Survival of Salmonella enterica serovar Heidelberg in pine shavings used as broiler litter

Wood shavings is the most common bedding material used around the world to raise broiler. Therefore, wood shavings can be a vehicle for the transmission of pathogens to live birds. In this study, we performed an in-depth genomic characterization of three Salmonella enterica serovar Heidelberg (S. Heidelberg) strains recovered after their inoculation into fresh pine shaving. The three strains used for the microcosm study were previously isolated from broiler feces (SH-AAFC), broiler carcass (SH-ARS) and chicken thigh (SH-FSIS) and differed one from another by 46 - 94 single nucleotide variants. The SH-AAFC strain harbored an antimicrobial resistant gene (ARG) (bla_CMY-2) on an IncI1 plasmid while the SH-FSIS strain harbored multiple ARGs (floR, cmlA1, tet(A), bla_TEM-1B, ant(2'')-Ia, aph(6)-Id, aph(3'')-Ib and sul2) on an IncC plasmid. The SH-ARS isolate was pan susceptible to several antibiotics evaluated. We determined the abundance of Salmonella at days 0, 1, 7, 14 and 21 and performed antibiotic susceptibility testing and whole genome sequencing (on 77 randomly selected S. Heidelberg isolates. After 21 days of incubation, Salmonella abundance decreased by 4.4 logs. Salmonella with high minimum inhibitory concentrations (MICs) against ampicillin showed a significantly higher abundance and survival rate compared to Salmonella with high MIC against gentamicin (P< 0.05). Clonal SH-AAFC was the most prevalent strain in the microcosms (48/77), followed by the strain SH-ARS (25/77). Only 4/77 isolates were determined to be clones of SH-FSIS. We identified recombination events and plasmid copy number changes that were associated with the fitness of S. Heidelberg strain carrying IncI1 and Col plasmids. Lastly, we found that litter physicochemical variables including water activity could explain up to 85% of the variability in our data.