Detection of SARS-CoV-2 in Terrestrial Animals in Southern Nigeria: Potential Cases of Reverse Zoonosis.

Since SARS-CoV-2 caused the COVID-19 pandemic, records have suggested the occurrence of reverse zoonosis of pets and farm animals in contact with SARS-CoV-2-positive humans in the Occident. However, there is little information on the spread of the virus among animals in contact with humans in Africa. Therefore, this study aimed to investigate the occurrence of SARS-CoV-2 in various animals in Nigeria. Overall, 791 animals from Ebonyi, Ogun, Ondo, and Oyo States, Nigeria were screened for SARS-CoV-2 using RT-qPCR (n = 364) and IgG ELISA (n = 654). SARS-CoV-2 positivity rates were 45.9% (RT-qPCR) and 1.4% (ELISA). SARS-CoV-2 RNA was detected in almost all animal taxa and sampling locations except Oyo State. SARS-CoV-2 IgGs were detected only in goats from Ebonyi and pigs from Ogun States. Overall, SARS-CoV-2 infectivity rates were higher in 2021 than in 2022. Our study highlights the ability of the virus to infect various animals. It presents the first report of natural SARS-CoV-2 infection in poultry, pigs, domestic ruminants, and lizards. The close human-animal interactions in these settings suggest ongoing reverse zoonosis, highlighting the role of behavioral factors of transmission and the potential for SARS-CoV-2 to spread among animals. These underscore the importance of continuous monitoring to detect and intervene in any eventual upsurge.

Serological evidence of SARS-CoV-2 infection in dromedary camels and domestic bovids in Oman.

SARS-CoV-2 has demonstrated the ability to infect a wide range of animal species. Here, we investigated SARS-CoV-2 infection in livestock species in Oman and provided serological evidence of SARS-CoV-2 infection in cattle, sheep, goats, and dromedary camel using the surrogate virus neutralization and plaque reduction neutralization tests. To better understand the extent of SARS-CoV-2 infection in animals and associated risks, "One Health" epidemiological investigations targeting animals exposed to COVID-19 human cases should be implemented with integrated data analysis of the epidemiologically linked human and animal cases.

Safety and Immunogenicity of Recombinant Bacille Calmette-Guérin Strain VPM1002 and Its Derivatives in a Goat Model.

A more effective vaccine against tuberculosis than Bacille Calmette-Guérin (BCG) is urgently needed. BCG derived recombinant VPM1002 has been found to be more efficacious and safer than the parental strain in mice models. Newer candidates, such as VPM1002 Δpdx1 (PDX) and VPM1002 ΔnuoG (NUOG), were generated to further improve the safety profile or efficacy of the vaccine. Herein, we assessed the safety and immunogenicity of VPM1002 and its derivatives, PDX and NUOG, in juvenile goats. Vaccination did not affect the goats' health in regards to clinical/hematological features. However, all three tested vaccine candidates and BCG induced granulomas at the site of injection, with some of the nodules developing ulcerations approximately one month post-vaccination. Viable vaccine strains were cultured from the injection site wounds in a few NUOG- and PDX- vaccinated animals. At necropsy (127 days post-vaccination), BCG, VPM1002, and NUOG, but not PDX, still persisted at the injection granulomas. All strains, apart from NUOG, induced granuloma formation only in the lymph nodes draining the injection site. In one animal, the administered BCG strain was recovered from the mediastinal lymph nodes. Interferon gamma (IFN-γ) release assay showed that VPM1002 and NUOG induced a strong antigen-specific response comparable to that elicited by BCG, while the response to PDX was delayed. Flow cytometry analysis of IFN-γ production by CD4+, CD8+, and γδ T cells showed that CD4+ T cells of VPM1002- and NUOG-vaccinated goats produced more IFN-γ compared to BCG-vaccinated and mock-treated animals. In summary, the subcutaneous application of VPM1002 and NUOG induced anti-tuberculous immunity, while exhibiting a comparable safety profile to BCG in goats.

