Rapid in situ generation of DNA restriction endonuclease patterns for Neisseria gonorrhoeae

Abstract

Restriction endonuclease (RE) digestion patterns of 26 isolates of Neisseria gonorrhoeae representing different serovars of serogroups WI, WII, and WIII were generated by agarose pellet entrapment and in situ digestion with HinfI and BglII. The method was fast, simple, and reproducible, and stable RE patterns were produced from subsequent in vitro passages. The cost of culture materials was reduced considerably, and no toxic or flammable solvents needed to be used. Excellent resolution of DNA fragments of higher molecular weights was obtained as a result of minimal mechanical shearing of the DNA. The REs HinfI and BglII were discriminative in the fragment length ranges of 2 to 6.5 and 2.5 to 21.5 kilobases, respectively. On the basis of densitometric scanning of electrophoretograms generated by HinfI digestion, the 26 isolates representing 12 serovars were divided into seven groups. BglII was found to be more discriminative; 15 RE patterns were established among the 26 isolates. Patterns generated by both REs showed that there was no correlation between a particular RE pattern and a serovar, since strains with identical RE patterns were from different serovars. With the exception of two strains (D3 and D14), which demonstrated positive correlation when both enzymes were used, all strains with identical serovar patterns had different RE patterns.