Molecular detection using hybridization capture and next-generation sequencing reveals cross-species transmission of feline coronavirus type-1 between a domestic cat and a captive wild felid.

Feline coronavirus (FCoV) infection normally causes mild or subclinical signs and is common in domestic cats. However, in some cats, FCoV infection can also lead to the development of feline infectious peritonitis (FIP)-a typically lethal disease. FCoV has two serotypes or genotypes, FCoV-1 and FCoV-2, both of which can cause FIP. The main difference between the genotypes is the viral spike (S) protein that determines tropism and pathogenicity, crucial mechanisms in the development of FIP. Subclinical infection and FIP have both been reported in wild felids, including in threatened species. Due to the high genetic variability of the S gene and the technical challenges to sequencing it, detection and characterization of FCoV in wild felids have mainly centered on other more conserved genes. Therefore, the genotype causing FIP in most wild felids remains unknown. Here, we report a retrospective molecular epidemiological investigation of FCoV in a zoological institution in the U.Ss. In 2008, a domestic cat (Felis catus) and a Pallas' cat (Otocolobus manul) sharing the same room succumbed to FIP. Using in situ hybridization, we detected FCoV RNA in different tissues of both felids. Using hybridization capture and next-generation sequencing, we detected, sequenced, and characterized the whole genome of the FCoV infecting both felids. Our data show for the first time that FCoV-1 can be transmitted between domestic and wild felids and extends the known host range of FCoV-1. Our findings highlight the importance of identifying the genotype causing FIP, to develop effective control measures. IMPORTANCE: Feline coronavirus (FCoV) is highly prevalent in domestic cats worldwide and has also been reported in wild felids, including endangered species, in which it has caused substantial population declines. Characterizing the genetic diversity of FCoV is crucial due to recent reports of novel pathogenic recombinant variants causing high mortality in feral cats in Cyprus. In this retrospective molecular epidemiology study, we used archived samples collected in a zoological institution in the U.S. in which a domestic and a wild felid succumbed to FCoV. Using hybridization capture (HC) and next-generation sequencing, we show for the first time that FCoV can be naturally transmitted between domestic and wild felids. We demonstrate the efficacy of HC for detecting and sequencing the whole genome of FCoV, which is essential to characterize its different genotypes.

Two Lineages of Papillomaviruses Identified from Caracals (Caracal caracal) in South Africa.

Papillomaviruses (PV) infect epithelial cells and can cause hyperplastic or neoplastic lesions. In felids, most described PVs are from domestic cats (Felis catus; n = 7 types), with one type identified in each of the five wild felid species studied to date (Panthera uncia, Puma concolor, Leopardus wiedii, Panthera leo persica and Lynx rufus). PVs from domestic cats are highly diverse and are currently classified into three genera (Lambdapapillomavirus, Dyothetapapillomavirus, and Taupapillomavirus), whereas those from wild felids, although diverse, are all classified into the Lambdapapillomavirus genus. In this study, we used a metagenomic approach to identify ten novel PV genomes from rectal swabs of five deceased caracals (Caracal caracal) living in the greater Cape Town area, South Africa. These are the first PVs to be described from caracals, and represent six new PV types, i.e., Caracal caracal papillomavirus (CcarPV) 1-6. These CcarPV fall into two phylogenetically distinct genera: Lambdapapillomavirus, and Treisetapapillomavirus. Two or more PV types were identified in a single individual for three of the five caracals, and four caracals shared at least one of the same PV types with another caracal. This study broadens our understanding of wild felid PVs and provides evidence that there may be several wild felid PV lineages.

Identification and genomic characterization of feline calicivirus from a leopard cat (Prionailurus bengalensis) in Taiwan.

The leopard cat (Prionailurus bengalensis) is an endangered wildlife that is protected under Taiwan's regulations. The body of a road-killed leopard cat was found to contain sequences of feline calicivirus (FCV), designated W109-1443. Analysis of the complete genomic sequence revealed that it shared approximately 81% similarity with a Chinese strain of FCV found in a domestic cat. Phylogenetic analysis of the VP1 gene indicated that the W109-1443 isolate belonged to genogroup II. Recombination analysis revealed that the W109-1443 isolate may have resulted from recombination between two FCV strains. Given the potential impact of FCV on the health and survival of wild felids, further investigation is necessary to assess its pathogenicity in the leopard cat population.

