Identification and Genetic Characterization of a Novel Respirovirus in Alpine Chamois (rupicapra rupicapra rupicapra).

The Respirovirus genus, family Paramamixoviridae, includes respiratory viral pathogens. Here we report the identification and genetic characterization of a respirovirus in an Alpine chamois showing interstitial pneumonia associated with catarrhal bronchopneumonia. The full-genome characterization of this respirovirus, named ChamoisRV/IT2014, revealed low similarities to caprine respirovirus (77.1%), bovine respirovirus (74.5%) and human respirovirus (72.0%). The phylogenetic analyses based on the full-length genome sequence of the novel isolate and reference respirovirus strains showed that ChamoisRV/IT2014 clustered with caprine respirovirus but formed a separate branch. The phylogenetic tree topology of complete large protein amino acid sequences, representing the current species demarcation criterion for Respirovirus genus, showed a 0.05 branch length of ChamoisRV/IT2014 sequence between the nearest node and the tip of the branch, suggesting that this virus belongs to a novel species. This new isolate in a new host species raises several questions to be addressed on the epidemiological role of chamois and the risks of cross-transmission between wild ruminants and livestock.

BVDV permissiveness and lack of expression of co-stimulatory molecules on PBMCs from calves pre-infected with BVDV.

Bovine viral diarrhea virus (BVDV) has been detected in peripheral blood mononuclear cells (PBMCs) of immunocompetent animals, not being clear whether the development of a specific humoral immune response can prevent BVDV infection. The aim of this study was to evaluate the ability of non-cytopathic BVDV to replicate and produce infectious virus in PBMCs from calves pre-infected with BVDV and to elucidate the immunomodulatory effect of BVDV on these cells in an in vitro model. Quantification of virus was by quantitative PCR, while its replicative capacity and shedding into the extracellular environment was evaluated by viral titration. Apoptosis was assessed by flow cytometry analysis of annexin V and propidium iodide, and by expression of caspase-3/7. Flow cytometry was used to analyze the expression of CD14/CD11b/CD80, CD4/CD8/CD25, MHC-I/MHC-II and B-B2 markers. Our results showed that PBMCs from cattle naturally infected with BVDV were more susceptible to in vitro BVDV infection and showed a more severe apoptosis response than those from naïve animals. Non-cytopathic BVDV in vitro infection also resulted in a lack of effect in the expression of antigen presentation surface markers. All these findings could be related to the immunosuppressive capacity of BVDV and the susceptibility of cattle to this infection.

Presence of organic halogenated compounds, organophosphorus insecticides and polycyclic aromatic hydrocarbons in meat of different game animal species from an Italian subalpine area.

The exposure to several compounds such as polychlorinated biphenyls (PCBs), organochlorine pesticides (OCPs), organophosphorus pesticides (OPs), polycyclic aromatic hydrocarbon (PAHs), perfluoroalkyl substances (PFASs) and polybrominated diphenyl ethers (PBDEs) is a public health issue. The European Union (EU) recommended that its member states monitor the presence of emerging contaminants, like PBDEs and PFASs, in food and in the environment to obtain an accurate estimation of exposure. The tissues of wild animals exposed to these compounds can represent a suitable indicator of environmental pollution. The aim of this work is to evaluate: (i) the occurrence of PCBs, PBDEs, PFASs, PAHs, OCPs and OPs in four game animals' meat (chamois, red deer, wild boar and roe deer); (ii) interspecies differences and (iii) human exposure. Muscle samples from seventy-nine animals were collected during the hunting season in a Northern Italy mountain area at altitudes ranging from 300 to 2500 meters above sea level. The analyses were performed with gas chromatography-mass spectrometry (GC-MS/MS) and ultra-performance liquid chromatography - tandem mass spectrometry (UHPLC-MS/MS). No PBDEs were found in the samples. OCPs, OPs and PCBs were detected in almost all samples at different concentration ranges, showing higher frequency in ungulate species than in wild boar. PFAs were found only in wild boar. Anthracene and benzopyrene, among PAHs, were found only in chamois, at low concentrations. The lack of an accurate pattern of exposure as well as variable consumption by hunters does not allow accurate risk characterisation. However, a low risk for consumers can be indicated due to the frequent detection of contaminants only at trace levels, the scarce prevalence of high concentrations of some contaminants and the low consumption of game animal meat. In conclusion, the organisation of a control plan on residues in game animals would be advisable.

Quality parameters of hunted game meat: Sensory analysis and pH monitoring.

