Distinct adaptation and epidemiological success of different genotypes within Salmonella enterica serovar Dublin

<i>Salmonella</i> Dublin is a host-adapted, invasive nontyphoidal <i>Salmonella</i> (iNTS) serovar that causes bloodstream infections in humans and demonstrates increasing prevalence of antimicrobial resistance (AMR). Using a global dataset of 1303 genomes, coupled with in vitro assays, we examined the evolutionary, resistance, and virulence characteristics of <i>S</i>. Dublin. Our analysis revealed strong geographical associations between AMR profiles and plasmid types, with highly resistant isolates confined predominantly to North America, linked to IncC plasmids co-encoding AMR and heavy metal resistance. By contrast, Australian isolates were largely antimicrobial-susceptible, reflecting differing AMR pressures. We identified two phylogenetically distinct Australian lineages, ST10 and ST74, with a small number of ST10 isolates harbouring a novel hybrid plasmid encoding both AMR and mercuric resistance. Whereas the ST10 lineage remains globally dominant, the ST74 lineage was less prevalent. ST74 exhibited unique genomic features including a larger pan genome compared to ST10 and the absence of key virulence loci, including <i>Salmonella</i> pathogenicity island (SPI)-19 which encodes a type VI secretion system (T6SS). Despite these genomic differences, the ST74 lineage displayed enhanced intracellular replication in human macrophages and induced less pro-inflammatory responses compared with ST10, suggesting alternative virulence strategies that may support systemic dissemination of ST74. The Vi antigen was absent in all ST10 and ST74 genomes, highlighting challenges for serotyping and vaccine development, and has implications for current diagnostic and control strategies for <i>S.</i> Dublin infections. Collectively, this study represents the most comprehensive investigation of <i>S</i>. Dublin to date and, importantly, has revealed distinct adaptations of two genotypes within the same serovar, leading to different epidemiological success. The regional emergence and evolution of distinct <i>S.</i> Dublin lineages highlight the need to understand the divergence of intra-serovar virulence mechanisms which may impact the development of effective control measures against this important global pathogen.