Genetic trends in parrot Bornavirus: a clinical analysis.

Parrot Bornavirus (PaBV) has been reported to cause indigestion and other wasting symptoms such as weight loss and lethargy. The pathogenesis of PaBV has yet to be fully elucidated. This study reports PaBV infections in South Korea and suggests a trend in the genetic information gathered from clinical cases. A total of 487 birds with or without clinical symptoms were tested for bornavirus. Twelve of 361 asymptomatic birds tested positive for bornavirus, while 15 of 126 birds with various symptoms tested positive. A segment of approximately 1,540 bps including the N, X, P and M proteins were obtained from 23 of the positive strains and analyzed with other strains found on GenBank that had clinical information. PaBV was type 2 and 4 in South Korea, and certain amino acid sequences showed a difference between symptom presenting animals and asymptomatic animals in the X protein and P protein. When considering that some asymptomatic cases may have been latent infections at the time of examination, it is plausible these trends may grow stronger with time. Majority of PaBV was type 4 in South Korea. If these trends are confirmed, diagnosis of potentially pathogenic PaBVs in a clinical manner will be possible during the early stages of infection.

Detection of bornavirus-reactive antibodies and BoDV-1 RNA only in encephalitis patients from virus endemic areas: a comparative serological and molecular sensitivity, specificity, predictive value, and disease duration correlation study.

PURPOSE: Human Borna disease virus (BoDV-1) encephalitis is an emerging disease in Germany. This study investigates the spectrum of human BoDV-1 infection, characterizes anti-BoDV-1-antibodies and kinetics, and compares laboratory test performances. METHODS: Three hundred four encephalitis cases, 308 nation-wide neuropsychiatric conditions, 127 well-defined psychiatric cases from Borna disease-endemic areas, and 20 persons with contact to BoDV-1 encephalitis patients or animals were tested for BoDV-1 infections by serology and PCR. RESULTS: BoDV-1 infections were only found in encephalitis patients with residence in, or recent travel to, virus-endemic areas. Antibodies were detected as early as 12 days after symptom onset. Serum antibody levels correlated with disease duration. Serology was ordered after 50% of the disease duration had elapsed, reflecting low awareness. BoDV-1-antibodies were of IgG1 subclass, and the epitope on BoDV-1 antigens was determined. Specificity of the indirect immunofluorescence antibody test (IFAT) and lineblot (LB) from serum and cerebrospinal fluid (CSF), as well as PCR testing from CSF, was 100%. Sensitivity, depending on first or all samples, reached 75-86% in serum and 92-94% in CSF for the IFAT, and 33-57% in serum and 18-24% in CSF for the LB. Sensitivity for PCR in CSF was 25-67%. Positive predictive values were 100% each, while negative predictive values were 99% (IFAT), 91-97% (LB), and 90% (PCR). CONCLUSIONS: There is no hint that BoDV-1 causes other diseases than encephalitis in humans. Awareness has to be increased in virus-endemic areas. Tests are robust but lack sensitivity. Detection of IgG1 against specific peptides may facilitate diagnosis. Screening of healthy individuals is likely not beneficial.

Experimental infection of aquatic bird bornavirus 1 (ABBV-1) in Canada geese (Branta canadensis).

