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Meningococcal Reference Unit user manual

1. Health Protection Agency Meningococcal Reference Unit Part of HPA Respiratory and Systemic Infections Department RSID User Manual April 2012 Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 1 of 26 Contains Information on References Services for gt Neisseria meningitidis isolate characterisation gt Polysaccharide antigen detection gt Neisseria meningitidis Meningococcal DNA detection by PCR Streptococcus pneumoniae detection by PCR gt Vaccine response pre and post immunisation Postal address Meningococcal Reference Unit Manchester Medical Microbiology Partnership PO Box 209 Clinical Sciences Building 2 Manchester Royal Infirmary Oxford Road Manchester UK M13 9WZ Hays address DX 6962410 Manchester 90 M Telephone 44 0 161 276 6757 Fax 44 0 161 276 5744 Out of hours Telephone 44 0 161 276 1234 and ask for Medical Microbiologist on call Authorised By Dr E Kaczmarski Head of Unit Effective Date April 2012 Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 2 of 26 Introduction gs cectesace 5a teraacteacetcace cede deseaweneasaadecseeeinnece te unaeseeeenesanceacces 4 Summary of Services and ReESOUICES rererere 6 Services AvAIADIE ccccceeecceeeececeaeeueeeseeeeeeeeeeeeeeeseaeansnseees 7 Key Factors affecting specimen performance cccce 18 MRU Price List sixcccdeccteeteeatentcstnceeaotaeseeie
2. Road Downend Bristol BS16 2SF Tel 44 0 0117 373 73 73 Fax 44 0 0117 373 73 74 Email information meningitisUK org Web www meningitisUK org Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 26 of 26
3. confirm N meningitidis and determine epidemiological markers The identification of Neisseria species other than N meningitidis N lactamica and N gonorrhoeae is problematic Please do not submit isolates or organisms that are very unlikely to be N meningitidis or lactamica N gonorrhoeae other Neisseria spp and Moraxella spp should be referred to Cfl Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 18 of 26 Meningococcal Reference Unit 2012 2013 price list Turn round time for Provisional result working days Turn round time to Final result working days Meningococcal cultures outside Provisional results are telephoned within 24 72 hours Printed reports issued within England amp Wales where FOC 1 2 weeks 2 weeks Meningococcal PCR outside England amp Wales where FOC 24 hours 48 72 hours Meningococcal serology serum bactericidal assay per target 28 days 28 days Meningococcal serogroup specific IgG 28 days 28 days Contact Mr Richard Mallard richard mallard hpa org uk for details of current prices MENINGOCOCCAL REFERENCE UNIT Edition no 05 Issue date April 2012 Page 19 of 26 SPECIMEN ACCEPTANCE POLICY GUIDANCE FOR LABORATORIES AND HEALTH PROTECTION UNITS LABELLING YOUR SPECIMENS MATTERS Specimens must be correctly labelled and request forms adequately completed Minimise specimen rejection confusion delay by Please f
4. NGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 8 of 26 Transport containers and documentation It is the responsibility of senders to comply with the current transport legislation and safety recommendations Refer to http www dft gov uk 426155 425453 800 300 infectioussubstances pdf for current guidelines Complaints Should it be necessary to submit a formal complaint to the MRU about our service please contact either Dr Ed Kaczmarski or Dr Steve Gray Turnaround times for isolate characterisation Optimal turnaround times are conditional on receiving established pure cultures with appropriate documentation Serogroup results for clinical isolates will be telephoned to the sending laboratory as soon as available usually later on the day of receipt Monday to Friday The telephone report is logged but not printed A final report comprising the serogroup and phenotypic characterisation serotyping and serosubtyping and antibiotic MIC profile are reported within 7 working days Contact the MRU for urgent weekend reporting or if more rapid results are required e g for cluster investigation The determination of molecular subtyping porA sequencing variable regions VR1 VR2 and VR3 is reported when available 7 10 days usually as an additional report Urgent culture specimens In circumstances where urgent characterisation of an isolate is required a provisional serogroup result can be available withi
