Listeria (L.) monocytogenes is an opportunistic pathogen causing life-threatening infections in diverse mammalian species including humans and ruminants. As little is known on the link between strains and clinicopathological phenotypes, we studied potential strain-associated virulence and organ tropism in L. monocytogenes isolates from well-defined ruminant cases of clinical infections and the farm environment. The phylogeny of isolates and their virulence-associated genes were analyzed by multilocus sequence typing (MLST) and sequence analysis of virulence-associated genes. Additionally, a panel of representative isolates was subjected to in vitro infection assays. Our data suggest the environmental exposure of ruminants to a broad range of strains and yet the strong association of sequence type (ST) 1 from clonal complex (CC) 1 with rhombencephalitis, suggesting increased neurotropism of ST1 in ruminants, which is possibly related to its hypervirulence. This study emphasizes the importance of considering clonal background of L. monocytogenes isolates in surveillance, epidemiological investigation and disease control. Listeria (L.) monocytogenes is an opportunistic pathogen that may cause life-threatening infections in many mam-malian species upon ingestion 1,2. Listeriosis is of major importance in humans and in farmed ruminants 3 , in which it is associated with gastroenteritis, abortions, bacteremia, mastitis and central nervous system (CNS) infections (neurolisteriosis) 4-7. In humans, L. monocytogenes has the highest hospitalization and mortality rates amongst food-borne pathogens 8,9. Surveillance and control prove to be challenging due to the environmental lifestyle of L. monocytogenes, which colonizes diverse environments including soil, water, food processing plants, mammalian intestinal tracts and faeces 8,10-12. In consequence, L. monocytogenes is frequently found as a contaminant of human food and animal feed. L. monocytogenes is divided into 13 serotypes 13 and four phylogenetic lineages 14-17. Regulations consider all L. monocytogenes strains an equally serious threat for public health 18 , although an increasing number of studies indicate differences in ecology and virulence between L. monocytogenes strains 7,16,19-22. Most lineage I strains are overrepresented in human clinical infections and in ruminant neurolisteriosis, while a majority of lineage II strains are associated with food contaminations and the environment 7,14,23,24. The minor lineages III and IV are rarely isolated and have been linked to animals 14. Multilocus sequence typing (MLST), which categorizes isolates into sequence types (STs) and clonal complexes (CCs, hereafter equated to clones) based on sequence data of