Effective conservation of subterranean-roosting bats.

Bats frequently inhabit caves and other subterranean habitats and play a critical role in subterranean food webs. With escalating threats to subterranean ecosystems, identifying the most effective measures to protect subterranean-roosting bats is critical. We conducted a meta-analysis to evaluate the effectiveness of conservation and management interventions for subterranean-roosting bats. We used network analyses to determine to what extent interventions for bats overlap those used for other subterranean taxa. We conducted our analyses with data extracted from 345 papers recommending a total of 910 conservation interventions. Gating of roost entrances was applied to preserve bat populations in 21 studies, but its effectiveness was unclear. Habitat restoration and disturbance reduction positively affected bat populations and bat behavior, respectively, in ≤4 studies. Decontamination was assessed in 2 studies and positively affected bat populations, particularly in studies focused on reducing fungal spores associated with white-nose syndrome in North America. Monitoring of bat populations as an effective conservation strategy was unclear and infrequently tested. Only 4% of bat studies simultaneously considered other subterranean organisms. However, effective interventions for bat conservation had similarities with all other organisms. If other subterranean organisms are considered when applying interventions to conserve bats, they might also benefit.

Conservación eficiente de murciélagos subterráneos Resumen Es común que los murciélagos habiten en cuevas y otros hábitats subterráneos y contribuyan a las redes alimenticias bajo tierra. Ya que estos ecosistemas cada vez se enfrentan a más amenazas, es importante identificar las medidas más efectivas para proteger a los murciélagos subterráneos. Realizamos un metaanálisis para evaluar la eficiencia de la conservación y las intervenciones de manejo para estos mamíferos. Usamos un análisis de redes para determinar el grado al que las intervenciones en pro de los murciélagos se traslapan con aquellas usadas para otros taxones subterráneos. Realizamos nuestros análisis con datos extraídos de 345 artículos que recomendaban 910 intervenciones de conservación. Se aplicó la colocación de compuertas en la entrada de los dormideros para conservar la población de murciélagos en 21 estudios, pero no quedó clara su efectividad. La restauración del hábitat y la reducción de las perturbaciones afectaron, respectivamente, a las poblaciones y al comportamiento de los murciélagos en ≤ 4 cuatro estudios. Se evaluó a la desinfección en dos estudios y ésta tuvo un efecto positivo sobre las poblaciones, particularmente en los estudios enfocados en la reducción de esporas micóticas asociadas con el síndrome de nariz blanca en América del Norte. La eficiencia del monitoreo de las poblaciones de murciélagos como una estrategia de conservación no fue clara y casi nunca se evaluó. Sólo el 4% de los estudios sobre murciélagos consideró simultáneamente a otros organismos subterráneos. Sin embargo, las intervenciones eficientes para la conservación de murciélagos tuvieron similitudes con las de todos los demás organismos. Otros organismos pueden beneficiarse si se les considera cuando se aplican las intervenciones para conservar a los murciélagos.

Whole-genome Analysis Reveals Contrasting Relationships Among Nuclear and Mitochondrial Genomes Between Three Sympatric Bat Species.

Understanding mechanisms involved in speciation can be challenging, especially when hybridization or introgression blurs species boundaries. In bats, resolving relationships of some closely related groups has proved difficult due subtle interspecific variation both in morphometrics and molecular data sets. The endemic South American Histiotus bats, currently considered a subgenus of Eptesicus, harbor unresolved phylogenetic relationships and of those is a trio consisting of two closely related species: Eptesicus (Histiotus) macrotus and Eptesicus (Histiotus) montanus, and their relationship with a third, Eptesicus (Histiotus) magellanicus. The three sympatric species bear marked resemblance to each other, but can be differentiated morphologically. Furthermore, previous studies have been unable to differentiate the species from each other at a molecular level. In order to disentangle the phylogenetic relationships of these species, we examined the differentiation patterns and evolutionary history of the three Eptesicus (H.) species at the whole-genome level. The nuclear DNA statistics between the species suggest strong gene flow and recent hybridization between E. (H.) montanus and E. (H.) macrotus, whereas E. (H.) magellanicus shows a higher degree of isolation. In contrast, mitochondrial DNA shows a closer relationship between E. (H.) magellanicus and E. (H.) montanus. Opposing patterns in mtDNA and nuclear markers are often due to differences in dispersal, and here it could be both as a result of isolation in refugia during the last glacial maximum and female philopatry and male-biased dispersal. In conclusion, this study shows the importance of both the nuclear and mitochondrial DNA in resolving phylogenetic relationships and species histories.

