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Neisseria gonorrhoeae Multi-Antigen Sequence Typing using non-cultured clinical specimens
  1. David M Whiley1,*,
  2. Namraj Goire1,
  3. E. Sanghamitra Ray2,
  4. Athena Limnios2,
  5. Stephen Lambert1,
  6. MIchael Nissen1,
  7. Theo Sloots1,
  8. John Tapsall2
  1. 1 Queensland Paediatric Infectious Diseases Laboratory, Sir Albert Sakzewski Virus Research Centre, Australia;
  2. 2 WHO Collaborating Centre for STD and HIV, Microbiology Department, South Eastern Area Laboratory Ser, Australia
  1. Correspondence to: David M Whiley, Royal, Children's Hospital, Clinical Virology Research Unit, SASVRC, Clinical Virology Research Unit, SASVRC, Royal, Children's Hospital, Hertson Road, Herston, Brisbane, 4029, Australia; d.whiley{at}uq.edu.au

Abstract

Objectives: The N. 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. We sought to examine the performance on the NG-MAST system on a range of non-cultured samples.

Methods: Nucleic acid extracts of 73 N. gonorrhoeae positive samples, comprising 8 cervical swabs, 9 urethral swabs, 35 urine samples, 1 vaginal swab, 13 rectal swabs and 7 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 further investigate 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 uro-genital specimens, 11 of 13 rectal swabs and 4 of 7 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 problematic for NG-MAST typing of throat swabs specimens.

Conclusions: The NG-MAST system can successfully be applied directly to non-cultured uro-genital samples, however, is less suitable for extra-genital specimens, particularly throat swabs, due to cross-reaction with commensal Neisseria species.

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