Immunophenotype of the inflammatory response in the central and enteric nervous systems of cockatiels (Nymphicus hollandicus) experimentally infected with parrot bornavirus 2

Proventricular dilatation disease is a lethal disease of psittacine birds. In this study, we characterized the local cellular immune response in the brain, proventriculus, and small intestine of 27 cockatiels ( Nymphicus hollandicus) experimentally infected with parrot bornavirus 2 (PaBV-2). Perivascular cuffs in the brain were composed of CD3+ T-lymphocytes and Iba1+ macrophages/microglia in most cockatiels (n = 26). In the ganglia of the proventriculus, CD3+ T-lymphocytes (n = 17) and Iba1+ macrophages (n = 13) prevailed. The ganglia of the small intestine had a more homogeneous distribution of these leukocytes, including PAX5+ B-lymphocytes (n = 9), CD3+ T-lymphocytes (n = 8), and Iba1+ macrophages (n = 8). Our results indicate that perivascular cuffs in the brain and the inflammatory infiltrate in the proventriculus of PaBV-2-infected cockatiels is predominately composed of T-lymphocytes, while the inflammatory infiltrates in the ganglia of the small intestine are characterized by a mixed infiltrate composed of T-lymphocytes, B-lymphocytes, and macrophages.

Seeing beyond a Dilated Proventriculus: Diagnostic Tools for Proventricular Dilatation Disease in Psittacine Birds

Proventricular dilatation disease (PDD) is a life-threatening neurological disease caused by parrot bornaviruses (PaBVs) that affects several species worldwide. PDD can be clinically manifested as either a central nervous system condition or a gastrointestinal condition if the nerves and ganglia of the gastrointestinal tract are compromised. We intend to provide a concise review for veterinary clinicians and diagnosticians with focus on the main tools available for PDD diagnosis, including gross and histopathology, immunohistochemistry, molecular techniques and serology. We suggest that a combination of different strategies can increase the success of diagnostic outcomes, as tools such as reverse transcription polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) can be implemented for identification of bornaviral infections in live patients, and gross pathology, histopathology, immunohistochemistry and RT-PCR can provide reliable results for postmortem diagnosis of PDD.

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.

The Pathogenesis of Proventricular Dilatation Disease Caused by Parrot Bornaviruses: A Possible Role For Neuropeptide Y (Npy)

Background Psittacine Bornaviruses cause a unique disease syndrome in parrots and related birds. Known as proventricular dilatation disease (PDD), it is characterized by massive dilatation of their proventriculus as a result of excessive food accumulation within that organ. This leads to gastric obstruction and eventually, to death by starvation. Results In a preliminary study on the transcriptome of psittacine bornavirus-infected human astroglia it was noted that the gene encoding neuropeptide Y was significantly upregulated. In a subsequent study of cockatiels experimentally infected with the same strain of psittacine bornavirus, their brains were examined by RNA-seq to determine which genes were being actively transcribed. It was confirmed that among the genes whose expression was significantly increased relative to control, uninfected birds was that for neuropeptide Y. Conclusions Neuropeptide Y is known to cause overeating in birds. We hypothesize therefore that the clinical manifestations of proventricular dilatation disease are a result of the excessive production of neuropeptide Y by bornavirus-infected brain cells acting in association with damage to the neurons of the proventricular myenteric plexus.

Recombinant Modified Vaccinia Virus Ankara (MVA) Vaccines Efficiently Protect Cockatiels Against Parrot Bornavirus Infection and Proventricular Dilatation Disease

Parrot bornaviruses (PaBVs) are the causative agents of proventricular dilatation disease (PDD), a chronic and often fatal neurologic disorder in Psittaciformes. The disease is widely distributed in private parrot collections and threatens breeding populations of endangered species. Thus, immunoprophylaxis strategies are urgently needed. In previous studies we demonstrated a prime-boost vaccination regime using modified vaccinia virus Ankara (MVA) and Newcastle disease virus (NDV) constructs expressing the nucleoprotein and phosphoprotein of PaBV-4 (MVA/PaBV-4 and NDV/PaBV-4, respectively) to protect cockatiels (Nymphicus hollandicus) against experimental challenge infection. Here we investigated the protective effect provided by repeated immunization with either MVA/PaBV-4, NDV/PaBV-4 or Orf virus constructs (ORFV/PaBV-4) individually. While MVA/PaBV-4-vaccinated cockatiels were completely protected against subsequent PaBV-2 challenge infection and PDD-associated lesions, the course of the challenge infection in NDV/PaBV-4- or ORFV/PaBV-4-vaccinated birds did not differ from the unvaccinated control group. We further investigated the effect of vaccination on persistently PaBV-4-infected cockatiels. Remarkably, subsequent immunization with MVA/PaBV-4 and NDV/PaBV-4 neither induced obvious immunopathogenesis exacerbating the disease nor reduced viral loads in the infected birds. In summary, we demonstrated that vaccination with MVA/PaBV-4 alone is sufficient to efficiently prevent PaBV-2 challenge infection in cockatiels, providing a suitable vaccine candidate against avian bornavirus infection and bornavirus-induced PDD.

