J Comp Physiol A (2014) 200:799–809 DOI 10.1007/s00359-014-0921-3

Original Paper

Prey pursuit strategy of Japanese horseshoe bats during an in‑flight target‑selection task Yuki Kinoshita · Daiki Ogata · Yoshiaki Watanabe · Hiroshi Riquimaroux · Tetsuo Ohta · Shizuko Hiryu 

Received: 16 December 2013 / Revised: 23 April 2014 / Accepted: 3 June 2014 / Published online: 24 June 2014 © Springer-Verlag Berlin Heidelberg 2014

Abstract  The prey pursuit behavior of Japanese horseshoe bats (Rhinolophus ferrumequinum nippon) was investigated by tasking bats during flight with choosing between two tethered fluttering moths. Echolocation pulses were recorded using a telemetry microphone mounted on the bat combined with a 17-channel horizontal microphone array to measure pulse directions. Flight paths of the bat and moths were monitored using two high-speed video cameras. Acoustical measurements of returning echoes from fluttering moths were first collected using an ultrasonic loudspeaker, turning the head direction of the moth relative to the loudspeaker from 0° (front) to 180° (back) in the horizontal plane. The amount of acoustical glints caused by moth fluttering varied with the sound direction, reaching a maximum at 70°–100° in the horizontal plane. In the flight experiment, moths chosen by the bat fluttered within or moved across these angles relative to the bat’s pulse direction, which would cause maximum dynamic changes in the frequency and amplitude of acoustical glints during flight. These results suggest that echoes with acoustical glints containing the strongest frequency and amplitude modulations appear to attract bats for prey selection. Keywords  Acoustical glints · Fluttering moths · Pulse direction

Y. Kinoshita · D. Ogata · T. Ohta  Faculty of Life and Medical Sciences, Doshisha University, Kyotanabe 610‑0321, Japan Y. Watanabe · H. Riquimaroux · S. Hiryu (*)  Faculty of Life and Medical Sciences, Neurosensing and Bionavigation Research Center, Doshisha University, Kyotanabe 610‑0321, Japan e-mail: [email protected]

Abbreviations CF Constant frequency CF2 Constant frequency component with a second harmonic FM Frequency modulated iFM Initial frequency modulated PI Pulse interval TF Terminal frequency tFM Terminal frequency modulated

Introduction Bats in the Rhinolophidae, Hipposideridae, and Mormoopidae families emit pulses consisting of a long constant frequency (CF) component [e.g., R. ferrumequinum: 50– 60 ms (Tian and Schnitzler 1997), Hipposideros terasensis: 5–10 ms (Hiryu et al. 2005)] followed, and sometimes also preceded, by short (1–2 ms) initial upward and terminal downward frequency modulated (FM) components. These bat species are called CF-FM bats, and some previous studies have suggested that CF-FM bats use the acoustical information obtained from fluttering insects that is encoded in both frequency and amplitude modulations (acoustical glints) of the echoes for detecting prey insects (Schnitzler and Flieger 1983; Schnitzler and Ostwald 1983; Link et al. 1986; von der Emde and Menne 1989; von der Emde and Schnitzler 1990; Schnitzler and Kalko 2001; Schnitzler and Denzinger 2011). Acoustical measurements demonstrated that echoes from fluttering insects are both species specific and orientation specific, which may enable bats even to identify or classify insects (Schuller 1984; Kober and Schnitzler 1990; von der Emde and Schnitzler 1990). In some CF-FM bat species, the beginning of the approach phase for capturing a fluttering moth has been reported to

13

800

be characterized by an increase in the duration of the CF portion of the pulse [Pteronotus parnellii (Novick 1963); R. ferrumequinum (Vogler and Neuweiler 1983; Schnitzler et al. 1985; Neuweiler et al. 1987; Mantani et al. 2012)]. Given that the CF component of the signal is better suited than FM portions for detecting acoustical glints, the bat can enhance detection of acoustical glints by extending the duration of the CF component. These facts also suggest that the acoustical glints provide the bats with detailed information on potential target insects (Schnitzler and Denzinger 2011) as well as important cues for target selection by echolocation. In this study, echoes from fluttering moths (Goniocraspidum pryeri) were first investigated using artificial sounds to estimate how the acoustical features of the echoes coming back from fluttering moths changed with the incident angle of echolocation pulses emitted by the bats during prey capture flight. We then observed the prey pursuit behavior of Japanese horseshoe bats R. ferrumequinum nippon by tasking the bats with choosing between two tethered fluttering moths during flight in a flight chamber. Echolocation sounds were recorded using a telemetry microphone (Telemike) that was mounted on the bats, combined with a microphone array system that was arranged in the horizontal plane in the chamber because more significant behavioral responses by bats were expected in the horizontal plane than in the vertical plane. We investigated how the bats changed the direction and beam width of their emitted pulses during the target selection task, considering changes in the acoustical glints of echoes coming back from the moths.

