Identification and characterization of Highlands J virus from a Mississippi sandhill crane using unbiased next-generation sequencing.

Advances in massively parallel DNA sequencing platforms, commonly termed next-generation sequencing (NGS) technologies, have greatly reduced time, labor, and cost associated with DNA sequencing. Thus, NGS has become a routine tool for new viral pathogen discovery and will likely become the standard for routine laboratory diagnostics of infectious diseases in the near future. This study demonstrated the application of NGS for the rapid identification and characterization of a virus isolated from the brain of an endangered Mississippi sandhill crane. This bird was part of a population restoration effort and was found in an emaciated state several days after Hurricane Isaac passed over the refuge in Mississippi in 2012. Post-mortem examination had identified trichostrongyliasis as the possible cause of death, but because a virus with morphology consistent with a togavirus was isolated from the brain of the bird, an arboviral etiology was strongly suspected. Because individual molecular assays for several known arboviruses were negative, unbiased NGS by Illumina MiSeq was used to definitively identify and characterize the causative viral agent. Whole genome sequencing and phylogenetic analysis revealed the viral isolate to be the Highlands J virus, a known avian pathogen. This study demonstrates the use of unbiased NGS for the rapid detection and characterization of an unidentified viral pathogen and the application of this technology to wildlife disease diagnostics and conservation medicine.

West nile virus transmission in winter: the 2013 great salt lake bald eagle and eared grebes mortality event.

West Nile Virus (WNV) infection has been reported in over 300 species of birds and mammals. Raptors such as eagles, hawks and falcons are remarkably susceptible, but reports of WNV infection in Bald Eagles (Haliaeetus leucocephalus) are rare and reports of WNV infection in grebes (Podicipediformes) even rarer. We report an unusually large wild bird mortality event involving between 15,000-20,000 Eared Grebes (Podiceps nigricollis) and over 40 Bald Eagles around the Great Salt Lake, Utah, in November-December 2013. Mortality in grebes was first reported in early November during a period when the area was unseasonably warm and the grebes were beginning to gather and stage prior to migration. Ten out of ten Eared Grebes collected during this period were WNV RT-PCR and/or isolation positive. This is the first report of WNV infection in Eared Grebes and the associated mortality event is matched in scale only by the combined outbreaks in American White Pelican (Pelecanus erythrorhynchos) colonies in the north central states in 2002-2003. We cannot be sure that all of the grebes were infected by mosquito transmission; some may have become infected through contact with WNV shed orally or cloacally from other infected grebes. Beginning in early December, Bald Eagles in the Great Salt Lake area were observed to display neurological signs such as body tremors, limb paralysis and lethargy. At least 43 Bald Eagles had died by the end of the month. Nine of nine Bald Eagles examined were infected with WNV. To the best of our knowledge, this is the largest single raptor mortality event since WNV became endemic in the USA. Because the majority of the eagles affected were found after onset of below-freezing temperatures, we suggest at least some of the Bald Eagles were infected with WNV via consumption of infected Eared Grebes or horizontal transmission at roost sites.

Avian botulism and avian chlamydiosis in wild water birds, Benton Lake National Wildlife Refuge, Montana, USA.

In 1999, the U.S. Geological Survey (USGS) National Wildlife Health Center, Madison, Wisconsin, conducted a diagnostic investigation into a water bird mortality event involving intoxication with avian botulism type C and infection with avian chlamydiosis at the Benton Lake National Wildlife Refuge in Montana, USA. Of 24 carcasses necropsied, 11 had lesions consistent with avian chlamydiosis, including two that tested positive for infectious Chlamydophila psittaci, and 12 were positive for avian botulism type C. One bird tested positive for both avian botulism type C and C. psittaci. Of 61 apparently healthy water birds sampled and released, 13 had serologic evidence of C. psittaci infection and 7 were, at the time of capture, shedding infectious C. psittaci via the cloacal or oropharyngeal route. Since more routinely diagnosed disease conditions may mask avian chlamydiosis, these findings support the need for a comprehensive diagnostic investigation when determining the cause of a wildlife mortality event.

Pathology and virus detection in tissues of nestling house sparrows naturally infected with Buggy Creek virus (Togaviridae).

Alphaviruses (Togaviridae) infect wild birds, but clinical illness and death attributable to virus in naturally infected birds is rarely reported, particularly for small passerine species or nestlings. Buggy Creek virus is a unique alphavirus in the Western equine encephalomyelitis virus (WEEV) complex that is vectored by the cimicid swallow bug (Oeciacus vicarius), an ectoparasite of the colonially nesting Cliff Swallow (Petrochelidon pyrrhonota) and the introduced House Sparrow (Passer domesticus). While sampling birds for Buggy Creek virus (BCRV) during the summers of 2007 and 2008, we discovered large numbers of clinically ill or dead House Sparrow nestlings. Ill nestlings exhibited ataxia, torticollis, paresis, and lethargy. Histologic examination revealed that encephalitis was the most common finding, followed by myositis, myocarditis, and hepatic changes, but pathology was highly variable. We isolated BCRV from brain tissue in most of the ill or dead nestlings, and from blood, liver, kidney, spleen, lung, feather pulp, and skin in some birds. To our knowledge, this is the first report of clinical illness, gross pathology, and histopathology for a WEEV-complex alphavirus in a field-collected passerine species.

Corvidae feather pulp and West Nile virus detection.

We evaluated cloacal swab, vascular pulp of flight feather, and kidney and spleen pool samples from carcasses of members of the family Corvidae as sources of West Nile virus (WNV). The cloacal swab, kidney and spleen pool, and feather pulp were the source of WNV in 38%, 43%, and 77%, respectively, of the carcasses.