5. R specimens These should be discussed with a member of the MRU staff who will liaise with colleagues performing the assays and make arrangements for the earliest possible testing Contact details will be required especially any out of hours contact at the sending laboratory relevant CCDC or HPU Please do not send urgent samples likely to arrive out of hours without first discussing with MRU staff Refer to out of hours section on page 8 Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 15 of 26 Antigen detection non culture confirmation Polysaccharide antigen detection Meningococcal antigen detection using commercial latex agglutination kits is available on request Please discuss with a member of MRU staff before the sample is submitted For acute investigations PCR is preferred as it is more sensitive and if positive additional molecular typing can be performed NB Antigen detection will reduce material available for PCR and could compromise the integrity of the sample What specimens to send for polysaccharide antigen detection CSF and serum a minimum sample volume of 200uL is needed Turnaround time Telephone reports are available on day of receipt Printed reports normally sent out on the following working day Urgent specimen These can be processed and results telephoned within two hours of receipt at the MRU Please discuss with the MRU if urgent antigen tests are require
6. USA e Kuipers B van den Dopplesteen G Wedege E amp van Alphen L 2001 Serological characterization In Meningococcal Disease methods and protocols pp131 145 Edited by Pollard A J amp Maiden M C J Humana Press Inc Totowa New Jersey USA e Rosenavist E Wedege E Hoiby E A amp Froholm L O 1990 Serogroup determination of Neisseria meningitidis by whole cell ELISA dot blotting and agglutination APMIS 1990 98 501 506 e Suker J Feavers IM and Maiden MC Monoclonal antibody recognition of members of the meningococcal P1 10 variable region family implications for serological typing and vaccine design Microbiology 1996 142 63 69 e Wedege E Hoiby E A Rosenqvist E amp Froholm L O Serotyping and serosubtyping of Neisseria meningitidis isolates by co agglutination dot blotting and ELISA J Med Microbiol 1990 31 195 201 Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 25 of 26 Meningitis Trust Head Office Fern House Bath Road STROUD Gloucestershire GL5 3TJ United Kingdom 24 hour free phone helpline 0800 028 18 28 Tel 44 0 1453 768000 Fax 44 0 1453 768001 E mail info meningitis trust org http www meningitis trust org Meningitis Research Foundation Midland Way Thornbury Bristol BS25 2BS Tel 44 0 01454 281811 Fax 44 0 01454 281094 Email info meningitis org http www meningitis org Meningitis UK 25 Cleeve Wood
7. amsay ME Guiver M Fox AJ Borrow R Mallard RH and Kaczmarski EB Epidemiology of meningococcal disease in England and Wales 1993 94 to 2003 04 contribution and experiences of the Meningococcal Reference Unit J Med Microbiol 2006 55 887 896 e Gray SJ Borrow R Kaczmarski EB 2001 Meningococcal serology In Meningococcal Disease Methods in Molecular Medicine eds Pollard AJ and Maiden MCJ pp 61 87 Humana Press Totowa New Jersey e Borrow R Carlone GM 2001 Seogroup B and C serum bactericidal assays In Meningococcal Vaccines Methods in Molecular Medicine eds Pollard AJ and Maiden MCV pp 289 308 Humana Press Totowa New Jersey e Corless CE Guiver M Borrow R Edwards Jones V Fox AJ Kaczmarski EB Simultaneous detection of Neisseria meningitidis Haemophilus influenzae and Streptococcus pneumoniae in suspected cases of meningitis and septicemia using real time PCR J Clin Microbiol 2001 39 1553 8 e Cartwright K Meningococcal Disease J Wiley amp Sons 1995 e Feavers IM Fox AJ Gray SJ Jones DM and Maiden MC Antigenic diversity of meningococcal outer membrane protein porA has implications for epidemiological analysis and vaccine design Clin Diagn Lab Immunol 1996 3 4 444 450 e Jolley K Urwin R Suker J and Gray SJ 2006 Methods for meningococcal typing in Handbook of Meningococcal Disease Infection biology and Clinical Management Editors M Frosch and MCJ Maiden Chpt3 37 51 Wiley VCH
8. atories supplied their telephone and secure fax numbers see page 21 The following information is important for accurate patient data reconciliation and assists provision of meaningful local statistics date of birth home post code health district of residence Isolates Please send cultures from all positive sites For all isolates e Presenting clinical features i e meningitis septicaemia both if other please give details Where relevant e Names of other possibly related cases e In contact tracing the name of the index case and location school town etc e Recent travel details if there is a possibility of the disease being contracted abroad Meningococcal PCR e Type of specimen EDTA Heparin Serum CSF e Time elapsed since illness onset e Whether and when parenteral antibiotics have been given in relation to sample collection Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 21 of 26 How to send isolates transport Only submit viable isolate samples in approved packaging UN3373 which are suitable for Royal Mail post airfreight or commercial couriers such as HAYS DX Agar slopes where possible pure viable cultures inoculated on chocolate heated blood agar blood agar or Dorset egg slopes after establishing growth by overnight incubation at 37 C On occasion it may be necessary to submit an unincubated culture This can save time but requires a heavy inoculum to ens
