LABORATORY PROCEDURE
Contents
1. AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST ID AST ID AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST ID AST AST Page 19 of 30 Salmonella haardt Salmonella hadar Salmonella hamburg Salmonella hartford Salmonella heidelberg Salmonella illinois Salmonella infantis Salmonella inverness Salmonella java Salmonella javiana Salmonella kentucky Salmonella kirkee Salmonella kunduchi Salmonella kvittingfoss Salmonella lansing Salmonella litchfield Salmonella liverpool Salmonella london Salmonella luciana Salmonella manhattan Salmonella mbandaka Salmonella meleagridis Salmonella memphis Salmonella michigan Salmonella minneapolis Salmonella minnesota Salmonella montevideo Salmonella muenchen Salmonella muenster Salmonella newington Salmonella newport Salmonella nottingham Salmonella ohio Salmonella onderstepoort Salmonella oranienburg AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST AST Page 20 of 30 Salmonella panama Salmonella paratyphi A Salmonella paratyphi B Salmonella poona Salmonella pullorum Salmonella quinhon Salmonella rubislaw Salmonella saintpaul Salmonella schwarzengrund Salmonella seftenberg Salmonella species Salmonella tallahassee Salmonella thompson Salmonella typhi Salmonella typhimurium Salmonella virginia Salmonella w2. ID AST ID AST ID ID AST AST ID AST ID AST AST ID AST ID AST ID AST ID AST AST ID AST AST ID AST ID Page 22 of 30 Gram Negative 0 25 McFarland Gram Negative Taxa Achromobacter species Acinetobacter baumannii calcoaceticus complex Acinetobacter haemolyticus Acinetobacter Iwoffii Actinobacillus lignieresii Actinobacillus suis Actinobacillus ureae Aeromonas caviae Aeromonas hydrophila Aeromonas salmonicida ssp masoucida Aeromonas salmonicida ssp salmonicida Aeromonas salmonicida ssp smithia Aeromonas schubertii Aeromonas sobria Aeromonas veronii Alcaligenes faecalis Bergeyella zoohelcum Bordetella bronchiseptica Brevundimonas diminuta Brevundimonas vesicularis Burkholderia cepacia Burkholderia gladioli Cardiobacterium hominis CDC group EF 4a CDC group EF 4b CDC group EO 2 CDC group Vb 3 Cedecea davisae Cedecea lapagei Cedecea neteri Chromobacterium violaceum ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID ID ID AST ID AST ID AST ID AST ID AST ID AST ID Page 23 of 30 Chryseobacterium gleum Chryseobacterium indologenes Chryseobacterium meningosepticum Citrobacter amalonaticus Citrobacter braakii Citrobacter farmeri Citrobacter freundii Citrobacter koseri Citrobacter sedlakii Citrobacter werkmanii Citrobacter youngae Comamonas terrigena Comamonas testosteroni Delftia acidovorans
3. growth in the presence of an antimicrobial agent Continuous measurements of changes to the indicator as well as bacterial turbidity are used in the determination of bacterial growth Each AST panel configuration contains several antimicrobial agents with a range of two fold doubling dilution concentrations Organism identification is used in the interpretation of the MIC values of each antimicrobial agent producing Susceptible Intermediate or Resistant SIR result classifications A complete list of taxa for which the Phoenix system can provide AST results is provided in Table A The list of antimicrobial agents and concentrations available for susceptibility testing in the Phoenix system is provided under Performance Characteristics There are antimicrobial agents for use with the Phoenix System that are not proven to be effective for treating infections for all organisms listed in the taxa For interpreting and reporting results of antimicrobial agents that have been shown to be active against organism groups both in vitro and in clinical infections refer to the individual pharmaceutical antimicrobial agent labeling Alternatively refer to the most recent CLSI M100 Performance Standard Table 1 Suggested Groupings of US FDA Approved Antimicrobial Agents That Should Be Considered for Routine Testing and Reporting on Organisms by Clinical Microbiological Laboratories The components required for testing using the Phoenix system include 1
4. AST testing refer to Table C for recommended media When swabs are used only cotton tipped applicators should be used to prepare the inoculum suspensions Some polyester swabs may cause problems with inoculation of the panels The usefulness of the Phoenix system or any other diagnostic procedure performed on clinical specimens is directly influenced by the quality of the specimens themselves It is strongly recommended that laboratories employ methods discussed in the Manual of Clinical Microbiology for specimen collection transport and placement on primary isolation media Inoculum for use on the Phoenix system is prepared by the CLSI recommended direct colony suspension method Due to variations in inoculum concentrations prepared with McFarland standards use of the BBL CrystalSpec or BD PhoenixSpec nephelometer is required for adjusting the test inoculum prior to use in the Phoenix system It is highly recommended that the purity of the inoculum be checked by preparing a purity plate See Purity Check below MATERIALS REQUIRED Materials Provided Phoenix Panels Phoenix ID Broth Phoenix AST Broth Phoenix AST Indicator Solution Phoenix Inoculation Station Phoenix Panel Caddy BBL CrystalSpec or BD PhoenixSpec Nephelometer and Standards 25 uL pipettor and sterile tips 50 uL pipettor and sterile tips 2 Pipette stands Materials Required But Not Provided Gram stain reagents Sterile cotton swabs Nonselective culture p
5. Phoenix panels with panel closures 2 Phoenix ID Broth 3 Phoenix AST Broth 4 Phoenix AST Indicator solution 5 Phoenix Inoculation Station 6 Phoenix Panel Caddy 7 BBL CrystalSpec or BD PhoenixSpec nephelometer 8 25 uL pipettor and sterile tips and 9 Miscellaneous lab supplies listed under Materials Required But Not Provided Prior to inoculation the Phoenix panel is placed on the Inoculation Station with the inoculation ports at the top for filling Separate inocula are added manually to the ID and AST ports The Page 2 of 30 inocula flow down the panel in serpentine fashion filling the panel wells as the liquid front progresses toward the pad The pad absorbs excess inoculum Closures are manually inserted in the fill ports An air admittance port is located in the divider area of the panel lid to ensure adequate oxygen tension in the panel for the duration of the test INGREDIENTS For a listing of biochemical substrates used in the Phoenix panel refer to Table B The package insert enclosed in the panel box provides a listing of the specific antimicrobial agents and concentrations found in the panel PRECAUTIONS For in vitro Diagnostic Use All patient specimens and microbial cultures are potentially infectious and should be treated with universal precautions Please refer to CDC manual Bio safety in Microbiological and Biomedical Laboratories 4 Edition 1999 as well as other recommended literature Prior to discardi
