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ARS Home » Plains Area » Clay Center, Nebraska » U.S. Meat Animal Research Center » Meat Safety and Quality » Research » Research Project #429923

Research Project: Assessment of Genotypic and Phenotypic Factors for Foodborne Pathogen Transmission and Development of Intervention Strategies

Location: Meat Safety and Quality

2017 Annual Report

Objective 1. Determine the genotypic and/or phenotypic factors associated with the levels and persistence of pathogens and antibiotic resistance in the host animal and the livestock production environment. Sub-objective 1.1 - Determine genotypic or phenotypic factors associated with persistence of E. coli O157:H7 in cattle production. Sub-objective 1.2 - Determine animal host genotypes that confer resistance/susceptibility to pathogen infection. Sub-objective 1.3 - Evaluate the influence of spatiotemporal, environmental, and wildlife factors on pathogen and antibiotic resistance occurrence and transmission dynamics in cattle and waterways in a pasture-based production system. Objective 2. Develop and evaluate intervention strategies that reduce or eliminate the occurrence, transmission or persistence of foodborne pathogens in host animals, including cattle and swine, and the environment. Sub-objective 2.1 - Develop a high-throughput procedure to identify and measure antibiotic resistance genes. Sub-objective 2.2 - Identify alternatives to antibiotics for use as growth promoters in production animals. Sub-objective 2.3 - Determine the effect of calcium hydroxide application to feedlot pens on E. coli O157:H7, total E. coli, and antibiotic resistance on feedlot pen surfaces and in cattle.

The overall goal of this project is to reduce the risk of human foodborne illness, by providing information that can be used to reduce the transmission of zoonotic pathogens and antibiotic resistance from cattle and swine production to food, water, and the environment. Primary targets of the work include Escherichia coli O157:H7 and other Shiga-toxigenic E. coli (STEC), Salmonella, Campylobacter, and antibiotic resistant bacteria in cattle and swine. Approaches for reducing these organisms include reduction of colonization and shedding by livestock, as well as reduction of pathogens and antibiotic resistant bacteria present in the manure and production environment. Persistent shedding, shedding in high numbers, and environmental persistence have been identified as important reasons for the prevalence and maintenance of zoonotic pathogens in livestock, and therefore are intervention targets to reduce these organisms. This project will focus on determining microbial, host, and environmental factors that contribute to these persistence mechanisms, and identify potential strategies for reduction of persistence. Furthermore, this project will develop molecular approaches to assess antibiotic resistance genes in cattle and swine production, and utilize these strategies to evaluate their distribution and abundance. In addition, understanding the potential sources and transmission dynamics of pathogens in livestock production environments is critical for identifying management strategies to reduce their introduction and dissemination. Expected outcomes are scientific information and procedures that will be used to reduce or eliminate foodborne pathogens and antibiotic resistance in livestock production, thus contributing to a safer food and water supply and a lower risk of human foodborne illness.

Progress Report
Reducing pathogens in livestock and their production environment will require the identification of genotypic and/or phenotypic factors that are associated with their occurrence, levels, or persistence (Objective 1). In 2017, we continued work to extract and sequence bacterial DNA from the bovine fecal sample repository, for the identification of bacterial populations that are associated with E. coli O157:H7 colonization and shedding. Bioinformatic pipelines have been developed and validated for these sequence analyses. Also in 2017, experiments were initiated to phylotype E. coli in bovine feces, as a means to determine if particular phylotypes of commensal E. coli are associated with the propensity for an animal to shed E. coli O157:H7. Studies were continued to determine if there are associations between bovine host genotypes and chronic Salmonella colonization of cattle. Experiments were approved and sampling was initiated in swine experiments that seek to determine if host genotypes and performance influence the shedding of antibiotic resistant bacteria and zoonotic pathogens. In 2017, studies were initiated to assess the effects of environmental and other factors on the occurrence and transmission of pathogens and antibiotic resistance in water and cattle in pasture-based cattle production. Also, a study was initiated to determine the immediate and long-term effects of flash grazing of cattle on the microbiological water quality of a riparian stream. Reducing pathogen and antibiotic resistance persistence and transmission from host animals will require the development of intervention strategies that reduce their prevalence and persistence in manure and the production environment (Objective 2). Assessing intervention effects on antibiotic resistance genes will require the development of novel approaches to assay those genes that are of concern to both animal and human health. In 2017, research was continued to develop a high-throughput sequence-based assay for detecting and quantifying the broad spectrum of bacterial antibiotic resistance genes. The genes to be targeted have been compiled into databases and primer sets have been identified and validated. Also in 2017, we initiated studies to compare conventionally-raised cattle to cattle that have never been treated with antibiotics, with the objective to discern the contribution of conventional feedlot practices to the development, transmission, and persistence of antibiotic resistance. Studies that examined the use of Lactobacillus fermentation extracts as alternatives to antibiotic growth promotants in swine were completed in 2017. In addition, we initiated a collaboration with scientists at Kansas State University and the University of Otago in New Zealand, to better understand the ecology of Fusobacterium in feedlot cattle. Fusobacterium is a bacterial pathogen that causes liver abscesses, and reducing this microorganism is the main reason for feeding the macrolide antibiotic tylosin to beef cattle. Developing alternative antimicrobials targeted to control Fusobacterium in the rumen of cattle is needed to reduce dietary antibiotic use and the potential for antibiotic resistance development. A separate project examined the use of limonene as a potential alternative to tylosin, by testing its ability to reduce liver abscesses, as well as its ability to reduce E. coli O157:H7. In addition, we began planning with collaborators to identify indigenous ruminal flora that are associated with cattle production efficiency, with the intent to identify potential probiotic microorganisms that can function as alternatives to antimicrobial use.

