Comparisons with free-flying bats
At the end of the experiment in late hibernation, we sampled free-flying bats found roosting near the cages within each persisting site. At Persisting 1 (Cold + Dry), we found that free-flying bats had significantly lower tissue invasion compared to the bats within cages (Fig. 5; Supplemental Table 7). Conversely, free-flying bats at Persisting 2 (Cold + Wet) displayed a similar degree of infection severity to caged bats within the same site. Free-flying bats had higher late hibernation body mass than caged bats in both persisting sites (Supplemental Fig. 6; Supplemental Table 8), and this difference was more pronounced in Persisting 1 (Cold + Dry).
DISCUSSION
Our data suggest that environmental conditions interact with host traits to jointly drive persistence of host populations. We found evidence for elevated on-host growth of P. destructans with increasing roosting temperature in bats that originated in Persisting 1 (Cold + Dry), but no relationship in bats that originated in Persisting 2 (Cold + Wet). Infection severity, host body condition, and survival also appeared to be influenced by site humidity, with higher disease severity and lower survival associated with over-winter exposure to the driest conditions in Persisting 1 (Cold + Dry). However, within the dry conditions, bats sampled from outside of cages in late hibernation displayed significantly lower infection severity compared to their caged counterparts, whereas no such difference was detected in Persisting 2 (Cold + Wet). This suggests that bats within Persisting 1 (Cold + Dry) may be utilizing a variety of microclimates and not remaining in these dry environments for the entire winter period, as the caged bats experienced. Importantly, the survival we observed during this experiment was significantly higher than survival during the initial epidemic within the same sites in all cases, as well as during a similar experiment in 2009. Furthermore, we observed declines in pathogen loads on 10 individuals, nine of which were in persisting sites. These data collectively suggest that persisting little brown bat colonies in the northeast United States have evolved traits beneficial to surviving WNS88,89, but that these host traits interact with environmental conditions such that protection against severe disease and mortality depends to a strong degree on temperature and humidity.
We detected a positive relationship between average roosting temperature during hibernation and on-host pathogen growth rate. This is corroborated by data from the initial WNS epidemic, where the most severely impacted colonies were those hibernating in relatively warm hibernacula3. Infection severity and over-winter weight loss showed a similar trend when humidity was high, with lower values occurring at Persisting 2 (Cold + Wet) compared to Extirpated (Warm + Wet). However, despite the coldest ambient conditions, infection severity and host weight loss were high under the dry conditions within Persisting 1 (Cold + Dry) and were comparable to the extirpated site. Under unfavorable ambient conditions, fungal pathogens may forgo reproduction and instead commit resources to within-host growth and the formation of spores that can survive stressful conditions. For example,Metarhizium anisopliae is a fungal pathogen of tick eggs that invades the egg tissue and undergoes growth90,91. Under humid, favorable conditions, the fungus will emerge from the egg to undergo asexual reproduction. However, under dry, unfavorable conditions, the fungus will instead remain within the egg host, continue to undergo growth, and produce environmentally resistant spores91. We suggest that, similarly, exposure to dry conditions of Persisting 1 (Cold + Dry) over winter were unfavorable to the survival of P. destructans in superficial infections, and that the pathogen augmented tissue invasion to satisfy moisture requirements, resulting in a high degree of infection severity. Additionally, evaporative water loss from hibernating bats is highest in dry conditions92,93 and is exacerbated by infection with P. destructans 78, resulting in dehydration and increased arousal frequency to re-hydrate78–84. Increased frequency of arousal from torpor drives the premature depletion of fat reserves during hibernation, resulting in weight loss and starvation58,94. Therefore, the increased tissue invasion and evaporative water loss in Persisting 1 (Cold + Dry) may have operated synergistically to result in severe disease and ultimately the lowest observed survival.