6. Solution to warm to room temperature before dispensing into AST broth The unused portion of the indicator should be returned to 2 8 C as soon as possible Do not store at room temperature for more than 2 hours Opened bottles should be discarded after 14 days from initial opening If volume other than one drop is added inadvertently discard the tube and use a fresh tube of AST broth After the addition of the Indicator to AST broth the mixed solution can be stored in the dark at room temperature for as long as 8 hours Tubes must be used within 2 hours after the addition of the indicator solution if exposed to light If an inoculum density of 0 50 0 60 was used transfer 25 uL of the bacterial suspension from the ID tube into the AST broth tube If an inoculum density of 0 20 0 30 was used transfer 50 uL use 2 shots if utilizing a 25 uL pipettor of the bacterial suspension from the ID tube into the AST broth tube Note Panels must be inoculated within 30 minutes of the time that the AST inoculum is prepared Cap the AST tube and invert several times to mix Do not vortex Wait a few seconds for air bubbles to surface Tap the tube gently to aid in eliminating bubbles Pour the ID tube inoculum into the fill port on the ID side of the panel 51 well side Allow the fluid to traverse down the tracks before moving the panel If using an AST only panel DO NOT inoculate the ID side of the panel Retain the ID or AST tube for a purity
7. check Pour the AST tube inoculum into the fill port on the AST side of the panel 85 well side Allow the fluid to traverse down the tracks before moving the panel Before placing panel closure check for residual droplets of inoculum on the edge of the fill ports If a droplet is present remove the droplet with absorbent material The used absorbent material must be discarded along with your biohazard waste Snap on the panel closure Make sure that the closure is fully seated Visually inspect panels to be sure each of the wells is full Look at both sides of the panel Make certain that the wells are not overfilled If any of the wells are unfilled or overfilled inoculate a new panel Note Panels must be loaded into the instrument within 30 minutes of inoculation Panels must be kept in the inoculation station after inoculation until the excess fluid has been completely absorbed by the pad Panels Page 6 of 30 should stay vertical in the transport caddy until loaded into the instrument Inoculated panels should be handled with care Avoid knocking or jarring the panel Purity Check 1 Using asterile loop recover a small drop from the inoculum fluid tube either before or after inoculating the panel Inoculate an agar plate any appropriate medium for a purity check Discard inoculum fluid tube and cap in a biohazard disposal container Incubate the plate for 24 48 hours at 35 C under appropriate conditions PoP ID Inocul
8. the Phoenix disposable The combination panel includes an ID side with dried substrates for bacterial identification an AST side with varying concentrations of antimicrobial agents and growth and fluorescent controls at appropriate well locations The Phoenix system utilizes an optimized colorimetric redox indicator for AST and a variety of colorimetric and fluorometric indicators for ID The AST Broth is cation adjusted e g Ca and Mg to optimize susceptibility testing performance Page 1 of 30 The Phoenix panel is comprised of a 51 well ID side and an 85 well AST side The ID side contains 45 wells with dried biochemical substrates and 2 fluorescent control wells The AST side contains 84 wells with dried antimicrobial agents and 1 growth control well Panels are available as ID only Phoenix NID Panels and Phoenix PID Panels AST only Phoenix NMIC Panels and Phoenix PMIC Panels or ID AST combination Phoenix NMIC ID Panels and Phoenix PMIC ID Panels Unused wells are reserved for future se Phoenix panels are inoculated with a standardized inoculum Organism suspensions must be prepared only with the BBL CrystalSpec or BD PhoenixSpec nephelometer Once inoculated panels are placed into the instrument and continuously incubated at 35 C The instrument tests panels every 20 minutes on the hour at 20 minutes past the hour and again at 40 minutes past the hour up to 16 hours if necessary Phoenix panels are
9. 16 of 30 Hafnia alvei Kingella denitrificans Kingella kingae Klebsiella granulomatis Klebsiella oxytoca Klebsiella pneumoniae ssp ozaenae Klebsiella pneumoniae ssp pneumoniae Klebsiella pneumoniae ssp rhinoscleromatis Kluyvera ascorbata Kluyvera cryocrescens Kluyvera georgiana Leclercia adecarboxylata Leminorella grimontii Leminorella richardii Mannheimia haemolytica Methylobacterium extorquens Moellerella wisconsensis Moraxella Branhamella catarrhalis Moraxella species Morganella morganii Myroides odoratus odoratimimus Ochrobactrum anthropi Oligella ureolytica Oligella urethralis Pantoea agglomerans Pantoea ananatis Pantoea dispersa Pantoea stewartii ssp indologenes Pantoea stewartii ssp stewartii Pasteurella aerogenes Pasteurella multocida Pasteurella pneumotropica Photobacterium damselae Plesiomonas shigelloides Pragia fontium ID AST ID ID AST ID AST ID AST ID AST ID AST ID AST ID AST AST ID AST ID AST ID ID ID AST ID AST ID AST ID ID ID AST AST AST AST AST Page 17 of 30 Proteus hauseri Proteus mirabilis Proteus myxofaciens Proteus penneri Proteus vulgaris Providencia alcalifaciens Providencia heimbachae Providencia rettgeri Providencia rustigianii Providencia stuartii Pseudomonas aeruginosa Pseudomonas alcaligenes Pseudomonas fluorescens Pseudomonas luteola Pseudomonas mendocina Pseudomonas monteilii Pseudomonas oryzihabitans Pseudomonas pertucinog
