A proof of principle of a multi-target assay for genotyping Salmonella has been developed targeting 62 genomic marker sequences of Salmonella related to pathogenicity. The assay is based on multiplex ligation detection reaction (LDR) followed by customized ArrayTube® microarray detection. The feasibility of the developed assay was verified in a method comparison study with conventional PCR using 16 Salmonella ‘test’ strains comprising eight serovars. Subsequently, the feasibility of the LDR microarray assay was also tested by analyzing 41 strains belonging to 23 serovars. With the exception of four serovars each serovar was characterized by a unique virulence associated gene repertoire. The LDR microarray platform proved to be a convenient, rapid and easy to use tool with potential in tracing a Salmonella contamination in the food chain, for outbreak studies, and to provide data for risk assessors that support bio-traceability models.