Migratory bats are sensitive to magnetic inclination changes during the compass calibration period.

The Earth's magnetic field is used as a navigational cue by many animals. For mammals, however, there are few data to show that navigation ability relies on sensing the natural magnetic field. In night-time migrating bats, experiments demonstrating a role for the solar azimuth at sunset in the calibration of the orientation system suggest that the magnetic field is a candidate for their compass. Here, we investigated how an altered magnetic field at sunset changes the nocturnal orientation of the bat Pipistrellus pygmaeus. We exposed bats to either the natural magnetic field, a horizontally shifted field (120°), or the same shifted field combined with a reversal of the natural value of inclination (70° to -70°). We later released the bats and found that the take-off orientation differed among all treatments. Bats that were exposed to the 120° shift were unimodally oriented northwards in contrast to controls which exhibited a bimodal north-south distribution. Surprisingly, the orientation of bats exposed to both a 120° shift and reverse inclination was indistinguishable from a uniform distribution. These results suggest that these migratory bats calibrate the magnetic field at sunset, and for the first time, they show that bats are sensitive to the angle of magnetic inclination.

Sense of doubt: inaccurate and alternate locations of virtual magnetic displacements may give a distorted view of animal magnetoreception ability.

Virtual magnetic displacements are used to examine the magnetoreceptive ability of animals by changing the local magnetic field to emulate one that exists elsewhere. This technique can be used to test whether animals use a magnetic map. The viability of a magnetic map is dependant upon which magnetic parameters an animal's coordinate system is composed of, and how sensitive they are to those parameters. Previous research has not considered the degree to which sensitivity can change an animal's impression of where a virtual magnetic displacement is located. We re-assessed all published studies that use virtual magnetic displacements assuming the highest likely level of sensitivity to magnetic parameters in animals. The vast majority are susceptible to the existence of alternate possible virtual locations. In some cases, this can cause results to become ambiguous. We present a tool for visualising all possible virtual magnetic displacement alternative locations (ViMDAL) and propose changes to how further research on animal magnetoreception is conducted and reported.

High vulnerability of juvenile Nathusius' pipistrelle bats (Pipistrellus nathusii) at wind turbines.

Large numbers of bats are killed by wind turbines globally, yet the specific demographic consequences of wind turbine mortality are still unclear. In this study, we compared characteristics of Nathusius' pipistrelles (Pipistrellus nathusii) killed at wind turbines (N = 119) to those observed within the live population (N = 524) during the summer migration period in Germany. We used generalized linear mixed-effects modeling to identify demographic groups most vulnerable to wind turbine mortality, including sex (female or male), age (adult or juvenile), and geographic origin (regional or long-distance migrant; depicted by fur stable hydrogen isotope ratios). Juveniles contributed with a higher proportion of carcasses at wind turbines than expected given their frequency in the live population suggesting that juvenile bats may be particularly vulnerable to wind turbine mortality. This effect varied with wind turbine density. Specifically, at low wind turbine densities, representing mostly inland areas with water bodies and forests where Nathusius' pipistrelles breed, juveniles were found more often dead beneath turbines than expected based on their abundance in the live population. At high wind turbine densities, representing mostly coastal areas where Nathusius' pipistrelles migrate, adults and juveniles were equally vulnerable. We found no evidence of increased vulnerability to wind turbines in either sex, yet we observed a higher proportion of females than males among both carcasses and the live population, which may reflect a female bias in the live population most likely caused by females migrating from their northeastern breeding areas migrating into Germany. A high mortality of females is conservation concern for this migratory bat species because it affects the annual reproduction rate of populations. A distant origin did not influence the likelihood of getting killed at wind turbines. A disproportionately high vulnerability of juveniles to wind turbine mortality may reduce juvenile recruitment, which may limit the resilience of Nathusius' pipistrelles to environmental stressors such as climate change or habitat loss. Schemes to mitigate wind turbine mortality, such as elevated cut-in speeds, should be implemented throughout Europe to prevent population declines of Nathusius' pipistrelles and other migratory bats.

Corneal sensitivity is required for orientation in free-flying migratory bats.

The exact anatomical location for an iron particle-based magnetic sense remains enigmatic in vertebrates. For mammals, findings from a cornea anaesthesia experiment in mole rats suggest that it carries the primary sensors for magnetoreception. Yet, this has never been tested in a free-ranging mammal. Here, we investigated whether intact corneal sensation is crucial for navigation in migrating Nathusius' bats, Pipistrellus nathusii, translocated from their migratory corridor. We found that bats treated with corneal anaesthesia in both eyes flew in random directions after translocation and release, contrasting bats with a single eye treated, and the control group, which both oriented in the seasonally appropriate direction. Using a Y-maze test, we confirmed that light detection remained unaffected by topical anaesthesia. Therefore our results suggest the cornea as a possible site of magnetoreception in bats, although other conceivable effects of the anaesthetic are also explored. Furthermore, we demonstrate that the corneal based sense is of bilateral nature but can function in a single eye if necessary.

