Betanodavirus meningoencephalitis in an Atlantic blue marlin.

Viral nervous necrosis (viral encephalopathy and retinopathy) is caused by piscine nodavirus (Nodaviridae, Betanodavirus). Since 1986, this highly infectious virus has caused mass mortalities of up to 100% in farmed saltwater and freshwater fish around the world (with the exception of South America and Antarctica), affecting >60 species across 10 orders. The Atlantic blue marlin (Makaira nigricans Lacépède, 1802) is a top-level predator found throughout the tropical waters of the Atlantic and Indo-Pacific oceans. Despite their popularity as a sportfish, relatively little is known about the Atlantic blue marlin and other billfish. We describe here chronic betanodavirus infection in a juvenile Atlantic blue marlin, which is, to our knowledge, the first report of disease in M. nigricans.

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.

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.

Vasculitis Associated with Parrot Bornavirus 4 Infection in a Rose-Crowned Parakeet (Pyrrhurarhodocephala).

A 3-month-old, female rose-crowned parakeet (Pyrrhura rhodocephala) was found dead after a 24-h course of lethargy and passing blood-tinged faeces. Fine white streaks were seen in the pectoral muscles on necropsy. Microscopic examination revealed typical lesions of avian ganglioneuritis and vascular necrosis in the pectoral muscles, myocardium, kidneys, air sacs, adrenal glands, pancreas and thyroid gland. These lesions were characterized by mural fibrinoid necrosis of small and medium-calibre arteries and arterioles, associated with lymphoplasmacytic inflammation, necrosis, atrophy and fibrosis of the surrounding tissues. Parrot bornavirus (PaBV) nucleoprotein was demonstrated by immunohistochemistry in smooth muscle and endothelial cells of many vessels. An avian bornavirus was isolated from kidney tissue and its identity confirmed as PaBV-4 by sequencing and phylogenetic analysis. We postulate that the vascular lesions could have been immune-mediated and that PaBV-4 may have played a role in its pathogenesis.

Bornavirus Encephalitis Shows a Characteristic Magnetic Resonance Phenotype in Humans.

OBJECTIVE: The number of diagnosed fatal encephalitis cases in humans caused by the classical Borna disease virus (BoDV-1) has been increasing, ever since it was proved that BoDV-1 can cause human infections. However, awareness of this entity is low, and a specific imaging pattern has not yet been identified. We therefore provide the first comprehensive description of the morphology of human BoDV-1 encephalitis, with histopathological verification of imaging abnormalities. METHODS: In an institutional review board-approved multicenter study, we carried out a retrospective analysis of 55 magnetic resonance imaging (MRI) examinations of 19 patients with confirmed BoDV-1 encephalitis. Fifty brain regions were analyzed systematically (T1w, T2w, T2*w, T1w + Gd, and DWI), in order to discern a specific pattern of inflammation. Histopathological analysis of 25 locations in one patient served as correlation for MRI abnormalities. RESULTS: Baseline imaging, acquired at a mean of 11 ± 10 days after symptom onset, in addition to follow-up scans of 16 patients, revealed characteristic T2 hyperintensities with a predilection for the head of the caudate nucleus, insula, and cortical spread to the limbic system, whereas the occipital lobes and cerebellar hemispheres were unaffected. This gradient was confirmed by histology. Nine patients (47.4%) developed T1 hyperintensities of the basal ganglia, corresponding to accumulated lipid phagocytes on histology and typical for late-stage necrosis. INTERPRETATION: BoDV-1 encephalitis shows a distinct pattern of inflammation in both the early and late stages of the disease. Its appearance can mimic sporadic Creutzfeldt-Jakob disease on MRI and should be considered a differential diagnosis in the case of atypical clinical presentation. ANN NEUROL 2020;88:723-735.

Parrot bornavirus in naturally infected Brazilian captive parrots: Challenges in viral spread control.

