Endogenous retrovirus ERV-DC8 highly integrated in domestic cat populations is a replication-competent provirus.

Endogenous retroviruses (ERVs) are remnants of ancient retroviral infections in vertebrate genomes and are inherited by offspring. ERVs can produce pathogenic viruses through gene mutations or recombination. ERVs in domestic cats (ERV-DCs) generate feline leukemia virus subgroup D (FeLV-D) through viral recombination. Herein, we characterized the locus ERV-DC8, on chromosome B1, as an infectious replication-competent provirus. ERV-DC8 infected several cell lines, including human cells. Transmission electron microscopy of ERV-DC8 identified the viral release as a Gammaretrovirus. ERV-DC8 was identified as the FeLV-D viral interference group, with feline copper transporter 1 as its viral receptor. Insertional polymorphism analysis showed high ERV-DC8 integration in domestic cats. This study highlights the role, pathogenicity, and evolutionary relationships between ERVs and their hosts.

Feline Foamy Virus Transmission in Tsushima Leopard Cats (Prionailurus bengalensis euptilurus) on Tsushima Island, Japan.

Tsushima leopard cats (TLC; Prionailurus bengalensis euptilurus) only inhabit Tsushima Island, Nagasaki, Japan and are critically endangered and threatened by infectious diseases. The feline foamy virus (FFV) is widely endemic in domestic cats. Therefore, its transmission from domestic cats to TLCs may threaten the TLC population. Thus, this study aimed to assess the possibility that domestic cats could transmit FFV to TLCs. Eighty-nine TLC samples were screened, and FFV was identified in seven (7.86%). To assess the FFV infection status of domestic cats, 199 domestic cats were screened; 14.07% were infected. The phylogenetic analysis revealed that the FFV partial sequence from domestic cats and TLC sequences clustered in one clade, suggesting that the two populations share the same strain. The statistical data minimally supported the association between increased infection rate and sex (p = 0.28), indicating that FFV transmission is not sex dependent. In domestic cats, a significant difference was observed in FFV detection in feline immunodeficiency virus (p = 0.002) and gammaherpesvirus1 infection statuses (p = 0.0001) but not in feline leukemia virus infection status (p = 0.21). Monitoring FFV infection in domestic cats and TLC populations is highly recommended as part of TLC surveillance and management strategies.

Distribution of infectious endogenous retroviruses in mixed-breed and purebred cats.

Endogenous retroviruses of domestic cats (ERV-DCs) are members of the genus Gammaretrovirus that infect domestic cats (Felis silvestris catus). Uniquely, domestic cats harbor replication-competent proviruses such as ERV-DC10 (ERV-DC18) and ERV-DC14 (xenotropic and nonecotropic viruses, respectively). The purpose of this study was to assess invasion by two distinct infectious ERV-DCs, ERV-DC10 and ERV-DC14, in domestic cats. Of a total sample of 1646 cats, 568 animals (34.5%) were positive for ERV-DC10 (heterozygous: 377; homozygous: 191), 68 animals (4.1%) were positive for ERV-DC14 (heterozygous: 67; homozygous: 1), and 10 animals (0.6%) were positive for both ERV-DC10 and ERV-DC14. ERV-DC10 and ERV-DC14 were detected in domestic cats in Japan as well as in Tanzania, Sri Lanka, Vietnam, South Korea and Spain. Breeding cats, including Singapura, Norwegian Forest and Ragdoll cats, showed high frequencies of ERV-DC10 (60-100%). By contrast, ERV-DC14 was detected at low frequency in breeding cats. Our results suggest that ERV-DC10 is widely distributed while ERV-DC14 is maintained in a minor population of cats. Thus, ERV-DC10 and ERV-DC14 have invaded cat populations independently.

Reduced Folate Carrier: an Entry Receptor for a Novel Feline Leukemia Virus Variant.

Feline leukemia virus (FeLV) is horizontally transmitted among cats and causes a variety of hematopoietic disorders. Five subgroups of FeLV, A to D and T, each with distinct receptor usages, have been described. Recently, we identified a new FeLV Env (TG35-2) gene from a pseudotyped virus that does not belong to any known subgroup. FeLV-A is the primary virus from which other subgroups have emerged via mutation or recombination of the subgroup A env gene. Retrovirus entry into cells is mediated by the interaction of envelope protein (Env) with specific cell surface receptors. Here, phenotypic screening of a human/hamster radiation hybrid panel identified SLC19A1, a feline reduced folate carrier (RFC) and potential receptor for TG35-2-phenotypic virus. RFC is a multipass transmembrane protein. Feline and human RFC cDNAs conferred susceptibility to TG35-2-pseudotyped virus when introduced into nonpermissive cells but did not render these cells permissive to other FeLV subgroups or feline endogenous retrovirus. Moreover, human cells with genomic deletion of RFC were nonpermissive for TG35-2-pseudotyped virus infection, but the introduction of feline and human cDNAs rendered them permissive. Mutation analysis of FeLV Env demonstrated that amino acid substitutions within variable region A altered the specificity of the Env-receptor interaction. We isolated and reconstructed the full-length infectious TG35-2-phenotypic provirus from a naturally FeLV-infected cat, from which the FeLV Env (TG35-2) gene was previously isolated, and compared the replication of the virus in hematopoietic cell lines with that of FeLV-A 61E by measuring the viral RNA copy numbers. These results provide a tool for further investigation of FeLV infectious disease.IMPORTANCE Feline leukemia virus (FeLV) is a member of the genus Gammaretrovirus, which causes malignant diseases in cats. The most prevalent FeLV among cats is FeLV subgroup A (FeLV-A), and specific binding of FeLV-A Env to its viral receptor, thiamine transporter feTHTR1, is the first step of infection. In infected cats, novel variants of FeLV with altered receptor specificity for viral entry have emerged by mutation or recombination of the env gene. A novel FeLV variant arose from a subtle mutation of FeLV-A Env, which altered the specific interaction of the virus with its receptor. RFC, a folate transporter, is a potential receptor for the novel FeLV variant. The perturbation of specific retrovirus-receptor interactions under selective pressure by the host results in the emergence of novel viruses.

