Evaluation of a toxoid fusion protein vaccine produced in plants to protect poultry against necrotic enteritis

Abstract

BackgroundNecrotic enteritis (NE) is caused by type A strains of the bacteriumClostridium perfringens. Total global economic losses to the poultry industry due to NE is estimated to be over two billion dollars annually. Traditionally, NE has been effectively controlled by inclusion of antibiotics in the diet of poultry. However, recent concerns regarding the impact of this practice on increasing antibiotic resistance in human pathogens have led us to consider alternative approaches, such as vaccination, for controlling this disease. NE strains ofC. perfringensproduce two major toxins, a-toxin and NetB. Immune responses against either toxin can provide partial protection against NE.MethodsWe have developed a fusion protein combining a non-toxic carboxyl-terminal domain of a-toxin (PlcC) and an attenuated, mutant form of NetB (NetB-W262A) for use as a vaccine antigen to immunize poultry against NE. We utilized a DNA sequence that was codon-optimized forNicotiana benthamianato enable high levels of expression. The 6-His tagged PlcC-NetB fusion protein was synthesized inN. benthamianausing a geminiviral replicon transient expression system, purified by metal affinity chromatography, and used to immunize broiler birds.ResultsImmunized birds produced a strong serum IgY response against both the plant produced PlcC-NetB protein and against bacterially produced His-PlcC and His-NetB. Immunized birds were significantly protected against a subsequent in-feed challenge with virulentC. perfringenswhen treated with the fusion protein. These results indicate that a plant-produced PlcC-NetB toxoid is a promising vaccine candidate for controlling NE in poultry.