J You1, Y Xu, H Li, L Wang, F Wu, F Xu, L Jin, S Li, X Li. 1. School of Life Science and Biotechnology, Dalian University of Technology, Dalian, Liaoning, China; Postdoctoral Working Station, Dalian SEM Bio-Engineering Technology Co., Ltd., Dalian, Liaoning, China.
Abstract
AIM: To obtain a recombinant enterotoxigenic Escherichia coli (ETEC) fusion enterotoxin protein LTB-STa-STb (Bab) that can express the immunogenicity of the haptens STa and STb and induce their corresponding neutralizing antibodies. METHODS AND RESULTS: The three important ETEC enterotoxin genes coding LTB, STa and STb were PCR-amplified, and the amplified products were fused to construct the trivalent enterotoxin expression vector pET30-Bab. SDS-PAGE and Western blot were used to verify the expression of the fusion protein Bab by E. coli BL21 carrying plasmid pET30-Bab. Laying hens immunized with Bab developed high egg yolk immunoglobulin (IgY) titres specific to LTB, STa and STb, and all were significantly higher than those in the control group (P < 0·01). A suckling mouse assay showed that anti-Bab IgY can neutralize the natural toxicity of STa and STb with the highest dilution of 1/2 and 1/32, respectively. CONCLUSIONS: Genetically constructed Bab induced significant antibody responses against STa and STb in chickens, and the resulting IgY had the capacity to neutralize the toxicity of ST. SIGNIFICANCE AND IMPACT OF THE STUDY: The recombinant Bab protein containing three important ETEC enterotoxins may serve as an effective and convenient polyvalent toxoid that can be used to produce multiple antitoxin IgYs to prevent colibacillosis caused by ETEC with various fimbriae in young animals.
AIM: To obtain a recombinant enterotoxigenic Escherichia coli (ETEC) fusion enterotoxin protein LTB-STa-STb (Bab) that can express the immunogenicity of the haptens STa and STb and induce their corresponding neutralizing antibodies. METHODS AND RESULTS: The three important ETEC enterotoxin genes coding LTB, STa and STb were PCR-amplified, and the amplified products were fused to construct the trivalent enterotoxin expression vector pET30-Bab. SDS-PAGE and Western blot were used to verify the expression of the fusion protein Bab by E. coli BL21 carrying plasmid pET30-Bab. Laying hens immunized with Bab developed high egg yolk immunoglobulin (IgY) titres specific to LTB, STa and STb, and all were significantly higher than those in the control group (P < 0·01). A suckling mouse assay showed that anti-Bab IgY can neutralize the natural toxicity of STa and STb with the highest dilution of 1/2 and 1/32, respectively. CONCLUSIONS: Genetically constructed Bab induced significant antibody responses against STa and STb in chickens, and the resulting IgY had the capacity to neutralize the toxicity of ST. SIGNIFICANCE AND IMPACT OF THE STUDY: The recombinant Bab protein containing three important ETEC enterotoxins may serve as an effective and convenient polyvalent toxoid that can be used to produce multiple antitoxin IgYs to prevent colibacillosis caused by ETEC with various fimbriae in young animals.