Abstract Foodborne diseases represent a link between environmental, animal, and human health interfaces from the One Health perspective. Whole genome sequencing (WGS) is becoming the gold standard in foodborne surveillance, worldwide. WGS than provides precision data from pathogens allows laboratories to resolve the genetic relations among all sources from One Health perspective, especially during outbreak investigations, getting insights into their transmission routes and pathogenicity. Salmonella spp. is the most prevalent foodborne bacteria in Colombia, in 2020, 268 foodborne outbreaks were reported to National health institute (INS) and 3079 invasive and faecal salmonellosis samples from 81% of the Colombian geographical regions. Several studies in different food sources exhibited an increase of contamination with Salmonella , a public health concern due to the steady development of antimicrobial resistance associated to specific serovars. However, integration of Salmonella spp . data including food-chain supply and clinical interfaces is very scarce in Colombia. This study carried out a national comparison of Salmonella isolates collected from food-chain supply and clinical sources. Using an epidemiological and phenotypic approach, we demonstrated the higher resolution of WGS compared with PFGE, routinely used in Salmonella surveillance in Colombia. For example, the resolution of PFGE allowed the description of two main clusters of food Salmonella Enteritidis isolates which were expanded to eight clades by using WGS. Virulence factors and antimicrobial determinant genes observed in the foodborne clades should be considered a public health concern in Colombia. WGS is a technology that provides precise and valid evidence for the establishment of dissemination routes of foodborne high-risk Salmonella clades, but it requires an integrative and continued collaboration between the stakeholders across the One Health interfaces to promote and support integrated real-time actions in public health.