Seasonal roost selection and activity of a remnant population of northern myotis in Pennsylvania.

The decline in northern myotis (Myotis septentrionalis) populations due to the disease white-nose syndrome (WNS) has led to the species receiving federal protection in the United States and Canada, requiring conservation of critical habitats. However, considerably more is known about summer habitat preferences of northern myotis compared to late summer through winter. Our goal was to describe the seasonal presence and habitat use of a remnant colony of northern myotis in central Pennsylvania. We radio-tagged 31 northern myotis and established 6 acoustic monitoring stations to document activity from 2017-2021. We found that roost trees used during the maternity season by reproductive females were occupied by bats during both summer (21 June-14 August) and autumn (15 August-31 October), indicating similar habitat use patterns between seasons. During this time, both males and females preferred to roost in dead and declining trees. No other variable influenced male use, but females also preferred trees located close to water and in forest stands with higher basal area than randomly located trees. Northern myotis with active transmitters never left the study area and were tracked to roosts until early November. During October and November, a female and male were tracked to an underground network of air-filled voids (the Milieu Souterrain Superficiel) we presume to be a hibernaculum. Northern myotis calls were recorded outside this roost between March and October, and bats were observed emerging from this roost during spring and autumn but not summer. Acoustic activity at this site exhibited a seasonal pattern that differed from acoustic activity near roost trees and foraging areas, with a peak of activity during late summer when northern myotis are known to swarm. These data show that northern myotis maternity roosts are used extensively outside of summer and may be vulnerable to forestry practices that occur even outside of the pup-rearing season. These data also support the growing evidence that some northern myotis hibernate outside of caves and mines.

Towards evidence-based conservation of subterranean ecosystems.

Subterranean ecosystems are among the most widespread environments on Earth, yet we still have poor knowledge of their biodiversity. To raise awareness of subterranean ecosystems, the essential services they provide, and their unique conservation challenges, 2021 and 2022 were designated International Years of Caves and Karst. As these ecosystems have traditionally been overlooked in global conservation agendas and multilateral agreements, a quantitative assessment of solution-based approaches to safeguard subterranean biota and associated habitats is timely. This assessment allows researchers and practitioners to understand the progress made and research needs in subterranean ecology and management. We conducted a systematic review of peer-reviewed and grey literature focused on subterranean ecosystems globally (terrestrial, freshwater, and saltwater systems), to quantify the available evidence-base for the effectiveness of conservation interventions. We selected 708 publications from the years 1964 to 2021 that discussed, recommended, or implemented 1,954 conservation interventions in subterranean ecosystems. We noted a steep increase in the number of studies from the 2000s while, surprisingly, the proportion of studies quantifying the impact of conservation interventions has steadily and significantly decreased in recent years. The effectiveness of 31% of conservation interventions has been tested statistically. We further highlight that 64% of the reported research occurred in the Palearctic and Nearctic biogeographic regions. Assessments of the effectiveness of conservation interventions were heavily biased towards indirect measures (monitoring and risk assessment), a limited sample of organisms (mostly arthropods and bats), and more accessible systems (terrestrial caves). Our results indicate that most conservation science in the field of subterranean biology does not apply a rigorous quantitative approach, resulting in sparse evidence for the effectiveness of interventions. This raises the important question of how to make conservation efforts more feasible to implement, cost-effective, and long-lasting. Although there is no single remedy, we propose a suite of potential solutions to focus our efforts better towards increasing statistical testing and stress the importance of standardising study reporting to facilitate meta-analytical exercises. We also provide a database summarising the available literature, which will help to build quantitative knowledge about interventions likely to yield the greatest impacts depending upon the subterranean species and habitats of interest. We view this as a starting point to shift away from the widespread tendency of recommending conservation interventions based on anecdotal and expert-based information rather than scientific evidence, without quantitatively testing their effectiveness.

