Management factors influence Salmonella persistence in reused poultry litter over three successive flocks

Salmonella infections are a leading cause of bacterial food-borne illness worldwide. Infections are highly associated with the consumption of contaminated food, and in particular, chicken meat. Understanding how management practices and environmental factors influence Salmonella populations in broiler chicken production may aid in reducing the risk of food-borne illness in humans. Utilizing whole genome sequencing with antimicrobial and heavy metal resistance, virulence factor and plasmid identification, we have characterized the genetic diversity of Salmonella enterica isolates (n = 55) obtained from broiler chicken litter. S. enterica isolates were recovered from the litter of broiler chickens over three consecutive flocks in four broiler houses on a single integrated farm in Georgia, USA. The chickens were raised under a newly adopted "No Antibiotics Ever" program and copper sulfate was administered via drinking water. In-silico serovar prediction identified three S. enterica serovars: Enteritidis (n = 12), Kentucky (n = 40) and Senftenberg (n = 3). Antimicrobial susceptibility testing revealed that only one S. Kentucky isolate was resistant to streptomycin, while the remaining isolates were susceptible to all antibiotics tested. Metal resistance operons, including copper and silver, were identified chromosomally and on plasmids in serovar Senftenberg and Kentucky isolates, respectively. Serovar Kentucky isolates harboring metal resistance operons were the only Salmonella isolates recovered from the litter of third flock cohort. These results suggest the addition of copper sulfate to drinking water may have selected for S. Kentucky isolates harboring plasmid-borne copper resistance genes and may explain their persistence in litter from flock to flock. ImportanceSalmonella foodborne illnesses are the leading cause of hospitalizations and deaths, resulting in a high economic burden on the healthcare system. Globally, chicken meat is one of the highest consumed meats and is a predominant source of foodborne illness. The severity of Salmonella infections depends on the presence of antimicrobial resistance genes and virulence factors. While there are many studies which have investigated Salmonella strains isolated from post-harvest chicken samples, there is a gap in our understanding of the prevalence and persistence of Salmonella in pre-harvest and in particular their makeup of antibiotic resistance genes, virulence factors and metal resistance genes. The objective of this study was to determine how on-farm management practices and environmental factors influence Salmonella persistence, as well as the antimicrobial resistance genes and virulence factors they harbor. In this study we demonstrate that broiler chickens raised without antibiotics are less likely to harbor antibiotic resistance, however the practice of adding acidified copper sulfate to drinking water may select for strains carrying metal resistant genes.