|LEE, KW - US Department Of Agriculture (USDA)
|KIM, WH - US Department Of Agriculture (USDA)
Submitted to: Avian Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/15/2020
Publication Date: 6/17/2020
Citation: Lee, K., Kim, W., Li, C.Z., Lillehoj, H.S. 2020. Detection of Necrotic Enteritis B-like toxin secreted by Clostridium perfringens using capture Enzyme-Linked Immunosorbent Assay. Avian Diseases. 64(4):490-495. https://doi.org/10.1637/0005-2086-64.4.490.
Interpretive Summary: Necrotic enteritis (NE) is a devastating enteric disease caused by Clostridium perfringens type A/G which impacts global poultry industry by compromising performance, health and welfare of the chickens. The causative main virulent factor responsible for NE pathogenesis has been identified as NE B-like (NetB) toxin, but there is no diagnostic assay for NE. In this paper, ARS scientists and collaborators in Korea developed twenty monoclonal antibodies which specifically identify recombinant NetB toxin protein. These antibodies were used to develop a high throughput ELISA method that can be used to screen chicken samples from poultry farm to identify NE infection. In conclusion, this paper report the successful development of monoclonal antibody-based antigen capture ELISA to detect native NetB protein that will allow the accurate measurement of C. perfringens virulence in poultry samples to monitor flock health status or to make early detection of NE outbreak in NE-susceptible farms.
Technical Abstract: Necrotic enteritis (NE) is a devastating enteric disease caused by Clostridium perfringens type A/G which impacts global poultry industry by compromising performance, health and welfare of the chickens. The causative main virulent factor responsible for NE pathogenesis has been shifted from a-toxin, which is a phospholipase C, to NE B-like (netB) toxin, plasmid-encoded pore-forming heptameric protein, in NE development. The fact that netB protein can only be detected by western blot analysis using polyclonal anti-netB antiserum attempted us to develop netB-specific monoclonal antibody (mAb)-based capture enzyme-linked immunosorbent assay (ELISA). Twenty mAbs reacting against E. coli expressed netB protein were selected, isotyped, and conjugated with horseradish peroxidase for antibody pair test. Multiple mAb pairs were found to detect E. coli netB protein and the mAbs could detect native netB protein originated from C. perfringens using western blot assay. The developed capture ELISA could quantify in vitro production of native netB protein secreted from netB-positive C. perfringens. Here, we first report that native NetB toxin can be detected in C. perfringens netB-specific monoclonal antibody-based capture ELISA.