Detection of Neisseria gonorrhoeae from air-dried genital samples by single-tube nested PCR

Abstract

A single-tube nested PCR method was developed for the detection of Neisseria gonorrhoeae. The optimized assay had a detection limit of less than 0.3 cell. Five different storage conditions for gonococcal specimens were compared with respect to the PCR detection of bacteria. For air-dried gonococcal slides containing three bacteria, DNA was detected after 8 weeks at ambient temperature, and for slides containing 300 bacteria, DNA could be detected after 24 weeks at ambient temperature. Air-dried storage combined with analysis by the single-tube nested PCR and a commercially available PCR (Amplicor) was used to test 350 cervical specimens from women in the West African island nation of Cape Verde. The in-house PCR detected 17 cases of N. gonorrhoeae infection, while the Amplicor system detected 14 cases of N. gonorrhoeae infection. No specimen was negative by the in-house PCR assay and positive by the Amplicor PCR. This sensitive nested PCR assay, combined with air-dried storage, allows for the detection of gonococci when specimen storage and transport times are extended and freezing conditions are not available.