SARS-CoV-2 Research in Dogs (Canis lupus familiaris) and Felines (Felis silvestris catus) Domiciled in an International Border Region (Paraguay and Brazil).

Introduction: COVID-19 is an infectious disease caused by SARS-CoV-2 that has become a serious threat to public health owing to its rapid spread from aerosols from infected people. Despite being considered a strictly human disease, there are reports in the literature about animals with confirmed presence of the virus. Aim: Owing to the scarcity of scientific literature on the potential for infection of animals and their importance for One Health, the objective of this work was to research SARS-CoV-2 RNA in felines (Felis silvestris catus) and dogs (Canis lupus familiaris) domiciled. Materials and Methods: Oropharyngeal swabs were collected from domestic dogs and cats belonging to patients diagnosed with COVID-19 from August to October 2021 and residents of the northwest and west regions of Paraná, Brazil. Results: Of the 34 samples collected, 14 were from dogs and 20 from cats. Three of these samples tested positive in real-time PCR, and two of them were also positive in the immunochromatographic test. After testing positive in real-time PCR, the samples underwent genetic sequencing using the Illumina COVIDSeq test. Of the 34 samples collected, three (9%), all of them female and from the feline species, tested positive in real-time PCR, with two of these (67%) also testing positive in the immunochromatographic test. Regarding sequencing, it was possible to sequence the three samples aligned with the AY.101 lineage, corresponding to the Delta variant. Conclusion: The occurrence of SARS-CoV-2 infection in dogs and cats is seen as an unintended event with significant implications for public health, including its potential transmission to other animal species. Further research is required to enhance our understanding of how this disease spreads among these animals and its broader impact on One Health initiatives.

One Health surveillance of West Nile and Usutu viruses: a repeated cross-sectional study exploring seroprevalence and endemicity in Southern France, 2016 to 2020.

BackgroundWest Nile virus (WNV) and Usutu virus (USUV), two closely related flaviviruses, mainly follow an enzootic cycle involving mosquitoes and birds, but also infect humans and other mammals. Since 2010, their epidemiological situation may have shifted from irregular epidemics to endemicity in several European regions; this requires confirmation, as it could have implications for risk assessment and surveillance strategies.AimTo explore the seroprevalence in animals and humans and potential endemicity of WNV and USUV in Southern France, given a long history of WNV outbreaks and the only severe human USUV case in France in this region.MethodsWe evaluated the prevalence of WNV and USUV in a repeated cross-sectional study by serological and molecular analyses of human, dog, horse, bird and mosquito samples in the Camargue area, including the city of Montpellier, between 2016 and 2020.ResultsWe observed the active transmission of both viruses and higher USUV prevalence in humans, dogs, birds and mosquitoes, while WNV prevalence was higher in horses. In 500 human samples, 15 were positive for USUV and 6 for WNV. Genetic data showed that the same lineages, WNV lineage 1a and USUV lineage Africa 3, were found in mosquitoes in 2015, 2018 and 2020.ConclusionThese findings support existing literature suggesting endemisation in the study region and contribute to a better understanding of USUV and WNV circulation in Southern France. Our study underlines the importance of a One Health approach for the surveillance of these viruses.

A novel hepadnavirus in domestic dogs.

Hepadnaviruses have been identified in several animal species. The hepadnavirus prototype, human hepatitis B virus (HBV), is a major public health problem associated with chronic liver diseases and hepatocellular carcinoma. Recently, a novel hepadnavirus, similar to HBV, was identified in domestic cats. Since several pathogens can be shared between cats and dogs, we hypothesized that dogs could also harbor hepadnaviruses and we tested a collection of canine sera with multiple molecular strategies. Overall, hepadnavirus DNA was identified in 6.3% (40/635) of canine serum samples, although the viral load in positive sera was low (geometric mean of 2.70 × 102 genome copies per mL, range min 1.36 × 102-max 4.03 × 104 genome copies per mL). On genome sequencing, the canine hepadnaviruses revealed high nucleotide identity (about 98%) and similar organization to the domestic cat hepadnavirus. Altered hepatic markers were found in hepadnavirus-positive dogs, although the role of hepadnavirus in canine health remains to be elucidated.

