Trichinella britovi in wild boar meat from Italy, 2015-2021: A citizen science approach to surveillance.

As a result of the increase of game meat intended for human consumption through Europe, a plethora of food-borne diseases, including trichinellosis, may occur in consumers, posing a relevant public health threat. Thus, this study aims to a citizen science approach to monitor the occurrence of Trichinella spp. in wild boar meat intended for human consumption, evaluating the risk of infection for consumers. Following the European Regulation 2015/1375 (laying down specific rules on official controls for Trichinella in meat), from 2015 to 2021, hunters (n = 478) were involved to collect diaphragm pillar samples of wild boars from mainland southern Italy, which were tested for Trichinella spp. L1 larvae via HCl-pepsin digestion and Multiplex PCR. Overall, 139,160 animals were collected (average of 19,880 per year), being 14 (i.e., 0.01%) tested positive to Trichinella britovi by the combined biochemical and molecular approach. An average larval burden of 28.4 L1 per gram of meat was found (minimum 3.2 - maximum 132.6). A statistically significant difference was found in the prevalence according to hunting seasons (p < 0.01, with higher values in 2016 and 2021) and regions of the study area (p < 0.01). No statistically significant decrease in the prevalence of T. britovi throughout the study period was found (p = 0.51), except in Apulia region (p < 0.01). These findings revealed a stable prevalence of T. britovi in wild boar meat intended for human consumption, suggesting a risk of infection for consumers, especially hunters and local markets users. Citizen science surveillance models could be promoted to improve trichinellosis control and prevention in a One Health perspective.

Droplet Digital PCR (ddPCR) Analysis for the Detection and Quantification of Cow DNA in Buffalo Mozzarella Cheese.

Buffalo mozzarella cheese is one of the most appreciated traditional Italian products and it is certified as a Protected Designation of Origin (PDO) product under the European Commission Regulation No. 1151/2012. It is obtained exclusively from buffalo milk. If made from cow milk, or a mixture of buffalo and cow milk, buffalo mozzarella cheese does not qualify as a PDO product. In order to maximize their profits, some producers market buffalo mozzarella that also contains cow milk as a PDO product, thus defrauding consumers. New methods for revealing this fraud are therefore needed. One such method is the droplet digital Polymerase Chain Reaction (ddPCR). Thanks to its high precision and sensitivity, the ddPCR could prove an efficacious means for detecting the presence of cow milk in buffalo mozzarella cheese that is marketed as a PDO product. ddPCR has proved able to detect the DNA of cow and/or buffalo milk in 33 buffalo mozzarella cheeses labelled as PDO products, and experimental evidence could support its application in routine analyses.

The microbiota of water buffalo milk during mastitis.

The aim of this study was to define the microbiota of water buffalo milk during sub-clinical and clinical mastitis, as compared to healthy status, by using high-throughput sequencing of the 16S rRNA gene. A total of 137 quarter samples were included in the experimental design: 27 samples derived from healthy, culture negative quarters, with a Somatic Cell Count (SCC) of less than 200,000 cells/ml; 27 samples from quarters with clinical mastitis; 83 samples were collected from quarters with subclinical mastitis, with a SCC number greater of 200,000 cells/ml and/or culture positive for udder pathogens, without clinical signs of mastitis. Bacterial DNA was purified and the 16S rRNA genes were individually amplified and sequenced. Significant differences were found in milk samples from healthy quarters and those with sub-clinical and clinical mastitis. The microbiota diversity of milk from healthy quarters was richer as compared to samples with sub-clinical mastitis, whose microbiota diversity was in turn richer as compared to those from clinical mastitis. The core microbiota of water buffalo milk, defined as the asset of microorganisms shared by all healthy milk samples, includes 15 genera, namely Micrococcus, Propionibacterium, 5-7N15, Solibacillus, Staphylococcus, Aerococcus, Facklamia, Trichococcus, Turicibacter, 02d06, SMB53, Clostridium, Acinetobacter, Psychrobacter and Pseudomonas. Only two genera (Acinetobacter and Pseudomonas) were present in all the samples from sub-clinical mastitis, and no genus was shared across all in clinical mastitis milk samples. The presence of mastitis was found to be related to the change in the relative abundance of genera, such as Psychrobacter, whose relative abundance decreased from 16.26% in the milk samples from healthy quarters to 3.2% in clinical mastitis. Other genera, such as SMB53 and Solibacillus, were decreased as well. Discriminant analysis presents the evidence that the microbial community of healthy and clinical mastitis could be discriminated on the background of their microbiota profiles.

Recovery of Anisakid larvae by means of chloro-peptic digestion and proposal of the method for the official control.

Anisakidae larvae belonging to the genera Anisakis and Pseudoterranova, are the most responsible for zoonosis transmitted by fish products (anisakidosis). Acquired by the consumption of raw or undercooked marine fish or squid, the anisakid larvae may cause pathogenic diseases like gastric or intestinal anisakiasis and gastro-allergic disorders. In accordance with current EU legislation, the fresh fish products must be inspected visually in order to detect the possible presence of visible parasites. It is recognized that the visual method is not accurate enough to detect the larvae of parasites in food preparations containing raw or practically raw seafood and it clearly emerges that the official system of control needs to be able to utilise an most efficient analytical technique. In this work, the authors have drawn up and validated an analytical method, which involves artificial digestion and the use of a heated magnetic stirrer, based on the EU Regulation n. 2075/2005. The larvae isolated are then subjected to morphological identification at genus level by using optical microscope. The method, proved to be suitable for the detection of live and dead larvae of anisakidae in ready-to-eat foodstuffs containing raw fish or cephalopods and it is fast and accurate. The method showed high levels of sensitivity and specificity, and the suitability of its use in official food control was confirmed. Its use should be incorporated systematically into specific monitoring programs for the control of foodstuffs containing raw fish products.