Multiplex-PCR Detection of Clostridium tyrobutyricum, Clostridium butyricum, and Clostridium sporogenes in Raw Milk for Cheesemaking.

Late blowing defects in semi-hard and hard cheeses caused by spore-forming clostridia (e.g., Clostridium tyrobutyricum, Clostridium butyricum, Clostridium sporogenes) pose a major issue for the dairy industry. With this study, we applied a multiplex PCR for the rapid and simultaneous detection of clostridia in raw milk for cheese production. Spore detection in milk usually relies on culture-dependent methods, among which the most probable number (MPN) technique is sensitive but time-consuming and nonspecific. We tested two PCR-based protocols: the one entailed direct milk analysis with results obtained within 24 h; the other included an enrichment step and gave results within 72 h. The second protocol was found to be more sensitive; it detected concentrations as low as 100 cells/L for C. sporogenes and C. butyricum and 800 cells/L for C. tyrobutyricum. Both protocols were applied to field samples (211 samples underwent protocol no. 1; 117 samples underwent protocol no. 2) collected from four dairy processing plants in Piedmont. The prevalence of C. butyricum (protocol no. 1: 9.5%; protocol no. 2: 23%) was higher than either C. sporogenes (0%; 9.4%) or C. tyrobutyricum (0%; 6.8%). Protocol no. 2 detected multiple targets in eight samples, indicating that more than one microorganism was present. Our findings underscore the importance of implementing preventive measures and early detection strategies to mitigate the risk of cheese spoilage due to clostridial contamination.

Shiga Toxin-Producing Escherichia coli Isolated from Wild Ruminants in Liguria, North-West Italy.

Wildlife may represent an important source of infectious diseases for humans and other wild and domestic animals. Wild ruminants can harbour and transmit Shiga toxin-producing Escherichia coli (STEC) to humans, and some strains even carry important antimicrobial resistance. In this study, 289 livers of wild roe deer, fallow deer, red deer and chamois collected in Liguria, north-west Italy, from 2019 to 2023 were analysed. Overall, 44 STEC strains were isolated from 28 samples. The characterisation of serogroups showed the presence of O104, O113, O145 and O146 serogroups, although for 28 colonies, the serogroup could not be determined. The most prevalent Shiga toxin gene in isolated strains was Stx2, and more specifically the subtype Stx2b. The other retrieved subtypes were Stx1a, Stx1c, Stx1d and Stx2g. The isolated strains generally proved to be susceptible to the tested antimicrobials. However, multi-drug resistances against highly critical antimicrobials were found in one strain isolated from a roe deer. This study highlights the importance of wildlife monitoring in the context of a "One Health" approach.

Detection and Characterization of Zoonotic Pathogens in Game Meat Hunted in Northwestern Italy.

Wildlife can represent a reservoir of zoonotic pathogens and a public health problem. In the present study, we investigated the spread of zoonotic pathogens (Salmonella spp., Yersinia enterocolitica, Listeria monocytogenes, Shiga-toxin-producing Escherichia coli (STEC), and hepatitis E virus (HEV)) considering the presence of virulence and antibiotic resistance genes in game meat from animals hunted in northwest Italy. During two hunting seasons (2020 to 2022), samples of liver and/or muscle tissue were collected from chamois (n = 48), roe deer (n = 26), deer (n = 39), and wild boar (n = 35). Conventional microbiology and biomolecular methods were used for the detection, isolation, and characterization of the investigated pathogens. Two L. monocytogenes serotype IIa strains were isolated from wild boar liver; both presented fosfomycin resistance gene and a total of 22 virulence genes were detected and specified in the text. Eight Y. enterocolitica biotype 1A strains were isolated from chamois (2), wild boar (5), and deer (1) liver samples; all showed streptogramin and beta-lactam resistance genes; the virulence genes found were myfA (8/8 strains), ymoA (8/8), invA (8/8), ystB (8/8), and ail (4/8). Our data underscore the potential role of wildlife as a carrier of zoonotic and antibiotic-resistant pathogens in northwest Italy and a food safety risk for game meat consumers.

Food Safety Monitoring of Salmonella spp. in Northern Italy 2019-2021.

Salmonella is the second most frequent bacterial pathogen involved in human gastrointestinal outbreaks in the European Union; it can enter the food-production chain from animal or environmental sources or from asymptomatic food operators. European food legislation has established microbiological criteria to ensure consumer protection. Salmonella is listed under both process hygiene criteria and food safety criteria. Each EU member state designates an agency to organize or perform controls and other official activities. This paper describes the official control plans performed by competent authorities in Northern Italy in the three-year period 2019-2021. A total of 4413 food samples were delivered to the IZS Food Safety laboratories for Salmonella detection, of which 36 (0.8%) tested positive. Salmonella was most frequently detected in poultry meat samples (25/36 positive samples) followed by other meat products and pork products. The official controls for the protection of consumer health apply the EU's farm-to-fork approach: the samples were collected during production (food production plants), from products on the market, and from collective catering (restaurants, cafeterias, canteens). This manuscript will provide information about the presence of Salmonella in foodstuffs that can help competent authorities to set control plans based on risk assessments.