Cooling of bat hibernacula to mitigate white-nose syndrome.

White-nose syndrome (WNS) is a fungal disease that has caused precipitous declines in several North American bat species, creating an urgent need for conservation. We examined how microclimates and other characteristics of hibernacula have affected bat populations following WNS-associated declines and evaluated whether cooling of warm, little-used hibernacula could benefit bats. During the period following mass mortality (2013-2020), we conducted 191 winter surveys of 25 unmanipulated hibernacula and 6 manipulated hibernacula across Pennsylvania (USA). We joined these data with additional datasets on historical (pre-WNS) bat counts and on the spatial distribution of underground sites. We used generalized linear mixed models and model selection to identify factors affecting bat populations. Winter counts of Myotis lucifugus were higher and increased over time in colder hibernacula (those with midwinter temperatures of 3-6 °C) compared with warmer (7-11 °C) hibernacula. Counts of Eptesicus fuscus, Myotis leibii, and Myotis septentrionalis were likewise higher in colder hibernacula (temperature effects = -0.73 [SE 0.15], -0.51 [0.18], and -0.97 [0.28], respectively). Populations of M. lucifugus and M. septentrionalis increased most over time in hibernacula surrounded by more nearby sites, whereas Eptesicus fuscus counts remained high where they had been high before WNS onset (pre-WNS high count effect = 0.59 [0.22]). Winter counts of M. leibii were higher in hibernacula with high vapor pressure deficits (VPDs) (particularly over 0.1 kPa) compared with sites with lower VPDs (VPD effect = 15.3 [4.6]). Counts of M. lucifugus and E. fuscus also appeared higher where VPD was higher. In contrast, Perimyotis subflavus counts increased over time in relatively warm hibernacula and were unaffected by VPD. Where we manipulated hibernacula, we achieved cooling of on average 2.1 °C. At manipulated hibernacula, counts of M. lucifugus and P. subflavus increased over time (years since manipulation effect = 0.70 [0.28] and 0.51 [0.15], respectively). Further, there were more E. fuscus where cooling was greatest (temperature difference effect = -0.46 [SE 0.11]), and there was some evidence there were more P. subflavus in hibernacula sections that remained warm after manipulation. These data show bats are responding effectively to WNS through habitat selection. In M. lucifugus, M. septentrionalis, and possibly P. subflavus, this response is ongoing, with bats increasingly aggregating at suitable hibernacula, whereas E. fuscus remain in previously favored sites. Our results suggest that cooling warm sites receiving little use by bats is a viable strategy for combating WNS.

