The leading cause of food poisoning in both Taiwan and Japan is Vibrio parahaemolyticus infection, whose mechanism of enteropathogenesis is still unclear. To evaluate whether surface components are responsible for the intestinal adhesion of V . parahaemolyticus , we have developed a novel method for isolating the capsular polysaccharide (CPS) from V . parahaemolyticus (serotype O4:K8). We found that culturing of V . parahaemolyticus in broth for 1 week or more changed the colony form of the bacteria on an agar plate from opaque to translucent. The translucent colonies of V . parahaemolyticus contained little CPS and exhibited a much lower level of adherence to epithelial cells (Int-407) than the opaque colonies of the bacteria. Incubation of V . parahaemolyticus in medium supplemented with bile increased the levels of CPS and adherence. Treatment of V . parahaemolyticus with anti-CPS but not anti-LPS serum decreased the level of bacterial adherence. In addition, purified CPS bound to epithelial cells in a dose-dependent manner. Intranasal administration of CPS to mice in the presence of adjuvants such as immunostimulatory sequence oligodeoxynucleotides or cholera toxin elicited CPS-specific mucosal and systemic immune responses. These results indicate that CPS plays an important role in the adherence of V . parahaemolyticus to its target cells and may be considered a potential target for the development of a vaccine against this pathogen.

Background: Plant viruses can be employed as versatile vectors for the production of vaccines by expressing immunogenic epitopes on the surface of chimeric viral particles. Although several viruses, including tobacco mosaic virus, potato virus X and cowpea mosaic virus, have been developed as vectors, we aimed to develop a new viral vaccine delivery system, a bamboo mosaic virus (BaMV), that would carry larger transgene loads, and generate better immunity in the target animals with fewer adverse environmental effects. Methods: We engineered the BaMV as a vaccine vector expressing the antigenic epitope(s) of the capsid protein VP1 of foot-and-mouth disease virus (FMDV). The recombinant BaMV plasmid (pBVP1) was constructed by replacing DNA encoding the 35 N-terminal amino acid residues of the BaMV coat protein with that encoding 37 amino acid residues (T¹²⁸-N¹⁶⁴) of FMDV VP1. Results: The pBVP1 was able to infect host plants and to generate a chimeric virion BVP1 expressing VP1 epitopes in its coat protein. Inoculation of swine with BVP1 virions resulted in the production of anti-FMDV neutralizing antibodies. Real-time PCR analysis of peripheral blood mononuclear cells from the BVP1-immunized swine revealed that they produced VP1-specific IFN-γ. Furthermore, all BVP1-immunized swine were protected against FMDV challenge. Conclusion: Chimeric BaMV virions that express partial sequence of FMDV VP1 can effectively induce not only humoral and cell-mediated immune responses but also full protection against FMDV in target animals. This BaMV-based vector technology may be applied to other vaccines that require correct expression of antigens on chimeric viral particles.

Bacterial polysaccharides are known to induce the immune response in macrophages. Here we isolated a novel extracellular polysaccharide from the biofilm of Thermus aquaticus YT-1 and evaluated its structure and immunomodulatory effects. The size of this polysaccharide, TA-1, was deduced by size-exclusion chromatography as 500 kDa. GC-MS, high performance anion-exchange chromatography with pulsed amperometric detection, electrospray ionization-MS/MS, and NMR revealed the novel structure of TA-1. The polysaccharide is composed of tetrasaccharide-repeating units of galactofuranose, galactopyranose, and N-acetylgalactosamine (1:1:2) and lacked acidic sugars. TA-1 stimulated macrophage cells to produce the cytokines TNF-α and IL-6. Screening of Toll-like receptors and antibody-blocking experiments indicated that the natural receptor of TA-1 in its immunoactivity is TLR2. Recognition of TA-1 by TLR2 was confirmed by TA-1 induction of IL-6 production in peritoneal macrophages from wild-type mice but not from TLR2^−/− mice. TA-1, as a TLR2 agonist, could possibly be used as an adjuvant and could enhance cytokine release, which increases the immune response. Furthermore, TA-1 induced cytokine release is dependent on MyD88/TIRAP.