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Background Listeria monocytogenes is a foodborne pathogen with low incidence but severe manifestations in at risk categories (up to 30% fatality rate in YOPI). L. monocytogenes can form biofilm and persist in food processing environments for decades. In the Lombardy Region up to 50% of listeriosis cases have been attributed to dairy products. This study aimed at investigating a L. monocytogenes clone widely present in one of the most important Italian cheese production chain. Methods Through a retrospective analysis of the Lombardy Region PFGE database a cluster comprising isolates from eight PDO Gorgonzola and Taleggio dairies (2013-2019) was identified. A representative subset of 33 isolates was typed with MLVA and MLST. All isolates were further typed with WGS and compared to all Lombardy clinical isolates (n = 9) that shared the same Sequence Type (ST). Genomic relationship among isolates were investigated with cgMLST and SNP analysis. Results All isolates had the same MVLA/MLST profile (ST325). cgMLST and SNP analysis divided the isolates in two clusters that however shared a very high similarity (99.4%). Interestingly, isolates collected from same production plants clustered separately. Conversely, isolates collected in different years and plants clustered together. By querying the L. monocytogenes MLVA database it was possible to identify isolates of the same production chain with identical profile back to the year 2005. ST325 clinical isolates represented 2% of all typed clinical isolates and have an incidence of 0.007 cases/1 inhabitants. Due to the high genomic similarity, it was not possible to link any clinical isolate to a putative source. Conclusions Our data suggest a persistent contamination by a L. monocytogenes ST325 clone traced back to the year 2005 present in the dairy production chain of Lombardy. Given the high similarity among the isolates further analyses with a larger dataset and are needed to elucidate the epidemiology of this clone. Key messages A highly clonal contamination by L. monocytogenes is widespread in the Lombardy Region dairy production chain. The low incidence of ST325 among the population suggests a low pathogenicity of the strain, however the occurrence of an ongoing long-term outbreak cannot be excluded.

A Listeria monocytogenes ST325 clone is widespread in the Lombardy Region dairy processing plants