Aquatic bird bornavirus 1 (ABBV-1) has a high prevalence of infection in certain North American populations of Canada geese (Branta canadensis), suggesting a possible role of these birds as an ABBV-1 reservoir. The goal of this study was to evaluate the ability of Canada geese to become experimentally infected with ABBV-1, develop lesions, and transmit the virus to conspecifics. One-week-old Canada geese (n, 65) were inoculated with ABBV-1 through the intramuscular (IM) or cloacal (CL) routes, with the control group receiving carrier only. An additional 6 geese were added to each group to test horizontal transmission (sentinel birds). Geese were monitored daily, and selected birds were euthanized at 1, 8, and 15-weeks post infection (wpi) to assess virus replication in tissues and lesion development. At 15 wpi, over 70% of IM birds were infected, while the CL route yielded only 1 infected goose. Of the infected IM geese, 26% developed encephalitis and/or myelitis after 8 wpi. No clinical signs were observed, and no sentinel birds became infected in any group. Only 1 oropharyngeal swab (IM group) tested positive for ABBV-1 RNA, while the water from the enclosures was consistently negative for virus RNA. This study documents successful experimental infection of Canada geese with ABBV-1, with findings comparable to what is described in infection trials with other waterfowl species. However, minimal shedding and lack of environmental dispersal indicate that Canada geese have little potential to disseminate the virus among wild waterfowl, and that other species could be better suited to act as chronic ABBV-1 shedders in the wild.

Detection of a new avian bornavirus in barn owl (Tyto alba) by pan-viral microarray.

A barn owl (Tyto alba) died with neurological signs compatible with a viral infection. After discarding other possible infections caused by circulating viruses in the area, analysis of the central nervous system using a pan-viral microarray revealed hybridization to canary bornavirus 2 (CnBV-2). Subsequent sequence analysis confirmed the presence of a virus sharing more than 83% identity with CnBV-2. Surprisingly, the new sequence corresponds to a new virus, here named Barn owl Bornavirus 1 (BoBV-1), within the Orthobornavirus serini species. Moreover, it is the first member of this species that has been detected in a non-passerine bird, indicating that Orthobornavirus serini species comprises viruses with a wider range of hosts than previously presumed. The use of this microarray has proven to be an excellent tool for viral detection in clinical samples, with capacity to detect new viral variants. This allows the diagnosis of a great range of viruses, which can cause similar disease symptoms and which identification by PCR methods might be tedious, probably unsuccessful and, in the long run, expensive. This platform is highly useful for a fast and precise viral detection, contributing to the improvement of diagnostic methods.

Molecular detection of bornavirus in parrots imported to China in 2022.

BACKGROUND: Avian bornavirus (ABV) is a neurotropic virus, it has been established as the primary causative agent of proventricular dilatation disease (PDD). However, substantial international trade and transnational trafficking of wild birds occur, potentially enabling these birds to harbor and transmit pathogens to domestic poultry, adversely affecting their well-being. Real-time RT-PCR was employed to detect the presence of PaBV-4 in parrots imported to China in 2022. RESULTS: In 2022, a total of 47 cloacal swabs from 9 distinct species of parrots were collected at the Wildlife Rescue Monitoring Center in Guangdong, China. The purpose of this collection was to detect the presence of PaBV-4. Using real-time PCR techniques, it was determined that the positive rate of PaBV-4 was 2.12% (1 out of 47) in parrots. The PaBV-4 virus was detected in a Amazona aestiva that had been adopted for one month. Conversely, all other species tested negative for the virus. Subsequently, the whole genome of the PaBV-4 GD2207 strains was sequenced, and the homology and genetic evolution between these strains and previously published PaBV-4 strains on GenBank were analyzed using DNAStar and MEGA7.0 software. The findings revealed that the full-length genome of PaBV-4 consisted of 8915 nucleotides and encoded six proteins. Additionally, it exhibited the highest nucleotide similarity (99.9%) to the GZ2019 strain, which causes death and severe clinical symptoms in Aratinga solstitialis. Furthermore, when compared to other strains of PaBV-4, the GD2207 strain demonstrated the highest amino acid homology with GZ2019. The phylogenetic analysis demonstrated that the GD2207 strain clustered with various strains found in Japanese, American, and German parrots, indicating a close genetic relationship with PaBV-4, but it revealed a distant relationship with PaBV-5 Cockg5 from America. Notably, the GD2207 was closely associated with the GZ2019 strain from Aratinga solstitialis in China. CONCLUSION: This study presents the preliminary identification of PaBV-4 in Amazona aestiva parrots, emphasizing its importance as the predominant viral genotype linked to parrot infections resulting from trade into China. Through genetic evolution analysis, it was determined that the GD2207 strain of PaBV-4 exhibits the closest genetic relationship with GZ 2019 (Aratinga solstitialis, China), M14 (Ara macao, USA), AG5 (Psittacus erithacus, USA) and 6758 (Ara ararauna, Germany) suggesting a shared ancestry.

