Frequent Arousal from Hibernation Linked to Severity of Infection and Mortality in Bats with White-Nose Syndrome

White-nose syndrome (WNS), an emerging infectious disease that has killed over 5.5 million hibernating bats, is named for the causative agent, a white fungus (Geomyces destructans (Gd)) that invades the skin of torpid bats. During hibernation, arousals to warm (euthermic) body temperatures are normal but deplete fat stores. Temperature-sensitive dataloggers were attached to the backs of 504 free-ranging little brown bats (Myotis lucifugus) in hibernacula located throughout the northeastern USA. Dataloggers were retrieved at the end of the hibernation season and complete profiles of skin temperature data were available from 83 bats, which were categorized as: (1) unaffected, (2) WNS-affected but alive at time of datalogger removal, or (3) WNS-affected but found dead at time of datalogger removal. Histological confirmation of WNS severity (as indexed by degree of fungal infection) as well as confirmation of presence/absence of DNA from Gd by PCR was determined for 26 animals. We demonstrated that WNS-affected bats aroused to euthermic body temperatures more frequently than unaffected bats, likely contributing to subsequent mortality. Within the subset of WNS-affected bats that were found dead at the time of datalogger removal, the number of arousal bouts since datalogger attachment significantly predicted date of death. Additionally, the severity of cutaneous Gd infection correlated with the number of arousal episodes from torpor during hibernation. Thus, increased frequency of arousal from torpor likely contributes to WNS-associated mortality, but the question of how Gd infection induces increased arousals remains unanswered.

Altered Behavior in Bats Affected by White-Nose Syndrome

WNS-affected bats did so over similar time frames as WNSunaffected bats. The behaviors of bats with WNS did not change as drastically as expected. Thereseems to be little to no effect on their ability to fly/forage until much later stages of the disease when they are likely near death. WNS-affected bats are grooming more which could be altering the way they use energy reserves during hibernation possibly leading tostarvation and eventually death. The decreased likelihood of arousals in response to external cues may be the result of spending more energy during previous and increasingly frequent arousals. While it is clear that WNS does result in changes in behavior whether these changes are directly in response to fungal skin infection or to some other component of the syndrome such as decreased energy availability or loss of homeostasis is unknown. bat behavior, white-nose syndrome, behavior, video surveillance, arousal patterns White-Nose Syndrome (WNS) is a disease of hibernating bats caused by the fungal pathogen Geomyces destructans. The fungus, which was first noted in 2006, invades bats wings and other exposed membranes, eventually resulting in death. Researchers have yet to understand many aspects of this disease, including basic etiology and epidemiology. There is also a lack of information on how fungal infection may change the behavior of healthy bats during hibernation or how changes in behavior may influence disease progression. Based upon the physiological changes that are known to occur in affected bats, and upon anecdotal observations of aberrant behavior in these bats, I hypothesized that WNS would significantly change the behavior of the little brown myotis (Myotis lucifugus). My research examined the behavior of hibernating bats during arousals from torpor. I compared WNS-affected and unaffected bats, in the field and incaptivity, using motion-sensitive infrared cameras. Flight maneuverability and echolocation were also tested between WNS-affected and unaffected bats during arousalsfrom hibernation to detect changes in the bats' ability to perform basic locomotion or potentially catch insect prey. Lastly, hibernating bats were artificially disturbed and theirarousal patterns were monitored to examine changes in the response to external stimuli between WNS-affected and unaffected bats.Bats with WNS groomed for longer periods of time after arousing from torpor, both in the field and in captivity. They also engaged in longer periods of any sort of activity during these arousals. There were no changes in acoustical signaling during flight tests and changes in flight maneuverability were only found in bats were seen staging" near the entrance of the mine which is itself a unique behavior exhibited by affected bats. At this point these bats were likely near death and could barely fly at all. In response toexternal stimuli bats with WNS were less likely to arouse than unaffected bats. However when they did arouse WNS-affected bats did so over similar time frames as WNSunaffected bats. The behaviors of bats with WNS did not change as drastically as expected. Thereseems to be little to no effect on their ability to fly/forage until much later stages of the disease when they are likely near death. WNS-affected bats are grooming more which could be altering the way they use energy reserves during hibernation possibly leading tostarvation and eventually death. The decreased likelihood of arousals in response to external cues may be the result of spending more energy during previous and increasingly frequent arousals. While it is clear that WNS does result in changes in behavior whetherthese changes are directly in response to fungal skin infection or to some other component of the syndrome such as decreased energy availability or loss of homeostasis is unknown."

