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WHO EQAP for the detection of influenza A virus subtype by PCR

WHO EQAP for the detection of influenza A virus subtype by PCR. Wilina Lim Centre for Health Protection Hong Kong SAR, China. Aims/ o bjectives. T o monitor quality and standards of performance To facilitate information exchange To identify problems with assays

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WHO EQAP for the detection of influenza A virus subtype by PCR

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  1. WHO EQAP for the detection of influenza A virus subtype by PCR Wilina Lim Centre for Health Protection Hong Kong SAR, China

  2. Aims/objectives • To monitor quality and standards of performance • To facilitate information exchange • To identify problems with assays • To help develop testing strategies • To provide mechanisms to remedy any deficiencies revealed • Requirement of lab accreditation in accordance with international standard such as ISO 15189

  3. Benefits identified by NICs To monitor • feasibility to ship samples to countries • laboratory capability • timeliness of reporting

  4. QAP Process 1A.Invitation 1B.NewParticipants 2.Preparation of Panels 3.Panel Distribution 4.Data Collection 5.Preliminary report The EQAP has been accredited in accordance With ISO 17043 6.Data Analysis 7.Final report

  5. Preparation of panels • Include different subtypes/clades Dried RNA of influenza AH5, H1, H3, H1v and influenza B viruses Gamma-ray inactivated seasonal influenza samples • Verify sample content • Verify sufficient homogeneity Samples in final packaged form selected and tested • Assure sufficient stability Test over a range of storage conditions prior to distribution Samples were tested after 7 days of storage at 37oC using both conventional and real-time PCR assays

  6. Temperature survey • Monitor the temperature change during shipment • Target participants • With temperature record higher than 37℃ during sample dispatch period • Panel 9: Jan-Mar 2011

  7. Temperature survey on EQAP panel 9 shipment Longest duration with temperature higher than 37℃ was 6 hours

  8. Temperature above 37℃ were recorded during the transit in daytime

  9. Composition of panels

  10. Response of participants

  11. Problems encountered • No PCR capacity • No reagents • Delay in obtaining import permit • Varying requirements at the customs • Shipment detained at customs for prolonged period

  12. Reasons for laboratories not receiving panels among total invited

  13. Performance of participants

  14. Performance of SEAR participants % of all correct

  15. Performance of WPR participants % of all correct

  16. Performance in Panel 8

  17. Performance in Panel 9

  18. Problems identified in EQAP • Inconsistent technical performance • Positive control not used appropriately • Lab contamination • Misinterpretation of results • Primers and probes mismatch • Transcriptional error

  19. False negative results due to probe mis-matches • An example of a H5 real-time PCR primer/probe set • Forward primer: 1 bp mis-match • Probe: 2 bp mis-matches • Reverse primer: none

  20. GLPsurvey • 2007 survey composed of 73 questions • 2008 survey composed of 25 questions • 2010 survey composed of 32 questions • Questions on the following seven categories: • personnel • quality managements • design, equipment and consumables • pre-analytical procedures • analytical procedures • post-analytical procedures • reporting and record keeping • safety

  21. Molecular diagnosis (PCR) & Good laboratory practice (GLP) Laboratories returning completed GLP survey forms 2007 64 ( 96%) 2008 94(82%) 2010 142 (89%)

  22. Good laboratory practices More than 80% of laboratories Less than 80% of laboratories Separate work room for molecular diagnosis (99%) Separate set of equipment and consumables in each working area (94%) Equipment maintenance programme (87%) Control materials for molecular diagnosis (100%) Standard operating procedures (94%) Evaluation of the reagents used for molecular tests (67%) Evaluation of the sensitivity/specificity of the molecular tests (59%) Internal audit programme (54%) Accredited by international/national scheme (34%) Countercheck results (78%) Data from GLP survey 2010

  23. GLP and EQAP performance • Compare GLP with EQAP results • Group A (laboratories returned correct answers for all 10 samples) • Group B (laboratories returned less than 10 corrects answers) • Laboratories with less good performance (Group B) Lab did not return all correct results tends to meet less quality parameters; significantly more likely (p < 0.05) not having - audits of personnel - separate rooms for all steps involving PCR - programme to monitor equipment - more samples in recent representative month - SOP on preparation of in-house controls - established test turn-around-time

  24. Reagents/tests evaluation, internal audit and accreditation in laboratories of different WHO regions Data from GLP survey 2010

  25. The way forward • Review - materials for simulated specimens/scope - type/subtype/clade to include in the panel - frequency of shipment • Enhance performance through training • Accreditation by recognized authority

  26. http://www.wpro.who.int/sites/htl/documents/Laboratory+Quality+Standards+and+Implementation.htmhttp://www.wpro.who.int/sites/htl/documents/Laboratory+Quality+Standards+and+Implementation.htm

  27. Thank You

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