4-hydroxy-2-nonenal as a marker of the oxidative stress in brains of dogs with canine distemper / 4-hidroxi-2-nonenal como marcador de estresse oxidativo no cérebro de cães com cinomose

Canine Distemper is a disease caused by Canine morbillivirus (CM), a pantropic virus that can affect the central nervous system (CNS), causing demyelination. However, the pathogenesis of this lesion remains to be clarified. Brain samples of 14 naturally infected dogs by CM were analyzed to evaluate the presence of oxidative stress and demyelination. RT-PCR assay was performed to confirm a diagnosis of canine distemper in the brain, immunohistochemistry anti-CM was used to localize the viral proteins in the tissue, and anti-4-hydroxy-2-nonenal (4-HNE) was a marker of a product of lipid peroxidation. The results showed the presence of viral proteins in the demyelinated area with the presence of 4-HNE. Our results suggest that the CM virus infection causes oxidative stress leading to lipid peroxidation, which causes tissue damage and demyelination. In conclusion, oxidative stress plays a significant role in canine distemper pathogenesis in the CNS.(AU)

A cinomose canina é uma doença causada pelo Morbilivírus canino (CM), um vírus pantrópico que pode afetar o sistema nervoso central (SNC), causando desmielinização. No entanto, a patogênese dessa lesão não está totalmente esclarecida. RT-PCR e imuno-histoquímica foram realizadas para confirmação do diagnóstico de cinomose em amostras de encéfalo de 14 cães naturalmente infectados. Após confirmação, foi realizada uma avaliação do estresse oxidativo por imuno-histoquímica com uso de anti-4-hidroxi-nonenal (4HNE) como marcador de produtos resultantes da peroxidação lipídica. Os resultados sugerem que a infecção pelo CM causa estresse oxidativo no tecido, levando a peroxidação lipídica, a qual causa danos ao tecido, culminando com desmielinização. Conclui-se que o estresse oxidativo tem papel importante na patogênese da cinomose canina no sistema nervoso central.(AU)

Molecular Epidemiology of Rabies in Wild Canidae in Tunisia.

Rabies is a viral zoonosis that is transmissible to humans via domestic and wild animals. There are two epidemiological cycles for rabies, the urban and the sylvatic cycles. In an attempt to study the epidemiological role of wild canidae in rabies transmission, the present study aimed to analyze the genetic characteristics of virus isolates and confirm prior suggestions that rabies is maintained through a dog reservoir in Tunisia. Virus strains isolated from wild canidae were subject to viral sequencing, and Bayesian phylogenetic analysis was performed using Beast2 software. Essentially, the virus strains isolated from wild canidae belonged to the Africa-1 clade, which clearly diverges from fox-related strains. Our study also demonstrated that genetic characteristics of the virus isolates were not as distinct as could be expected if a wild reservoir had already existed. On the contrary, the geographic landscape is responsible for the genetic diversity of the virus. The landscape itself could have also acted as a natural barrier to the spread of the virus.

Cell Entry of Animal Coronaviruses.

Coronaviruses (CoVs) are a group of enveloped positive-sense RNA viruses and can cause deadly diseases in animals and humans. Cell entry is the first and essential step of successful virus infection and can be divided into two ongoing steps: cell binding and membrane fusion. Over the past two decades, stimulated by the global outbreak of SARS-CoV and pandemic of SARS-CoV-2, numerous efforts have been made in the CoV research. As a result, significant progress has been achieved in our understanding of the cell entry process. Here, we review the current knowledge of this essential process, including the viral and host components involved in cell binding and membrane fusion, molecular mechanisms of their interactions, and the sites of virus entry. We highlight the recent findings of host restriction factors that inhibit CoVs entry. This knowledge not only enhances our understanding of the cell entry process, pathogenesis, tissue tropism, host range, and interspecies-transmission of CoVs but also provides a theoretical basis to design effective preventive and therapeutic strategies to control CoVs infection.

Predicting hosts based on early SARS-CoV-2 samples and analyzing the 2020 pandemic.

