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Pathophysiological Effects of Vibrio Cholerae and Enterotoxigenic Escherichia Coli and Their Exotoxins on Eucaryotic Cells KAREN L

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Pathophysiological Effects of Vibrio Cholerae and Enterotoxigenic Escherichia Coli and Their Exotoxins on Eucaryotic Cells KAREN L MICROBIOLOGICAL REVIEWs, Sept. 1978, p. 592-613 Vol. 42, No.3 0146-0749/78/0042-0592$02.00/0 Copyright i 1978 American Society for Microbiology Printed in U.S.A. Pathophysiological Effects of Vibrio cholerae and Enterotoxigenic Escherichia coli and Their Exotoxins on Eucaryotic Cells KAREN L. RICHARDS AND STEVEN D. DOUGLAS* Departments ofMicrobiology and Medicine, University ofMinnesota Medical School, Minneapolis, Minnesota 55455 iNTRODUCTION .5.9.2......592 PATHOPHYSIOLOGY ...................................... 593 Etiology ...........3.................. ... ........3....593 Factors in Pathogenesis ..................................................... 593 Genetic and Physiological Basis for Toxin Production .59.......3.53 THE TOXINS ................................................................. 594 Structure 594 Antigenic Relatedness of Toxins .594 Binding Site 606 Action of Toxins on Adenylate Cyclase .596 Solubilized Adenylate Cyclase and Toxins .597 Cofactors Required for Toxin Activity .597 Involvement of Guanosine 5'-Monophosphate (GMP) ........................ 597 Effect of Increased Intracellular Cyclic Adenosine 3',5'-Monophosphate . 598 Variability in Activation of Adenylate Cyclase .598 Similarities to Glycoprotein Hormones and Other Bacterial Toxins ......... 598 Vaccines and Immunity .......5...............................599 ENTEROTOXINS IN IN VIVO SYSTEMS ... 600 Whole-Animal Models ........... ................................. 600 Ileal Loop Assay ......................................... 600 Skin Permeability Assay ......... 601 ENTEROTOXINS IN IN VITRO SYSTEMS ...... 601 Erythrocyte Ghosts ............................................... 601 Adrenal Cells ....... .... ........ ... 602 Isolated Fat Cells .......................... 603 Lymphocyte and Lymphoid Cell Lines and the Immune Response ..... ...... 603 Fibroblasts ................................................................. 604 Other Cell Systems .................................... 604 DISCUSSION ........6..0.5..................605 CONCLUSIONS .60...................................606 LITERATURE CITED .606 INTRODUCTION differences constitute the basis for further stud- Vibrio cholerae and enterotoxigenic strains of ies on the structural and functional relatedness Escherichia coli cause diarrheal disease in man ofthe toxins and aspects ofmechanisms ofaction and several animal species. E. coli strains can (63, 79, 80, 128). This review will analyze factors produce two enterotoxins; a low-molecular- that are requisite for localization of these orga- weight heat-stable toxin (ST) and a heat-labile nisms within the intestinal tract and toxin pro- toxin (LT) that has many properties similar to ductibn. The physicochemical properties of the those of cholera toxin (CT). Although the mech- toxins and the effects of intestinal cells will be anism of action of E. coli ST is not known, discussed as well as the response of other organ clinical symptoms due to cholera and E. coli LT and cellular systems. In addition, analogies will are the result of action of the toxins on mucosal be drawn between these toxins and other biolog- cells of the small intestine. The major action of ically active molecules. The role they may play V. cholerae and E. coli heat-labile enterotoxins in the growth of the organisms and their use as is to activate adenylate cyclase followed by in- probes into the molecular biology of eucaryotic creases in intracellular levels of cyclic 3',5'-aden- cells will be discussed. osine monophosphate (cAMP) and hypersecre- (Part of this review was presented at the 77th tion of salts and water into the intestinal lumen. Annual Meeting of the American Society for There are a number offeatures distinctive to the Microbiology, 8-13 May 1977, New Orleans, exotoxins of V. cholerae and E. coli. These La.). 592 VOL. 42, 1978 EFFECTS OF E. COLI AND V. CHOLERAE TOXINS 593 PATHOPHYSIOLOGY after centrifugation. Motile vibrios show little or no movement after adherence (116). Vibrio spe- Etiology cies also possess pili (14, 197) which may be Cholera is characterized clinically by an acute involved in adhesion. Adhesion to brush borders enteritis after bacterial colonization of the small occurs at 37 and 220C but not at 40C and is intestine. Massive fluid loss frequently leads to optimum at calcium concentrations of 1 to 10 severe dehydration and shock. V. cholerae is a mM (114), although strontium is also effective. gram-negative, motile, curved rod that grows on Cholera vibrios also agglutinate human group simple nutrient media at alkaline pH (5, 181). O erythrocytes (115). This