Reverse genetics of parrot bornavirus 4 reveals a unique splicing of the glycoprotein gene that affects viral propagation.

Viruses can utilize host splicing machinery to enable the expression of multiple genes from a limited-sized genome. Orthobornaviruses use alternative splicing to regulate the expression level of viral proteins and achieve efficient viral replication in the nucleus. Although more than 20 orthobornaviruses have been identified belonging to eight different viral species, virus-specific splicing has not been demonstrated. Here, we demonstrate that the glycoprotein (G) transcript of parrot bornavirus 4 (PaBV-4; species Orthobornavirus alphapsittaciforme), a highly virulent virus in psittacines, undergoes mRNA splicing and expresses a soluble isoform termed sGP. Interestingly, the splicing donor for sGP is not conserved in other orthobornaviruses, including those belonging to the same orthobornavirus species, suggesting that this splicing has evolved as a PaBV-4-specific event. We have also shown that exogenous expression of sGP does not affect PaBV-4 replication or de novo virion infectivity. In this study, to investigate the role of sGP in viral replication, we established a reverse genetics system for PaBV-4 by using avian cell lines and generated a recombinant virus lacking the spliced mRNA for sGP. Using the recombinant viruses, we show that the replication of the sGP-deficient virus is significantly slower than that of the wild-type virus and that the exogenous expression of sGP cannot restore its propagation efficiency. These results suggest that autologous or controlled expression of sGP by splicing may be important for PaBV-4 propagation. The reverse genetics system for avian bornaviruses developed here will be a powerful tool for understanding the replication strategies and pathogenesis of avian orthobornaviruses. IMPORTANCE Parrot bornavirus 4 (PaBV-4) is the dominant cause of proventricular dilatation disease, a severe gastrointestinal and central nervous system disease among avian bornaviruses. In this study, we discovered that PaBV-4 expresses a soluble isoform of glycoprotein (G), called sGP, through alternative splicing of the G mRNA, which is unique to this virus. To understand the role of sGP in viral replication, we generated recombinant PaBV-4 lacking the newly identified splicing donor site for sGP using a reverse genetics system and found that its propagation was significantly slower than that of the wild-type virus, suggesting that sGP plays an essential role in PaBV-4 infection. Our results provide important insights not only into the replication strategy but also into the pathogenesis of PaBV-4, which is the most prevalent bornavirus in captive psittacines worldwide.

Magnetic resonance imaging of human variegated squirrel bornavirus 1 (VSBV-1) encephalitis reveals diagnostic pattern indistinguishable from Borna disease virus 1 (BoDV-1) encephalitis but typical for bornaviruses.

Human bornavirus encephalitis is an emerging disease caused by the variegated squirrel bornavirus 1 (VSBV-1) and the Borna disease virus 1 (BoDV-1). While characteristic brain magnetic resonance imaging (MRI) changes have been described for BoDV-1 encephalitis, only scarce diagnostic data in VSBV-1 encephalitis exist. We systematically analysed brain MRI scans from all known VSBV-1 encephalitis patients. Initial and follow-up scans demonstrated characteristic T2 hyperintense lesions in the limbic system and the basal ganglia, followed by the brainstem. No involvement of the cerebellar cortex was seen. Deep white matter affection occurred in a later stage of the disease. Strict symmetry of pathologic changes was seen in 62%. T2 hyperintense areas were often associated with low T1 signal intensity and with mass effect. Sinusitis in three patients on the first MRI and an early involvement of the limbic system suggest an olfactory route of VSBV-1 entry. The viral spread could occur per continuitatem to adjacent anatomical brain regions or along specific neural tracts to more distant brain regions. The number and extent of lesions did not correlate with the length of patients' survivals. The overall pattern closely resembles that described for BoDV-1 encephalitis. The exact bornavirus species can thus not be deduced from imaging results alone, and molecular testing and serology should be performed to confirm the causative bornavirus. As VSBV-1 is likely of tropical origin, and MRI investigations are increasingly available globally, imaging techniques might be helpful to facilitate an early presumptive diagnosis of VSBV-1 encephalitis when molecular and/or serological testing is not available.