Materials and methods Subjects Four adult horseshoe bats (R. ferrumequinum nippon, body length 6.0–8.0 cm, body mass 20–30 g) were used (two females and two males). The bats were captured under license and in compliance with current Japanese laws from a natural cave in Hyogo prefecture in Japan. The animals were housed in a temperature- and humidity-controlled colony room [4 (L) × 3 (W) × 2 m (H)] at Doshisha University in Kyoto, Japan. In the colony room, the bats were allowed to fly freely and had ad libitum access to water and food (mealworms). The day and night cycle of the room was set to 12 h dark:12 h light. On the day before the experiment, the amount of food provided to the bats was reduced by 50 % to ensure motivation for capturing moths during the experiment. Figure 1a shows representative echolocation pulses of R. ferrumequinum nippon. Rhinolophus ferrumequinum nippon uses compound echolocation signals,

13

J Comp Physiol A (2014) 200:799–809

each consisting of a CF component with a second harmonic (CF2), strongest around 68–70 kHz, plus an accompanying initial short upward FM sweep (2–8 kHz, ending at 68–70 kHz for the second harmonic: iFM2) and a terminal short downward FM sweep (beginning at 68–70 kHz and extending 8–12 kHz lower for the second harmonic: tFM2) (Hiryu et al. 2008). Noctuid moths (Goniocraspidum pryeri, body length approximately 20 mm, wingspan 40–44 mm) were used as target prey. The moths were caught as adults in a cave in Osaka Prefecture, Japan. The moths were stored in a plastic rearing cage [50 (L) × 35 (W) × 30 cm (H)] in the bat room under the same light cycle; 12 h dark and 12 h light. Goniocraspidum pryeri are widely distributed in Japan, and they diapause from summer to the next spring in caves that are used by R. ferrumequinum nippon as day roosts (Sano 2006). Goniocraspidum pryeri have ears, but their auditory characteristics with respect to the echolocation sounds of bats have not been investigated. In our previous results, however, G. pryeri sometimes showed distinct evasive flight patterns, such as a sudden changes in flight direction with increased flight velocity when a bat approached within approximately 1 m of the moth (Matsuta et al. 2013). This suggests that G. pryeri may have a hearing frequency range that enables them to respond to bat ultrasound. Acoustical measurements of echoes from fluttering moth To record the echoes returning from the moths, the moths were suspended 0.5 m in front of a loudspeaker (Pioneer Corporation, PT-R7 III, Kanagawa, Japan), which was positioned in the center of the flight chamber [8 (L)  × 3 (W) × 2 (H)]. The recording procedure was the same as that used in a previous study (Mantani et al. 2012). The ventral part of the moth body was carefully impaled on the tip of a thin steel wire (0.5-mm diameter, 10-cm length) which did not interfere with fluttering, so that the moth remained in place but kept fluttering. The opposite end of the wire was attached to a fixed pole attached to the center of a turntable, so that the fluttering moth could be turned on a horizontal plane while remaining suspended in front of the center of the loudspeaker (top panel in Fig. 2a). The walls of the chamber were coated with sound-absorbing materials to adequately reduce surrounding echoes. An ultrasonic microphone (Titley Electronics, Ltd., ANABAT II, Ballina, Australia) was mounted on the top of the loudspeaker in the same orientation, i.e., pointing toward the moths. CF sounds (100-ms duration, 67 kHz), which were generated using a function generator (Agilent Technologies, 33220A, Tokyo, Japan), were emitted from the loudspeaker at 114 dB re 20 μPa (peak-to-peak) measured 0.5 m in front of the loudspeaker. Echoes returning from the moths were picked up by the microphone and then band-pass filtered