10. 5 McFarland Gram Negative Taxa ID AST ID AST Achromobacter piechaudii AST Achromobacter species ID AST Achromobacter xylosoxidans ssp denitrificans AST Achromobacter xylosoxidans ssp xylosoxidans AST Acinetobacter baumannii ID AST Acinetobacter baumannii calcoaceticus complex ID AST Acinetobacter calcoaceticus AST Acinetobacter haemolyticus ID AST Acinetobacter johnsonii AST Acinetobacter junii AST Acinetobacter Iwoffii ID AST Acinetobacter radioresistens AST Acinetobacter species ID AST Actinobacillus lignieresii ID Actinobacillus suis ID Actinobacillus ureae ID Aeromonas allosaccharophila AST Aeromonas caviae ID AST Aeromonas eucrenophila AST Aeromonas hydrophila ID AST Aeromonas jandaei AST Aeromonas media AST Aeromonas salmonicida AST Aeromonas salmonicida ssp achromogenes AST Page 14 of 30 Aeromonas salmonicida ssp masoucida Aeromonas salmonicida ssp pectinolytica Aeromonas salmonicida ssp salmonicida Aeromonas salmonicida ssp smithia Aeromonas schubertii Aeromonas sobria Aeromonas trota Aeromonas veronii Alcaligenes faecalis Bergeyella zoohelcum Bordetella bronchiseptica Brevundimonas diminuta Brevundimonas vesicularis Burkholderia cepacia Burkholderia gladioli Cardiobacterium hominis CDC group EF 4a CDC group EF 4b CDC group EO 2 CDC group Vb 3 Cedecea davisae Cedecea lapagei Cedecea neteri Cedecea species 3 Cedecea species 5 Chromobacterium violaceum Chryseobacterium gleum Chryseoba
11. 68 96 3 1068 Cephem Cephalothin CF 1 64 2025 96 4 2025 Folate Antagonist Trimethoprim TMP 0 5 16 1856 955 1856 Folate Antagonist Trimethoprim Sulfamethoxazole SXT 0 5 9 5 16 304 2212 96 0 2212 Monobactam Aztreonam ATM 0 5 64 1470 96 2 1470 Nitrofurantoin Nitrofurantoin FM 8 512 2130 958 2130 Quinolone Nalidixic Acid NA 2 32 2103 96 2 2103 Tetracycline Tetracycline TE 0 5 16 2837 95 5 2837 REFERENCES 1 Bronfenbrenner J and Schlesigner M J 1918 A Rapid Method for the Identification of Bacteria Fermenting Carbohydrates Am J Public Health 8 922 923 2 Arnold W M Jr and Weaver R H 1948 Quick Microtechniques for Identification of Cultures Indole production J Lab Clin Med 33 1334 1337 3 Bachmann B and Weaver R H 1951 Rapid Microtechnics for Identification of Cultures V Reduction of Nitrates to Nitrites Am J Clin Pathol 21 195 196 4 Hannan J and Weaver R H 1948 Quick Microtechniques for the Identification of Cultures Il Fermentations J Lab Clin Med 33 1338 1341 5 Hartman P A 1968 Paper strip and disc methods p 123 132 Miniaturized microbiological methods Academic Press New York 6 Sanders A C Faber J E and Cook T M 1957 A Rapid Method for the Characterization of Enteric Pathogen Using Paper Discs Appl Microbiol 5 36 40 7 Synder M L 1954 Paper Discs Containing Entire Culture Medium for the Differentiation of Bacteria Pathol Bact
12. A KETOGLUTARIC ACID C KGA TIGLIC ACID C TIG FLUORESCENT POSITIVE FLR CTL CONTROL Control to standardize fluorescent substrate FLUORESENT POSTIVE FLR CTL results CONTROL L PROLINE NA N LPROT Enzymatic hydrolysis of the colorless amide substrate releases yellow p nitroaniline GAMMA L GLUTAMYL NA N LGGH BIS PNP PHOSPHATE P BPHO Enzymatic hydrolysis of the colorless aryl substituted glycoside releases yellow PNP BD GLUCOSIDE P BDGLU p nitrophenol Page 28 of 30 Substrate Name Code Principle BETA ALLOSE R_BALL N ACETYL GALACTOSAMINE R_NGA N ACETYL GLUCOSAMINE R_NGU SORBITOL R_DSBT SUCROSE R_DSUC GALACTURONIC ACID R_GRA MALTULOSE R_MTU L RHAMNOSE R_LRHA Utilization of carbohydrate results in lower pH BETA GENTIOBIOSE R BGEN and change in indicator phenol red DEXTROSE R_DEX D GALACTOSE R_DGAL D FRUCTOSE R_DFRU D GLUCONIC ACID R_DGUA D MELIBIOSE R_DMLB L ARABINOSE R_LARA METHYL B GLUCOSIDE R_MBGU ORNITHINE S_ORN Utilization of ornithine results in pH rise and change in fluorescent indicator UREA S_URE Hydrolysis of urea and the resulting ammonia change results in pH rise and change in fluorescent indicator ESCULIN T_ESC Hydrolysis of esculin results in a black precipitate in the presence of ferric ion Page 29 of 30 Table C Recommended Media and Approved Use Recommend
13. Edwardsiella hoshinae Edwardsiella ictaluri Edwardsiella tarda Eikenella corrodens Empedobacter brevis Enterobacter aerogenes Enterobacter amnigenus biogroup 1 Enterobacter amnigenus biogroup 2 Enterobacter asburiae Enterobacter cancerogenus Enterobacter cloacae Enterobacter gergoviae Enterobacter hormaechei Enterobacter intermedius Enterobacter sakazakii Escherichia coli Escherichia fergusonii Escherichia hermannii Escherichia vulneris Ewingella americana Hafnia alvei ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID ID ID AST ID AST ID AST ID AST ID ID ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID ID AST Page 24 of 30 Klebsiella oxytoca Klebsiella pneumoniae ssp ozaenae Klebsiella pneumoniae ssp pneumoniae Klebsiella pneumoniae ssp rhinoscleromatis Kluyvera ascorbata Kluyvera cryocrescens Leclercia adecarboxylata Leminorella grimontii Leminorella richardii Mannheimia haemolytica Moellerella wisconsensis Morganella morganii Myroides odoratus odoratimimus Ochrobactrum anthropi Oligella ureolytica Oligella urethralis Pantoea agglomerans Pasteurella aerogenes Pasteurella multocida Pasteurella pneumotropica Photobacterium damselae Plesiomonas shigelloides Pragia fontium Proteus mirabilis Proteus penneri Proteus vulgaris Providencia alcalifaciens Providencia rettgeri Providencia ru
14. Fluoroquinolone Ciprofloxacin CIP 0 25 4 2853 98 8 2853 95 1 5 Fluoroquinolone Gatifloxacin GAT 0 25 8 2213 988 2213 95 8 5 Fluoroquinolone Levofloxacin LVX 0 25 8 2934 98 5 2934 95 8 5 Fluoroquinolone Moxifloxacin MXF 0 12 8 2202 298 3 2202 97 6 5 Fluoroquinolone Norfloxacin NOR 0 25 16 2792 975 2792 943 5 Fluoroquinolone Ofloxacin OFX 0 25 8 2926 98 5 2926 94 6 Aminoglycoside Amikacin AN 0 5 64 2598 94 7 2598 96 7 Aminoglycoside Gentamicin GM 0 25 16 2751 96 2 2751 96 3 Aminoglycoside Tobramycin NN 0 12 16 2658 93 3 2658 95 3 B Lac B Lac Inh Amoxicillin Clavulanate AMC 0 5 0 25 32 16 2249 96 7 2249 90 9 B Lac B Lac Inh Ampicillin Sulbactam SAM _ 0 5 0 25 32 16 1305 97 2 1305 87 5 B Lac B Lac Inh Ticarcillin Clavulanate TIM 1 2 128 2 1527 925 1527 89 7 B Lactam Pen Ampicillin AM 0 5 32 1712 97 0 1712 94 6 B Lactam Pen Piperacillin PIP 0 5 128 1781 943 1781 93 8 B Lac B Lac Inh Piperacillin Tazobactam TZP 0 5 4 128 4 1546 932 1546 94 9 B Lactam Pen Ticarcillin TIC 1 128 2882 94 7 2882 92 7 Page 11 of 30 Carbapenem Imipenem IPM 1 16 2680 97 2 2680 Carbapenem Meropenem MEM 0 25 16 2905 97 6 2905 Cephem Cefazolin CZ 0 5 32 1331 96 7 1331 Cephem Cefepime FEP 0 5 64 1789 95 2 1789 Cephem Cefotaxime CTX 0 5 64 2268 295 0 2268 Cephem Cefotetan CTT 2 64 1175 966 1175 Cephem Cefoxitin FOX 0 5 64 1397 96 9 1397 Cephem Ceftazidime CAZ 0 5 64 1796 96 5 1796 Cephem Ceftriaxone CRO 0 5 64 1872 958 1872 Cephem Cefuroxime CXM 1 64 10