In situ novel environment assay reveals acoustic exploration as a repeatable behavioral response in migratory bats.

Integrating information on species-specific sensory perception with spatial activity provides a high-resolution understanding of how animals explore environments, yet frequently used exploration assays commonly ignore sensory acquisition as a measure for exploration. Echolocation is an active sensing system used by hundreds of mammal species, primarily bats. As echolocation call activity can be reliably quantified, bats present an excellent model system to investigate intraspecific variation in environmental cue sampling. Here, we developed an in situ roost-like novel environment assay for tree-roosting bats. We repeatedly tested 52 individuals of the migratory bat species, Pipistrellus nathusii, across 24 h, to examine the role of echolocation when crawling through a maze-type arena and test for consistent intraspecific variation in sensory-based exploration. We reveal a strong correlation between echolocation call activity and spatial activity. Moreover, we show that during the exploration of the maze, individuals consistently differed in spatial activity as well as echolocation call activity, given their spatial activity, a behavioral response we term 'acoustic exploration'. Acoustic exploration was correlated with other exploratory behaviors, but not with emergence latency. We here present a relevant new measure for exploration behavior and provide evidence for consistent (short-term) intra-specific variation in the level at which wild bats collect information from a novel environment.

Identifying migratory pathways of Nathusius' pipistrelles (Pipistrellus nathusii) using stable hydrogen and strontium isotopes.

RATIONALE: Identifying migratory corridors of animals is essential for their effective protection, yet the exact location of such corridors is often unknown, particularly for elusive animals such as bats. While migrating along the German coastline, Nathusius' pipistrelles (Pipistrellus nathusii) are regularly killed at wind turbines. Therefore, we explored the paths taken on their annual journey. METHODS: We used isotope ratio mass spectrometry to measure stable hydrogen and strontium isotope ratios in fur keratin of 59 Nathusius' pipistrelles captured on three offshore islands. Samples were pre-treated before analysis to report exclusively stable isotope ratios of non-exchangeable hydrogen. We generated maps to predict summer origins of bats using isoscape models. RESULTS: Bats were classified as long-distance migrants, mostly originating from Eastern Europe. Hydrogen analysis suggested for some bats a possible Fennoscandian origin, yet additional information from strontium analysis excluded this possibility. Instead, our data suggest that most Nathusius' pipistrelles migrating along the German coastline were of continental European summer origin, but also highlight the possibility that Nathusius' pipistrelles of Baltorussian origin may travel offshore from Fennoscandia to Germany. CONCLUSIONS: Our findings demonstrate the benefit of using complementary isotopic tracers for analysing the migratory pathways of bats and also potentially other terrestrial vertebrate species. Furthermore, data from our study suggest an offset of fur strontium isotope ratios in relation to local bedrock.

The immune response of bats differs between pre-migration and migration seasons.

Maintaining a competent immune system is energetically costly and thus immunity may be traded against other costly traits such as seasonal migration. Here, we tested in long-distance migratory Nathusius' pipistrelles (Pipistrellus nathusii), if selected branches of immunity are expressed differently in response to the energy demands and oxidative stress of aerial migration. During the migration period, we observed higher baseline lymphocyte and lower neutrophil levels than during the pre-migration period, but no stronger response of cellular effectors to an antigen challenge. Baseline plasma haptoglobin, as a component of the humoral innate immunity, remained similar during both seasons, yet baseline plasma haptoglobin levels increased by a factor of 7.8 in migratory bats during an immune challenge, whereas they did not change during the pre-migration period. Oxidative stress was higher during migration than during pre-migration, yet there was no association between blood oxidative status and immune parameters, and immune challenge did not trigger any changes in oxidative stress, irrespective of season. Our findings suggest that humoral effectors of the acute phase response may play a stronger role in the first-line defense against infections for migrating bats compared to non-migrating bats. We conclude that Nathusius' pipistrelles allocate resources differently into the branches of their immune system, most likely following current demands resulting from tight energy budgets during migration.

Migratory flight imposes oxidative stress in bats.

