The genome sequence of Tadarida brasiliensis I. Geoffroy Saint-Hilaire, 1824 [Molossidae; Tadarida].

We present a genome assembly from an individual male Tadarida brasiliensis (The Brazilian free-tailed bat; Chordata; Mammalia; Chiroptera; Molossidae). The genome sequence is 2.28 Gb in span. The majority of the assembly is scaffolded into 25 chromosomal pseudomolecules, with the X and Y sex chromosomes assembled.

The prefrontal cortex of the Mexican free-tailed bat is more selective to communication calls than primary auditory cortex.

In this study, we examined the auditory responses of a prefrontal area, the frontal auditory field (FAF), of an echolocating bat (Tadarida brasiliensis) and presented a comparative analysis of the neuronal response properties between the FAF and the primary auditory cortex (A1). We compared single-unit responses from the A1 and the FAF elicited by pure tones, downward frequency-modulated sweeps (dFMs), and species-specific vocalizations. Unlike the A1, FAFs were not frequency tuned. However, progressive increases in dFM sweep rate elicited a systematic increase of response precision, a phenomenon that does not take place in the A1. Call selectivity was higher in the FAF versus A1. We calculated the neuronal spectrotemporal receptive fields (STRFs) and spike-triggered averages (STAs) to predict responses to the communication calls and provide an explanation for the differences in call selectivity between the FAF and A1. In the A1, we found a high correlation between predicted and evoked responses. However, we did not generate reasonable STRFs in the FAF, and the prediction based on the STAs showed lower correlation coefficient than that of the A1. This suggests nonlinear response properties in the FAF that are stronger than the linear response properties in the A1. Stimulating with a call sequence increased call selectivity in the A1, but it remained unchanged in the FAF. These data are consistent with a role for the FAF in assessing distinctive acoustic features downstream of A1, similar to the role proposed for primate ventrolateral prefrontal cortex.NEW & NOTEWORTHY In this study, we examined the neuronal responses of a frontal cortical area in an echolocating bat to behaviorally relevant acoustic stimuli and compared them with those in the primary auditory cortex (A1). In contrast to the A1, neurons in the bat frontal auditory field are not frequency tuned but showed a higher selectivity for social signals such as communication calls. The results presented here indicate that the frontal auditory field may represent an additional processing center for behaviorally relevant sounds.

Singing strategies are linked to perch use on foraging territories in heart-nosed bats.

Acoustic communication allows animals to coordinate and optimize resource utilization in space. Cardioderma cor, the heart-nosed bat, is one of the few species of bats known to sing during nighttime foraging. Previous research found that heart-nosed bats react aggressively to song playback, supporting the territorial defense hypothesis of singing in this species. We further investigated the territorial defense hypothesis from an ecological standpoint, which predicts that singing should be associated with exclusive areas containing a resource, by tracking 14 individuals nightly during the dry seasons in Tanzania. We quantified the singing behavior of individuals at all perches used throughout the night. Using home range analysis tools, we quantified overall use, night ranges and singing ranges, as well as areas used in early and later time periods at night. Males sang back and forth from small ( x ¯  = 3.48 ± 2.71 ha), largely exclusive areas that overlapped with overall night ranges used for gleaning prey. Individuals varied in singing effort; however, all sang significantly more as night progressed. Subsequently, areas used earlier at night and overall use areas were both larger than singing areas. Individuals varied in singing strategies. Some males sang for long periods in particular trees and had smaller core areas, while others moved frequently among singing trees. The most prolific singers used more perches overall. Our results support the hypothesis that acoustic communication repertoires evolved in support of stable foraging territory advertisement and defense in some bats.

Faster Repetition Rate Sharpens the Cortical Representation of Echo Streams in Echolocating Bats.