9. boration with its customers Customers are invited to review our arrangements in conjunction with the Caldicott Guardian Customers are also asked to draw to the Caldicott Guardian s attention any instances where PID security has been threatened or has broken down Uses that PID are put to outside clinical diagnostic services generally allow patient identifiers to have been removed before hand and when PID is used for research purposes the proposals are considered first by the HPA Research Ethics Committee All enquiries about the security and use of PID should be addressed to the Caldicott Guardian Prof F J Bolton Tel 0161276 5699 e mail eric bolton hpa org uk Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 24 of 26 e Kaczmarski EB Cartwright KAV Control of meningococcal disease guidance for microbiologists Comm Dis Rep Rev 1997 5 R196 R198 e Kaczmarski EB Meningococcal disease in England and Wales 1995 Comm Dis Rep Rev 1997 7 R55 R59 e Guiver M Borrow R Marsh J Gray SJ Kaczmarski EB Howells D Boseley P Fox Au Evaluation of the Applied Biosystems automated Taqman polymerase chain reaction system for the detection of meningococcal DNA FEMS Immunol Med Microbiol 2000 28 173 179 e Guiver M and Borrow R 2001 PCR diagnostics In Meningococcal Disease Methods in Molecular Medicine eds Pollard AJ and Maiden MCJ pp 23 39 Humana Press Totowa New Jersey e Gray SJ Trotter CL R
10. d Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 16 of 26 Meningococcal Serology a Serodiagnosis Serodiagnosis of meningococcal disease is not routinely available b Pre and post vaccine response The following services are available from the HPA Vaccine Evaluation Unit VEU based at the MMMP Functional total immunoglobulin and isotype specific antibody levels for immunogenicity studies by internationally standardised assays are available Samples of clotted blood or serum should be collected three to eight weeks post vaccination A minimum sample volume of 500uL is preferred There will be a charge for these investigations unless they are part of an MRU or HPA instigated epidemiological investigation 1 Quantitation of total IgG to serogroups C Y W135 or A polysaccharides 2 Functional antibody to serogroup C Y W135 or A meningococci by internationally standardised serum bactericidal assays SBAs 3 Novel assays bactericidal and ELISA for other meningococcal serogroups such as B are available on request Charges Requests for vaccine response testing if not initiated as part of an MRU or HPA epidemiological or case investigation will be charged for Turnaround Times Serogroup C vaccine response results are available within 28 working days of submission Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 17 of 26 What specimens to send All submitted sample
11. e MRU meningococcal ctrA screening assay is performed as part of a four component multiplex assay also comprising meningococcal serogroup B siaDe confirmatory assay ply pneumolysin for Streptococcus pneumoniae screen and an in house Internal Control IC assay process control The ctrA PCR assay confirms the presence of N meningitidis DNA specifically from capsulated meningococci as ctrA is involved in the transfer of polysaccharide to the cell surface The addition of another reverse ctrA primer to the originally published assay Corless et al 2001 since 2003 has allowed for the additional detection of a small subset of meningococci not confirmed by the initial assay currently unpublished but data on file Specific serogroup confirmatory assays are based on the sialylation siaD of the polysaccharide capsule for serogroups B C Y and W135 Serogroup A polysaccharide is chemically distinct and not sialylated hence the specific mynA assay Charges Note The HPA MRU meningococcal PCR assays are reference services for England and Wales epidemiology and when performed are free of charge The pneumococcal assays are currently free of charge as part of the HPA enhanced surveillance of pneumococcal disease post pneumococcal conjugate Prevenar vaccine introduction As part of the enhanced surveillance programme of pneumococcal disease pneumococcal DNA positive CSFs bloods and empyema fluids from patients under the age
12. el 44 0 20 8200 6868 E mail mary ramsay hpa org uk MENINGOCOCCAL REFERENCE UNIT Edition no 05 Issue date April 2012 Page 5 of 26 e Clinical advice for case and outbreak investigation and management e Meningococcal isolate confirmation and characterisation e Meningococcal DNA detection by PCR for non culture case confirmation e Molecular characterisation of meningococcal isolates and non culture DNA positive only material e Technical laboratory advice and support for large scale investigations e Meningococcal vaccine evaluation e Determination of response to meningococcal vaccination e Collection of gt 50 000 phenotypically characterised meningococcal isolates e Computerised database of laboratory confirmed cases e Collection of sera from laboratory proven cases of meningococcal disease e Support for collaborative scientific projects and audits Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 6 of 26 Routine Investigations Neisseria meningitidis isolate characterisation Species confirmation Phenotypic confirmation of Neisseria meningitidis isolates based on morphology and biochemical reactions Epidemiological characterisation of strains Phenotype a b c Serogroup identification of capsular polysaccharide antigens by serological reactions coagglutination using polyclonal antibodies in house commercial slide agglutination commercial latex antig