Many animal species migrate over long distances, but the physiological challenges of migration are poorly understood. It has recently been suggested that increased molecular oxidative damage might be one important challenge for migratory animals. We tested the hypothesis that autumn migration imposes an oxidative challenge to bats by comparing values of 4 blood-based markers of oxidative status (oxidative damage and both enzymatic and nonenzymatic antioxidants) between Nathusius' bats Pipistrellus nathusii that were caught during migration flights with those measured in conspecifics after resting for 18 or 24 h. Experiments were carried out at Pape Ornithological Station in Pape (Latvia) in 2016 and 2017. Our results show that flying bats have a blood oxidative status different from that of resting bats due to higher oxidative damage and different expression of both nonenzymatic and enzymatic antioxidants (glutathione peroxidase). The differences in oxidative status markers varied between sampling years and were independent from individual body condition or sex. Our work provides evidence that migratory flight might impose acute oxidative stress to bats and that resting helps animals to recover from oxidative damage accrued en route. Our data suggest that migrating bats and birds might share similar strategies of mitigating and recovering from oxidative stress.

Experienced Migratory Bats Integrate the Sun's Position at Dusk for Navigation at Night.

From bats to whales, millions of mammals migrate every year. However, their navigation capacity for accomplishing long-distance movements remains remarkably understudied and lags behind by five decades compared to other animals [1, 2]-partly because, unlike for other taxa, such as birds and sea turtles, no small-scale orientation assay has so far been developed. Yet recently, bats became a model to investigate which cues mammals use for long-range navigation, and, surprisingly for nocturnal animals, sunset cues, and particularly polarized-light cues, appear to be crucial for calibration of the magnetic-compass system in non-migratory bats [3-5]. This does not appear to hold for a species of migratory bat, however [6], and thus the nature of the information used by migratory bats for navigation remains unclear. Here, we asked whether the position of the solar disk per se is relevant for compass orientation in a migratory bat, Pipistrellus pygmaeus. Using a new experimental assay that measures takeoff orientation, we tested the orientation of bats exposed to a shifted sunset azimuth using a mirror at dusk. Bats exposed to a 180°-rotated azimuth of the setting sun and released after translocation during the same night shifted their heading direction by ∼180° compared to control bats. However, first-year migrants had no clear orientation either as controls or after the same treatment. This suggests that learning the migratory direction is a key component in the navigational system of naive bats in this species. Our study provides rare evidence for the specific cues and mechanisms that migratory mammals use for navigation.

Migratory bats are attracted by red light but not by warm-white light: Implications for the protection of nocturnal migrants.

The replacement of conventional lighting with energy-saving light emitting diodes (LED) is a worldwide trend, yet its consequences for animals and ecosystems are poorly understood. Strictly nocturnal animals such as bats are particularly sensitive to artificial light at night (ALAN). Past studies have shown that bats, in general, respond to ALAN according to the emitted light color and that migratory bats, in particular, exhibit phototaxis in response to green light. As red and white light is frequently used in outdoor lighting, we asked how migratory bats respond to these wavelength spectra. At a major migration corridor, we recorded the presence of migrating bats based on ultrasonic recorders during 10-min light-on/light-off intervals to red or warm-white LED, interspersed with dark controls. When the red LED was switched on, we observed an increase in flight activity for Pipistrellus pygmaeus and a trend for a higher activity for Pipistrellus nathusii. As the higher flight activity of bats was not associated with increased feeding, we rule out the possibility that bats foraged at the red LED light. Instead, bats may have flown toward the red LED light source. When exposed to warm-white LED, general flight activity at the light source did not increase, yet we observed an increased foraging activity directly at the light source compared to the dark control. Our findings highlight a response of migratory bats toward LED light that was dependent on light color. The most parsimonious explanation for the response to red LED is phototaxis and for the response to warm-white LED foraging. Our findings call for caution in the application of red aviation lighting, particularly at wind turbines, as this light color might attract bats, leading eventually to an increased collision risk of migratory bats at wind turbines.

Simulated bacterial infection disrupts the circadian fluctuation of immune cells in wrinkle-lipped bats (Chaerephon plicatus).

BACKGROUND: Leukocyte concentrations follow a circadian pattern in mammals, with elevated values at times of potential contact with pathogens and parasites. We hypothesized that this pattern is disturbed after an immune challenge. METHODS: In Thailand, we captured wrinkle-lipped bats (Chaerephon plicatus), when they returned to their colony at dawn. We challenged half of the animals (experimental group) with bacterial lipopolysaccharides and treated the others only with the carrier liquid (control group). We then compared body mass changes and differences in circulating immune cell counts at 8 h post-treatment. RESULTS: In experimental animals, we observed an increase in total leukocyte and neutrophil numbers of 17% and 95%, respectively. In control animals, concentrations of leukocytes decreased by 44% and those of neutrophils remained constant. Experimental treatment had no effect on lymphocytes, yet changes in eosinophil numbers were explained by sex. Eosinophils decreased by 66% in females and by 62% in males. Basophils and monocytes were rarest among all observed cell types and analysis was either impossible because of low numbers or yielded no significant effects, respectively. DISCUSSION: Our findings show that a simulated bacterial infection triggered a neutrophil-associated immune response in wrinkle-lipped bats, indicating a disruption of the diurnal fluctuation of immune cells. Our study suggests that bats exhibit circadian rhythms in immune cell counts. The magnitude of these fluctuations may vary across species according to specific-specific infection risks associated with colony sizes or specific roosting habits.

