| Summary: | Brachyspira are the causative agent of avian intestinal spirochaetosis, a gastrointestinal disease common in layer hens and broiler breeders. This disease costs the UK laying industry approximately £18 million per annum, resulting from reduced egg production and poor egg quality. Prevalence of Brachyspira is increasing, and due to the poor understanding of this pathogen, mitigation strategies have been largely unsuccessful. Therefore, preventative measures are essential. These studies aimed to improve the understanding of Brachyspira pathobiology and investigate Lactobacillus probiotics as a suitable mitigation strategy. Brachyspira and Lactobacillus species were characterised using phenotypic and genotypic methods. Four Lactobacillus isolates were selected for their inhibition of Brachyspira in vitro and demonstrated inhibition by a number of mechanisms. Secreted metabolites in Lactobacillus cell free supernatant inhibited Brachyspira (p value < 0.05) and metabolomic studies identified the production of organic acids to be a major contributor to inhibition. Protein denaturation in cell free supernatants significantly reduced Brachyspira inhibition (p value < 0.05), suggesting the role of bacteriocins in inhibition. Furthermore, L. reuteri isolates co-aggregated with Brachyspira in vitro, reducing pathogen viability (p value < 0.05). Pro-inflammatory responses to Brachyspira in HD11 avian macrophages were dominated by upregulation of IFNg (p value < 0.01) and pre-treatment of cells with Lactobacillus significantly reduced this response (p value < 0.0001), demonstrating the ability of probiotics to alter immune responses to Brachyspira. Galleria mellonella were utilised to study Brachyspira virulence and probiotic intervention. G. mellonella exhibited a varied response to Brachyspira iv infection and Lactobacillus isolates were able to protect against the mortality associated with Brachyspira isolates (p value < 0.05). The studies here demonstrated that Lactobacillus probiotics are a suitable mitigation strategy against Brachyspira. A number of mechanisms were identified, however future studies are required to explore these mechanisms in a more relevant in vivo chicken model.
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