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In the dairy industry, the late blowing defect in medium and long ripened cheeses causes important economic losses. This cheese defect, caused mainly by the anaerobic spore former Clostridium tyrobutyricum, is characterized by gas production resulting in holes formation in the paste, and in unpleasant aromas related to the butyrate production. Since longtime, the reference method used to detect the presence of total spore number in milk is the Most Probable Number (MPN), a semi-quantitative method which measures the bacterial growth and the gas production after heat treatment of the samples. This method often gives uncertain results, it requires long incubation periods and fails to differentiate between different species of spore-forming bacteria. A TaqMan real-time quantitative PCR targeting the phosphotransacetylase gene (pta) for C. tyrobutyricum was previously developed as an alternative technique to the traditional method, faster and more specific, and therefore of great interest to preventively determine the contamination of milk by C. tyrobutyricum. The aim of this work was the validation of this quantitative real-time PCR protocol for the C. tyrobutyricum detection in milk. Following the method validation, 120 samples of UHT and raw milk artificially contaminated with C. tyrobutyricum spores were analyzed in increasing concentrations, relating the results of the molecular method with those obtained by the MPN. Finally, 144 raw milk samples with a possible natural contamination were analyzed and demonstrated a positivity rate of 15.28% (CI95% 10.31–22.05%).

Italy has one of the highest incidence rates of foodborne botulism in Europe. It is a neuro-paralytic disease caused by ingesting ready-formed botulinum neurotoxins (BoNTs) produced by some spore-forming clostridia. Due to its severity and epidemic potential, foodborne botulism is included in the list of notifiable diseases, and botulism cases are managed based on collaboration among public health authorities providing food inspections, formal analysis, and supporting hospitals in rapid diagnosis. This study aimed to conduct a survey for investigating BoNT-producing clostridia occurrences in foods from Northern Italy between 2013 and 2020. In total, 2187 food samples were provided by food business operators for their own control protocols, and the presence of BoNT-producing clostridia was investigated using multiplex real-time PCR. Sixteen food samples (0.73%, 95% CI 0.45–1.19) were found positive for the presence of BoNT-producing clostridia, including dairy products, fruit, vegetables, sauces, bakery products, gastronomic preparations, meat products, and spices and flavourings, with the target genes bont/A and bont/B being the only targets found in this survey. Collaboration with local hospitals was carried out through analysis of biological samples in suspected foodborne botulism cases using a mouse bioassay for identifying the toxin involved. When available, food residues related to the suspected cases were also analysed. Among 52 patients with suspected symptoms of foodborne botulism, 18 patients were confirmed to be positive, showing the presence of BoNT/A, BoNT/B, and BoNT/E. In five cases, food residues were identified, mostly as homemade products. Our study highlights that foodborne botulism continues to be a public health concern in both industrial processing and homemade products, stressing the importance of consumer education to avoid hazardous botulism outbreaks.

In the three-years period 2012–2014, 160 cow milk samples from farms located in Lombardy and Emilia Romagna regions (Italy) were analyzed during the implementation of the Italian National Residues Monitoring Plan to assess the presence of PCDD/F, DL-PCB and NDL-PCB residues. The obtained contamination data were combined with cow milk consumption data from the Italian national dietary survey to estimate PCDD/F, DL-PCB and NDL-PCB human dietary exposure through the consumption of whole, semi skimmed and skimmed bovine milk. The exposure assessment was carried out separately for children, teenagers, adults and elderly. Average contamination levels of the analyzed samples were found to be 1.26 pg WHO-TEQ/g fat for the sum of PCDD/Fs and DL-PCBs and 9.30 ng/g fat for the sum of the 6 NDL-PCB indicators. PCB 126 was found to be the main contributor to the total WHO-TEQ. Using the upper bound approach, the estimated mean dietary intakes ranged from 0.07 pg WHO-TEQ/kg bw per day to 0.39 pg WHO-TEQ/kg bw per day for the sum of PCDD/Fs and DL-PCBs and considering exposure from whole milk. NDL-PCB mean dietary intakes resulted between 0.52 ng/kg bw per day and 2.86 ng/kg bw per day for consumption of whole milk. Children and teenagers were found to be the most exposed groups. This is the first time that Italian consumers exposure to NDL-PCBs is assessed using contamination data of cow milk produced in Italy.

The recent outbreak of Salmonella Agona linked to the consumption of infant formula (powdered formula) has rekindled the attention about the correct procedures for preparation and use of these products. International guidelines have already been published so far, particularly in association with Cronobacter sakazakii in early 2000s. FAO/WHO suggested to reconstitute formula with water at no less than 70°C. We therefore contaminated powdered formula with low levels of Salmonella spp and C sakazakii to evaluate the pathogens inactivation during the formula preparation using water at 70°C. In these conditions we observed a survival of both pathogens, indicating that the suggested recommendations may be not enough to guarantee the safety of this product. Higher temperatures are needed to reduce the biological risk, even if it may be not easily realized in actual domestic conditions. Moreover, the impact on the nutritional value of reconstituted formulas should be evaluated.

