methods When using Erythromycin 15 ug and Clindamycin 2 ug the distance between disks must be lt 15 mm 40 Plate 13 3 a Plate 13 3 b Demonstration of the presence of inducible clindamycin resistens in Staphylococcus aureus inducible MLS resistance Note the flattening of the Clindamycin zones Plate 13 3 b near the Erythromycin Neo Sensitabs ERYTR and the big circular zone Plate 13 3 a around the Clindamycin Neo Sensitabs CLIND when not induced CA MRSA are often susceptible to non beta lactam drugs althoug they may show resistance to macrolides by an efflux mechanism These strains should be tested for inducible clindamycin resistance 27 Differentiation of phenotypes of macrolide resistance in staphylococci may be possible by the following criteria Phenotypes of macrolide resistance in staphylococci 34 35 Phenotype Genotype Induction CLINDA ERY Remarks test type Inducible MLS erm A D S gt R Blunted D shaped clear zone around CLINDA tablet near the ERY tablet Report CLINDA resistant Inducible MLS erm C D S gt R Blunted D shaped zone around CLINDA tablet near the ERY tablet and small colonies growing into the CLINDA zone in an otherwise clear zone Report CLINDA resistant Constitutive MLS erm A Hazy D R R S
255. ours at room temperature the tablet disc is removed by knocking the plate agianst the table and the plate is maintained at room temperature for further 18 hours overnight The plate is now inoculated with the strain to be tested using a McFarland 0 5 inoculum Additional antibiotic discs Neo Sensitabs may be added now using a dispenser if MH agar is used and thereafter the plate is incubated overnight at 35 37 The zones of inhibition are then measured Zone breakpoints are tentative and for research use only Notice In the laboratory the prediffusion plate can be prepared the day before it is inoculated in which case there is no loss of time and results are obtained within 24 hours Interpretation IA Detection of Visa GISA hVISA strains medium BHI 5 blood inoculum McF 0 5 VISA GISA hVISA strains will show the following zones of inhibition Teicoplanin 30 ug inhibition zone lt 20 mm and or Vancomycin5 ug inhibition zone lt 22 mm IB Detection of GISCN hGISCN strains medium BHI 5 blood inoculum McF 0 5 Teicoplanin 30 ug inhibition zone lt 20 mm GISCN Glycopeptide intermediate staphylococci coagulase negative 2 Strains showing zones of inhibition 20 mm around Teicoplanin 30 ug should be reported as heteroresistant to both teicoplanin and vancomycin IC Detection of vanA vanB and vanC in enterococci Use vancomyicn 5 ug 24 18 h prediffusion MH agar and McF 0 5 inoculum Susceptible zone
256. performed rountinely by laboratories 5 The susceptibility of the strains tested is determined according to the interpretation table below Interpretative zones for Campylobacter Mueller Hinton Agar 5 Blood Inoculum McFarland 1 0 Incubation Campylobacter atmosphere catalysator Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Amoxycillin 30 ug AMOXY gt 28 27 24 lt 23 lt 2 gt 8 Amoxycillin Clavulanate 30 15ug AM CL 228 27 24 23 2 28 Ampicillin 33 ug AMP33 228 27 24 lt 23 lt 2 28 Azithromycin 30ug AZITR 223 22 20 lt 19 lt 1 gt 4 Cefotaxime 30 ug CFTAX gt 28 27 24 lt 23 lt 4 gt 16 Cephalothin identification 66 ug CLOTN gt 18 i lt 16 ID gt 32 Chloramphenicol 60 ug CLR60 gt 28 27 24 lt 23 lt 4 gt 16 Ciprofloxacin 8 10 ug CIP10 gt 24 23 21 lt 20 lt 1 gt 4 Ciprofloxacin 8 0 5 ug CIP L gt 16 15 13 lt 12 lt 1 gt 4 Clarithromycin 30 ug CLARI gt 23 22 20 lt 19 lt 1 gt 2 Clindamycin 25 ug CLIND gt 28 27 24 lt 23 lt 1 gt 2 Doxycycline 80 ug DOXYC gt 28 27 24 23 lt 2 gt 8 Erythromycin 8 78 ug ERYTR 222 21 15 lt 14 lt 8 232 Furazolidone 50 ug FURAZ gt 28 27 24 lt 23 lt 2 24 Gentamicin 40 ug GEN40 gt 28 27 24 lt 23 lt 2 gt 4 Imipenem 15 ug IMIPM gt 28 27 24 lt 23 lt 1 gt 2 Levofloxacin 5ug LEVOF gt 23 22 20 lt 19 lt 1 gt 2 Meropenem 10 ug MEROP gt 28 27 24 lt 23 lt 1 gt 2 Moxifloxacin 5ug MOXIF 226 25 23 22 lt 0
257. phalosporins and aztreonam Clavulanate is an inhibitor of class A carbapenemases and therefore synergy with imipenem is useful to detect these enzymes 1 2 3 4 5 The KPC family of enzymes confer greater resistance to third gen cephalosporins than to carbapenems 3 5 Carbapenemase IMI 2 is the first inducible and plasmid encoded carbapenemase Class D carbapenemases correspond to the enzymes classified as OXA types oxacillinase activity They hydrolyze imipenem and meropenem weakly and do not hydrolyze third gen cephalosporins and aztreonam although MICs against the later drugs are often increased due to the presence of other beta lactamases Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 139 of 170 Clavulanate is a progressive inhibitor of most OXA carbapenemases but not all The synergy test clavulanate and imipenem may have value for the detection of these enzymes KPC possessing Enterobacter spp and K pneumoniae were reported as falsely susceptible to carbapenems using automated systems Vitek MIC microdilution using standard inocula of 10 or 10 CFU ml did not detect carbapenem resistance while diffusion methods E test using inocula of 10 CFU ml detected resistance 5 7 12 pneumoniae intermediate or resistant to ertapenem or meropenem should be considered resistant to all carbapenems 7 Clinicians should be aware of the potential for clinical failure Class D OXA 55
258. pneumoniae gt 36 35 26 lt 26 lt 0 06 gt 0 5 Copyright Diagnostica A S Chapter 10 NEO SENSITABS Zone diameter 09 2007 2008 Chapter 10 Page 42 of 170 Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S R S R f Erythromycin 78 ug ERYTR j S pneumoniaelstrep gt 28 lt 28 lt 0 5 gt 0 5 Moraxella catarrhalis gt 32 31 22 lt 22 lt 0 5 gt 4 C jeikeium urealyticum gt 32 31 22 lt 22 lt 0 5 gt 4 f Clarithromycin 30 ug CLARI S pneumoniae strep test ERY Moraxella catarrhalis 226 25 17 lt 17 lt 0 5 gt 4 f Azithromycin 30 ug AZITR S pneumoniae strep test ERY Moraxella catarrhalis gt 26 25 21 lt 21 lt 0 5 gt 2 Telithromycin ug S pneumoniae strep gt 26 25 18 lt 18 lt 0 125 gt 1 Moraxella catarrhalis gt 24 23 18 lt 18 lt 0 5 gt 1 Clindamycin 25 ug CLIND S pneumoniae strep gt 32 31 26 lt 26 lt 0 5 gt 2 Chloramphenicol 10 ug CLR10 Haemophilus spp gt 23 22 20 lt 20 lt 2 gt 2 S pneumoniae gt 16 15 14 lt 14 8 gt 8 Chloramphenicol 60 ug CLR60 Haemophilus spp gt 34 33 30 lt 30 lt 2 gt 2 S pneumoniae gt 26 25 23 lt 23 lt 8 gt 8 Tetracyclines 80 ug TET80 Haemophilus spp gt 28 27 23 lt 23 lt 2 gt 2 S pneumoniae strep gt 28 27 23 lt 23 lt 2 gt 2 Moraxella catarrhalis gt 28 27 23 lt 23 lt 9 gt 2 Rifampicin 30 ug RIFAM pneumoniae strep gt 32 31 28 lt 28 lt 1 gt 1 Linezolid 30 ug LINEZ S pneumoniae strep gt
259. points inmm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R c Cephalothin 66 ug CLOTN gt 23 22 20 lt 19 8 232 c Cephradine 60 ug CFRAD gt 23 22 20 lt 19 lt 8 gt 32 Chloramphenicol 60 ug CLR60 225 24 21 lt 20 lt 8 gt 32 a Cinoxacin U 30 ug CINOX gt 16 15 14 lt 13 lt 16 gt 64 q Ciprofloxacin 10 ug CIP10 gt 20 19 17 lt 16 lt 1 24 Ciprofloxacin 05ug Salmonella spp gt 24 E lt 23 lt 0 06 gt 0 12 Clarithromycin 30 ug CLARI gt 18 17 15 lt 14 lt 2 gt 8 j Clindamycin 25 ug CLIND gt 26 25 23 lt 22 lt 0 5 24 Cloxacillin 500ug X CL500 Detection of plasmid mediated AmpC beta lactamases 5 Colistin 10 ug CO 10 2 18 h prediffusion gt 15 14 11 lt 10 lt 2 gt 8 v Daptomycin 30 ug DAPCa Staphylococcus spp 2 18 h prediffusion 222 lt 1 Enterococcus faecalis S 2 18h pred 212 lt 4 Doxycycline 80 ug DOXYC 220 19 17 lt 16 lt 4 gt 16 q Enrofloxacin Vet 10 ug ENROF gt 23 22 17 lt 16 lt 0 5 24 c Ertapenem 10 ug ERTAP gt 19 18 16 lt 15 lt 2 gt 8 j Erythromycin 78 ug ERYTR gt 26 25 19 lt 18 lt 0 5 gt 8 Flumequine Vet 30 ug FLUME 220 19 17 lt 16 I Fosfomycin U 70 40 ug FOSFO gt 16 15 14 lt 13 lt 64 gt 256 i Fucidin 100ug FUCID 228 27 24 lt 23 lt 1 gt 4 Furazolidone 50 ug FURAZ gt 23 22 20 lt 19 lt 4 gt 8 q Gatifloxacin 5ug GATIF gt 18 17 15 lt 14 lt 2 gt 8 Staphylococcus spp gt 23 22 20 lt 19 lt 0 5 gt 2 r Gent
260. possible that qnrB contributes to the widespread sistribution of DHA 1 plasmid mediated AmpC in areas where 3rd generation cephalosporins and fluoroquinolones are widely used O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 17 Page 150 of 170 Screening procedure Perform antibiogram as usual standard procedure MH agar inoculum McF 0 5 incubation at 35 37 C for 18 24 hours Strains of Enterobacteriaceae should be suspected of plasmid mediated quinolone resistance when showing unusual multiresistance phenotypes such as Neo Sensitabs Ampicillin 33 ug no zone HLR Sulphonamides no zone HLR Trimethroprim no zone HLR Trimethroprim Sulfa no zone HLR Streptomycins no zone HLR Nalidixan no zone or zone lt 20 mm Ceftazidime Zone lt 20 mm Chloramphenicol may show resistance Gentamicin may show resistance Tetracyclines may show resistance Suspected strains can be tested for the presence of the Qnr gen by PCR It should be noted that strains showing the above mentioned resistance phenotypes are most probably integron carrying Enterobacteriaceae and barrier precaution should be established to prevent further spread In a selected group of ciprofloxacin and ceftazidime resistant Enterobacteriaceae mainly K pneumonae and E cloacae carriage of QnrA gene was 32 9 From those 73 were ESBL positive References 1 Minggui Wang et al Emerging plasmid mediated qu
261. pp U gt 26 25 22 lt 22 lt 1 gt 1 Others Nitrofurantoin U 260ug NITRO 224 lt 24 lt 32 gt 32 Sulfonamides U 240ug SULFA Enterobacteriaceae gt 26 25 20 lt 20 lt 64 gt 128 Staphylococcus spp Trimethoprim U 5 2 ug TRIME gt 20 19 16 lt 16 lt 2 gt 4 Trimethoprim Sulfa 5 24240 ug TR SU 228 27 23 lt 23 lt 2 38 gt 8 152 Colistin Polymyxins 150ug C0150 220 lt 20 y Colistin 10 pg CO 10 2 2 18 h prediffusion 215 15 lt 2 22 y Tigecycline 15 ug TIG15 Enterobacteriaceae 223 23 lt 1 gt 2 Staphylococcus spp gt 25 25 lt 0 5 gt 0 5 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 47 of 170 Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S R Fucidin 100ug FUCID r Staphylococcus spp gt 32 lt 32 lt 0 5 gt 0 5 Rifampicin 30 ug RIFAM Staphylococcus spp gt 32 lt 32 lt 1 gt 1 Enterococcus spp Linezolid 30 ug LINEZ Staphylococcus spp 226 25 22 22 lt 4 gt 4 Enterococcus spp Quinupristin Dalfopristin 15 ug SYNI5 5 Staphylococcus spp gt 18 17 16 lt 16 lt 2 gt 2 Enterococcus spp U urine Remarks a With these MIC breakpoints the normal population of E coli is categorized as intermediate to ampicillin amoxycillin Klebsiella spp should always be reported as resistant to ampicillin no matter the size of the inhibition zone b Reduced sensitivity to ampicillin
262. pp Vancomycin Trimethroprim Polymyxins Lincomycin Novobiocin Aztreonam Helicobacter pylori Vancomycin Polymyxins Nalidixic acid Trimetoprim Sulfonamides Corynebacterium in general Fosfomycin Mupirocin Polymyxins Nalidixic acid C jeikeium urealyticum Anaerobes in generel All Penicillins 1 2 and 3 gen Cephalosporins Amoxicillin Clavulanate Imipenem Meropenem Aminoglycosides Chloramphenicol Nalidixic acid Trim Sulfa Polymyxins Fosfomycin Mupirocin Macrolides Aminoglycosides Aztreonam exept Fusobacteria Trimethoprim Nalidixic acid Bacteroides fragilis group Aminoglycosides Vancomycin Aminopenicillins 1 and 2 gen Cephalosporins Polymyxins Glycopeptides Fosfomycin Aztreonam Oxgall Clostridium spp Kanamycin Trimethrprim Aztreonam Polymyxins Fosfomycin Fusobacteria spp Nalidixic acid Vancomycin Macrolides low level Porphyromonas spp Polymyxins Fosfomycin Aminoglycosides Prevotella spp Glycopeptides Fosfomycin Aminoglycosides O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 17 Page 155 of 170 NATURAL RESISTANCE Actinomyces Propionibacterium 1 and 2 gen Cephalosporins Polymyxins Metronidazole Mobiluncus spp Metronidazole Peptostreptococcus Eubacterium Polymyxins Fosfomycin Veillonella spp Macrolides low level Glycopeptides References 1
