Parallel evolution of KCNQ4 in echolocating bats

High-frequency hearing is required for echolocating bats to locate, range and identify objects, yet little is known about its molecular basis. The discovery of a high-frequency hearing-related gene, KCNQ4, provides an opportunity to address this question. Here, we obtain the coding regions of KCNQ4 from 15 species of bats, including echolocating bats that have higher frequency hearing and non-echolocating bats that have the same ability as most other species of mammals. The strongly supported protein-tree resolves a monophyletic group containing all bats with higher frequency hearing and this arrangement conflicts with the phylogeny of bats in which these species are paraphyletic. We identify five parallel evolved sites in echolocating bats belonging to both suborders. The evolutionary trajectories of the parallel sites suggest the independent gain of higher frequency hearing ability in echolocating bats. This study highlights the usefulness of convergent or parallel evolutionary studies for finding phenotype-related genes and contributing to the resolution of evolutionary problems.     

Dynamics of jamming avoidance in echolocating bats

Animals using active sensing systems such as echolocation or electrolocation may experience interference from the signals of neighbouring conspecifics, which can be offset by a jamming avoidance response (JAR). Here, we report JAR in one echolocating bat (Tadarida teniotis: Molossidae) but not in another (Taphozous perforatus: Emballonuridae) when both flew and foraged with conspecifics. In T. teniotis, JAR consisted of shifts in the dominant frequencies of echolocation calls, enhancing differences among individuals. Larger spectral overlap of signals elicited stronger JAR. Tadarida teniotis showed two types of JAR: (i) for distant conspecifics: a symmetric JAR, with lower- and higher-frequency bats shifting their frequencies downwards and upwards, respectively, on average by the same amount; and (ii) for closer conspecifics: an asymmetric JAR, with only the upper-frequency bat shifting its frequency upwards. In comparison, 'wave-type' weakly electric fishes also shift frequencies of discharges in a JAR, but unlike T. teniotis, the shifts are either symmetric in some species or asymmetric in others. We hypothesize that symmetric JAR in T. teniotis serves to avoid jamming and improve echolocation, whereas asymmetric JAR may aid communication by helping to identify and locate conspecifics, thus minimizing chances of mid-air collisions.     

Foraging ecology and audition in echolocating bats

The types of echolocation signal and the auditory capacities of echolocating bats are adapted to specific acoustical constraints of the foraging areas. Bats hunting insects above the canopy use low frequencies for echolocation; this is an adaptation to prey detection over long distances. Bats foraging close to and within foliage avoid masking of insect echoes by specializing on 'fluttering target' detection. 'Gleaning' bats are adapted to the auditory detection of very faint noises generated by ground-dwelling prey, and are capable of analysing fine changes in the echo spectrum, which may indicate a stationary prey changing its posture on a substrate. This review of recent research demonstrates that, in bats, foraging ecology and audition are intricately interrelated and interdependent.     

Neural coding of echo-envelope disparities in echolocating bats

The effective use of echolocation requires not only measuring the delay between the emitted call and returning echo to estimate the distance of an ensonified object. To locate an object in azimuth and elevation, the bat’s auditory system must analyze the returning echoes in terms of their binaural properties, i.e., the echoes’ interaural intensity and time differences (IIDs and ITDs). The effectiveness of IIDs for echolocation is undisputed, but when bats ensonify complex objects, the temporal structure of echoes may facilitate the analysis of the echo envelope in terms of envelope ITDs. Using extracellular recordings from the auditory midbrain of the bat, Phyllostomus discolor, we found a population of neurons that are sensitive to envelope ITDs of echoes of their sonar calls. Moreover, the envelope-ITD sensitivity improved with increasing temporal fluctuations in the echo envelopes, a sonar parameter related to the spatial statistics of complex natural reflectors like vegetation. The data show that in bats envelope ITDs may be used not only to locate external, prey-generated rustling sounds but also in the context of echolocation. Specifically, the temporal fluctuations in the echo envelope, which are created when the sonar emission is reflected from a complex natural target, support ITD-mediated echolocation.     

Object-oriented echo perception and cortical representation in echolocating bats

Echolocating bats can identify three-dimensional objects exclusively through the analysis of acoustic echoes of their ultrasonic emissions. However, objects of the same structure can differ in size, and the auditory system must achieve a size-invariant, normalized object representation for reliable object recognition. This study describes both the behavioral classification and the cortical neural representation of echoes of complex virtual objects that vary in object size. In a phantom-target playback experiment, it is shown that the bat Phyllostomus discolor spontaneously classifies most scaled versions of objects according to trained standards. This psychophysical performance is reflected in the electrophysiological responses of a population of cortical units that showed an object-size invariant response (14/109 units, 13%). These units respond preferentially to echoes from objects in which echo duration (encoding object depth) and echo amplitude (encoding object surface area) co-varies in a meaningful manner. These results indicate that at the level of the bat's auditory cortex, an object-oriented rather than a stimulus-parameter-oriented representation of echoes is achieved.     

