Neisseria gonorrhoeae multi-antigen sequence typing using non-cultured clinical specimens

Abstract

Objectives: The Neisseria gonorrhoeae multi-antigen sequence typing (NG-MAST) system, based on PCR amplification and sequence analysis of the gonococcal porB and tbpB genes, is widely used for molecular typing of gonococcal isolates but is not validated for non-cultured clinical samples. This study sought to examine the performance of the NG-MAST system on a range of non-cultured samples. Methods: Nucleic acid extracts of 73 N gonorrhoeae-positive samples, comprising eight cervical swabs, nine urethral swabs, 35 urine samples, one vaginal swab, 13 rectal swabs and seven throat swabs, were analysed by NG-MAST. For 27 specimens, corresponding gonococcal isolates were also analysed and the results compared. A panel of 44 non-gonococcal Neisseria strains and 100 N gonorrhoeae-negative clinical samples were used to investigate further the specificity of the NG-MAST PCR reactions. Results: PCR amplification and DNA sequencing of gonococcal porB and tbpB genes was successful for all N gonorrhoeae-positive urogenital specimens, 11 of 13 rectal swabs and four of seven throat swabs. For the 27 N gonorrhoeae-positive specimens with corresponding gonococcal isolates, the porB and tbpB sequences obtained from the non-cultured specimen were identical to those obtained from the isolate. Cross-reaction with non-gonococcal Neisseria species was observed for both the porB and tbpB PCR reactions, and proved to be problematical for NG-MAST typing of throat swab specimens. Conclusions: The NG-MAST system can successfully be applied directly to non-cultured urogenital samples, but is less suitable for extragenital specimens, particularly throat swabs, due to cross-reaction with commensal Neisseria species. Show more