Enteropathy and bone marrow hypoplasia associated with presumptive albendazole toxicosis in a juvenile Boer goat.

Albendazole is a widely used anthelmintic drug that is labeled for the treatment of specific nematodes and flukes in ruminants. Albendazole is approved for the treatment of liver flukes in goats (10 mg/kg PO for a single dose), but is commonly used extra-label in situations in which parasite resistance is an issue. Albendazole toxicosis has been reported in pigeons, doves, alpacas, humans, dogs, and cats. Here we report an adverse event in a 6-mo-old goat associated with extra-label use of albendazole (35.7 mg/kg PO daily for 3 d). Clinicopathologic findings included severe diarrhea and death, with small intestinal crypt necrosis and dysplasia, and severe bone marrow hypoplasia. Microbial and molecular testing and transmission electron microscopy ruled out infectious organisms. The described pathologic changes are similar to those reported in other species that have experienced toxicosis associated with albendazole. To our knowledge, bone marrow and intestinal lesions associated with albendazole use in the goat have not been reported previously. Veterinarians should be aware of potential adverse events and toxicoses associated with anthelmintic drugs, especially as parasite resistance increases, and extra-label usage, and the use of such drugs without veterinary supervision, becomes more common.

Tick-Borne Encephalitis Virus RNA Found in Frozen Goat's Milk in a Family Outbreak.

Tick-borne encephalitis (TBE) is one of the commonest arthropod-borne viral diseases in Middle-East Europe and North Asia. The main reservoir of the virus is comprised of small rodents and domestic mammals with the common tick (Ixodes) being the usual vector. The clinical spectrum of TBE ranges from mild meningitis to severe meningoencephalomyelitis. This disease can lead to severe sequelae and has a mortality up to 2% in Europe. Even though the majority of cases are transmitted through bites of infected ticks, infections through ingestion of contaminated milk and dairy products from farms in endemic areas have been reported. We report a family outbreak of a febrile disease, initially suggestive of human-to-human infection, during the early summertime in Austria. Tick-borne encephalitis was diagnosed following consumption of unpasteurised goat's milk and the virus was subsequently detected in frozen milk samples. Although this is a rare manifestation of TBE, this case series shows that TBE should be included in the differential diagnosis of an outbreak of febrile disease, and a careful clinical history with reference to unpasteurized dairy products is crucial in order to prevent further disease spread. The best preventive measure is active immunisation of people living in, or travelling to, endemic areas.

Crimean-Congo Hemorrhagic Fever Outbreak in Refugee Settlement during COVID-19 Pandemic, Uganda, April 2021.

Crimean-Congo hemorrhagic fever (CCHF) was detected in 2 refugees living in a refugee settlement in Kikuube district, Uganda. Investigations revealed a CCHF IgG seroprevalence of 71.3% (37/52) in goats within the refugee settlement. This finding highlights the need for a multisectoral approach to controlling CCHF in humans and animals in Uganda.

Susceptibility of Domestic Goat (Capra aegagrus hircus) to Experimental Infection with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) B.1.351/Beta Variant.

A wide range of animal species are susceptible to the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Natural and/or experimental infections have been reported in pet, zoo, farmed and wild animals. Interestingly, some SARS-CoV-2 variants, such as B.1.1.7/Alpha, B.1.351/Beta, and B.1.1.529/Omicron, were demonstrated to infect some animal species not susceptible to classical viral variants. The present study aimed to elucidate if goats (Capra aegagrus hircus) are susceptible to the B.1.351/Beta variant. First, an in silico approach was used to predict the affinity between the receptor-binding domain of the spike protein of SARS-CoV-2 B.1.351/Beta variant and angiotensin-converting enzyme 2 from goats. Moreover, we performed an experimental inoculation with this variant in domestic goat and showed evidence of infection. SARS-CoV-2 was detected in nasal swabs and tissues by RT-qPCR and/or immunohistochemistry, and seroneutralisation was confirmed via ELISA and live virus neutralisation assays. However, the viral amount and tissue distribution suggest a low susceptibility of goats to the B.1.351/Beta variant. Therefore, although monitoring livestock is advisable, it is unlikely that goats play a role as SARS-CoV-2 reservoir species, and they are not useful surrogates to study SARS-CoV-2 infection in farmed animals.