Mortality associated with SARS-CoV-2 in nondomestic felids.

Between September and November 2021, 5 snow leopards (Panthera uncia) and 1 lion (Panthera leo) were naturally infected with severe acute respiratory coronavirus 2 (SARS-CoV-2) and developed progressive respiratory disease that resulted in death. Severe acute respiratory syndrome coronavirus 2 sequencing identified the delta variant in all cases sequenced, which was the predominant human variant at that time. The time between initial clinical signs and death ranged from 3 to 45 days. Gross lesions in all 6 cats included nasal turbinate hyperemia with purulent discharge and marked pulmonary edema. Ulcerative tracheitis and bronchitis were noted in 4 cases. Histologically, there was necrotizing and ulcerative rhinotracheitis and bronchitis with fibrinocellular exudates and fibrinosuppurative to pyogranulomatous bronchopneumonia. The 4 cats that survived longer than 8 days had fungal abscesses. Concurrent bacteria were noted in 4 cases, including those with more acute disease courses. Severe acute respiratory syndrome coronavirus 2 was detected by in situ hybridization using probes against SARS-CoV-2 spike and nucleocapsid genes and by immunohistochemistry. Viral nucleic acid and protein were variably localized to mucosal and glandular epithelial cells, pneumocytes, macrophages, and fibrinocellular debris. Based on established criteria, SARS-CoV-2 was considered a contributing cause of death in all 6 cats. While mild clinical infections are more common, these findings suggest that some SARS-CoV-2 variants may cause more severe disease and that snow leopards may be more severely affected than other felids.

Complex evolutionary history of felid anelloviruses.

Anellovirus infections are highly prevalent in mammals, however, prior to this study only a handful of anellovirus genomes had been identified in members of the Felidae family. Here we characterise anelloviruses in pumas (Puma concolor), bobcats (Lynx rufus), Canada lynx (Lynx canadensis), caracals (Caracal caracal) and domestic cats (Felis catus). The complete anellovirus genomes (n = 220) recovered from 149 individuals were diverse. ORF1 protein sequence similarity network analysis coupled with phylogenetic analysis, revealed two distinct clusters that are populated by felid-derived anellovirus sequences, a pattern mirroring that observed for the porcine anelloviruses. Of the two-felid dominant anellovirus groups, one includes sequences from bobcats, pumas, domestic cats and an ocelot, and the other includes sequences from caracals, Canada lynx, domestic cats and pumas. Coinfections of diverse anelloviruses appear to be common among the felids. Evidence of recombination, both within and between felid-specific anellovirus groups, supports a long coevolution history between host and virus.

FELINE CORONAVIRUS AND FELINE INFECTIOUS PERITONITIS IN NONDOMESTIC FELID SPECIES.

Feline coronavirus (FCoV) is reported worldwide and known to cause disease in domestic and nondomestic felid species. Although FCoV often results in mild to inapparent disease, a small subset of cats succumb to the fatal, systemic disease feline infectious peritonitis (FIP). An outbreak of FIP in Cheetahs (Acinonyx jubatus) in a zoological collection demonstrated the devastating effect of FCoV introduction into a naïve group of animals. In addition to cheetahs, FIP has been described in European wildcats (Felis silvestris), a tiger (Panthera tigris), a mountain lion (Puma concolor), and lion (Panthera leo). This paper reviews the reported cases of FIP in nondomestic felid species and highlights the surveys of FCoV in populations of nondomestic felids.

Natural infection of parvovirus in wild fishing cats (Prionailurus viverrinus) reveals extant viral localization in kidneys.