Identification of Felis catus Gammaherpesvirus 1 in Tsushima Leopard Cats (Prionailurus bengalensis euptilurus) on Tsushima Island, Japan.

Felis catus gammaherpesvirus 1 (FcaGHV1) is a widely endemic infection of domestic cats. Current epidemiological data identify domestic cats as the sole natural host for FcaGHV1. The Tsushima leopard cat (TLC; Prionailurus bengalensis euptilurus) is a critically endangered species that lives only on Tsushima Island, Nagasaki, Japan. Nested PCR was used to test the blood or spleen of 89 TLCs for FcaGHV1 DNA; three (3.37%; 95% CI, 0.70⁻9.54) were positive. For TLC management purposes, we also screened domestic cats and the virus was detected in 13.02% (95% CI, 8.83⁻18.27) of 215 cats. Regarding phylogeny, the partial sequences of FcaGHV1 from domestic cats and TLCs formed one cluster, indicating that similar strains circulate in both populations. In domestic cats, we found no significant difference in FcaGHV1 detection in feline immunodeficiency virus-infected (p = 0.080) or feline leukemia virus-infected (p = 0.163) cats, but males were significantly more likely to be FcaGHV1 positive (odds ratio, 5.86; 95% CI, 2.27⁻15.14) than females. The higher frequency of FcaGHV1 detection in domestic cats than TLCs, and the location of the viral DNA sequences from both cats within the same genetic cluster suggests that virus transmission from domestic cats to TLCs is likely.

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.

Existence of Two Distinct Infectious Endogenous Retroviruses in Domestic Cats and Their Different Strategies for Adaptation to Transcriptional Regulation.

UNLABELLED: Endogenous retroviruses (ERVs) are the remnants of ancient retroviral infections of germ cells. Previous work identified one of the youngest feline ERV groups, ERV-DC, and reported that two ERV-DC loci, ERV-DC10 and ERV-DC18 (ERV-DC10/DC18), can replicate in cultured cells. Here, we identified another replication-competent provirus, ERV-DC14, on chromosome C1q32. ERV-DC14 differs from ERV-DC10/DC18 in its phylogeny, receptor usage, and, most notably, transcriptional activities; although ERV-DC14 can replicate in cultured cells, it cannot establish a persistent infection owing to its low transcriptional activity. Furthermore, we examined ERV-DC transcription and its regulation in feline tissues. Quantitative reverse transcription-PCR (RT-PCR) detected extremely low ERV-DC10 expression levels in feline tissues, and bisulfite sequencing showed that 5' long terminal repeats (LTRs) of ERV-DC10/DC18 are significantly hypermethylated in feline blood cells. Reporter assays found that the 5'-LTR promoter activities of ERV-DC10/DC18 are high, whereas that of ERV-DC14 is low. This difference in promoter activity is due to a single substitution from A to T in the LTR, and reverse mutation at this nucleotide in ERV-DC14 enhanced its replication and enabled it to persistently infect cultured cells. Therefore, ERV-DC LTRs can be divided into two types based on this nucleotide, the A type or T type, which have strong or attenuated promoter activity, respectively. Notably, ERV-DCs with T-type LTRs, such as ERV-DC14, have expanded in the cat genome significantly more than A-type ERV-DCs, despite their low promoter activities. Our results provide insights into how the host controls potentially infectious ERVs and, conversely, how ERVs adapt to and invade the host genome. IMPORTANCE: The domestic cat genome contains many endogenous retroviruses, including ERV-DCs. These ERV-DCs have been acquired through germ cell infections with exogenous retroviruses. Some of these ERV-DCs are still capable of producing infectious virions. Hosts must tightly control these ERVs because replication-competent viruses in the genome pose a risk to the host. Here, we investigated how ERV-DCs are adapted by their hosts. Replication-competent viruses with strong promoter activity, such as ERV-DC10 and ERV-DC18, were suppressed by promoter methylation in LTRs. On the other hand, replication-competent viruses with weak promoter activity, such as ERV-DC14, seemed to escape strict control via promoter methylation by the host. Interestingly, ERV-DCs with weak promoter activity, such as ERV-DC14, have expanded in the cat genome significantly more than ERV-DCs with strong promoter activity. Our results improve the understanding of the host-virus conflict and how ERVs adapt in their hosts over time.

Novel Feline Leukemia Virus Interference Group Based on the env Gene.

Feline leukemia virus (FeLV) subgroups have emerged in infected cats via the mutation or recombination of the env gene of subgroup A FeLV (FeLV-A), the primary virus. We report the isolation and characterization of a novel env gene, TG35-2, and report that the TG35-2 pseudotype can be categorized as a novel FeLV subgroup. The TG35-2 envelope protein displays strong sequence identity to FeLV-A Env, suggesting that selection pressure in cats causes novel FeLV subgroups to emerge.