Molecular survey for tick-borne pathogens and associated risk factors in sheep and goats in Kano Metropolis, Nigeria.

Tick-borne pathogens (TBPs) pose an increased health and productivity risk to livestock in sub-Saharan Africa. Information regarding TBPs infecting small ruminants in Kano metropolis is scarce. Therefore, we investigated the molecular epidemiology of tick-borne pathogens of economic importance from sheep and goats in Kano, Nigeria using Polymerase chain reaction (PCR). A total of 346 blood DNA samples were collected from small ruminants and analyzed for TBPs using PCR and sequencing. Risk of infection was determined for age, sex, breed and animal species. Our results indicate the absence of piroplasmids (Babesia/Theileria) and Rickettsia spp. infections. The overall prevalence for Anaplasma spp. was 9.25% (32/346) with a higher prevalence in goats 13.59% (25/184) compared with sheep 4.32% (7/162). With respect to age of animals, goats >4 years had the highest prevalence of 32.45% (11/37) which differs significantly (P = 0.0059) compared with other age categories. Cross breed goats had a prevalence of 15.63% (5/32) compared with Kano brown breed 14.08 (20/142). Sex significant difference (P = 0.029) was observed in the goats with females having the highest prevalence 20.89% (14/67) compared with males 9.40% (11/117). Furthermore, with regards to sheep, no significant difference (P > 0.05) was observed with respect to age and breed. Finally, no significant difference (P > 0.05) was observed with the prevalence of Anaplasma spp. due to Body condition score (BCS) in both sheep and goats. Conclusively, the occurrence of TBPs in small ruminants is low. Continuous efforts in tick control must be sustained to ensure high productive yield and reduced disease burden associated with TBPs of sheep and goats in Kano metropolis.

Research Relevant Background Lesions and Conditions: Ferrets, Dogs, Swine, Sheep, and Goats.

Animal models provide a valuable tool and resource for biomedical researchers as they investigate biological processes, disease pathogenesis, novel therapies, and toxicologic studies. Interpretation of animal model data requires knowledge not only of the processes/diseases being studied but also awareness of spontaneous conditions and background lesions in the model that can influence or even confound the study results. Species, breed/stock, sex, age, anatomy, physiology, diseases (noninfectious and infectious), and neoplastic processes are model features that can impact the results as well as study interpretation. Here, we review these features in several common laboratory animal species, including ferret, dog (beagle), pig, sheep, and goats.

Susceptibility of livestock to SARS-CoV-2 infection.

We report pilot studies to evaluate the susceptibility of common domestic livestock (cattle, sheep, goat, alpaca, rabbit, and horse) to intranasal infection with SARS-CoV-2. None of the infected animals shed infectious virus via nasal, oral, or faecal routes, although viral RNA was detected in several animals. Further, neutralizing antibody titres were low or non-existent one month following infection. These results suggest that domestic livestock are unlikely to contribute to SARS-CoV-2 epidemiology.

Prevalence of Cryptosporidium Infection in Sheep and Goat Flocks in China During 2010-2019: A Systematic Review and Meta-Analysis.

