Identification of genetic targets specific to Neisseria gonorrhoeae for use in molecular detection methods has been a challenge. The porA pseudogene in N gonorrhoeae has been commonly used but recently gonococcal isolates giving a negative result in these PCRs have been reported. Here we describe the characterisation of two such gonococcal isolates received by the reference service at the Health Protection Agency, London, England.
Phenotypic characterisation was achieved using conventional biochemical and immunological tests, matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF-MS), antimicrobial susceptibility testing, serovar determination and detection of meningococcal PorA using monoclonal antibody 4BG4-E7. Genetic species confirmation was determined using commercial and in house PCRs and 16S rRNA gene sequencing. Molecular typing using the N gonorrhoeae multi-antigen sequence typing (NG-MAST) and multilocus sequence typing (MLST) was performed. The DNA sequence of the full-length gonococcal porA pseudogene was determined and compared with published sequences.
Both isolates were confirmed, biochemically and immunologically as N gonorrhoeae, but repeatedly gave negative results with two in house real-time PCR assays for the porA pseudogene. Further characterisation of these isolates identified the presence of a meningococcal porA sequence and showed these isolates belong to serovar Bropyst, and to NG-MAST sequence type (ST) 5967 and MLST ST1901.
Gonococcal isolates that give false negative results with porA pseudogene PCR assays have now been identified in four countries, three of which are in Europe, and do not appear clonal. This report highlights the genetic diversity of N gonorrhoeae, which remains a challenge for the molecular detection methods.
In many settings, human papillomavirus (HPV) DNA testing already plays an important role in cervical cancer screening. It is unclear whether hormonal fluctuations associated with menstrual phase or oral contraceptive (OC) use have any effect on HPV detection. We evaluated the effects of OC use and timing of cervical sampling in relation to women's last menstrual period (LMP) on HPV detection, and viral load in the Brazilian Ludwig–McGill cohort study.
Women in the cohort were followed every 4–6 months, and at each clinic visit they were asked to complete a questionnaire and to provide a cervical sample for HPV testing. Specimens from 6093 patient visits (n=2209 women) were categorised according to date of LMP into four distinct phases: follicular (days 5–9), midcycle (days 10–15), luteal (days 16–22), or late luteal (days 23–31).
Compared with follicular phase (referent group), HPV detection did not differ according to reported LMP for midcycle (OR=1.14, 95% CI 0.95 to 1.37), luteal (OR=1.03, 95% CI 0.85 to 1.25), or late luteal menstrual phase (OR=1.01, 95% CI 0.83 to 1.24), and was also not influenced by OC use. Analyses restricted to high-risk HPV types (grouped) and HPVs 16 and 18 (separately), produced similar non-significant associations. For HPV-positive samples, we found that the menstrual phase did not influence the total viral load.
These results indicate HPV detection is not associated with menstrual phase. Our findings suggest that standardising the timing of specimen collection for HPV testing is not necessary.