El síndrome de nariz blanca (SNB) es una enfermedad fúngica que ha causado declinaciones precipitadas en varias especies de murciélagos norteamericanos, creando una necesidad urgente por conservarlas. Analizamos cómo los microclimas y otras características de los hibernáculos han afectado a las poblaciones de murciélagos después de declinaciones asociadas al SNB y evaluamos si el enfriamiento de hibernáculos cálidos con poco uso podría beneficiar a los murciélagos. Durante el periodo posterior a una mortalidad masiva (2013 - 2020), realizamos 191 censos invernales en 25 hibernáculos sin manipulación y en seis hibernáculos manipulados localizados en Pensilvania (EUA). Juntamos estos datos con conjuntos adicionales de datos de los conteos históricos (previos WNS) de murciélagos y de la distribución espacial de sitios subterráneos. Usamos modelos mixtos lineales generalizados y selección de modelos para identificar los factores que afectan a las poblaciones de murciélagos. Los conteos invernales de Myotis lucifugus fueron más altos e incrementaron con el tiempo en los hibernáculos fríos (aquellos con temperaturas de 3 - 6° C registradas a mitad del invierno) en comparación con los hibernáculos cálidos (7 - 11° C). Los conteos Eptesicus fuscus, M. leibii, y M. septentrionalis fueron igualmente más altos en los hibernáculos fríos (efectos de la temperatura = -0.73 [ES 0.15], -0.51 [0.18], y -0.97 [0.28], respectivamente). Las poblaciones de M. lucifugus y M. septentrionalis fueron las que más incrementaron con el tiempo en los hibernáculos rodeados por más sitios cercanos, mientras que los conteos de E. fuscus permanecieron altos en donde ya habían sido altos antes del comienzo del SNB (el efecto del conteo alto previo al SNB = 0.59 [0.22]). Los conteos invernales de M. leibii fueron más altos en los hibernáculos con altos déficits de presión de vapor (DPV) (particularmente por encima de los 0.1 kPa) en comparación con los sitios con un DPV menor (efecto del VPD = 15.3 [4.6]). Los conteos de M. lucifugus y E. fuscus también fueron más altos en donde el DPV era alto. Al contrario, los conteos de Perimyotis subflavus incrementaron con el tiempo en hibernáculos relativamente cálidos y no se vieron afectados por el DPV. En donde alcanzamos un promedio de enfriamiento de 2.1° C de los hibernáculos, los conteos de M. lucifugus y P. subflavus incrementaron con el tiempo (años desde el efecto de manipulación = 0.70 [0.28] y 0.51 [0.15], respectivamente). Además, encontramos más E. fuscus en donde el enfriamiento fue mayor (efecto de la diferencia en temperatura = −0.46 [ES 0.11]), y hubo algunas evidencias de que había mayor cantidad de P. subflavus en las secciones del hibernáculo que permanecieron cálidas después de la manipulación. Estos datos muestran que los murciélagos están respondiendo efectivamente al SNB mediante la selección de hábitat. En el caso de M. lucifugus, M. septentrionalis y posiblemente P. subflavus, esta respuesta es persistente, con los murciélagos agrupándose cada vez más en hibernáculos adecuados, mientras que E. fuscus permanece en sitios favorecidos previamente. Nuestros resultados sugieren que el enfriamiento de los sitios cálidos que reciben poco uso por parte de los murciélagos es una estrategia viable para combatir al SNB. Enfriamiento de los Hibernáculos de Murciélagos para Mitigar el Síndrome de Nariz Blanca.

Ten-year projection of white-nose syndrome disease dynamics at the southern leading-edge of infection in North America.

Predicting the emergence and spread of infectious diseases is critical for the effective conservation of biodiversity. White-nose syndrome (WNS), an emerging infectious disease of bats, has resulted in high mortality in eastern North America. Because the fungal causative agent Pseudogymnoascus destructans is constrained by temperature and humidity, spread dynamics may vary by geography. Environmental conditions in the southern part of the continent are different than the northeast, where disease dynamics are typically studied, making it difficult to predict how the disease will manifest. Herein, we modelled WNS pathogen spread in Texas based on cave densities and average dispersal distances of hosts, projecting these results out to 10 years. We parameterized a predictive model of WNS epidemiology and its effects on bat populations with observed cave environmental data. Our model suggests that bat populations in northern Texas will be more affected by WNS mortality than southern Texas. As such, we recommend prioritizing the preservation of large overwintering colonies of bats in north Texas through management actions. Our model illustrates that infectious disease spread and infectious disease severity can become uncoupled over a gradient of environmental variation and highlight the importance of understanding host, pathogen and environmental conditions across a breadth of environments.

Social networks based on frequency of roost cohabitation do not reflect association rates of Myotis lucifugus within their roosts.

Bats are a group of mammals well known for forming dynamic social groups. Studies of bat social structures are often based upon the frequency at which bats occupy the same roosts because observing bats directly is not always possible. However, it is not always clear how closely bats occupying the same roost associate with each other, obscuring whether associations result from social relationships or factors such as shared preferences for roosts. Our goal was to determine if bats cohabitating buildings were also found together inside roosts by using anti-collision technology for PIT tags, which enables simultaneous detection of multiple tags. We PIT-tagged 293 female little brown myotis (Myotis lucifugus) and installed antennas within two buildings used as maternity roosts in Yellowstone National Park. Antennas were positioned at roost entryways to generate cohabitation networks and along regions of attic ceilings in each building to generate intraroost networks based on proximity of bats to each other. We found that intraroost and cohabitation networks of buildings were significantly correlated, with the same bats tending to be linked in both networks, but that bats cohabitating the same building often roosted apart, leading to differing assessments of social structure. Cohabitation rates implied that bats associate with a greater number of their roost-mates than was supported by observations within the roost. This caused social networks built upon roost cohabitation rates to be denser, smaller in diameter, and contain nodes with higher average degree centrality. These results show that roost cohabitation does not reflect preference for roost-mates in little brown myotis, as is often inferred from similar studies, and that social network analyses based on cohabitation may provide misleading results.