J Comp Physiol A (2014) 200:799–809 Fig.  1  a Representative echolocation sounds of R. ferrumequinum nippon during flight, recorded with the Telemike. b Schematic diagram of the microphone array system in a flight chamber for measuring the direction and beam width of pulses emitted by the bats during moth capture flight. Seventeen microphones were arranged in a U-shape on the X–Y plane 1.2 m from the floor. Microphones were spaced at 0.5-m intervals on the x-axis and 0.8-m intervals on the y-axis. c Top view of the horizontal pulse direction and beam width, which were determined based on sounds received by the microphones. The pulse direction was calculated by summing the vectors, which were proportional to the sound pressure level of each microphone. The beam width was calculated by the −6 dB off-axis angle from the pulse direction on the pulse directivity pattern. d Definitions of the direction components; the x-axis and y-axis were defined as 0° and 90° in the horizontal plane, respectively. The target direction was determined using the 3D coordinates of the bat and the moth. The flight direction of the bat was calculated as the time derivative of the coordinates of the bat’s flight trajectory

801

a

b

Bat

C

from 20 to 150 kHz (NF Corporation, Model 3625, Yokohama, Japan), digitized by a DAT recorder (Sony, Model SIR-1000W, Tokyo, Japan, 16-bit, 384 kHz), and stored as files on the hard disk of a personal computer. The returning echoes were picked up every 10° over the range from 0° to 180° in the horizontal plane while the moths were rotated (top panel in Fig. 2a). In this case, 0° was defined as the angle whereby the moths’ heads pointed directly toward the loudspeaker. Three moths were used in this study. Echoes returning from the fluttering G. pryeri included several periodic peaks in amplitude (amplitude glints;

d

middle panel Fig. 2a) and a spectrogram (spectral glints: bottom panel Fig. 2a), both caused by the fluttering of the moth. The peak-to-peak voltage of each amplitude glint (echo amplitude) was analyzed from the amplitude patterns of recorded echoes using custom Matlab routines on a personal computer. Each echo usually included 3–5 glints, and for further analysis, the glint with the largest echo amplitude was chosen from each returning echo. A total of 15 amplitude glints from 3 moths (5 glints from each individual) were averaged for each incident angle of the sound. Additionally, the positive and negative Doppler shifts of each spectral glint were analyzed. The glint with the

13

802

J Comp Physiol A (2014) 200:799–809

In this study, the detailed acoustic measurements of echoes from fluttering moths were only conducted in the horizontal plane because changes in the extent of acoustical glints in the vertical plane were small compared to changes in the horizontal plane during preliminary measurements. Therefore, during the subsequent behavioral experiments of moth-capture flight, we assumed that more significant behavioral changes by bats would be observed when responding to horizontal movements of fluttering moths than changes in the vertical plane.

a

Behavior experiments during moth‑capture flights b

c

Fig. 2  Echo recordings from fluttering G. pryeri. a Amplitude pattern (middle panel) and spectrogram (bottom panel) of the echoes returning from the fluttering moth using the artificial CF sound (100ms duration, 67 kHz) emitted from an ultrasound loudspeaker located 0.5 m in front of the moth. The moth was oriented at 0°, 100°, or 180° relative to the sound source (top panel). Positive and negative Doppler shifts were repeated in synchronization with the wing beat cycle of the moth (marked with arrows, 26–27 ms). b Changes in the amount and amplitude of glints from the echoes of the fluttering moth as a function of the incident angle of the sound. c Changes in positive and negative Doppler shifts as a function of the incident angle of sounds. Data for b and c were taken from three G. pryeri (five glints from each individual). The angle at which the moth’s head was directed toward the loudspeaker was defined as 0°

maximum Doppler shift was chosen from each echo, and then the 15 spectral glints (5 glints from each individual) were averaged.