There is consensus that primary auditory cortex (A1) utilizes a combination of rate codes and temporally precise population codes to represent discreet auditory objects. During the response to auditory streams, forward suppression constrains cortical rate coding strategies, but it may also be well positioned to enhance temporal coding strategies that rely on synchronized firing across neural ensembles. Here, we exploited the rapid temporal dynamics of bat echolocation to investigate how forward suppression modulates the cortical ensemble representation of complex acoustic signals embedded in echo streams. We recorded from auditory cortex of anesthetized free-tailed bats while stimulating the auditory system with naturalistic biosonar pulse-echo sequences covering a range of pulse emission rates. As expected, increasing pulse repetition rate significantly reduced the number of spikes per echo stimulus, but it also increased spike timing precision and doubled the information gain. This increased spike-timing precision translated into more robust inter-neuronal synchronization patterns with >10-dB higher signal-to-noise ratios (SNRs) at the ensemble level. We propose that forward suppression dynamically mediates a trade-off between the sensitive detection of isolated sounds versus precise spatiotemporal encoding of ongoing sound sequences in auditory cortex.

Flying bats use serial sampling to locate odour sources.

Olfactory tracking generally sacrifices speed for sensitivity, but some fast-moving animals appear surprisingly efficient at foraging by smell. Here, we analysed the olfactory tracking strategies of flying bats foraging for fruit. Fruit- and nectar-feeding bats use odour cues to find food despite the sensory challenges derived from fast flight speeds and echolocation. We trained Jamaican fruit-eating bats (Artibeus jamaicensis) to locate an odour reward and reconstructed their flight paths in three-dimensional space. Results confirmed that bats relied upon olfactory cues to locate a reward. Flight paths revealed a combination of odour- and memory-guided search strategies. During 'inspection flights', bats significantly reduced flight speeds and flew within approximately 6 cm of possible targets to evaluate the presence or absence of the odour cue. This behaviour combined with echolocation explains how bats maximize foraging efficiency while compensating for trade-offs associated with olfactory detection and locomotion.

Olfactory tracking strategies in a neotropical fruit bat.

Many studies have characterized olfactory-tracking behaviors in animals, and it has been proposed that search strategies may be generalizable across a wide range of species. Olfaction is important for fruit- and nectar-feeding bats, but it is uncertain whether existing olfactory search models can predict the strategies of flying mammals that emit echolocation pulses through their nose. Quantitative assessments of how well echolocating bats track and localize odor sources are lacking, so we developed a behavioral assay to characterize the olfactory detection and tracking behavior of crawling northern yellow-shouldered bats (Sturnira parvidens), a common neotropical frugivore. Trained bats were presented with a choice between control and banana-odor-infused solutions in a series of experiments that confirmed that bats are able to locate a reward based on odor cues alone and examined the effect of odor concentration on olfactory search behaviors. Decision distance (the distance from which bats made their change in direction before directly approaching the target) was distinctly bimodal, with an observed peak that coincided with an inflection point in the odor concentration gradient. We observed two main search patterns that are consistent with both serial sampling and learned route-following strategies. These results support the hypothesis that bats can combine klinotaxis with spatial awareness of experimental conditions to locate odor sources, similar to terrestrial mammals. Contrary to existing models, bats did not display prominent head-scanning behaviors during their final approach, which may be due to constraints of nasal-emitted biosonar for orientation.

Temporal coding of echo spectral shape in the bat auditory cortex.

Echolocating bats rely upon spectral interference patterns in echoes to reconstruct fine details of a reflecting object's shape. However, the acoustic modulations required to do this are extremely brief, raising questions about how their auditory cortex encodes and processes such rapid and fine spectrotemporal details. Here, we tested the hypothesis that biosonar target shape representation in the primary auditory cortex (A1) is more reliably encoded by changes in spike timing (latency) than spike rates and that latency is sufficiently precise to support a synchronization-based ensemble representation of this critical auditory object feature space. To test this, we measured how the spatiotemporal activation patterns of A1 changed when naturalistic spectral notches were inserted into echo mimic stimuli. Neurons tuned to notch frequencies were predicted to exhibit longer latencies and lower mean firing rates due to lower signal amplitudes at their preferred frequencies, and both were found to occur. Comparative analyses confirmed that significantly more information was recoverable from changes in spike times relative to concurrent changes in spike rates. With this data, we reconstructed spatiotemporal activation maps of A1 and estimated the level of emerging neuronal spike synchrony between cortical neurons tuned to different frequencies. The results support existing computational models, indicating that spectral interference patterns may be efficiently encoded by a cascading tonotopic sequence of neural synchronization patterns within an ensemble of network activity that relates to the physical features of the reflecting object surface.

