ABSTRACTWhole-genome sequencing (WGS) has revolutionized surveillance of infectious diseases. Disease outbreaks can now be detected with high precision, and correct attribution of infection sources has been improved. Listeriosis, caused by the bacteriumListeria monocytogenes, is a foodborne disease with a high case fatality rate and a large proportion of outbreak-related cases. Timely recognition of listeriosis outbreaks and precise allocation of food sources are important to prevent further infections and to promote public health. We report the WGS-based identification of a large multinational listeriosis outbreak with 55 cases that affected Germany, Austria, Denmark, and Switzerland during 2020 and 2021. Clinical isolates formed a highly clonal cluster (called Ny9) based on core genome multilocus sequence typing (cgMLST). Routine andad hocinvestigations of food samples identifiedL. monocytogenesisolates from smoked rainbow trout filets from a Danish producer grouping with the Ny9 cluster. Patient interviews confirmed consumption of rainbow trout as the most likely infection source. The Ny9 cluster was caused by a MLST sequence type (ST) ST394 clone belonging to molecular serogroup IIa, forming a distinct clade within molecular serogroup IIa strains. Analysis of the Ny9 genome revealedclpY,dgcB, andrecQinactivating mutations, but phenotypic characterization of several virulence-associated traits of a representative Ny9 isolate showed that the outbreak strain had the same pathogenic potential as other serogroup IIa strains. Our report demonstrates that international food trade can cause multicountry outbreaks that necessitate cross-border outbreak collaboration. It also corroborates the relevance of ready-to-eat smoked fish products as causes for listeriosis.IMPORTANCEListeriosis is a severe infectious disease in humans and characterized by an exceptionally high case fatality rate. The disease is transmitted through consumption of food contaminated by the bacteriumListeria monocytogenes. Outbreaks of listeriosis often occur but can be recognized and stopped through implementation of whole-genome sequencing-based pathogen surveillance systems. We here describe the detection and management of a large listeriosis outbreak in Germany and three neighboring countries. This outbreak was caused by rainbow trout filet, which was contaminated by aL. monocytogenesclone belonging to sequence type ST394. This work further expands our knowledge on the genetic diversity and transmission routes of an important foodborne pathogen.