13

Video recordings Figure 1b shows a schematic diagram of the measurement system employed in this study. The experiments were conducted in a flight chamber under long-wavelength lighting with filters (>650 nm) to avoid any visual effects. The flight chamber was constructed of steel plates to minimize interference from external electromagnetic noises and waves used by commercial FM radio stations. A thin polyester string (FUJIX, Shappespun, Kyoto, Japan, the diameter 0.1–0.2 mm, the length 1 m) was carefully attached to the dorsal portion of the moth’s body using a drop of beeswax (Tree of Life Co., Ltd., Tokyo, Japan), tethering the moth from the ceiling while allowing it to move freely within the length of the string during the experiment. Thus, the moth could move freely within 1 m of the tether during recordings. We observed the in-flight prey pursuit behavior of bats that were tasked with making a choice between two tethered fluttering moths (Fig. 1b). Two moths were placed 2 m from the front wall of the chamber, corresponding to 3–4 m from the bat’s starting point. Moths were positioned at 0.5 m to the left and right of the centerline of the chamber, resulting in a space of 1 m between individuals. The experimenter released an individual bat from one end of the flight chamber. Flight behaviors of the bats and moths were recorded using two digital high-speed video cameras (IDT Japan, Inc., MotionPro X3, Tokyo, Japan) located behind the bat at the left and right corners of the flight chamber so as not to interfere with the bat’s flight path. The video cameras recorded 125 frames/s, and three-dimensional (3D) coordinates of the bat and moth flight paths were reconstructed from the video images using motion-analysis software (Ditect Corporation, DIPPMotionPro ver.2.2.1.0, Tokyo, Japan). Prior to recording the bat flights, a 3D reference frame with known coordinates was positioned in the center of the flight chamber and briefly recorded by the two video cameras. The analysis software then calibrated the 3D flight path reconstruction system using the cameras’ stereo view

J Comp Physiol A (2014) 200:799–809

of the reference frame. Based on a technique of direct linear transformation from the reference frame’s coordinates, successive positions of the flying bat and moths as well as the locations of other objects were reconstructed from video-scene coordinates measured from a pair of 2D video images. Using the 3D coordinate data, the flight trajectories of the bat and the moths were determined in conjunction with the acoustic characteristics of the bat’s echolocation sounds. Telemike recordings Echolocation sounds emitted by the flying bat were recorded using a custom-made telemetry microphone (Telemike) mounted on the bat. The recording procedure was the same as that used in a previous study (Hiryu et al. 2008). The Telemike consisted of a 1/8-in. omni-directional condenser microphone (Knowles, Model FG-3329, Itasca, IL, USA), a miniature custom-designed FM transmitter unit, a 1.5 V hearing-aid battery (Sony, Type SR521SW, Tokyo, Japan), and a transmitting antenna. The total weight of the Telemike was approximately 0.6 g. The Telemike was attached to the back of the bat using a piece of double-sided adhesive tape. The microphone pointed forward and was positioned approximately 1 cm above the noseleaf, centered between the right and left pinnae. The Telemike’s transmitter produced radio signals that were received by an FM antenna (RadioShack Corporation, Model 15-1859, TX, USA) that was tethered to the ceiling of the flight chamber. The received signals were demodulated to recover the bat’s ultrasonic broadcasts using a custom-made FM receiver. The total frequency response of the Telemike system was flat within ±4 dB between 20 and 100 kHz. The signals from the receiver were then band-pass filtered from 20 to 150 kHz (NF Corporation, Model 3625, Yokohama, Japan) and digitized by a DAT recorder (SONY, Model SIR-1000W, Tokyo, Japan, 16 bit, 384 kHz) with the control signal that triggered video recordings. All digitized data were stored as files on the hard disk of a personal computer so that the sound recordings could be synchronized with flight coordinates. Microphone array recordings To measure the horizontal direction and beam width of pulses emitted by the bat during moth-capture, a U-shaped 17-channel (ch) horizontal microphone array was arranged along the x–y plane of the chamber 1.2 m above the floor (blue frame in Fig. 1b). The recording procedure was the same as that used in a previous study (Matsuta et al. 2013). Microphones in the arrays were spaced at 0.8-m intervals on the x-axis and 0.5-m intervals on the y-axis of the flight chamber. The microphone circuit board consisted of a