Functional organization of the primary auditory cortex of the free-tailed bat Tadarida brasiliensis.

The Mexican free-tailed bat, Tadarida brasiliensis, is a fast-flying bat that hunts by biosonar at high altitudes in open space. The auditory periphery and ascending auditory pathways have been described in great detail for this species, but nothing is yet known about its auditory cortex. Here we describe the topographical organization of response properties in the primary auditory cortex (AC) of the Mexican free-tailed bat with emphasis on the sensitivity for FM sweeps and echo-delay tuning. Responses of 716 units to pure tones and of 373 units to FM sweeps and FM-FM pairs were recorded extracellularly using multielectrode arrays in anesthetized bats. A general tonotopy was confirmed with low frequencies represented caudally and high frequencies represented rostrally. Characteristic frequencies (CF) ranged from 15 to 70 kHz, and fifty percent of CFs fell between 20 and 30 kHz, reflecting a hyper-representation of a bandwidth corresponding to search-phase echolocation pulses. Most units showed a stronger response to downward rather than upward FM sweeps and forty percent of the neurons interspersed throughout AC (150/371) showed echo-delay sensitivity to FM-FM pairs. Overall, the results illustrate that the free-tailed bat auditory cortex is organized similarly to that of other FM-type insectivorous bats.

Role of ecology in shaping external nasal morphology in bats and implications for olfactory tracking.

Many animals display morphological adaptations of the nose that improve their ability to detect and track odors. Bilateral odor sampling improves an animals' ability to navigate using olfaction and increased separation of the nostrils facilitates olfactory source localization. Many bats use odors to find food and mates and bats display an elaborate diversity of facial features. Prior studies have quantified how variations in facial features correlate with echolocation and feeding ecology, but surprisingly none have asked whether bat noses might be adapted for olfactory tracking in flight. We predicted that bat species that rely upon odor cues while foraging would have greater nostril separation in support of olfactory tropotaxis. Using museum specimens, we measured the external nose and cranial morphology of 40 New World bat species. Diet had a significant effect on external nose morphology, but contrary to our predictions, insectivorous bats had the largest relative separation of nostrils, while nectar feeding species had the narrowest nostril widths. Furthermore, nasal echolocating bats had significantly narrower nostrils than oral emitting bats, reflecting a potential trade-off between sonar pulse emission and stereo-olfaction in those species. To our knowledge, this is the first study to evaluate the evolutionary interactions between olfaction and echolocation in shaping the external morphology of a facial feature using modern phylogenetic comparative methods. Future work pairing olfactory morphology with tracking behavior will provide more insight into how animals such as bats integrate olfactory information while foraging.

Laminar Organization of FM Direction Selectivity in the Primary Auditory Cortex of the Free-Tailed Bat.

We studied the columnar and layer-specific response properties of neurons in the primary auditory cortex (A1) of six (four females, two males) anesthetized free-tailed bats, Tadaridabrasiliensis, in response to pure tones and down and upward frequency modulated (FM; 50 kHz bandwidth) sweeps. In addition, we calculated current source density (CSD) to test whether lateral intracortical projections facilitate neuronal activation in response to FM echoes containing spectrally distant frequencies from the excitatory frequency response area (FRA). Auditory responses to a set of stimuli changing in frequency and level were recorded along 64 penetrations in the left A1 of six free-tailed bats. FRA shapes were consistent across the cortical depth within a column and there were no obvious differences in tuning properties. Generally, response latencies were shorter (<10 ms) for cortical depths between 500 and 600 µm, which might correspond to thalamocortical input layers IIIb-IV. Most units showed a stronger response to downward FM sweeps, and direction selectivity did not vary across cortical depth. CSD profiles calculated in response to the CF showed a current sink located at depths between 500 and 600 µm. Frequencies lower than the frequency range eliciting a spike response failed to evoke any visible current sink. Frequencies higher than the frequency range producing a spike response evoked layer IV sinks at longer latencies that increased with spectral distance. These data support the hypothesis that a progressive downward relay of spectral information spreads along the tonotopic axis of A1 via lateral connections, contributing to the neural processing of FM down sweeps used in biosonar.

