Large-scale genomic survey of multidrug-resistant Salmonella enterica serovar Enteritidis clones in a one health approach

The global burden of antimicrobial resistance (AMR) continues to rise, with Salmonella enterica serovar Enteritidis remaining a leading cause of foodborne infections worldwide. In the context of Saudi Arabia, the increasing reliance on imported food products, coupled with intensive poultry production, has created a favorable environment for the emergence and dissemination of multidrug-resistant (MDR) strains. This study presents a comprehensive genomic investigation of 188 S. Enteritidis isolates, comprising 93 clinical and 95 food-derived strains, collected over a two-year period as part of a national effort to assess AMR within a One Health framework. Whole-genome sequencing (WGS) was performed for all isolates, followed by in-depth bioinformatic analyses to characterize resistance determinants, virulence factors, plasmid content, and phylogenetic relationships. The isolates were distributed across several phylogenetic clusters, with three dominant groups identified based on core genome SNP analysis. Among these, Cluster 3 emerged as the most concerning in terms of both its resistance profile and virulence potential. This cluster was enriched in MDR determinants, including genes conferring resistance to aminoglycosides (e.g., aadA, aph), beta-lactams (notably bla_TEM-1, bla_SHV, and bla_CTX-M), and fluoroquinolones (qnrS1), in addition to efflux systems implicated in decreased susceptibility.

Plasmid analysis revealed that Cluster 3 isolates consistently harbored an IncFII(S)-type plasmid. This plasmid was variably associated with bla_CTX-M and qnrS genes and frequently carried composite mobile elements, suggesting its role as a key vector in the horizontal transfer of resistance traits. Furthermore, Cluster 3 isolates exhibited a conserved virulence gene repertoire, including factors encoded within Salmonella pathogenicity islands SPI-3 and SPI-4. Of particular note was the consistent presence of the spv operon (spvB, spvC, spvD), which is known to enhance intracellular survival and systemic dissemination in hosts. Additional virulence- associated elements such as pefBC, mig-5, and rck were also identified, supporting the hypothesis that these isolates possess enhanced pathogenic potential. The geographic distribution of Cluster 3 isolates indicated a strong association with Saudi Arabia, spanning both clinical and food-related contexts. Comparative analysis with international genomes revealed phylogenetic proximity to strains from neighboring countries, including Egypt and Lebanon, suggesting a broader regional lineage with transboundary transmission potential.

The close SNP distances observed between food and clinical isolates further support the zoonotic capacity of these strains and the likelihood of direct or indirect transmission along the food chain. Taken together, the findings highlight the emergence of a dominant MDR S. Enteritidis lineage in Saudi Arabia, characterized by the co-localization of clinically significant resistance genes and conserved virulence determinants on mobile genetic elements. The integration of resistance and virulence within a single clonal group represents a serious public health threat, particularly in light of the limited therapeutic options available for invasive salmonellosis. These results emphasize the importance of sustained genomic surveillance and coordinated One Health interventions to monitor the spread of high-risk clones and inform evidence-based policy decisions. Moreover, the study aligns with national health priorities and the broader goals of Saudi Vision 2030 in addressing AMR and enhancing food and health security through advanced molecular epidemiology.