Production of Egg Yolk Immunoglobulin (IgY) Against Recombinant LTB of Enterotoxigenic Escherichia coli (ETEC) and Evaluation of Its Protective Effect in Animal Model

Abstract

Background and Aim: Enterotoxigenic Escherichia coli is one of the most common bacterial causes of diarrhea worldwide. One of the most important ETEC colonization factors is B subunit of the heat labile enterotoxin, which forms the connecting part of the toxin. In the present study, egg yolk immunoglobulin (IgY) against recombinant LTB protein was produced and its protective effect was evalutated in the animal model. Material and methods: Hens were injected intramuscularly with 100 μg of recombinant LTB protein. Antibody titers of the sera were estimated and the eggs were collected. IgYs were purified from egg yolk and the antibody titers were subsequently estimated by ELISA method. Effecacy of different concentrations of IgY against the effect of LT toxin on Y1 cells was also studied. In order to investigate the effect of IgY-treated bacteria on intestinal epithelial cells we used standard Ileal loop technique. Results: Induction of protein expression led to the production of recombinant LTB with molecular weight of about 14 kDa. Concentration of purified protein was 4.5mg/ml. Immunization of the hens induced serum antibody rise. IgY against LTB was purified from egg yolk at a concentration of 12mg/ml. 125 μg per ml of IgY against LTB prevented the effects of heat labile enterotoxin on Y1. In the Ileal loop test, 1.5 mg / ml of IgY neutralized the toxin effect of LTB on the intestine. Accumulation of fluid in the test loops decreased by 74.8% compared to that in the untreated control loops. Conclusion: The results obtained indicated that specific egg yolk immunoglobulin was effective against recombinant LTB protein and can be used as a preventive antibody to inhibit the heat labile enterotoxin function of ETEC bacteria.