Animal Model Alternatives in Filovirus and Bornavirus Research.

The order Mononegavirales contains a variety of highly pathogenic viruses that may infect humans, including the families Filoviridae, Bornaviridae, Paramyxoviridae, and Rhabodoviridae. Animal models have historically been important to study virus pathogenicity and to develop medical countermeasures. As these have inherent shortcomings, the rise of microphysiological systems and organoids able to recapitulate hallmarks of the diseases caused by these viruses may have enormous potential to add to or partially replace animal modeling in the future. Indeed, microphysiological systems and organoids are already used in the pharmaceutical R&D pipeline because they are prefigured to overcome the translational gap between model systems and clinical studies. Moreover, they may serve to alleviate ethical concerns related to animal research. In this review, we discuss the value of animal model alternatives in human pathogenic filovirus and bornavirus research. The current animal models and their limitations are presented followed by an overview of existing alternatives, such as organoids and microphysiological systems, which might help answering open research questions.

Viral Sequences Are Repurposed for Controlling Antiviral Responses as Non-Retroviral Endogenous Viral Elements.

Eukaryotic genomes contain numerous copies of endogenous viral elements (EVEs), most of which are considered endogenous retrovirus (ERV) sequences. Over the past decade, non-retroviral endogenous viral elements (nrEVEs) derived from ancient RNA viruses have been discovered. Several functions have been proposed for these elements, including antiviral defense. This review summarizes the current understanding of nrEVEs derived from RNA viruses, particularly endogenous bornavirus-like elements (EBLs) and endogenous filovirus-like elements (EFLs). EBLs are one of the most extensively studied nrEVEs. The EBL derived from bornavirus nucleoprotein (EBLN) is thought to function as a non-coding RNA or protein that regulates host gene expression or inhibits virus propagation. Ebolavirus and marburgvirus, which are filoviruses, induce severe hemorrhagic fever in humans and nonhuman primates. Although the ecology of filoviruses remains unclear, bats are believed to be potential reservoirs. Based on the knowledge from EBLs, it is postulated that EFLs in the bat genome help to maintain the balance between filovirus infection and the bat's defense system, which may partially explain why bats act as potential reservoirs. Further research into the functions of nrEVEs could reveal novel antiviral systems and inspire novel antiviral approaches.

Experimental infection of aquatic bird bornavirus 1 in domestic chickens.

Aquatic bird bornavirus 1 (ABBV-1), classified in the Orthobornavirus genus, is a neurotropic virus that infects wild waterfowl causing persistent infection of the nervous system. Given the conspicuous presence of wild waterfowl in urban areas and farmlands, spillover of this virus into domesticated poultry species is a concern. The goal of this study was to test the ability of ABBV-1 to infect and cause disease in chickens. Two day-old, White Leghorn chickens (n, 176) were inoculated with ABBV-1 through the oral, intramuscular, or intracranial routes, and sampled at 1, 4, 8, and 12-weeks post infection (wpi) to assess virus replication and lesion development. Chickens became infected only through the intracranial and intramuscular routes, developing earliest infection in the brain by 1 wpi (intracranial group), and spinal cord by 8 wpi (intramuscular group). Except for the kidney of one bird in the intracranial group, no other tissues (including choanal and cloacal swabs) tested positive for the virus. Therefore, while the virus could reach the central nervous tissue (CNS) from the muscle in approximately 20% of birds (centripetal spread), it inefficiently reached peripheral sites after replication in the CNS (centrifugal spread). Inflammation in the CNS was observed in the intracranial and intramuscular groups starting at 8 and 12 wpi, respectively, and consisted of mononuclear perivascular cuffing. This is the first study to document the susceptibility of chickens to ABBV-1 infection, and indicates that this species can become infected with ABBV-1, although less extensively than what is observed in waterfowl. This suggests that ABBV-1 replication is partially restricted in gallinaceous birds.