The SARS-CoV-2 pandemic has raised concerns in the identification of the hosts of the virus since the early stages of the outbreak. To address this problem, we proposed a deep learning method, DeepHoF, based on extracting viral genomic features automatically, to predict the host likelihood scores on five host types, including plant, germ, invertebrate, non-human vertebrate and human, for novel viruses. DeepHoF made up for the lack of an accurate tool, reaching a satisfactory AUC of 0.975 in the five-classification, and could make a reliable prediction for the novel viruses without close neighbors in phylogeny. Additionally, to fill the gap in the efficient inference of host species for SARS-CoV-2 using existing tools, we conducted a deep analysis on the host likelihood profile calculated by DeepHoF. Using the isolates sequenced in the earliest stage of the COVID-19 pandemic, we inferred that minks, bats, dogs and cats were potential hosts of SARS-CoV-2, while minks might be one of the most noteworthy hosts. Several genes of SARS-CoV-2 demonstrated their significance in determining the host range. Furthermore, a large-scale genome analysis, based on DeepHoF's computation for the later pandemic in 2020, disclosed the uniformity of host range among SARS-CoV-2 samples and the strong association of SARS-CoV-2 between humans and minks.

Serological Evidence of Phleboviruses in Domestic Animals on the Pre-Apennine Hills (Northern Italy).

Phleboviruses are arboviruses transmitted by sand flies, mosquitoes and ticks. Some sand fly-borne phleboviruses cause illnesses in humans, such as the summer fevers caused by the Sicilian and Naples viruses or meningitis caused by the Toscana virus. Indeed, traces of several phleboviral infections have been serologically detected in domestic animals, but their potential pathogenic role in vertebrates other than humans is still unclear, as is the role of vertebrates as potential reservoirs of these viruses. In this study, we report the results of a serological survey performed on domestic animals sampled in Northern Italy, against four phleboviruses isolated from sand flies in the same area. The sera of 23 dogs, 165 sheep and 23 goats were tested with a virus neutralization assay for Toscana virus, Fermo virus, Ponticelli I virus and Ponticelli III virus. Neutralizing antibodies against one or more phleboviruses were detected in four out of 23 dogs, 31 out of 165 sheep and 12 out of 23 goats. This study shows preliminary evidence for the distribution pattern of phleboviral infections in different animal species, highlighting the potential infection of the Toscana virus in dogs and the Fermo virus in goats.

Current State of Knowledge about Role of Pets in Zoonotic Transmission of SARS-CoV-2.

Pets play a crucial role in the development of human feelings, social life, and care. However, in the era of the prevailing global pandemic of COVID-19 disease caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), many questions addressing the routes of the virus spread and transmission to humans are dramatically emerging. Although cases of SARS-CoV-2 infection have been found in pets including dogs, cats, and ferrets, to date there is no strong evidence for pet-to-human transmission or sustained pet-to-pet transmission of SARS-CoV-2. However, an increasing number of studies reporting detection of SARS-CoV-2 in farmed minks raises suspicion of potential viral transmission from these animals to humans. Furthermore, due to the high susceptibility of cats, ferrets, minks and hamsters to COVID-19 infection under natural and/or experimental conditions, these animals have been extensively explored as animal models to study the SARS-CoV-2 pathogenesis and transmission. In this review, we present the latest reports focusing on SARS-CoV-2 detection, isolation, and characterization in pets. Moreover, based on the current literature, we document studies aiming to broaden the knowledge about pathogenicity and transmissibility of SARS-CoV-2, and the development of viral therapeutics, drugs and vaccines. Lastly, considering the high rate of SARS-CoV-2 evolution and replication, we also suggest routes of protection against the virus.

First complete genome sequence and molecular characterization of Canine morbillivirus isolated in Central Brazil.

The Brazilian regions are still highly endemic areas for Canine morbillivirus [canine distemper virus (CDV)]. However, little is known regarding the genetic variability of the strain circulating in several Brazilian regions. Here, we report the first full-length genome and molecular characterization of CDV isolated from domestic dogs in the Brazilian Center-West region. Sequence alignment and phylogenetic analyses based on deduced amino acid and nucleotide sequences showed that the isolated strain is characterized as the South America-I/Europe genotype. However, it segregates into a CDV subgenotype branch. Interestingly, both H and F proteins have a gain of a potential N-glycosylation sites compared to the Onderstepoort vaccine strain. Therefore, this study provides a reference to further understand the epidemic and molecular characteristics of the CDV in Brazil.

