Wildlife Disease Association conference abstracts
Authors: David S. Blehert, Anne E. Ballmann, Brenda M. Berlowski-Zier, Jeffrey M. Lorch, and Carol U. Meteyer
Key words: Bat, cutaneous infection, fungal infection, Geomyces, white-nose syndrome
A previously undescribed fungus, Geomyces sp., is associated with white-nose syndrome, an emergent disease linked with the estimated mortality of over 400,000 cave-hibernating bats of the northeastern and mid-Atlantic United States.
White-nose syndrome (WNS) is a condition associated with an unprecedented bat mortality event in the northeastern and mid-Atlantic US. Since the winter of 2006-2007 bat population declines ranging from 80-97% have been documented at surveyed hibernacula. This presentation summarizes evidence indicating an association between WNS and a newly described fungus.
Following necropsy, bat muzzle and wing skin samples were submitted for histologic and culture analyses. Histology sections were stained with PAS and fungal cultures (Sabouraud dextrose agar) were incubated at 7?C. Fungal isolates were identified by microscopy and by PCR amplification/sequencing of the rRNA gene internal transcribed spacer region using primers ITS4 and ITS5. Other techniques under development include fungus-specific PCR for screening environmental and tissue samples, and in situ hybridization for identification of fungal hyphae in histologic sections.
Histopathological analyses demonstrated that 90% of necropsied bats (n=117) submitted from WNS-suspect sites exhibited a cutaneous fungal infection. Direct microscopy, culture, and PCR/sequencing analyses confirmed that the skin of WNS-affected bats was colonized by genetically identical isolates of a psychrophilic (cold-loving) fungus with a unique conidial morphology.
There is a growing body of evidence supporting a strong association between WNS and cutaneous infection by a newly discovered psychrophilic fungus. The growth temperature requirements of the fungus are consistent with the core temperatures of hibernating cave-bat species throughout temperate regions of the world. Given the hundreds of thousands of hibernating bats found throughout the WNS-affected region, this condition represents an unprecedented threat to bats of the northeastern United States and potentially beyond.
Coral reefs can undergo relatively rapid changes in the dominant biota, a phenomenon referred to as phase shift. Various reasons have been proposed to explain this phenomenon including increased human disturbance, pollution, or changes in coral reef biota that serve a major ecological function such as depletion of grazers. However, pinpointing the actual factors potentially responsible can be problematic. Here we show a phase shift from coral to the corallimorpharian Rhodactis howesii associated with a long line vessel that wrecked in 1991 on an isolated atoll (Palmyra) in the central Pacific Ocean. We documented high densities of R. howesii near the ship that progressively decreased with distance from the ship whereas R. howesii were rare to absent in other parts of the atoll. We also confirmed high densities of R. howesii around several buoys recently installed on the atoll in 2001. This is the first time that a phase shift on a coral reef has been unambiguously associated with man-made structures. This association was made, in part, because of the remoteness of Palmyra and its recent history of minimal human habitation or impact. Phase shifts can have long-term negative ramification for coral reefs, and eradication of organisms responsible for phase shifts in marine ecosystems can be difficult, particularly if such organisms cover a large area. The extensive R. howesii invasion and subsequent loss of coral reef habitat at Palmyra also highlights the importance of rapid removal of shipwrecks on corals reefs to mitigate the potential of reef overgrowth by invasives.
Laysan ducks are endemic to the Hawaiian archipelago and are one of the world’s most endangered waterfowl. For 150 years, Laysan ducks were restricted to an estimated 4 km2 of land on Laysan Island in the Northwestern Hawaiian Islands. In 2004 and 2005 42 Laysan ducks were translocated to Midway Atoll, and the population increased to approximately 200 by 2007. In August 2008 mortality due to botulism type C was identified, and 181 adult, fledgling, and duckling carcasses were collected between August-October. Diseased birds were found on two islands within Midway Atoll at multiple wetlands; however, one wetland contributed the most carcasses. The epizootic was discovered approximately 14-21 days after the mortality started and lasted for 30 days. Based on necropsy findings 3 of 84 recovered Laysan duck carcasses were also infected with a proventricular worm suspected to be Echinuria uncinata which is known to be highly pathogenic to Laysan ducks on Laysan Island. This worm was either moved to Midway during the 2004-2005 translocations of ducks from Laysan despite antihelminthic treatment of all founding birds or it arrived with migratory waterfowl. The details of this epizootic highlight the disease risk to birds restricted to small island populations and the challenges associated with managing newly translocated endangered species. Frequent population monitoring for early disease detection and comprehensive wetland monitoring and management will be needed to mitigate the potential effects of avian botulism and parasitism on endangered Laysan ducks.
