Project : USDA ARS

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Research Project: Identification of the Ecological Niches and Development of Intervention Strategies to Reduce Pathogenic Foodborne Pathogens in Poultry

Location: Food and Feed Safety Research

2018 Annual Report

Objectives
Objective 1: Define the differential host-pathogen interactions between Salmonella and chicken and poultry mucosal immune systems using genomic technologies. Determine the relationship between foodborne pathogens and the mucosal innate immune response, focusing on epigenetic reprogramming of host immune genes in persistent infections. Objective 2: Identify and develop key strategies including waste, vaccination (using innate immunity), and lighting management strategies for use at animal production facilities that mitigate and reduce the bacterial load of Salmonella and other foodborne pathogens without the use of antibiotics during pre-harvest production in broiler chickens and turkeys. Objective 3: Analyze and characterize both host and Salmonella proteins that are modulated in expression during infection using quantitative proteomic. Develop strategies to reduce foodborne pathogens by targeting host immune-metabolic signaling pathways affected by Salmonella and Campylobacter virulence factors. Objective 4: Investigate potential alternatives to antibiotics, such as chitosan preparations and other commercially available products, on the cecal levels of Salmonella and Campylobacter using an experimental model and metagenomics. Investigate the potential for use and the mechanism used by specific nutritional supplements to inhibit the transfer of genetic resistance elements, such as plasmids, by conjugation between commensal and foodborne bacteria. Objective 5: Investigate the interaction between yeast and fungi and foodborne bacteria to determine their role as commensals and inhibitors or their use as alternatives to antibiotics as pre-and probiotics. Objective 6: Identify ecological reservoirs of pathogens and the potential role of dispersal of animal waste that enable the retention of foodborne pathogens within animal production facilities and the surrounding environments.

Approach
The Centers for Disease Control and Prevention continues to monitor multistate foodborne outbreaks that impact health of the nation over the last 10 years. One area of concern is the reduction of Salmonella as a foodborne pathogen. Despite control efforts that cost over a half a billion dollars annually, foodborne illnesses due to Salmonella continues to impact the consumer. Poultry are commonly identified as a major source of Salmonella. To develop urgently needed new control strategies against Salmonella, we will take a multi-faceted, but integrated approach to identify and evaluate factors at the pre-harvest level that can be used. Based on previous research and collaborations with industry, we will identify and modify management practices that may decrease foodborne pathogen load, as well as environmental conditions associated with higher risk that would be conducive to pathogen survival and growth. Cost effective alternatives will be suggested throughout the poultry production phase. Environmental areas of concern, such as poultry waste and insect vectors will be included. At a more micro-level, interactions among fungi, protozoa, and other microbes will be evaluated under commercial production practices with the outcome of proposed new strategies for pathogen reduction. Campylobacter, a foodborne pathogen in poultry, has become an increasing concern due to the development of antibiotic resistance, especially to fluoroquinolones. The proposed research will investigate strategies to reduce pre-harvest Campylobacter, which will enhance the microbiological safety of poultry. This is important for food safety, but also for the reduction of potential antimicrobial resistance in animal agriculture and public health. Immune modulation is one approach for new anti-infective therapies, whereby natural mechanisms in the host can be exploited to strengthen therapeutic benefits. The stimulation of innate immunity has considerable potential to induce a profound and rapid cross-protection against multiple serovars of bacteria. Using "omic" techniques, including functional genomics, epigenetics, proteomics, and metabolomics, we will identify effective modulators of innate immunity to control infections, especially in situations where vaccination is not appropriate. Furthermore, metabolism and host immunity are essential requirements for survival. Mounting an immune response requires major changes to metabolic processes. Thus, the integration of central metabolic pathways and nutrient sensing with antibacterial immunity alters cellular energy homeostasis and contributes to the prevention or resolution of infectious diseases. Hence, immune and metabolic response processes govern infectious diseases. Research taken will focus on obtaining a greater understanding of the critical nodes of immunometabolism during Salmonella and Campylobacter infection.

Progress Report
Work under the project during FY 2018 concentrated on investigating the interaction of chicken intestinal tissues and chicken macrophages with Salmonella using kinomic peptide array. The work has identified many key host kinases and pathways which play critical roles in determining the fate of the intracellular Salmonella (Objectives 3 and 4). Additional work in FY 2018 identified crucial elements of the mucosal immune system that provide a remarkable ability to respond and modify diverse encounters in the intestines via two distinct functions: The ability to respond to pathobionts (potential pathogenic microbes), invasive pathogens, and microbial products while also maintaining a state of tolerance to the diverse and beneficial commensal intestinal microbes (Objective 2). The development of the different divisions of the immune response was found to correspond with the acquisition and maintenance of a symbiotic microbiota. This microbiota train stimulates and functionally adjusts the different features of the immune system. Together, the immune system, the microbiota, and the host nutritional/metabolic systems form the "intestinal ménage a trois" which provides the maintenance of optimal gut health.

Accomplishments
1. Alternative to antibiotics. Antimicrobials are provided to poultry and livestock for disease prophylaxis, treatment, and growth promotion. Persistent low dose administration of antimicrobials in animal feed is thought to lead to selection, emergence, and dissemination of drug resistant microorganisms. Alternatives to antimicrobials are actively being sought to replace antimicrobials currently used in agriculture. Methylsulfonylmethane is a dietary supplement that exhibits anti-inflammatory properties and is used for the treatment of osteoarthritis in horses, dogs, and humans. It has been generally regarded as safe by the Federal Drug Administration for the treatment of osteoarthritis. Methylsulfonylmethane was found to be inhibitory to several antimicrobial resistant Escherichia coli during initial studies. ARS researchers in College Station, Texas, have done several growth studies to determine parameters of inhibition by methylsulfonylmethane and to determine if it was bacteriostatic or bactericidal. Methylsulfonylmethane at 0, 3, 5, 7, 10, 12, and 16% was evaluated on E. coli, Salmonella Kinshasa, vancomycin-resistant Enterococcus faecium, and Staphylococcus aureus strains. Methylsulfonylmethane was bacteriostatic to all strains. Methylsulfonylmethane demonstrated bacteriostatic inhibition on bacterial growth in short-term growth studies, but was bactericidal in long-term growth studies under the conditions used during this study. Methylsulfonylmethane provided to food animals as a nutritional supplement may reduce the need for antimicrobial use in agriculture. If true, this would have a global impact with regard to agriculture's contribution to antimicrobial resistance.

2. Genetic selection for resistance to foodborne pathogens in poultry. Breeding chickens resistant to Salmonella and Campylobacter infection is considered to be a potential long-term intervention in controlling these bacteria in broiler chicken production. ARS scientists at College Station, Texas, developed an innovative selection strategy based on a phenotype of inherently higher pro-inflammatory mediators that showed the profile of the sire was passed onto progeny. This approach is very different from other selection strategies that are seeking to improve resistance against single pathogens. A population of sires and dams from two lines of broiler chickens have been screened, and individuals with naturally high and low levels of key immune markers (IL6, CXCLi2, and CCLi2) have been identified. Selection based on pro-inflammatory mediators could be valuable in light of stricter regulations with respect to antibiotic use, and may provide the poultry industry with a viable option to enhance selection for improved robustness, livability, and resistance against a broad range of poultry and foodborne pathogens.

U.S. DEPARTMENT OF AGRICULTURE

Food and Feed Safety Research: College Station, TX