Nonlethal Screening of Bat-Wing Skin with the Use of Ultraviolet Fluorescence to Detect Lesions Indicative of White-Nose Syndrome

Definitive diagnosis of the bat disease white-nose syndrome (WNS) requires histologic analysis to identify the cutaneous erosions caused by the fungal pathogen Pseudogymnoascus [formerly Geomyces] destructans (Pd). Gross visual inspection does not distinguish bats with or without WNS, and no nonlethal, on-site, preliminary screening methods are available for WNS in bats. We demonstrate that long-wave ultraviolet (UV) light (wavelength 366-385 nm) elicits a distinct orange yellow fluorescence in bat-wing membranes (skin) that corresponds directly with the fungal cupping erosions in histologic sections of skin that are the current gold standard for diagnosis of WNS. Between March 2009 and April 2012, wing membranes from 168 North American bat carcasses submitted to the US Geological Survey National Wildlife Health Center were examined with the use of both UV light and histology. Comparison of these techniques showed that 98.8% of the bats with foci of orange yellow wing fluorescence (n=80) were WNS-positive based on histologic diagnosis; bat wings that did not fluoresce under UV light (n=88) were all histologically negative for WNS lesions. Punch biopsy samples as small as 3 mm taken from areas of wing with UV fluorescence were effective for identifying lesions diagnostic for WNS by histopathology. In a nonlethal biopsy-based study of 62 bats sampled (4-mm diameter) in hibernacula of the Czech Republic during 2012, 95.5% of fluorescent (n=22) and 100% of nonfluorescent (n=40) wing samples were confirmed by histopathology to be WNS positive and negative, respectively. This evidence supports use of long-wave UV light as a nonlethal and field-applicable method to screen bats for lesions indicative of WNS. Further, UV fluorescence can be used to guide targeted, nonlethal biopsy sampling for follow-up molecular testing, fungal culture analysis, and histologic confirmation of WNS.

Experimental Infection of Bats with Geomyces destructans Causes White-nose Syndrome

White-nose syndrome (WNS) has caused recent catastrophic declines among multiple species of bats in eastern North America1, 2. The disease’s name derives from a visually apparent white growth of the newly discovered fungus Geomyces destructans on the skin (including the muzzle) of hibernating bats1, 3. Colonization of skin by this fungus is associated with characteristic cutaneous lesions that are the only consistent pathological finding related to WNS4. However, the role of G. destructans in WNS remains controversial because evidence to implicate the fungus as the primary cause of this disease is lacking. The debate is fuelled, in part, by the assumption that fungal infections in mammals are most commonly associated with immune system dysfunction5, 6, 7. Additionally, the recent discovery that G. destructans commonly colonizes the skin of bats of Europe, where no unusual bat mortality events have been reported8, 9, 10, has generated further speculation that the fungus is an opportunistic pathogen and that other unidentified factors are the primary cause of WNS11, 12. Here we demonstrate that exposure of healthy little brown bats (Myotis lucifugus) to pure cultures of G. destructans causes WNS. Live G. destructans was subsequently cultured from diseased bats, successfully fulfilling established criteria for the determination ofG. destructans as a primary pathogen13. We also confirmed that WNS can be transmitted from infected bats to healthy bats through direct contact. Our results provide the first direct evidence that G. destructans is the causal agent of WNS and that the recent emergence of WNS in North America may represent translocation of the fungus to a region with a naive population of animals8. Demonstration of causality is an instrumental step in elucidating the pathogenesis14 and epidemiology15 of WNS and in guiding management actions to preserve bat populations against the novel threat posed by this devastating infectious disease.