263. ptible to Amoxycillin Clavulanate Ticarcillin Clavulanate or Piperacillin Tazobactam do not report it as susceptible because resistant mutants may be selected during therapy Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 128 of 170 For Q C use Klebsiella pneumoniae ATCC 700603 zone of Ceftazidime Clavulanate and Cefepime Clavulanate is gt 5 mm larger than Ceftazidime Cefepime Neo Sensitabs see chapter 16 2 Detection of ESBLs using Neo Sensitabs ENTEROBACTERIACEAE Strains showing cefotaxime and or ceftazidime MICs gt 1 ug ml showing reduced susceptibility to amoxicillin clavulanate should be tested further for the presence of ESBLs Procedure Mueller Hinton agar plates are inoculated with the strain to be tested and Neo Sensitabs applied onto the agar Cefotaxime Ceftazidime and Cefepime Neo Sensitabs at a distance of 15 20 mm edge to edge from Amoxycillin Clavulanate Neo Sensitabs or using their combinations Cefotaxime Clavulanate Ceftazidime Clavulanate and Cefepime Clavulanate Neo Sensitabs Interpretation A key hole or ghost zone between Amoxycillin Clavulanate and any of Cefotaxime Ceftazidime or Cefepime Neo Sensitabs indicates the presence of an ESBL When using the combination disks 2 5 mm larger zone for any of the combinations compared to the corresponding single antimicrobial indicates the presence of an ESBL Cefpodoxime and Cefpodoxime Clavul
264. r Ceftazidime Clav 2 Aminoglycosides Kanamycin 100 ug Amikacin and Isepamicin APH 35 ANT 4 Staphylococci zone lt 25 mm resistance Gentamicin 40 ug Resistance to aminoglycosides APH 2 AAC 6 Staphylococci zone 23 mm except streptomycin Kanamycin 500 ug zone 14 mm HLR to amikacin no synergy with penicillins APH 3 ANT 4 Enterococci HLR Gentamicin 250 ug zone lt 14 mm HLR to all aminoglycosides APH 2 AAC 6 Enterococci HLR Streptomycin 500 ug zone 14 mm Streptomycin resistance Enterococci HLR Amikacin Tobramycin Resistance to aminoglycosides APH 3 VI Acinetobacter Netilmicin Tobramycin Resistance to aminoglycosides AAC 3 Pseudomonas 3 Others Erythromycin Inducible MLS resistance Ribosomal Staphylococci Clindamycin antagonism methylation Streptococci Nalidixan Reduced sensitivity to DNA gyrase Enterobacteriaceae zone lt 25 mm quinolones Vibrio cholerae Haemophilus Nalidixan Reduced sensitivity to DNA gyrase Gonococci zone lt 28 mm quinolones Meningococci Ciprofloxacin 0 5 ug Quinolone resistance DNA gyrase Gonococci zone lt 20 mm Haemophilus Vancomycin 5 ug Vancomycin resistance Van A Van B Enterococci Teicoplanin 30 2 18 hours prediffusion VISA hVISA Staphylococci Metronidazole Imidazole resistance Reductase Anaerobes a for non beta lactamase producing enterococci b Synergy AM
265. r Hinton plain or 5 blood Inoculum McFarland 0 5 Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R a Amoxycillin 30 ug AMOXY compare zone to AMC beta lactamase Amoxycillin Clav 30 15 ug AM CL gt 24 lt 23 4 2 8 4 a Ampicillin 33 ug AMP33 compare zone to AMC beta lactamase a Ampicillin 2 5 ug AMP L gt 26 25 21 lt 20 lt 0 12 gt 1 Azithromycin 30 ug AZITR 220 19 17 16 2 gt 8 Cefaclor 30 ug CCLOR 220 19 17 lt 16 lt 8 gt 32 Cefotaxime 30 ug CFTAX gt 30 lt 29 lt 2 Ceftazidime 30 ug CEZDI gt 30 lt 29 lt 2 Ceftriaxone 30 ug CETRX gt 30 lt 29 lt 2 Cefuroxime 60 ug CEFUR gt 28 27 24 lt 23 lt 4 gt 16 Cefuroxime oral 60 ug CEFUR gt 32 31 27 lt 26 lt 1 gt 4 Chloramphenicol 60 ug CLR60 232 31 27 26 lt 2 gt 8 Chloramphenicol 10 ug CLR10 gt 20 19 17 lt 16 lt 2 gt 8 Ciprofloxacin 10 ug CIP10 gt 24 lt 23 lt 1 x Clarithromycin 30 ug CLARI gt 20 19 17 lt 16 lt 2 gt 8 Clindamycin 25 ug CLIND gt 28 27 24 lt 23 lt 0 5 gt 4 Doxycycline 80 ug DOXYC gt 27 26 23 lt 22 lt 2 28 Erythromycin 78 ug ERYTR gt 28 27 21 lt 20 lt 0 5 28 Gatifloxacin 5 ug GATIF gt 30 29 27 lt 26 lt 0 25 20 5 Gentamicin 40 ug GEN40 gt 28 27 24 lt 23 lt 2 28 Imipenem 15 ug IMIPM gt 26 25 21 lt 20 lt 1 24 Levofloxacin 5ug LEVOF 222 21 lt 2 Meropenem 10 ug MEROP gt 24 23 21 lt 20 lt 1 gt 2 Moxifloxacin 5 ug MOXIF gt 30 29 27 lt 26 lt
266. r comparative zone results with Imipenm EDTA due to the intrinsic antibacterial activity of EDTA difference of 2 5 mm larger zone with Ceftazidime DPA compared to Ceftazidime alone indicates the presence of a MBL Strains showing no zone of inhibition around Ceftazidime Neo Sensitabs and a zone of inhibition 2 13 mm around Ceftazidime DPA Neo Sensitabs synergism should indicate the presence of a MBL e Plate 16 6 a Pseudomonas aeruginosa CCUG 51971 producing metallo beta lactamases Note the large zone around Imipenem EDTA Neo Sensitabs IM ED reflecting the production of metallo beta lactamases and no zone around Imipenem Neo Sensitabs IMIPM visible at the left and right side of IM ED there is synergism between both Imipenem Neo Sensitabs different potencies and the EDTA component of the combination of IM ED Plate 16 6 b Plate 16 6 c Demonstration of the presence of metallo beta lactamases in Pseudomonas aeruginosa CCUG 51971 using discs with dipicolinic acid D P A the combination ceftazidime plus D P A or D P A alone Plate 16 6 b Note the large zone around Ceftazidime D P A CAZ D and no zone around Ceftazidime CAZ30 reflecting the production of metallo beta lactamases Plate 16 6 c Note the synergism between Ceftazidime CAZ30 and D P A and no zone around Ceftazidime CAZ30 or D P A reflecting the production of metallo beta lactamases O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 20
267. ransfer by conjugation 0 0 0 Resistance induced by Vancomycin 0 or ND 0 Teicoplanin 0 ND 0 Location Chromosome Chromosome Microorganisms E gallinarum E casseliflavus Lactobacillus detected E flavescens Leuconostoc spp Pediococcus spp E rhusiopathiae Lactococcus spp If vancomycin is used for serious enterococcal infections such as endocarditis combined therapy with an amino glycoside is usually indicated 4 Hospital outbreaks of VRE van A van B are almost exclusively caused by a specific genogroup of VR E faecium that can easily be characterized by co resistance to ampicillin Vanco I R Ampi R and the presence of the variant esp gene 18 Ampicillin resistance MIC gt 16 ug ml and or zone 20 mm with Ampicillin 33 ug Neo S appears to bea specific and sensitive marker for this genogroup In vivo transfer of a Van A resistance gene from an animal isolate of E facecium into a human isolate of E faecium has been achieved in the intestines of human volunteers 23 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 14 Page 95 of 170 14 3 High level Aminoglycoside Resistance HLR High level resistance to aminoglycosides is an indication that an enterococcal isolate will not be affected synergistically by a combination of a penicillin or glycopeptide plus an aminoglycoside 4 Screening for high level gentamicin and streptomycin resistance should be performed on enterococcal isolates f
268. ration cephalosporins Include Cefepime and Cefpirome Neo Sensitabs O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 8 Page 23 of 170 Penems and Carbapenems Imipenem and Meropenem Neo Sensitabs have a very broad spectrum of activity and high stability against beta lactamases New members of this group are Ertapenem and Doripenem Monobactams Aztreonam is active only against gram negative aerobic bacteria Macrolides Streptogramines Ketolides Drugs in the macrolide group Azithromycin Clarithromycin Erythromycin are closely related and with few exceptions only Erythromycin may need to be tested routinely Clindamycin and Lincomycin have a similar spectrum of activity against aerobes and only one needs to be tested routinely Against anaerobes clindamycin has a broader spectrum of activity than lincomycin Quinupristin Dalfopristin Synercid Neo Sensitabs and Telithromycin Neo Sensitabs ketolid should be tested separately Oxazolidinones Linezolid Neo Sensitabs has activity against gram positive bacteria Aminoglycosides This class includes members affected by aminoglycoside inactivating enzymes which results in some differences in spectrum between the agents Include Streptomycins 100 ug Kanamycin 100 ug Neomycin Amikacin Gentamicin 40 ug Netilmicin Isepamicin and Tobramycin Neo Sensitabs Besides Streptomycins 500 ug Kanamycin 500 ug and Gentamicin 250 ug Neo Sensitabs for testing
269. re J Antimicr Chemother 55 131 2 2005 14 Zebruh M et al Evaluation of a new E test method for antimicrobial susceptibility testing of P aeruginosa isolates from cystic fibrosis Pathologie Biologie 53 490 4 2005 French 15 Kronvall G et al Extended antimicrobial resistance screening of the dominant faecal E coli and of rare resistant clones Intl J Antimicrobial Agents 26 473 8 2005 16 Bartolini A et al Multidrug resistant commensal E coli in children Peru and Bolivia Emerg Infect Dis 12 907 913 2006 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 6 Page 17 of 170 6 Inoculum Standardization and Prediffusion Method 6 1 Inoculum Standardization ICS and CLSI Kirby Bauer The specimen must be fully typical of the site of infection i e every effort should be made to obtain a representative sample of the relevant pathogenic bacteria The density of the inoculum is one of the variables that will have a great influence on the size of the zones of inhibition With primary cultures the inoculum may vary to a large degree i e from few colonies to a dense growth Our studies with Neo Sensitabs have shown that too sparse an inoculum will give too large zones and too dense an inoculum will give too small zones compared to the interpretation charts 6 1 1 Inoculum according to ICS 1 When secondary pure cultures are used the inoculum may be standardized as recommended by the IC
270. reduced susceptibility to carbapenems in Enterobacteriaceae they may remain underestimated because they are not detected in the laboratory Acquired carbapenemases are increasingly reported worldwide and consequently it is important to be able to detect them in the laboratory Carbapenemases classification 1 MICs pg ml Inhibited by Ambler 3rd gen classification Enzymes cepha AZTRM IMIPM MEROP CLAV EDTA Inducible A NmcA S 4 gt 16 2 8 wk no yes Sme 1 to Sme 3 S 4 64 gt 16 0 25 8 wk no yes IMI 1 to IMI 2 S S gt 64 4 32 no yes KPC 1 to KPC 4 gt 32 gt 64 4 16 4 16 Tor wk no no GES 2 to GES 5 232 163R 0 2516 0 5 16 0 1 16 gt 32 SOR 0 5 128 0 25s3R no yes no Metallo beta VIM 1 12 gt 64 SOR 15R 0 53R no yes no lactamases SPM 1 gt 256 4 R R no yes no GIM 1 16 32 8 16 28 28 no yes no SIM 1 2256 128 8 16 16 no yes no D OXA 23 27 gt 256 gt 256 4 64 4 128 wk no Oxacillinases OXA 40 48 SOR SoR 2 64 0 25 64 wk no no OXA 54 55 S S 4 0 25 wk no OXA 60 S R 0 5 2 no no yes OXA 58 4 128 232 3 32 2364 no no no wk weak References 1 Nordmann P et al Emerging carbapenemases in gram negative aerobes Clin Microbiol Infect 8 321 331 2002 16 6 1 Detection of acquired carbapenemases Ambler classes A and D Class A carbapenemases are penicillinases with greater activity against imipenem than meropenem and they also give resistance to penicillins ce
271. reduced susceptibility to ciprofloxacin MIC gt 0 125 ug ml show decreased susceptibility to all quinolones Meningococci g Oxacillin 1 ug or 5 ug are used routinely for the detection of reduced sensitivity to penicillins in meningococci chromosomal resistance h Rifampicin Used for prophylaxis only not treatment 1 Nalidixan is useful to screen for strains with reduced susceptibility to quinolones Gonococci k Nalidixan is useful to detect strains with reduced susceptibility to quinolones Ciprafloxacin resistant gonococci should presumably be resistant to all quinolones A positive beta lactamase test predicts resistance to penicillin amoxycillin ampicillin piperacillin and ticarcillin D Oxacillin 1 ug and 5 ug Neo Sensitabs are useful to detect betal lactamase negative gonococci with decreased susceptibility to penicillin chromosomal resistance Campylobacter For Campylobacter spp the absence of zone of inhibition around B lactams aminoglycosides macrolides or quinolones indicates high level resistance Anaerobes Vancomycin 5 ug Kanamycin 500 ug and Colistin 10 Neo Sensitabs are very useful for the identification of the most important gram negative bacilli B fragilis group are resistant to Vancomycin 5 ug Kanamycin 500 ug and Colistin 10 ug Prevotella is resistant to Kanamycin 500 ug and Vancomycin 5 ug zone 18 mm while it is variable to Colistin 10 ug Porphyromonas is sensitive to Vancomycin 5 ug zo
