Population Genomics of Salmonella Enteritidis in Saudi Arabia Reveals Globally Circulating Food- and Human-Associated Lineages and Plasmid-Mediated Antimicrobial Resistance

Background: Salmonella enterica serovar Enteritidis (S. Enteritidis) is one of the leading causes of foodborne gastroenteritis worldwide and an increasingly important driver of antimicrobial-resistant infections. Despite its public health importance, genomic surveillance of S. Enteritidis remains sparse across the Middle East, where integrated One Health studies linking human, food, and animal reservoirs are limited. This lack of regional genomic data has hindered understanding of the population structure, transmission dynamics, and dissemination of antimicrobial-resistant lineages. We therefore performed a large-scale One Health genomic investigation of S. Enteritidis in the Kingdom of Saudi Arabia, integrating human, food, poultry, and global genomic datasets to characterize its evolutionary history, transmission dynamics, and antimicrobial resistance. Methods: A total of 220 Salmonella enterica serovar Enteritidis (S. Enteritidis) isolates obtained in Saudi Arabia from human clinical infections and food products between 2020 and 2023 underwent whole-genome sequencing. To position these isolates within a global evolutionary context, they were integrated with all publicly available S. Enteritidis genomes, yielding a comparative dataset of 397 genomes. A comprehensive genomic analysis framework was employed, encompassing population genomic, phylogenetic, and phylodynamic analyses to define population structure, infer evolutionary history, and investigate transmission dynamics. In addition, the genomic determinants of antimicrobial resistance and virulence were characterized, with particular emphasis on plasmid architecture. Results: Isolates from Saudi Arabia belonged predominantly to a single globally distributed epidemiological cluster of S. Enteritidis and were embedded within globally circulating transmission networks rather than forming a distinct endemic population. Within this cluster, the Saudi isolates were distributed across seven Bayesian Analysis of Population Structure (BAPS) groups, representing distinct genomic sub-clones. Multiple lineages contained closely related isolates from Europe, North America, Oceania, and the Middle East, indicating repeated international introductions and extensive global connectivity. Human and food isolates were highly intermixed throughout the phylogeny, with food-associated isolates frequently occupying central positions within transmission networks, consistent with food reservoirs acting as dissemination hubs linking local and international transmission. Phylodynamic analyses indicated that the dominant Saudi-associated lineages emerged within the past 10-15 years and underwent rapid demographic expansion following introduction into the country. Recently emerged epidemic lineages carried significantly higher antimicrobial resistance burdens than ancestral populations and were characterized by near fixation of blaTEM-1 and tetA on IncX1 plasmids, whereas clinically important ESBL genes (blaCTX-M-8 and blaSHV-12) were detected exclusively in human-associated isolates on hybrid resistance-virulence plasmids. Conclusions: The population of S. Enteritidis in Saudi Arabia is shaped by repeated introductions of globally circulating clones, transmission between food and human reservoirs, and the dissemination of resistance plasmids across lineages. The emergence of successful epidemic clones is linked to plasmid-mediated acquisition of antimicrobial resistance determinants. These findings emphasize the importance of integrated One Health surveillance strategies for controlling foodborne transmission and monitoring the emergence of antimicrobial-resistant epidemic lineages.