Confirmatory assays are essential when using molecular testing for Neisseria gonorrhoeae in low-prevalence settings: insights from the third National Survey of Sexual Attitudes and Lifestyles (Natsal-3).

OBJECTIVES: To investigate the occurrence of unconfirmed positive gonorrhoea results when using molecular testing within a large population-based survey. DESIGN, SETTING AND PARTICIPANTS: Between 2010 and 2012, we did a probability sample survey of 15,162 men and women aged 16-74 years in Britain. Urine from participants aged 16-44 years reporting ≥1 lifetime sexual partner was tested for Neisseria gonorrhoeae and Chlamydia trachomatis using the Aptima Combo 2 (AC2) assay, with positive or equivocal results confirmed with molecular assays using different nucleic acid targets. RESULTS: A total of 4550 participants aged 16-44 years had urine test results (1885 men; 2665 women). For gonorrhoea, 18 samples initially tested positive and eight were equivocal. Only five out of 26 confirmed, giving a positive predictive value (PPV) for the initial testing of 19% (95% CI 4% to 34%). Most (86% (18/21)) participants with unconfirmed positive results for gonorrhoea reported zero or one sexual partner without condoms in the past year and none had chlamydia co-infection, whereas all five with confirmed gonorrhoea reported at least two recent sexual partners without condoms, and four had chlamydia co-infection. The weighted prevalence for gonorrhoea positivity fell from 0.4% (0.3% to 0.7%) after initial screening to <0.1% (0.0% to 0.1%) after confirmatory testing. By comparison, 103 samples tested positive or equivocal for chlamydia and 98 were confirmed (PPV=95% (91% to 99%)). CONCLUSIONS: We highlight the low PPV for gonorrhoea of an unconfirmed reactive test when deploying molecular testing in a low-prevalence population. Failure to undertake confirmatory testing in low-prevalence settings may lead to inappropriate diagnoses, unnecessary treatment and overestimation of population prevalence.

Global evaluation of lineage-specific human papillomavirus capsid antigenicity using antibodies elicited by natural infection.

Human Papillomavirus (HPV) type variants have been classified into lineages and sublineages based upon their whole genome sequence. Here we have examined the specificity of antibodies generated following natural infection with lineage variants of oncogenic types (HPV16, 18, 31, 33, 45, 52 and 58) by testing serum samples assembled from existing archives from women residing in Africa, The Americas, Asia or Europe against representative lineage-specific pseudoviruses for each genotype. We have subjected the resulting neutralizing antibody data to antigenic clustering methods and created relational antigenic profiles for each genotype to inform the delineation of lineage-specific serotypes. For most genotypes, there was evidence of differential recognition of lineage-specific antigens and in some cases of a sufficient magnitude to suggest that some lineages should be considered antigenically distinct within their respective genotypes. These data provide compelling evidence for a degree of lineage specificity within the humoral immune response following natural infection with oncogenic HPV.

Still no evidence of new variant Chlamydia trachomatis in England and Wales.

BACKGROUND: New variant Chlamydia trachomatis (nvCT) remains an important public health concern and in 2008 four cases of nvCT were reported in Scotland. The present study set out to determine whether nvCT was present in England and Wales. METHODS: 1054 clinical specimens, which had been confirmed as chlamydia positive (using an nvCT unaffected platform) at nine different diagnostic laboratories throughout England and Wales, were examined for the presence of the 377 bp nvCT deletion. RESULTS: 92% (968/1054) of specimens examined were confirmed as wild-type C. trachomatis. The remaining 86 specimens were found to be untypeable, which was probably due to low levels of DNA. No nvCT specimens were identified. CONCLUSIONS: There is currently no evidence that nvCT is present in England and Wales; however, laboratories using nvCT-affected platform should remain vigilant.

Evaluation of strategies for confirming Neisseria gonorrhoeae nucleic acid amplification tests.

The implementation of widespread unselected screening for Neisseria gonorrhoeae in England, using nucleic acid amplification tests (NAATs), has raised concerns regarding the potential increase in misdiagnoses. To increase the positive predictive value, confirmatory testing of positive specimens has been recommended; however, in practice this can be difficult to perform. This study examined the role of two different testing strategies for confirming the N. gonorrhoeae status of specimens that had been examined by the ProbeTec Strand Displacement Amplification (SDA) assay (Becton Dickinson). A total of 227 residual clinical specimens in SDA assay collection tubes were sent for confirmatory testing using two different testing approaches: (i) examination using two in-house real-time PCR assays (opa and porA pseudogene) and (ii) examination using the APTIMA Combo 2 (AC2) assay and the APTIMA Monospecific N. gonorrhoeae (AGC) tests (Gen-Probe). Of the 113 SDA-positive specimens (including low positives) examined, 93 % were confirmed as N. gonorrhoeae-positive using either one or both real-time PCR assays. In contrast, only 34 % were confirmed using the AC2 and/or the AGC assays. All 114 SDA-negative specimens were confirmed as negative using all four confirmatory tests. Clearly the AC2 and AGC assays cannot reliably be used to confirm residual specimens in SDA assay transportation buffers, due to the incompatibility of different platform chemistries. Although high rates of confirmation (93 %) can be achieved when examining residual SDA assay specimens using independent real-time PCR assays, establishing well-validated in-house real-time PCR assays for diagnostic use is a large undertaking for many primary laboratories and so such tests may be better confined to specialist laboratory services.