Habitat use of migratory bats killed during autumn at wind turbines.

The killing of large numbers of migratory bats at wind turbines is a pressing conservation problem. Even though avoidance and mitigation measures could benefit from a better knowledge of the species' migratory habits, we lack basic information about what habitats and corridors bats use during migration. We studied the isotopic niche dimensions of three bat species that are frequently killed at wind turbines in Germany: non-migratory Pipistrellus pipistrellus, mid-distance migratory Nyctalus noctula, and long- distance migratory Pipistrellus nathusii. We measured stable carbon and nitrogen isotope ratios (δ¹³C, δ¹5N) in five tissues that differed in isotopic retention time (fur, wing membrane tissue, muscle, liver, blood) to shed light on the species-specific habitat use during the autumn migration period using standard ellipse areas (SEAc). Further, we used stable isotope ratios of non-exchangeable hydrogen (δ²H(K)) in fur keratin to assess the breeding origin of bats. We inferred from isotopic composition (δ¹³C, δ¹5N) of fur keratin that isotopic niche dimensions of P. nathusii was distinct from that of N. noctula and P. pipistrellus, probably because P. nathusii was using more aquatic habitats than the other two species. Isoscape origin models supported that traveled distances before dying at wind turbines was largest for P. nathusii, intermediate for N. noctula, and shortest for P. pipistrellus. Isotopic niche dimensions calculated for each sample type separately reflected the species' migratory behavior. Pipistrellus pipistrellus and N. noctula showed similar isotopic niche breadth across all tissue types, whereas SEAc values of P. nathusii increased in tissues with slow turnaround time. Isotopic data suggested that P. nathusii consistently used aquatic habitats throughout the autumn period, whereas N. noctula showed a stronger association with terrestrial habitats during autumn compared to the pre-migration period.

Polarized skylight does not calibrate the compass system of a migratory bat.

In a recent study, Greif et al. (Greif et al. Nat Commun 5, 4488. (doi:10.1038/ncomms5488)) demonstrated a functional role of polarized light for a bat species confronted with a homing task. These non-migratory bats appeared to calibrate their magnetic compass by using polarized skylight at dusk, yet it is unknown if migratory bats also use these cues for calibration. During autumn migration, we equipped Nathusius' bats, Pipistrellus nathusii, with radio transmitters and tested if experimental animals exposed during dusk to a 90° rotated band of polarized light would head in a different direction compared with control animals. After release, bats of both groups continued their journey in the same direction. This observation argues against the use of a polarization-calibrated magnetic compass by this migratory bat and questions that the ability of using polarized light for navigation is a consistent feature in bats. This finding matches with observations in some passerine birds that used polarized light for calibration of their magnetic compass before but not during migration.

Wind farm facilities in Germany kill noctule bats from near and far.

Over recent years, it became widely accepted that alternative, renewable energy may come at some risk for wildlife, for example, when wind turbines cause large numbers of bat fatalities. To better assess likely populations effects of wind turbine related wildlife fatalities, we studied the geographical origin of the most common bat species found dead below German wind turbines, the noctule bat (Nyctalus noctula). We measured stable isotope ratios of non-exchangeable hydrogen in fur keratin to separate migrants from local individuals, used a linear mixed-effects model to identify temporal, spatial and biological factors explaining the variance in measured stable isotope ratios and determined the geographical breeding provenance of killed migrants using isoscape origin models. We found that 72% of noctule bat casualties (n = 136) were of local origin, while 28% were long-distance migrants. These findings highlight that bat fatalities at German wind turbines may affect both local and distant populations. Our results indicated a sex and age-specific vulnerability of bats towards lethal accidents at turbines, i.e. a relatively high proportion of killed females were recorded among migratory individuals, whereas more juveniles than adults were recorded among killed bats of local origin. Migratory noctule bats were found to originate from distant populations in the Northeastern parts of Europe. The large catchment areas of German wind turbines and high vulnerability of female and juvenile noctule bats call for immediate action to reduce the negative cross-boundary effects of bat fatalities at wind turbines on local and distant populations. Further, our study highlights the importance of implementing effective mitigation measures and developing species and scale-specific conservation approaches on both national and international levels to protect source populations of bats. The efficacy of local compensatory measures appears doubtful, at least for migrant noctule bats, considering the large geographical catchment areas of German wind turbines for this species.