Adjuvants can enhance vaccine immunogenicity and induce long-term enhancement of immune responses. Thus, adjuvants are important for vaccine research. Polysaccharides isolated from select Chinese herbs have been demonstrated to possess various beneficial functions and excellent adjuvant abilities. In the present study, the polysaccharides IIP-A-1 and IIP-2 were isolated from Isatis indigotica root and compared with the common vaccine adjuvant aluminum hydroxide via intramuscular co-administration of inactivated rabies virus rCVS-11-G into mice. Blood was collected to determine virus neutralizing antibody (VNA) titers and B and T lymphocyte activation status. Inguinal lymph node samples were collected and used to measure B lymphocyte proliferation. Splenocytes were isolated, from which antigen-specific cellular immune responses were detected via ELISpot, ELISA and intracellular cytokine staining. The results revealed that both types of polysaccharides induce more rapid changes and higher VNA titers than aluminum hydroxide. Flow cytometry assays revealed that the polysaccharides activated more B lymphocytes in the lymph nodes and more B and T lymphocytes in the blood than aluminum hydroxide. Antigen-specific cellular immune responses showed that IIP-2 strongly induced T lymphocyte proliferation in the spleen and high levels of cytokine secretion from splenocytes, whereas aluminum hydroxide induced proliferation in only a small number of lymphocytes and the secretion of only small quantities of cytokines. Collectively, these data suggest that the polysaccharide IIP-2 exhibits excellent adjuvant activity and can enhance both cellular and humoral immunity.
Ruminants are the major reservoirs of Escherichia coli O157:H7 and its fecal shedding mainly act as a source of entry of this pathogen into the human food chain. In humans, E. coli O157:H7 infection causes diarrhea, hemorrhagic colitis and hemolytic uremic syndrome. Intimate adherence of E. coli O157:H7 is mediated by Translocated intimin receptor (Tir) to which intimin binds in the host cell. Since E. coli O157:H7 colonizes intestinal epithelium, the mucosal vaccine has a potential to prevent its colonization. Zonula occludens toxin (Zot) of Vibrio cholerae transiently, reversibly alters epithelial tight junction structure to increase mucosal permeability of macromolecules via paracellular route. The C-terminal region of Zot (?G) responsible for this function could be used for mucosal antigen delivery. Therefore, we employed individual (Tir), cocktail (?G + Tir), fusion protein (?G-Tir) and assessed the efficacy of its intranasal immunization on immunogenicity and fecal shedding of E. coli O157:H7 in streptomycin treated mouse model. Compared to control, ?G + Tir, ?G-Tir immunized mice elicited significant antigen specific antibody titers in serum (IgG, IgA) and feces (IgA), whereas Tir immunized mice induced only serum IgG titer. Cytokine analysis revealed mixed Th1/Th2 type immune response in case of ?G + Tir, ?G-Tir group while that of Tir group was solely Th2 type. Tir, ?G + Tir and ?G-Tir immunized mice showed reduction in shedding of E. coli O157:H7 compared to control group. However, ?G-Tir immunized group performed better than ?G + Tir, Tir group in reducing fecal shedding. Overall, our results demonstrate that intranasal immunization of ?G-Tir induces effective systemic, mucosal, cellular immune responses and represents a promising mucosal subunit vaccine to prevent E. coli O157:H7 colonization.
