https://mmbr.asm.org/content/mmbr/47/4/510.full.pdf

New knowledge on pathogenesis of bacterial enteric infections as applied to vaccine development.

https://mmbr.asm.org/content/mmbr/60/1/167.full.pdf

Enteric bacterial toxins: mechanisms of action and linkage to intestinal secretion.

Vibrio cholerae O1 A-B- vaccine strain JBK 70 and A-B+ CVD 101 prepared by recombinant DNA techniques from pathogenic EI Tor Inaba N16961 and classical Ogawa 395, respectively, were fed to 38 volunteers in single doses of 10(4) to 10(10). Although severe diarrhea did not occur in any vaccine, more than one-half developed mild diarrhea. These attenuated strains colonized well and elicited prominent vibriocidal and antitoxic (CVD 101) antibody responses. Recipients of a single dose of JBK 70 were significantly protected when challenged with 10(6) wild-type N16961. Diarrhea occurred in 7 of 8 controls but in only 1 of 10 vaccines (P less than 0.003, 89% vaccine efficacy), demonstrating the potency of immune mechanisms that do not involve cholera antitoxin. Further derivatives were prepared to explore the pathogenesis of the residual diarrhea, considering that either intestinal colonization by the vaccine itself or accessory toxins might be responsible. CVD 102, an auxotrophic mutant of CVD 101, did not cause diarrhea but colonized poorly and elicited feeble immune responses. Derivatives of JBK 70 and CVD 101 (CVD 104 and 105) deleted of genes encoding the EI Tor hemolysin still caused mild diarrhea. Genetically engineered strains can be colonizing, highly immunogenic, and protective single-dose oral vaccines, but they must be further attenuated before they can be considered for use as public health tools.

Volunteer studies of deletion mutants of Vibrio cholerae O1 prepared by recombinant techniques.

Vibrio cholerae causes the potentially lethal disease cholera through the elaboration of the intestinal secretogen cholera toxin. A second toxin of V. cholerae, Zot, decreases intestinal tissue resistance by modifying intercellular tight junctions. In this report, a third toxin of V. cholerae, Ace (accessory cholera enterotoxin), is described. Ace increases short-circuit current in Ussing chambers and causes fluid secretion in ligated rabbit ileal loops. The predicted protein sequence of Ace shows striking similarity to eukaryotic ion-transporting ATPases, including the product of the cystic fibrosis gene. The gene encoding Ace is located immediately upstream of the genes encoding Zot and cholera toxin. The ctx, zot, and ace genes, which are located on a dynamic sector of the chromosome, comprise a V. cholerae "virulence cassette."

https://www.pnas.org/content/pnas/90/11/5267.full.pdf

Accessory cholera enterotoxin (Ace), the third toxin of a Vibrio cholerae virulence cassette

Live oral candidate cholera vaccines have previously been constructed by deletion of Vibrio cholerae sequences encoding the enzymatically active A subunit of the cholera toxin. However, volunteer studies have shown that these non-cholera toxin-producing strains still provoke mild to moderate diarrhea in some individuals. We recently reported the identification of a second toxin produced by V. cholerae which may be responsible for this residual diarrhea (A. Fasano, B. Baudry, D. W. Pumplin, S. S. Wasserman, B. D. Tall, J. M. Ketley, and J. B. Kaper, Proc. Natl. Acad. Sci. USA 88:5242-5246, 1991). This new toxigenic factor increases the permeability of rabbit ileal mucosa by affecting the structure of the intercellular tight junctions (zonula occludens). We now report the identification and cloning of the gene encoding this new toxin. This gene, named zot (for zonula occludens toxin), consists of a 1.3-kb open reading frame which could potentially encode a 44.8-kDa polypeptide. The location of the zot gene encoding the new toxin is immediately upstream of the ctx operon encoding cholera toxin.

https://iai.asm.org/content/iai/60/2/428.full.pdf

Cloning of a gene (zot) encoding a new toxin produced by Vibrio cholerae.

Nonenterotoxigenic strains no. 1196-78 and no. 1074-78 of Vibrio cholerae serogroup O1 (biotype El Tor, serotype Ogawa) were isolated from sewage water in Brazil and fed to 20 volunteers. Neither strain caused diarrhea. None of the seven volunteers who ingested Ogawa strain no. 1074-78 (10(6) organisms) excreted the organism whereas eight of the 13 volunteers who ingested Ogawa strain no. 1196-78 (10(6) or 10(8) organisms) did excrete the organism in their stools. None of 114 stool-culture isolates yielded cholera enterotoxin, and none of the 20 volunteers had significant increases in serum titers of antitoxin as measured by enzyme-linked immunosorbent assay although six of the volunteers had slightly elevated vibriocidal antibody levels. Six volunteers used as controls and four volunteers who had stool cultures positive for strain no. 1196-78 of V. cholerae were challenged with pathogenic El Tor Ogawa strain no. E7946 (10(6) organisms) to determine if previous ingestion of the Brazilian strain would induce protective immunity. All 10 of the volunteers developed diarrhea, and the severity of the illness was similar in both groups.

D. M. Gill

Papers

The Pathogenicity of Nonenterotoxigenic Vibrio cholerae Serogroup 01 Biotype EI Tor Isolated from Sewage Water in Brazil