13. en kits and monoclonal antibodies Serotype identification of class 2 3 PorB outer membrane proteins by a dot blot ELISA using monoclonal antibodies Sero subtype identification of class 1 PorA outer membrane proteins by a dot blot ELISA using monoclonal antibodies supplied by HPA NIBSC Genotype a b c Serogroup use of PCR based serogroup confirmation enables identification of non viable organisms porA Serosubtype genetic characterisation of serosubtype by DNA sequencing Routinely tested and reported on all clinical isolates since October 2007 Multi Locus Sequence Typing MLST performed and reported on strains collected for epidemiological monitoring and some outbreak investigation Molecular subtyping of isolates Molecular characterisation of other potential typing targets such as fetA and fHbp a component of newly developed vaccines is undergoing evaluation Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 7 of 26 Antibiotic susceptibility testing Minimum Inhibitory concentrations MICs The MICs routinely determined on submitted isolates are penicillin cefotaxime rifampicin ciprofloxacin and sulphonamide sulphamethoxazole using Etest Biomerieux gradient diffusion methodology Other antibiotic susceptibility tests may be performed on request How to obtain MRU services Telephone contact For general enquiries 0161 276 6757 The MRU laboratory i
14. g assay but not confirmed with the specific siaD serogroup assays Hence menoingococcal DNA may be detected but serogroup genogroup not determined Very rarely non capsulated ctrA negative meningococci have been isolated from sterile sites and may be a cause of infection Pneumococcal PCR assays The samples found positive with the pneumolysin PCR ply screen in the four component multiplex assay are referred for confirmation using a pneumococcal specific autolysin ytA PCR Samples found to be positive with both ply and lytA are reported as Streptococcus pneumoniae or pneumococcal PCR positive Samples that are only ply positive alone may indicate a streptococcal species possibly an oral streptococcus species but are not reported as such They are reported as pneumococcal PCR Negative Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 13 of 26 Positive pneumococcal reports from blood in children under 2 years of age It should be noted that there have been reports in the literature of pneumococcal DNA detected PCR positive in the blood samples of children under the age of 2 years who are perfectly well and healthy It is for that reason that all positive pneumococcal reports from bloods of children less than two years of age recommend clinical interpretation of the molecular results Hib PCRs An additional and separate PCR assay to detect Haemophilus influenzae type b is available on request Other mo
15. gococcal specific screening ctrA assay for capsulated N meningitidis the meningococcal serogroup B siaDe confirmatory assay ply pneumolysin for Streptococcus pneumoniae screen and an in house Internal Control IC assay process control Samples positive with the Internal Control IC assay are considered suitable for testing and reporting The IC is added before nucleic acid extraction and assesses the sample for inhibitory qualities Such as may occur with badly lysed blood samples or often with blood culture bottle fluids If DNA extracts and not clinical samples are submitted to the MRU the process control is not added and potential inhibition cannot be assessed Meningococcal PCR Assays Samples positive with the ctrA PCR assay confirm the presence of N meningitidis DNA from capsulated meningococci To improve turnaround times the simultaneous confirmation of serogroup B by siaDg PCR assay is used Samples positive for ctrA but negative for siaDg are referred for specific serogroup C siaDc Y siaDY and W135 siaDy assays If serogroup A is being considered a vaery rare finding in the UK a specific mynA assay is used It should be noted that other serogroups such as X and 29E can cause invasive disease and are not currently confirmed molecularly by the HPA MRU It is not uncommon for low level positives i e samples with low numbers of genome copies organisms to be weakly positive with the sensitive ctrA screenin
