Colonization factor of Vibrio cholerae O1

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Sugar chain of LPS, OmpU, FSHA and NAGV14 pili of Vibrio cholerae were investigated to identify colonization factor(s) and protective antigen(s) of cholera vibrios.1) LPS sugar chain: When cholerae vibrios (V. cholerae O1 and O139) were treated with anti-LPS Fab, the adhesive ability was lowered but the level was not significant. Heat- or formaline-killed organisms maintaining the sugar chain did not adhere to the intestinal epithelium. A few strains with normal LPS which did not adhere to the intestinal epithelium were found. From these findings, LPS sugar chain was eliminated from the candidates of colonization factors of cholerae vibrios.2) OmpU: OmpU was purified and characterized. The purified OmpU adhered to the intestinal epithelium, however, the adhesion was not inhibited by treating the organisms with anti-OmpU and treating the intestine with purified OmpU. The antibody did not agglutinate the organisms, suggesting OmpU is not surface exposed. Therefore, OmpU is not supposed to function as a colonization factor.3) FSHA: We failed to purify L-fucose sensitive hemagglutinin (FSHA), therefore, genetic approach was made. Transposon (Tn5) was randomly inserted to the chromosome of Vibrio cholerae O1 strain 86B3, and hemagglutinin negative mutant was screened. Now we are on the process of identifying the position of hemagglutination gene.4) NAGV14 pili: NAGV14 pilus is a sole element which adhere to the intestinal epithelium found in V. cholerae. Gene analysis was carried out by using degenerate PCR, inverse PCR, PCR walking, and LA PCR cloning kit, based on the N-terminal amino acid sequence of the subunit protein of the pili. In conclusion, the gene encoding NAGV14 pili was not found in cholera vibrios but some restricted strains of Vibrio cholerae which are likely enteropathogenic.