1. Beta-agonists do not impact E. coli O157:H7 fecal shedding in cattle. Beta-agonists are fed to cattle to increase red meat production, and previous research had proposed that these compounds reduced the fecal shedding of E. coli O157:H7 from cattle. ARS scientists at Clay Center, Nebraska conducted a three-year study in which E. coli O157:H7 in bovine feces was evaluated using both enrichment (presence/absence) and enumeration testing. No significant effect of feeding either of beta-agonists zilpaterol or ractopamine on E. coli O157:H7 fecal shedding was observed. Beta-agonists had no impact on the shedding of E. coli O157:H7 in cattle at harvest, and therefore do not improve the microbial safety of beef.

2. Source of dietary protein affects fecal shedding of E. coli O157:H7 in cattle. Cattle diets are supplemented with crude protein sources to improve diet quality and animal growth. Previous studies determined that cattle diets with high levels of distillers grains led to increased fecal shedding of E. coli O157:H7, but low levels did not have any effect. ARS scientists in Clay Center, Nebraska, conducted a series of studies to determine if increased shedding was due to high protein or to distillers grains in the diet. Supplementation of even moderate levels of wet distillers grain with solubles increased E. coli O157:H7 shedding from cattle, whereas cattle fed diets supplemented with soybean meal appeared to have reduced shedding for E. coli O157:H7. Dietary protein level does not appear to be a factor in fecal shedding of E. coli O157:H7, but supplementation with soybean meal may be beneficial to reducing fecal shedding of this pathogen by cattle.

3. Feeding monensin at high levels affects fecal shedding of E. coli O157:H7 in cattle. Monensin is fed to cattle to improve animal growth and feed efficiency, and to reduce methane production from the rumen. Previous research has reported conflicting results regarding the effects of feeding monensin to cattle on E. coli O157:H7 fecal shedding. ARS scientists at Clay Center, Nebraska conducted a series of studies over two years to evaluate the potential effects of feeding different levels of monensin on fecal shedding of E. coli O157:H7. Dietary monensin did not appear to impact fecal presence or absence for E. coli O157:H7 in the studies. However, feeding monensin at the highest allowable level increased the percentage of animals that were shedding E. coli O157:H7 at high levels. This research indicates that feeding ionophores like monensin is not inhibitory to E. coli O157:H7 in the bovine gut, but at high levels may alter colonization and shedding in cattle.

4. E. coli O157:H7 on feedlot pen surfaces of cattle fed distillers grains. Previous studies have shown that feeding wet distillers grains with solubles (WDGS) to cattle can increase the load of the foodborne pathogen E. coli O157:H7 in feces and on hides, but the reasons are not fully understood. ARS scientists in Clay Center, Nebraska, conducted research to determine if environmental persistence contributes to increasing E. coli O157:H7 in cattle fed WDGS. The survival of E. coli O157:H7 on feedlot pen surfaces of cattle that were fed dry-rolled corn-based diets with 0 and 40% WDGS was determined. Greater persistence of E. coli O157:H7 on the pen surfaces of animals fed 40% WDGS was not demonstrated, however these pens had higher prevalence of E. coli O157:H7 in feedlot surface manure after cattle were removed. Either higher environmental numbers or greater environmental persistence can affect the potential risk of transmission and maintenance of this pathogen in cattle production. Hence, attention to factors affecting both shedding and environmental contamination may be needed to reduce E. coli O157:H7 in cattle, and to reduce the risk of human foodborne illness associated with beef consumption.

Review Publications
Berry, E.D., Wells, J. 2016. Reducing foodborne pathogen persistence and transmission in animal production environments: Challenges and Opportunities. Microbiology Spectrum. 4(4):1-18. doi:10.1128/microbiolspec.PFS-0006-2014.
Wells, J., Berry, E.D. 2017. Pathogens affecting beef. In: Acuff, G.R., Dickson, J.S., editors. Ensuring safety and quality in the production of beef Volume 1, Cambridge, UK, Burleigh Dodds Science Publishing. p. 1-16.