16. ion porA sequencing and susceptibility testing of isolates Antigen detection PCR Requests for molecular epidemiology Initial contact for most MRU enquiries Tel 0161 276 6757 Dr Stephen J Gray Lead BMS Tel 44 0 161 276 6757 steve gray hpa org uk Mr Anthony Carr BMS2 tony carr hpa org uk Dr Lynne Newbold BMS2 lynne newbold hpa org uk Patient investigation and clinical advice Interpretation of results Outbreak investigation and management advice Vaccine evaluation research and development Vaccine response assessment Proposed research projects Other Key S Dr Edward Kaczmarski Head of MRU Tel 44 0 161276 5699 Mobile 07774243886 ed kaczmarski hpa org uk taff Professor Ray Borrow Deputy Unit Head of MRU Head of Vaccine Evaluation Unit VEU Tel 44 0 161 276 6793 ray borrow hpa org uk PCR diagnosis of N meningitidis Service and molecular research projects Dr Malcolm Guiver Head of Molecular Diagnostics Tel 44 0 161 276 8833 malcolm guiver hpa org uk Mr John Marsh Deputy Lead BMS Tel 44 0 161 276 5685 john marsh cmft nhs uk Database management Mr Richard Mallard Head of Laboratory Operations HPA North West Tel 44 0 161 276 5747 richard mallard hpa org uk Other sources of information Dr Mary Ramsay Consultant Epidemiologist Immunisation Hepatitis and Blood Safety Department HPA Centre for Infections 61 Colindale Avenue London NW9 5EQ T
17. lecular detection assays Situated within the MMMP molecular diagnostics department the MRU is able to request a variety of additional PCR based assays including viral causes of meningitis eg Herpes simplex enterovirus The additional assays are not part of the free meningococcal service but may be added to requests at the time of submission or later Should only limited amounts of unrepeatable samples be available this may be a cost effective option Nucleic acid extracts containing both DNA and RNA are available for rapid testing Additional viral PCRs will be invoiced If the additional viral PCRs are not requetsed on the original form it is necessary to provide the requesting clinician s name at the time of the telephone request Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 14 of 26 Availability of results Results on specimens received up to 10 00 on Monday Friday are normally available between 16 30 and 17 00 on the same day Positive results will be telephoned following serogroup confirmation up to 17 30pm or as soon as possible on the morning of the next working day when printed reports will also be sent out It is useful to telephone the MRU where a result is of particular urgency NB Although copy results are sent to the HPU it is the responsibility of the requesting laboratory to inform their local CCDC HPU of positive meningococcal PCR results in an appropriate timely fashion Urgent PC
18. me CSFs must be submitted in an appropriate sized container or tube Whole CSF i e an uncentrifuged specimen should be sent in small sterile containers such as a sterile 2mL screw capped vial rather than universal containers Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 11 of 26 Submission of minimum volumes is not preferred as repeat extraction is required to confirm positivity or the addition of molecular epidemiology assays Original CSF uncentrifuged or re suspended CSF deposits are preferred to CSF supernatants in order to increase sensitivity of detection Collection and timing of samples for PCR testing The likelihood of a positive PCR result decreases as the interval of sampling after starting antibiotics lengthens Blood samples for PCR taken more than 48 hours after commencement of antibiotic therapy are unlikely to give useful results CSF may remain positive for longer periods Any specimens for PCR tests should be stored at 4 C and not frozen prior to transport Freeze thawing may reduce the likelinood of positivity with low genome copy samples and can result in cracked or broken containers Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 12 of 26 MRU PCR investigations performed Meningococcal and Pneumococcal PCR investigations All suitable submitted samples are tested using a four component real time ABI Taqman multiplex assay comprising the menin
19. n two hours of receipt of an established culture When additional information of epidemiological importance such as serotyping and serosubtyping is needed rapidly a provisional phenotypic result can be available later the same working day if isolates are received before 10 00am Monday to Friday Arrangements to expedite urgent specimens should be made by telephone request to the MRU This is particularly important if samples are likely to arrive at the MRU later than 17 00 Monday Friday Molecular subtyping porA sequencing requires a minimum of 3 working days from receipt Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 9 of 26 Meningococcal DNA detection by PCR The MRU uses real time PCR ABI Taqman assays to confirm N meningitidis meningococcal disease and determine the infecting serogroup where possible The MRU meningococcal ctrA screening assays are performed as part of a multiplex assay combined with a pneumolysin PCR for Streptococcus pneumoniae detection Meningococcal detection by PCR The MRU uses real time PCR ABI Taqman assays to confirm N meningitidis meningococcal disease and determine the infecting serogroup where possible The MRU uses specific PCR assays to confirm the presence of N meningitidis and determine the genogroup the potential serogroup and also screens for the presence of pneumococcal DNA by two specific PCR assays pneumolysin ply and autolysin ytA Th