Isolation and genetic characterization of canine adenovirus type 2 from a domestic dog showing neurological symptoms.

Canine adenoviruses (CAVs) are of two types: canine adenovirus type 1 (CAV-1), which causes infectious canine hepatitis, and canine adenovirus type 2 (CAV-2), which is mainly associated with the respiratory type of disease in dogs. Due to the widespread use of modified live vaccines to control canine adenoviral infections and subsequently reduced disease incidence, CAVs are often neglected by clinicians. Although a number of studies are available about CAV-1 prevalence in India, only meagre information is available about CAV-2. This study reports the CAV-2 infection in a vaccinated dog with neurological and respiratory symptoms which was found negative for other canine pathogens like canine distemper virus and canine parvovirus. The virus was successfully isolated from rectal swab in MDCK cells and characterized by immunofluorescence assay and virus neutralization test. On phylogenetic analysis of partial E3 region, the Indian CAV-2 grouped in a separate clade different from established subgroups. An insertion of "G" nucleotide was reported at nucleotide (nt.) position 1077 in the E3 gene of Indian CAV-2 isolates which led to a frameshift in the coding region of E3 gene thereby imparting additional eleven amino acids to its C-terminal end in comparison to isolates from other parts of the world. This may have an implication on the functional role of E3 protein inside the cell. This study reinforces the unique signature insertion in the E3 gene of Indian CAV-2 and is the second study in the world to report the association of CAV-2 with neurological disease in dogs.

SARS-CoV-2 Infections and Viral Isolations among Serially Tested Cats and Dogs in Households with Infected Owners in Texas, USA.

Understanding the ecological and epidemiological roles of pets in the transmission of SARS-CoV-2 is critical for animal and human health, identifying household reservoirs, and predicting the potential enzootic maintenance of the virus. We conducted a longitudinal household transmission study of 76 dogs and cats living with at least one SARS-CoV-2-infected human in Texas and found that 17 pets from 25.6% of 39 households met the national case definition for SARS-CoV-2 infections in animals. This includes three out of seventeen (17.6%) cats and one out of fifty-nine (1.7%) dogs that were positive by RT-PCR and sequencing, with the virus successfully isolated from the respiratory swabs of one cat and one dog. Whole-genome sequences of SARS-CoV-2 obtained from all four PCR-positive animals were unique variants grouping with genomes circulating among people with COVID-19 in Texas. Re-sampling showed persistence of viral RNA for at least 25 d-post initial test. Additionally, seven out of sixteen (43.8%) cats and seven out of fifty-nine (11.9%) dogs harbored SARS-CoV-2 neutralizing antibodies upon initial sampling, with relatively stable or increasing titers over the 2-3 months of follow-up and no evidence of seroreversion. The majority (82.4%) of infected pets were asymptomatic. 'Reverse zoonotic' transmission of SARS-CoV-2 from infected people to animals may occur more frequently than recognized.

Trend and epidemiological patterns of animal bites in Golestan province (Northern Iran) between 2017 and 2020.

BACKGROUND: Rabies is one of the oldest zoonosis viral diseases, which still remains as one of the most important threats to public health in the 21st century. METHODS: This cross-sectional study examined epidemiologic features of all 33,996 cases of persons bitten by animals and referred to the rabies prophylaxis centers in Golestan province between March 2017 and March 2020. Factors included demographic information of the victim (age, gender, and occupation), type of invasive animals (dog, cat, and other types), time of bite (year, month, and hour), place of residence (urban or rural), and injury and treatment statuses. We also obtained national and provincial animal bite incidence data for all of Iran and for Golestan province for the longer interval 2013-2020 to examine broader time trends. We used SPSS version 19, QGIS version 3.1, and Excel 2013 to generate frequency distributions and descriptive statistics. RESULTS: The incidence rates of animal bites in Golestan province and Iran as a whole both increased smoothly. The latest incidence rate of animal bites in Golestan was 652 per 100,000 people, almost three times the overall national figure for 2020. Most cases of animal bites (67.6%) occurred in rural areas, and 36% of the victims aged under 19 years old. Dog and cat bites accounted for the great majority of cases (89% and 8%, respectively). The highest rate of animal bites was reported in the spring (30.8%). The lower limb was the most commonly bitten area in these individuals (64.6%). Of note, 87% of the cases received incomplete prophylactic post-exposure treatment, and 18% received immunoglobulin. CONCLUSION: The increasing rates of animal bites in the study area as well as the higher rate compared to the national average indicates the need for further review of animal bite control programs.

