Global transmission of prolyliminopeptidase-negative Neisseria gonorrhoeae strains: implications for changes in diagnostic strategies

Abstract

Background: Species confirmation of Neisseria gonorrhoeae is commonly performed with biochemical kits, rely on the activity of the enzyme prolyliminopeptidase (PIP). This enzyme has previously been considered to be almost universally present in N gonorrhoeae. However, increasing numbers of N gonorrhoeae isolates lacking PIP activity have been identified. Objectives: To investigate the possibility of a widespread transmission of one or several N gonorrhoeae PIPnegative strains among several countries worldwide. Methods: PIP-negative N gonorrhoeae isolates cultured from 2001 to 2004 in Australia, New Zealand and Scotland were comprehensively characterised and compared with previous data from England and Denmark. All isolates were characterised by antibiotic susceptibility testing, serovar determination, pulsed-field gel electrophoresis (PFGE), opa-typing, sequencing of the entire porB gene and N gonorrhoeae multiantigen sequence typing (NG-MAST). Results: Most (83%) of the viable Australian isolates, and all the New Zealand and Scottish isolates were assigned serovar IB-4, with similar antibiograms, nearly identical porB1b gene sequences, identical (ST210) or highly related (ST292, ST1259) NG-MAST STs, and indistinguishable or related PFGE fingerprints as well as opa-types. The isolates showed characteristics indistinguishable or highly related to the previously described English and Danish outbreak strain. Conclusions: A comprehensive characterisation indicates a widespread dissemination, mainly among men who have sex with men (MSM), of indistinguishable and highly related genotypes that have evolved from a single N gonorrhoeae PIP-negative serovar IB-4 strain among several countries worldwide. An increased awareness of PIP-negative N gonorrhoeae strains is crucial and changes in the diagnostic strategies may need to be considered.

Review and International Recommendation of Methods for Typing Neisseria gonorrhoeae Isolates and Their Implications for Improved Knowledge of Gonococcal Epidemiology, Treatment, and Biology

“…Opa typing is based on the PCR amplification of 11 opa genes followed by restriction endonuclease digestion with TaqI and HhaI, separation of fragments by gel electrophoresis, and subsequent visualization of the banding patterns (118). The method imparts high typeability and reproducibility and excellent discriminatory ability, even in comparison with highly discriminatory DNA sequence-based approaches (19,20,26,49,65,71,74,93,96,104,118,121,127,174,182,186,189). However, Opa typing has the same disadvantages as RFLP methods, including labor-intensiveness, subjectiveness, and a need for pronounced standardization for interlaboratory comparisons.…”

“…Therefore, PFGE is particularly a useful method to increase discrimination between isolates in specific situations, especially those involving extreme microepidemiology (49,54,82,135,153,172,174,178). PFGE is reproducible, and all N. gonorrhoeae isolates are typeable by this method (24,54,71,80,87,105,111,113,132,133,135,153,154,156,170,172,174,178,179,182,196,197). The distinct disadvantages of RFLP and PFGE analysis include the requirement for a high level of technical and interpretive expertise, the potentially subjective interpretation of banding patterns on gels, the time involved for typing (several days), the lack of high-throughput analysis, and high cost.…”

“…Digestion of genomic DNA with enzymes such as SpeI and/or BglII has proven to be highly discriminatory for N. gonorrhoeae. PFGE has been used to define the gonococcal strain populations in a particular region, to identify clusters of circulating strains, including antibiotic resistant strains (3,24,49,51,54,71,80,87,105,111,113,132,133,135,153,154,156,170,172,174,179,182,196,197), and in forensic evaluations (30; M. Unemo, unpublished data). This method can distinguish subtypes within A/S classes and within genotypes determined with other highly discriminatory DNA sequence-based methods, such as full-or extended-length porB sequencing and NG-MAST.…”

Direct Bacterial Profiling by Matrix-Assisted Laser Desorption−Ionization Time-of-Flight Mass Spectrometry for Identification of Pathogenic Neisseria

“…3 As taxonomic differences between members of the Neisseria genus are small, the identification of these pathogens remains problematic and as such, no single method is currently recommended for a definitive identification. Prolyliminopeptidase was previously considered to be almost universally present in N. gonorrhoeae, but the widespread occurrence of Prolyliminopeptidase-negative N. gonorrhoeae strains 4 potentially results in incorrect, doubtful, or delayed species identification. Additionally, N. meningitidis strains producing Prolyliminopeptidase like N. gonorrhoeae have been reported.…”

Molecular Epidemiological Identification of Neisseria gonorrhoeae Clonal Clusters with Distinct Susceptibility Profiles Associated with Specific Groups at High Risk of Contracting Human Immunodeficiency Virus and Syphilis

“…Molecular typing methods such as pulsed-field gel electrophoresis-based methods, antimicrobial susceptibility patterns, porB gene sequencing, and opa typing have been used to assess the clonal relatedness of N. gonorrhoeae strains (9,15,16). Recently, N. gonorrhoeae multiantigen sequence typing (NG-MAST) of two genes (por and tbpB) has been developed to assess the genetic diversity of N. gonorrhoeae (4).…”