Psittaciform orthobornaviruses are currently considered to be a major threat to the psittacine bird population worldwide. Parrot bornavirus (PaBV) was identified recently in Brazil and, since then, few studies have been conducted to understand the epidemiology of PaBV in captive psittacine birds. In the present study, natural infections by PaBV in South American parrots were investigated in two breeding facilities: commercial (A) and conservationist (B). Thirty-eight psittacine of 21 different species were presented for postmortem examination. Tissue samples were collected and investigated for the presence of PaBV-RNA using RT-PCR. In addition, clinical information about these birds was used when available. PaBV infection was detected in 73.7% of all birds investigated, indicating a wide dissemination of this virus in both facilities. From birds investigated in aviary A, 66.7% showed clinical signs, 100% had typical lesions of proventricular dilatation disease (PDD), 100% had mild to severe proventricular dilatation and 88.9% were PaBV-positive. In birds from aviary B, 27.6% showed clinical signs, 65.5% had typical lesions of PDD, 62% had mild to severe proventricular dilatation and 69% were PaBV-positive. Neurological disease was observed more frequently than gastrointestinal disease. Sequencing analysis of the matrix gene fragment revealed the occurrence of genotype 4 (PaBV-4) in both places. About 15.8% of birds in this study are threatened species. We discussed the difficulties and challenges for controlling viral spread in these aviaries and implications for South American psittacine conservation. These results emphasize the urgent need to develop a national regulatory and health standard for breeding psittacine birds in the country.

Update on Avian Bornavirus and Proventricular Dilatation Disease: Diagnostics, Pathology, Prevalence, and Control.

Avian bornavirus (ABV) is a neurotropic virus that can cause gastrointestinal and/or neurologic signs of disease in birds. The disease process is called proventricular dilatation disease (PDD). The characteristic lesions observed in birds include encephalitis and gross dilatation of the proventriculus. ABV is widely distributed in captive and wild bird populations. Most birds infected do not show clinical signs of disease. This article is an update of the Veterinary Clinics of North America article from 2013: Avian Bornavirus and Proventricular Dilatation Disease: Diagnostics, Pathology, Prevalence, and Control.

Cardiac Lesions of Natural and Experimental Infection by Parrot Bornaviruses.

Lymphoplasmacytic inflammation associated with bornavirus N protein occurs in the epicardial ganglia, myocardium and endocardium of birds diagnosed with proventricular dilatation disease (PDD). These pathological findings suggest that sudden death in psittacine birds might stem from cardiac compromise due to parrot bornavirus (PaBV) infection. Therefore, we investigated cardiac lesions in cases of PDD, searching databases from 1988 to 2019, and reviewed three experimental studies of PaBV infection. Fifty cases of PDD in birds infected naturally with PaBV and 27 cases of PDD in birds infected experimentally with PaBV (all having descriptions of inflammatory cardiac lesions) were reviewed. For each case, five regions of the heart were evaluated by light microscopy and immunohistochemistry (IHC). These regions were the epicardial ganglia/nerves, the endocardium, the myocardium, the Purkinje fibres and the great vessels. Sudden death was documented in 17/50 naturally infected cases, while 23/50 had digestive signs, and only 12/50 had neurological signs. Grossly, only five naturally-infected and five experimentally-infected cases had cardiomegaly or hydropericardium. Epicardial ganglioneuritis was the most consistent microscopical finding in natural (46/50) and experimental cases (26/27), followed by myocarditis (34/50) for naturally-infected and endocarditis for experimentally-infected birds (6/27). PaBV-2 antigen was detected most frequently by IHC in the epicardial ganglia (54/77) compared with the other tissues. This retrospective study demonstrates the presence of PaBV protein and inflammation in the heart of birds infected with PaBV and suggests a link between PaBV and cardiac disease and sudden death in psittacine birds.

Analysis of exotic squirrel trade and detection of human infections with variegated squirrel bornavirus 1, Germany, 2005 to 2018.

Following the discovery in 2015 of the variegated squirrel bornavirus 1 (VSBV-1) in fatal encephalitis cases among exotic squirrel breeders and a zoo animal caretaker in Germany, a case definition was developed. It was employed during trace-back animal trade investigations and sero-epidemiological studies among breeders and zoo animal caretakers of holdings with VSBV-1 infected squirrels. During the investigation, two possible human cases who had died of encephalitis were identified retrospectively among the squirrel breeders. Moreover, one probable human case was detected among the breeders who had a positive memory T-cell response to VSBV-1 antigen and antibodies against VSBV-1. The low rate of seropositivity found among living persons in risk groups that handle exotic squirrels privately or at zoos may reflect rareness of exposure to VSBV-1 during animal contact, a high lethality of infection or a combination of these factors. As a precaution against human exposure, testing of exotic squirrels for VSBV-1 infection and/or avoiding direct contact with exotic squirrels in zoos and private holdings is strongly advised.

Distribution of Viral Antigen and Inflammatory Lesions in the Central Nervous System of Cockatiels ( Nymphicus hollandicus) Experimentally Infected with Parrot Bornavirus 2.