15. L test resident on the Phoenix panels were compared to the results obtained from the CLSI Page 10 of 30 reference confirmatory ESBL test For Challenge organisms this result is an expected result and for Clinical isolates this result was obtained from concurrent testing in the CLSI reference broth microdilution method Additionally a challenge set of 30 previously characterized organisms was tested at one site Positive Percent Agreement 183 189 96 8 Negative Percent Agreement 780 812 96 1 Overall Percent Agreement 963 1001 96 2 Gram Negative Susceptibility Clinical stock and challenge isolates were tested across multiple clinical sites to determine Essential Agreement EA and Category Agreement CA of the Phoenix system to the CLSI broth microdilution reference method Essential Agreement occurs when the MIC of the Phoenix system and the reference method agree exactly or is within 1 dilution of each other Category Agreement occurs when the Phoenix system results agree with the reference method with respect to the CLSI categorical interpretative criteria susceptible intermediate resistant The table below summarizes the data from these studies Additionally testing performed at multiple clinical sites demonstrated at least 95 reproducibility or greater within 1 doubling dilution for all antimicrobial agents listed in the table below DRUG CLASS DRUG NAME DRUG DRUG EA EA CA CA CODE RANGE N N ug mL 5
16. LABORATORY PROCEDURE BD Phoenix NMIC ID Panels BD Phoenix NMIC Panels BD Phoenix NID Panels INTENDED USE The BD Phoenix Automated Microbiology System is intended for the in vitro rapid identification ID and quantitative determination of antimicrobial susceptibility by minimal inhibitory concentration MIC of Gram Negative aerobic and facultative anaerobic bacteria belonging to the family Enterobacteriaceae and non Enterobacteriaceae SUMMARY AND EXPLANATION OF THE TEST Micromethods for the biochemical identification of microorganisms were reported as early as 1918 Several publications reported on the use of the reagent impregnated paper discs and micro tube methods for differentiating enteric bacteria The interest in miniaturized identification systems led to the introduction of several commercial systems in the late 1960s and they provided advantages in requiring little storage space extended shelf life standardized quality control and ease of use Many of the tests used in the Phoenix ID panels are modifications of the classical methods These include tests for fermentation oxidation degradation and hydrolysis of various substrates In addition to these the Phoenix system utilizes chromogenic and fluorogenic substrates as well as single carbon source substrates in the identification of organisms The modern broth microdilution test used today has origins in the tube dilution test used in 1942 by Rammelkamp and Maxo
17. Patrick R et al ed Manual of Clinical Microbiology 8 Edition ASM Press Washington D C 2003 18 CLSI M7 A6 Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically Approved Standard Sixth Edition January 2003 Manufactured by Becton Dickinson and Company 7 Loveton Circle Sparks MD 21152 USA 800 638 8663 Made in USA TECHNICAL INFORMATION Approved by Date Effective Supervisor Date Director Date Reviewed 9 2006 Phoenix BDXpert BBL CrystalSpec PhoenixSpec Trypticase and BD are trademarks of Becton Dickinson and Company ATCC is a trademark of American Type Culture Collection Sharpie is a trademark of Sanford CHROMagar is a trademark of Dr A Rambach Page 13 of 30 Table A Taxa for ID AST Determination There are antimicrobial agents for use with the Phoenix system that are not proven to be effective for treating infections for all organisms listed in this section For interpreting and reporting results of antimicrobial agents that have shown to be active against organism groups both in vitro and in clinical infections refer to the individual pharmaceutical antimicrobial agent labeling Alternatively refer to the most recent CLSI M100 Performance Standard Table 1 Suggested Groupings of US FDA Approved Antimicrobial Agents That Should be Considered for Routine Testing and Reporting on Organisms by Clinical Microbiological Laboratories Gram Negative 0
18. amicin Escherichia coli 1 16 Piperacillin Morganella morganii 4 128 Achromobacter species 4 128 Piperacillin Achromobacter species 2 4 128 4 Tazobactam Serratia marcescens 4 4 128 4 Serratia species 4 4 128 4 Tetracycline Morganella morganii 1 16 Ticarcillin Achromobacter species 4 128 Alcaligenes species 4 128 Brevundimonas species 4 128 Chryseobacterium species 4 128 Delftia acidoverans 4 128 Myroides species 4 128 Ochrobactrum anthropi 4 128 Providencia species 4 128 Ralstonia species 4 128 Salmonella species 4 128 Serratia species 4 128 Shewanella species 4 128 Shingobacterium species 4 128 Wautersia species 4 128 Ticarcillin Citrobacter freundii 4 2 128 2 Clavulanate Morganella morganii 4 2 128 2 Tobramycin Enterobacter aerogenes 0 5 16 Trimethoprim Enterobacter aerogenes 1 16 Proteus mirabilis 1 16 LIMITATIONS OF THE PROCEDURE See the package insert shipped with the panel for specific organism antimicrobial limitations General A Gram stain test is required for the selection of the appropriate Phoenix panel types Accurate identification and or AST results may not be made without this test Use only well isolated bacterial colonies from one of the recommended primary isolation media See Table C Media containing esculin should not be used Use of mixed colonies could result in inaccurate identification and or AST interpretations Page 9 of 30 If the instrument inoculum density is configured to 0 5 for