272. reus 216 15 14 lt 13 1 22 Coag neg staph 220 lt 19 lt 0 25 gt 0 5 i a Ampicillin 33 ug AMP33 228 27 21 lt 20 lt 2 gt 16 i a Amoxycillin 30 ug AMOXY 228 27 21 lt 20 lt 2 gt 16 a Ticarcillin 75 ug TICAR gt 28 27 21 lt 20 lt 8 gt 64 a Piperacillin 100ug PIPRA gt 28 27 21 lt 20 lt 4 gt 64 b Ampicillin Sulbactam 30 30 ug AM SU gt 28 27 21 lt 20 lt 248 gt 16 8 b Amoxycillin Clav 30415 ug AM CL gt 28 27 21 lt 20 lt 242 21642 b Ticarcillin Clavulanate 75415 ug TI CL 228 27 21 20 lt 842 26442 b Piperacillin Tazobactam 100 10ug PI TZ gt 28 27 21 lt 20 lt 444 gt 64 4 b Cefaclor 30 ug CCLOR gt 30 29 24 lt 23 lt 1 gt 8 b Cefuroxime oral 60 ug CEFUR 232 31 24 23 1 28 b Cefuroxime parenteral 60 ug CEFUR 228 27 21 lt 20 lt 4 gt 16 g b Cefpodoxime 30 ug CFPOX 230 29 24 23 1 gt 8 b Cefixime 30 ug CFFIX gt 30 29 27 lt 26 lt 1 gt 4 b Cephalothin 66 ug CLOTN gt 30 29 21 lt 20 lt 2 gt 16 b Cefazolin 60 ug CFZOL gt 28 27 21 lt 20 lt 4 gt 16 b Cefoxitin 60 ug CFOXT gt 28 27 21 lt 20 lt 4 gt 16 S aureus gt 30 29 28 lt 27 OxaS Apos Coag neg staph gt 34 33 32 lt 31 OxaS Ceftazidime 30 ug CEZDI gt 26 25 19 lt 18 lt 4 gt 32 b Cefotaxime 30 ug CFTAX 228 27 21 lt 20 lt 2 gt 16 b Ceftizoxime 30 ug CEZOX gt 28 27 21 lt 20 lt 2 gt 16 b Ceftriaxone 30 ug CETRX gt 26 25 19 lt 18 lt 4 gt 32 b Cefepime
273. rfor aminoglykosider og bgr kun testes for HLR high level resistens overfor Gentamicin 40 ug og Streptomycins 100 ug Neo Sensitabs Gentamicin 40 ug zone lt 20 mm HLR MIC gt 500 ug ml Streptomycins 100 ug zone lt 20 mm HLR MIC gt 1000 pg ml For enterokokker anvendes Vancomycin 5 ug og Mueller Hinton Agar uden blod med kraftig inokulum McFarland 0 5 P Dansk Blod Agar kan man ikke p vise VRE van B Afl sning S gt 15 mm I 14 13 mm R lt 12 mm G lder ogs for 2418 t pr diffusion Ved testning af pneumokokker med Oxacillin 1 ug anvendes kraftig inokulum McFarland 0 5 Afl sning S gt 20 mm I R x 19 mm Hvis v ksten p pladen kun er semikonfluerende anvendes S gt 25 mm I R lt 24 mm Ved testning af Methicillin med Staph aureus anvendes S gt 28 mm R x 27 mm For Koagulase negative stafylokokker screening anvendes kraftig inokulum McFarland 0 5 og gt 28 mm R lt 27 mm Methicillin Neo S Hvis stammen KNS er multi resistent og methicillin f lsom p testes den med Oxacillin I ug p Mueller Hinton Agar uden blod og kraftigt inokulum McFarland 0 5 S gt 18 mm MIC x 0 25 pg ml R 17 mm MIC gt 0 5 ug ml Breakpoints fra NCCLS M2 A7 2000 Cefoxitin m testes sidel bende Enterobacter og Proteus spp b r rapporteres R mod Nitrofurantoin uanset zonest rrelse Bem rk nye MIC breakpoints for Kinoloner skandinavisk model Tidligere MIC breakpoints var fra CLSI
274. rom blood or CSF Special high content Neo Sensitabs Gentamicin 250 ug Kanamycin 500 ug and Streptomycin 500 ug are used to screen for this type of resistance Inhibition zones 14 mm indicates high level resistance HLR and zones gt 15 mm indicates a lack of HLR The test is performed on plain Mueller Hinton agar using McFarland 0 5 inoculum If the inoculum used results in semiconfluent growth the zone diameter breakpoint is 17 mm instead of 14 mm Aminoglycosides Zone diameters in mm Equivalent Break point High level resistant HLR MIC pg ml Gentamicin 250 ug 14 mm HLR gt 500 Kanamycin 500 ug 14 mm HLR 1000 Streptomycin 500 ug 14 mm HLR 1000 e Ifthe strain is HLR to streptomycin This aminoglycoside cannot be used in combination with a penicillin or glycopeptide e If the strain is HLR to kanamycin Then kanamycin isepamicin and amikacin cannot be used Ifthe strain is HLR to gentamicin Then the strain is HLR to all aminoglycosides gentamicin tobramycin sisomicin netilmicin kanamycin isepamicin and amikacin except streptomycin HLR to gentamicin in enterococci is known to be mediated by the bifunctional enzyme AAC 6 APH 2 E faecium is known to harbour a chromosomally mediated enzyme AAC 6 1 and consequently is naturally resistant to penicillin tobramycin penicillin netilmicin penicillin sisomicin and penicillin kanamycin synergy 6 For E faecium report HLR to all ami
275. romycin 30 ug CLARI gt 24 23 21 lt 20 lt 0 25 2 Clindamycin 25 ug CLIND gt 28 27 24 lt 23 lt 0 25 gt 1 Doxycycline 80 ug DOXYC gt 30 29 27 lt 26 lt 2 gt 8 Erythromycin 78 ug ERYTR gt 28 27 24 lt 23 lt 0 25 gt 1 h Fosfomycin 70 40 ug FOSFO 224 lt 23 lt 8 Gatifloxacin 5 ug GATIF gt 21 20 18 lt 17 lt 1 gt 4 d Imipenem 15 ug IMIPM gt 30 29 27 lt 26 lt 0 12 gt 1 Levofloxacin 5 ug LEVOF gt 18 17 15 lt 14 lt 2 gt 8 Linezolid 30 ug LINEZ gt 21 A lt 2 d Meropenem 10 ug MEROP gt 28 27 24 lt 23 lt 0 25 2 d Moxifloxacin 5ug MOXIF gt 18 17 15 lt 14 lt 1 gt 4 9 Norfloxacin 10 ug NORFX lt 12 Reduced susceptibility to quinolones Ofloxacin 10 ug OFLOX gt 20 19 17 lt 16 lt 2 gt 8 a Oxacillin lug OXA 1 220 19 lt 19 lt 0 06 gt 0 12 penicillin pen R pen Quinupristin Dalfopristin 15 ug SYNI5 gt 19 18 16 lt 15 lt 1 gt 4 e Rifampicin 30 ug RIFAM gt 28 27 24 lt 23 lt 1 gt 4 Teicoplanin 60 ug TEICO gt 18 E lt 1 Telithromycin 15 ug TEL15 gt 19 18 16 lt 15 lt 1 gt 4 Tetracyclines 80 ug TET80 gt 30 29 27 lt 26 lt 2 gt 8 Tetracyclines 10 ug TET10 220 19 17 16 lt 2 gt 8 Tigecycline 15 ug TIG15 gt 19 lt 0 25 Trimethoprim Sulfa 5 2 240 ug TR SU gt 32 31 27 lt 26 0 5 9 5 gt 4 76 5 ug VAN 5 gt 17 lt 1 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15
276. s C freundii S marcescens M morganii Prov rettgeri P stuartii Hafnia alvei except P vulgaris AMC S Aminopenicillins Amoxicillin Clavulanate 1 gen Cephalosporins Citrobacter freundii Aminopenicillins Amoxicillin Clavulanate 1 gen Cephalosporins Cefoxitin Citrobacter koseri diversus Aminopenicillins Carboxypenicillins Enterobacter aerogenes E cloacae Aminopenicillins Amoxicillin Clavulanate Cefoxitin 1 gen Cephalosporins Nitrofurantoin Klebsiella pneumoniae K oxytoca Aminopenicillins Carboxypenicillins Morganella morganii Aminopenicillins Amoxicillin Clavulanate 1 and 2 gen Cephalosporins Cefoxitin Polymyxins Tetracyclines Nitrofurantoin Proteus mirabilis Polymyxins Tetracyclines Nitrofurantoin Proteus vulgaris P penneri Aminopenicillins Carboxypenicillins Cefuroxime Polymyxins Tetracyclines Nitrofurantoin Providencia rettgeri Aminopenicillins Polymyxins Tetracyclines Nitrofurantoin Amoxycillint Clavulanate Providencia stuartii Aminopenicillins Amoxicillin Clavulanate Polymyxins Tetracyclines Nitrofurantoin Gentamicin Tobramycin Netilmicin Salmonella spp 1 and 2 gen Cephalosporins Cefuroxime active in vitro not active in vivo Aminoglycosides in vivo Serratia marcescens Aminopenicillins Amoxicillin Clavulanate 1 and 2 gen Cephalosporins Polymyxins Shigella spp 1 and 2 gen Cephalosp
277. s for Yeasts modified Shadomy agar The following zone size interpretation should be used with using this procedure with MIC breakpoints according the CLSI M44 A O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 21 Page 165 of 170 21 2 1 1 Interpretation according to CLSI breakpoints Shadomy Modified Agar Inoculum McF 0 5 diluted 1 1 flooding MIG breakpoints according to CLSI M44A Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE R S R Amphotericin B 10 ug AMPHO gt 15 14 10 lt 10 lt 1 gt 2 Caspofungin 5 ug CASP5 gt 15 14 12 lt 11 lt 2 22 Fluconazole 25 ug FLUCZ 222 21 15 DD x14 8 264 Fluorocytosine 1 ug FLU 1 220 19 12 lt 11 lt 4 lt 32 Fluorocytosine 10 ug FLU10 gt 30 29 23 lt 22 lt 4 lt 32 Itraconazole 8 ug ITRAC 223 22 14 DD lt 13 lt 0 12 gt 1 Ketoconazole 15 ug KETOC gt 30 29 23 lt 22 lt 0 12 20 5 Posaconazole 5ug POSAC 217 16 14 DD lt 13 lt 1 gt 4 Voriconazole lug VOR 1 gt 17 16 14 DD lt 13 lt 1 gt 4 DD Dosis dependent 21 2 1 2 Interpretation according to EUCAST MIC breakpoints MIC breakpoints according to EUCAST Shadomy Modified Agar Inoculum McF 0 5 diluted 1 1 flooding Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S R Amphotericin B 10 ug AMPHO gt 18 5 lt 0 25 Caspofungin 5 ug CASP5 gt 15 lt 1 Fluconazole 25 ug FLUCZ gt 28
278. s in Europe J Antimicrob Chemother 38 409 424 1996 6 Cormican M G et al Detection of extended spectrum beta lactamases ESBL producing strains by the E test ESBL Screen J Clin Microbiol 34 1880 1884 1996 7 Wahaboglu et al Resistance to extended spectrum cephalosporins casued by PER 1 beta lactamase in Salm typhimurium from Istanbul Turkey J Med Microbiol 43 294 299 1995 8 Karas J A et al Treatment failure due to extended spectrum beta lactamase J Antimicrob Chemother 37 203 204 1996 9 Performance standards for Antimicrobial Susceptibility Testing 15th Inf Suppl M 100 S15 2005 10 Vercauteren E et al Comparison of screening methods for detection of Extended Spectrum Beta Lactamases and their prevalence among blood isolates of E coli and Klebsiella spp in a Belgian Teaching Hospital J Clin Microbiol 35 2191 2197 1997 11 Thomson K S Controversies about extended spectrum and AmpC beta lactamases Emerg Infect Dis 7 March April 2001 12 Steward C D et al Characterization of clinical isolates of Kl pneumoniae from 19 laboratories using the NCCLS ESBL detection methods J Clin Microbiol 39 2864 2872 2001 13 de Gheldre Y et al National epidemiologic survey of Enterobacter aerogenes Belgian hospitals from 1996 to 1998 J Clin Microbiol 39 889 896 2001 14 Livermore D M Detection of beta lactamase mediated resistance J Antimior Chemother 48 Suppl S1 59 64 20
279. sceptibility Testing 8th Ed M2 A8 January 2003 2 del Cuerpo M et al Stability of beta lactam antibiotics in paper disks and tablets used in the diffusion test Rev Esp Quimioter 10 nr 3 1997 Spanish 3 Steward C D et al Antimicrobial susceptibility testing of carbepenems multicenter validity testing and accuracy levels of 5 antimicrobial test methods for detecting resistance in Enterobacteriaceae and Pseudomonas aeruginosa isolates J Clin Microbiol 41 351 358 2003 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 3 Page 9 of 170 NEO SENSITABS Range 3 1 Antibacterials Identification Diffusible code amount of Neo Sensitabs Neo Sensitabs Antimicrobial A Penicillins Penams PENICILLIN LOW PEN L 5ug AMPICILLIN 33 ug AMP33 33 ug AMPICILLIN 2 5 ug AMP L 2 5 ug AMOXYCILLIN AMOXY 30 ug METHICILLIN METHI 29 ug OXACILLIN 1 ug OXA 1 lug OXACILLIN 5 ug OXA 5 5 CLOXACILLIN 500 ug AmpC test Diatabs CL500 500 ug MECILLINAM Amdinocillin MECIL 33 ug TICARCILLIN TICAR 75 ug PIPERACILLIN PIPER 100 ug TEMOCILLIN TEMOC 30 ug B Beta lactam Beta lactamase Inhibitor Combinations AMOXYCILLIN CLAVULANATE AM CL 30 15 ug Augmentin AMPICILLIN SULBACTAM AM SU 30 30 ug TICARCILLIN CLAVULANATE TI CL 75415 ug Timentin PIPERACILLIN TAZOBACTAM PI TZ 100 10 ug CEFTAZIDIME CLAVULANATE CZ CL 30 10 ug CEFEPIME CLAVULANATE CP CL 30 10 ug IMIPENEM EDTA IM ED 15 750 ug BORONIC ACID Diata
280. smid mediated AmpC 6 14 Poirel et al 10 have shown in vivo selection of fluoroquinolone resistant E coli isolates expressing plasmid mediated quinolone resistance and ESBL and physical linkage between ESBL and QnrA encoding genes in the same integron Although QnrA QnrB QnrS produce low levels of quinolone resistance it facilitates selection for a high level of quinolone resistance OnrB another plasmid mediated gene for quinolone resistance has been discovered in plasmids encoding the ESBL CTA 15 from a K pneumoniae These strains show low level resistance to quinolones and MIC of 16 ug ml towards nalidixic acid and show similar multiresistance phenotypes as Qnr A containing strains 11 Lavigne et al 12 screened for qnr genes 112 clinical isolates of ESBL producing E coli from French hospitals in 2004 7 7 of CTX M producing E coli presented a plasmid mediated resistance to quinolones All strains harboured a qnrA gene located on a class 1 integron Poirel et al 13 listed 186 ESBL positive Enterobacteriaceae From them 2 2 and 1 6 96 carried a QnrA1 and a QnrS1 determinant respectively The association of the QnrA gene with class 1 integrons was confirmed Hyunjoo Pai et al 14 screened E coli and pneumoniae producing ESBLs or plasmid mediated AmpC beta Icatamases for the presence of qnrA and qnrB genes QnrB was present in 54 of 54 DHA 1 producing K pneumoniae isolates and 10 of 45 SHV 12 producing isolates It is
