Binding specificities of heat-labile enterotoxins isolated from porcine and human enterotoxigenic Escherichia coli for different gangliosides

1989 ◽  
Vol 35 (6) ◽  
pp. 670-673 ◽  
Author(s):  
Shunji Sugii ◽  
Takao Tsuji
Keyword(s):  
Escherichia Coli ◽  
Competitive Binding ◽  
Enterotoxigenic Escherichia Coli ◽  
Type B ◽  
Binding Assays ◽  
B Subunit ◽  
Human Type ◽  
Heat Labile ◽  
Sepharose 4B ◽  
Competitive Binding Assays

The binding specificities of heat-labile enterotoxins (LTp and LTh) isolated from porcine and human enterotoxigenic Escherichia coli on human erythrocytes were studied by competitive binding assays using different gangliosides as inhibitors. The binding of 125I–labeled LTp to neuraminidase-treated human type A erythrocytes was most effectively inhibited by ganglioside GM1 Ganglioside GM1 was 11 and 105 times more potent than gangliosides GD1b and GM2, respectively. Gangliosides GD1a, GT1b, and GM3 were much less potent. Similar results were also obtained in competitive binding assays with the 125I-labeled B subunit of LTh and neuraminidase-treated human type B erythrocytes, and in those with 3H-labeled ganglioside GM1 and LTp-coupled Sepharose 4B. The binding of 3H-labeled ganglioside GM1 to LTp was not effectively inhibited by galactose-β(1 → 3)N-acetyl-D-galactosamine at the highest concentration used. These findings suggest that the combining sites of LTp and LTh may be specific for at least the galactose-N-acetyl-D-galactosamine-galactose (N-acetyl-neuraminic acid) portion of ganglioside GM1.Key words: binding specificity, heat-labile enterotoxin, enterotoxigenic Escherichia coli, ganglioside.

Expression and Immunogenicity of Enterotoxigenic Escherichia coli Heat-Labile Toxin B Subunit in Transgenic Rice Callus

2009 ◽  
Vol 44 (1) ◽  
pp. 14-21 ◽  
Author(s):  
Tae-Geum Kim ◽  
Bang-Geul Kim ◽  
Mi-Young Kim ◽  
Jae-Kwon Choi ◽  
Eun-Sun Jung ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Transgenic Rice ◽  
Enterotoxigenic Escherichia Coli ◽  
Rice Callus ◽  
Toxin B ◽  
B Subunit ◽  
Heat Labile

scholarly journals Evaluation of the Immunogenicity and Protective Efficacy of an Enterotoxigenic Escherichia coli CFA/I Adhesin–Heat-Labile Toxin Chimera

2020 ◽  
Vol 88 (11) ◽  
Author(s):  
Aisling O'Dowd ◽  
Milton Maciel ◽  
Steven T. Poole ◽  
Michael G. Jobling ◽  
Julianne E. Rollenhagen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Protective Efficacy ◽  
Antibody Responses ◽  
Enterotoxigenic Escherichia Coli ◽  
Content Type ◽  
B Subunit ◽  
Colonization Factor ◽  
Heat Labile ◽  
Intestinal Adhesion ◽  
Colonization Factors

ABSTRACT Recent efforts to develop an enterotoxigenic Escherichia coli (ETEC) vaccine have focused on the antigenically conserved tip adhesins of colonization factors. We showed previously that intranasal immunization with dsc19CfaE, a soluble variant of the in cis donor strand-complemented tip adhesin of a colonization factor of the class 5 family (CFA/I) fimbria, is highly immunogenic and protects against oral challenge with CFA/I-positive (CFA/I+) ETEC strain H10407 in the Aotus nancymaae nonhuman primate. We also reported a cholera toxin (CT)-like chimera (called dsc19CfaE-CTA2/CTB) in which the CTA1 domain of CT was replaced by dsc19CfaE that was strongly immunogenic when administered intranasally or orogastrically in mice. Here, we evaluate the immunogenicity and protective efficacy (PE) of a refined and more stable chimera comprised of a pentameric B subunit of ETEC heat-labile toxin (LTB) in lieu of the CTB pentamer and a donor strand truncation (dsc14) of CfaE. The refined chimera, dsc14CfaE-sCTA2/LTB, was highly immunogenic in mice when administered intranasally or intradermally, eliciting serum and fecal antibody responses against CfaE and LTB, as well as strong hemagglutination inhibition titers, a surrogate for neutralization of intestinal adhesion mediated by CfaE. Moreover, the chimera was safe and highly immunogenic when administered intradermally to guinea pigs. In A. nancymaae, intradermal (i.d.) immunization with chimera plus single-mutant heat-labile toxin [LT(R192G)] elicited strong serum anti-CfaE and anti-LTB antibody responses and conferred significant reduction of diarrhea compared to phosphate-buffered saline (PBS) controls (PE = 84.1%; P < 0.02). These data support the further evaluation of dsc14CfaE-sCTA2/LTB as an ETEC vaccine in humans.