The aim of the present research is to propose a new, quick and objective method for the certification of hunted and/or culled wild game meat quality and to monitor its origin and the hunting practices adopted by hunters. The expected deliverable is a new labelling scheme for Italian hunted wild game meat that will guarantee high quality and safety standards for consumers and will decrease transaction costs of the supply chains. During the 2015, 2016 and 2017 hunting seasons, 1,056 hunted wild ungulates were sampled. Specifically, alpine chamois (n=537), roe deer (n=113), red deer (n=342) and wild boar (n=64), which were all hunted in the VCO2-Ossola Nord hunting district (Verbania Province, Piedmont, Italy). Samples of the longissimus dorsi were collected to evaluate the nutritional parameters and the acid profiles of the products. As a measure of meat quality, pH values have been recorded after slaughtering by inserting a probe in the semimembranosus muscle. The results were categorized as DFD (pH≥6,2), intermediate DFD (5,8≤pH<6,2) and high-quality meat (pH <5,8). As explanatory variables for the quality of wild game meat, differences based on age, gender and hunting practices were considered. Concerning the latter variables, measures were collected from animals received at hunting districts control centers by trained technicians who also collected information on the hunting practices, i.e., bleeding and evisceration of the carcasses and number of shots. Nutritional values showed low fat (<3 g per 100 g), low saturated fat (<1,5 g per 100 g) and high protein contents. Furthermore, wild game meat has high values of ω3 and CLA, ensuring a positive ω6/ω3 ratio. Differences were found in the concentrations of fat between age and gender, considering that during the mating season, adult males' weight loss can exceed 40%. Hunting practices seem to affect meat quality.

Host range of mammalian orthoreovirus type 3 widening to alpine chamois.

Mammalian orthoreoviruses (MRV) type 3 have been recently identified in human and several animal hosts, highlighting the apparent lack of species barriers. Here we report the identification and genetic characterization of MRVs strains in alpine chamois, one of the most abundant wild ungulate in the Alps. Serological survey was also performed by MRV neutralization test in chamois population during five consecutive years (2008-2012). Three novel MRVs were isolated on cell culture from chamois lung tissues. No respiratory or other clinical symptoms neither lung macroscopic lesions were observed in the chamois population. MRV strains were classified as MRV-3 within the lineage III, based on S1 phylogeny, and were closely related to Italian strains identified in dog, bat and diarrheic pig. The full genome sequence was obtained by next-generation sequencing and phylogenetic analyses showed that other segments were more similar to MRVs of different geographic locations, serotypes and hosts, including human, highlighting genome reassortment and lack of host specific barriers. By using serum neutralization test, a high prevalence of MRV-3 antibodies was observed in chamois population throughout the monitored period, showing an endemic level of infection and suggesting a self-maintenance of MRV and/or a continuous spill-over of infection from other animal species.

Clonal diversity, virulence-associated genes and antimicrobial resistance profile of Staphylococcus aureus isolates from nasal cavities and soft tissue infections in wild ruminants in Italian Alps.

Staphylococcus aureus is a commensal and a pathogenic bacterium that causes a wide variety of diseases in humans and animals with a high impact on public health and the livestock industry. S. aureus virulence pattern, antimicrobial resistance profile and host specialization are of great concern both in livestock and in companion animals. Concerning wild animals, S. aureus carriage and antimicrobial resistance profile has been recently investigated in free-ranging species both in aquatic and terrestrial environment. Here we report genotyping (spa typing, Multilocus Sequence Typing and SCCmec typing), virulence and antimicrobial resistance profile of four S. aureus isolated in Alpine chamois (Rupicapra r. rupicapra) and roe deer (Capreolus capreolus), euthanized due to walking impairment and signs of disorientation. S. aureus was isolated from nasal cavities in both wild ruminant species and in soft tissue infections in chamois. A marked S. aureus genetic heterogeneity was detected: spa type t1523, sequence type 45 (Clonal Complex 45), and spa type t1328, ST22 (CC22) from the nasal cavities and the liver of a chamois kid respectively, t1773, ST700 (CC130) from an adult chamois abscess, and a new sequence type, ST2712, belonging to CC97 from the roe deer nasal cavities. One of the main findings was the confirmation that the t1328, ST22 isolate, obtained from the liver of the chamois kid, was a methicillin-resistant S. aureus (MRSA) harbouring a SCCmec cassette type IV. The set of virulence marker and toxin genes investigated showed profiles characteristic of the S. aureus lineages detected, including those of the human adapted ST (CC) 22 and ST (CC) 45 isolates.