Feasibility and Effectiveness Studies with Oral Vaccination of Free-Roaming Dogs against Rabies in Thailand.

(1) Background: Thailand has made significant progress in reducing the number of human and animal rabies cases. However, control and elimination of the last remaining pockets of dog-mediated rabies have shown to be burdensome, predominantly as a result of the large numbers of free-roaming dogs without an owner that cannot be restrained without special efforts and therefore remain unvaccinated. To reach these dogs, the feasibility, and benefits of oral rabies vaccination (ORV) as a complementary tool has been examined under field conditions. (2) Methods: ORV of dogs was tested in five study areas of four provinces in Thailand. In these areas, sites with free-roaming dogs were identified with the support of local municipal workers and dog caretakers. ORV teams visited each of five study areas and distributed rabies vaccine (SPBN GASGAS) in three bait formats that were offered to the dogs using a hand-out and retrieval model. The three bait types tested included: egg-flavored baits, egg-flavored baits pasted with commercially available cat liquid snack, and boiled-intestine baits. A dog offered a vaccine bait was considered vaccinated when the discarded sachet was perforated or if a dog chewed vaccine bait at least 5 times before it swallowed the bait, including the sachet. (3) Results: A total of 2444 free-roaming dogs considered inaccessible for parenteral vaccination were identified at 338 sites. As not all dogs were approachable, 79.0% were offered a bait; of these dogs, 91.6% accepted the bait and subsequently 83.0% were considered successfully vaccinated. (4) Conclusion: Overall, 65.6% of the free-roaming dogs at these sites were successfully vaccinated by the oral route. Such a significant increase of the vaccination coverage of the free-roaming dog population could interrupt the rabies transmission cycle and offers a unique opportunity to reach the goal to eliminate dog-mediated human rabies in Thailand by 2030.

Awareness and knowledge of canine rabies: A state-wide cross-sectional study in Nigeria.

Rabies is a highly fatal disease that is endemic in Nigeria. The poor community awareness and knowledge of canine rabies have thwarted the realization of zero deaths from dog mediated human rabies. This study aimed to assess the awareness and knowledge of canine rabies in Kwara state. A total of 1,460 questionnaires were administered to respondents in the three senatorial zones of the state using open data kit (ODK) on mobile phones between September 2019 to January 2020. The rabies awareness rate was 38.1%. The mean knowledge score was 3.78 ± 2.15. Only 10.6% (n = 59/557) of the respondents had satisfactory knowledge of canine rabies. Respondents had poor knowledge of the mode of transmission, symptoms, prevention, and the control measures needed to eliminate canine rabies. Only 20.1% of respondents owned at least a dog. Dog owners were 3.85× (95% CI: 2.89, 5.13; p < 0.01) more likely to be aware of canine rabies and were 1.78× (95% CI: 1.22-2.60; p = 0.003) more likely to have satisfactory knowledge about canine rabies than non-dog owners. Respondents with tertiary education were at least 6.81× (95% CI: 4.24, 10.92; p < 0.01) more likely to be aware of rabies than respondents with no formal education. The findings of this study showed very low awareness and knowledge of canine rabies among residents of Kwara state. Mass sensitization of the populace on the dangers of rabies should be intensified. Such interventions should be targeted at the general public and dog owners.

Outbreak of Severe Vomiting in Dogs Associated with a Canine Enteric Coronavirus, United Kingdom.

The lack of population health surveillance for companion animal populations leaves them vulnerable to the effects of novel diseases without means of early detection. We present evidence on the effectiveness of a system that enabled early detection and rapid response a canine gastroenteritis outbreak in the United Kingdom. In January 2020, prolific vomiting among dogs was sporadically reported in the United Kingdom. Electronic health records from a nationwide sentinel network of veterinary practices confirmed a significant increase in dogs with signs of gastroenteric disease. Male dogs and dogs living with other vomiting dogs were more likely to be affected. Diet and vaccination status were not associated with the disease; however, a canine enteric coronavirus was significantly associated with illness. The system we describe potentially fills a gap in surveillance in neglected populations and could provide a blueprint for other countries.