scholarly journals Specificity of the Type II Secretion Systems of Enterotoxigenic Escherichia coli and Vibrio cholerae for Heat-Labile Enterotoxin and Cholera Toxin

2010 ◽  
Vol 192 (7) ◽  
pp. 1902-1911 ◽  
Author(s):  
Benjamin Mudrak ◽  
Meta J. Kuehn
Keyword(s):  
Escherichia Coli ◽  
Cholera Toxin ◽  
Vibrio Cholerae ◽  
Enterotoxigenic Escherichia Coli ◽  
Type Ii Secretion ◽  
Type Ii ◽  
Wild Type ◽  
Secretion Systems ◽  
B Subunit ◽  
Heat Labile

ABSTRACT The Gram-negative type II secretion (T2S) system is a multiprotein complex mediating the release of virulence factors from a number of pathogens. While an understanding of the function of T2S components is emerging, little is known about what identifies substrates for export. To investigate T2S substrate recognition, we compared mutations affecting the secretion of two highly homologous substrates: heat-labile enterotoxin (LT) from enterotoxigenic Escherichia coli (ETEC) and cholera toxin (CT) from Vibrio cholerae. Each toxin consists of one enzymatic A subunit and a ring of five B subunits mediating the toxin's secretion. Here, we report two mutations in LT's B subunit (LTB) that reduce its secretion from ETEC without global effects on the toxin. The Q3K mutation reduced levels of secreted LT by half, and as with CT (T. D. Connell, D. J. Metzger, M. Wang, M. G. Jobling, and R. K. Holmes, Infect. Immun. 63:4091-4098, 1995), the E11K mutation impaired LT secretion. Results in vitro and in vivo show that these mutants are not degraded more readily than wild-type LT. The Q3K mutation did not significantly affect CT B subunit (CTB) secretion from V. cholerae, and the E11A mutation altered LT and CTB secretion to various extents, indicating that these toxins are identified as secretion substrates in different ways. The levels of mutant LTB expressed in V. cholerae were low or undetectable, but each CTB mutant expressed and secreted at wild-type levels in ETEC. Therefore, ETEC's T2S system seems to accommodate mutations in CTB that impair the secretion of LTB. Our results highlight the exquisitely fine-tuned relationship between T2S substrates and their coordinate secretion machineries in different bacterial species.

scholarly journals Intradermal or Sublingual Delivery and Heat-Labile Enterotoxin Proteins Shape Immunologic Responses to a CFA/I Fimbria-Derived Subunit Antigen Vaccine against Enterotoxigenic Escherichia coli

2019 ◽  
Vol 87 (11) ◽  
Author(s):  
Milton Maciel ◽  
David Bauer ◽  
Robin L. Baudier ◽  
Jacob Bitoun ◽  
John D. Clements ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibody Responses ◽  
Enterotoxigenic Escherichia Coli ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cell Agglutination ◽  
Local Skin ◽  
Content Type ◽  
B Subunit ◽  
Heat Labile ◽  
Functional Antibodies

ABSTRACT Enterotoxigenic Escherichia coli (ETEC) is a major cause of infectious diarrhea in children, travelers, and deployed military personnel. As such, development of a vaccine would be advantageous for public health. One strategy is to use subunits of colonization factors combined with antigen/adjuvant toxoids as an ETEC vaccine. Here, we investigated the intradermal (i.d.) or sublingual (s.l.) delivery of CFA/I fimbrial antigens, including CfaEB and a CfaE-heat-labile toxin B subunit (LTB) chimera admixed with double mutant heat-labile toxin (LT) LT-R192G/L211A (dmLT). In addition, we compared dmLT with other LT proteins to better understand the generation of adjuvanted fimbrial and toxoid immunity as well as the influence on any local skin reactogenicity. We demonstrate that immunization with dmLT admixed with CfaEB induces robust serum and fecal antibody responses to CFA/I fimbriae and LT but that i.d. formulations are not optimal for s.l. delivery. Improved s.l. vaccination outcomes were observed when higher doses of dmLT (1 to 5 μg) were admixed with CfaEB or, even better, when a CfaE-LTB chimera antigen was used instead. Serum anti-CFA/I total antibodies, detected by enzyme-linked immunosorbent assay, were the best predictor of functional antibodies, based on the inhibition of red blood cell agglutination by ETEC. Immunization with other LT proteins or formulations with altered B-subunit binding during i.d. immunization (e.g., by addition of 5% lactose, LTA1, or LT-G33D) minimally altered the development of antibody responses and cytokine recall responses but reduced skin reactogenicity at the injection site. These results reveal how formulations and delivery parameters shape the adaptive immune responses to a toxoid and fimbria-derived subunit vaccine against ETEC.

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