title: Immunization with recombinant Lactobacillus casei strains producing K99, K88 fimbrial protein protects mice against enterotoxigenic Escherichia coli.
abstract: To exploit a safe and effective vaccine for the prevention against K99 or K88 infections of enterotoxigenic Escherichia coli (ETEC), we have developed a mucosal delivery vehicle based on Lactobacillus casei CICC 6105 using poly-γ-glutamate synthetase A (PgsA) as an anchoring matrix. To evaluate the immunization effect of the recombinant strains (harboring plasmids pLA-K99-K88-LTB, pLA-K99, and pLA-K88), anti-ETEC K99 or K88 antibody responses, T-cell proliferation, and cytokines by intracellular staining (ICS) were investigated after specific pathogen-free (SPF) C57BL/6 mice orally inoculated with these recombinant strains. After oral vaccination into C57BL/6 mice, all recombinant strains were proved to be immunogenic and able to elicit high levels of mucosal immunoglobulin A (IgA) titers in bronchoalveolar lavage fluids, intestinal fluids and prominent systemic immunoglobulin G and IgG subclasses (IgG1, IgG2b, and IgG2a) responses in sera. Using the T-cell proliferation assay, the stimulation index (SI) of groups immunized with pLA-K99/L. casei and pLA-K88/L. casei reached to 2.73 and 2.64, respectively, versus 2.56 in a group immunized with pLA-K99-K88-LTB/L. casei. A detailed analysis of the cell-mediated immune responses by ICS showed the number of specific CD8(+) T cells expressing cytokines (IFN-γ, TNF-α, and IL-2) and granule-associated proteins (CD107a) was higher than that of specific CD4(+) T cells secreted by immune spleen cells upon restimulation in vitro with peptides. Next, the results showed that DCs activated in vitro with recombinant L. casei enhance specific T-cell proliferation and promote T cells to produce both Th1 and Th2 cytokines. More than 80% of the vaccinated mice were protected after challenge with a lethal dose of standard strains C83912 and C83902. These results demonstrate that recombinant L. casei can induce specific humoral and mucosal antibodies and cellular immune response against protective antigens upon oral administration.
authors.affiliation: College of Life Science and Technology, Heilongjiang Bayi Agricultural University, Daqing, Heilongjiang Province, 163319, China.
authors.forename: Li-Juan
authors.initials: LJ
authors.lastname: Wen
authors.forename: Xi-Lin
authors.initials: XL
authors.lastname: Hou
authors.forename: Gui-Hua
authors.initials: GH
authors.lastname: Wang
authors.forename: Li-Yun
authors.initials: LY
authors.lastname: Yu
authors.forename: Xiao-Man
authors.initials: XM
authors.lastname: Wei
authors.forename: Jian-Kui
authors.initials: JK
authors.lastname: Liu
authors.forename: Qinfang
authors.initials: Q
authors.lastname: Liu
authors.forename: Chun-Hua
authors.initials: CH
authors.lastname: Wei
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000032:analysis / Q000097:blood; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology*; D000954:Antigens, Surface / Q000235:genetics / Q000276:immunology*; D001427:Bacterial Toxins / Q000235:genetics / Q000276:immunology*; D001992:Bronchoalveolar Lavage Fluid / Q000276:immunology; D049109:Cell Proliferation; D016207:Cytokines / Q000096:biosynthesis; D004195:Disease Models, Animal; D004337:Drug Carriers; D054307:Enterotoxigenic Escherichia coli / Q000235:genetics / Q000276:immunology*; D004927:Escherichia coli Infections / Q000276:immunology / Q000401:mortality / Q000517:prevention & control*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000276:immunology*; D022361:Escherichia coli Vaccines / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology*; D005260:Female; D036781:Fimbriae Proteins / Q000235:genetics / Q000276:immunology*; D005822:Genetic Vectors; D018928:Immunity, Mucosal; D007114:Immunization / Q000379:methods; D007070:Immunoglobulin A / Q000032:analysis; D007074:Immunoglobulin G / Q000097:blood; D007780:Lacticaseibacillus casei / Q000201:enzymology / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D010453:Peptide Synthases / Q000235:genetics; D016019:Survival Analysis; D013601:T-Lymphocytes / Q000276:immunology; D014614:Vaccines, Synthetic / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D000954:Antigens, Surface; D001427:Bacterial Toxins; D016207:Cytokines; D004337:Drug Carriers; D029968:Escherichia coli Proteins; D022361:Escherichia coli Vaccines; D007070:Immunoglobulin A; D007074:Immunoglobulin G; C026888:K88 antigen, E coli; C031429:K99 antigen; D014614:Vaccines, Synthetic; D036781:Fimbriae Proteins; D010453:Peptide Synthases; C445299:gamma-polyglutamic acid synthetase