Combined Vaccine Regimen Based on Parenteral Priming with a DNA Vaccine and Administration of an Oral Booster Consisting of a RecombinantSalmonella entericaSerovar Typhimurium Vaccine Strain for Immunization against Infection with Human-Derived EnterotoxigenicEscherichia coliStrains

Abstract

Repeated evidence has demonstrated that combined primer-booster immunization regimens can improve both secreted and humoral immune responses to antigens derived from viral, bacterial, and parasitic pathogens. For the present work, we evaluated the synergic serum immunoglobulin G (IgG) and fecal IgA antibody responses elicited in BALB/c mice who were intramuscularly primed with a DNA vaccine, pRECFA, followed by oral boosting with an attenuated Salmonella enterica serovar Typhimurium vaccine (HG3) strain, with both vaccines encoding the structural subunit (CfaB) of the CFA/I fimbriae produced by human-derived enterotoxigenic Escherichia coli (ETEC) strains. The immunological properties of the vaccine regimen were evaluated according to the order of the administered vaccines, the nature of the oral antigen carrier, the age of the vaccinated animals, the interval between the priming and boosting doses, and the amount of injected DNA. The production of gamma interferon and the IgG2a subclass in serum indicated that mice immunized with the primer-booster regimen developed prevailing type 1 T-cell-dependent immune responses. The synergic effect of the vaccine regimen on the induced antibody responses was also revealed by its ability to block the adhesive properties of CFA/I fimbriae expressed by live bacteria, as shown by the inhibition of Caco-2 cell and human erythrocyte binding. Moreover, DBA2 newborn mice were protected from lethal challenges with a CFA/I ؉ ETEC strain after the incubation of live bacteria with serum samples harvested from mice who were subjected to the primer-booster regimen. We propose, therefore, that the DNA primer-Salmonella booster regimen represents an alternative for the development of vaccines requiring both mucosal and systemic antibody responses for immunological protection.

Maternal Vaccination with a Fimbrial Tip Adhesin and Passive Protection of Neonatal Mice against Lethal Human Enterotoxigenic Escherichia coli Challenge

“…Establishment of challenge conditions and the LD 50 for H10407 enabled the incorporation of this CFA/ I-ETEC strain into the dam vaccination-pup challenge experiments, in harmonization with volunteer studies in which H10407 has been widely used (38). Demonstration of the protective efficacy of antifimbrial antibodies in the neonatal mouse model is concordant with a prior neonatal mouse study (32) and, more importantly, with a volunteer study in which hyperimmune anti-CFA/I bovine milk IgG conferred passive protection against challenge with H10407 (10), lending credibility to the innovative finding that dscCfaE vaccination also conferred passive protection in the neonatal mouse challenge model. It is important to note that dam vaccination with LTK63 alone by either i.n.…”

“…The neonatal mouse ETEC challenge model was described by Duchet-Suchaux et al (36,46) and was subsequently adapted to test the protective passive immunity conferred by CFA/I-based vaccines (31)(32)(33). In keeping with a prior report, neonatal CBA mice were found to be the most susceptible of several inbred mouse strains to intragastric CFA/I-ETEC challenge in screening experiments, but high rates of cannibalism after CBA pup handling confounded outcomes (unpublished observations).…”

“…Lastly, preincubation of CFA/I-ETEC strain 258909-3 with anti-CFA/I and anti-CfaE bovine colostral IgG over a range of concentrations prior to use in intragastric challenge of pups resulted in a dosedependent increase in survival compared to controls in which the inoculum was incubated with skim milk. Its use here provided a bridge back to previous neonatal mouse challenge studies where this strain, distinct from H10407 in its O:H serotype, had been used to test CfaB-based ETEC vaccines (31)(32)(33).…”

“…LTK63-specific serum IgG and mucosal IgA titers were determined by GM1-ELISA (44), using purified LTK63 as GM1-binding molecule, whereas anti-CFA/I and CfaE antibodies levels were evaluated by conventional ELISA, using plates coated with CFA/I fimbriae or dscCfaE (32,33). Serum anti-LTK63 and anti-CFA/I and anti-CfaE IgG (total IgG and IgG1/IgG2a subclasses) titers were measured with a horseradish per-oxidase-conjugated rabbit anti-mouse IgG antibody (Sigma) diluted to 1:3,000.…”

“…A lethal neonatal mouse model has been used to study the pathogenesis of bovine ETEC and more recently to assess virulence features and test vaccines of human ETEC (30)(31)(32)(33)(34)(35)(36)(37). In the present study, we developed a lethal neonatal mouse challenge with ETEC strain H10407, a CFA/I-ETEC challenge strain that has been extensively used in volunteer challenges (38) and then applied it to test the hypothesis that the dscCfaE fimbrial tip adhesin serves as a protective antigen.…”