20. of 16 are routinely referred to HPA Colindale RSIL for serotype confirmation The pneumolysin PCR is also performed free of charge as part of the enhanced surveillance of pneumococcal disease post pneumococcal conjugate Prevenar vaccine introduction _Pneumolysin PCR positive samples are currently confirmed as Sir pneumoniae by an additional autolysin PCR assay As part of the enhanced surveillance pneumococcal DNA positive blood and empyema Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 10 of 26 fluids from patients under 16 years are referred to Cfl for serotypying What specimens to send for Meningococcal PCR MRU meningococcal PCR assays have been validated on EDTA whole blood CSF coagulated whole blood serum plasma and joint fluids EDTA whole blood and CSF are the preferred specimens Plasma or serum can be examined howver sensitivity may be compromised If coagulated bloods are submitted it is only possible to test the serum fraction EDTA blood 2 5 5 mL sample collected on admission should be sent routinely to the MRU in the event that PCR confirmation is required Smaller volumes 0 5 1mL from infants and babies can also be examined Heparinised or citrated samples can be tested but EDTA is preferred CSF samples if available should be sent in addition to an EDTA blood sample Definitive laboratory confirmation of meningococcal meningitis can only be made by analysis of a CSF sam
21. ollow these rules Specimens MUST be labelled with the following Surname Sender reference number PLUS any two out of three of the following Forename Full Date of Birth NHS Number AND Date of Collection of Specimen Request forms MUST match the information on the sample PLUS Address for the report requesting laboratory Patient address with postcode Consultant GP CSCDC Name of requestor Tests required Sender Reference Number Request forms SHOULD have Time and Date collected Sex Contact number for requestor Relevant clinical information Postcode If you have any problems queries contact Dr Steve Gray BMS3 MRU steve gray hpa org uk Tel 0161 2766757 Fax 0161 276 5744 Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 20 of 26 Information required The MRU request form MUST be used whenever specimens are submitted This can be downloaded from the HPA website at the following link http www hpa org uk ProductsServices I nfectiousDiseases LaboratoriesAndReferenceFacilities MeningococcalReferenceUnit Completion of the form ensures that the relevant investigations are carried out and reported back to sending laboratories with minimum delay If important information is missing sending laboratories may be contacted to supply details before testing is performed Please see the MRU specimen acceptance policy on the previous page It would be helpful if all requesting labor
22. ple Other specimens from normally sterile sites may be examined after prior consultation with the MRU and a blood and or CSF specimen should accompany them if available The nucleic acid extraction processes are designed for fluid samples so there will be limited experience for unusual sample types Positive results may be determined for such samples on the understanding that these should be considered unvalidated particularly with regards to negative results Enhanced surveillance of pneumococcal disease has included the successful screening of empyema fluid and other respiratory samples by pneumolysin PCR If tissue samples or blocks require examination they should only be submitted following specific consultation with MRU staff They are not currently considered a routine investigation as they require bespoke manual processing with concomitant increases in turnaround time Minimum volumes for PCR testing DNA extraction Blood or fluids the routine use of automated nucleic acid extraction systems requires a minimum 400 uL of blood but a larger volume is preferred in case repeat testing is required If smaller samples are submitted the fluid volume should be at least 100 uL Small volumes require separate extraction and this will increase turnaround times CSF 400 uL or more is preferred but small samples 50 uL can be tested The small volumes require specific extraction and will likely increase turnaround times Low volu