Cryptosporidiosis is an extensively contagious zoonotic waterborne disease caused by the genus Cryptosporidium and poses to be a danger to public health. Sheep and goats are an intermediate host of Cryptosporidium. Consequently, a first systematic review and meta-analysis are performed to assess the burden of the infection relative to the Cryptosporidium in sheep and goat flocks in China. Five databases were searched for relevant literature in accordance with the inclusion criteria until January 30, 2020. At last, a total of 33 qualified documents were included. We calculate the overall prevalence of Cryptosporidium (4.9%) in sheep and goats in China with the random-effects model. The prevalence after 2014 (4.6%) was higher than that before or in 2014 (2.8%). The pooled prevalence of Cryptosporidium in sheep and goats from Northern China (12.3%) was significantly higher (p < 0.05) than other regions. The infection rate of modified acid-fast staining (14.3%) was the highest among the detection methods. In age subgroups, the prevalence of Cryptosporidium in sheep and goats in 3 months or before was the highest (20.8%). Goats had a higher infection rate (5.9%) in species. The prevalence of large-scale farms (2.8%) was lower than free-ranging farms (4.4%). The medium quality level (6.4%) was the highest. Besides, geographical factors (such as latitude, longitude, height, precipitation, humidity, mean temperature, etc.) were further analyzed as potential risk factors of Cryptosporidium in sheep and goats. This meta-analysis indicates that the Cryptosporidium infection of Chinese sheep and goat flocks is general. Thus, it is necessary to further monitor the prevalence of Cryptosporidium, and the reasonable preventive strategy should be formulated on the basis of the geographical factors of different regions and the differences in sheep and goats' growth stages to reduce the prevalence of Cryptosporidium in sheep and goats.

MERS-CoV in sheep, goats, and cattle, United Arab Emirates, 2019: Virological and serological investigations reveal an accidental spillover from dromedaries.

The recent COVID-19 pandemic has demonstrated again the global threat posed by emerging zoonotic coronaviruses. During the past two decades alone, humans have experienced the emergence of several coronaviruses, such as SARS-CoV in 2003, MERS-CoV in 2012, and SARS-CoV-2 in 2019. To date, MERS-CoV has been detected in 27 countries, with a case fatality ratio of approximately 34.5%. Similar to other coronaviruses, MERS-CoV presumably originated from bats; however, the main reservoir and primary source of human infections are dromedary camels. Other species within the Camelidae family, such as Bactrian camels, alpacas, and llamas, seem to be susceptible to the infection as well, although to a lesser extent. In contrast, susceptibility studies on sheep, goats, cattle, pigs, chickens, and horses obtained divergent results. In the present study, we tested nasal swabs and/or sera from 55 sheep, 45 goats, and 52 cattle, collected at the largest livestock market in the United Arab Emirates, where dromedaries are also traded, for the presence of MERS-CoV nucleic acid by RT-qPCR, and for specific antibodies by immunofluorescence assay. All sera were negative for MERS-CoV-reactive antibodies, but the nasal swab of one sheep (1.8%) repeatedly tested positive for MERS-CoV nucleic acid. Next generation sequencing (NGS) of the complete N gene of the sheep-derived MERS-CoV revealed >99% nucleotide identity to MERS-CoV sequences of five dromedaries in nearby pens and to three reference sequences. The NGS sequence of the sheep-derived MERS-CoV was confirmed by conventional RT-PCR of a part of the N gene and subsequent Sanger sequencing. All MERS-CoV sequences clustered within clade B, lineage 5. In conclusion, our study shows that noncamelid livestock, such as sheep, goats, and cattle do not play a major role in MERS-CoV epidemiology. The one sheep that tested positive most likely reflects an accidental viral spillover event from infected dromedaries in nearby pens.

Evaluating angiotensin-converting enzyme 2-mediated SARS-CoV-2 entry across species.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic represents a global threat, and the interaction between the virus and angiotensin-converting enzyme 2 (ACE2), the primary entry receptor for SARS-CoV-2, is a key determinant of the range of hosts that can be infected by the virus. However, the mechanisms underpinning ACE2-mediated viral entry across species remains unclear. Using infection assay, we evaluated SARS-CoV-2 entry mediated by ACE2 of 11 different animal species. We discovered that ACE2 of Rhinolophus sinicus (Chinese rufous horseshoe bat), Felis catus (domestic cat), Canis lupus familiaris (dog), Sus scrofa (wild pig), Capra hircus (goat), and Manis javanica (Malayan pangolin) facilitated SARS-CoV-2 entry into nonsusceptible cells. Moreover, ACE2 of the pangolin also mediated SARS-CoV-2 entry, adding credence to the hypothesis that SARS-CoV-2 may have originated from pangolins. However, the ACE2 proteins of Rhinolophus ferrumequinum (greater horseshoe bat), Gallus gallus (red junglefowl), Notechis scutatus (mainland tiger snake), or Mus musculus (house mouse) did not facilitate SARS-CoV-2 entry. In addition, a natural isoform of the ACE2 protein of Macaca mulatta (rhesus monkey) with the Y217N mutation was resistant to SARS-CoV-2 infection, highlighting the possible impact of this ACE2 mutation on SARS-CoV-2 studies in rhesus monkeys. We further demonstrated that the Y217 residue of ACE2 is a critical determinant for the ability of ACE2 to mediate SARS-CoV-2 entry. Overall, these results clarify that SARS-CoV-2 can use the ACE2 receptors of multiple animal species and show that tracking the natural reservoirs and intermediate hosts of SARS-CoV-2 is complex.

