Assessment of plasmids for relating the 2020 Salmonella enterica serovar Newport onion outbreak to farms implicated by the outbreak investigation

Abstract

Abstract Background The Salmonella enterica serovar Newport red onion outbreak of 2020 was the largest foodborne outbreak of Salmonella in over a decade. The epidemiological investigation suggested two farms as the likely source of contamination. However, single nucleotide polymorphism (SNP) analysis of the whole genome sequencing data did not find any Salmonella isolates from the farm regions that were closely related to the clinical isolates—preventing the use of phylogenetics in source identification. Here, we explored an alternative method for analyzing the whole genome sequencing data driven by the hypothesis that if the outbreak strain had come from the farm regions, then the clinical isolates would disproportionately contain plasmids found in isolates from the farm regions due to recent horizontal transfer. Results SNP analysis confirmed that the clinical isolates formed a highly related clade with evidence for ancestry in California going back a decade. The clinical isolates not only had a large and highly conserved core genome (4,399 genes), but also 2,577 sparsely distributed accessory genes—at least 64% of which were carried on plasmids. Amongst the clinical isolates and Salmonella isolates from the farm regions were 2,187 and 503 putative plasmids, respectively. High similarity was observed between 17 plasmids from 8 farm isolates and 14 plasmids from 13 clinical isolates. Phylogenetic analysis suggested the highly similar plasmids shared a recent common ancestor and might have been transferred via intermediary species, but the seeming promiscuity of the plasmids prevented any conclusions about geographic location, isolation source, and time since transfer. Our sampling analysis suggested that observing a similar number and combination of highly similar plasmids in random samples of environmental Salmonella enterica within NCBI Pathogen Detection database was unlikely, supporting a connection between the outbreak strain and the farms implicated by the epidemiological investigation. Conclusion Horizontally transferred plasmids provided evidence for a connection between clinical isolates and the farms implicated as the source of the outbreak. Our case study suggests that such analyses might add a new dimension to source tracking investigations, but highlights the need for detailed and accurate metadata, more extensive environmental sampling, and a better understanding of plasmid molecular evolution.