Carnivore protoparvovirus-1 (CPPV-1), a viral species containing feline panleukopenia virus (FPV) and canine parvovirus (CPV) variants, are widely spread among domestic and wild carnivores causing systemic fatal diseases. Wild fishing cats (Prionailurus viverrinus), a globally vulnerable species, have been found dead. Postmortem examination of the carcasses revealed lesions in intestine, spleen and kidney. CPPV-1 antigen identification in these tissues, using polymerase chain reaction (PCR) and immunohistochemistry (IHC), supported the infection by the virus. PCR- and IHC-positivity in kidney tissues revealed atypical localization of the virus while in situ hybridization (ISH) and transmission electron microscopy (TEM) with the pop-off technique confirmed the first description of viral localization in kidneys. Complete genome characterization and deduced amino acid analysis of the obtained CPPV-1 from the fishing cats revealed FPV as a causative agent. The detected FPV sequences showed amino acid mutations at I566M and M569R in the capsid protein. Phylogenetic and evolutionary analyses of complete coding genome sequences revealed that the fishing cat CPPV-1 genomes are genetically clustered to the FPV genomes isolated from domestic cats in Thailand. Since the 1970s, these genomes have also been shown to share a genetic evolution with Chinese FPV strains. This study is the first evidence of CPPV-1 infection in fishing cats and it is the first to show its localization in the kidneys. These findings support the multi-host range of this parvovirus and suggest fatal CPPV-1 infections may result in other vulnerable wild carnivores.

Identification of Novel Feline Paramyxoviruses in Guignas (Leopardus guigna) from Chile.

The family of paramyxoviruses has received growing attention as several new species have been identified recently, notably two different clusters in domestic cats, designated as feline morbillivirus (FeMV) and feline paramyxovirus (FPaV). Their phylogenetic origin and whether wild felids also harbor these viruses are currently unknown. Kidney samples from 35 guignas (Leopardus guigna), a wild felid from Chile, were investigated for paramyxoviruses using consensus-RT-PCR. In addition, thirteen serum samples of guignas were screened for the presence of FeMV-specific antibodies by an immunofluorescence assay (IFA). Viral RNA was detected in 31% of the kidney samples. Phylogenetic analyses revealed two well-supported clusters, related to isolates from domestic cats, rodents and bats. No significant histopathology changes were recorded in infected guignas. Serology identified two samples which were positive for FeMV-specific antibodies. Our study highlights the diversity of paramyxovirus infections in felids with special emphasis on guignas from Chile.

Leopardus wiedii Papillomavirus type 1, a novel papillomavirus species in the tree ocelot, suggests Felidae Lambdapapillomavirus polyphyletic origin and host-independent evolution.

The limited knowledge on Papillomavirus diversity (particularly in wild animal species) influences the accuracy of PVs phylogeny and their evolutionary history, and hinders the comprehension of PVs pathogenicity, especially the mechanism of virus - related cancer progression. This study reports the identification of Leopardus wiedii Papillomavirus type 1 (LwiePV1), the first PV type within Lambdapapillomavirus in a Leopardus host. LwiePV1 full genome sequencing allowed the investigation of its taxonomic position and phylogeny. Based on results, LwiePV1 should be assigned to a novel PV species providing evidence for a polyphyletic origin of feline lambda PVs, and representing an exception to codivergence between feline lambda PVs and their hosts. Results improve our knowledge on PV diversity and pave the way to future studies investigating biological and evolutionary features of animal PVs.

Immunohistochemical identification of antigens of canine distemper virus in neotropical felids from Southern Brazil.

The pathologic and immunohistochemical findings associated with infections due to canine distemper virus (CDV) are described in the cougar (Puma concolor), margay (Leopardus wiedii) and jaguarundi (Herpailurus yagouaroundi) from Southern Brazil. Tissue sections of the neotropical felids (n = 3) that died at the Bela Vista Sanctuary, Paraná, Southern Brazil were routinely processed for histopathology to identify possible histopathologic patterns associated with infections due to CDV. Selected formalin-fixed paraffin embedded tissue sections of the lungs and urinary bladder were used in immunohistochemical assays designed to identify the antigens of CDV. The main histopathologic patterns identified were interstitial pneumonia in the margay and jaguarundi, while ballooning degeneration of the transitional epithelium of the urinary bladder was observed in the cougar. Positive immunoreactivity to antigens of CDV was identified within intralesional sections of the lungs of the two wild felids with interstitial pneumonia and in the degenerated urothelium of the cougar. These findings indicate that these neotropical cats were infected by a viral infectious disease pathogen common to the domestic dog and add to the few documented descriptions of CDV-induced infections in wildlife from Brazil.

