Evaluation of surface components of Vibrio cholerae as protective immunogens

Surface components of a motile Inaba strain (CA401) were removed from washed cells by low speed shearing. Flagella contaminated with a vesicular material (designated as crude flagella [CF]) were obtained by differential centrifugation of the shear fluid. Vesicles were obtained from a nonflagellated mutant by the same procedure. Homogeneous small vesicles were obtained in diminished yield from CsCl gradients of CF preparations. Treatment of CF with sodium deoxycholate removed the vesicular material and flagellar sheaths and yielded naked flagella (NF). The ability of these preparations to passively protect infant mice suckled by CFW mothers that had been immunized at the time of mating was compared, on a dry weight basis, with commercial vaccine (CV). Eight day old mice were challenged orally with more than 1,000 50% lethal doses of either the homologous or a heterologous (Ogawa CA411) strain. The most effective immunogen was CF, which provided complete protection a 1 μg against both challenges. CF and vesicles provided 50 to 100 fold greater protection than CV against homologous challenge. With heterologous challenge, vesicles were 10 fold more protective than CV, but markedly less protective than CF. The NF offered only slightly greater protection than CV against both challenges. Immunoelectrophoresis revealed an antigen in CF distinct from vesicles, cell wall lipopolysaccharide or NF. This antigen is not present in the nonflagellated mutant and is apparently associated with motility.