Colonies of little brown bats in dry hibernacula may be persisting because of the availability of different microclimates. Microclimatic conditions are not uniform throughout an entire hibernation site, but vary with factors such as depth, air flow, and the height of the ceiling95. Recent evidence suggests that bats may arouse and move to different roosting locations periodically during hibernation96, possibly in response to shifting costs associated with hibernation97, which could expose them to a variety of microclimates98. For example, some data suggest that bats may transition from roosting in relatively warm sections of hibernacula in early hibernation to the relatively cold sections by late winter96. In our study, bats were unable to select varying microclimates over the course of hibernation. However, we observed hundreds of little brown bats roosting in the area surrounding the cages during late winter in Persisting 1 (Cold + Dry), whereas less than a dozen individuals appeared to use that specific location in early hibernation, suggesting that bats do not roost in the same location for the entirety of hibernation in this site. Given that disease severity is highly dependent on environmental conditions within hibernacula, this movement behavior may have been pre-adaptive to surviving WNS if bats utilize microclimates that mitigate disease severity for at least part of hibernation. For example, movement within hibernacula may reduce the growth of P. destructans in late hibernation if bats move to the relatively cold conditions that slow pathogen growth, potentially affording them enough time to emerge from hibernation in spring and clear infection. Within Persisting 1 (Cold + Dry), free-flying bats sampled at the end of hibernation had significantly lower infection severity than caged bats, which is the expected pattern if movement within hibernacula is indeed beneficial to mitigating disease severity. Furthermore, free-flying bats in both persisting sites had higher late hibernation body masses, and this was more pronounced in Persisting 1 (Cold + Dry). Behavioral responses that moderate the severity of disease have also been proposed for snake populations impacted by snake fungal disease99, caused by the fungal pathogen Ophidiomyces ophiodiicola 100. Snakes infected with O. ophiodiicola exhibit changes to their behavior that include increased surface activity and more time spent in exposed environments compared to their disease-free conspecifics99,100, potentially a sign of a behavioral fever response to infection101. However, we make the important distinction here that because movement within hibernacula was observed in bats prior to the WNS epidemic, this behavior may have been pre-adaptive to surviving the disease rather than a direct response to the disease itself. Future research should investigate how the availability and utilization of varying environmental conditions can influence the dynamics of WNS, and how this may scale up to a population-level response.
During the initial epidemic, the cold conditions within hibernacula utilized by colonies may have prevented total colony collapse, allowing standing genetic variation for favorable host traits to propagate3,24,68,70,102. Previous research has also found genetic evidence from persisting little brown bat colonies indicative of a selective sweep following the invasion of P. destructans 89,103. However, our data suggest that populations that appear to have evolved adaptive host traits are only afforded protection within a narrow environmental space. These processes have the potential to result in local adaptation, in which the evolutionary response of populations to WNS and the resulting dominant phenotype is specific to the local environmental conditions of hibernacula104. We detected an effect of origin site on pathogen growth rate within Extirpated (Warm + Wet), potentially a signature of local adaptation to differing conditions in source hibernacula. However, for local adaptation to occur, the strength of selection must be high enough to combat the homogenizing effects of gene flow105–108, and current genetic evidence suggests a panmictic genetic landscape for bat populations109–113, but see114,115.
Environmental conditions within hibernacula are sensitive to conditions present aboveground95, and global climate change has the potential to alter the host-pathogen interaction in this system116,117. Disease severity is strongly linked to temperature and humidity conditions, as is the protection afforded by unique host traits, as this study suggests. Therefore, even slight changes to the environmental conditions within hibernacula may alter host survival within a given site, potentially resulting in more severe disease and higher mortality if conditions deviate from the environmental space within which bats coexist with the pathogen. Future research should explore the potential for climate change to impact the disease dynamics of WNS, as well as the potential for bat populations to respond to shifting conditions.
As P. destructans continues to spread throughout North America, bat population declines and regional extirpations will continue to occur. However, this study strongly suggests that prior to the invasion of P. destructans , host traits conducive to surviving WNS circulated in little brown bat populations, which now offer some colonies imperfect protection from the disease. These host traits do not operate independently to promote population persistence with WNS, but rather interact strongly with environmental conditions, specifically temperature and humidity, to ultimately drive host-pathogen coexistence. Therefore, we should not expect to see all little brown bat populations across North America stabilize or rebound from declines, but rather the persistence of colonies with the correct combination of host traits and environmental conditions.
Host population response to the invasion of a virulent pathogen will not be predictable by a single aspect of the host, environment, or pathogen. Rather, host-pathogen interactions and coexistence will be strongly mediated by environmental conditions, the result of which may be as variable as the environment itself118,119. Underlying variation in host and pathogen populations will set the stage for subsequent coevolutionary processes and the likelihood of coexistence102,120, but this interaction and the resulting host population response may be influenced by environmental conditions that vary over space and time118,121, as illustrated by this study. Therefore, to achieve predictability in how emerging infectious diseases will impact host populations, it is essential to disentangle host-environment-pathogen interactions across a geographic and temporal mosaic of host-pathogen coevolution.
MATERIALS AND METHODS