803

1/8-in. omni-directional condenser microphone (Knowles, Model FG-3329, Itasca, IL, USA), a custom-designed differential amplifier circuit (+46 dB) and a custom-designed band-pass filter (10–250 kHz). A urethane acoustic absorption material (20 cm × 20 cm) was attached to the rear of each condenser microphone to reduce echoes from the walls of the chamber. All of the signals recorded by the microphone array system were digitized using a high-speed data acquisition card (National Instruments, Model NI PXI6250, Tokyo, Japan, 16 bit, fs  = 200 kHz). The digitized signals of all channels were stored as files on the hard disk of a personal computer using a custom program in LabVIEW (NI, Model NI LabVIEW 8.0, Tokyo, Japan) with the control signal of the video recording so that the microphone array data were synchronized with both flight coordinates and the sound recordings obtained by the Telemike. Sound analysis The acoustic characteristics of the flying bat’s broadcast sounds (excluding the direction and beam width of the sound) were analyzed from spectrograms made through the Telemike recordings using custom Matlab routines on a personal computer. Each pulse was extracted from the recording and then the second harmonic component of the pulse was analyzed for pulse duration and pulse interval (PI: the interval between the onsets of successive calls). In this study, PI was defined as the time from the beginning of one pulse to the beginning of the subsequent pulse. The pulse duration was determined from the spectrogram at −25 dB relative to the peak intensity of each pulse. To analyze the direction and beam width of pulses emitted by the bat during moth-capture flight, tFM2 components were used because the CF2 components of the pulses recorded by the microphone array were temporally overlapped with echoes from the surrounding walls of the chamber (Matsuta et al. 2013). The energy maximum in the tFM2 component spectrogram was measured to quantify changes in the sound pressure levels in order to reconstruct the directivity pattern of the emitted pulse. The sound pressure levels of the pulses were corrected for sound propagation losses between the bat and each microphone in the air and for sensitivity differences among microphones in the array. For propagation loss, a spreading loss was calculated based on the distance between the bat and each microphone. The time at which each bat emitted a pulse was determined from the Telemike recording. Sound absorption was calculated based on a measured absorption coefficient at 65 kHz (α  = 2.4 dB/m), which corresponded to the average frequency at the peak energy in the tFM2 component of the pulse emitted by R. ferrumequinum nippon. The sensitivity of each microphone in the array system was measured by presenting a tone burst at 65 kHz using

13

804

an ultrasonic loudspeaker (Pioneer Corporation, PT-R7 III, Kanagawa, Japan) to compensate for sensitivity differences among microphones in the array. The corrected sound pressure levels of each microphone in the array were converted to vectors (red arrows in Fig.  1c), and the horizontal pulse direction was computed by summing these vectors (blue arrow for horizontal pulse direction, Fig. 1c). The beam width was defined by −6 dB off-axis angles from the direction of the emitted pulse on the pulse directivity pattern (Fig. 1c, green double-headed arrow). Figure 1d shows the definitions of the angular components in this study. We defined the directions of the x-axis and y-axis as 0° and 90°, respectively, in the horizontal plane. The direction of the moth from the bat (the target direction) was determined using the 3D coordinates of each animal. The flight direction of the bat was calculated as the time derivative of the coordinates of the bat’s flight trajectory. The angular difference between the pulse and target directions was defined as misalignment Δϕ (Fig. 1d). Prior to the experiments, the measurement errors in pulse direction and beam width created by our microphone array system were investigated when the ultrasound tone burst (65 kHz with 3 ms duration; 107 dB SPL at 1 m from the loudspeaker) was presented from the loudspeaker (Pioneer Corporation, PT-R7 III, Kanagawa, Japan) that was set up in the chamber. Horizontal pulse direction and beam width were measured while moving the distance of the loudspeaker from the front wall between 0.5 and 6 m. The measurement errors in pulse direction and beam width were less than approximately ±3° and ±5°, respectively, in the horizontal plane and did not change with distance of 1–6 m from the front wall (Matsuta et al. 2013). Hence, we placed the target moths >2 m from the front wall to maintain measurement accuracy. For statistical comparisons, either Student’s t tests, one-way factorial analysis of variance (Kruskal–Wallis tests) were used, as appropriate, to test for significant differences in call parameters between data sets.

Results

J Comp Physiol A (2014) 200:799–809

G. pryeri were amplitude and frequency modulated and showed periodic peaks every 26–27 ms corresponding to the fluttering period of G. pryeri (Mantani et al. 2012). Figure 2b shows changes in the peak-to-peak voltage of amplitude glints of the echoes from a fluttering moth (echo amplitude) as a function of the incident angles of sound. The mean peak-to-peak amplitude ranged from approximately 0.25 to 0.4 V between 0° and 60°, after which the amplitude glints reached a maximum at 70°–100°. However, once the incident angle increased to 110°, the echo amplitude suddenly decreased to 0.15–0.25 V, resulting in an 11–6 dB decrease for angles of 110°–180° compared with those of 70°–100° (double-headed arrow). When the incident angles were divided into three equal ranges (0°–60°, 60°–120° and 120°–180°), there were significant differences in the peak-to-peak voltages of the amplitude glints (Kruskal–Wallis test; P 60 ms), the bat changed its target from moth 2 to moth 1 (−1.5 s before the capture, Fig. 3d), and then began its approach phase for capturing moth 1 with decreases in the pulse duration and PI (Fig. 3c). During the approach phase, the horizontal angular difference between the pulse and target (moth 1) directions (misalignment Δϕ) was approximately 10°–15° (Fig. 3d). The beam widths of the pulses emitted during the approach phase (mean ± SD: ±21.7°  ± 2.5°) covered the misalignment, indicating that the bat could retain the moving moth 1 within its echolocation window during the approach. In another example of moth-capture flight by a different bat, shown in Fig. 4, the bat initially directed its pulses toward both moth 1 and moth 2, before finally turning left to capture moth 1. The consecutive emissions of long-duration pulses were also observed when the distances to the moths were approximately 4 m (−1.4 s before capture, Fig. 4c). During this period, the bat tracked both of the moving two