281. solate demonstrating extraordinary high level resistance against various aminoglycosides Antimicr Ag Chemother 50 178 184 2006 8 LeeS etal Dissemination of 16S rRNA methylase in AmpC producing E cloacae C freundii and S marcescens in Korea Clin Microbiol Infect 12 Suppl 4 P1274 2006 9 Bogaerts P et al Emergence of ArmA and RmtB aminoglycoside resistance rRNA methylases in Belgium JAC 59 459 464 2007 17 2 Screening for Plasmid mediated Quinolone Resistance QnrA QnrB QnrS Enterobacteriaceae Integrons The plasmid gene responsible for quinolone resistance QnrA QnrB QnrS is carried on class 1 integrons of the In 4 family an efficient mechanism for rapid horizontal and vertical dissemination og antibiotic resistance determinants among bacteria The plasmid mediated mechanisms have led to resistance to almost all clinical important antimicrobials such as B lactams aminoglycosides macrolides phenicols sulphonamides and trimethoprim The identification in the US of qnr in clinical strains of K pneumoniae isolates besides producing plasmidic B lactamases and ESBL s 7 and its discovery in strains of E coli from Southeast Asia and Salmonella in Hong Kong indicates the emergence of this new mechanism of quinolones resistance in clinical strains Itis important to indicate that a significant relation exists between quinolone resistance and resistance to 3 gen cephalosporins co resistance ESBL and or pla
282. spp gt 26 25 22 22 lt 2 gt 4 Streptococcus spp gt 22 21 18 lt 18 lt 4 gt 16 h N meningitidis 232 lt 0 25 Anaerobes 222 21 18 18 lt 4 gt 16 C2 Norfloxacin 10 ug NORFX Reduced susceptibility to quinolones S pneumoniae lt 12 gt 8 Ciprofloxacin 10 ug CIP10 Haemophilus spp 228 Use nalidixan i lt 0 5 gonorrhoeae gt 36 Use nalidixan lt 36 lt 0 06 gt 0 12 Campylobacter gt 28 27 23 lt 23 lt 0 5 gt 1 Helicobacter pylori gt 20 lt 20 1 gt 1 Ofloxacin 10 ug OFLOX Haemophilus spp 228 Use nalidixan lt 0 5 gonorrhoeae 234 Usenalidixan 34 0 12 gt 0 25 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 71 of 170 Zone diameter Concentrations inmm critiques NEO SENSITABS POTENCY CODE S l R S R Levofloxacin 5 ug LEVOF Haemophilus spp gt 24 Usenalidixan lt 1 z c2 S pneumoniae gt 18 lt 18 lt 2 gt 2 Streptococcus Spp gt 20 19 17 lt 16 lt 1 gt 2 Gatifloxacin 5ug GATIF Haemophilus spp gt 24 Use nalidixan lt 1 C2 S pneumoniae I Streptococcus spp gt 21 20 18 18 lt 1 gt 2 Moxifloxacin 5ug MOXIF Haemophilus spp gt 28 Use nalidixan lt 0 5 C2 S pneumoniae gt 24 lt 24 lt 0 5 gt 0 5 Streptococcus spp gt 24 23 21 lt 20 lt 0 5 gt 1 Anaerobes gt 21 20 18 18 lt 1 gt 2 Nalidixan 130ug NALID f Haemophilus spp 28 decreased susceptibility
283. spp and Cryptococcus neoformans with Posaconazole and Amphtohericin B J Clin Microbio 44 3616 22 2006 Espinel Ingroff A et al Correlation of Neo Sensitabs Tablet diffusion assay on 3 media with CLSI broth microdilution M27 A2 and Disk Diffusioin M44A methods for susceptibilty testing of Candida spp and Cryptococcus neoformans with Amphothericin B Caspofungin Fluconazole Itraconazole and Voriconazole In Press Carrillo Munoz A J et al Activity of Caspofungin and Voriconazole against clinical isolates of Candida and other medically important yeasts by the CLSI M44A disk diffusion method with Neo Sensitabs tablets Chemotherapy 2007 in press Espenel Ingroff A et al Multicenter evaluation of a new disk agar diffusion method for susceptibility testing of filamentous fungi with Voriconazole Posaconazole Itraconazole Amphothericin B and Caspofungin J Clin Microbiol 45 1811 20 2007 Espinel Ingroff A et al Standardized disk diffusion method for yeasts Clin Microbiol Newsletter 29 97 100 2007 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 22 Page 168 of 170 22 Veterinary Practice CLSI Interpretation of the Antibiogramme with Neo Sensitabs MIC break points according to CLSI M31 A3 2007 Inoculum according to Kirby Bauer confluent colonies Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S R S R Amoxicillin Clav 30 15 ug AM CL Staphylococcus spp g
284. standardization of the method showing that the interpretative zone diameters correspond to MIC values in agreement with internationally accepted standards The availability of regression lines makes it possible for the microbiologist to convert the results obtained by the diffusion method susceptible inter mediate resistant into more quantitative results i e approximated MIC values An accurate diffusion method requires well standardized technique including control strains e g Staphylococcus aureus ATCC 25923 Escherichia coli ATCC 25922 Pseudomonas aeruginosa ATCC 27853 and Enterococcus faecalis ATCC 29212 enabling control of inoculum and media variations Well traced regression lines which can be controlled because there is a correlation between the slope of the regression line and the molecular weight of the antimicrobial tested Separate regression lines for rapidly growing bacteria and slow growing species Regression lines for Neo Sensitabs have been prepared using the agar dilution method to determine the MIC values For the aminoglycosides amikacin gentamicin netilmicin tobramycin and polymyxins colistin we have also determined MIC s by the broth dilution method in order to correct for the amount of antimicrobial bound to the agar The above mentioned antimicrobials are strongly bound to the sulphate groups of the agar and consequently different MIC values as well as zone sizes may be obtained depending on the pur
285. t Ceftriaxone 30 ug CETRX 226 lt 0 5 Penicillin Low 5 ug PEN L gt 22 lt 0 12 Note The beta haemolytic group includes the large colony forming pyogenic strains of streptococci with group A S pyogenes C G antigens and strains with group B S agalactiae antigen Penicillin resistant strains of Group A and Group B streptococci have not yet been recognized Any strains found to be intermediate or resistant should be referred to a Reference Laboratory for confirmation NCCLS 2000 Group B streptococci are susceptible to penicillin ampicillin and cefazolin but may be resistant to clindamycin and or erythromycin When a group B streptococci is isolated from a pregnant woman with severe penicillin allergy clindamycin and erythromycin should be tested and reported See Double tablet induction test D Zone test page 117 CLSI recommend disk diffusion zone diameter interpretive standards for Ampicillin Amoxycillin and Penicillin against Beta haemolytic Streptococci Pen Low Neo Sensitabs S 2 22 mm Ampicillin 33 and Amoxycillin Neo Sensitabs S 2 28 mm Strains showing zones less than 22 mm Pen Low or less than 28 mm Ampi 33 Amox should be sent to a Reference Laboratory 1 CLSI recommend specific zone diameter interpretative criteria for Cefotaxime Ceftriaxone and Cefepime against Beta Haemolytic Streptococci Cefotaxime Neo Sensitabs S gt 26 mm MIC x 0 5 ug ml Ceftriaxone Neo Sensitabs S gt 26
286. t 26 lt 25 lt 4 2 gt 8 4 Other organisms 220 19 17 lt 16 lt 8 4 gt 32 16 Amikacin 40 ug AMIKA gt 20 19 17 lt 16 lt 16 gt 64 Amoxycillin 30 ug AMOXY Enterococcus spp 220 lt 19 lt 8 gt 16 Other organisms 220 19 17 lt 16 lt 8 232 Ampicillin 33 ug AMP33 Enterobacteriaceae 220 19 17 lt 16 lt 8 gt 32 Staphylococcus spp use Pen Low Enterococcus spp gt 20 lt 19 lt 8 gt 16 Streptococcus spp not pneumoniae gt 30 29 21 lt 20 lt 0 25 gt 8 Listeria monocytogenes 226 25 2 gt 4 Mannheimia haemolytica gt 32 31 26 lt 25 lt 0 25 gt 1 c Apramycin 40 ug APRAM 223 22 20 lt 19 4 gt 16 c Bacitracin 40U BACIT 220 19 17 lt 16 c e h Cefadroxil 30 ug CFDRO gt 23 22 20 lt 19 8 232 e h Cefazolin 60 ug CFZOL 223 22 20 lt 19 lt 8 232 f Cefoxitin 60 ug CFOXT S aureus 225 lt 24 OxaS MecA pos Coag neg staph gt 28 lt 27 OxaS MecA pos a c h Ceftriaxone 30 ug CETRX gt 20 19 17 lt 16 lt 8 gt 32 Cefoperazone Cefpodoxime 30 ug CFPOX 225 24 21 lt 20 lt 2 gt 8 c h Cefquinome 30 ug CFQUI gt 23 22 20 lt 19 lt 2 gt 8 h Ceftiofur 30 ug CFTIO 223 22 20 19 2 28 c h Cefuroxime 60 ug CEFUR 223 22 20 lt 19 lt 8 232 c e h Cephalexin 30 ug CFLEX gt 20 19 17 lt 16 8 gt 32 c e h Cephalothin 60 ug CLOTN 223 22 20 lt 19 lt 8 gt 32 Chloramphenicol 60 ug CLR60 S pneumoniae gt 28 27 lt 4 gt Streptoccoccus spp 228 27 21 20 lt 4 gt 1
287. t 28 mm Klebsiella Enterobacter and Proteus spp should be reported R to nitrofurantoin Carbapenem testing in Iso sensitest Agar may give falsely susceptibility for isolates that harbour metallo beta lactamases testing on Mueller Hinton Agar may be preferred BSAC 2001 Nalidixan is a good screening for the detection of decreased fluoroquinolone susceptibility in Salmonella spp Hakanen A et al J Clin Microbiol 37 3572 77 1999 Strains resistant to Nalidixan Neo S zone 28 mm show a decreased susceptibility to quinolones ciprofloxacin MIC 0 125 ug ml Strains showing zone 24 mm with Cefpodoxime Neo Sensitabs should be suspected of producing ESBL E coli Klebsiella Salmonella For confirmatory tests use Ceftazidime Clavulanate and Cefepime Clavulanate compared to Ceftazidime and Cefepime Neo Sensitabs see ESBL O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 40 of 170 j For the detection of methicillin oxacillin resistance in staphylococci use Mueller Hinton Agar Iso sensitest will not reliably detect resistance in these organisms BSAC 2001 Cefoxitin is very useful to detect mec A positive strains in staphylococci k Staphylococci resistant to gentamicin should be reported as resistant to netilmicin and tobramycin Gentamicin is recommended as the test drug for staphylococci because netilmicin might give false sensitive results with coagulase negative staphylococci Isola
288. t 20 lt 1 gt 2 Quinu Dalfopristin 15 ug SYNIS5 225 24 19 lt 18 lt 0 5 gt 2 Rifampicin 30 ug RIFAM Staphylococcus spp gt 28 27 24 lt 23 lt 0 5 gt 4 other gt 23 22 18 lt 17 lt 4 gt 16 Sparfloxacin 10 ug gt 23 22 20 lt 19 lt 1 gt 2 Spiramycin 200 ug SPIRA gt 26 25 23 lt 22 lt 1 gt 4 Streptomycin 100 ug 100 gt 25 24 21 lt 20 lt 8 gt 16 h Streptomycin HNR 500ug ST500 gt 14 lt 14 lt 250 gt 500 Sulphonamides U 240 ug gt 23 22 20 19 lt 100 gt 350 k s Teicoplanin 30 ug TPN30 gt 16 15 14 lt 14 lt 4 gt 8 2 18 hours prediffusion 22 lt 2 gt 4 VISA GISA Telithromycin 15 ug TELI5 21 20 17 lt 16 lt 0 5 gt 2 Temocillin 30 ug gt 18 17 15 lt 14 lt 16 232 Tetracyclines 80 ug TET80 223 22 20 lt 19 lt 4 gt 8 a Ticarcillin 75 ug TICAR gt 23 22 20 lt 19 lt 16 gt 64 d o TicarcillinzClavulanate 75 15 ug TI CL 223 22 20 lt 19 lt 16 2 gt 64 2 Tigecycline 15 ug TIG15 gt 19 18 15 lt 15 lt 2 gt 8 Tobramycin 40 ug TOBRA gt 24 23 21 lt 20 lt 2 gt 4 Staphylococcus spp gt 26 lt 26 lt 1 gt 1 P aeruginosa Acinetobacter spp 223 22 20 19 lt 4 gt 8 Trimethoprim U 5 2 ug TRIME gt 20 19 17 lt 16 lt 4 gt 8 i Trimethoprim Sulfa 5 2 240 ug TR SU gt 28 27 24 lt 23 lt 2 38 gt 8 152 k s Vancomycin 5 ug VAN 5 gt 16 lt 4 gt 8 2 18 hours prediffusion staph lt 20 gt 2 VISA GISA 2 18 hours predif
289. t mutations of PBP4 It should be emphasized that bactericidal therapy of serious enterococcal infections such as endocarditis can only be achieved with combined drug therapy high dose penicillin ampicillin or vancomycin aminoglycoside but combined therapy is not required for less serious infections e g UTI 4 14 2 Glycopeptide Resistance VRE Accurate detection of vancomycin resistant enterococci by the diffusion method requires the following e Use of Vancomycin 5 ug Neo Sensitabs on Mueller Hinton agar without blood and McFarland 0 5 inoculum e Use the 2418 hours prediffusion method e Examine carefully the vancomycin zone of apparent inhibition with transmitted light for evidence of small colonies or light film growing within the zone If found report resistance to vancomycin e Use the interpretation S gt 16 mm lt 16 mm e tis important to examine the zone edges when reading the zone diameters around Vancomycin Neo Sensitabs Vancomycin susceptible strains show sharp edges while vancomycin resistant strains show diffuse edges e See Quality Control Limits with E faecalis ATCC 51299 see chapter 14 5 Warning Vancomycin testing on Iso sensitest Agar and Danish Blood Agar will not detect vanB resistance and therefore should be avoided O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 14 Page 93 of 170 Resistance to glycopeptides in enterococci has been detected in di