Borna Disease Virus 1 Phosphoprotein Forms a Tetramer and Interacts with Host Factors Involved in DNA Double-Strand Break Repair and mRNA Processing.

Determining the structural organisation of viral replication complexes and unravelling the impact of infection on cellular homeostasis represent important challenges in virology. This may prove particularly useful when confronted with viruses that pose a significant threat to human health, that appear unique within their family, or for which knowledge is scarce. Among Mononegavirales, bornaviruses (family Bornaviridae) stand out due to their compact genomes and their nuclear localisation for replication. The recent recognition of the zoonotic potential of several orthobornaviruses has sparked a surge of interest in improving our knowledge on this viral family. In this work, we provide a complete analysis of the structural organisation of Borna disease virus 1 (BoDV-1) phosphoprotein (P), an important cofactor for polymerase activity. Using X-ray diffusion and diffraction experiments, we revealed that BoDV-1 P adopts a long coiled-coil α-helical structure split into two parts by an original ß-strand twist motif, which is highly conserved across the members of whole Orthobornavirus genus and may regulate viral replication. In parallel, we used BioID to determine the proximal interactome of P in living cells. We confirmed previously known interactors and identified novel proteins linked to several biological processes such as DNA repair or mRNA metabolism. Altogether, our study provides important structure/function cues, which may improve our understanding of BoDV-1 pathogenesis.

Canary Bornavirus (Orthobornavirus serini) Infections Are Associated with Clinical Symptoms in Common Canaries (Serinus canaria dom.).

While parrot bornaviruses are accepted as the cause of proventricular dilatation disease (PDD) in psittacine birds, the pathogenic role of bornaviruses in common canaries is still unclear. To answer the question of whether canary bornaviruses (species Orthobornavirus serini) are associated with a PDD-like disease in common canaries (Serinus canaria f. dom.), the clinical data of 201 canary bird patients tested for bornaviruses using RT-PCR assays, were analyzed for the presence of PDD-like gastrointestinal or central nervous system signs and for other viruses (mainly circovirus and polyomavirus), yeasts and trichomonads. Canary bornavirus RNA was detected in the clinical samples of 40 out of 201 canaries (19.9%) coming from 28 of 140 flocks (20%). All nucleotide sequences obtained could unequivocally be determined as canary bornavirus 1, 2, or 3 supporting the current taxonomy of the species Orthobornavirus serini. PDD-like signs were found associated with canary bornavirus detection, and to a lesser extent, with circoviruses detection, but not with the detection of polyomaviruses, yeasts or trichomonads. The data indicate that canary bornaviruses contribute to a PDD-like disease in naturally infected canaries, and suggest a promoting effect of circoviruses for the development of PDD-like signs.

Transcriptome Analysis of Duck and Chicken Brains Infected with Aquatic Bird Bornavirus-1 (ABBV-1).

