Application of a Quantitative Real-Time PCR Assay for Early Detection of Salmonella enterica Serovar Enteritidis on Poultry Farms During an Outbreak in New South Wales, Australia (2018-2020).

Salmonella spp. are a significant cause of human foodborne illness globally, with ingestion of contaminated eggs a major vehicle for infection. Salmonella enterica serovar Enteritidis (S. Enteritidis, SE) is the serovar most linked to egg-related foodborne salmonellosis in most developed countries. Until 2018, the Australian egg industry was considered free of SE. This report documents the diagnostic testing performed on samples from egg layer farms across New South Wales (NSW), Australia, as part of a SE outbreak response between 2018 and 2020. Testing was undertaken following a cluster of cases of SE infection in humans traced to the consumption of eggs originating from a single contaminated poultry farm. Quantitative real-time polymerase chain reaction (qPCR) testing was used to screen environmental and animal samples (n = 2058) from 29 different properties identified through contact tracing. Confirmatory bacterial culture (n = 717) was performed on any SE qPCR-positive samples and a subset of qPCR-negative and qPCR-inconclusive samples. In total, 13/29 (45%) of egg layer farms were SE-positive by qPCR testing, with 12/13 (92%) of these farms confirmed SE-positive by bacterial culture and serotyping. Both environmental and animal samples produced SE-positive results, in particular surface swabs, boot covers, feces, and eggs. When qPCR testing and bacterial culture were performed side-by-side, qPCR testing to detect SE compared to bacterial culture had sensitivity of 100% (43/43) and specificity of 94.1% (238/253; 95% confidence interval[CI] 91.4-96.8). SE isolates obtained during the outbreak were predominantly phage type (PT)1b and PT12. Whole genome sequencing (WGS) of SE isolates from 9 of 12 culture-positive properties confirmed that they were all sequence type 11, Clade B, and derived from a single source. As a result of rapid qPCR detection of SE on contaminated farms, appropriate biosecurity responses were implemented, and NSW commercial layer farms were again considered SE-free in August 2020. This report highlights the utility of high-throughput molecular testing for SE in outbreak situations.