CMTM3 overexpression promotes cell apoptosis while DHT promotes cell proliferation in hair follicle stem cells (HFSCs).

In Yangtze River Delta white goat, hypermethylation of CMTM3 leads to a decreased expression level in high quality brush hair. However, the regulation of CMTM3 expression and its function in hair follicle stem cells (HFSCs) remains largely unknown. In this study, we investigated the regulation of CMTM3 expression, function, and molecular mechanism in HFSCs. The re-expression of CMTM3 significantly suppressed the proliferation of HFSCs by inducing G1 cell cycle arrest and promoting apoptosis. Moreover, the downregulation of CMTM3 promoted HFSC proliferation. Treatment with sh_CMTM3 and incubation in a DHT culture medium had the most significant growth-promoting effect. It was hypothesized that transcriptome analysis using RNA sequencing (RNA-seq) in samples would enable the identification of unique protein-coding and non-coding genes that may help uncover the role of CMTM3. Multiple genes and pathways were involved in this process, including 168 common DEGs, such as CXCL8 and E-selectin, which is reportedly involved in multiple regulatory pathways. These results indicated that CMTM3 can function as HFSCs through the induction of a G1 cell cycle arrest and promoted apoptosis by mediating crosstalk between several pathways and transcription factors. Our data is available in the National Center for Biotechnology Information (NCBI) database with the accession number PRJNA657430.

In-silico Analysis of Angiotensin-converting Enzyme 2 (ACE2) of Livestock, Pet and Poultry Animals to Determine its Susceptibility to SARS-CoV- 2 Infection.

BACKGROUND: Novel coronavirus SARS-CoV-2 is responsible for the COVID-19 pandemic. It was first reported in Wuhan, China, in December 2019, and despite the tremendous efforts to control the disease, it has now spread almost all over the world. The interaction of SARSCoV- 2spike protein and its acceptor protein ACE2 is an important issue in determining viral host range and cross-species infection, while the binding capacity of spike protein to ACE2 of different species is unknown. OBJECTIVE: The present study has been conducted to determine the susceptibility of livestock, poultry and pets to SARS-CoV-2. METHODS: We evaluated the receptor-utilizing capability of ACE2s from various species by sequence alignment, phylogenetic clustering and protein-ligand interaction studies with the currently known ACE2s utilized by SARS-CoV-2. RESULT: In-silico study predicted that SARS-CoV-2 tends to utilize ACE2s of various animal species with varied possible interactions. The probability of the receptor utilization will be greater in horse and poor in chicken, followed by ruminants. CONCLUSION: Present study predicted that SARS-CoV-2 tends to utilize ACE2s of various livestock and poultry species with greater probability in equine and poor in chicken. The study may provide important insights into the animal models for SARS-CoV-2 and animal management for COVID- 19 control.

Molecular-based cross-species evaluation of bovine coronavirus infection in cattle, sheep and goats in Ghana.

