EVALUATION OF FORMALIN-INACTIVATED CLOSTRIDIUM-DIFFICILE VACCINES ADMINISTERED BY PARENTERAL AND MUCOSAL ROUTES OF IMMUNIZATION IN HAMSTERS

Clostridium difficile produces toxins that cause inflammation, necrosi s, and fluid in the intestine and is the most important cause of nosoc omial antibiotic-associated diarrhea and colitis. We evaluated C. diff icile antigens as vaccines to protect against systemic and intestinal disease in a hamster model of clindamycin colitis. Formalin-inactivate d culture filtrates from a highly toxigenic strain were administered b y mucosal routes (intranasal, intragastric, and rectal) with cholera t oxin as a mucosal adjuvant. A preparation of culture filtrate and kill ed whole cells was also tested rectally. The toroid was also tested pa renterally (subcutaneously and intraperitoneally) and by a combination of three intranasal immunizations followed by a combined intranasal-i ntraperitoneal boost. Serum antibodies against toxins A and B and whol e-cell antigen were measured by enzyme-linked immunosorbent assay, neu tralization of cytotoxic activity, and bacterial agglutination. The tw o rectal immunization regimens induced low antibody responses and prot ected only 20% of hamsters against death and 0% against diarrhea. The intragastric regimen induced high antibody responses but low protectio n, 40% against death and 0% against diarrhea. Hamsters immunized by th e intranasal, intraperitoneal, and subcutaneous routes were 100% prote cted against death and partially protected (40, 40, and 20%, respectiv ely) against diarrhea. Among the latter groups, intraperitoneally immu nized animals had the highest serum anticytotoxic activity and the hig hest agglutinating antibody responses. Hamsters immunized intranasally and revaccinated intraperitoneally were 100% protected against both d eath and diarrhea, Protection against death and diarrhea correlated wi th antibody responses to all antigens tested. The results indicate tha t optimal protection against C. difficile disease can be achieved with combined parenteral and mucosal immunization.