19. atory Quality Control Refer to the Package Insert that accompanies the Phoenix panels for expected ID and AST results for QC organisms For instructions for QC panel login and loading refer to the BD Phoenix System User s Manual Panel Login and Inserting Panels in the Instrument ID NMIC ID and NID panels Escherichia coli ATCC 25922 Pseudomonas aeruginosa ATCC 27853 AST NMIC ID NMIC panels Page 7 of 30 Escherichia coli ATCCTM 25922 Pseudomonas aeruginosa ATCC 27853 Escherichia coli ATCC 35218 Klebsiella pneumoniae ATCC 700603 For the most reliable results it is recommended that the QC organisms be subcultured at least twice on two consecutive days onto TSA II with 5 Sheep Blood Agar before use in the Phoenix system Compare recorded results to those listed in the Package Insert If discrepant results are obtained review test procedures as well as confirm purity of the quality control strain used before contacting BD Diagnostics Technical Services Department Unacceptable QC results are documented as Fail and acceptable QC results are documented as Pass on the QC Report RESULTS Organism identification will appear on the Phoenix Report Form with a probability percentage from the Phoenix database based on the substrate reaction profile Results from each substrate will appear as V or X for each reaction The MIC results and Interpretive Categorical Results SIR will be shown for the a
20. blue color Store Phoenix AST Indicator Solution at 2 8 C Each bottle contains enough solution to test up to 100 panels Expiration dating is shown on the box pouch and bottle label and is for unopened bottles An opened bottle is stable for up to 14 days if stored at 2 8 C Be sure the bottle is held vertically when dispensing the AST Indicator Solution SPECIMEN COLLECTION AND PROCESSING The Phoenix system is not for use directly with clinical specimens Only pure culture isolates of aerobic and or facultatively anaerobic Gram Negative organisms are acceptable for testing The test isolate must be a pure culture It is recommended that cultures be no more than 24 hours old unless additional incubation is required to achieve sufficient growth Isolates must be tested with a Gram stain test to assure the appropriate selection of Phoenix panel type Once the Gram stain reaction is confirmed select the appropriate Phoenix panel for Page 3 of 30 inoculation e g NMIC ID panel for use with Gram Negative organisms Selection of the incorrect panel type could lead to incorrect results For AST testing in the Phoenix system isolates recovered from non selective media are recommended It is recommended that media containing antibiotics not be used for organisms to be tested in the Phoenix system Selective media may inhibit some strains of bacteria therefore caution must be used when selecting isolated colonies from these media For ID and
21. cterium indologenes Chryseobacterium meningosepticum Chryseobacterium scophthalmum Citrobacter amalonaticus Citrobacter braakii Citrobacter farmeri Citrobacter freundii Citrobacter gillenii ID AST AST ID AST ID AST ID AST ID AST AST ID AST ID AST ID ID ID AST ID AST ID AST ID AST ID AST ID AST AST AST ID ID AST ID AST ID AST AST ID AST ID AST ID AST ID AST AST Page 15 of 30 Citrobacter koseri Citrobacter murliniae Citrobacter rodentium Citrobacter sedlakii Citrobacter werkmanii Citrobacter youngae Comamonas terrigena Comamonas testosteroni Delftia acidovorans Edwardsiella hoshinae Edwardsiella ictaluri Edwardsiella tarda Eikenella corrodens Empedobacter brevis Enterobacter aerogenes Enterobacter amnigenus Enterobacter amnigenus biogroup 1 Enterobacter amnigenus biogroup 2 Enterobacter asburiae Enterobacter cancerogenus Enterobacter cloacae Enterobacter cowanii Enterobacter dissolvens Enterobacter gergoviae Enterobacter hormaechei Enterobacter intermedius Enterobacter kobei Enterobacter nimipressuralis Enterobacter sakazakii Escherichia blattae Escherichia coli Escherichia fergusonii Escherichia hermannii Escherichia vulneris Ewingella americana ID AST AST AST ID AST ID AST ID AST ID ID ID AST ID AST ID AST ID AST ID ID ID AST AST ID AST ID AST ID AST ID AST ID AST AST AST ID AST ID AST ID AST AST AST ID AST AST ID AST ID AST ID AST ID AST ID Page
22. e of resistant strains precludes defining any result categories other than susceptible For strains yielding results suggestive of a nonsusceptible category organism identification and antimicrobial susceptibility test results should be confirmed Subsequently the isolates should be saved and submitted to a reference laboratory that will confirm the result using the CLSI reference dilution method PERFORMANCE CHARACTERISTICS Gram Negative Identification In two internal studies the performance of the Phoenix Gram Negative identification was evaluated The 0 5 inoculum density configuration and the 0 25 inoculum density configuration were tested with 721 strains 0 5 and 784 strains 0 25 respectively Enteric and non enteric results were evaluated against commercial and non commercial methods The Phoenix Gram Negative identification performance is outlined below McFarland Agreement No Agreement No ID Species Level 0 5 95 6 3 6 0 8 0 25 98 1 1 4 0 5 An internal study was performed to simulate inter site reproducibility The identification results obtained using the Phoenix system were compared with expected results This performance testing demonstrated intra site and inter site reproducibility of at least 95 or greater Confirmatory ESBL Test To determine the accuracy of the Phoenix Confirmatory ESBL test accuracy testing was performed at multiple sites using Clinical and Challenge isolates The results from the ESB
23. ed Media Approved Use ID AST Trypticase Soy Agar with 5 Sheep Blood Yes Yes Bromthymol Blue BTB Lactose Agar Yes Yes BBL CHROMagar Orientation Yes Yes Chocolate Agar Yes Yes Columbia Agar with 5 Horse Blood Yes Yes Columbia Agar with 5 Sheep Blood Yes Yes Cystine Lactose Electrolyte Deficient CLED Agar Yes Yes Dey Egley D E Neutralizing Agar Yes No Eosin Methylene Blue Yes Yes Hektoen Enteric Agar Yes No MacConkey Agar Yes Yes Trypticase Soy Agar without Blood Yes No Trypticase Soy Agar with Lecithin and Tween Yes No 80 Xylose Lysine Desoxycholate Agar Yes No Page 30 of 30