Summer day-roost selection by eastern red bats varies between areas with different land-use histories.

The eastern red bat (Lasiurus borealis) is widely considered to be in decline, inspiring interest in identifying important habitats for conservation in the eastern United States. Unfortunately, knowledge of important day-roosting habitats is lacking for much of the species' range. We examined patterns of day-roost selection by male and female eastern red bats at two study sites in southeastern Ohio, U. S. A, to help fill this information gap. We radio-tagged 28 male and 25 female bats during the summers of 2016-2019 and located 53 male and 74 female roosts. Day-roost selection differed between sexes and study areas. In a mostly even-aged forest with significant historical disturbance, we found males and females roosting in trees located at higher elevations, with no clear selection based on tree or stand characteristics. Specifically, males selected trees with larger diameters located at lower, cooler elevations than females, which selected smaller diameter trees found at higher, warmer elevations. However, in a forest with less historical disturbance and more structural diversity, we found sexes differed in how they selected from available habitats. These data show that heterogeneity in environmental conditions can lead to different patterns in selection, even between sites located within a small geographic area. They also show that eastern red bats sexually segregate on the local landscape in the presence of diverse forest conditions but may not do so in the absence of such diversity. We recommend managing forests to maintain structural diversity across an elevational gradient to provide male and female eastern red bats with suitable day-roosting habitat in southeast Ohio.

Population Connectivity Predicts Vulnerability to White-Nose Syndrome in the Chilean Myotis (Myotis chiloensis) - A Genomics Approach.

Despite its peculiar distribution, the biology of the southernmost bat species in the world, the Chilean myotis (Myotis chiloensis), has garnered little attention so far. The species has a north-south distribution of c. 2800 km, mostly on the eastern side of the Andes mountain range. Use of extended torpor occurs in the southernmost portion of the range, putting the species at risk of bat white-nose syndrome, a fungal disease responsible for massive population declines in North American bats. Here, we examined how geographic distance and topology would be reflected in the population structure of M. chiloensis along the majority of its range using a double digestion RAD-seq method. We sampled 66 individuals across the species range and discovered pronounced isolation-by-distance. Furthermore, and surprisingly, we found higher degrees of heterozygosity in the southernmost populations compared to the north. A coalescence analysis revealed that our populations may still not have reached secondary contact after the Last Glacial Maximum. As for the potential spread of pathogens, such as the fungus causing WNS, connectivity among populations was noticeably low, especially between the southern hibernatory populations in the Magallanes and Tierra del Fuego, and more northerly populations. This suggests the probability of geographic spread of the disease from the north through bat-to-bat contact to susceptible populations is low. The study presents a rare case of defined population structure in a bat species and warrants further research on the underlying factors contributing to this. See the graphical abstract here. https://doi.org/10.25387/g3.12173385.

Resistance is futile: RNA-sequencing reveals differing responses to bat fungal pathogen in Nearctic Myotis lucifugus and Palearctic Myotis myotis.

Resistance and tolerance allow organisms to cope with potentially life-threatening pathogens. Recently introduced pathogens initially induce resistance responses, but natural selection favors the development of tolerance, allowing for a commensal relationship to evolve. Mycosis by Pseudogymnoascus destructans, causing white-nose syndrome (WNS) in Nearctic hibernating bats, has resulted in population declines since 2006. The pathogen, which spread from Europe, has infected species of Palearctic Myotis for a longer period. We compared ecologically relevant responses to the fungal infection in the susceptible Nearctic M. lucifugus and less susceptible Palearctic M. myotis, to uncover factors contributing to survival differences in the two species. Samples were collected from euthermic bats during arousal from hibernation, a naturally occurring phenomenon, during which transcriptional responses are activated. We compared the whole-transcriptome responses in wild bats infected with P. destructans hibernating in their natural habitat. Our results show dramatically different local transcriptional responses to the pathogen between uninfected and infected samples from the two species. Whereas we found 1526 significantly upregulated or downregulated transcripts in infected M. lucifugus, only one transcript was downregulated in M. myotis. The upregulated response pathways in M. lucifugus include immune cell activation and migration, and inflammatory pathways, indicative of an unsuccessful attempt to resist the infection. In contrast, M. myotis appears to tolerate P. destructans infection by not activating a transcriptional response. These host-microbe interactions determine pathology, contributing to WNS susceptibility, or commensalism, promoting tolerance to fungal colonization during hibernation that favors survival.