A systematic review of Toxoplasma gondii genotypes and feline: Geographical distribution trends.

Toxoplasma gondii (T. gondii) is well known for its ability to virtually infect all warm-blooded vertebrates. Although felines as the definitive hosts have an important role in the epidemiology of toxoplasmosis, there are few descriptions of genetic diversity in the world. The present review study aimed to describe the population structure of T. gondii in these animal species. For the purpose of the study, five English language databases reporting data on T. gondii genotyping in cats were searched within March-June 2019. This study is registered on the site of CAMARADES-NC3Rs (15-Jan-2018). The searching process resulted in the inclusion of 50 reports published from 1992 to June 2019. The data revealed that 47,390 samples were genotyped into 662 T. gondii DNA/isolates. Globally, atypical genotypes were predominant (47.7%, n = 316); in addition, Type II clonal strains were the second most common genotype (37%, n = 244). These results suggested an epidemic population structure in America and Asia, and a clonal population structure in Europe and Africa. Genotype #3 was found to be dominant in Africa, Europe and Oceania continents. Furthermore, genotypes #9 and #5 were prevalent in Asia and America, respectively. Additionally, genotypes #2, #3, #5 and #20 were common genotypes in domestic and sylvatic cycles from family Felidae. Collectively, this systematic review indicated a large degree of genetic diversity and circulation of mouse-virulent T. gondii strains in this family. However, further studies are necessary to better understand the population structure of T. gondii in these animal species and determine the significance of their features.

Distribution of Carnivore protoparvovirus 1 in free-living leopard cats (Prionailurus bengalensis chinensis) and its association with domestic carnivores in Taiwan.

Carnivore protoparvovirus 1 (CPPV-1) is widespread among free-living carnivores, and CPPV-1 infection may directly or indirectly impact on the population of endangered carnivore species. In this study, we used molecular screening of viral capsid protein 2 (VP2) from 2015 to 2017, to assess the prevalence of CPPV-1 infection in 9 live-trapped (LT) and 17 vehicle collision (VC)-affected free-living leopard cats (Prionailurus bengalensis chinensis). In addition, we conducted the phylogenetic analysis to evaluate the possible transmission of CPPV-1 between domestic carnivores and leopard cats. We identified the circulation of feline parvovirus and variants of canine parvovirus (CPV), including CPV-2a, CPV-2b, and CPV-2c, in the free-living leopard cat population. The partial sequences of different variants of VP2 obtained from the leopard cats were identical with those obtained from the domestic dogs and cats in Taiwan. Our result suggested that CPPV-1 was currently transmitted between domestic carnivores and leopard cats in Taiwan. A plan of conservation measures based on vaccination program for domestic carnivores, strict controls on populations of free-living dogs and cats and limiting road development only to low-risk areas for leopard cats should be encouraged.

Serological evidence of influenza virus infection in captive wild felids, Thailand.

Influenza virus is known to affect wild felids. To explore the prevalence of influenza viruses in these animal species, 196 archival sera from 5 felid species including Panthera tigris (N=147), Prionailurus viverrinus (N=35), Panthera leo (N=5), Pardofelis temminckii (N=8) and Neofelis nebulosa (N=1) collected between 2011 and 2015 in 10 provinces of Thailand were determined for the presence of antibody to avian and human influenza viruses. Blocking enzyme-linked immunosorbent (ELISA) assay and hemagglutination inhibition (HI) assay were employed as the screening tests, which the serum samples with HI antibody titers ≥20 were further confirmed by cytopathic effect/hemagglutination based-microneutralization (CPE/HA-based microNT) test. Based on HI and microNT assays, the seropositive rates of low pathogenic avian influenza (LPAI) H5 virus, highly pathogenic avian influenza (HPAI) H5 virus and human H1 virus were 1.53% (3/196), 2.04% (4/196) and 6.63% (13/196), respectively. In addition, we also found antibody against both LPAI H5 virus and HPAI H5 virus in 2 out of 196 tested sera (1.02%). Evidences of influenza virus infection were found in captive P. tigris in Kanchanaburi, Nakhon Sawan and Ratchaburi provinces of Thailand. The findings of our study highlights the need of a continuous active surveillance program of influenza viruses in wild felid species.