Classification of insects by echolocating greater horseshoe bats

Summary Echolocating greater horseshoe bats (Rhinolophus ferrumequinum) detect insects by concentrating on the characteristic amplitude- and frequency modulation pattern fluttering insects impose on the returning echoes. This study shows that horseshoe bats can also further analyse insect echoes and thus recognize and categorize the kind of insect they are echolocating.Four greater horseshoe bats were trained in a twoalternative forced-choice procedure to choose the echo of one particular insect species turning its side towards the bat (Fig. 1). The bats were able to discriminate with over 90% correct choices between the reward-positive echo and the echoes of other insect species all fluttering with exactly the same wingbeat rate (Fig. 4).When the angular orientation of the reward-positive insect was changed (Fig. 2), the bats still preferred these unknown echoes over echoes from other insect species (Fig. 5) without any further training. Because the untrained bats did not show any prey preference, this indicates that the bats were able to perform an aspect-anglein-dependent classification of insects.Finally we tested what parameters in the echo were responsible for species recognition. It turned out that the bats especially used the small echo-modulations in between glints as a source of information (Fig. 7). Neither the amplitudenor the frequencymodulation of the echoes alone was sufficient for recognition of the insect species (Fig. 8). Bats performed a pattern recognition task based on complex computations of several acoustic parameters, an ability which might be termed cognitive.Abbreviations AM amplitude modulation - CF constant frequency - FM frequency modulation - S+ positive stimulus - S- negative stimulus     

Neural ensemble coding of target identity in echolocating bats

Most insectivorous bats use echolocation to determine the identity of flying insects. Among the many target features that are so extracted, the insect's wingbeat pattern and frequency appear to serve as useful cues for identification. Biosonar pulses impinging on the fluttering wings of an insect are returned as echoes whose amplitudes vary with time, thus providing a characteristic signature of the insect. It has been shown previously that neurons in the inferior colliculus, a midbrain auditory nucleus, of the little brown bat respond to sound stimuli that mimic echoes from fluttering targets. To examine the manner in which target identity is represented in the inferior colliculus, an ensemble coding analysis using a filter-based approach was undertaken. The analysis indicates that a discrete subset of neurons in the inferior colliculus, the onset units, are strongly tuned to wingbeat frequencies of targets that the bat hunts, and that ensemble response reaches a maximum at a distinct phase of the prey capture maneuver: the late approach stage. On the basis of the analysis it is hypothesized that inferior colliculus neurons may play an important role in target detection-identification processing. Although ensemble coding of temporally sequenced information has not been analyzed in the auditory system so far, this study indicates that this method of coding may provide the information necessary to detect and identify targets during prey capture. Received: 4 December 1995 / Accepted in revised form: 19 April 1996     

Experimental evidence for group hunting via eavesdropping in echolocating bats

Group foraging has been suggested as an important factor for the evolution of sociality. However, visual cues are predominantly used to gain information about group members'' foraging success in diurnally foraging animals such as birds, where group foraging has been studied most intensively. By contrast, nocturnal animals, such as bats, would have to rely on other cues or signals to coordinate foraging. We investigated the role of echolocation calls as inadvertently produced cues for social foraging in the insectivorous bat Noctilio albiventris. Females of this species live in small groups, forage over water bodies for swarming insects and have an extremely short daily activity period. We predicted and confirmed that (i) free-ranging bats are attracted by playbacks of echolocation calls produced during prey capture, and that (ii) bats of the same social unit forage together to benefit from passive information transfer via the change in group members'' echolocation calls upon finding prey. Network analysis of high-resolution automated radio telemetry confirmed that group members flew within the predicted maximum hearing distance 94±6 per cent of the time. Thus, echolocation calls also serve as intraspecific communication cues. Sociality appears to allow for more effective group foraging strategies via eavesdropping on acoustical cues of group members in nocturnal mammals.     

