Evaluation of PorB variable region typing of Neisseria gonorrhoeae using PCR‐ELISA in samples collected from men who have sex with men

Abstract

A polymerase chain reaction (PCR)-based enzyme-linked immunosorbent assay (ELISA) methodology was developed to characterize Neisseria gonorrhoeae porB gene variable regions (VR); the methodology was evaluated in comparison to porB VR typing by checkerboard hybridization. Clinical noncultured samples from 35 men who have sex with men (MSM), positive by nucleic amplification assays for N. gonorrhoeae, were typed using a panel of 40 oligonucleotide probes to porB VRs and compared to checkerboard hybridization. Complete concordance was observed between the two methods at PIB VRs 1, 3, and 7. At the more degenerate VRs 5 and 6, PCR ELISA resulted in obtaining more typeable VRs than checkerboard hybridization due to single nucleotide mismatches. By PCR ELISA, two predominant PIB porB types were identified in 58% of the samples and the remaining 16 samples had one of six other porB types. Both PCR ELISA and checkerboard hybridization methods of porB VR typing allowed characterization of N. gonorrhoeae from noncultured clinical samples including throat and rectal swabs and discriminated N. gonorrhoeae from N. meningitidis present in some of the samples. PCR ELISA is a rapid, relatively inexpensive and alternative molecular typing method for N. gonorrhoeae, suitable for use in conjunction with molecular diagnostic tests.

Quantitative Proteomics of the 2016 WHO Neisseria gonorrhoeae Reference Strains Surveys Vaccine Candidates and Antimicrobial Resistance Determinants

“…Expression and Homologs of Gonorrhea Vaccine Candidates-We first evaluated expression profiles of extensively studied gonorrhea vaccine candidates including MtrE (91-93), PorB (94,95), PilQ (96), TbpA (97,98), Opa (99,100), and AniA (19,(101)(102)(103). MtrE was upregulated in 8 isolates (Table II).…”

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

“…The sensitivity (i.e., ability to detect small quantities of DNA) of DNA sequencebased methods has the potential to make them useful for direct strain typing of samples obtained noninvasively, such as urine. The disadvantages of DNA sequence-based methods include their relative expense in low-resource settings, their nonoptimization or validation for use on specimens obtained noninvasively and samples with nonviable N. gonorrhoeae (85,191), and the lack of phenotypic information produced, such as information on immunologically important epitopes which might be valuable for future vaccine development. In a population with a high level of sexual mixing, mixed gonococcal infections, in which an individual is concurrently colonized with more than one strain of N. gonorrhoeae, have been identified through the use of highly discriminatory genotypic techniques (88,94,178,179).…”

A SERS‐active enzymatic product used for the quantification of disease‐related molecules

“…Therefore, the ELISA method typically involves a chromophore reporter and substrates that produce an observable color that indicates the presence of the target. More recently developed ELISA‐like techniques use fluorogenic, electrochemoluminescent, and real‐time polymerase chain reaction (PCR) reporters to produce quantifiable signals . The choice of substrate depends upon the required assay sensitivity and the instrumentation available for signal detection (spectrophotometer, fluorometer, or luminometer) .…”