Listeria monocytogenes is a widespread Gram-positive pathogenic bacterium that causes listeriosis, a rather rare but severe foodborne disease. Pregnant women, infants, the elderly, and immunocompromised individuals are considered particularly at risk. L. monocytogenes can contaminate food and food-processing environments. In particular, ready-to-eat (RTE) products are the most common source associated with listeriosis. L. monocytogenes virulence factors include internalin A (InlA), a surface protein known to facilitate bacterial uptake by human intestinal epithelial cells that express the E-cadherin receptor. Previous studies have demonstrated that the presence of premature stop codon (PMSC) mutations naturally occurring in inlA lead to the production of a truncated protein correlated with attenuate virulence. In this study, 849 L. monocytogenes isolates, collected from food, food-processing plants, and clinical cases in Italy, were typed and analyzed for the presence of PMSCs in the inlA gene using Sanger sequencing or whole-genome sequencing (WGS). PMSC mutations were found in 27% of the isolates, predominantly in those belonging to hypovirulent clones (ST9 and ST121). The presence of inlA PMSC mutations in food and environmental isolates was higher than that in clinical isolates. The results reveal the distribution of the virulence potential of L. monocytogenes circulating in Italy and could help to improve risk assessment approaches.

Listeria monocytogenes is a major human foodborne pathogen responsible for listeriosis. The virulence factor Internalin A (inlA) has a key role in the invasion of L. monocytogenes into the human intestinal epithelium, and the presence of premature stop-codons (PMSC) mutations in the inlA gene sequence is correlated with attenuated virulence. The inlA sequencing process is carried out by dividing the gene into three sections which are then reassembled to obtain the full gene. The primers available however were only able to entirely amplify the lineage II isolates. In this study, we present a set of new primers which allow inlA sequencing of isolates belonging to both lineages, since lineage I isolates are the ones most frequently associated to clinical cases. Using newly designed primers, we assessed the presence of inlA PMSCs in food, food processing environments and clinical isolates.

Over the last few decades, several pestiviruses have been discovered in ruminants, pigs, and, more recently, in non-ungulate hosts. Consequently, the nomenclature and taxonomy of pestiviruses have been updated. The Tunisian sheep-like pestivirus (TSV, Pestivirus N) is an additional ovine pestivirus genetically closely related to classical swine fever virus (CSFV). In this study, during a survey of pestivirus infections in ovine farms in the Lombardy region of Northern Italy, we identified and isolated a pestivirus strain from a sheep that was found to belong to Pestivirus N species based on its genomic nucleotide identity. The sheep itself and its lamb were found to be persistently infected. We performed molecular characterization and phylogenetic analysis of three viral genomic regions (a fragment of 5′-UTR, partial N^pro, and the whole E2 region). In conclusion, these results confirmed circulating TSV in Northern Italy after notification in Sicily, Italy, and France. Correlation with Italian, Tunisian, and French strains showed that detection might have resulted from the trading of live animals between countries, which supports the need for health control measures.

Foodborne botulism is a potentially fatal disease caused by Clostridium botulinum neurotoxins (BoNTs). These spore-forming bacteria are ubiquitous in the environment and can contaminate various food products, especially raw vegetables. During the preparation of home-made preserves, favorable conditions of anaerobiosis, temperature, salinity, and pH can lead to spore germination and toxin production. BoNTs can reach neuromuscular junctions where they block the release of acetylcholine. In this study, we present a case of foodborne botulism associated with the consumption of chili peppers preserve containing BoNT/B. The isolated strains were characterized through Whole Genome Sequencing, confirming the strains involved in the outbreak. This work increases the understanding of the epidemiology and the ecology of C. botulinum, highlighting the importance of raising medical awareness and making timely clinical diagnoses for the effective management of botulism outbreaks.

Background: Listeria monocytogenes is a widespread foodborne pathogen that causes listeriosis, a human infection with low morbidity but high mortality. Internalin A (InlA), a L. monocytogenes virulence factor, has a key role in the invasion of human intestinal epithelium through the interaction with E-cadherin receptor. Truncated InlA caused by premature stop-codons (PMSC) have been correlated with attenuated virulence. To date, 29 mutations leading to a PMSC have been reported. In this study, we identified a rare PMSC mutation widespread in isolates from the production chain of the Gorgonzola and Taleggio PDO cheeses. Methods: Within an ongoing monitoring plan for the characterization of L. monocytogenes, a total of 81 isolates collected in Lombardy (Northern Italy) from food, food processing environments, and clinical cases, between 2013-2021, were characterized using multilocus sequence tying (MLST) and then screened for the presence of PMSC by sequencing the 2400 pb inlA gene. Results: A mutation leading to PMSC was identified on the position 277 (PMSC mutation type 26). To date, this mutation has been reported only on a CC7 strain from a clinical case. In our study, all isolates harbouring this mutation (n = 21) belong to ST325 (CC31, serotype 1/2a). Among these, 86% (n = 18) belong to the production chain of the Gorgonzola and Taleggio PDO cheeses. Conclusions: To the best of our knowledge, this is the first report of PMSC mutation type 26 on food and environmental isolates. Although PMSC 26 is rarely found worldwide, our findings suggest that ST325 with this mutation is widespread in the production chain of the two PDO cheeses, and the presence of this PMSC may be correlated with the low incidence of ST325 clinical cases. Key messages: A rare mutation leading to truncated InlA has been identified in a cluster of isolates correlated to the production chain of two PDO cheeses. Truncated inlA is suggestive of adaptation to the niche to the detriment of virulence.

Background: S. aureus is a widespread pathogen responsible for mild to severe human and animals’ infections. The abuse of antimicrobials provides the potential for selection of resistant strains in livestock, which represents a public health concern. S. aureus can carry several virulence factors of which staphylococcal enterotoxins (SEs) play a key role during food poisoning in human populations. The aim of this study is to monitor the prevalence of antimicrobial resistance factors in S. aureus isolates and their ability to produce enterotoxins. Methods: Within an ongoing monitoring plan for the assessment of antimicrobial resistance in S. aureus strains, a total of 83 isolates collected from food, and swine and dairy farms, between 2020-2022, were characterized using MLST and then screened for the presence of methicillin resistance and SEs genes. The isolates were tested for susceptibility to a panel of 14 antimicrobial agents using the disc agar diffusion method on Mueller-Hinton agar. Results: Among 83 S. aureus isolates, 53% carried at least one SEs gene. Eighteen isolates were methicillin-resistant of which 17 were no-enterotoxigenic strains belonging to ST398, and one was a food origin ST8 strain and harbored SEs genes. Among the ST398 isolates, only one was a food origin strain, while the others were from swine farms. The antibiogram showed that a few isolates were susceptible to nalidixic acid, and 42% resulted multidrug-resistant. Conclusions: Our results showed that more than half of S. aureus isolates were enterotoxigenic, the majority belonging to food industries. Numerous tested isolates resulted multidrug-resistant, confirming that antimicrobial resistance is a critical public health threat in a food safety perspective. Key messages: • The antimicrobial resistance profiles of S. aureus isolates underlines the importance of monitoring plans with a One Health perspective. • The prevalence of enterotoxins genes in S. aureus strains in Lombardy confirms the relevance of the microorganism as a foodborne pathogen.