Forward masking enhances the auditory brainstem response in the free-tailed bat, Tadarida brasiliensis, during a critical time window for sonar reception.

Forward masking is a widespread auditory phenomenon in which the response to one sound transiently reduces the response to a succeeding sound. This study used auditory brainstem responses to measure temporal masking effects in the free-tailed bat, Tadarida brasiliensis. A digital subtraction protocol was used to isolate responses to the second of a pair of pulses varying in interval, revealing a suppression phase lasting <4 ms followed by an enhancement phase lasting 4-15 ms during which the ABR waveform was amplified up to 100%. The results suggest echolocating bats possess adaptations for enhancing sonar receiver gain shortly after pulse emission.

Suppression of emission rates improves sonar performance by flying bats.

Echolocating bats face the challenge of actively sensing their environment through their own emissions, while also hearing calls and echoes of nearby conspecifics. How bats mitigate interference is a long-standing question that has both ecological and technological implications, as biosonar systems continue to outperform man-made sonar systems in noisy, cluttered environments. We recently showed that perched bats decreased calling rates in groups, displaying a behavioral strategy resembling the back-off algorithms used in artificial communication networks to optimize information throughput at the group level. We tested whether free-tailed bats (Tadarida brasiliensis) would employ such a coordinated strategy while performing challenging flight maneuvers, and report here that bats navigating obstacles lowered emission rates when hearing artificial playback of another bat's calls. We measured the impact of acoustic interference on navigation performance and show that the calculated reductions in interference rates are sufficient to reduce interference and improve obstacle avoidance. When bats flew in pairs, each bat responded to the presence of the other as an obstacle by increasing emissions, but hearing the sonar emissions of the nearby bat partially suppressed this response. This behavior supports social cohesion by providing a key mechanism for minimizing mutual interference.

Infectivity of attenuated poxvirus vaccine vectors and immunogenicity of a raccoonpox vectored rabies vaccine in the Brazilian Free-tailed bat (Tadarida brasiliensis).

Bats (Order Chiroptera) are an abundant group of mammals with tremendous ecological value as insectivores and plant dispersers, but their role as reservoirs of zoonotic diseases has received more attention in the last decade. With the goal of managing disease in free-ranging bats, we tested modified vaccinia Ankara (MVA) and raccoon poxvirus (RCN) as potential vaccine vectors in the Brazilian Free-tailed bat (Tadarida brasiliensis), using biophotonic in vivo imaging and immunogenicity studies. Animals were administered recombinant poxviral vectors expressing the luciferase gene (MVA-luc, RCN-luc) through oronasal (ON) or intramuscular (IM) routes and subsequently monitored for bioluminescent signal indicative of viral infection. No clinical illness was noted after exposure to any of the vectors, and limited luciferase expression was observed. Higher and longer levels of expression were observed with the RCN-luc construct. When given IM, luciferase expression was limited to the site of injection, while ON exposure led to initial expression in the oral cavity, often followed by secondary replication at another location, likely the gastric mucosa or gastric associated lymphatic tissue. Viral DNA was detected in oral swabs up to 7 and 9 days post infection (dpi) for MVA and RCN, respectively. While no live virus was detected in oral swabs from MVA-infected bats, titers up to 3.88 x 104 PFU/ml were recovered from oral swabs of RCN-infected bats. Viral DNA was also detected in fecal samples from two bats inoculated IM with RCN, but no live virus was recovered. Finally, we examined the immunogenicity of a RCN based rabies vaccine (RCN-G) following ON administration. Significant rabies neutralizing antibody titers were detected in the serum of immunized bats using the rapid fluorescence focus inhibition test (RFFIT). These studies highlight the safety and immunogenicity of attenuated poxviruses and their potential use as vaccine vectors in bats.

The origins and diversity of bat songs.