24. ena Pseudomonas pseudoalcaligenes Pseudomonas putida Pseudomonas species Pseudomonas stutzeri Pseudomonas veronii Rahnella aquatilis Ralstonia pickettii Ralstonia solanacearum Ralstonia species Raoultella ornithinolytica Raoultella planticola Raoultella terrigena Rhizobium radiobacter Salmonella aberdeen Salmonella abortus equi Salmonella adelaide Salmonella aderike AST ID AST AST ID AST ID AST ID AST AST ID AST ID AST ID AST ID AST AST ID AST ID AST ID AST AST ID AST AST ID AST ID AST ID AST ID AST AST ID ID AST AST AST ID AST AST AST ID AST AST AST AST Page 18 of 30 Salmonella agona Salmonella alachua Salmonella anatum Salmonella arizonae Salmonella avana Salmonella bahrenfeld Salmonella blockley Salmonella bongori Salmonella braenderup Salmonella bredeney Salmonella bunn Salmonella california Salmonella carrau Salmonella cerro Salmonella champaign Salmonella chittagong Salmonella cholerasuis Salmonella choleraesuis ssp arizonae Salmonella choleraesuis ssp choleraesuis Salmonella choleraesuis ssp diarizonae Salmonella choleraesuis ssp houtenae Salmonella choleraesuis ssp indica Salmonella choleraesuis ssp salamae Salmonella cubana Salmonella dakar Salmonella daressalaam Salmonella derby Salmonella dessau Salmonella DT Salmonella dublin Salmonella duesseldorf Salmonella enteritidis Salmonella fresno Salmonella gallinarum Salmonella give AST
25. eriol 67 217 226 8 Soto O B 1949 Fermentation Reactions with Dried Paper Discs Containing Carbohydrate and Indicator Puerto Rican J Publ Hith Trop Med 25 96 100 9 Weaver R H 1954 Quicker Bacteriological Results Am J Med Technol 20 14 26 10 K mpfer P Rauhoff O and Dott W 1991 Glycosidase Profiles of Members of the Family Enterobacteriaceae J Clin Microbiol 29 2877 2879 Page 12 of 30 96 8 98 3 94 4 92 9 92 7 96 7 93 3 94 4 90 9 93 3 89 0 98 7 97 7 96 2 84 4 98 6 92 3 11 Manafi M Kneifel W and Bascomb S 1991 Fluorogenic and Chromogenic Substrates Used in Bacterial Diagnostics Microbiol Rev 55 335 348 12 Rammelkamp C H and Maxon T 1942 Resistance of Staphylococcus aureus to the Action of Penicillin Proc Soc Biol and Med 51 386 389 13 Marymont J H and Wentz R M 1966 Serial Dilution Antibiotic Sensitivity Testing with the Microtitrator System Am J Clin Pathol 45 548 551 14 Gavan T L and Town M A 1970 A Microdilution Method for Antibiotic Susceptibility Testing An Evaluation Am J Clin Pathol 53 880 885 15 Lancaster M V and Fields R D 1996 Antibiotic and Cytotoxic Drug Susceptibility Assays Using Resazurin and Poising Agents U S Patent 5 501 959 16 CLSI M100 S15 Performance Standards for Antimicrobial Susceptibility Testing Fifteenth Informational Supplement January 2005 17 Murray
26. esterstede Salmonella worthington Serratia entomophilia Serratia ficaria Serratia fonticola Serratia grimesii Serratia liquifaciens Serratia marcescens Serratia odorifera Serratia odorifera 1 Serratia odorifera 2 Serratia plymuthica Serratia proteamaculans ssp proteamaculans Serratia proteamaculans ssp quinovora Serratia rubidaea Shewanella algae Shewanella putrefaciens Shigella boydii Shigella dysenteriae AST ID AST AST AST ID AST AST AST AST AST AST ID AST AST AST ID AST AST AST AST AST AST ID AST ID AST AST ID AST ID AST AST ID AST ID AST ID AST AST AST ID AST AST ID AST ID AST ID AST Page 21 of 30 Shigella flexneri Shigella sonnei Shigella species Sphingobacterium multivorum Sphingobacterium spiritivorum Sphingobacterium thalpophilum Sphingomonas paucimobilis Stenotrophomonas maltophilia Suttonella indologenes Tatumella ptyseos Vibrio alginolyticus Vibrio cholerae Vibrio fluvialis Vibrio hollisae Vibrio metschnikovii Vibrio mimicus Vibrio parahaemolyticus Vibrio vulnificus Wautersia gilardii Wautersia paucula Weeksella virosa Yersinia aldovae Yersinia bercovieri Yersinia enterocolitica Yersinia frederiksenii Yersinia intermedia Yersinia kristensenii Yersinia mollaretii Yersinia pseudotuberculosis Yersinia rohdei Yersinia ruckeri Yokenella regensburgei Not all species encountered during clinical performance evaluations ID AST ID AST ID AST ID AST
27. h 3 Place the panel on the Inoculation Station with ports at the top and pad on the bottom 4 Label a Phoenix ID Broth tube with the patients specimen number Using aseptic technique pick colonies of the same morphology with the tip of a sterile cotton swab do not use a polyester swab or a wooden applicator stick from one of the recommended media See Table C 5 Suspend the colonies in the Phoenix ID Broth 4 5 mL Cap the tube and vortex for 5 seconds 7 Allow approximately ten seconds for air bubbles to surface Tap the tube gently to aid in eliminating bubbles 8 Confirm default settings for inoculum density before inoculating panels Insert the tube into the BBL CrystalSpec or BD PhoenixSpec Nephelometer Make sure the tube is inserted as far as it will go Note Only the BD PhoenixSpec Nephelometer can be used to make inoculum densities of 0 25 McFarland Refer to the BBL CrystalSpec Nephelometer or BD PhoenixSpec product insert for correct usage instructions and calibration verification 9 If the inoculum density is set to 0 5 McFarland for the panel type being run then a range of 0 50 0 60 is acceptable If the inoculum density is set to 0 25 for the panel type being run then a range of 0 20 0 30 is acceptable If the density of organisms is low you can add colonies from the isolate Re vortex the sample and reread to confirm that the correct density has been achieved If the density of organisms exceeds 0 6 McFarland fol
28. io hollisae Vibrio metschnikovii Vibrio mimicus Vibrio parahaemolyticus Vibrio vulnificus Wautersia paucula Weeksella virosa Yersinia enterocolitica Yersinia frederiksenii Yersinia intermedia Yersinia kristensenii Yersinia pseudotuberculosis Yersinia ruckeri Yokenella regensburgei Not all species encountered during clinical performance evaluations ID AST ID ID AST ID AST ID AST ID AST ID AST ID AST ID Page 27 of 30 Table B List of Reagents and Principles Employed in the Phoenix System Substrate Name Code Principle L PHENYLALANINE AMC A_LPHET 4MU N ACETYL BD GLUCOSAM A_NAG INIDE L GLUTAMIC ACID AMC A_LGTA L TRYPTOPHAN AMC A_LTRY L PYROGLUTAMIC ACID AMC A LPYR Enzymatic hydrolysis of the amide or L PROLINE AMC A LPROB glycosidic bond results in the release of a L ARGININE AMC A_LARGH fluorescent coumarin or ARGININE ARGININE AMC A ARARR 4 methylumbelliferone derivative GLYCINE AMC A GLYB L LEUCINE AMC A LLEUH LYSINE ALANINE AMC A LYALD GLUTARYL GLYCINE ARGININE A GUGAH AMC GLYCINE PROLINE AMC A GLPRB COLISTIN C CLST Resistance to the antimicrobial agents results in a reduction of resazurin based indicator POLYMYXIN B C PXB D MANNITOL C DMNT CITRATE C CIT ACETATE C ACT ADONITOL C ADO Utilization of a carbon source results in a MALONATE C MLO reduction of the resazurin based indicator ALPH