290. t of the macrolide group The normal population of Haemophilus spp is categorized as intermediate to erythromycin The breakpoints are chosen in such a way that the normal population of pneumococci is categorized as intermediate to ciprofloxacin ofloxacin For detection of inducible MLSp resistance we refer to Neo Sensitabs User s Guide O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 52 of 170 10 5 V Danish Blood Agar Interpretation Valid for Denmark Va Danish blood agar Interpretation valid for Denmark Afl sningsskema for Neo Sensitabs hurtigvoksende bakterier DIREKTE METODE Resistensplade DBA Semikonfluerende v kst 3 gt 30 mm F lsom 3 3 gt 22 mm 2 21 29 Moderat f lsom 2 2 19 21 mm 0 lt 20 mm Relativ resistent 1 1 13 18 mm Resistent 0 lt 12 Ciprofloxacin 10 ug i 9 m Gatifloxacin i Vancomycin 5 ug Imipenem Pseud 30 23 2418 t pr diff 22 21 staph s Levofloxacin i 3 gt 28 mm Azithromycin Moxifloxacin i 2 20 27 im Bacitracin i 1 15 19 mm Clarithromycin Spiramycin 30 26 0 Zimm Polymyxins 150 ug Ertapenem 30 26 r Telithromycin 22 16 _ Amoxycillin a Trimethoprim 3 gt 28 mm Amoxycillin Clav b Cefoxitin 10 ug 2 23 27 mm Aztreonam S aureus 22 22 1 15 22 mm Cefepime b c Daptomycin 0 lt 14 mm Cefotaxime b c 2418 t pr diff staph 22 21 s PA Cefpirome b c _ Ampicillin 33 ug a Cefsulodi
291. ta lactamases both chromosomal and plasmidic DHA 1 DHA 2 will show antagonism distorted zone between Cefoxitin and Ceftazidime Strains of Klebsiella spp Salmonella spp and P mirabilis that show synergism using Boronic acid and or Cloxacillin 500 ug possess presumptively plasmid mediated AmpC beta lactamases 10 Strains of coli showing synergism with Boronic acid and or Cloxacillin 500 ug may possess plasmidic or chromosomal AmpC bata lactamases In order to differentiate plasmidic from chromosomal AmpC beta lactamases in E coli 9 look at the zones around Cefotaxime Ceftazidime and Aztreonam Neo Sensitabs The presence of scattered colonies resistant mutants at the edge of the zones indicates plasmid mediated AmpC while the presence of well defined zone edges indicates chromosomal AmpC beta lactamase O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 16 Page 137 of 170 References 1 Smith Moland E et al Occurrence of newer beta lactamases in Klebsiella pneumoniae isolates from 24 US Hospitals Antimicr Ag Chemother 46 3837 3842 2002 2 Philippon A et al Plasmid determined AmpC type beta lactamases Antimicr Ag Chemother 46 1 11 2002 3 Gupta et al Emergence of multidrug resistant Salmonella enterica serotype Newport resistant to expanded spectrum cephalosporins in the US J Infect Dis 188 1707 16 2003 4 Hyunjeo Pai et al Epidemiology and clinical features of b
292. tation D 226 ICAAC september 2007 Chicago USA Borda N et al Comparison of methods diffusion DF prediffusion PDF and E test on isolates of Ac baumannii calcoaceticus complex Abc against colistin 2007 in press O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 7 Page 20 of 170 7 Dispensers and Templates Schabelons The NEO SENSITABS Dispensers The Neo Sensitabs Dispensers have been designed for uniform and accurate simultaneous positioning of the tablets on an agar plate in any combination desired The application is one operation easy accurate and time saving Today we have 2 different kinds of models available from Rosco 1 Neo Sensitabs Dispensers mobile These dispensers are to be used with the range of Neo Sensitabs in cartridges without a spring Models available 1 adaptable to 8 10 cm petri dishes delivering up to 7 Neo Sensitabs at a time 2 for 10 cm petri dishes delivering up to 9 Neo Sensitabs at a time 3 for 14 15 cm petri dishes delivering up to 12 Neo Sensitabs at a time 4 for square petri dishes 12x12 cm delivering up to 16 Neo Sensitabs at a time The mobile Neo Sensitabs dispensers are made from transparent acrylic plastic and the agar surface is plainly visible while the tablets are being transferred The tablets come packed in cartridges tubes matching the dispenser top plate holes Each cartridge accommodates 50 Neo Sensitabs For easy identific
293. tent will reduce zone sizes whereas low cation content may result in unacceptable large zones of inhibition 1 Ca and Mg should be available in the medium in the form of soluble salts Later differences were found in thymidine content affecting testing of trimethoprim and methicillin resistant staphylococci 2 3 Most agar media contain small amounts of sulphonamide and trimethoprim antagonists that may affect the results of susceptibility testing especially if blood is not added when low content disks are used while Neo Sensitabs are less affected 4 Susceptibility test media should contain less than 0 03 mg l thymidine otherwise small colonies are seen inside the trimethoprim zone If the medium contains slightly more thymidine than recommended it is possible to reduce the concentration by adding thymidinephosphorylase 0 025 to 0 1 IU enzyme ml medium or 5 haemolyzed horse blood which contains the same enzyme Reducing the thymidine content of Mueller Hinton agar for clearer sulphonamide and trimethoprim zones might have an inhibitory effect on the growth of some staphylococci This phenomenon may also be related to problems in detecting resistance to oxacillin methicillin in staphylococci Consequently to make sure that a certain Mueller Hinton agar is useful for susceptibility testing it must fulfil the following criteria t shows acceptable batch to batch reproducibility for susceptibility testing Zone diameters with th
294. ter spp Imipenem EDTA 15 750ug IM ED detection of metallo B lactamases Levofloxacin 5ug LEVOF 223 22 20 19 2 gt b Meropenem 10ug MEROP gt 23 22 19 lt 18 lt 4 gt 16 B cepacia Acinetobacter spp Minocycline 80 ug MINOC 226 25 23 lt 22 lt 4 gt 16 Acinetobacter spp 222 21 19 lt 18 lt 4 gt 16 Moxifloxacin 5 ug MOXIF gt 23 22 20 lt 19 lt 2 gt 8 Ofloxacin 10 ug OFLOX gt 23 22 20 lt 19 lt 2 gt 8 Piperacillin 100 ug PIPRA gt 26 25 21 lt 20 lt 16 gt 128 B cepacia Acinetobacter spp Piperacillin Tazo 100 10ug PI TZ gt 26 25 21 lt 20 lt 16 4 gt 128 4 B cepacia Acinetobacter spp Polymyxins 150 150ug 150 222 21 lt 4 gt 8 S maltophilia Acinetobacter spp Rifampicin 30 ug RIFAM 223 22 20 lt 19 lt 4 gt 16 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 15 Page 124 of 170 Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE S R S R Temocillin 30 ug TEMOC gt 18 17 15 lt 14 lt 16 gt 32 B cepacia Tetracyclines 80 ug TET80 gt 26 25 23 lt 22 lt 4 gt 16 Acinetobacter spp 222 21 19 lt 20 lt 4 gt 16 Ticarcillin Clavulanate 75 15 ug TI CL gt 26 25 21 lt 20 lt 16 2 gt 128 2 S maltophilia Acinetobacter spp Tigecycline 15 ug TIG15 gt 16 z lt 2 Acinetobacter spp Burkholderia spp 18 Trimethoprim Sulfa 5 2 240 ug TR SU gt 30 29 24 lt 23 lt 2 38 gt 8 152 Interpr
295. teria Interpretation according to the MIC break points recommended by the Swedish Reference Group for Antibiotics SRGA 1 Media Iso sensitest Inoculum acc to ICS Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Beta lactams Penicillin Low 5 ug PEN L Staphylococcus spp gt 28 lt 28 lt 0 12 Penicillinase Enterococcus spp gt 26 25 10 lt 10 lt 0 25 gt 4 Oxacillin lug OXA 1 j S aureus gt 16 lt 16 lt 1 gt 1 j Coag neg staph gt 20 lt 20 lt 0 25 gt 0 5 CLSD Cloxacillin 500 ug CL500 Detection of plasmid mediated AmpC Ampicillin 33 ug AMP33 a Enterobacteriaceae gt 32 31 18 18 lt 1 gt 8 Enterococcus spp Pr mirabilis gt 26 25 18 lt 18 lt 2 gt 8 d Amoxycillint Clav 30 15 ug AM CL gt 28 27 18 lt 18 lt 2 1 gt 8 4 Mecillinam 33 ug MECIL gt 30 29 20 lt 20 lt 1 gt 8 Pr mirabilis gt 24 23 20 lt 20 Piperacillin 100ug PIPRA gt 28 27 24 lt 24 lt 16 gt 16 Piperacillin Tazobactam 100 10ug PI TZ gt 28 27 24 lt 24 lt 16 gt 16 Cephalothin 66 ug CLOTN gt 34 33 20 lt 20 lt 1 gt 8 Cephalexin 30 ug CFLEX 230 29 18 18 lt 1 gt 8 Cefadroxil 30 ug CFDRO 230 29 18 18 lt 1 gt 8 d Cefaclor 30 ug CCLOR 230 29 18 18 1 28 Cefuroxime inj 60 ug CEFUR 226 25 23 23 8 28 d Cefoxitin 60 ug CFOXT 226 25 20 20 lt 4 gt 4 j S aureus and S lugdunensis gt 30 29 28 lt 28 OxaS pos j Coag neg staph gt
296. tes sensitive to gentamicin will currently be sensitive to netilmicin I Iso sensitest cannot be used to detect vancomycin resistant enterococci van B Use Mueller Hinton Agar m Iso sensitest is not recommended for detection of metallo beta lactamases Use Mueller Hinton Agar n For detection of VISA GISA hVISA strains see chapter in User s Guide on detection of staphylococci with decreased susceptibility to vancomycin page 86 o When testing enterococci use Gentamicin 250 ug Neo Sensitabs to detect HLR Strains that are HLR to gentamicin are HLR to alle aminoglycosides except streptomycin May be tested with Streptomycin 500 ug p Prediffusion technique 2 18 hours described on page 18 References 1 RAF Referens och Metodbok Resistensbest mning av bakterier mot Antibiotika March 1990 2 Kompletering av RAF s Reference och Metodbok Juni 1995 3 RAF Nyheter 2006 O Copyright Rosco Diagnostica A S NEO SENSITABS III b 09 2007 2008 Page 41 of 170 Haemophilus spp Moraxella catarrhalis S pneumoniae Streptococcus spp and Corynebacterium jeikeium urealyticum Interpretation according to SRGA 2002 Media Iso sensitest 5 blood NAD and 5 CO Inoculum acc to ICS Zone diameter Break points in mm MIC ug ml NEO SENSITABS POTENCY CODE S l R S R Penicillin Low G 5ug PEN L Haemophilus spp 220 19 18 18 lt 1 gt 1 Streptococcus spp gt 26 25 20 lt 20 lt 0 25 gt 1 Moraxella
297. tes with a zone 12 mm should be subjected to MIC test Report results as fluoroquinolone S or R Fosfomycin to be used in combination with ceftriaxone or vancomycin 17 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Page 113 of 170 Chapter 15 15 5 1 Quality Control Limits for S pneumoniae ATCC 49619 Quality Control Limits for S pneumoniae ATCC 49619 MH 5 blood Inoculum McFarland 0 5 Incubation in 5 7 CO NEO SENSITABS POTENCY CODE Zone diameter in mm MIC pg ml Amoxycillin 30 ug AMOXY 36 42 0 06 0 12 Ampicillin 33 ug AMP33 36 42 0 06 0 12 Ceftobiprole BAL 9141 30 32 39 0 008 0 015 Cefepime 30 ug CFEPM 28 35 0 12 Cefotaxime 30 ug CFTAX 33 41 0 06 Ceftizoxime 30 ug CEZOX 28 34 0 25 Ceftriaxone 30 ug CETRX 33 40 0 06 Cefuroxime 60ug CEFU 32 38 0 25 0 5 Chloramphenicol 10 ug CLR10 18 24 4 Chloramphenicol 60 ug CLR60 28 35 4 Clindamycin 25 ug CLIND 29 36 0 06 0 12 Doripenem 10 ug DORIP 30 38 0 06 Doxycycline 80 ug DOXYC 27 34 0 06 Erythromycin 78 ug ERYTR 28 35 0 06 0 12 Faropenem 21 29 0 06 Gatifloxacin 5ug GATIF 24 31 0 25 Iclaprim 21 29 Levofloxacin 5ug LEVOF 20 25 1 Linezolid 30 ug LINEZ 28 34 1 Meropenem 10 ug MEROP 30 37 0 12 Moxifloxacin 5ug MOXIF 25 31 0 12 Ofloxacin 10 ug OFLOX 17 23 2 Oxacillin lug OXA 1 12 16 0 5 pen Penicillin Low 5ug PEN L 22 28 0 5 Quinupristin Dalfopristin 15 ug SYNI5 19 24 0 5 Telavancin 30ug 17 24 0 004 0 008 Telithromycin 15 ug T
298. than amoxycillin against several Entero bacteriaceae The group of strains Enterobacter C freundii Hafnia alvei M morganii Providencia Proteus indole positive and S marscenscens all produce an inducible chromosomal AmpC beta lactamase which is not inhibited by clavulanate In most cases there is an antagonism between amoxycillin and clavulanate smaller zone with the combination than with amoxycillin alone due to the presence of the inducible beta lactamase All these strains should be reported as resistant to ampicillin amoxycillin and to amoxycillin clavulanate except P vulgaris The presence of oval zones is in many cases an indication of heterogeneous cultures which in most cases are resistant to the particular antimicrobial Oval zones are measured by taking the mean between the two diameters Due to the above mentioned factors it is not infrequent that when different persons measure the same zone of inhibition they arrive to discordant results variations of 2 3 mm are currently seen It is therefore important to establish rules for the reading of inhibition zones A good procedure is regularly testing of control organisms see quality control O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 9 Page 26 of 170 References 1 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 January 2003 2 Annear D I Hudson J A An unusual zone surrounding colis