Singing plays an important role in the social lives of several disparate bat species, but just how significant the behavior may be among bats generally is unknown. Recent discoveries suggest singing by bats might be surprisingly more diverse and widespread than anticipated, but if true then two questions must be addressed: firstly why has singing been so rarely documented among bats, and secondly do bats sing for the same reasons as songbirds? We address the first question by reviewing how sampling bias and technical constraints may have produced a myopic view of bat social communication. To address the second question, we review evidence from 50 years of batsong literature supporting the supposition that bat singing is linked to the same constellation of ecological variables that favored birdsong, including territoriality, polygyny, metabolic constraints, migratory behaviors and especially powered flight. We propose that bats sing like birds because they fly like birds; flight is energetically expensive and singing reduces time spent flying. Factoring in the singular importance of acoustic communication for echolocating bats, it seems likely that singing may prove to be relatively common among certain groups of bats once it becomes clear when and where to look for it.

Groups of bats improve sonar efficiency through mutual suppression of pulse emissions.

How bats adapt their sonar behavior to accommodate the noisiness of a crowded day roost is a mystery. Some bats change their pulse acoustics to enhance the distinction between theirs and another bat's echoes, but additional mechanisms are needed to explain the bat sonar system's exceptional resilience to jamming by conspecifics. Variable pulse repetition rate strategies offer one potential solution to this dynamic problem, but precisely how changes in pulse rate could improve sonar performance in social settings is unclear. Here we show that bats decrease their emission rates as population density increases, following a pattern that reflects a cumulative mutual suppression of each other's pulse emissions. Playback of artificially-generated echolocation pulses similarly slowed emission rates, demonstrating that suppression was mediated by hearing the pulses of other bats. Slower emission rates did not support an antiphonal emission strategy but did reduce the relative proportion of emitted pulses that overlapped with another bat's emissions, reducing the relative rate of mutual interference. The prevalence of acoustic interferences occurring amongst bats was empirically determined to be a linear function of population density and mean emission rates. Consequently as group size increased, small reductions in emission rates spread across the group partially mitigated the increase in interference rate. Drawing on lessons learned from communications networking theory we show how modest decreases in pulse emission rates can significantly increase the net information throughput of the shared acoustic space, thereby improving sonar efficiency for all individuals in a group. We propose that an automated acoustic suppression of pulse emissions triggered by bats hearing each other's emissions dynamically optimizes sonar efficiency for the entire group.

Regulation of bat echolocation pulse acoustics by striatal dopamine.

The ability to control the bandwidth, amplitude and duration of echolocation pulses is a crucial aspect of echolocation performance but few details are known about the neural mechanisms underlying the control of these voice parameters in any mammal. The basal ganglia (BG) are a suite of forebrain nuclei centrally involved in sensory-motor control and are characterized by their dependence on dopamine. We hypothesized that pharmacological manipulation of brain dopamine levels could reveal how BG circuits might influence the acoustic structure of bat echolocation pulses. A single intraperitoneal injection of a low dose (5 mg kg(-1)) of the neurotoxin 1-methyl-4-phenylpyridine (MPTP), which selectively targets dopamine-producing cells of the substantia nigra, produced a rapid degradation in pulse acoustic structure and eliminated the bat's ability to make compensatory changes in pulse amplitude in response to background noise, i.e. the Lombard response. However, high-performance liquid chromatography (HPLC) measurements of striatal dopamine concentrations revealed that the main effect of MPTP was a fourfold increase rather than the predicted decrease in striatal dopamine levels. After first using autoradiographic methods to confirm the presence and location of D(1)- and D(2)-type dopamine receptors in the bat striatum, systemic injections of receptor subtype-specific agonists showed that MPTP's effects on pulse acoustics were mimicked by a D(2)-type dopamine receptor agonist (Quinpirole) but not by a D(1)-type dopamine receptor agonist (SKF82958). The results suggest that BG circuits have the capacity to influence echolocation pulse acoustics, particularly via D(2)-type dopamine receptor-mediated pathways, and may therefore represent an important mechanism for vocal control in bats.

Mapping vocalization-related immediate early gene expression in echolocating bats.