Correction: Winter habitats of bats in Texas.

[This corrects the article DOI: 10.1371/journal.pone.0220839.].

Winter habitats of bats in Texas.

Few studies have described winter microclimate selection by bats in the southern United States. This is of particular importance as the cold-adapted fungus, Pseudogymnoascus destructans, which causes the fatal bat disease white-nose syndrome (WNS), continues to spread into southern United States. To better understand the suitability of winter bat habitats for the growth of P. destructans in this region, we collected roost temperature and vapor pressure deficit from 97 hibernacula in six ecoregions in Texas during winter 2016-17 and 2017-18. We also measured skin temperature of Rafinesque's big-eared bats (Corynorhinus townsendii), Townsend's big-eared bats (C. townsendii), big-brown bats (Eptesicus fuscus), southeastern myotis (Myotis austroriparius), cave myotis (M. velifer), tri-colored bats (Perimyotis subflavus), and Mexican free-tailed bats (Tadarida brasiliensis) during hibernation to study their use of torpor in these habitats. We found that temperatures within hibernacula were strongly correlated with external air temperatures and were often within the optimal range of temperatures for P. destructans growth. Hibernacula and skin temperatures differed among species, with Rafinesque's big-eared bats, southeastern myotis, and Mexican free-tailed bats occupying warmer microclimates and having higher torpid skin temperatures. For species that were broadly distributed throughout Texas, hibernacula and skin temperatures differed within species by ecoregion; Tri-colored bats and cave myotis in colder, northern regions occupied colder microclimates within hibernacula and exhibited colder skin temperatures, than individuals of the same species in warmer, southern regions. These data illustrate the variability in microclimates used as hibernacula by bats in Texas and suggest similar variation in susceptibility to WNS in the state. Thus, monitoring microclimates at winter roosts may help predict where WNS may develop, and where management efforts would be most effective.

The Klingon batbugs: Morphological adaptations in the primitive bat bugs, Bucimex chilensis and Primicimex cavernis, including updated phylogeny of Cimicidae.

The Cimicidae is a family of blood-dependent ectoparasites in which dispersion capacity is greatly associated with host movements. Bats are the ancestral and most prevalent hosts for cimicids. Cimicids have a worldwide distribution matching that of their hosts, but the global classification is incomplete, especially for species outside the most common Cimicidae taxa. In this study, we place a little-studied cimicid species, Bucimex chilensis, within a comprehensive molecular phylogeny of Cimicidae by sequencing the genomic regions of this and other closely related species. For this study, we collected B. chilensis females from Myotis chiloensis in Tierra del Fuego, 1,300 km further south than previously known southernmost distribution boundary. We also sequenced COI regions from Primicimex cavernis, a species which together with B. chilensis comprise the entire subfamily Primiciminae. Using Bayesian posterior probability and maximum-likelihood approaches, we found that B. chilensis and P. cavernis clustered close to each other in the molecular analyses, receiving support from similar morphological features, agreeing with the morphology-based taxonomic placement of the two species within the subfamily Primiciminae. We also describe a previously unrecognized morphological adaptation of the tarsal structure, which allows the austral bat ectoparasite, B. chilensis, to cling on to the pelage of its known host, the Chilean myotis (Myotis chiloensis). Through a morphological study and behavioral observation, we elucidate how this tarsal structure operates, and we hypothesize that by clinging in the host pelage, B. chilensis is able to disperse effectively to new areas despite low host density. This is a unique feature shared by P. cavernis, the only other species in Primiciminae.

Effect of torpor on host transcriptomic responses to a fungal pathogen in hibernating bats.

