Investigations into Metabolically Active yet Non-Culturable (MAyNC) Clostridium perfringens to Control Necrotic Enteritis in Broiler Chickens

Clostridium perfringens-based Necrotic Enteritis (NE) is a disease that can cause significant morbidity and mortality in poultry, causing economic losses of approximately $6 billion annually. This disease was previously controlled using antibiotic growth promoters, which have been phased-out due to concerns of the antibiotic resistance genes. Vaccines against infectious diseases have been proposed as a solution to the use of antibiotics in livestock animals. Electron Beam (eBeam) technology is a commercial technology that inactivates microorganisms by causing significant damage to the nucleic acid. While the cell is unable to replicate, membrane integrity and metabolic activity are maintained. This state is termed a Metabolically Active, yet Non-culturable (MAyNC) and possess characteristics of a potential vaccine: MAyNC cells are unable to replicate and cause disease within a host, but because they are intact and metabolically active, they may retain their antigenicity and immunogenicity. In this study, the MAyNC state of eBeam inactivated C. perfringens (EBCP) was investigated as a potential control mechanism against NE in broiler chickens. The results demonstrated that when exposed to a target dose of 10 kGy, C. perfringens was irreversibly inactivated even when enriched in vitro and administered in a NE challenge model in vivo. The MAyNC state of EBCP was confirmed using metabolic activity and membrane integrity assays. Metabolomic analysis demonstrated that MAyNC C. perfringens had a distinct metabolomic profile immediately after inactivation with increased production of many cell-signaling metabolites. The protective ability of two EBCP cocktails was assessed by vaccinating birds with adjuvanted or non-adjuvanted formulations on day-of-hatch, and then challenging them with homologous live C. perfringens in a NE challenge model. EBCP cocktails alone were not protective upon challenge and birds vaccinated with adjuvanted EBCP had significantly more signs of disease than even unvaccinated control birds. The results of these studies suggest that eBeam inactivation induces a MAyNC state in C. perfringens that has the potential to be used to protect against NE challenge, although further optimization of the vaccine formulation is required