Mice protected by oral immunization with Lactobacillus reuteri secreting fusion protein of Escherichia coli enterotoxin subunit protein.
A green fluorescent protein (gfp) gene was ligated to the Lactobacillus reuteri-specific nisin-inducible expression-secretion vector pNIES, generating a pNIES-GFP vector capable of secreting the cloned gene as a GFP-fusion protein with fluorescent activity. To develop this system as a live vehicle carrying the heat-stable enterotoxin (ST) and heat-labile enterotoxin B (LT(B)) of the enterotoxigenic Escherichia coli (ETEC), a recombinant 5'-ST-LT(B)-3' DNA fragment was cloned into pNIES-GFP. The resulting L. reuteri/pNIES-GFP:STLT(B) system was found to possess the capability of adhering to the mice gut, secreting GFP:STLT(B) product at 0.14 and 0.026 pgcell(-1) under induced and noninduced conditions, respectively. Further analysis of the GFP:STLT(B) product confirmed its ganglioside-binding ability, LT(B) antigenicity and relative freedom from the ST-associated toxicity, making it suitable for use as an oral vaccine in mice. Oral inoculation of the L. reuteri/pNIES-GFP:STLT(B) culture in mice elicited significant (P<0.01) serum IgG and mucosal IgA antibodies against the STLT(B) antigen. These immunized mice were subsequently challenged with ETEC and showed full protection against the fluid influx response in the gut. This is the first report of using L. reuteri as a vaccine carrier to induce complete immunologic protection against ETEC.
Wu
Chi-Ming
CM
Institute of Veterinary Microbiology, National Chung-Hsing University, Taichung, Taiwan.
Chung
Tung-Ching
TC
D000284:Administration, Oral; D000818:Animals; D001427:Bacterial Toxins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D001428:Bacterial Vaccines / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D003001:Cloning, Molecular; D004768:Enterotoxins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000276:immunology; D004927:Escherichia coli Infections / Q000517:prevention & control*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D007114:Immunization; D007070:Immunoglobulin A / Q000097:blood; D007074:Immunoglobulin G / Q000097:blood; D007422:Intestines / Q000276:immunology; D052200:Limosilactobacillus reuteri / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D009561:Nisin / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
D001427:Bacterial Toxins; D001428:Bacterial Vaccines; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D007070:Immunoglobulin A; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; C034151:heat stable toxin (E coli); D009561:Nisin; D049452:Green Fluorescent Proteins; C027538:heat-labile enterotoxin, E coli