Neurotropism is a striking characteristic of bornaviruses, including parrot bornavirus 2 (PaBV-2). Our study evaluated the distribution of inflammatory foci and viral nucleoprotein (N) antigen in the brain and spinal cord of 27 cockatiels ( Nymphicus hollandicus) following experimental infection with PaBV-2 by injection into the pectoral muscle. Tissue samples were taken at 12 timepoints between 5 and 114 days post-inoculation (dpi). Each experimental group had approximately 3 cockatiels per group and usually 1 negative control. Immunolabeling was first observed within the ventral horns of the thoracic spinal cord at 20 dpi and in the brain (thalamic nuclei and hindbrain) at 25 dpi. Both inflammation and viral antigen were restricted to the central core of the brain until 40 dpi. The virus then spread quickly at 60 dpi to both gray and white matter of all analyzed sections of the central nervous system (CNS). Encephalitis was most severe in the thalamus and hindbrain, while myelitis was most prominent in the gray matter and equally distributed in the cervical, thoracic, and lumbosacral spinal cord. Our results demonstrate a caudal to rostral spread of virus in the CNS following experimental inoculation of PABV-2 into the pectoral muscle, with the presence of viral antigen and inflammatory lesions first in the spinal cord and progressing to the brain.

Two Neuropsychiatric Cases Seropositive for Bornavirus Improved by Ribavirin.

While we previously detected anti-bornavirus antibodies via radioligand assay in psychiatric patients, we did not examine the viral pathogenicity in these individuals. Herein, we present 2 psychiatric patients who were seropositive for bornavirus and whose treatment-resistant symptoms improved after oral administration of ribavirin, a broad-spectrum antiviral agent. Cerebrospinal fluid analysis indicated that ribavirin affected the central nervous system of these patients. Ribavirin ameliorated intermittent involuntary head shaking, which is reminiscent of a symptom observed in bornavirus-infected animals. Using radioligand assays to examine the serial sera of these patients, we found a relationship between the titers of anti-bornavirus antibodies and the change in the patients' symptoms. Our findings suggest there is a relationship between bornavirus infection and human symptoms and that ribavirin may be useful in suppressing chronic bornavirus infection in some neuropsychiatric patients. However, the possibility remains that some other known or unknown virus other than bornavirus that is sensitive to ribavirin may have caused the symptoms. Additional evidence that directly indicates the causative relationship between bornavirus infection and human symptoms is needed before establishing the pathogenesis and treatment for human bornavirus infection.

Avian Ganglioneuritis in Clinical Practice.

Avian ganglioneuritis (AG) comprises one of the most intricate pathologies in avian medicine and is researched worldwide. Avian bornavirus (ABV) has been shown to be a causative agent of proventricular dilatation disease in birds. The avian Bornaviridae represent a genetically diverse group of viruses that are widely distributed in captive and wild populations around the world. ABV and other infective agents are implicated as a cause of the autoimmune pathology that leads to AG, similar to human Guillain Barrè syndrome. Management of affected birds is beneficial and currently centered at reducing neurologic inflammation, managing secondary complications, and providing nutritional support.

From nerves to brain to gastrointestinal tract: A time-based study of parrot bornavirus 2 (PaBV-2) pathogenesis in cockatiels (Nymphicus hollandicus).

Parrot bornaviruses (PaBVs) are the causative agents of proventricular dilatation disease, however key aspects of its pathogenesis, such as route of infection, viral spread and distribution, and target cells remain unclear. Our study aimed to track the viral spread and lesion development at 5, 10, 20, 25, 35, 40, 60, 80, 95 and 114 dpi using histopathology, immunohistochemistry, and RT-PCR. After intramuscular inoculation of parrot bornavirus 2 (PaBV-2) in the pectoral muscle of cockatiels, this virus was first detected in macrophages and lymphocytes in the inoculation site and adjacent nerves, then reached the brachial plexus, centripetally spread to the thoracic segment of the spinal cord, and subsequently invaded the other spinal segments and brain. After reaching the central nervous system (CNS), PaBV-2 centrifugally spread out the CNS to the ganglia in the gastrointestinal (GI) system, adrenal gland, heart, and kidneys. At late points of infection, PaBV-2 was not only detected in nerves and ganglia but widespread in the smooth muscle and/or scattered epithelial cells of tissues such as crop, intestines, proventriculus, kidneys, skin, and vessels. Despite the hallmark lesion of PaBVs infection being the dilation of the proventriculus, our results demonstrate PaBV-2 first targets the CNS, before migrating to peripheral tissues such as the GI system.