Authors: Jeffrey M. Lorch, Carol U. Meteyer, Brenda M. Berlowski-Zier, and David S. Blehert
Key words: Bat, fungal infection, Geomyces, PCR, white-nose syndrome
A PCR method was developed to rapidly, selectively, and accurately detect the white-nose syndrome-associated Geomyces sp. fungus on bat wing tissue.
Skin infection by Geomyces sp. is a clinical indication of bat white-nose syndrome (WNS). Formerly, time-consuming fungal culture and/or histologic examination of bat skin were necessary to confirm infection. We developed a PCR test to specifically detect the WNS-associated fungus directly from bat wing skin in less than one day.
Bat carcasses were obtained from hibernacula within and outside of the WNS-affected region. A 4mm X 4mm piece of wing tissue was excised from each animal for genomic DNA extraction/PCR analysis, and additional wing skin from each animal was used for companion histology and culture analyses. PCR was conducted using primers specific for the Geomyces sp. rRNA gene internal transcribed spacer region. Generation of a 586 nucleotide band indicated that Geomyces sp. DNA was present. PCR, histology, and culture results from each bat were compared to determine the efficacy of the method.
Samples testing positive for Geomyces sp. infection by culture or histology (n=22) also tested positive by PCR 95% of the time. Due to competition by faster-growing microbes, culture can yield false-negative results. PCR had the capability to identify Geomyces sp. DNA in the absence of positive culture results.
WNS was formerly diagnosed using culture and/or histological techniques. However, those methods are time-consuming, require specialized skills, and have the potential to yield false negative results. We developed an accurate PCR-based test for detection of Geomyces sp. with a significantly faster turn-around time than the previous methods. This technique will be useful in delivering results promptly to field personnel and wildlife managers, for expanded WNS-surveillance efforts, and for environmental surveys.
Authors: Carol Meteyer, Doug Docherty, Hon Ip, Nathan Ramsay, Emi Saito, Lindsay Oaks
Key words: American crow, enteritis, mortality, Orthoreovirus
A novel Orthoreovirus is associated with fatal necrotizing enteritis in American crows and has been diagnosed in 14 Unites States, from New York to the state of Washington. Crow mortality associated with this syndrome has recurred annually in some states since it was first diagnosed at the National Wildlife Health Center during the winter of 2001-2002.
Fatal enteritis in American crows associated with reovirus was first detected during surveillance for West Nile virus in 2002. A focused diagnostic investigation was subsequently conducted on American crows dying from hemorrhagic enteritis to determine factors that contribute to this syndrome in crows.
American crows that were submitted to the National Wildlife Health Center for West Nile virus surveillance from Washington during September 2003 were necropsied using a uniform protocol. Intestine, spleen and kidney were cultured on Vero cells. Intestine was cultured for aerobic and anaerobic bacteria, for salmonella using XLT agar, and submitted for parasitology evaluation. Multiple tissues including intestine, spleen, and liver were examined for histopathology.
Necrotizing enteritis and splenic necrosis was present in 10/20 crows and an orthoreovirus was isolated from all ten. Of these 10 crows, liver necrosis was present in 4, Escherichia coli was isolated from 7, Clostridium perfringens positive for alpha toxin from 3, Clostridium sordelli from one, and intestinal coccidia from two.
The significance of this novel reovirus in the fatal necrotizing enteritis in American crows has yet to be established. Pathogenicity studies are ongoing to determine the relative importance of the multiple potential pathogens identified in these birds. The population significance of this syndrome compounded by the continued mortality in corvids due to West Nile virus is also unknown.
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U.S. Department of the Interior | U.S. Geological Survey
