Protective immunity and antibody-secreting cell responses elicited by combined oral attenuated Wa human rotavirus and intranasal Wa 2/6-VLPs with mutant Escherichia coli heat-labile toxin in gnotobiotic pigs

Two combined rotavirus vaccination regimens were evaluated in a gnotobiotic pig model of rotavirus infection and disease and were compared to previous ly tested rotavirus vaccination regimens. The first (AttHRV/VLP2x) involved oral inoculation with one dose of attenuated (Att) Wa human rotavirus (HRV ), followed by two intranasal (i.n.) doses of a rotavirus-like particle (2/ 6-VLPs) vaccine derived from Wa (VP6) and bovine RF (VP2) rotavirus strains . The 2/6-VLPs were coadministered with a mutant Escherichia coli heat-labi le toxin, LT-R192G (mLT) adjuvant. For the second regimen (VLP2x/AttHRV), t wo i.n. doses of 2/6-VLPs+mLT were given, followed by one oral dose of atte nuated Wa HRV. To compare the protective efficacy and immune responses indu ced by the combined vaccine regimens with individual rotavirus vaccine regi mens, we included in the experiments the following vaccine groups: one oral dose of attenuated Wa HRV (AttHRV1 x and Mock2x/AttHRV, respectively), thr ee oral doses of attenuated Wa HRV (AttHRV3x), three i.n. doses of 2/6-VLPs plus mLT (VLP3x), three i.n. doses of purified double-layered inactivated Wa HRV plus mLT (InactHRV3x), mLT alone, and mock-inoculated pigs. The isot ype, magnitude, and tissue distribution of antibody-secreting cells (ASCs) in the intestinal and systemic lymphoid tissues were evaluated using an enz yme-linked immunospot assay. The AttHRV/VLP2x regimen stimulated the highes t mean numbers of intestinal immunoglobulin A (IgA) ASCs prechallenge among all vaccine groups. This regimen induced partial protection against virus shedding (58%) and diarrhea (44%) upon challenge of pigs with virulent Wa H RV. The reverse VLP2x/AttHRV regimen was less efficacious than the AttHRV/V LP2x regimen in inducing IgA ASC responses and protection against diarrhea (25% protection rate) but was more efficacious than VLP3x or InactHRV3x (no protection). In conclusion, the AttHRV/VLP2x vaccination regimen stimulate d the strongest B-cell responses in the intestinal mucosal immune system at challenge and conferred a moderately high protection rate against rotaviru s disease, indicating that priming of the mucosal inductive site at the por tal of natural infection with a replicating vaccine, followed by boosting w ith a nonreplicating vaccine at a second mucosal inductive site, may be a h ighly effective approach to stimulate the mucosal immune system and induce protective immunity against various mucosal pathogens.