29. lated media e g Trypticase Soy Agar with 5 Sheep Blood Incubators Biohazard disposable container Markers etc Page 4 of 30 PHOENIX TEST PROCEDURE Note The Phoenix instrument should always be powered on If it is not power on the instrument and allow 2 hours for the instrument to warm up before loading panels Prepare the Phoenix instrument to receive new panels as described in the BD Phoenix System User s Manual Operation Daily System Maintenance Care should be exercised when handling Phoenix panels You should handle panels by the sides only to avoid marking smudging or obscuring the front or back of the panel in any way Accession barcode labels affixed to a Phoenix panel should Not be of fluorescent material Not cover any Phoenix panel reaction wells Not cover the Phoenix panel sequence number barcode Broth and Panel Preparation 1 Confirm the Gram stain reaction of the isolate before proceeding with the inoculum preparation for use in the Phoenix instrument Once the Gram stain reaction is confirmed select the appropriate Phoenix panel for inoculation Selection of the incorrect panel type could lead to incorrect results 2 Examine the pouch and do not use the panel if the pouch is punctured or opened Remove the panel from the pouch Discard the desiccant Do not use the panel if there is no desiccant or if the desiccant pouch is torn Note Panels must be used within 2 hours of being removed from the pouc
30. low the steps below to dilute the broth It is very important to accurately fill the wells in the panel Note The standardized bacterial suspension in ID broth must be used within 60 minutes of preparation a Using a marker mark the broth level in the over inoculated Phoenix ID Broth tube Page 5 of 30 10 11 12 13 14 15 16 17 18 b Using a sterile pipette aseptically add fresh Phoenix ID Broth to the inoculum Only Phoenix ID broth may be used to dilute the inoculum c Vortex the tube and allow to sit for 10 seconds d Place the tube in the nephelometer and remeasure the turbidity of the suspension e lf the reader is greater than 0 6 repeat steps b d e If the reading is 0 5 0 6 go to Step e e Using a sterile pipette aseptically remove excess broth to the original level indicated by the mark on the tube created in Step a Remove excess broth to avoid overfilling the panel Also do not removed too much broth as there may be insufficient broth to adequately fill the panel f Broth may now be used to inoculate the Phoenix AST Broth and or the Phoenix panel If you are performing identification only proceed to Step 15 and continue the procedure Label a Phoenix AST Broth tube 8 0 mL with the patient s specimen number Holding the AST Indicator Solution bottle vertically add one free falling drop of AST indicator solution to the AST broth tube Invert to mix DO NOT VORTEX Note Allow AST Indicator
31. n to determine in vitro antimicrobial susceptibility testing of bacterial isolates from clinical specimens The broth dilution technique involves exposing bacteria to decreasing concentrations of antimicrobial agents in liquid media by serial two fold dilutions The lowest concentration of an antimicrobial agent in which no visible growth occurs is defined as the minimal inhibitory concentration MIC The introduction in 1956 of a microtitrator system using calibrated precision spiral wire loops and droppers for making accurate dilutions rapidly allowed Marymont and Wentz to develop a serial dilution antimicrobial susceptibility test AST 3 The microtitrator system was accurate and allowed the reduction in volumes of antimicrobial agents The term microdilution appeared in 1970 to describe the MIC tests performed in volumes of 0 1 mL or less of antimicrobial solution The Phoenix AST test is a modified miniaturized version of the micro broth doubling dilution technique Susceptibility testing in the Phoenix system is performed through determination of bacterial growth in the presence of various concentrations of the antimicrobial agent tested PRINCIPLES OF THE PROCEDURE A maximum of 100 identification and antimicrobial susceptibility tests can be performed in the Phoenix instrument at a time using Phoenix ID AST combination panels A sealed and self inoculating molded polystyrene tray with 136 micro wells containing dried reagents serves as
32. ng sterilize specimen containers and other contaminated materials by autoclaving Panels once inoculated should be handled carefully until placed in the instrument STORAGE AND HANDLING Phoenix Panels Panels are individually packaged and must be stored unopened at room temperature 15 25 C Do not refrigerate or freeze Visually inspect the package for holes or cracks in the foil package Do not use if the packaging or panel appears to be damaged If stored as recommended the panels will retain expected reactivity until the date of expiration Phoenix ID Broth Tubes are packaged as 100 tube packs Visually inspect the tubes for cracks leaks etc Do not use if there appears to be a leak tube or cap damage or visual evidence of contamination i e haziness turbidity Store Phoenix ID Broth tubes at 2 25 C Expiration dating is shown on the tube label Phoenix AST Broth Tubes are packaged as 100 tube packs Visually inspect the tubes for cracks leaks etc Do not use if there appears to be a leak tube or cap damage or visual evidence of contamination i e haziness turbidity Store Phoenix AST Broth tubes at 2 25 C Expiration dating is shown on the tube label Phoenix AST Indicator Solution The indicator solution is individually pouched and packaged as a package of 10 dropper bottles Visually inspect the bottle for cracks leaks etc Do not use if there appears to be a leak bottle or cap damage or any change from a dark
