Performing Department
(N/A)
Non Technical Summary
Necrotic Enteritis (NE), caused byClostridium perfringens, is costing the global poultry industry $6 billion annually. With no current effective mitigation strategies, it is imperative to develop novel antibiotic alternative solutions. Our lab has been investigating the role of commensal bacteria in mediating colonization resistance against NE. Following a recent screening of a library of intestinal bacteria, we identified a Gram-negative commensal bacterium with a robust capacity to inhibitC. perfringensgrowth and enhance innate immunity by stimulating the synthesis of host defense peptides (HDP). Importantly, oral administration of the bacterium provided significant protection to chickens against NE, implying its potential for NE mitigation. The overall goal of this project is to reveal the major antibacterial and immune-boosting metabolites produced by the bacterium and their mechanisms of action in resisting NE. We hypothesize that the commensal bacterium produces antimicrobial metabolites and modulates intestinal microbiota, mucosal immunity, and barrier integrity to limit the colonization ofC. perfringens. Two objectives are proposed: 1) identify major antibacterial and HDP-inducing metabolites and 2) elucidate the mechanisms by which major metabolites resist C. perfringens bothin vitroandin vivo. Upon project completion, several metabolites are expected to be identified with potent anti-C. perfringensand HDP-inducing activities. Additionally, the metabolites are anticipated to accelerate the maturation of the intestinal microbiota, strengthen barrier function, and enhance mucosal immunity. The outcomes will facilitate the development of the commensal bacterium and its metabolites as a novel strategy against NE. Importantly, the project will support an up-and-coming scientist as she pursues a career in animal health research.
Animal Health Component
100%
Research Effort Categories
Basic
10%
Applied
90%
Developmental
(N/A)
Goals / Objectives
The primary aim of this research is to develop a cost-effective antibiotic-free solution for mitigating necrotic enteritis (NE). We recently discovered a chicken cecal commensal bacterium to be capable of inhibitingClostridium perfringensgrowth and inducing host defense peptide gene expression. Importantly, oral administration of the bacterium showed robust protection of chickens from NE. The goal of the current project is to understand the mechanisms underlying the protective action of the bacterium against NE in chickens in order to explore its potential as a probiotic for disease control and prevention against NE. The first objective is toIdentify major antibacterial and HDP-inducing metabolites produced by the bacterium.The 50 most abundantmetabolites will be identified from the overnight culture through quantitative untargeted metabolomics and short-chain fatty acid assays or through fractionation withhigh-performance liquid chromatography. These metabolites responsible for anti-C. perfringensand/or HDP-inducing activity will be identified through individual evaluations and mass spectrometry. Subsequently, the second objective is to elucidate the mechanism through which the major metabolites resistC. perfringenscolonization bothin vitroandin vivo.The antibacterial mechanisms of major metabolites will be assessed by observing their effects on the membrane structure ofC. perfringensusing electron microscopy. Additionally, the impact of the metabolites ontranscriptional response ofC. perfringenswill be analyzed. Additionally, the influence of the bacterial metabolites on the development of intestinal microbiota, immunity, and barrier function will be evaluated in healthy chickens and in a chicken model of NE using a range of multi-omics techniques.
Project Methods
The accomplish our goals, the following two objectives are proposed:Objective 1: Identify major antibacterial and host defense peptide-inducing metabolites produced by the protective commensal bacterium. The cell-free supernatant of the bacterium will be subjected to quantitative untargeted metabolomic assays. The top 50 most abundant metabolites will be evaluated further. To determine if the top metabolites have activity againstClostridium perfringens, an agar well diffusion assay will be used. Further, to determine if the abundant metabolites are responsible for the host defense peptide (HDP)-inducing activity ofMs, a recently developed reporter cell line will be used. The success of this objective will be evaluated by our ability to identify key metabolites produced byMsthat exhibit inhibitory effects onC. perfringensgrowth and stimulate HDP gene transcription.Objective 2: Elucidate the mechanisms by which major metabolites resistC. perfringensbothin vitroandin vivo.Theinfluence of main antibacterial and HDP-inducing metabolites on the membrane structure of C. perfringens will be assessed using both scanning and transmission electron microscopy. Additionally, the transcriptomic responses of C. perfringens to the metabolites will be analyzed. Furthermore, a multi-omics approach will be employed to investigate the impact of the metabolites on the development of intestinal microbiota, immunity, and barrier function in both healthy and NE-infected chickens.