The voltage-gated potassium channel subfamily KQT member 4 (KCNQ4) displays parallel evolution in echolocating bats

Bats are the only mammals that use highly developed laryngeal echolocation, a sensory mechanism based on the ability to emit laryngeal sounds and interpret the returning echoes to identify objects. Although this capability allows bats to orientate and hunt in complete darkness, endowing them with great survival advantages, the genetic bases underlying the evolution of bat echolocation are still largely unknown. Echolocation requires high-frequency hearing that in mammals is largely dependent on somatic electromotility of outer hair cells. Then, understanding the molecular evolution of outer hair cell genes might help to unravel the evolutionary history of echolocation. In this work, we analyzed the molecular evolution of two key outer hair cell genes: the voltage-gated potassium channel gene KCNQ4 and CHRNA10, the gene encoding the α10 nicotinic acetylcholine receptor subunit. We reconstructed the phylogeny of bats based on KCNQ4 and CHRNA10 protein and nucleotide sequences. A phylogenetic tree built using KCNQ4 amino acid sequences showed that two paraphyletic clades of laryngeal echolocating bats grouped together, with eight shared substitutions among particular lineages. In addition, our analyses indicated that two of these parallel substitutions, M388I and P406S, were probably fixed under positive selection and could have had a strong functional impact on KCNQ4. Moreover, our results indicated that KCNQ4 evolved under positive selection in the ancestral lineage leading to mammals, suggesting that this gene might have been important for the evolution of mammalian hearing. On the other hand, we found that CHRNA10, a gene that evolved adaptively in the mammalian lineage, was under strong purifying selection in bats. Thus, the CHRNA10 amino acid tree did not show echolocating bat monophyly and reproduced the bat species tree. These results suggest that only a subset of hearing genes could underlie the evolution of echolocation. The present work continues to delineate the genetic bases of echolocation and ultrasonic hearing in bats.     

Does body mass restrict call peak frequency in echolocating bats?

1. Echolocation is the ability of some animals to orient themselves through sound emission and interpretation of the echoes. This is bats’ main sense for orientation and recognising biotopes that provide food, water, and roosts. It is widely accepted that echolocation call frequency is related to body mass, and this relationship has been described as the ‘allometric hypothesis’, which proposes a negative correlation between these variables.
2. There is evidence that, in many cases, the allometric hypothesis does not apply. Additionally, studies supporting this hypothesis were done at the family level, resulting in a broad range of correlation values with r ranging from −0.36 to −0.76, and only insectivorous bats were included. Due to the notable exceptions and the lack of a quantitative synthesis of this hypothesis including all echolocating bats, we evaluated the allometric hypothesis of echolocation calls for this group.
3. Using a meta-analysis and phylogenetic generalised least-squares techniques, we evaluated the relationship between echolocation call peak frequency and the body mass of bats.
4. We found a negative relationship between body mass and echolocation call peak frequency for the 85 bat species that were included in our analysis (r = −0.3, p = 0.005). The relationship was consistent when we analysed the data at the insectivorous guild level, and in bats belonging to the families Vespertilionidae, Rhinolophidae, Emballonuridae, and the genus Myotis. However, the wide range of r values suggests that the strength of the relationship between peak frequency and body mass varies within the order Chiroptera.
5. Our results support the allometric hypothesis of sound production in echolocating bats. However, the low coefficient we found suggests that factors other than body mass may influence the peak frequency of echolocation calls produced by bats.

     

Variability of the approach phase of landing echolocating Greater Mouse-eared bats

The approach phase of landing vespertilionid bats ends with a group of calls, which either consists of buzz I alone or buzz I and buzz II. To understand the possible role of buzz II, we trained Myotis myotis to land on a vertical grid, and compared the flight and echolocation behavior during approach in trials with and without buzz II. During the approach, we did not find any differences in the echolocation behavior until the end of buzz I which indicated whether buzz II was emitted or not. However, bats flying from the periphery of the flight channel, such that they had to make a small turn at the very last moment, finished the sequence with a buzz II. Bats flying on a rather stereotyped trajectory near the center of the flight channel without last instant corrections emitted buzz I alone. Our results indicate that buzz II occurred only on trajectories that implied a higher risk to fail at landing. The information delivered by buzz II reaches the bat too late to be used for landing. Therefore, we hypothesize that buzz II may help the bats to evaluate unsuccessful attempts and to eventually react adequately.     

Peripheral control of acoustic signals in the auditory system of echolocating bats.

Many species of echolocating bats emit intense orientation sounds. If such intense sounds directly stimulated their ears, detection of faint echoes would be impaired. Therefore, possible mechanisms for the attenuation of self-stimulation were studied with Myotis lucifugus. The acoustic middle-ear-muscle reflex could perfectly and transiently regulate the amplitude of an incoming signal only at its beginning. However, its shortest latency in terms of electromyograms and of the attenuation of the cochlear microphonic was 3-4 and 4-8 msec, respectively, so that these muscles failed to attenuate orientation signals by the reflex. The muscles, however, received a message from the vocalization system when the bat vocalized, and contracted synchronously with vocalization. The duration of the contraction-relaxation was so short that the self-stimulation was attenuated, but the echoes were not. The tetanus-fusion frequency of tha stapedium muscle ranged between 260 and 320/sec. Unlike the efferent fibres in the lateral-line and vestibular systems, the olivo-cochlear bundle showed no sign of attenuation of self-stimulation.