por Variable-Region Typing by DNA Probe Hybridization Is Broadly Applicable to Epidemiologic Studies of Neisseria gonorrhoeae

Abstract

The porin gene (porB) of Neisseria gonorrhoeae encodes the major outer membrane protein identified as PI or Por. To examine the utility of por variable-region (VR) typing, porB from 206 isolates was characterized by using oligonucleotide probes in a checkerboard hybridization assay that identifies the sequence types of five VRs of both PIA and PIB porB alleles. The strains represented temporally and geographically distinct isolates, isolates from a large cluster, epidemiologically linked partner isolates, and a collection of strains from disseminated gonococcal infections. By using rigorous epidemiologic criteria for transmission of infection between sex partners, por VR typing was more discriminatory than serovar typing in classifying isolates from both members of 43 epidemiologically linked pairs: 39 of 43 pairs were classified as coinciding by por VR typing compared to 43 of 43 by serovar determination (P ‫؍‬ 0.058). porB sequence data confirmed the accuracy of the por VR method. Relationships between VR type and serovar typing monoclonal antibodies were observed for all six PIB and three of six PIA antibodies. por VR typing is a molecular tool that appears to have broad applicability. This method can be adapted to a wide range of technologies from simple hybridization to microarray and may allow for typing from noncultured clinical specimens.

Comparison of Neisseria gonorrhoeae Multiantigen Sequence Typing and porB Sequence Analysis for Identification of Clusters of N. gonorrhoeae Isolates

“…Among isolates with identified epidemiological links, both methods were congruent with epidemiological findings. These results validate the use of porB DNA sequence analysis for epidemiological studies, in agreement with data from previous studies (1,5,8,11,13,15,(18)(19)(20)(21)23). NG-MAST further differentiated porB types due to the sequence variations present in tbpB alleles (6), which contributed to a different NG-MAST ST.…”

“…DNA sequence analysis of various genes is currently the method of choice for distinguishing N. gonorrhoeae strains, as it provides unambiguous and reproducible information and high discriminatory power, and data can be stored or shared electronically, permitting reliable comparisons to be made between laboratories (10,11,23). porB DNA sequence analysis is now commonly used for studying the molecular epidemiology of N. gonorrhoeae (1,9,13,15,16,19,20,23) and involves the sequencing of either the entire gene or various regions of porB (8,(17)(18)(19). The N. gonorrhoeae multiantigen sequence typing (NG-MAST) methodology, which has been applied since 2004 (2,9,13,20,21), is based on limited DNA sequence analyses of two highly polymorphic loci, porB and tbpB (11).…”

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

“…For each VR probe tested, positive and negative bacterial control strains were chosen based on the presence or absence of the most similar VR typing sequence, respectively, as determined by checkerboard hybridization and the porB sequences for each strain (26). Based on evaluations of other PCR ELISA assays (data not shown), optical density (OD) values equal or greater than 2.5 times the OD value obtained for the negative control for that VR, were considered positive for that VR.…”

“…A panel of 40 probes corresponding to loops 1, 2, 3, 6, and 7 for the PIA porin, and 1, 3, 5, 6, and 7 for the PIB porin were employed to characterize VR types of the porB gene (26). Hybridization temperature was 591C for all probes.…”

Extragenital manifestations of Neisseria gonorrhoeae

“…DGI has been associated with strains that have similar porin proteins (PI or PorB) identified either serologically or by molecular typing [22][23][24]. DGI strains are resistant to the bactericidal effects of normal human serum.…”