Investigation of Different Infection Routes of Parrot Bornavirus in Cockatiels.

The aim of this study was to determine the natural infection route of parrot bornavirus (PaBV), the causative agent of proventricular dilatation disease (PDD) in psittacines. For this purpose, nine cockatiels ( Nymphicus hollandicus ) were inoculated orally, and nine cockatiels were inoculated intranasally, with a PaBV-4 isolate. To compare the results of the trials, the same isolate and the same experimental design were used as in a previous study where infection was successful by intravenous as well as intracerebral inoculation. After inoculation, the birds were observed for a period of 6 mo and tested for PaBV RNA shedding, virus replication, presence of inflammatory lesions, and PaBV-4 antigen in tissues, as well as specific antibody production. In contrast to the previous study involving intravenous and intracerebral infections, clinical signs typical for PDD were not observed in this study. Additionally, anti-PaBV antibodies and infectious virus were not detected in any investigated bird during the study. Parrot bornavirus RNA was detected in only four birds early after infection (1-34 days postinfection). Furthermore, histopathologic examination did not reveal lesions typical for PDD, and PaBV antigen was not detected in any organ investigated by immunohistochemistry. In summary, oral or nasal inoculation did not lead to a valid infection with PaBV in these cockatiels. Therefore it seems to be questionable that the formerly proposed fecal-oral transmission is the natural route of infection in immunocompetent adult or subadult cockatiels.

The pathogenesis of proventricular dilatation disease.

Bornaviruses cause neurologic diseases in several species of birds, especially parrots, waterfowl and finches. The characteristic lesions observed in these birds include encephalitis and gross dilatation of the anterior stomach - the proventriculus. The disease is thus known as proventricular dilatation disease (PDD). PDD is characterized by extreme proventricular dilatation, blockage of the passage of digesta and consequent death by starvation. There are few clinical resemblances between this and the bornaviral encephalitides observed in mammals. Nevertheless, there are common virus-induced pathogenic pathways shared across this disease spectrum that are explored in this review. Additionally, a review of the literature relating to gastroparesis in humans and the control of gastric mobility in mammals and birds points to several plausible mechanisms by which bornaviral infection may result in extreme proventricular dilatation.

A Variegated Squirrel Bornavirus Associated with Fatal Human Encephalitis.

Between 2011 and 2013, three breeders of variegated squirrels (Sciurus variegatoides) had encephalitis with similar clinical signs and died 2 to 4 months after onset of the clinical symptoms. With the use of a metagenomic approach that incorporated next-generation sequencing and real-time reverse-transcriptase quantitative polymerase chain reaction (RT-qPCR), the presence of a previously unknown bornavirus was detected in a contact squirrel and in brain samples from the three patients. Phylogenetic analyses showed that this virus, tentatively named variegated squirrel 1 bornavirus (VSBV-1), forms a lineage separate from that of the known bornavirus species. (Funded by the Federal Ministry of Food and Agriculture [Germany] and others.).

No contact transmission of avian bornavirus in experimentally infected cockatiels (Nymphicus hollandicus) and domestic canaries (Serinus canaria forma domestica).

Avian bornaviruses (ABV) are the causative agents of proventricular dilatation disease (PDD), a widely distributed disease of parrots. Distinct ABV lineages were also found in various non-psittacine avian species, such as canaries, but the pathogenic role of ABV in these species is less clear. Despite the wide distribution of ABV in captive parrots and canaries, its mode of transmission is poorly understood: both horizontal transmission via the urofaecal-oral route and vertical transmission are discussed to play a role. In this study we investigated pathology and horizontal transmission of ABV in domestic canaries (Serinus canaria forma domestica) and cockatiels (Nymphicus hollandicus), two natural host species commonly used for experimental ABV infections. ABV inoculation resulted in persistent infection of all inoculated animals from both species. ABV-infected cockatiels exhibited PDD-like symptoms, such as neurologic signs or shedding of undigested seeds. In contrast, infected domestic canaries did not develop clinical disease. Interestingly, we did not detect viral RNA in cloacal swabs and organ samples or ABV-specific antibodies in serum samples of contact-exposed sentinel birds from either species at any time during a four months observation period. Our results strongly indicate that horizontal transmission of ABV by direct contact is inefficient in immunocompetent fully fledged domestic canaries and cockatiels.

