2. Universidad Cardenal Herrera-CEU

Severe Acute Respiratory Syndrome Coronavirus 2 is the causative agent of Coronavirus Disease 2019 in humans. Among domestic animals, cats are more susceptible to SARS-CoV-2 than dogs. The detection of anti-SARS-CoV-2 antibodies in seemingly healthy cats and/or infected cats which are in close contact with infected humans has been described. The presence of animals that tested positive by serology or molecular techniques could represent a potential transmission pathway of SARS-CoV-2 that can spill over into urban wildlife. This study analyses the seroprevalence variation of SARS-CoV-2 in stray cats from different waves of outbreaks in a geographical area where previous seroepidemiological information of SARS-CoV-2 was available and investigate if SARS-CoV-2-seropositive cats were exposed to other co-infections causing an immunosuppressive status and/or a chronic disease that could lead to a SARS-CoV-2 susceptibility. For this purpose, a total of 254 stray cats from Zaragoza (Spain) were included. This analysis was carried out by the enzyme-linked immunosorbent assay using the receptor binding domain of Spike antigen and confirmed by serum virus neutralization assay. The presence of co-infections including Toxoplasma gondii, Leishmania infantum, Dirofilaria immitis, feline calicivirus, feline herpesvirus type 1, feline leukemia virus and feline immunodeficiency virus, was evaluated using different serological methods. A seropositivity of 1.57% was observed for SARS-CoV-2 including the presence of neutralizing antibodies in three cats. None of the seropositive to SARS-CoV-2 cats were positive to feline coronavirus, however, four SARS-CoV- 2-seropositive cats were also seropositive to other pathogens such as L. infantum, D. immitis and FIV (n = 1), L. infantum and D. immitis (n = 1) and L. infantum alone (n = 1).Considering other pathogens, a seroprevalence of 16.54% was detected for L. infantum, 30.31% for D. immitis, 13.78%, for T. gondii, 83.86% for feline calicivirus, 42.52% for feline herpesvirus type 1, 3.15% for FeLV and 7.87% for FIV. Our findings suggest that the epidemiological role of stray cats in SARS-CoV-2 transmission is scarce, and there is no increase in seropositivity during the different waves of COVID-19 outbreaks in this group of animals. Further epidemiological surveillances are necessary to determine the risk that other animals might possess even though stray cats do not seem to play a role in transmission.

We used anonymous questionnaires to assess the hygienic and sanitary aspects of game meat self-consumption in Eastern Spain as the first step towards a health risk assessment. The survey yielded 472 valid interviews from active hunters. The maximum possible score was 65 points (average 29 8; range 1–52). Most participants were men (95%), but women achieved significantly better scores (p = 0.003). Hunters above 65 years old scored significantly lower results than younger groups (p = 0.007). The score increased with the educational level (p = 0.046). A 92% of the collaborators consumed game meat. Veterinary inspection and freezing were irregular among the participants. Most respondents declared carrying the animals in their personal vehicles. Of the dressing process, 61% of sites were outdoors, 68% of the participants declared using specific knives, 64% used the same clothes as in the field, and 42% used disposable gloves. The most usual way to dispose of the remains was garbage containers (41%); offal abandonment in the field was 33%, and 13% fed domestic animals using the remains. We conclude that public health authorities should increase their interest in the self-consumption of game meat. Clear guidelines about domestic dressing facilities and hygienic habits should be published, these being essential when looking for synergies with hunter associations.

Animals have been involved in the three known outbreaks of severe respiratory syndromes due to coronaviruses (years 2005, 2012, and 2019). The pandemic nature of the SARS-CoV-2 outbreak increases the likelihood of infection from humans of susceptible animal species that, thus, could become secondary viral hosts and even disease reservoirs. We present evidence of spillover infection of wild mustelids by reporting the presence of SARS-CoV-2 in a Eurasian river otter found near a water reservoir in the Valencian Community (Spain). We detected the virus using two different commercial RTqPCR assays on RNA extracted from the nasopharynx (swabbing) and from lung tissue and mediastinal lymph node homogenates. The corresponding samples from two additional otters from distant sites tested negative in identical assays. The diagnosis in the positive otter was confirmed by two-tube RT-PCR assay in which RNA was first retrotranscribed, and then specific regions of the spike (S), nucleocapsid (N), and ORF10 genes were separately amplified from the produced cDNA, followed by electrophoretic visualization and Sanger sequencing. The sequences of the amplified products revealed some non-synonymous changes in the N and ORF10 partial sequences, relative to the consensus sequence. These changes, identified already in human patient samples, point to human origin of the virus, although their specific combination was unique. These findings, together with our previous report of SARS-CoV-2 infection of feral American mink, highlight the need for SARS-CoV-2 surveillance of wild or feralmustelids to evaluate the risk that these animals could become SARS-CoV-2 reservoirs.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causal agent of COVID-19, is considered a pathogen of animal origin that is mainly transmitted from human to human. Several animal species can be naturally or experimentally infected by SARS-CoV-2, with compelling evidence that mink is highly susceptible to SARS-CoV-2 infection. Human-to-mink infection cases have been reported and there are also suggestions that mink-to-human infection occurs. Mink infections have been reported to date only on fur farms, except for one infected freeranging wild mink near a Utah (USA) fur farm, which suggests a transmission pathway from farms to wild mink. We now report the detection of SARS-CoV-2 in 2 of 13 feral dark brown American mink (Neovison vison) trapped in the Valencian Community (Eastern Spain), during an invasive species trapping campaign. They were trapped in riverbeds in sparsely inhabited rural areas known to harbor self-sustained feral mink populations. The closest fur farm is about 20 km away. SARS-CoV-2 RNA was detected by two-step RT-PCR in these animals’ mesenteric lymph nodes and was confirmed by sequencing a 397-nucleotide amplified region of the S gene, yielding identical sequences in both animals. A molecular phylogenetic analysis was run on this sequence, which was found to correspond to the consensus SARS-CoV-2 sequence fromWuhan. Our findings appear to represent the first example of SARS-CoV-2 acquired in the wild by feral mink in self-sustained populations.

The appearance of methicillin-resistant strains of Staphylococcus aureus (MRSA) in several animal species (including rabbits) has set o alarms for their capacity to act as reservoirs for this bacterium. This is especially important in wild animals given its epidemiological implications. The objectives of this study were to identify and characterize S. aureus, specifically MRSA, strains in wild lagomorph high-density areas. Ten hares and 353 wild rabbits from 14 towns with a high rabbit density in the Valencian region (eastern Spanish coast) were sampled. Swabs from the nasal cavity, ears, perineum and lesions (when present) were taken for microbiological studies. The detection of di erent genes and antibiotic susceptibility studies were also carried out. Of all the animals, 41.3% were positive for S. aureus, of which 63.3% were MRSA. Ears were the anatomical location with more S. aureus and MRSA strains. The more frequently identified MLST type was ST1945 (97.1%, 136/140). The mecA gene was found only in one sample. The rest (n = 139) carried the mecC gene and were included in CC130, except one. Penicillin resistance was detected in 28 mec-negative isolates and, in one case, bacitracin resistance. mecA isolate presented resistance to enrofloxacin and tetracycline, and 10 mecC isolates also showed bacitracin resistance. No MRSA isolate was positive for genes chp, sea, tst and PVL. Two ST1945 isolates contained IEC type E (comprising genes scn and sak). mecA-isolate was positive for blaZ. Of the 28 MSSA strains showing resistance to penicillin, 22 carried the blaZ gene. These surprising results highlight the marked presence of MRSA strains in wild rabbits in high-density areas.