FEDERAL PUBLIC SERVICE, HEALTH, FOOD CHAIN SECURITY AND ENVIRONMENT CLINICAL BIOLOGY COMMISSION CLINICAL BIOLOGY SECTION

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1 IPH J. Wytsmanstreet 14 B-1050 Brussels FEDERAL PUBLIC SERVICE, HEALTH, FOOD CHAIN SECURITY AND ENVIRONMENT CLINICAL BIOLOGY COMMISSION CLINICAL BIOLOGY SECTION External Quality Assessment for Molecular Microbiology Survey 2008/04 Chlamydia trachomatis detection in urine and swabs panel B This report is available on our webpage: This report may not be reproduced, published or distributed without permission of IPH

2 QUALITY CONTROL for MOLECULAR DIAGNOSTICS Block 4, Kelvin Campus, West of Scotland Science Park, Glasgow, G20 0SP Scotland Tel: +44 (0) Fax: +44 (0) BEIPH Final Report QCMD 2008 Chlamydia trachomatis (CTDNA08B) EQA Programme William G MacKay on behalf of QCMD and its Scientific Advisory Board October 2008 Not to be reproduced or quoted without permission of QCMD. Any queries about this report should be addressed to the QCMD Neutral Office. The QCMD programme is organised in collaboration with the European Society for Clinical Virology and the European Society for Clinical Microbiology & Infectious Diseases. Registered in Scotland Reg No: SC Registered Office: 39 Castle Street, Edinburgh EH2 3BH

3 1. Programme aims The primary aim of this External Quality Assessment Programme was to determine the qualitative and quantitative performance of participants testing for Chlamydia trachomatis using molecular technologies. The secondary aims of the programme were: 1) To determine the incidence of false positive results and to compare performance with previous distributions. 2) To determine the performance of molecular diagnostics for C. trachomatis in samples with a high level of N. gonorrhoeae. 3) To determine the proficiency of laboratories in the correct detection of the Swedish variant of C. trachomatis. 2. Programme details Table 1: Programme Details CTDNA08B Date of panel distribution 17/06/2008 Number of participants 52 Number of countries 1 Number of respondents 48 (92%) Number of datasets submitted 50 Number of qualitative datasets submitted 50 (100%) Number of qualitative and quantitative datasets submitted 0 (0%) Four participants did not return results. Two of these withdrew officially citing 'assay under development' (n=1) and 'internal issues' (n=1). 3. Panel composition This EQA panel for the detection of Chlamydia trachomatis consisted of ten samples containing various concentrations of C. trachomatis and two samples negative for C. trachomatis in either urine or simulated swab. C. trachomatis (LGV L2) was cultured from a clinical isolate. Table 2: Panel composition Sample Sample Sample * Sample conc. Sample content matrix Copies/vial status CTB08-01 Chlamydia trachomatis Urine 570 Strong positive CTB08-02 Chlamydia trachomatis Urine 5700 Strong positive CTB08-03 Chlamydia trachomatis Urine 57 Weak Positive CTB08-04 C. t Negative Urine Urine Negative CTB08-05 Chlamydia trachomatis Urine 57 Weak Positive CTB08-06 Chlamydia trachomatis Urine 570 Positive CTB08-07 Chlamydia trachomatis ß Urine 2.27x10-2 Diln Weak Positive CTB08-08 Chlamydia trachomatis Swab 57 Positive CTB08-09 Chlamydia trachomatis Swab 5700 Strong positive CTB08-10 C. t Negative 'Swab' Swab Negative * Urine: Chlamydia trachomatis -negative urine collected from healthy adult volunteers. Swab: 1/10 dilution of TE buffer. Diln: Dilution from a stock. ''Sample concentration' and/or 'Target values' are for QCMD internal purposes only. They should not be used by participants for method comparison or as a target for individual laboratory assessment. A sample status was assigned to each panel sample and consisted of 'Strong positive', 'Positive', 'Weak positive' and 'Negative'. C. trachomatis positive sample containing Neisseria gonorrhoeae at a concentration of CFU/ml. ß Swedish variant, missing 377 base pairs of the cryptic plasmid (Rippa & Nilsson 2007). 2

