Proventricular dilatation disease (PDD) is considered by many to be the greatest threat to aviculture of psittacine birds (parrots). This disease has been documented in multiple continents in over 50 different species of psittacines as well as captive and free-ranging species in at least 5 other orders of birds [1
]. Most, if not all major psittacine collections throughout the world have experienced cases of PDD. It has been particularly devastating in countries like Canada and northern areas of the United States where parrots are housed primarily indoors. However, it is also problematic in warmer regions where birds are typically bred in outdoor aviaries. Moreover, captive breeding efforts for at least one psittacine which is thought to be extinct in the wild, the Spix's macaw (Cyanopsitta spixii
), have been severely impacted by PDD.
PDD is an inflammatory disease of birds, first described in the 1970s as Macaw Wasting Disease during an outbreak among macaws (reviewed in [3
]). PDD primarily affects the autonomic nerves of the upper and middle digestive tract, including the esophagus, crop, proventriculus, ventriculus, and duodenum. Microscopically, the disease is recognized by the presence of lymphoplasmacytic infiltrates within myenteric ganglia and nerves. Similar infiltrates may also be present in the brain, spinal cord, peripheral nerves, conductive tissue of the heart, smooth and cardiac muscle, and adrenal glands. Non-suppurative leiomyositis and/or myocarditis may accompany the neural lesions [6
]. Clinically, PDD cases present with GI tract dysfunction (dysphagia, regurgitation, and passage of undigested food in feces), neurologic symptoms (e.g. ataxia, abnormal gait, proprioceptive defects), or both [3
]. Although the clinical course of the disease can vary, it is generally fatal in untreated animals [3
The cause of PDD is unknown, but several studies have raised the possibility that PDD may be caused by a viral pathogen. Evidence for an infectious etiology stems from the initial outbreaks of Macaw Wasting Disease, and other subsequent outbreaks of PDD [2
]. Reports of pleomorphic virus-like particles of variable size (30–250 nm) observed in tissues of PDD affected birds [8
] led to the proposal that paramyxovirus (PMV) may cause the disease; however, serological data has shown that PDD affected birds lack detectable antibodies against PMV of serotypes 1–4, 6, and 7, as well as against avian herpes viruses, polyomavirus, and avian encephalitis virus [3
]. Similarly, a proposed role for equine encephalitis virus in PDD has been ruled out [11
]. Enveloped virus-like particles of approximately 80 nm in diameter derived from the feces of affected birds have been shown to produce cytopathic effect in monolayers of macaw embryonic cells [12
], but to date no reports confirming these results or identifying this possible agent have been published. Likewise, adeno-like viruses, enteroviruses, coronaviruses and reoviruses have also been sporadically documented in tissues or excretions of affected birds [3
] yet in each case, follow-up evidence for reproducible isolation specifically from PDD cases or identification of these candidate agents has not been reported. Thus, the etiology of PDD has remained an open question.
To address this question, we have turned to a comprehensive, high throughput strategy to test for the presence of known or novel viruses in PDD affected birds. We employed the Virus chip, a DNA microarray containing representation of all viral taxonomy to interrogate 2 PDD case/control series independently collected on two different continents for the presence of viral pathogens. We report here the detection of a novel bornavirus signature in 62.5% of the PDD cases and none of the controls. These bornavirus-positive samples were confirmed by virus-specific PCR testing, and the complete genome sequence has been recovered by ultra-high throughput sequencing combined with conventional PCR-based cloning.
Bornaviruses are a family of negative strand RNA viruses whose prototype member is Borna Disease Virus (BDV), an agent of encephalitis whose natural reservoir is primarily horses and sheep [15
]. Although experimental transmission of BDV to many species (including chicks [16
]) has been described, there is little information on natural avian infection, and existing BDV isolates are remarkable for their relative sequence homogeneity. The agent reported here, which we designate avian bornavirus (ABV) is highly diverged from all previously identified members of the Bornaviridae
family and represents the first full-length bornavirus genome cloned directly from avian tissue. Subsequent PCR screening for similar ABVs confirmed a detection rate of approximately 70% among PDD cases and none among the controls. Sequence analysis of a single complete genome and all of the additional partial sequences that we have recovered directly from the PDD case specimens suggests that the viruses detected in cases of PDD form a new, genetically diverse clade of the Bornaviridae