Vibrio cholerae (Vc) causes cholera. Its success as a pathogen depends on appropriate transcriptional regulation in different surroundings, such as the human intestinal tract or the aquatic environment, where growth conditions are sub-optimal. Gene regulation results in expression of operons that are critical for generation of a biofilm. Biofilms are complex structures composed of bacterial cells, proteins, and polysaccharides which allow the bacteria to exist in hostile environments. Compared to smooth Vc, the infectious rugose Vc variant is characterized by corrugated colonies, the capacity to develop extensive biofilms, and increased resistance to osmotic and oxidative stresses. Compared to smooth Vc, rugose Vc biofilms have enhanced survival in the presence of bile which may explain why recently shed Vc biofilm aggregates, that have biofilm characteristics, are more infectious for human. The rugosity and biofilm structure modulator gene cluster (rbmA-F) has been identified, which along with several other secreted proteins, are required for the formation of El Tor Vc rugose biofilms. The protective role of antibodies (Abs) to the Rbm proteins has not been studied. Rbm proteins are induced by bile and thus are expressed as part of the infecting inoculum and during the stages of colonization;RbmA, RbmC, and Bap-1 are required for wild type colonization. We hypothesize that the RbmA, RbmC, and Bap-1 (a homolog to RbmC) secreted proteins, known to regulate the biofilm matrix, are protective antigens able to induce antibodies (Abs) that modulate Vc biofilms and thus pathogenesis. These studies are important to determine if biofilm antigens are relevant targets for the immune system and thus cholera vaccines. The proposed study brings together the expertise of two scientists, Drs. Wade and Yildiz who both have extensive research experience in different aspects of cholera research. Dr. W. Wade is a molecular immunologist who pioneered the development of the vaccination protocols for synthetic Vc LPS antigens conjugated to carrier proteins. Dr. Wade is an established immunologist with an extensive track record in generating and characterizing protective Abs to Vc antigens (TcpA, LPS). Dr. F. Yildiz is a recognized molecular biologist who isolated and characterized the Rbm proteins (also referred to in the application as biofilm matrix or matrix proteins) and their requirement for Vc biofilm biology.
Cholera is still a disease that sickens millions and kills thousands yearly. We propose to formulate a new subunit cholera vaccine based on identification of protective epitopes in proteins known to be required for V. cholerae biofilm generation. Previous work from Dr. Wade's laboratories has shown that anti-TcpA antibodies can modify biofilm growth. Other work by Dr. Yildiz's group has identified proteins required for biofilm development that will serve as immunogens to induce antibodies.
