Involvement of the hap gene (mucinase) in the survival of Vibrio cholerae O1 in association with the blue-green alga, Anabaena sp.

Abstract

Mucinase is a soluble haemagglutinin protease, which may be important for the survival of Vibrio cholerae in association with mucilaginous blue-green algae (cyanobacteria). A comparative survival study was carried out with an Anabaena sp. and a wild-type V. cholerae O1 strain hap+ gene (haemagglutinin-protease), together with its isogenic mutant hap (hap-deleted gene). A simple spread plate technique was followed to count culturable V. cholerae O1 on taurocholate tellurite gelatin agar plate. The fluorescent antibody technique of Kogure et al. (1979) was used for the microscopical viable count of V. cholerae O1. Polymerase chain reaction (PCR) and Southern blot hybridization were carried out to detect a lower number of viable but nonculturable (VBNC) V. cholerae O1 from the laboratory-based experiments. The wild and mutant V. cholerae O1 strains survived in culturable form for 22 and 10 days, respectively, in association with the Anabaena sp., with the difference being statistically significant (P < 0.01). The fluorescent antibody technique, PCR, and hybridization results also showed that the wild strain survived better in the VBNC state than did the mutant VBNC strain in association with an Anabaena sp. These results indicate that the enzyme mucinase may play an important role in the association and long-term survival of V. cholerae O1 with a mucilaginous blue-green alga, Anabaena sp.Key words: Vibrio cholerae, Anabaena, mucinase, reservoir.