The aims of this study were to (1) study the optimal growth condition for two Lactococcus lactis strains to produce the anti-listeria compounds and (2) evaluate the effect of films activated with a twofold concentrated cell-free supernatant of La. lactis to inhibit Listeria monocytogenes. For the growth conditions, skimmed milk added to either Ml7 or MRS provided the highest levels of anti-listeria activity assayed (800 arbitrary unit per milliliter) for both La. lactis strains. A significant decrease of Li. monocytogenes counts was observed in cheese packed in films treated with an average decrease of 2.12 log colony-forming unit per gram after 15 days of storage. Practical Applications The antimicrobial film for food packaging can be used for the preservation of perishable foods such as the sliced cheese. This film developed in our study inhibits Listeria monocytogenes growth in sliced cheese, and does not have an adverse effect on the lactic microbiota, contributing to the preservation of the quality of the product. Thus, it could be a useful tool for the control of the development of pathogens in foods during storage of dairy products.

The aim of this study was to evaluate the behaviour of Escherichia coli, Salmonella Typhimurium and Listeria monocytogenes in an innovative semi-dry reduced fat Italian salami. The product is made from pork meat and it is characterized by less than 20% fat, lactose-, gluten- and milk protein-free. It is developed in Italy according to EC Regulation No. 1924/2006 “on nutrition and health claims made on foods”. Multi-strain cocktails of each pathogen were used to inoculate (5 log cfu g−1) separately the salami batter. During the manufacture and ripening, E. coli, S. Typhimurium, and L. monocytogenes decreased by 2.5, 1.65 and 0.5 log cfu g−1 respectively from the initial inoculated levels. Experimental data indicated that, during the shelf life in a condition of moderate thermal abuse (8–12 °C), the portioned and vacuum packed low fat salami are not able to support the growth of L. monocytogenes.

West Nile virus (WNV) is a recently re-emerged health problem in Europe. In Italy, an increasing number of outbreaks of West Nile disease, with occurrences of human cases, have been reported since 2008. This is particularly true in northern Italy, where entomological surveillance systems have been implemented at a regional level. The aim of this study was to use, for the first time, all the entomological data collected in the five regions undergoing surveillance for WNV in northern Italy to characterize the viral circulation (at a spatial and temporal scale), identify potential mosquito vectors, and specify relationships between virus circulation and meteorological conditions. In 2013, 286 sites covering the entire Pianura Padana area were monitored. A total of 757,461 mosquitoes were sampled. Of these, 562,079 were tested by real-time PCR in 9,268 pools, of which 180 (1.9%) were positive for WNV. The largest part of the detected WNV sequences belonged to lineage II, demonstrating that, unlike those in the past, the 2013 outbreak was mainly sustained by this WNV lineage. This surveillance also detected the Usutu virus, a WNV-related flavivirus, in 241 (2.6%) pools. The WNV surveillance systems precisely identified the area affected by the virus and detected the viral circulation approximately two weeks before the occurrence of onset of human cases. Ninety percent of the sampled mosquitoes were Culex pipiens, and 178/180 WNV-positive pools were composed of only this species, suggesting this mosquito is the main WNV vector in northern Italy. A significantly higher abundance of the vector was recorded in the WNV circulation area, which was characterized by warmer and less rainy conditions and greater evapotranspiration compared to the rest of the Pianura Padana, suggesting that areas exposed to these conditions are more suitable for WNV circulation. This observation highlights warmer and less rainy conditions as factors able to enhance WNV circulation and cause virus spillover outside the sylvatic cycle.

Long-chain n − 3 polyunsaturated fatty acids (PUFAs), particularly docosahexaenoic acid (DHA), have been shown to be essential during peri-natal growth and development of mammals. The study was conducted to determine whether the inclusion of fish oil in dairy goats diet during late gestation and early lactation would increase the proportion of n − 3 PUFAs in colostral and mature milk. From 3 weeks before kidding throughout 42 days of lactation, 14 dairy goats were fed either a control diet or a diet containing 1.1% fish oil (FO). Dietary treatment did not affect main components of colostrum, but lowered percentages of fat and lactose in mature milk. In colostrum, FO supplementation lowered the proportions of C18:0, C18:2, C18:3 and elevated those of C16:1 and very long-chain n − 3 PUFAs, mainly EPA (eicosapentaenoic acid) and DHA. In mature milk, FO supplementation lowered the proportion of C18:0 and raised C16:1 and n − 3 PUFAs (C18:3, EPA and DHA). Estimated transfer efficiencies for EPA and DHA into mature milk were 14 and 7%, respectively. Results suggest that goat's colostrum and milk can be enriched in n − 3 PUFAs by maternal dietary fish oil supplementation during late gestation and early lactation.