Safety, biodistribution and viral shedding of oncolytic vaccinia virus TG6002 administered intravenously in healthy beagle dogs.

Oncolytic virotherapy is an emerging strategy that uses replication-competent viruses to kill tumor cells. We have reported the oncolytic effects of TG6002, a recombinant oncolytic vaccinia virus, in preclinical human xenograft models and canine tumor explants. To assess the safety, biodistribution and shedding of TG6002 administered by the intravenous route, we conducted a study in immune-competent healthy dogs. Three dogs each received a single intravenous injection of TG6002 at 105 PFU/kg, 106 PFU/kg or 107 PFU/kg, and one dog received three intravenous injections at 107 PFU/kg. The injections were well tolerated without any clinical, hematological or biochemical adverse events. Viral genomes were only detected in blood at the earliest sampling time point of one-hour post-injection at 107 PFU/kg. Post mortem analyses at day 35 allowed detection of viral DNA in the spleen of the dog which received three injections at 107 PFU/kg. Viral genomes were not detected in the urine, saliva or feces of any dogs. Seven days after the injections, a dose-dependent antibody mediated immune response was identified. In conclusion, intravenous administration of TG6002 shows a good safety profile, supporting the initiation of clinical trials in canine cancer patients as well as further development as a human cancer therapy.

Rabies in Tunisia: A spatio-temporal analysis in the region of CapBon-Nabeul.

Human rabies is a significant public health concern in Tunisia. However, the spatiotemporal spread pattern of rabies in dogs, the major reservoir and vector, and its determinants are poorly understood. We collected geographic locations and timeline of reported animal rabies cases in the region of CapBon (study area), for the years 2015-2019 and integrated them in Geographical Information System (GIS) approach to explore the spatio-temporal pattern of dog rabies. The results show that roads and irrigated areas can act as ecological corridors to viral spread. Our study showed that there was a significant seasonal variation in the number of cases of rabies recorded, with a strong peak in spring and lower peak in winter and summer. These findings may play a role in updating and directing public health policy, as well as providing opportunities for authorities to explore control options in time and space. A better knowledge of the ecology and dog population dynamics is still necessary and important to achieve an effective rabies control.

Rabies control in Liberia: Joint efforts towards zero by 30.

Despite declaration as a national priority disease, dog rabies remains endemic in Liberia, with surveillance systems and disease control activities still developing. The objective of these initial efforts was to establish animal rabies diagnostics, foster collaboration between all rabies control stakeholders, and develop a short-term action plan with estimated costs for rabies control and elimination in Liberia. Four rabies diagnostic tests, the direct fluorescent antibody (DFA) test, the direct immunohistochemical test (dRIT), the reverse transcriptase polymerase chain reaction (RT-PCR) assay and the rapid immunochromatographic diagnostic test (RIDT), were implemented at the Central Veterinary Laboratory (CVL) in Monrovia between July 2017 and February 2018. Seven samples (n=7) out of eight suspected animals were confirmed positive for rabies lyssavirus, and molecular analyses revealed that all isolates belonged to the Africa 2 lineage, subgroup H. During a comprehensive in-country One Health rabies stakeholder meeting in 2018, a practical workplan, a short-term action plan and an accurately costed mass dog vaccination strategy were developed. Liberia is currently at stage 1.5/5 of the Stepwise Approach towards Rabies Elimination (SARE) tool, which corresponds with countries that are scaling up local-level interventions (e.g. dog vaccination campaigns) to the national level. Overall an estimated 5.3 - 8 million USD invested over 13 years is needed to eliminate rabies in Liberia by 2030. Liberia still has a long road to become free from dog-rabies. However, the dialogue between all relevant stakeholders took place, and disease surveillance considerably improved through implementing rabies diagnosis at the CVL. The joint efforts of diverse national and international stakeholders laid important foundations to achieve the goal of zero dog-mediated human rabies deaths by 2030.