Hibernation, the use of prolonged torpor to depress metabolism, is employed by mammals to conserve resources during extended periods of extreme temperatures and/or resource limitation. Mammalian hibernators arouse to euthermy periodically during torpor for reasons that are not well understood, and these arousals may facilitate immune processes. To determine whether arousals enable host responses to pathogens, we used dual RNA-Seq and a paired sampling approach to examine gene expression in a hibernating bat, the little brown myotis (Myotis lucifugus). During torpor, transcript levels differed in only a few genes between uninfected wing tissue and adjacent tissue infected with Pseudogymnoascus destructans, the fungal pathogen that causes white-nose syndrome. Within 70-80 min after emergence from torpor, large changes in gene expression were observed due to local infection, particularly in genes involved in pro-inflammatory host responses to fungal pathogens, but also in many genes involved in immune responses and metabolism. These results support the hypothesis that torpor is a period of relative immune dormancy and arousals allow for local immune responses in infected tissues during hibernation. Host-pathogen interactions were also found to regulate gene expression in the pathogen differently depending on the torpor state of the host. Hibernating species must balance the benefits of energy and water conservation achieved during torpor with the costs of decreased immune competence. Interbout arousals allow hibernators to optimize these, and other, trade-offs during prolonged hibernation by enabling host responses to pathogens within brief, periodic episodes of euthermy.

BATS RECOVERING FROM WHITE-NOSE SYNDROME ELEVATE METABOLIC RATE DURING WING HEALING IN SPRING.

Host responses to infection with novel pathogens are costly and require trade-offs among physiologic systems. One such pathogen is the fungus Pseudogymnoascus destructans (Pd) that causes white-nose syndrome (WNS) and has led to mass mortality of hibernating bats in eastern North America. Although infection with Pd does not always result in death, we hypothesized that bats that survive infection suffer significant consequences that negatively impact the ability of females to reproduce. To understand the physiologic consequences of surviving infection with Pd, we assessed differences in wing damage, mass-specific resting metabolic rate, and reproductive rate between little brown myotis ( Myotis lucifugus) that survived a winter in captivity after inoculation with Pd (WNS survivors) and comparable, uninfected bats. Survivors of WNS had significantly more damaged wing tissue and displayed elevated mass-specific metabolic rates compared with Pd-uninfected bats after emergence from hibernation. The WNS survivors and Pd-uninfected bats did not significantly differ in their reproductive capacity, at least in captivity. However, our metabolic data demonstrated greater energetic costs during spring in WNS survivors compared with uninfected bats, which may have led to other consequences for postpartum fitness. We suggest that, after surviving the energetic constraints of winter, temperate hibernating bats infected with Pd faced a second energetic bottleneck after emerging from hibernation.

Energy conserving thermoregulatory patterns and lower disease severity in a bat resistant to the impacts of white-nose syndrome.

The devastating bat fungal disease, white-nose syndrome (WNS), does not appear to affect all species equally. To experimentally determine susceptibility differences between species, we exposed hibernating naïve little brown myotis (Myotis lucifugus) and big brown bats (Eptesicus fuscus) to the fungus that causes WNS, Pseudogymnoascus destructans (Pd). After hibernating under identical conditions, Pd lesions were significantly more prevalent and more severe in little brown myotis. This species difference in pathology correlates with susceptibility to WNS in the wild and suggests that survival is related to different host physiological responses. We observed another fungal infection, associated with neutrophilic inflammation, that was equally present in all bats. This suggests that both species are capable of generating a response to cold tolerant fungi and that Pd may have evolved mechanisms for evading host responses that are effective in at least some bat species. These host-pathogen interactions are likely mediated not just by host physiological responses, but also by host behavior. Pd-exposed big brown bats, the less affected species, spent more time in torpor than did control animals, while little brown myotis did not exhibit this change. This differential thermoregulatory response to Pd infection by big brown bat hosts may allow for a more effective (or less pathological) immune response to tissue invasion.

The White-Nose Syndrome Transcriptome: Activation of Anti-fungal Host Responses in Wing Tissue of Hibernating Little Brown Myotis.