Aquatic bird bornavirus 1 (ABBV-1) is a neurotropic virus that infects waterfowls, resulting in persistent infection. Experimental infection showed that both Muscovy ducks and chickens support persistent ABBV-1 infection in the central nervous system (CNS), up to 12 weeks post-infection (wpi), without the development of clinical disease. The aim of the present study was to describe the transcriptomic profiles in the brains of experimentally infected Muscovy ducks and chickens infected with ABBV-1 at 4 and 12 wpi. Transcribed RNA was sequenced by next-generation sequencing and analyzed by principal component analysis (PCA) and differential gene expression. The functional annotation of differentially expressed genes was evaluated by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. The PCA showed that the infected ducks sampled at both 4 and 12 wpi clustered separately from the controls, while only the samples from the chickens at 12 wpi, but not at 4 wpi, formed a separate cluster. In the ducks, more genes were differentially expressed at 4 wpi than 12 wpi, and the majority of the highly differentially expressed genes (DEG) were upregulated. On the other hand, the infected chickens had fewer DEGs at 4 wpi than at 12 wpi, and the majority of those with high numbers of DEGs were downregulated at 4 wpi and upregulated at 12 wpi. The functional annotation showed that the most enriched GO terms were immune-associated in both species; however, the terms associated with the innate immune response were predominantly enriched in the ducks, whereas the chickens had enrichment of both the innate and adaptive immune response. Immune-associated pathways were also enriched according to the KEGG pathway analysis in both species. Overall, the transcriptomic analysis of the duck and chicken brains showed that the main biological responses to ABBV-1 infection were immune-associated and corresponded with the levels of inflammation in the CNS.

Human infections by Bornaviruses: are they real? / Quelle est la réalité des infections humaines par les Bornavirus ?

The reality of human infections by Bornaviridae (and particularly by mammalian Orthobornaviruses BoDV-1 and BoDV-2) has long been the centre of debate and controversies. New data, however, have profoundly modified the game by providing strong and unambiguous pieces of evidence, even if many points still need to be clarified. This review aims at presenting the current state of the question, based on today's knowledge.

La question de la réalité des infections humaines par les Bornaviridae (et plus précisément par les Orthobornavirus des mammifères BoDV-1 ou BoDV-2) a longtemps constitué un point de controverse. Des données récentes ont cependant profondément remanié les cartes et permettent désormais d'avoir quelques données solides en la matière. Il n'en reste pas moins que plusieurs aspects restent mal compris. Cette revue vise à faire le point, au vu de nos connaissances à ce jour.

Avian Bornavirus Research-A Comprehensive Review.

Avian bornaviruses constitute a genetically diverse group of at least 15 viruses belonging to the genus Orthobornavirus within the family Bornaviridae. After the discovery of the first avian bornaviruses in diseased psittacines in 2008, further viruses have been detected in passerines and aquatic birds. Parrot bornaviruses (PaBVs) possess the highest veterinary relevance amongst the avian bornaviruses as the causative agents of proventricular dilatation disease (PDD). PDD is a chronic and often fatal disease that may engulf a broad range of clinical presentations, typically including neurologic signs as well as impaired gastrointestinal motility, leading to proventricular dilatation. It occurs worldwide in captive psittacine populations and threatens private bird collections, zoological gardens and rehabilitation projects of endangered species. In contrast, only little is known about the pathogenic roles of passerine and waterbird bornaviruses. This comprehensive review summarizes the current knowledge on avian bornavirus infections, including their taxonomy, pathogenesis of associated diseases, epidemiology, diagnostic strategies and recent developments on prophylactic and therapeutic countermeasures.

A New Multiplex Real-Time RT-PCR for Simultaneous Detection and Differentiation of Avian Bornaviruses.