33. ppropriate organism antimicrobial agent combinations Special messages will be shown when the BDXpert System detects results that are of particular clinical interest Further information concerning results obtained from the Phoenix system can be found in the BD Phoenix System User s Manual Obtaining Results Messages Error messages may appear if the system detects unexpected reactivity due to inappropriate procedure or instrument malfunction For a complete listing of error codes and their meaning refer to the BD Phoenix System User s Manual System Alerts Needs Attention and Troubleshooting Special Notes In general the Phoenix System provides a MIC for all organisms at any of the concentrations defined on a specific panel For certain antimicrobic organism combinations a specific minimum or maximum MIC is reported even if there is a lower or higher concentration on the panel These MIC values are applied by the software and are reported out as less than or equal to lt for the minimum MIC or greater than gt for the maximum MIC The table below provides the range for these special antimicrobic organism combinations Antimicrobial Agent Organism s Applied Range ug mL Amikacin Morganella morganii 2 64 Proteus penneri 2 64 Proteus vulgaris 2 64 Providencia species 2 64 Aztreonam Providencia stuartii 2 64 Page 8 of 30 Cefotaxime Providencia species 2 64 Cefotetan Proteus mirabilis 4 64 Gent
34. read only by the instrument Phoenix panels cannot be read manually Bacterial Identification The ID portion of the Phoenix panel utilizes a series of conventional chromogenic and fluorogenic biochemical tests to determine the identification of the organism Both growth based and enzymatic substrates are employed to cover the different types of reactivity in the range of taxa The tests are based on microbial utilization and degradation of specific substrates detected by various indicator systems Acid production is indicated by a change in the phenol red indicator when an isolate is able to utilize a carbohydrate substrate Chromogenic substrates produce a yellow color upon enzymatic hydrolysis of either p nitrophenyl or p nitroanilide compounds Enzymatic hydrolysis of fluorogenic substrates results in the release of a fluorescent coumarin derivative Organisms that utilize a specific carbon source reduce the resazurin based indicator In addition there are other tests that detect the ability of an organism to hydrolyze degrade reduce or otherwise utilize a substrate A complete list of taxa that comprises the Phoenix ID Database is provided in Table A Reactions employed by various substrates and the principles employed in the Phoenix ID reactions are described in Table B Antimicrobial Susceptibility Testing The Phoenix AST method is a broth based microdilution test The Phoenix system utilizes a redox indicator for the detection of organism
35. stigianii Providencia stuartii Pseudomonas aeruginosa Pseudomonas fluorescens Pseudomonas luteola Pseudomonas mendocina Pseudomonas oryzihabitans ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID ID ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST Page 25 of 30 Pseudomonas putida Pueudomonas stutzeri Rahnella aquatilis Ralstonia pickettii Raoultella ornithinolytica Rhizobium radiobacter Salmonella choleraesuis ssp arizonae Salmonella choleraesuis ssp choleraesuis Salmonella gallinarum Salmonella paratyphi A Salmonella pullorum Salmonella species Salmonella typhi Serratia ficaria Serratia fonticola Serratia liquifaciens Serratia marcescens Serratia odorifera 1 Serratia odorifera 2 Serratia plymuthica Serratia rubidaea Shewanella putrefaciens Shigella boydii Shigella dysenteriae Shigella flexneri Shigella sonnei Sphingobacterium multivorum Sphingobacterium spiritivorum Sphingobacterium thalpophilum Sphingomonas paucimobilis Stenotrophomonas maltophilia Suttonella indologenes Tatumella ptyseos Vibrio alginolyticus Vibrio cholerae ID AST ID AST ID ID AST ID AST ID ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID AST ID ID AST Page 26 of 30 Vibrio fluvialis Vibr
36. the panel type being used an inoculum density of 0 50 0 60 must be met Only the BBL CrystalSpec or BD Phoenix Spec Nephelometer can be used to measure the inoculum density If the instrument inoculum density is configured to 0 25 for the panel type being used an inoculum density of 0 20 0 30 must be met Only the BD PhoenixSpec Nephelometer can be used to measure inoculum density for this range Phoenix panels can be read only by the Phoenix instrument Visual interpretation of the Phoenix panels is not possible Any attempt to manually interpret results from the panel may lead to misidentification and or inaccurate AST interpretations Identification The unique panel environment combined with the shortened incubation time may result in Phoenix panel reactions varying from those obtained using conventional biochemical media Antimicrobial Susceptibility Testing After the addition of Phoenix AST Indicator Solution to the AST broth tubes mix by inversion DO NOT VORTEX Vortexing may cause air bubbles to form in the AST broth which can result in inappropriate filling of the Phoenix panel during inoculation Because of the low probability of occurrence or special growth requirements some organisms included in the ID taxa are not included in the AST database These organisms will display the message Organism not included in the AST database perform alternate method For some organism antimicrobial combinations the absenc
37. um Density Flexibility You may run the ID portion of a panel in the opposite mode from what is configured by darkening well A17 on the back of the panel before placing the panel in the instrument This allows you to run a panel at an inoculum density of 0 20 0 30 even if you are configured for a density of 0 5 for that particular panel type Likewise you can run a panel at an inoculum density of 0 50 0 60 if you are configured for a density of 0 25 There is no way to alter the density setting during Panel Login To use a panel in the opposite density mode using a black Sharpie permanent marker blacken the entire well See the BD Phoenix System User s Manual Operation ID Inoculum Density Flexibility for position of well A17 For instructions for panel login and loading refer to the BD Phoenix System User s Manual Panel Login and Inserting Panels in the Instrument Current Instrument modum inceulunkDensit Concentration Amount of ID Inoculum to Well A 17 ERO RA y Desired for Test Add to AST Broth 9 Panel 0 50 0 25 50 uL Blackened 0 25 0 50 25 uL Blackened If also running AST USER QUALITY CONTROL In order to ensure appropriate set up procedure and acceptable performance of the system the following organisms are recommended for testing The user is advised to review the individual AST panel formats to determine if all test strains need to be tested for routine labor
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