White-nose syndrome (WNS) in North American bats is caused by an invasive cutaneous infection by the psychrophilic fungus Pseudogymnoascus destructans (Pd). We compared transcriptome-wide changes in gene expression using RNA-Seq on wing skin tissue from hibernating little brown myotis (Myotis lucifugus) with WNS to bats without Pd exposure. We found that WNS caused significant changes in gene expression in hibernating bats including pathways involved in inflammation, wound healing, and metabolism. Local acute inflammatory responses were initiated by fungal invasion. Gene expression was increased for inflammatory cytokines, including interleukins (IL) IL-1ß, IL-6, IL-17C, IL-20, IL-23A, IL-24, and G-CSF and chemokines, such as Ccl2 and Ccl20. This pattern of gene expression changes demonstrates that WNS is accompanied by an innate anti-fungal host response similar to that caused by cutaneous Candida albicans infections. However, despite the apparent production of appropriate chemokines, immune cells such as neutrophils and T cells do not appear to be recruited. We observed upregulation of acute inflammatory genes, including prostaglandin G/H synthase 2 (cyclooxygenase-2), that generate eicosanoids and other nociception mediators. We also observed differences in Pd gene expression that suggest host-pathogen interactions that might determine WNS progression. We identified several classes of potential virulence factors that are expressed in Pd during WNS, including secreted proteases that may mediate tissue invasion. These results demonstrate that hibernation does not prevent a local inflammatory response to Pd infection but that recruitment of leukocytes to the site of infection does not occur. The putative virulence factors may provide novel targets for treatment or prevention of WNS. These observations support a dual role for inflammation during WNS; inflammatory responses provide protection but excessive inflammation may contribute to mortality, either by affecting torpor behavior or causing damage upon emergence in the spring.

Sex and hibernaculum temperature predict survivorship in white-nose syndrome affected little brown myotis (Myotis lucifugus).

White-nose syndrome (WNS), an emerging infectious disease caused by the novel fungus Pseudogymnoascus destructans, has devastated North American bat populations since its discovery in 2006. The little brown myotis, Myotis lucifugus, has been especially affected. The goal of this 2-year captive study was to determine the impact of hibernacula temperature and sex on WNS survivorship in little brown myotis that displayed visible fungal infection when collected from affected hibernacula. In study 1, we found that WNS-affected male bats had increased survival over females and that bats housed at a colder temperature survived longer than those housed at warmer temperatures. In study 2, we found that WNS-affected bats housed at a colder temperature fared worse than unaffected bats. Our results demonstrate that WNS mortality varies among individuals, and that colder hibernacula are more favourable for survival. They also suggest that female bats may be more negatively affected by WNS than male bats, which has important implications for the long-term survival of the little brown myotis in eastern North America.

Antibodies to Pseudogymnoascus destructans are not sufficient for protection against white-nose syndrome.

White-nose syndrome (WNS) is a fungal disease caused by Pseudogymnoascus destructans (Pd) that affects bats during hibernation. Although millions of bats have died from WNS in North America, mass mortality has not been observed among European bats infected by the fungus, leading to the suggestion that bats in Europe are immune. We tested the hypothesis that an antibody-mediated immune response can provide protection against WNS by quantifying antibodies reactive to Pd in blood samples from seven species of free-ranging bats in North America and two free-ranging species in Europe. We also quantified antibodies in blood samples from little brown myotis (Myotis lucifugus) that were part of a captive colony that we injected with live Pd spores mixed with adjuvant, as well as individuals surviving a captive Pd infection trial. Seroprevalence of antibodies against Pd, as well as antibody titers, was greater among little brown myotis than among four other species of cave-hibernating bats in North America, including species with markedly lower WNS mortality rates. Among little brown myotis, the greatest titers occurred in populations occupying regions with longer histories of WNS, where bats lacked secondary symptoms of WNS. We detected antibodies cross-reactive with Pd among little brown myotis naïve to the fungus. We observed high titers among captive little brown myotis injected with Pd. We did not detect antibodies against Pd in Pd-infected European bats during winter, and titers during the active season were lower than among little brown myotis. These results show that antibody-mediated immunity cannot explain survival of European bats infected with Pd and that little brown myotis respond differently to Pd than species with higher WNS survival rates. Although it appears that some species of bats in North America may be developing resistance to WNS, an antibody-mediated immune response does not provide an explanation for these remnant populations.