Location: Poultry Microbiological Safety and Processing Research Unit
2021 Annual Report
Objectives
1. Identify and determine the presence and contributing factors for antimicrobial resistant foodborne bacteria in poultry and poultry-associated environments.
1.1. Determine the association of antimicrobial resistance (AR) in foodborne bacteria with resistance to biocides, metals, coccidiostats, and ionophores used in poultry husbandry and processing.
1.2. Evaluate the bacterial metagenome of retail poultry.
1.3. Identify and evaluate markers (resistance genes, genetic elements, virulence genes) to define outbreak and persistent foodborne bacteria in poultry.
1.4. Identify antimicrobial resistance gene cassettes (ARCs) and accumulation on plasmids.
2. Identify and evaluate biological and chemical intervention products and alternatives to antimicrobials to control or reduce foodborne pathogens in poultry.
2.1. Develop, validate, and produce multi-subunit vaccines to control Salmonella and Campylobacter in broiler chickens.
2.2. Develop antimicrobial peptides (AMP) as alternatives to antibiotics to reduce foodborne pathogens associated with poultry.
2.3. Identify and develop broad-spectrum bacteriocins to eliminate foodborne pathogens in poultry.
2.4. Utilize phage isolation, whole-genome sequencing (WGS), and metagenomics to identify lytic phage that target Salmonella and pathogenic Escherichia coli.
Approach
Microbial contamination of food products from poultry continues to be a leading cause of foodborne illness. Antibiotics have been used to treat bacterial infections since the mid-twentieth century. Because of their efficacy in treating and preventing disease, antimicrobials have also been widely used in poultry production contributing to antimicrobial resistance (AR) in foodborne pathogens and commensal bacteria. AR among these bacteria has the potential to compromise therapy and remains a global threat to human health. This research project represents a merger of two teams of scientists to provide solutions to colonization of poultry with human pathogens and AR in foodborne pathogens and commensal bacteria from poultry. Two major approaches will be employed: 1) development of alternatives to antibiotics for use in combating foodborne pathogens, and 2) investigations to accurately understand attributes of antimicrobial resistant foodborne pathogens and commensals. Alternatives to antibiotics include vaccines to control foodborne pathogens in live birds while innovative antimicrobial peptides, bacteriocins, and lytic phage will modulate the poultry microbiome to reduce or eliminate colonization by harmful bacteria from poultry to minimize AR and reduce risk to human health. Data generated on resistance to biocides, metals, coccidiostats, and ionophores used in poultry production and processing is a specific concern to the USDA Food Safety and Inspection Service (FSIS). Research designed to determine ecological niches of foodborne bacteria and identify genetic characteristics facilitating transfer of resistance or a fitness advantage will also benefit FSIS. According to FSIS, increased knowledge of the microbial ecology of antimicrobial resistant pathogens on poultry will result in data that the poultry industry can utilize in development of improved pathogen management strategies. Identification of genetic markers which support survival, persistence, and dissemination of foodborne pathogens, especially those that are resistant to antimicrobials, is critical to this research priority. Data and technology from the proposed research will be used to assist other Federal agencies and the poultry and agricultural biotechnology industry in addressing AR in poultry resulting in safer products for the consumer.
Progress Report
This project is new commencing in April 2021 replacing 6040-32000-009-00D “Monitoring and Molecular Characterization of Antimicrobial Resistance in Foodborne Bacteria” and 6040-32000-071-00D “Novel Pre-Harvest Interventions and Alternatives to Antibiotics to Reduce Foodborne Pathogens” and focuses on the development of alternatives to antibiotics and attributes of antimicrobial resistant foodborne pathogens and commensals from poultry.
Production of a susceptibility plate for testing bacterial resistance to biocides and heavy metals continued under Sub-objective 1.1. The biocide panel was used to assay multidrug-resistant Salmonella Infantis isolates to determine if the pESI plasmid (plasmid for Emerging Salmonella Infantis) in this strain conferred resistance to any biocides that would explain the persistence of S. Infantis in U.S. poultry production. The metal susceptibility panel was also further tested. The problem of precipitation of some metals when bacterial growth media was added to the assay was remedied with the use of minimal media.
For Sub-objective 1.2, research was conducted to investigate quinolone resistance in 250 Campylobacter spp. isolates from chicken livers. The gyrase A gene of Campylobacter isolates was amplified by PCR, purified, and sequenced. Results showed that in addition to the previously known amino acid residues, several new point mutations were identified as compared to the reference sequence.
Under Sub-objective 1.3, research to define markers in outbreak Salmonella serotype Infantis plasmid pESI strains in poultry was expanded. Prevalence of S. Infantis with the plasmid in U.S. isolates was determined and compared to the other countries where this strain and plasmid combination is a major issue globally. Approximately 90 percent of the genes found in pESI were conserved across all isolates, indicating that conserved regions such as virulence genes, iron acquisition genes, and metal resistance genes may be required for Infantis persistence in U.S. poultry. This information was presented in a publication through a collaboration with Centers for Disease Control and Prevention and the Food Safety and Inspection Service. Continuing work on this issue determined that Infantis with the plasmid is found in almost 30% of U.S. chicken isolates in 2020 and is also found in turkeys. In late 2020, the pESI plasmid was also detected in two new serotypes of Salmonella isolated from turkeys. This research was presented to poultry stakeholders, and a focus group of ARS scientists from Athens, Georgia, College Station, Texas, Ames, Iowa, and Fayetteville, Arkansas was formed to address this issue in U.S. poultry.
A collaborative effort with the Georgia Institute of Technology and Emory University was initiated to develop and produce an mRNA-based vaccine against Salmonella for use in poultry for Sub-objective 2.1. Evaluation of potential antigenic proteins in Salmonella for the vaccine targets has begun.
Accomplishments