Fatal proventricular dilatation disease in captive native psittacines in Brazil.

An outbreak of proventricular dilatation disease (PDD), a fatal inflammatory disease of psittacines (Aves: Psittaciformes), is described in native Brazilian psittacines. Twenty captive psittacines that died of suspected PDD were necropsied and 10 were submitted to histopathology, reverse transcriptase PCR (RT-PCR), and immunohistochemistry (IHC) for avian bornavirus (ABV). Examined species were one pileated parrot (Pionopsitta pileata), three vinaceous-breasted parrots (Amazona vinacea), two blue-winged macaws (Primolius maracana), one scarlet macaw (Ara macao), one chestnut-fronted macaw (Ara severa), one scaly-headed parrot (Pionus maximiliani), and one red-browed Amazon parrot (Amazona rhodocorytha). Gross examination and histopathology revealed typical PDD lesions in all birds. The presence of ABV was confirmed in four psittacines including one red-browed Amazon parrot, one blue-winged macaw, one scarlet macaw, and one chestnut-fronted macaw. In the red-browed Amazon parrot and in one blue-winged macaw, IHC demonstrated ABV antigens in the nucleus and cytoplasm of cells in various organs. This is the first description of PDD by ABV in Brazilian psittacines and indicates the necessity for adopting a strategic control plan for reducing its impact in native birds.

Avian bornaviruses are widely distributed in canary birds (Serinus canaria f. domestica).

Avian bornavirus (ABV) was identified in 2008 as the causative agent of proventricular dilatation disease (PDD) in psittacine birds. In addition, ABV variants were detected in wild waterfowl and in a canary bird. PDD-like diseases were also reported in various other avian species, but it remains unknown whether ABV is involved. In this study we detected ABV in 12 of 30 tested canary bird flocks (40%), indicating a wide distribution of ABV in captive canary birds in Germany. Sequence analysis identified several distinct ABV genotypes which differ markedly from the genotypes present in psittacine birds. Some canaries naturally infected with ABV exhibited gastrointestinal and neurological symptoms which resembled PDD in psittacines, while others did not show signs of disease. Canaries experimentally inoculated with ABV developed infections of the brain and various other organs. The experimentally infected canaries transmitted the virus to sentinel birds kept in the same aviary, but did not show any clinical signs during a five month observation period. Embryonated eggs originating from ABV-infected hens contained ABV-specific RNA, but virus could not be re-isolated from embryonic tissue. These results indicate that ABV is widely distributed in canary birds and due to its association to clinical signs should be considered as a potential pathogen of this species.

Pathology and diagnosis of avian bornavirus infection in wild Canada geese (Branta canadensis), trumpeter swans (Cygnus buccinator) and mute swans (Cygnus olor) in Canada: a retrospective study.

Nine hundred and fifty-five pathology cases collected in Ontario between 1992 and 2011 from wild free-ranging Canada geese, trumpeter swans and mute swans were retrospectively evaluated for the pathology associated with avian bornavirus (ABV) infection. Cases were selected based on the presence of upper gastrointestinal impaction, central nervous system histopathology or clinical history suggestive of ABV infection. The proportion of birds meeting at least one of these criteria was significantly higher at the Toronto Zoo (30/132) than elsewhere in Ontario (21/823). Central, peripheral and autonomic nervous tissues were examined for the presence of lymphocytes and plasma cells on histopathology. The presence of virus was assessed by immunohistochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR) on frozen brains and on formalin-fixed paraffin-embedded tissues. Among selected cases, 86.3% (44/51) were considered positive on histopathology, 56.8% (29/51) were positive by immunohistochemistry, and RT-PCR was positive on 88.2% (15/17) of the frozen brains and 78.4% (40/51) of the formalin-fixed paraffin-embedded samples. Histopathological lesions included gliosis and lymphoplasmacytic perivascular cuffing in brain (97.7%), spinal cord (50%), peripheral nerves (55.5%) and myenteric ganglia or nerves (62.8%), resembling lesions described in parrots affected with proventricular dilatation disease. Partial amino acid sequences of the nucleocapsid gene from seven geese were 100% identical amongst themselves and 98.1 to 100% identical to the waterfowl sequences recently described in the USA. Although ABV has been identified in apparently healthy geese, our study confirmed that ABV can also be associated with significant disease in wild waterfowl species.