BACKGROUND: Apart from the huge worldwide economic losses often occasioned by bovine coronavirus (BCoV) to the livestock industry, particularly with respect to cattle rearing, continuous surveillance of the virus in cattle and small ruminants is essential in monitoring variations in the virus that could enhance host switching. In this study, we collected rectal swabs from a total of 1,498 cattle, sheep and goats. BCoV detection was based on reverse transcriptase polymerase chain reaction. Sanger sequencing of the partial RNA-dependent RNA polymerase (RdRp) region for postive samples were done and nucleotide sequences were compared with homologous sequences from the GenBank. RESULTS: The study reports a BCoV prevalence of 0.3%, consisting of 4 positive cases; 3 goats and 1 cattle. Less than 10% of all the animals sampled showed clinical signs such as diarrhea and respiratory distress except for high temperature which occurred in > 1000 of the animals. However, none of the 4 BCoV positive animals manifested any clinical signs of the infection at the time of sample collection. Bayesian majority-rule cladogram comparing partial and full length BCoV RdRp genes obtained in the study to data from the GenBank revealed that the sequences obtained from this study formed one large monophyletic group with those from different species and countries. The goat sequences were similar to each other and clustered within the same clade. No major variations were thus observed between our isolates and those from elsewhere. CONCLUSIONS: Given that Ghana predominantly practices the extensive and semi-intensive systems of animal rearing, our study highlights the potential for spillover of BCoV to small ruminants in settings with mixed husbandry and limited separation between species.

Quantitative shadowgraphy of aerosol and droplet creation during oscillatory motion of the microkeratome amid COVID-19 and other infectious diseases.

PURPOSE: To quantify the atomization of liquid over the cornea during flap creation using microkeratome using high-speed shadowgraphy. SETTING: Laboratory study. DESIGN: Laboratory investigational study. METHOD: In an experimental setup, flap creation was performed on enucleated goat's eyes (n = 8) mounted on a stand using One Use-Plus SBK Moria microkeratome (Moria SA) to assess the spread of aerosols and droplets using high-speed shadowgraphy. Two conditions were computed. A constant airflow assumed uniform air velocity throughout the room. A decaying jet assumed that local air velocity at the site of measurements was smaller than the exit velocity from the air duct. RESULTS: With the advancement of the microkeratome across the wet corneal surface, the atomization of a balanced salt solution was recorded on shadowgraphy. The minimum droplet size was ∼90 µm. The maximum distance traversed was ∼1.8 m and ∼1.3 m assuming a constant airflow (setting of refractive surgery theater) and decaying jet condition (setting of an operating theater with air-handling unit), respectively. CONCLUSIONS: The microkeratome-assisted LASIK flap creation seemed to cause spread of droplets. The droplet diameters and velocities did not permit the formation of aerosols. Therefore, the risk of transmission of the virus to the surgeon and surgical personnel due to the microkeratome procedure seemed to be low.

Sero-prevalence, cross-species infection and serological determinants of prevalence of Bovine Coronavirus in Cattle, Sheep and Goats in Ghana.

Cattle, goats and sheep are dominant livestock species in sub-Saharan Africa, with sometimes limited information on the prevalence of major infectious diseases. Restrictions due to notifiable epizootics complicate the exchange of samples in surveillance studies and suggest that laboratory capacities should be established domestically. Bovine Coronavirus (BCoV) causes mainly enteric disease in cattle. Spillover to small ruminants is possible. Here we established BCoV serology based on a recombinant immunofluorescence assay for cattle, goats and sheep, and studied the seroprevalence of BCoV in these species in four different locations in the Greater Accra, Volta, Upper East, and Northern provinces of Ghana. The whole sampling and testing was organized and conducted by a veterinary school in Kumasi, Ashanti Region of Ghana. Among sampled sheep (n = 102), goats (n = 66), and cattle (n = 1495), the seroprevalence rates were 25.8 %, 43.1 % and 55.8 %. For cattle, seroprevalence was significantly higher on larger farms (82.2 % vs 17.8 %, comparing farms with >50 or <50 animals; p = 0.027). Highest prevalence was seen in the Northern province with dry climate, but no significant trend following the north-south gradient of sampling sites was detected. Our study identifies a considerable seroprevalence for BCoV in Ghana and provides further support for the spillover of BCoV to small ruminants in settings with mixed husbandry and limited separation between species.