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Research Project: Monitoring and Molecular Characterization of Antimicrobial Resistance in Foodborne Bacteria


2016 Annual Report

1. Provide data and characterize pathogen prevalence, unique characteristics and trends on antibiotic resistance, subtyping and molecular characterization of foodborne pathogens in food animals. 2. Identify and characterize potential genetic markers within and across serotypes for Salmonella isolated from poultry for rapid identification and diagnosis. 3. Evaluate the role of innovative chemical and/or biological treatments including arsenicals, prebiotics, or ammonium compounds and how they impact the prevalence and type of antimicrobial resistant pathogens or resistance genes.

Under current funding, this research is designed to be conducted by a team of two scientists. Because objectives for this project are non-hypothesis driven, specific goals have been established for each objective. Data from this research will be used to assist other Federal agencies in assessing antimicrobial resistance in food animal populations as well as to address a direct need outlined by the National Action Plan for Combatting Antibiotic Resistant Bacteria (CARB) in evaluating potential alternatives to antimicrobials. Data generated on biocide resistance and resistance genes active against chemicals specific to poultry production and processing is a specific concern to USDA-FSIS. Development of technologies for detection of microbial contaminants is a critical need for Federal regulatory agencies.

Progress Report
This project is new commencing in April 2016 replacing 6040-32000-006-00D. This project focuses on characterization of Salmonella, Escherichia coli, and Enterococcus from poultry to provide data on the prevalence of antimicrobial resistance and associated phenotypic and genotypic characteristics. Antimicrobial susceptibility, genetic mechanisms of resistance, vehicles for the dissemination of resistance genes (e.g. plasmids, transposons, integrons) in those isolates will be determined. To date, the genomes of 194 multidrug resistant Salmonella from food animals and food production facilities have been sequenced to identify resistance genes for antimicrobials, metals, and biocides. A total of 913 antimicrobial resistance genes from nine different antibiotic classes were detected; some, such as the macrolide class, have been rarely reported in Salmonella. Metal and biocide resistance genes such as copper, zinc, silver, and arsenic were also detected in most isolates. These preliminary results suggest that the selection or maintenance of resistance in Salmonella isolated from food animal sources could be due to exposure to metals in addition to antimicrobials. Draft genome analysis of rare Salmonella serotypes Kiambu, Lille, and Widemarsh were completed. Plasmid analysis of 164 multidrug resistant enterococci were also completed. Using plasmid replicon typing multiplex PCR, E. faecalis and E. faecium were found to be positive for at least one plasmid replicon family and up to 14 different plasmids were found among the isolates. Antimicrobial susceptible and single-resistance isolates were also positive for plasmids indicating that some plasmids may contain few or no antimicrobial resistance genes. Multidrug resistant Salmonella sequenced above were also evaluated for resistance to non-antimicrobial chemicals/solutions (biocides) used in poultry production or processing. Most of the isolates showed similar susceptibilities to each chemical with minimum inhibitory concentrations within +/- one two-fold dilution from the median minimum inhibitory concentration. Variable minimum inhibitory concentrations were found for arsenate and arsenite which may indicate reduced susceptibility or hyper susceptibility in some Salmonella isolates to these compounds. Preliminary results suggest that there is no relationship between antibiotic resistance pattern and susceptibility to biocides. The susceptibility of the isolates to different biocides was also not related to the serotype of Salmonella. Defining the role of surface water as a reservoir and a potential vehicle for the development of resistant bacteria and transmission to humans or food animals is important to both food safety and public health. Seasonal monitoring of Salmonella, E. coli and Enterococcus was conducted by collecting water samples through a partnership with the Upper Oconee Watershed Network at 100 different locations along the Oconee River. Multidrug resistant Salmonella were isolated, some of which were the same serotypes isolated from humans. Few multidrug resistant E. coli were detected; however, of the E. coli isolated, one-third belonged to a potentially pathogenic group and both enteropathogenic and Shiga toxin genes were detected in some of the isolates. Various enterococcal species were detected and almost all the isolates were intermediate or resistant to more than two drugs. Widespread occurrence of Salmonella in the surface water suggests that water can serve as a reservoir for Salmonella and may play a role in its transmission.

1. Few antimicrobial resistance genes present in rare Salmonella serotypes. Salmonella enterica is an important foodborne pathogen that causes gastroenteritis and enteric fever in humans. The more than two thousand serovars of Salmonella differ greatly in their ability to cause disease in humans. To characterize antimicrobial resistance in rare Salmonella serotypes, ARS researchers in Athens, Georgia, have sequenced the genomes of Salmonella serovar Lille, Kiambu, and Widemarsh. Collectively, draft genomes contained few antimicrobial resistance genes, but genes conferring resistance to arsenic were detected which may be important in survival of the bacteria in the environment. Addition of draft genomes of rare serovars increases the diversity of genomes currently available for comparative analysis to better understand virulence as well as antimicrobial resistance in Salmonella.

Provided research experience for an African-American male student from Clarke Central High School participating in the Boule Program. Program is designed to increase the number of underrepresented males pursuing college degrees in Science, Technology, Engineering, and Mathematics. Supervised 2 female University of Georgia Ph.D. level students (including 1 student of Asian descent). Provided research experience for an Asian-American female undergraduate student from the University of Georgia. The research experience will aid her in learning key skills necessary to succeed in the medical and bioscience fields.

Review Publications
Gupta, S., Williams, L., McMillan, E., Jackson, C.R., Desai, P., Porwollik, S., McClelland, M., Hiott, L.M., Humayoun, S.B., Frye, J.G. 2016. Draft genome sequence of Salmonella enterica subsp. enterica serovar Kiambu Strain CRJJGF_00061 (Phylum Gammaproteobacteria). Genome Announcements. 4(4):e00588-16.
Gupta, S., McMillan, E., Jackson, C.R., Desai, P., Porwollik, S., McClelland, M., Hiott, L.M., Humayoun, S.B., Frye, J.G. 2016. Draft genome sequence of Salmonella enterica subsp. enterica serovar Widemarsh Strain CRJJGF_00058 (Phylum Gammaproteobacteria). Genome Announcements. 4(4):e00604-16.
Gupta, S., McMillan, E., Jackson, C.R., Desai, P., Porwollik, S., McClelland, M., Hiott, L.M., Humayoun, S.B., Frye, J.G. 2016. Draft genome sequence of Salmonella enterica subsp. enterica serovar Lille Strain CRJJGF_000101 (Phylum Gammaproteobacteria). Genome Announcements. 4(4):e00603-16.