Correlation of avian bornavirus-specific antibodies and viral ribonucleic acid shedding with neurological signs and feather-damaging behaviour in psittacine birds

Parrot bornaviruses (PaBV) are the causative agents of proventricular dilatation disease in psittacine birds, but have also been linked to other clinical signs, including behavioural disorders and neurological signs. The aim of this study was to correlate PaBV infection in birds showing feather-damaging behaviour or neurological signs for which no other cause of disease could be identified. Psittacine birds presented to a private practice were divided into three groups: birds with neurological signs (n=28), birds showing feather-damaging behaviour (n=42) and birds presented for routine examinations (n=56). Swabs of crop and cloaca were collected and investigated for the presence of PaBV-RNA using real time RT-PCR. Additionally, serum samples were taken and examined for the presence of anti-PaBV antibodies by immunofluorescence test. PaBV infection was detected in one of the test systems in 40.5 per cent of all birds (n=126) investigated. In the clinically healthy birds (n=56), 19.6 per cent of the birds were positive in at least one of the PaBV tests, compared with 52.38 per cent of the feather-damaging (n=42) and 64.28 per cent of the neurologically diseased birds (n=28). Interestingly, the anti-PaBV antibody titres in birds with neurological signs were highest up to 1:20 480. High antibody titres (up to 1:5120) were also found in the feather-damaging group, whereas the birds of the control group, if PaBV positive, had only very low titres. Similarly, the highest viral load was found in the group of the neurologically diseased birds, followed by feather-damaging birds, whereas PaBV-positive birds in the control group demonstrated only low viral RNA shedding. A clear correlation between severity of clinical signs, amount of viral shedding and high levels of antibody titres was observed for most of the neurologically diseased birds and also for few birds with feather-damaging behaviour. For the first time, these results clearly indicate a correlation between PaBV infection and neurological signs in birds without gastrointestinal signs presented to the veterinarian in practice. It also may demonstrate a possible correlation with feather-damaging behaviour and anti-PaBV antibody presence. The antibody titre seems to represent a diagnostic tool to correlate clinical signs to PaBV as a cause.

Proventricular dilatation disease (PDD) outbreak in blue-and-gold macaws (Ara ararauna) in the State of Santa Catarina, southern Brazil

ABSTRACT: Proventricular dilatation disease (PDD) is a lethal and important disease of captive psittacine birds, and affects a wide range of species, including endangered ones, and lacks an effective treatment. This report describes PDD in three blue-and-gold macaws (Ara ararauna) in southern Brazil. All three macaws originated from the same aviary and presented similar clinical signs including anorexia, apathy, emaciation and prostration. At necropsy, one of the macaws presented an enlarged proventriculus. Histologically, lymphoplasmacytic infiltrates was observed in the ganglia and nerves of the esophagus, crop, proventriculus, ventriculus, heart, adrenal glands, and adrenal medulla of all three cases. Two macaws had meningoencephalomyelitis and one had myocarditis. Immunohistochemistry identified PaBV antigen in the brain, proventricular, ventricular ganglia, and epicardial ganglia, and cardiomyocytes of all three macaws.

Could introducing confiscated parrots to zoological collections jeopardise conservation breeding programmes?

SummaryConfiscated parrots are frequently introduced to captive populations in zoological institutions, regularly with insufficient health screening. This short communication describes a case where 25 confiscated parrots, from four different locations, were brought to the same zoological institution within two years, where they were kept under quarantine conditions. A year after the last birds arrived, several birds died due to either proventricular dilatation disease or herpesvirus infection. As all individuals belonged to rare species, the surviving birds were transferred to the Justus-Liebig-University in Giessen, Germany, for thorough diagnostics including parrot bornavirus, psittacine herpesvirus 1, adenovirus, polyomavirus, circovirus, Chlamydia psittaci, and mycobacteria. Birds that tested negative for all pathogens were transferred to captive breeding programmes, whereas pathogen carriers were paired up in collections of a similar pathogen status. This case report highlights the dangers of latent infections with different pathogens and the importance of managed screening programmes if such populations are to be considered for conservation.