The aim of this work was to study the microbiological and physico-chemical changes throughout three cheesemaking replicates of Italian Formaggelle di capra cheese made from raw goat milk. Therefore, during the process, three samples of milk, curd and cheese at 3, 7, 11, 14, 21 and 30 days of ripening old cheese were taken from three cheesemaking replicates. The average of total mesophilic bacteria and Enterobacteriaceae count in raw milk was 5.27±0.57 and 3.8±1.02 Log cfu/mL, respectively. Lactic acid bacteria was the predominant bacterial group during the process, and they developed in different ways in each of the media used (M17 and MRS agar). Variability of microbial concentrations was observed between three cheesemaking replicates. A correlation between the presence of higher levels of Enterobacteriaceae in milk and the presence of other contaminants bacteria such as Escherichia coli β-glucuronidase-positive and coagulase-positive staphylococci was observed. In cheesemaking replicate n. 2, E. coli level was 5.07±0.03 Log cfu/mL and increased by about 1 log until the last week of ripening, when the level decreased to 5.69±0.2 Log cfu/mL. The milk used for the cheesemaking replicate n. 2 was found to be contaminated also by coagulase-positive staphylococci (3.18±0.06 Log cfu/mL), but the behaviour of this group appeared to be very variable. In this study a first step of process control and microbial groups study was performed and the cheesemaking process was registered in the website www.ars-alimentaria.it, the Italian site supported by the Italian Board of Health.

Consumption of raw or insufficiently cooked mussels contaminated with hepatitis A virus (HAV) is a major cause of infection to humans. The origin of mussels commonly used for the preparation of marinated seafood salads is often unknown, since different producers worldwide undergo a precooking treatment at the original collection site with methods and parameters not always indicated. These treatments could be insufficient for the inactivation of HAV, which is characterized by a high temperature resistance. Both high hydrostatic pressure (HHP) and marinade treatments have been shown to affect HAV vitality. In this study, two treatments (HHP and marinating) were combined in order to assess a potential synergistic effect on the virus vitality. A kinetic test was conducted by subjecting the experimentally-contaminated mussels (HAV titre: 10(6)/ml TCID50) to marinating, and to different HHP treatment (4,000; 5,000; and 6,000 bar for 1, 5, and 9 min). Virus post-treatment vitality was assessed by its ability to grow on cell cultures and by quantitative real-time RT-PCR to evaluate virus resistance under such conditions. Marinating treatment alone (final pH 4.3, and NaCl 2 %) did not inactivate the virus. On the other hand, the use of HHP treatment alone on non-marinated HAV-contaminated mussels was effective only above 5,000 bar for 5 min. The results of the present study elucidate the synergistic effect of a combination between marination and HHP treatments on the inactivation of the virus.

In order to simulate a contamination at the processing plant, one batch of freshlyprocessed salami batter (20 kg) was inoculated (1% v:w) with 5 log colony forming unit (CFU)/g of a multi-strain cocktail of two strains of Escherichia coli O157:H7 (registered and wild strain). Another batch was inoculated (1% v:w) with sterile physiological saline solution and used to check the lactic acid bacteria (Lab) behaviour and the changes of physicochemical parameters (pH and aw). Both batches were then processed to obtain a semi-dry salami (Hungarian-style): microbiological and physico-chemical properties were monitored during 94 days of ripening. During the manufacturing process, the levels of pathogen decreased of about 2.18 log CFU/g with respect to the initial inoculated levels. The behaviour of the indigenous bacteria such as Lab and the physico-chemical properties can help to determine the fate of pathogens throughout processing.

Pooled milk samples from 115 Bergamasca meat sheep were collected aseptically five times from lambing to weaning to determine the prevalence of intramammary infection, somatic cell counts and milk quality parameters (protein, fat and lactose), and effects of infection on lamb weight gain. The global prevalence of subclinical intramammary infection was 51·2%. The Staphylococcus genus was responsible for the greatest prevalence (53·3% among infected udders). Staphylococcus aureus was isolated in 8·4% of infected milk samples. Infection status had significant effects on fat and protein percentage and on somatic cell count. Lamb growth was greatest for lambs of ewes with no infection and decreased as the number of infected samples increased. No significant differences were detected in the growth of lambs with dams infected by different bacterial species.

The behaviour of Listeria monocytogenes and Escherichia coli O157:H7 was studied during the manufacture and ripening of two traditional Italian Alps cheeses. Each cheese type was manufactured in a pilot plan from raw cow milk (without the addition of starter cultures) artificially inoculated with L. monocytogenes and E. coli O157:H7 to a final concentration of about 4 log CFU/mL. The pathogens were enumerated throughout the cheese making and ripening processes to study their behaviour. When the milk was inoculated with 4 Log CFU/mL, the pathogens counts increased in the first time during the manufacturing process and then remained constant, until the end of ripening, or decreased significantly. Results indicate that the environment and nature of food borne pathogens affected the concentration of the bacteria during the manufacturing and ripening process. Thus, the presence of low cells numbers of L. monocytogenes and E. coli O157:H7 in milk destined for the production of raw milk cheeses characterized by a cooking of the curd less than 48°C can constitute a hazard for the consumer.