hemagglutination, as The organism has been classified into the class- well as intestinal adherence, is inhibited by L- ical and El Tor biotypes on the basis of geo- fucose and D-mannose. Nelson et al. (155) ob- graphical distribution and the ability to lyse served more rapid and uniform adhesion of V. sheep erythrocytes (61, 62). Two serotypes, In- cholerae to infant rabbit ileal loops than to adult aba and Ogawa, are distinguishable by heat-sta- rabbit ileal loops. Colonization and clearance of ble somatic antigens. the organisms were similar in the infant and Certain strains of E. coli, commonly found in adult rabbit ileum. great numbers in the large intestine, are capable Enterotoxigenic strains of E. coli adhere to of colonizing the small intestine and have been intestinal epithelium and agglutinate guinea pig identified as causative agents of a severe cholera- and human type 0 erythrocytes (15, 50, 94). like disease in man (6, 82, 175) and domestic Certain protein K antigens on the surface of animals (116). Although several studies have animal isolates are important for adhesion (77, attempted to relate toxigenic strains to particu- 116, 117, 146, 162). A fimbrial antigen demon- lar somatic antigens (84, 88), clinical disease strated in certain diarrheagenic human strains correlates more closely with the presence of acts as a colonization factor (50, 56, 84). In distinct protein capsular (K) antigens (109). In contrast to V. cholerae, adhesion is not temper- addition, certain antigenic types are often asso- ature dependent (77) and is mannose resistant ciated with a particular host species; those iso- (50). K88, the adhesin of E. coli strains patho- lated from pigs possess antigens different from genic for pigs, and K99, found on isolates from those isolated from calves, lambs, or humans calves and lambs, have been isolated and puri- (117, 161). fied (88, 117, 146, 161). Agglutination of guinea pig erythrocytes by K8 is inhibited by mucous glycoproteins with terminal beta-D-galactosyl Factors in Pathogenesis residues (77). Adherence to intestinal surfaces and toxin Certain protein K antigens of pathogenic ani- production are two factors necessary for patho- mal strains and colonization factor antigens of genesis by these two microorganisms. Motility is human isolates are thin, flexible pili that are also a virulence factor for V. cholerae (72, 115). coded for by transmissible plasmids (50, 56, 88, Motile vibrios are more virulent than nonmotile 116, 146). These are distinct from type I pili of organisms of the same strain (72, 85, 115) and E. coli, which are also involved in adhesion and are able to penetrate the intestinal mucus and are receptors for specific phages (14, 157). The enter villous crypts (85, 86). Adherence is amino acid composition of type I pili is different thought to be important to enable the organisms from that of purified K88, and adhesion by type both to resist removal by peristalsis and to mul- I pili to epithelial cells and erythrocytes is in- tiply near the site of action of the toxin (86). hibited in the presence ofmannose (15, 177, 197). Studies of V. cholerae adherence to intestinal Tissue receptors for K88 in pigs are postulated epithelium by Freter and Jones (72), Jones et al. by Sellwood et al. (183) to be genetically deter- (114), and Nelson et al. (155) have demonstrated mined. Two pig phenotypes were observed, and that the organisms penetrate the intestinal mu- the presence of receptors was a dominant char- cus and attach to microvilli at the brush border acteristic; 65% ofthe offspring ofseveral matings of the epithelial cells. Several factors are impor- had K88 receptors, and littermates of particular tant determinants of adhesion of V. cholerae matings were phenotypically identical. The K and include bacterial motility (72), temperature, antigens of E. coli have recently been reviewed and cations (71, 114, 115). The exact nature of in detail (162). the adhesive antigen (adhesin) is not known; Genetic and Physiological Basis for Toxin however, it may be associated with the bacterial Production flagellum. Nonmotile vibrios are unable to ad- here to intestinal brush border membranes and V. cholerae synthesizes a protein exotoxin. intestinal tissue slices (72); nonmotile mutants Genetic studies by Vasel et al. have demon- are unable to adhere to brush border membranes strated that toxin production is coded for by a 594 RICHARDS AND DOUGLAS MICROBIOL. REV. bacterial gene (201). Segregation analysis after plex is noncovalently bound to four to six B (or conjugation indicates that the tox locus is linked L) subunits (136, 143). The B subunit, which is to the locus for histidine synthesis. responsible for toxin binding to cell membranes, Finkelstein et al. (67) have reported artificially contains two cysteine residues which may form induced mutants of V. cholerae that differ from intrasubunit bridges (136). Upon storage or the wild type both
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