23. r up to one year after receipt should sufficient remain following initial processing Since 2006 post mortem samples or samples from the deceased patients known by the MRU to have died at the time of submission have been destroyed sensitively or returned if requested A minimal number of highly selected positive samples are retained for quality control assay development s or epidemiological investigation under the local HTA guidance Should it be necessary to contact the MRU regarding a HTA issue the Person Designated PD is Professor Ray Borrow tel 0161 276 6793 The recommendations of the Caldicott Report 1997 have been adopted by the Health Protection agency as by the National Health Service as a whole These recommendations relate to the security of patient identifying data PID and the uses to which they are put MRU as an integral part of Manchester Medical Microbiology Partnership observes Caldicott guidance in handling PID The MMMP has appointed its own Caldicott Guardian who advises on confidentiality issues and is responsible for monitoring the physical security of PID This also applies to the transfer of results of investigations to and from MMMP whether by mail services telephone or fax The value of safe haven arrangements or other means of the sender and receiver of information identifying themselves to each other before data are transferred is emphasized MMMP is anxious to audit the security of its PID in colla
24. s available Monday Friday 09 00 to 17 00 often 08 30 17 30 dependent on staffing arrangements If your call to the laboratory is not answered promptly please telephone 0161 276 6757 as staff may be unable to stop a procedure or could be working in one of several other areas Weekend enquiries For urgent clinical enquiries particularly those occurring out of hours weekends or on bank holidays please contact Dr Ed Kaczmarski on mobile contact available via the consultant medical microbiologist rota through Central Manchester Foundation Trust switchboard on 0161 276 1234 Out of hours specimens Specimens for PCR investigation must be received at the MRU by 10 00am weekdays to be tested the same working day Arrangements to accept couriered urgent samples for PCR or other investigations must be agreed with the MRU before the samples are sent Failure to do so may result in the specimen s not being tested in a timely fashion Urgent couriered specimens should be addressed to Microbiologist On call if out of hours Meningococcal Reference Unit URGENT SPECIMEN Manchester Medical Microbiology Partnership Clinical Sciences Building 2 Manchester Royal Infirmary Oxford Road Manchester M13 9WL If arriving after 5 30pm Monday Friday at weekends or on Bank Holidays they should be left at the Manchester Royal Infirmary Accident and Emergency Department in the On Call Virology box Edition no 05 MENI
25. s must comply with the sample acceptance policy and be accompanied by a completed MRU request form which can be downloaded from the HPA website Isolates for case confirmation epidemiology and cluster management 1 Please submit all available sterile site CSF blood and joint fluids isolates from cases 2 If available please submit throat and nose swab isolates from cases as well 3 Any isolates from case contacts nose or throat swabs should also be sent indicating which case they relate to A complete case sample set could include CSF blood joint fluid nose and throat isolates They are useful for molecular studies and validation of typing techniques Other non sterile sites 1 Invasive respiratory samples eg BALs samples obtained by surgical procedure 2 Respiratory soutum sample isolates if thought to be clinically significant 3 N meningitidis isolates with high MICs or unusual antibiograms Note that approximately 30 of N meningitidis isolates have penicillin MICs gt 0 06 mg L BSAC breakpoint and MICs up to 0 38 mg L are not unusual Isolates with penicillin MICs gt 0 5 mg L are worth investigating further GenitoUrinary Medicine GUM isolates 1 Please do not submit routine GUM isolates Only submit isolates from GUM patients if they appear resistant MICs of 2 0 25mg L or are epidemiologically linked to cases of invasive meningococcal infection Other Neisseria species The MRU is established to
26. teodeage WE 19 Specimen amp Sample Submission Guidelines s s sssrseeseeneeeeenn 20 Faxing and emailing reports containing patient s data 23 Compliance with the Human Tissue Act cccccceeeeeeeeeeeeeees 24 MRU Recognition of Caldicott Recommendations 24 Key References ATTE taal escesccceceencessssensessssssseeeeneenensenaes 25 Charity and Public Information Contact Details 0008 26 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 3 of 26 The HPA Meningococcal Reference Unit MRU for England and Wales has been situated in Manchester since 1978 Originally established to provide phenotypic characterisation of meningococci isolated in laboratories throughout the country the nature and scope of this activity has widened as has the range of tests available The MRU re located from Withington Hospital Manchester to Central Manchester Foundation Trust in March 2003 as an integral part of the Manchester Medical Microbiology Partnership MMMP The MRU is part of the Respiratory and Systemic Infections Department RSID and works closely with other parts of the HPA particularly the Immunisation Division and many other HPA colleagues in LARS to optimise meningococcal disease ascertainment through enhanced surveillance The MRU has been a world leader in developing and making nationally available tests for non culture case confirmation of meningococcal infection b
