title: Real-time PCR detection of Neisseria gonorrhoeae susceptibility to penicillin.
abstract: The objective of this study was to develop a real-time PCR assay targeting the gonococcal porB gene (PorB-PCR) for predicting susceptibility of Neisseria gonorrhoeae to penicillin. This complements a previously described PCR assay for detecting penicillinase-producing N. gonorrhoeae (PPNG) developed by our laboratory (PPNG-PCR).
  
  
  
  The PorB-PCR assay was designed using six probes to characterize various combinations of amino acids at positions 101 and 102 of the PorB1b class protein, including the WT G101/A102 and mutant G101K/A102D, G101K/A102N and G101K/A102G sequences, as well as the PorB1a sequence. The ability of these sequences to predict penicillin susceptibility was initially assessed using 2307 N. gonorrhoeae isolates from throughout Australia for which phenotypic susceptibility data were available. The assay was then applied to N. gonorrhoeae-positive clinical specimens (n = 70). Specificity was assessed by testing commensal Neisseria strains (n = 75) and N. gonorrhoeae-negative clinical specimens (n = 171).
  
  
  
  Testing of the 2307 N. gonorrhoeae isolates using PorB-PCR to detect G101/A102 and PorB1a sequences identified a total of 78.4% (61.2% and 17.2%, respectively) of penicillin-susceptible isolates with specificities of 97.4% and 99.3% and positive predictive values of 98.8% and 98.9%, where PPNG strains were simultaneously identified and excluded. Similar performance data were obtained when the PorB-PCR assay was applied to the N. gonorrhoeae-positive clinical specimens. No false-positive results were observed for the N. gonorrhoeae-negative samples and no cross-reactions were observed with the non-gonococcal species.
  
  
  
  When used in parallel with the previously described PPNG-PCR, the PorB-PCR approach has the potential to facilitate individualized treatment of gonorrhoea using penicillin.
authors: The University of Queensland, UQ Centre for Clinical Research (UQCCR), Herston, Queensland 4029, Australia.
authors: Cameron
authors: C
authors: Buckley
authors: The University of Queensland, UQ Centre for Clinical Research (UQCCR), Herston, Queensland 4029, Australia e.trembizki@uq.edu.au.
authors: Ella
authors: E
authors: Trembizki
authors: Kirby Institute, UNSW Australia, Sydney, New South Wales 2052, Australia.
authors: Basil
authors: B
authors: Donovan
authors: Melbourne Sexual Health Centre, Alfred Health, Carlton, Victoria 3053, Australia.|Central Clinical School, Monash University, Melbourne, Victoria 3181, Australia.
authors: Marcus
authors: M
authors: Chen
authors: Microbiology Laboratory, Pathology Department, Royal Darwin Hospital, Darwin, Northern Territory, Australia.
authors: Kevin
authors: K
authors: Freeman
authors: Kirby Institute, UNSW Australia, Sydney, New South Wales 2052, Australia.
authors: Rebecca
authors: R
authors: Guy
authors: WHO Collaborating Centre for STD, Microbiology Department, South Eastern Area Laboratory Services, Prince of Wales Hospital, Sydney, New South Wales 2031, Australia.
authors: Monica M
authors: MM
authors: Lahra
authors: WHO Collaborating Centre for STD, Microbiology Department, South Eastern Area Laboratory Services, Prince of Wales Hospital, Sydney, New South Wales 2031, Australia.
authors: Ratan L
authors: RL
authors: Kundu
authors: Kirby Institute, UNSW Australia, Sydney, New South Wales 2052, Australia.
authors: David G
authors: DG
authors: Regan
authors: Public Health Microbiology, Communicable Disease, Queensland Health Forensic and Scientific Services, Archerfield, Queensland, Australia.
authors: Helen V
authors: HV
authors: Smith
authors: The University of Queensland, UQ Centre for Clinical Research (UQCCR), Herston, Queensland 4029, Australia.|Pathology Queensland, Microbiology Department, Herston, Queensland, Australia.
authors: David M
authors: DM
authors: Whiley
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001315:Australia; D060005:Genotyping Techniques / Q000379:methods; D006801:Humans; D008826:Microbial Sensitivity Tests / Q000379:methods*; D009344:Neisseria gonorrhoeae / Q000187:drug effects* / Q000235:genetics*; D015345:Oligonucleotide Probes / Q000235:genetics; D010406:Penicillins / Q000494:pharmacology*; D018272:Porins / Q000235:genetics; D011237:Predictive Value of Tests; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D000900:Anti-Bacterial Agents; D015345:Oligonucleotide Probes; D010406:Penicillins; D018272:Porins; C057699:porin protein, Neisseria