Canine parvovirus detected from a serval (Leptailurus serval) in South Africa.

Canine parvovirus first emerged in domestic dogs (Canis familiaris), most likely as a variant of the feline panleucopaenia virus. Relatively recently, canine parvovirus-2a and canine parvovirus-2b infections have been identified in both symptomatic and asymptomatic domestic cats, while canine parvovirus infections have also been demonstrated in wild felids. This report documents the first known case of canine parvovirus-2b detected in unvaccinated serval (Leptailurus serval) from South Africa. The serval presented with clinical signs of vomiting, anorexia and diarrhoea that responded to symptomatic treatment. Two weeks later, severe leucopaenia, thrombocytopenia and death occurred. Typical enteric histological lesions of parvovirus infection were not observed on histopathological examination of the small intestine; however, histological lesions consistent with septicaemia were present. Canine parvovirus was detected in formalin-fixed paraffin-embedded small intestine using polymerase chain reaction. Phylogenetic analysis of the sequence of the canine parvovirus viral capsid protein gene showed similarities between the sample from the serval and canine parvovirus-2b isolates from domestic dogs in Argentina and South Africa. A case of canine parvovirus-2b in a domestic dog from South Africa in 2012 that fell within the same clade as the serval sample appears distantly related because of the long branch length. The significance of these findings is explored. More extensive surveys of canine parvovirus in domestic and wild felids and canids are needed to understand the epidemiology of canine parvovirus in non-domestic felids in South Africa.

Serological evidence of H5-subtype influenza A virus infection in indigenous avian and mammalian species in Korea.

In Korea, H5-subtype highly pathogenic avian influenza (HPAI) has caused huge economic losses in poultry farms through outbreaks of H5N1 since 2003, H5N8 since 2013 and H5N6 since 2016. Although it was reported that long-distance migratory birds may play a major role in the global spread of avian influenza viruses (AIVs), transmission from such birds to poultry has not been confirmed. Intermediate hosts in the wild also may be a potential factor in viral transmission. Therefore, a total of 367 serum samples from wild animals were collected near major migratory bird habitats from 2011 to 2016 and tested by AIV-specific blocking ELISA and hemagglutination inhibition (HI) test. Two mammalian and eight avian species were seropositive according to the ELISA test. Among these, two mammalian (Hydropotes inermis and Prionailurus bengalensis) and three avian (Aegypius monachus, Cygnus cygnus, and Bubo bubo) species showed high HI titres (> 1,280) against one or two H5-subtype AIVs. As H. inermis (water deer), P. bengalensis (leopard cat), and B. bubo (Eurasian eagle owl) are indigenous animals in Korea, evidence of H5-subtype AIV in these animals implies that continuous monitoring of indigenous animals should be followed to understand interspecies transmission ecology of H5-subtype influenza viruses.

The effects of demographic, social, and environmental characteristics on pathogen prevalence in wild felids across a gradient of urbanization.

Transmission of pathogens among animals is influenced by demographic, social, and environmental factors. Anthropogenic alteration of landscapes can impact patterns of disease dynamics in wildlife populations, increasing the potential for spillover and spread of emerging infectious diseases in wildlife, human, and domestic animal populations. We evaluated the effects of multiple ecological mechanisms on patterns of pathogen exposure in animal populations. Specifically, we evaluated how ecological factors affected the prevalence of Toxoplasma gondii (Toxoplasma), Bartonella spp. (Bartonella), feline immunodeficiency virus (FIV), and feline calicivirus (FCV) in bobcat and puma populations across wildland-urban interface (WUI), low-density exurban development, and wildland habitat on the Western Slope (WS) and Front Range (FR) of Colorado during 2009-2011. Samples were collected from 37 bobcats and 29 pumas on the WS and FR. As predicted, age appeared to be positively related to the exposure to pathogens that are both environmentally transmitted (Toxoplasma) and directly transmitted between animals (FIV). In addition, WS bobcats appeared more likely to be exposed to Toxoplasma with increasing intraspecific space-use overlap. However, counter to our predictions, exposure to directly-transmitted pathogens (FCV and FIV) was more likely with decreasing space-use overlap (FCV: WS bobcats) and potential intraspecific contacts (FIV: FR pumas). Environmental factors, including urbanization and landscape covariates, were generally unsupported in our models. This study is an approximation of how pathogens can be evaluated in relation to demographic, social, and environmental factors to understand pathogen exposure in wild animal populations.

