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

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 CFA/I fimbria, is highly immunogenic and protects against oral challenge with CFA/I+ ETEC strain H10407 in the Aotus nancymaae non-human 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 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, ID immunization with chimera + LT(R192G) elicited strong serum anti-CfaE and anti-LTB antibody responses and conferred significant reduction of diarrhea compared to PBS controls (PE = 84.1%, P<0.02). These data support the further evaluation of dsc14CfaE-sCTA2/LTB as an ETEC vaccine in humans.

The Immunogenicity and Properties of a Whole-Cell ETEC Vaccine Inactivated with Psoralen and UVA Light in Comparison to Formalin

“…LTB ELISA. The detection of LTB-specific antibodies was performed according to a published protocol [ 45 ]. Plates were coated with 100 µL of GM1 ganglioside diluted in PBS (0.5 µg/mL, Sigma-Aldrich #G7641) for 18–24 h at 23 °C.…”

“…LTB ELISA. The detection of LTB-specific antibodies was performed according to a published protocol [45]. Plates were coated with 100 µL of GM1 ganglioside diluted in PBS (0.5 µg/mL, Sigma-Aldrich #G7641) for 18-24 h at 23 • C. After 3 washes, the plates were blocked, and 100 µl of recombinant LTB (Sigma-Aldrich #E8656) diluted to 0.5 µg/mL in antibody buffer was added for 1 h at 23 • C. After 3 washes, 100 µL of mouse sera diluted serially in antibody buffer was added for 2 h at 23 • C. The rest of the protocol was as per the ETEC whole-cell ELISA.…”

Confronting Challenges to Enterotoxigenic Escherichia coli Vaccine Development

“…This tip adhesin complex was subsequently tested for immunogenicity when delivered transcutaneously with mutant LT (R192G) via patch in human volunteers (87). Although these studies indicate that additional effort will be required for optimal delivery of the fimbrial tip adhesin complex (121), recent studies of dscCfaE genetically fused in a chimeric complex with the A2 subunit of cholera toxin and the B subunit of LT (dscCfaE-sCTA2/LTB) resulted in substantial protection in an Aotus nancymaae model of infection, following intradermal vaccination with mLT(R192G) (89). Similar preclinical studies of CS6, an antigen present in~1/5 of ETEC strains have been ongoing.…”

The role of CFA/I in adherence and toxin delivery by ETEC expressing multiple colonization factors in the human enteroid model

“…Assembled by the chaperone-usher pathway, CFA/I extends from the bacteria in a peritrichous manner, creating hair-like projections across the surface. CFA/I is one of the most well studied CFs; it has been demonstrated to be important for pathogenesis in human clinical trials as well as in laboratory-based studies and is included as a target in multiple vaccine strategies [18][19][20][21][22].…”