Avian bornaviruses were first described in 2008 as the causative agents of proventricular dilatation disease (PDD) in parrots and their relatives (Psittaciformes). To date, 15 genetically highly diverse avian bornaviruses covering at least five viral species have been discovered in different bird orders. Currently, the primary diagnostic tool is the detection of viral RNA by conventional or real-time RT-PCR (rRT-PCR). One of the drawbacks of this is the usage of either specific assays, allowing the detection of one particular virus, or of assays with a broad detection spectrum, which, however, do not allow for the simultaneous specification of the detected virus. To facilitate the simultaneous detection and specification of avian bornaviruses, a multiplex real-time RT-PCR assay was developed. Whole-genome sequences of various bornaviruses were aligned. Primers were designed to recognize conserved regions within the overlapping X/P gene and probes were selected to detect virus species-specific regions within the target region. The optimization of the assay resulted in the sensitive and specific detection of bornaviruses of Psittaciformes, Passeriformes, and aquatic birds. Finally, the new rRT-PCR was successfully employed to detect avian bornaviruses in field samples from various avian species. This assay will serve as powerful tool in epidemiological studies and will improve avian bornavirus detection.

ICTV Virus Taxonomy Profile: Bornaviridae.

Members of the family Bornaviridae produce enveloped virions containing a linear negative-sense non-segmented RNA genome of about 9 kb. Bornaviruses are found in mammals, birds, reptiles and fish. The most-studied viruses with public health and veterinary impact are Borna disease virus 1 and variegated squirrel bornavirus 1, both of which cause fatal encephalitis in humans. Several orthobornaviruses cause neurological and intestinal disorders in birds, mostly parrots. Endogenous bornavirus-like sequences occur in the genomes of various animals. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Bornaviridae, which is available at ictv.global/report/bornaviridae.

Two novel bornaviruses identified in colubrid and viperid snakes.

We present the complete genome sequences of Caribbean watersnake bornavirus (CWBV) and Mexican black-tailed rattlesnake bornavirus (MRBV), which we identified in archived raw transcriptomic read data of a Caribbean watersnake (Tretanorhinus variabilis) and a Mexican black-tailed rattlesnake (Crotalus molossus nigrescens), respectively. The genomes of CWBV and MRBV have a length of about 8,900 nucleotides and comprise the complete coding regions and the untranslated regions. The overall genomic makeup and predicted gene content is typical for members of the genus Orthobornavirus within the family Bornaviridae. Alternative splicing was detected for the L and M genes. Based on a phylogenetic analysis of all viral proteins, we consider both viruses to be members of a single novel species within the genus Orthobornavirus. Both viruses form a distinct outgroup to all currently known orthobornaviruses. Based on the novel virus genomes, we furthermore identified closely related endogenous bornavirus-like nucleoprotein sequences in transcriptomic data of veiled chameleons (Chamaeleo calyptratus) and a common lancehead (Bothrops atrox).

100-My history of bornavirus infections hidden in vertebrate genomes.

Although viruses have threatened our ancestors for millions of years, prehistoric epidemics of viruses are largely unknown. Endogenous bornavirus-like elements (EBLs) are ancient bornavirus sequences derived from the viral messenger RNAs that were reverse transcribed and inserted into animal genomes, most likely by retrotransposons. These elements can be used as molecular fossil records to trace past bornaviral infections. In this study, we systematically identified EBLs in vertebrate genomes and revealed the history of bornavirus infections over nearly 100 My. We confirmed that ancient bornaviral infections have occurred in diverse vertebrate lineages, especially in primate ancestors. Phylogenetic analyses indicated that primate ancestors were infected with various bornaviral lineages during evolution. EBLs in primate genomes formed clades according to their integration ages, suggesting that bornavirus lineages infected with primate ancestors had changed chronologically. However, some bornaviral lineages may have coexisted with primate ancestors and underwent repeated endogenizations for tens of millions of years. Moreover, a bornaviral lineage that coexisted with primate ancestors also endogenized in the genomes of some ancestral bats. The habitats of these bat ancestors have been reported to overlap with the migration route of primate ancestors. These results suggest that long-term virus-host coexistence expanded the geographic distributions of the bornaviral lineage along with primate migration and may have spread their infections to these bat ancestors. Our findings provide insight into the history of bornavirus infections over geological timescales that cannot be deduced from research using extant viruses alone, thus broadening our perspective on virus-host coevolution.