Epidemiologic survey of feline leukemia virus in domestic cats on Tsushima Island, Japan: management strategy for Tsushima leopard cats.

The Tsushima leopard cat (TLC) Prionailurus bengalensis euptilurus, a subspecies of P. bengalensis, is designated a National Natural Monument of Japan, and lives only on Tsushima Island, Nagasaki Prefecture, Japan. TLCs are threatened by various infectious diseases. Feline leukemia virus (FeLV) causes a serious infectious disease with a poor prognosis in cats. Therefore, the transmission of FeLV from Tsushima domestic cats (TDCs) to TLCs may threaten the TLC population. We investigated the FeLV infection status of both TDCs and TLCs on Tsushima Island by screening blood samples for FeLV p27 antigen and using PCR to amplify the full-length FeLV env gene. The prevalence of FeLV was 6.4% in TDCs and 0% in TLCs. We also demonstrated that the virus can replicate in the cells of TLCs, suggesting its potential cross-species transmission. The viruses in TDCs were classified as genotype I/clade 3, which is prevalent on a nearby island, based on previous studies of FeLV genotypes and FeLV epidemiology. The FeLV viruses identified on Tsushima Island can be further divided into 2 lineages within genotype I/clade 3, which are geographically separated in Kamijima and Shimojima, indicating that FeLV may have been transmitted to Tsushima Island at least twice. Monitoring FeLV infection in the TDC and TLC populations is highly recommended as part of the TLC surveillance and management strategy.

Molecular detection of viral agents in free-ranging and captive neotropical felids in Brazil.

We describe molecular testing for felid alphaherpesvirus 1 (FHV-1), carnivore protoparvovirus 1 (CPPV-1), feline calicivirus (FCV), alphacoronavirus 1 (feline coronavirus [FCoV]), feline leukemia virus (FeLV), feline immunodeficiency virus (FIV), and canine distemper virus (CDV) in whole blood samples of 109 free-ranging and 68 captive neotropical felids from Brazil. Samples from 2 jaguars ( Panthera onca) and 1 oncilla ( Leopardus tigrinus) were positive for FHV-1; 2 jaguars, 1 puma ( Puma concolor), and 1 jaguarundi ( Herpairulus yagouaroundi) tested positive for CPPV-1; and 1 puma was positive for FIV. Based on comparison of 103 nucleotides of the UL24-UL25 gene, the FHV-1 sequences were 99-100% similar to the FHV-1 strain of domestic cats. Nucleotide sequences of CPPV-1 were closely related to sequences detected in other wild carnivores, comparing 294 nucleotides of the VP1 gene. The FIV nucleotide sequence detected in the free-ranging puma, based on comparison of 444 nucleotides of the pol gene, grouped with other lentiviruses described in pumas, and had 82.4% identity with a free-ranging puma from Yellowstone Park and 79.5% with a captive puma from Brazil. Our data document the circulation of FHV-1, CPPV-1, and FIV in neotropical felids in Brazil.

CANINE DISTEMPER IN A VACCINATED SNOW LEOPARD ( PANTHERA UNCIA).

A 6-yr-old male snow leopard ( Panthera uncia) presented with acute seizures, hyperthermia, and tachypnea. Because of a diagnosis of anuric renal failure, the animal was euthanized. On histopathologic examination, numerous intralesional intracytoplasmic and intranuclear inclusions were found in the lungs, lymph nodes, and stomach. Positive immunohistochemical staining for canine distemper virus (CDV) was found in the lungs and, to a lesser extent, in the lymph nodes and brain. Molecular testing yielded a CDV H gene sequence that was closely related to CDV isolates concurrently found in wild raccoons from adjacent forested areas. The leopard had been vaccinated once against CDV with the use of a recombinant canarypox-vectored live vaccine during a routine wellness examination 12 wk prior to death. Serial serum neutralization titers performed on banked serum collected between vaccination and death showed poor serologic response to the vaccine. This case demonstrates a probable failure of protection against naturally occurring CDV.