299. the mecA gene determinant but a resistant phenotype can also be achieved by 2 hyper production of beta lactamase and 3 the presence of PBP s with decreased affinity for beta lactams PBP 2a or PBP 2 1 For susceptibility testing of S aureus and S lugdunensis we recommend e Cefoxitin Neo Sensitabs e Oxacillin 1 ug Neo Sensitabs only S aureus e Mueller Hinton no NaCl added e Inoculum from direct colony suspension McFarland 0 5 standard e Incubate for full 24 hours at 33 35 e Interpretation S gt 25 mm R lt 24 mm with Cefoxitin 60 ug Neo Sensitabs S gt 13 mm I 12 11 mm R lt 10 mm with Oxacillin I ug Neo Sensitabs In case of discrepancy Oxa S and Cefox R the cefoxitin result should be used The cefoxitin test is the preferred method for testing S aureus S lugdunensis and coagulase negative staphylococci for resistance to the penicillinase stable penicillins CLSI 2005 43 According to Felten 14 the cefoxitin diffusion method is the best performing test for routine detection of all classes of MRSA According to the Working Group on staphylococci NCCLS CLSI 2005 the results of disk tablet diffusion using cefoxitin can be used to predict mecA mediated resistance in stapylococci Cefoxitin Neo Sensitabs should be used to detect MRSA heterogeneous resistance Isolates of staphylococci that carry the mecA gene or that produce PBP 2a the mecA gene product should be reported as oxacillin methicillin
300. this case If only criteria are specified For some organism antimicrobial combinations the absence of resistant strains precludes defining any category other than susceptible For strains yielding results suggestive of non susceptible organism identification and antimicrobial susceptibility test results should be confirmed Subsequently the isolates should be submitted to a Reference Laboratory for further testing References 1 CLSI Performance Standards for Antimicrobial Disk Susceptibility Testing 8th ed M2 A8 2003 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 12 Page 80 of 170 12 Quality Control Procedures The goals of a good Quality Control Program are To control precision and accuracy of the test procedure To control performance of the reagents used in the test To control the performance of the staff who carry out the tests and read the results Stock cultures of Staphylococcus aureus ATCC 25923 E coli ATCC 25922 Pseudomonas aeruginosa ATCC 27853 and Enterococcus faecalis ATCC 29212 should be obtained from a reliable source Quality control stock organisms may be obtained from the ROSCO representative in your country Other strains recommended as quality control strains are Streptococcus pneumoniae ATCC 49619 see chapter 15 5 1 Haemophilus influenzae ATCC 49247 see chapter 15 1 1 E faecalis ATCC 51299 see chapter 14 5 S aureus ATCC 43300 see chapter 13 4 and S
301. tifloxacin 5 ug GATIF gt 21 20 18 lt 17 lt 1 gt 4 Imipenem 15 ug IMIPM gt 30 29 27 lt 26 lt 0 25 gt 1 b Levofloxacin 5ug LEVOF 218 17 15 lt 14 lt 2 gt Linezolid 30 ug LINEZ 221 lt 2 Meropenem 10 ug MEROP gt 28 lt 0 5 Moxifloxacin 5 ug MOXIF gt 18 17 15 lt 14 lt 1 gt 4 b Ofloxacin 10 ug OFLOX 220 19 17 16 2 28 Oxacillin 1 ug OXA 1 gt 14 lt 13 lt 13 lt 012 pen 20 25 VR penicillin screening Oxacillin 5 ug 5 gt 20 lt 19 lt 19 X012 pen 20 25 VR penicillin screening a Penicillin Low viridans 5 ug PEN L 226 25 13 lt 12 lt 0 12 gt 4 Quinupristin Dalfopristin 15 ug SYNI5 gt 19 18 16 lt 15 lt 1 gt 4 d Rifampicin 30 ug RIFAM gt 28 27 24 lt 23 lt 1 gt 4 f Tigecycline 15 ug TIG15 gt 19 lt 0 25 Teicoplanin 60 ug TEICO 218 9 Telithromycin 15 ug TEL15 gt 19 18 16 lt 15 lt 1 gt 4 Tetracyclines 80 ug TET80 gt 26 25 23 lt 22 lt 2 gt 8 Tetracyclines 10 ug TET10 gt 18 17 15 lt 14 lt 2 gt 8 Trimethoprim Sulfa 5 2 240 ug TR SU gt 32 31 27 lt 26 lt 0 5 9 5 gt 4 76 c Vancomycin 5ug VAN 5 gt 17 lt 1 Breakpoints have not been established by the CLSI Note Viridans streptococci isolated from blood or CSF should be tested for penicillin or ampicillin susceptibility using an MIC method 1 Small colony forming beta haemolytic strains with group A C F or G antigens S anginosus previously termed S miller
302. tin Dalfopristin 15 ug SYNIS 15 21 4 Telithromycin 15 ug TEL15 17 23 2 Tetracyclines 10 ug TET10 9 14 8 16 Tetracyclines 80 ug TET80 17 23 8 16 Tigecycline 15 ug TIG15 23 31 0 12 0 25 Trimethoprim Sulfa 5 2 240 ug TR SU 30 41 0 03 0 6 as trim Note These quality control limits apply only to the tests conducted with H influenzae ATCC 49247 using Haemophilus Test Medium References 1 NCCLS Performance Standards for Antimicrobial Disk Susceptibility Tests 8th Ed M2 A8 section 6 1 page 11 12 2003 2 Performance Standards for Antimicrobial Susceptibility Testing 17th Inf Suppl M100 S17 2007 3 ZervaL et al Reevaluation of interpretative criteria for H influenzae by using Meropenem 10ug Imipenem 10 ug and Ampicillin 2 and 10 ug disks J Clin Microbiol 34 1970 1974 1996 4 Campos J et al Long term persistance of ciprofloxacin resistant H influenzae in patients with cystic fibrosis J I D 174 1345 1347 1996 5 Doern Gary V et al Antibiotic resistance among clinical isolates of H influenzae in the U S in 1994 and 1995 and detection of beta lactamase positive strains resistant to Amoxycillin Clavulanate Results of a National Multicenter Surveillance Study Antimicrob Agents Chemother 41 292 297 1997 6 Jacobs M R et al Effect of various test media on the activities of 21 antimicrobial agents against Haem influenzae J Clin Microbiol 40 3269 3276 2002 7 P rez V zquez
303. tin discs in sensitivity tests of Serratia marcescens The Med J Austral i 840 841 1970 3 Brumfitt W et al Phenomenon of resistance to Augmentin assosiated with sensitivity to ampicillin occurrence and explanation J Clin Pathol 36 670 673 1983 O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 27 of 170 10 Zone Diameter Interpretation Tables 10 1 I Inoculum and MIC Breakpoints according to the CLSI Kirby Bauer I CLSI method Kirby Bauer rapidly growing bacteria Mueller Hinton agar inoculum McFarland 0 5 and break points according to CLSI M2 A8 Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE S R S R Amikacin 40 ug AMIKA 220 19 17 lt 16 lt 16 232 b Amoxycillin 30 ug AMOXY 220 19 17 lt 16 lt gt 32 e Enterococcus spp gt 20 gt 16 d Amoxycillint Clav 30 15 ug AM CL gt 20 19 17 lt 16 lt 8 4 gt 32 16 Staphylococcus gt 26 lt 4 2 gt 8 4 33 ug AMP33 220 19 17 lt 16 lt 8 gt 32 e Enterococcus spp gt 20 gt 16 d Ampicillin Sulbactam 30 30 ug AM SU 220 19 17 lt 16 lt 8 4 gt 32 16 Azithromycin 30 ug AZITR gt 18 17 15 lt 14 lt 2 gt 8 Aztreonam 30 ug AZTRM gt 23 22 20 lt 19 lt 8 gt 32 0 ESBL screening lt 26 gt 1 Bacitracin 40U BACIT 220 19 17 16 Cefaclor 30 ug CCLOR gt 20 19 17 lt 16 lt 8 gt 32 Cefadroxil 30 ug CFDRO 220 19
304. ting at the tablet edge Thereafter incubation for 18 24 hours at 35 37 Alterations in shape of the zones of inhibition around the test organism is considered indication of carbapenemase activity References 1 Pottumarthy S et al NmcA carbapenem hydrolyzing enzyme in E cloacae in North America Emerg Infect Dis 9 999 1002 2003 2 Yigit H et al Novel carbapenem hydrolyzing beta lactamase KPC 1 from a carbapenem resistant strain of K pneumoniae Antimicr Ag Chemother 45 1151 61 2001 3 Smith Moland E et al Plasmid mediated carbapenem hydrolyzing beta lactamase KPC 2 in K pneumoniae isolates J Antimicr Chemother 51 711 14 2003 4 Aubron C et al Carbapenemase producing Enterobacteriaceae U S rivers Emerg Infect Dis 11 260 4 2005 5 Bratu S et al Detection of KPC carbapenem hydrolyzing enzymes in Enterobacter spp from Brooklyn N Y Antimicr Ag Chemother 49 776 8 2005 6 Lopez Otsoa F et al Endemic carbapenem resistance associated with OX A 40 carbapenemase among A baumanni isolates from a hospital in Northern Spain J Clin Microbiol 40 4741 3 2002 7 Bratu S et al Emergence of KPC possessing pneumoniae in Brooklyn N Y epidemiology and recommendations for detection Antimicr Ag Chemother 49 3018 20 2005 8 Brown S Amyes S OXA beta lactamases in Acinetobacter the story so far JAC 57 1 3 2006 9 Dong Min et al Treatment failure due to emergence of resistance to carb
305. ting of thermophilic Campylobacter species Clin Microbiol and Infect 5 580 584 1999 6 Koeth L et al Quality control study evaluating the performance of daptomycin and daptomycin calcium disks and Etest Clin Microbiol Infect 10 Supp 3 541 2004 7 Denis O et al Polyclonal emergence and importation of CA MRSA strains harbouring Panton Valentine leucocidin genes in Belgium JAC 56 1103 06 2005 2 1 Storage of NEO SENSITABS Neo Sensitabs are stored at room temperature both unopened and cartridges in use Very few exceptions Cefpodoxime Cefepime Cefepime Clavulanate Temocillin Ticarcillin Ticarcillin Clavulanate Amphotericin B Caspofungin must be stored in the refrigerator With these Neo Sensitabs labelled STORE AT 2 8 C the following precautions must be taken 1 Unopened cartridges stock Store in refrigerator 2 8 C Expiry date on the label and cartridge 2 Cartridges that have been opened Store at room temperature for up to 2 months 3 Cartridges placed in dispenser Store in dispenser at room temperature for up to 2 months If an opened cartridge labelled 2 8 C is not used up within 2 months it should be kept in the refrigerator Each time the cartridge is taken from the refrigerator it must be allowed time to reach room temperature 30 60 min before it is opened in order to avoid water condensation on the tablets Cartridges labelled 2 8 C that have been opened and are kept in the refrig
306. tion method in susceptibility categories in all cases with S aureus P aeruginosa and E coli Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 5 Page 16 of 170 For the bacteriological analysis of sputum from cystic fibrosis patiens colonised by P aeruginosa sputum is fluidized with acetylcystein and further diluted 1 20 with saline Inoculate MH plate 5 blood Add antimicrobial disks tablets Ticarcillin Ceftazidime Aztreonam Imipenem Tobramycin Ciprofloxacin Polymyxins colistin and C 390 After incubation overnight look for suspicious colonies inside the inhibition zones Identify and perform antibiograms 14 S maltophilia will show resistance to imipenem P aeruginosa resistance to C 390 A xylosoxydans resistance to tobramycin and B cepacia resistance to colistin References 1 Waterworth P M Delpiano M Dependability of susceptibility tests in primary culture J Clin Path 29 179 184 1976 2 Mirrett S Reller L B Comparison of direct and standard antimicrobial disk susceptibility testing for bacteria isolated from blood J Clin Microbiol 10 482 487 1979 3 Fay D Oldfather J E Standardization of direct susceptibility tests for blood cultures J Clin Microbiol 9 347 350 1979 4 Doern G V Scott D R Rashad A L Kim K S Evaluation of a direct blood culture disk diffusion antimicrobial susceptibility test Antimicrob Agents Chemoth 20 696 698 1981 5 J
307. tional and multicenter comparison of EUCAST and CLSI M27 A2 broth microdilution methods for testing susceptibilities of Candida spp to fluconazole itraconazole posaconazole and voriconazole J Clin Microbiol 43 3884 9 2005 Espinel Ingroff A Canton E Comparison of three antifungal susceptibility methods for testing Candida spp Cryptococcus neoformans with Caspofungin CAS and Amphothericin B AMB Abstract presentation M1604 45th ICAAC 2005 Espinel Ingroff A et al Quality Control and reference guidelines for CLSI broth microdilution susceptibility method M38 A document for amphothericin B itraconazole posaconazole and voriconazole J Clin Microbiol 43 5243 6 2005 S S et al Triazole cross resistance among Candida spp case report occurrence among bloodstream isolates and implications for antifungal therapy J Clin Microbiol 44 529 535 2006 Pfaller M A et al Correlation of MIC with outcome for Candida spp tested against viroconazole analysis and proposal for interpretative breakpoints J Clin Microbiol 44 819 26 2006 Sims C R et al Correlation between microdilution E test and disk diffusion methods for antifungal susceptibility testing of Posaconazole against Candida spp J Clin Microbiol 44 2105 8 2006 Espinel Ingroff A Comparison of 3 commercial assays and a modified disk diffusion assay with 2 broth microdilution Reference Assays for testing zygomycetes Aspergillus spp Candida