4 4. Programme results 4a. Qualitative analysis of the EQA data The number (percentage) of correct qualitative results are presented in Table 3. Qualitative data were returned by participants as 'positive', 'negative' or 'not determined'. Not detemined results were counted as incorrect for all panel samples (positive or negative). QCMD organises datasets according to commercial and in-house technology groups, which are Conventional PCR, Real time PCR, NASBA, SDA, TMA and bdna. Where datasets were reported as other for a technology or kit method this was reviewed by the QCMD Neutral Office and assigned to an appropriate group where possible. Table 3: Number of correct qualitative results per panel member and technology type PCR SDA f TMA g Other h Sample Sample Sample conc. Total Conventional Real time content Copies/vial datasets Comm. a Comm. c In-house d n=50 n=21 n=14 n=3 n=10 n=1 n=1 n % n % n % n % n % n % n % CTB08-02 Chlamydia trachomatis CTB08-01 Chlamydia trachomatis CTB08-06 Chlamydia trachomatis CTB08-03 Chlamydia trachomatis CTB08-05 Chlamydia trachomatis CTB08-07 Chlamydia trachomatis 2.27x10-2 Diln CTB08-04 C. t Negative Urine CTB08-09 Chlamydia trachomatis CTB08-08 Chlamydia trachomatis CTB08-10 C. t Negative 'Swab' C. trachomatis positive sample containing Neisseria gonorrhoeae at a concentration of CFU/ml. Swedish variant, missing 377 base pairs of the cryptic plasmid. a: Roche Amplicor CT/NG (n=10), Roche COBAS Amplicor CT/NG (n=11). c: Abbott Real time CT (n=2), Abbott Real time CT/NG (n=1), Nanogen Chlamydia tr. Q-PCR Alert Kit (n=1), QIAGEN artus C. trachomatis PCR Kit (TM) (n=2), QIAGEN artus C. trachomatis Plus PCR Kit (LC) (n=1), QIAGEN artus C. trachomatis Plus PCR Kit (RG) (n=2), Roche Cobas TaqMan CT (n=4), Shangai Bio-Tech IMTec CT Real time PCR Kit (n=1). d: In-house details are not presented. f: Becton, Dickinson ProbeTec ET (n=10). g: Gen-Probe APTIMA Combo 2 Assay (n=1). h: Gen-Probe PACE 2 CT (n=1). QCMD uses a colour-coded scheme for scoring based on the classification of results in relation to expected or consensus results. A more detailed explanation of the scoring system is provided in the appendix to this report and on the QCMD website ( 3

5 4b. Qualitative performance scores Table 4: Qualitative performance scores per technology type Sample Sample Status Total All technologies n=50 PCR CTB08-02 Strong positive CTB08-01 Strong positive CTB08-06 Positive CTB08-03 Weak Positive CTB08-05 Weak Positive CTB08-07 Weak Positive CTB08-04 Negative CTB08-09 Strong positive CTB08-08 Positive CTB08-10 Negative SDA f TMA g Conventional Real time Commercial a n=21 Commercial c n=14 In-house d n=3 n=10 n=1 Other h n=1 a: Roche Amplicor CT/NG (n=10), Roche COBAS Amplicor CT/NG (n=11). c: Abbott Real time CT (n=2), Abbott Real time CT/NG (n=1), Nanogen Chlamydia tr. Q-PCR Alert Kit (n=1), QIAGEN artus C. trachomatis PCR Kit (TM) (n=2), QIAGEN artus C. trachomatis Plus PCR Kit (LC) (n=1), QIAGEN artus C. trachomatis Plus PCR Kit (RG) (n=2), Roche Cobas TaqMan CT (n=4), Shangai Bio-Tech IMTec CT Real time PCR Kit (n=1). d: In-house details are not presented. f: Becton, Dickinson ProbeTec ET (n=10). g: Gen-Probe APTIMA Combo 2 Assay (n=1). h: Gen-Probe PACE 2 CT (n=1). Figure 1: Percentage of qualitative performance scores per technology type % a c d f g a c d f g a c d f g a c d f g a c d f g a c d f g a c d f g a c d f g a c d f g a c d f g CTB08-02 CTB08-01 CTB08-06 CTB08-03 CTB08-05 CTB08-07 CTB08-04 CTB08-09 CTB08-08 CTB08-10 Technology group per panel sample a: Conventional commercial PCR, c: Real time commercial PCR, d: Real time in-house PCR, f: SDA, g: TMA. 4

6 References Ripa T, Nilsson PA. A Chlamydia trachomatis strain with a 377-bp deletion in the cryptic plasmid causing false-negative nucleic acid amplification tests. Sexually Transmitted Diseases. 2007; 34(5): Acknowledgements QCMD wishes to acknowledge the support of the Laboratory for Infectious Diseases (Groningen) for production of the bulk materials and performing release testing. QCMD also wishes to acknowledge the support of Molecular Diagnostics, Health Protection Agency (Manchester Royal Infirmary, UK) and North Glasgow Hospitals NHS Trust, West of Scotland Specialist Virology Centre (Gartnavel General Hospital, Glasgow, UK) for performing independent testing. QCMD also wishes to thank Dr. Kenneth Persson (from the Department of Clinical Microbiology, Malmö University Hospital, Malmö, Sweden) for providing the C. trachomatis Swedish variant strain. QCMD The data and report documents provided are intended for the sole use of the participant. It is based on material in our possession or supplied to us, which we believe to be reliable. Whilst every effort has been made to ensure its accuracy, we cannot offer any warranty that factual errors have not occurred. We therefore take no responsibility for any damage or loss that may be suffered by reason of any such inaccuracies. 5

7 Appendix A Scoring system for qualitative EQA data The scores awarded for qualitative EQA data were based on the sample status (see Section 3). The scoring system is represented in the following table, where 0 is 'highly satisfactory' and 3 is 'highly unsatisfactory'. Colour has been included as an extra visual aid. Scoring system based on the assigned sample status Sample status Participant's result Negative Not determined Positive Strong Positive Positive Weak Positive Negative