Recent studies of spontaneously vocalizing primates, cetaceans, bats and rodents suggest these animals possess a limited but meaningful capacity to manipulate the timing and acoustic structure of their vocalizations, yet the neural substrate for even the simplest forms of vocal modulation in mammals remains unknown. Echolocating bats rapidly and routinely manipulate the acoustic structure of their outgoing vocalizations to improve echolocation efficiency, reflecting cognitive rather than limbic control of the vocal motor pathways. In this study, we used immunohistochemical localization of immediate early gene (c-fos) expression to map neural activity in the brains of spontaneously echolocating stationary Mexican free-tailed bats. Our results support the current model of vocal control obtained largely through microstimulation studies, but also provide evidence for the contributions of two novel regions, the dorsolateral caudate nucleus and mediodorsal thalamic nucleus, which together suggest a striatothalamic feedback loop may be involved in the control of echolocation pulse production. Additionally, we found evidence of a motivation pathway, including the lateral habenula, substantia nigra pars compacta, and raphe nuclei. These data provide novel insights into where and how mammalian vocalizations may be regulated by sensory, contextual and motivational cues.

A mechanism for antiphonal echolocation by Free-tailed bats.

Bats are highly social and spend much of their lives echolocating in the presence of other bats. To reduce the effects of acoustic interferences from other bats' echolocation calls, we hypothesized that bats might shift the timing of their pulse emissions to minimize temporal overlap with another bat's echolocation pulses. To test this hypothesis we investigated whether free-tailed bats (Tadarida brasiliensis) echolocating in the lab would shift the timing of their own pulse emissions in response to regularly repeating artificial acoustic stimuli. A robust phase-locked temporal pattern in pulse emissions was displayed by every bat tested which included an initial suppressive phase lasting more than 60 ms after stimulus onset, during which the probability of emitting pulses was reduced by more than fifty percent, followed by a compensatory rebound phase, the timing and amplitude of which were dependent on the temporal pattern of the stimulus. The responses were non-adapting and were largely insensitive to broad changes in the acoustic properties of the stimulus. Randomly occurring noise-bursts also suppressed calling for up to 60 ms, but the time-course of the compensatory rebound phase was more rapid than when the bats were responding to regularly repeating patterns of noise bursts. These findings provide the first quantitative description of how external stimuli may cause echolocating bats to alter the timing of subsequent pulse emissions.

Context-dependent effects of noise on echolocation pulse characteristics in free-tailed bats.

Background noise evokes a similar suite of adaptations in the acoustic structure of communication calls across a diverse range of vertebrates. Echolocating bats may have evolved specialized vocal strategies for echolocating in noise, but also seem to exhibit generic vertebrate responses such as the ubiquitous Lombard response. We wondered how bats balance generic and echolocation-specific vocal responses to noise. To address this question, we first characterized the vocal responses of flying free-tailed bats (Tadarida brasiliensis) to broadband noises varying in amplitude. Secondly, we measured the bats' responses to band-limited noises that varied in the extent of overlap with their echolocation pulse bandwidth. We hypothesized that the bats' generic responses to noise would be graded proportionally with noise amplitude, total bandwidth and frequency content, and consequently that more selective responses to band-limited noise such as the jamming avoidance response could be explained by a linear decomposition of the response to broadband noise. Instead, the results showed that both the nature and the magnitude of the vocal responses varied with the acoustic structure of the outgoing pulse as well as non-linearly with noise parameters. We conclude that free-tailed bats utilize separate generic and specialized vocal responses to noise in a context-dependent fashion.

Versatility and stereotypy of free-tailed bat songs.

In mammals, complex songs are uncommon and few studies have examined song composition or the order of elements in songs, particularly with respect to regional and individual variation. In this study we examine how syllables and phrases are ordered and combined, ie "syntax", of the song of Tadarida brasiliensis, the Brazilian free-tailed bat. Specifically, we test whether phrase and song composition differ among individuals and between two regions, we determine variability across renditions within individuals, and test whether phrases are randomly ordered and combined. We report three major findings. First, song phrases were highly stereotyped across two regions, so much so that some songs from the two colonies were almost indistinguishable. All males produced songs with the same four types of syllables and the same three types of phrases. Second, we found that although song construction was similar across regions, the number of syllables within phrases, and the number and order of phrases in songs varied greatly within and among individuals. Last, we determined that phrase order, although diverse, deviated from random models. We found broad scale phrase-order rules and certain higher order combinations that were highly preferred. We conclude that free-tailed bat songs are composed of highly stereotyped phrases hierarchically organized by a common set of syntactical rules. However, within global species-specific patterns, songs male free-tailed bats dynamically vary syllable number, phrase order, and phrase repetitions across song renditions.