308. tracyclines 80 ug TET80 Streptococcus spp S pneumoniae gt 28 27 24 lt 23 lt 2 gt 2 Haemophilus spp Moraxella catarrhalis gt 28 27 24 lt 23 lt 2 gt 2 Rifampicin 30 ug RIFAM Streptococcus spp S pneumoniae 232 31 29 lt 28 lt 1 gt 2 Linezolid 30 ug LINEZ Streptococcus spp S pneumoniae 228 27 24 lt 23 lt 2 gt 4 Vancomycin 5ug VAN 5 Streptococcus spp S pneumoniae gt 18 17 16 lt 15 lt 2 gt 2 Corynebacterium spp Listeria spp gt 18 17 16 lt 15 lt 4 gt 4 Teicoplanin 60 ug TEICO Streptococcus spp S pneumoniae gt 24 23 17 lt 16 lt 0 5 gt 4 Corynebacterium spp Listeria spp gt 18 17 16 lt 15 lt 4 gt 4 i Ciprofloxacin 10 ug CIP10 b Haemophilus spp gt 36 35 29 lt 28 lt 0 12 gt 0 25 Moraxella catarrhalis gt 36 35 29 lt 28 lt 0 12 gt 0 25 i Nalidixan 130ug NALID Nedsat fglsomhed for b Haemophilus spp lt 28 kinoloner Norfloxacin 10ug NORFX Nedsat f lsomhed for Pneumococcus spp lt 16 kinoloner i Moxifloxacin Sug MOXIF h Streptococcus spp S pneumoniae gt 26 25 21 lt 20 lt 1 gt 2 b Haemophilus spp Moraxella catarrhalis gt 33 32 29 lt 28 lt 0 25 gt 0 5 Trimethoprim 5 2 ug TRIME Streptococcus spp 222 21 19 lt 18 lt 2 gt 4 Trimethoprim Sulfa 5 2 240ug TR SU Streptococcus spp S pneumoniae 232 31 27 lt 26 lt 16 gt 32 Haemophilus spp Listeria spp gt 32 31 27 lt 26 lt 16 gt 32 Bem rkninger Haemophilus a Beta lactamase negative ampic
309. transparent acrylic plastic with 7 9 12 or 16 holes corresponding to the dispenser types Each hole can be filled with a tablet and the corresponding zone size interpretation in the form of self adhesive circles is placed around it Currently a template is prepared for each type of test or dispenser i e enterobacteriaceae from urine staphylococci from blood etc The template is used by superimposing the susceptibility plate over it or the template is held on the back of the petri dish illuminated with reflected light Templates are made to order according to the individual wishes of each laboratory Ask the ROSCO representative in your country O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 8 Page 22 of 170 8 Representative Antimicrobials to be Tested Routinely Selection of the most appropriate antimicrobials to test and report is a decision made by each laboratory in consultation with the infectious disease practitioners the pharmacy and infection control committees Testing of some agents may be useful for infection control or epidemiological purposes 1 antimicrobials should be reported by their generic names To emphasize the relatedness of the currently available drugs they may be grouped together by drug classes as follows The penicillin group of antibiotics With regard to antimicrobial effect it seems appropriate to classify them as 1 labile to penicillinase Penicillin G Penic
310. ty testing in vitro to sertaconazole in clinical isolates of yeast Rev Esp Quimioter 12 march 1999 Spanish Carrillo Mufioz A J et al Sertaconazole in vitro antifungal activity against vaginal and other superficial yeast isolates J of Chemother 13 555 562 2001 Carrillo Mufioz A J et al Ciclopiroxolamine in vitro antifungal activity against clinical yeast isolates Intl J Antimicr Ag 20 375 379 2002 Mycology online Antifungal susceptibility testing Disk diffusion and E Test methods www mycology adelaide edu au mycology Cuenca Estrella M Personal communication 2002 Vandenbossche I et al Susceptibility testing of fluconazole by the NCCLS broth macrodilution method E Test and Disk diffusion for application in the routine laboratory J Clin Microbiol 40 918 921 2002 Lee S et al Fluconazole disk diffusion test with methylene blue and glucose enriched Mueller Hinton Agar for determining susceptibility of Candida species J Clin Microbiol 39 1615 7 2001 Barry A L et al Precision and accuracy of Fluconazole susceptibility testing by broth microdilution E Test and disk diffusion methods Antimicr Ag Chemoter 46 1781 4 2002 McCullough M et al A longitudinal study of the change in resistance patterns and genetic relationship of oral Candida albicans from HIV infected patients J Med Vet Mycol 33 33 37 2002 NCCLS 2002 Reference method for broth dilution antifungal susceptibility testing of
311. ty to beta lactam antimicrobials is often found on Iso sensitest Agar 8 14 The susceptibility of the strains tested is determined according to the interpretation table below Stenotrophomonas maltophilia Burkholderia cepacia Acinetobacter spp Mueller Hinton Agar plain Inoculum Mcfarland 0 5 Incubation at 30 for 20 24 hours S maltophilia B cepacia Incubation at 35 for 16 18 hours Acinetobacter spp Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Amikacin 40 ug AMIKA gt 20 19 17 lt 16 lt 16 gt 32 Acinetobacter spp Ampicillin Sulbactam 30 30 ug AM SU gt 20 19 17 lt 16 lt 8 4 gt 32 16 Acinetobacter spp Aztreonam 30 ug AZTRM gt 23 22 20 lt 19 lt 8 gt 32 Acinetobacter spp Cefepime 30 ug CFEPM gt 20 19 17 lt 16 lt 8 gt 32 Acinetobacter spp Ceftazidime 30 ug CEZDI Acinetobacter spp 220 19 17 lt 16 8 232 B cepacia 222 21 17 lt 16 lt 8 gt 32 Chloramphenicol 60 ug CLR60 gt 25 24 21 lt 20 lt 8 gt 32 Ciprofloxacin 10 ug CIP10 gt 26 25 21 lt 20 lt 1 24 8 Colistin 10 ug CO 10 5 lt 2 gt 4 aeruginosa Acinetobacter spp a 2 18 h prediffusion gt 15 14 11 lt 10 lt 2 gt 8 Doxycycline 80 ug DOXYC 226 25 23 lt 22 lt 4 216 Acinetobacter spp gt 22 21 19 lt 18 lt 4 gt 16 Gentamicin 40 ug GEN40 gt 23 22 20 lt 19 lt 1 gt Acinetobacter spp b Imipenem 15 ug IMIPM 223 22 19 lt 18 lt 4 216 B cepacialAcinetobac
312. ug TET80 gt 28 27 23 lt 23 lt 2 gt 2 Chloramphenicol 60 ug CLR60 gt 26 lt 26 8 28 Rifampicin 30 ug RIFAM 232 32 lt 1 gt 1 Quinupristin Dalfopristin 15 ug SYNI5 gt 20 19 16 lt 16 lt 1 gt 2 Linezolid 30 ug LINEZ gt 26 25 22 22 lt 4 gt 4 I n Teicoplanin 30 ug TPN30 216 15 14 14 lt 4 gt 8 p 2 18 h prediffusion lt 20 VISA GISA I n Vancomycin 5ug VAN 5 gt 16 15 14 lt 14 lt 4 gt 8 p 2 18 h prediffusion staph lt 22 VISA GISA 2 18 h prediffusion enterococci lt 16 VRE f Nitrofurantoin U 260ug NITRO gt 24 lt 24 lt 32 232 h Nalidixan U 130ug NALID 228 27 23 lt 23 lt 8 gt 16 Enterobacteriaceae lt 28 Reduced susceptibility to quinolones Norfloxacin U 10 ug NORFX gt 28 27 24 lt 24 lt 0 5 gt 1 Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 39 of 170 Zone diameter Break points inmm MIC ug ml NEO SENSITABS POTENCY CODE S R S R Ciprofloxacin 10 ug CIP10 Enterobacteriaceae Acinetobacter spp 228 27 24 24 lt 0 5 1 gt 1 Pseudomonas spp gt 30 29 27 lt 27 lt 0 5 gt 1 10 ug OFLOX 228 27 24 lt 24 lt 0 5 gt 1 Levofloxacin 5ug LEVOF 226 25 23 23 lt 1 gt 2 Pseudomonas spp gt 28 27 24 lt 24 lt 1 gt 2 Moxifloxacin 5 ug MOXIF gt 28 27 24 lt 24 lt 0 5 gt 1 Staphylococcus spp gt 30 29 26 lt 26 lt 0 5 gt 1 p Colistin 10 ug CO 10 2 18 h prediffusion gt
313. ug TR SU gt 28 27 24 lt 23 lt 2 38 gt 8 152 9 5 VAN 5 gt 15 14 13 lt 12 lt 4 gt 32 S aureus gt 16 15 13 lt 12 lt 2 gt 16 v 2 18 hours prediffusion staph 22 hVISA VISA 24 18 hours prediffusion enterococci 16 VRE Virginiamycin 30 ug VIRGI 223 22 20 lt 19 lt 2 gt 6 SX Break points have not been established by the CLSI M2 A8 M100 S14 We recommend 32 ug ml as break point for resistance because we believe that some resistant strains may be falsely recorded as Intermediate when using the 64 ug ml break point recommended by the CLSI xxx See description of this technique on page 86 Remarks a b d e 8 h k For urinary tract infections only Staphylococci resistant to Penicillin Low beta lactamase producers should be reported as resistant to penicillin G penicillin V amoxycillin ampicillin azlocillin carbenicillin piperacillin and ticarcillin Cefoxitin is preferable to detect methicillin resistance in staphylococci because it is more likely to detect heteroresistance Strains that are resistant to Cefoxitin Neo S should be be reported resistant to all other beta lactams penicillins beta lactamase inhibitor combinations cephalosporins and carbapenems Amoxycillin Clavulanate and Ampicillin Sulbactam For staphylococci it is preferable to use Cefoxitin Staphylococci resistant to Oxacillin Methicillin
314. ug ml towards cefotaxime ceftriaxone susceptible while isolates resistant to ceftizoxime should be tested by an MIC method c Erythromycin Interpretation valid for azithromycin and clarithromycin C2 Screening of pneumococci for reduced sensitivity to fluoroquinolones is done using Norfloxacin 10 ug Neo S If the inhibition zone is lt 12 mm or the MIC is lt 16 ug ml there is a high risk of development of resistant mutants in vivo O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 10 Page 72 of 170 Streptococci Penicillin resistant strains of Group A and Group B streptococci have not yet been recognized Viridans streptococci isolated from blood or CSF should be tested for penicillin or ampicillin susceptiblity using an MIC method d Oxacillin 1 ug or 5 are useful for screening for penicillin susceptibility in viridans streptococci c Erythromycin Interpretation valid for azithromycin and clarithromycin H influenzae e Beta lactamase negative ampicillin resistant strains BLNAR are best detected using Ampicillin 2 5 ug Neo Sensitabs Cephalothin Neo Sensitabs is also useful to detect BLNAR strains zone 26 mm BLNAR isolates must be considered resistant to amoxycillin amox clav as well as first and second generation cephalosporins no matter the size of the inhibition zone f Strains resistant to nalidixan should be suspected of having reduced susceptibility to quinolones Strains with
315. uivalent to the 0 5 McFarland standard Use the suspension for plate inoculation within 15 min Not more than 9 Neo Sensitabs should be placed on a 150 mm agar plate or 4 Neo Sensitabs onto a 90 100 mm plate The plates are incubated at 35 C with 5 7 for 20 to 24 hours and the inhibition zones measured to the nearest millimetre The susceptibility of the strains tested is determined according to the interpretation table below The MIC breakpoints when available are according to recommendations from CLSI formerly NCCLS 8 Interpretative criteria are based upon population distributions of MIC s of various agents published data on pharmacokinetics and clinical experience in combination with knowledge from different national guidelines Meningococci Mueller Hinton 5 Blood Inoculum McFarland 0 5 Incubation in 5 7 CO Break points according to CLSI Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R Ampicillin 2 5 ug AMP L gt 28 27 24 gt 23 lt 0 12 22 c Azithromycin 30 ug AZITR gt 20 lt 19 2 Cefotaxime 30 ug CFTAX gt 36 lt 35 lt 0 12 Ceftriaxone 30 ug CETRX gt 36 5 lt 35 lt 0 12 Chloramphenicol 60 ug CLR60 232 31 27 26 lt 2 gt 8 Chloramphenicol 10 ug CLR10 gt 20 19 17 lt 16 lt 2 gt 8 c Ciprofloxacin 10 ug CIP10 gt 36 35 32 lt 31 lt 0 03 gt 0 12 c Doxycycline 80 ug DOXYC gt 30 29 27 lt 26 lt 2 gt 8 c Levofloxacin 5 ug LEVOF gt 35 34 33 lt
316. ultiple beta lactam resistance and aztreonam susceptibility may be helpful for identification of these strains in the laboratory If the strain is resistant to aztreonam it may be due to additional resistance mechanisms efflux other beta lactamases ESBL etc Their expression is not inducible The MBL enzymes are resistant to beta lactamase inhibitors and susceptible to chelating agents like EDTA 2 mercaptopropionic acid 2 MPA and Dipicolinic acid DPA Early detection of MBL producing microorganisms is essential to prevent dissemination of these organisms The enclosed tables including strains of Enterobacteriaceae and Non fermenters producing MBLs show that MPL producers particularly in Enterobacteriaceae may show low MIC values against carbapenems making it difficult for the laboratory to detect MBL positive isolates Suspicious isolates resistant to ceftazidime showing no synergy between clavulanate and third gen cephalosporins and possibly showing reduced susceptibility to carbapenems should be tested for carbapenemase activity using Imipenem Meropenem and EDTA and Dipicolinic acid tests The first metallo beta lactamase producing strain of E coli in Spain has been detected in Barcelona using Imipenem EDTA Neo Sensitabs and E test 3 8 The first metallo beta lactamase producing strain of pneumoniae was found in France 4 MBL producing gram negatives have now emerged in Australia 15 The resistance gene bla IMP