27. tient s name must be conveyed separately using a linking patient identifier The report must be sent to a safe haven fax machine This means that if the location is in general use consideration must be given to ensuring that unauthorised personnel are unable to read reports accidentally or otherwise Also the room housing the fax machine must be a secure location which is locked if it is likely to be unattended at the time the fax is sent Assurance must be sought from the intended recipient of the faxed report preferably in writing that the receiving fax machine is a safe haven Measures must be taken to minimise the risk of mis dialling either by double checking numbers or having frequently used numbers available on the fax machine s memory dial facility Confirmation must always be sought from the intended recipient that the fax is expected and has been received Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 23 of 26 The MMMP MRU adhere to the HTA and its application within the Central Manchester Foundation Trust site Tissue samples CSF whole blood EDTA blood etc from patients are submitted to the MRU with their consent obtained at time of sampling for disease confirmation epidemiological or public health investigations Samples are tested and retained in accordance with the MRU specimen retention policy Where original samples following nucleic acid extraction are kept frozen fo
28. ure survival in transport Please indicate on the request form if the material slope has not been incubated Short term storage of sloped cultures is optimal at 30 C if there are delays before submission Non viable cultures cultures which are no longer viable may still be considered for characterisation by molecular based methods after consultation with the MRU A heavy inoculum of the inert material on a slope may be submitted with an appropriate request form Additional tests Additional tests can be requested by telephone or letter on samples received by the laboratory up to 2 months after the receipt of the sample although it must be recognised that the archive sample available may have a limited volume Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 22 of 26 The following guidelines are prepared having taken into account the Code of Practice on reporting patients results by fax prepared by the DoH and Caldicott recommendations It is MMMP MRU policy that reports containing patients data wherever possible should not be sent by fax or e mail E mails cannot be relied on to guarantee security of patients data because they can be intercepted by a third party on route unless encryption is used In exceptional circumstances it may be necessary to send a result by fax but not by e mail In this case the following conditions must be adhered to after discussion with the laboratory The pa
29. y PCR Initially designed to identify the major disease causing serogroups A B C Y and W135 the test repertoire has been extended to provide more detailed additional characterisation utilising state of the art molecular techniques including DNA sequencing of genomic material from isolates and directly from clinical specimens where possible The optimised surveillance along with serological studies performed in the HPA Vaccine Evaluation Unit co located within the MMMP at MRI were key elements in supporting and monitoring the successful introduction of meningococcal serogroup C conjugate vaccine in the UK and have contributed significantly to establishing the international reputation of the MRU In addition to providing confirmatory laboratory services staff from the MRU advise on investigation and management of individual cases and outbreaks The MRU and Cfl have been active in the recent establishment of a network of national and regional reference laboratories which are collaborating to harmonise and optimise surveillance throughout Europe and sharing this experience with other interested groups in the Americas and Oceania This has resulted in the establishment of the European Meningococcal Disease Society EMGM Edition no 05 MENINGOCOCCAL REFERENCE UNIT Issue date April 2012 Page 4 of 26 General MRU Result enquiries Identification phenotypic characterisation serogroup serotyping subtyping molecular characterisat

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