Lipopolysaccharides of Vibrio cholerae. I. Physical and chemical characterization

Vibrio cholerae is divided into more than 130 O serogroups; however, only organisms of the O1 serogroup have so far been associated with cholera in humans. V. cholerae O1 strains of both biotypes have been further subdivided into three serotypes, designated Inaba, Ogawa, and Hikojima, grouped according to the structure of the O antigens on the lipopolysaccharide (LPS). The LPS of gram-negative bacteria is the most abundant molecule on the cell surface, where it provides a protective barrier to hydrophobic agents and detergents. The most common sugars found in the O polysaccharide are perosamine and quinovosamine. A number of studies designed to correlate the various O-antigen polysaccharides with particular antigenic specificities have been carried out. The genes involved in O-antigen biosynthesis in V. cholerae O1 strains 569B (Inaba, classical) and O17 (Ogawa, EI Tor) have been cloned and expressed in Escherichia coli K-12. The E. coli RfaD is an ADP-L-glycero-D-mannoheptose epimerase and is one of the critical proteins involved in the synthesis of the core oligosaccharide of the LPS in E. coli. Genetic complementation studies have suggested that the determinant responsible for Ogawa specificity lies at the distal end of the rfb region, an area in which no readily detectable differences could be discerned.

Molecular Basis for O-Antigen Biosynthesis in Vibrio cholerae O1: Ogawa-Inaba Switching

https://mmbr.asm.org/content/mmbr/51/2/285.full.pdf

Genetics of Vibrio cholerae and its bacteriophages.

A comparative study was carried out on the sugar composition of lipopolysaccharides (LPS) isolated from representative strains of members of the family Vibrionaceae including all of the constituting genera, ie, Vibrio, Aeromonas, Photobacterium, Plesiomonas, and Lucibacterium More than 100 strains were examined It was found that, with the exception of Vibrio parahaemolyticus 06, 2-keto-3-deoxyoctonate (KDO), known generally as a component sugar in the core region of usual gram-negative bacterial LPS, is virtually absent from LPS of the Vibrionaceae strains so far examined Furthermore, mannose was also lacking in LPS of Vibrionaceae strains with the exception of only one strain, A anaerogenes (ATCC 15467) Instead, some KDO-like substances were found in LPS from Vibrio (“Beneckea”) nereida (ATCC 25917) and Plesiomonas shigelloides including the type strain (ATCC 14029), the same as those found in LPS from V parahaemolyticus O7 and O12, and three strains of V alginolyticus These substances were strongly positive in the periodate-thiobarbituric acid test, yielding a color with maximum absorption at 549 nm The spectra were identical to that of KDO, whereas substances differed from KDO at least in behavior in high-voltage paper electrophoresis and thin-layer chromatography



A particularly interesting feature from the chemotaxonomical point of view was found in the sugar composition of LPS isolated from V cholerae Fructose was present exclusively in LPS of V cholerae (both O1 and non-O1 groups and classical and eltor biotypes) with the exception of one strain of Photobacterium phosphoreum (NCMB 844) In addition, a pair of rarely occurring amino sugars, perosamine and quinovosamine, was found in LPS from O1 group V cholerae regardless of either the biotype (classical or eltor) or the serotype (Inaba or Ogawa), whereas this pair was not present in non-O1 group V cholerae (the so-called NAG vibrios) This feature was confirmed with LPS from more than 30 additional strains of O1 group V cholerae isolated from patients



The virtual absence of KDO in LPS of the family Vibrionaceae was demonstrated for the first time in this study These results are compatible with the interpretation that the absence of KDO in LPS can be used as one of the taxonomical characteristics of Vibrionaceae in addition to (G+C) content, DNA (or RNA) homology, and numerical analysis data

Sugar composition of lipopolysaccharides of family Vibrionaceae. Absence of 2-keto-3-deoxyoctonate (KDO) except in Vibrio parahaemolyticus O6.

A NEW IMMUNO ASSAY OF CHOLERA TOXIN WITH STABLE POLYSTYRENE LATEX PARTICLES Takeshi Yokota, Teruyo Ito, and Shogo Kuwahara* Department of Bacteriology, School of Medicine Juntendo University, Tokyo 113 and * Department of Microbiology, School of Medicine, Toho University, Tokyo 143, Japan. Automatic and manual latex agglutination tests were employed to measure cholera toxin (CT), and their subunits A and B. Dow polystyrene latex particles with an average diameter of 0.22 J.IDl and stable polystyrenechlorostyrene latex particles with a diameter of about 1 J.IDl, both of which were sensitized by purified rabbit specific immunogloblin corresponding to each antigen, were employed for the former and latter methods, respectively. Automatic agglutination was carried out by a nephelometric assay system, in which portions of the suspension of sensitized latex particles and samples were mechanically mixed and the time-dependent differences of light scattering due to agglutination were recorded. Manual latex agglutination was performed in microtiter plates. As low as 1,000 and 31 pg/ml of purified CT were estimated by the automatic and manual latex agglutination tests, respectively. Using the latex agglutination method, we measured the amounts of CT produced by clinical isolates of Vibrio cholerae. Furthermore, fine estimation of CT in numbers of cyclic AMPdeficient mutants of V. cholerae revealed that cyclic AMP is not essential for the production of the toxin. Introduction Accurate measurement of amounts of enterotoxins produced by Vibrio cholerae or enterotoxigenic Escherichia coli is frequently required. This is essential for the identification of nontoxigenic strains of V. cholerae, for the comparison of toxigenicities of the clinical isolates obtained in different decades, and for confirmation of transformants of E. coli by cloned LT genes (13). Conventional assay methods for enterotoxins, however, are complex and timeconsuming. It is almost impossible to estimate accurately CT or LT in one hundred strains by the tissue culture method with the CHO-KI cells even though this method is sensitive enough. Conversely, the reverse passive hemagglutination test is insensitive and may be interfered with by non-specific reactions (7). After the latex agglutination test was first introduced by Singer and Plotz (12) for measurement of rheumatoid factors, it quickly became popular for measurement of various antigens and antibodies (10, 11). To ensure accurate and rapid determination

Archana Dutta

Papers

Structural investigations on the lipopolysaccharide isolated from Vibrio cholera ogawa G-2102

Studies on the partial structure of the O-antigen of Vibrio cholera, Inaba 569 B.

Studies on the partial structure of the O-antigen of Vibrio cholera Ogawa G-2102.