317. urveillance Study 5 If these strains become common it would be inappropriate to consider all beta lactamase positive strains to be uniformly susceptible to AM CL Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Page 101 of 170 Chapter 15 Matic et al 13 reported that AM CL resistance in Haemophilus is due to changes in PBP3 in combination with B lactamase production Cerquetti et al 19 describe the first heterogeneous resistance to imipenem in H influenzae using E test The standard broth dilution method was unable to detect resistance Differentiation of strains showing beta lactam resistance H influenzae AMP 2 5 AM CL Beta lactamase Cefpodoxime Clav Sensitive Zone gt 20 mm Zone S neg no syn BLNAR Zone 20 mm neg no syn Beta lactamase pos No zone Zone 28 mm S pos no syn AM CL resistant No zone Zone 28 mm pos no syn ESBL positive No Zone SorR pos neg synergism From South Africa we have the first report of an ESBL in Haemophilus spp 10 17 Besides it is important to detect other types of resistance in Haemophilus B Chloramphenicol The diffusion method should be able to detect chloramphenicol resistance of Haemophilus The most common type of chloramphenicol resistance of haemophilus is the production of a plasmidic acetyl transferase CAT that inactivates chloramphenicol CAT can be detected using clover leaf technique
318. us aureus zone 24 mm Ampicillin 33 ug a Resistance to penicillins and Altered PBPs Enterococci zone 20 mm beta lactams inhibitor comb Oxacillin 1 ug Penicillin resistance PBP alteration zone 20 mm Pneumococci zone lt 14 mm Streptococci zone lt 12 mm Gonococci Ceftizoxime Resistance to third generation PBP alteration Pneumococci zone lt 26 mm cephalosporins Ampicillin 2 5 ug Resistance to AMP AMOXY PBP alteration Haemophilus zone 20 mm AM CL CCLOR CEFUR BLNAR strains Amoxycillin and b Penicillin resistance Beta lactamase Moraxella catarrhalis Amoxycillint Clavulanate AM CL synergy BRO 1 BRO 2 Ampicillin 33 ug and Penicillin resistance and Penicillinase Enterobacteria Amoxycillint Clavulanate AM CL susceptibility Ceftazidime Ceftriaxone Resistance to all cephems Screening ESBL Klebsiella spp zone lt 24 mm and aztreonam E coli Salmonella Cefpodoxime Resistance to all cephems Screening ESBL E coli zone lt 20 mm and aztreonam Klebsiella Salmonella Cefotaxime Ceftriaxone Ceftazidime Cefepime and c Synergy between CFTAX CETRX CEZDI and Extended spectrum beta lactamase Enterobacteriaceae d Amoxycillin Clavulanate AM CL CP CL ESBL double disk synergy Ceftazidime Clavulanate CZ CL zone gt 5 mm ESBL Enterobacteriaceae than CEZDI alone Cefepime and Amoxycillin Synergy between CFEPM ESBL Enterobacter Clavu
319. usceptible to ceftazidime They may show synergism between other third gen cephalosporins and amoxicillin clavulanate and may be mistaken as ESBL producers Adopted from CDS 2005 with modifications References 1 Sanders C C and Sanders W E Beta lactam resistance in gram negative bacteria global trends and clinical impact Clin Infect Dis 15 824 39 1992 2 Livermore D M et al Detection of beta lactamase mediated resistance J Antimicr Chemother 48 Supp S1 59 64 2001 O Copyright Rosco Diagnostica A S 09 2007 2008 Page 134 of 170 Chapter 16 NEO SENSITABS 16 4 Plasmid mediated AmpC Beta lactamases Plasmid mediated beta lactamases represent a new threat since they confer resistance to aminopenicillins carboxypenicillins ureidopenicillins although they are generally susceptible in vitro to mecillinam and or temocillin The enzymes provide resistance to third generation cephalosporins and cefoxitin The enzymes are also active against aztreonam although for some strains the aztreonam MICs are in the susceptible range Susceptibility to cefepime is little affected inoculum effect and the carbapenems are not affected The enzymes are not affected by beta lactamase inhibitors except for CMY 8 and CMY 9 that are inactivated by tazobactam Their expression is generally constitutive nevertheless inducible plasmid AmpC ACT 1 DHA 1 DHA 2 CMY 13 have been reported 6 Plasmid mediated AmpC beta lactamas
320. vnrnvrrnrrrnrrerarerarvrarrneneevvensvesnvernvesnsesvsversssrasvreevresnen 20 8 Representative Antimicrobials to be Tested Routinely sseeeeseeeeeeeeee nennen nennen 22 9 Measuring the Inhibition Zones og iite ene r reen nennen rene 25 10 Zone Diameter Interpretation Tables passe eerie Per tree Ee cape Heo reete EE rere ep ip 27 10 1 I Inoculum and MIC Breakpoints according to CLSI Kirby Bauer eee 27 10 2 Interpretations according to MIC Breakpoints of the Dutch CRG sene 32 10 3 III Interpretation according to MIC Breakpoints of SRGA in Sweden esee 37 10 4 IV Interpretations according to MIC Breakpoints of the Norwegian AFA Group 44 10 5 V Danish Blood Agar Interpretation Valid for Denmark eese 52 10 5 1 Interpretations According to MIC Breakpoints of the Danish Reference Group for Susceptibility ette rete rp dtp eo concede e e rege deas 57 10 6 VI Interpretation according to MIC Breakpoints of SFM France eene 65 10 7 VII Interpretation according to MIC Breakpoints of DIN 58940 4 73 10 8 Interpretation according to the MIC break points recommended by 76 11 Interpretation of RESUlts ss oerte ate
321. wed that the addition of 0 5 ug ml methylene blue made the zones of inhibition clearer and easier to read particularly with the azole drugs Finally this technique was approved by the CLSI formerly NCCLS as the disk diffusion method of choice for performing antifungal susceptibility testing of yeasts CLSI document M44 A Lee 25 and Barry 26 using the CLSI technique could read the inhibition zones after 24 hours incubation 97 of results were in agreement with those of the reference test Vandenbosche et al 24 showed that the disk diffusion method using Neo Sensitabs had the best agreement with the CLSI reference method better than Etest In a study using 282 Candida spp Rementeria et al 37 comparing paper disks and Neo Sensitabs containing Fluconazole and Voriconazole found that the results were equivalent but the zones of inhibition were best defined and easier to read with Neo Sensitabs In a recent study by Espenel Ingroff 50 comparing the Neo Sensitabs disk method to both CLSI reference broth microdilution and disk diffusion for testing susceptibilities of 10 20 isolates each of Candida 8 species and C neoformans to 5 antifungal agents amphothericin B caspofungin fluconazole itraconazole and voriconazole found that agreement by breakpoint category of Neo Sensitabs results with CLSI method M27 A2 was 95 5 96 Even though Amphotericin testing was found to work on MH GMB agar 38 results with amphothericn B might be d
322. y of the bacterial population the resistant part of the population having an optimal growth temperature at 30 35 is not detected at 37 because of poor slower growth Strains showing a better growth rate at 30 C than at 37 C should be tested at 30 C optimal growth temperature The antibiotic diffusion from Neo Sensitabs is not affected significantly by using incubation temperatures below or above 37 The following species 1 may show a better growth at 30 Yersinia spp Klebsiella ozaenae certain non fermentative gram negative rods Stenotrophomonas maltophilia Pseudomonas putida Pseudomonas fluorescens some strains of Acinetobacter spp Burkholderia cepacia Aeromonas spp and some Moraxella spp b Nutritionally supplemented media 2 Some strains require supplemented media for their growth 1 Symbiotic streptococci responsible for bacterial endocarditis require pyridoxine thiol or Isovitalex 2 Strains of Enterobacteriaceae which form dwarf colonies on media e g E coli Citrobacter spp Klebsiella spp Proteus spp Salmonella spp require supplement nutrients for larger colony growth Among these are the thymineless variants most likely related to the therapeutic use of trimethoprim Supplementation of the medium with thymidine is required for correct testing of these organisms Other strains require CO thiamin glutamic acid etc 3 Strains of S aureus that form dwarf colonies in routine media require
323. yeasts Approved Standard M27 A2 NCCLS 2004 Method for antifungal disk diffusion susceptibility testing of yeasts Approved guideline M44 A Pfaller M A et al Evaluation of the Etest and disk diffusion methods for determining susceptibilities of 235 bloodstream isolates of C glabrata to Fluconazole and Voriconazole J Clin Microbiol 41 1875 80 2003 Pfaller M A et al Evaluation of the NCCLS M44 P Disk diffusion method for determining Fluconazole susceptibility of 276 clinical isolates of Cryptococcus neoformans ICAAC 2003 presentation M 1204 Barry A et al Quality Control for Fluconazole disk susceptibility tests on Mueller Hinton Agar with glucose and methylene blue J Clin Microbiol 41 3410 12 2003 Carrillo Mufioz A J et al In vitro antifungal activity of voriconazole against dermatophytes and superficial isolates of S brevicaulis Rev Iberoam Micol 22 110 3 2005 Spanish Defontaine A et al In vitro resistance to azoles associated with mitochondrial DNA deficiency in Candida glabrata J Med Microbiol 48 663 670 1999 Pfaller M A et al Q C limits for voriconazole disk susceptibility tests on MH agar with glucose and methylene blue J clin Microbiol 42 1716 8 2004 Carrillo Mufioz A J et al Is Amphotericin B active against dermatophites and Scopuloriosis brevicaulis Rev Esp Quimiot 17 244 9 2004 Spanish O Copyright Rosco Diagnostica A S NEO SENSITABS 09 2007 2008 Chapter 21 37
324. ylobacter jejuni subsp jejuni strains isolated from humans in 1998 to 2001 in Montreal Canada Antimicr Ag Chemother 47 2027 9 2003 6 Lucero C et al Campylobacter spp Comparison of disc diffusion and agar dilution methods for susceptibility testing ICAAC 2003 Presentation D 238 7 McDermott P F et al Development of a standardized susceptibility test for campylobacter with Q C ranges for Ciprofloxacin Doxycycline Erythromycin Gentamicin and Meropenem Microb Drug Resist 10 124 131 2004 8 Gaudreau et al Comparison of disk diffusion and agar dilution methods for erythromycin and ciprofloxacin susceptibility testing of C jejuni Antimicr Agents Chermother 51 1524 26 2007 Copyright Rosco Diagnostica A S NEO SENSITABS 15 8 Susceptibility Testing of Vibrio cholerae 09 2007 2008 Chapter 15 Page 121 of 170 The recommended medium is Mueller Hinton agar Inoculum is prepared by the growth method or by direct colony suspension in 0 946 saline to a density equivalent to 0 5 Mc Farland standard Incubate the plates at 35 C in ambient air for 16 18 hours before reading the inhibition zones Zone diameter interpretative standards with breakpoints according to CLSI are as follows Zone diameter Break points in mm MIC pg ml NEO SENSITABS POTENCY CODE S l R S R a Ampicillin 33 ug AMP33 gt 20 19 17 lt 16 lt 8 232 Chloramphenicol 60 ug CLR60 225 24 21 lt 20 lt 8 gt 32 b Ciprofloxacin
325. zole 13 Isolates with fluconazole MIC gt 4 ug ml in general less susceptible to other azole drugs but different patterns of decreased susceptibility are found On the other hand isolates with fluconazole MICs lt 2 never show decreased susceptibility to other azole drugs 13 C glabrata C tropicalis C albicans and C parapsilosis showing fluconazole MICs gt 64 ug ml R corresponding to zones of inhibition lt 14 mm with Fluconazole Neo Sensitabs showed cross resistance with voriconazole MICs of gt 2 ug ml 46 On the other hand C Krusei fluconazole R are currently voriconazole susceptible no cross resistance EUCAST is working on the harmonization of breakpoints however there is no agreement with MIC breakpoints from CLSI In studies comparing methods interpretive categories misclassification may be seen caused by differences in MIC breakpoints 41 43 21 1 1 2 Interpretation according to EUCAST MIC breakpoints MH Glucose Methylene Blue Agar Inoculum McFarland 0 5 undiluted MIC breakpoints according to EUCAST tentative Zone diameter Break points inmm MIC pg ml NEO SENSITABS POTENCY CODE R S R Amphotericin B 10 ug AMPHO gt 18 17 13 lt 12 lt 0 25 gt 2 Fluconazole 25 ug FLUCZ gt 28 27 23 lt 22 lt 2 gt 4 Itraconazole 50 8 ug ITRAC 223 22 14 lt 13 lt 0 12 2 Ketoconazole 15 ug KETOC gt 30 29 23 lt 22 lt 0 12 21 Posaconazole 52 5ug POSAC 222 21 16 lt 15 lt 0 25 gt 2

User's Guide Neo-Sensitabs 18th ed 10-2005-2006

Contents

Download Pdf Manuals

Related Search

Related Contents