Genetic and Epigenetic Diversity of Salmonella enterica Isolates from Clinical and Veterinary Sources in Kazakhstan

Salmonella enterica is a major cause of foodborne and invasive infections worldwide. This retrospective study used whole-genome sequencing to compare the genotypes and characterize the genomic diversity of bacterial strains preserved in an archived culture collection, originating from clinical and veterinary sources and predominantly from Kazakhstan. Twenty-two bacterial isolates were analyzed by Illumina whole-genome sequencing; subsequent characterization confirmed twenty as S. enterica and identified two as Escherichia coli. Six selected S. enterica strains representing different lineages were additionally sequenced using PacBio Revio SMRT technology for DNA methylation profiling. Genome assembly, plasmid reconstruction, MLST genotyping, and analyses of virulence genes, antimicrobial resistance determinants, restriction-modification (R-M) operons, and orphan methyltransferases were performed. Comparative genomic analysis revealed substantial genotypic diversity within the collection, while phylogenetic grouping was consistent with MLST classification. ST11 was the most represented sequence type among the selected strains, comprising 11 of the 20 S. enterica isolates, including all three recent clinical S. enterica isolates. All six PacBio-sequenced genomes showed >99% DAM-associated adenine methylation at GATC motifs, while DCM-mediated cytosine methylation occurred at approximately 35% of CCWGG motifs. Adenine methylation at CAGAG is controlled by additional type III R-M systems. Strain 19S showed a distinct type I-associated adenine methylation pattern together with several HsdM amino acid substitutions; however, the functional relationship between these observations requires experimental validation. The study demonstrates the value of retrospective genomic characterization of culture collections for revealing genetic and epigenetic diversity preserved among historical and recent bacterial isolates and provides a basis for future population-level and functional studies.