805

a

b

c

d

Fig. 3  Representative horizontal flight paths of the bat and moths during moth capture flight (Bat A). a Horizontal flight trajectories of the bat (red line) and two moths (orange and green plots). Blue lines and asterisks indicate the direction of each pulse and long-duration pulse, respectively. b Spectrogram of the echolocation pulses recorded by the Telemike mounded on the bat during the flight. Asterisks indicate long-duration pulses. c Change in the duration and pulse interval (PI) of the pulses during this flight. d Changes in flight, target, and pulse directions in the horizontal plane as a function of flight time. The length of the vertical blue line corresponds to the beam width (−6 dB) of each pulse

moths with accuracy within 5° (Fig. 4d). After the consecutive emissions of the long-duration pulses (0.6 s before capture), the bat changed its target from moth 2 to moth 1. Estimated acoustical glints from fluttering moths Figures 3 and 4 show examples of bats shifting the directional aim of the sonar beam between moth 1 and moth 2 before beginning their final approaches, i.e., some pulses

13

806

J Comp Physiol A (2014) 200:799–809

a

long-duration pulses 180 150

[deg.]

120

moth

b

90

Moth1

moth1

60

Moth 2

moth2

30

moth1

0

c

Pulse direction

-2.0

-1.5

-1.0

moth2

-0.5

0.0

Time [s]

Fig.  5  a Changes in θmoth of moth 1 (orange) and moth 2 (green) as a function of flight time in the flight shown in Fig. 3. Black arrows indicate the moth targeted by the bat. Black bar indicate the consecutive emissions of long-duration pulses. The definition of θmoth is shown in the inset. In this case, the bat aimed its pulse toward moth 1

60

d

Targeted moth

Number of pulses [%]

50

Not-targeted moth

40 30 20 10

Fig. 4  Representative horizontal flight paths of the bat and moths during moth capture flight (Bat B). a Horizontal flight trajectories of the bat (red line) and two moths (orange and green plots). Blue lines and asterisks indicate the direction of each pulse and long-duration pulse, respectively. b Spectrogram of the echolocation pulses recorded by the Telemike mounted on the bat during the flight. Asterisks indicate the long-duration pulses. c Change in the duration and pulse interval (PI) of the pulses during this flight. d Changes in the flight, target, and pulse directions on the horizontal plane as a function of flight time. The length of vertical blue line corresponds to the beam width (−6 dB) of each pulse

were emitted toward moth 2 before the bats locked onto their final targets (moth 1). We tracked changes in angle differences between the pulse direction and flight directions of the moths (angle θmoth) over the flight time so that the amount of acoustical glints returning from the moths to the bat could be estimated. Figure 5 shows an example of the changes in the θmoth of moths 1 and 2 during the flight shown in Fig. 3. In this case, the bat directed its pulse toward moth 2 (black arrows) before the consecutive emissions of long-duration pulses. And then, the θmoth of moth 1

13

0

0-60

60-120 moth

120-180

[deg.]

Fig. 6  Comparison of θmoth of the moth that was the target of the bat’s pulse and the moth that was not targeted at the time of pulse emission. Data were taken from a total of 94 pulses emitted before approach phase (from 12 flight recordings of 4 bats, 2–4 flights from each individual)

increase whereas the θmoth of moth 2 decreased. From the acoustical measurements of echoes from fluttering moths shown in Fig. 2, these observed changes in the θmoth of moths 1 and 2 suggest that there were increase in the extent of the acoustical glints of echoes from moth 1 but decrease in the glints from moth 2. During this period, the bat shifted its pulse direction from moth 2 to moth 1. Here, we offered a hypothesis that the bats chose their targeted moths depending on the amount of acoustical glints. Figure 6 shows the comparison of θmoth in moths who were targets of bat’s pulses and those that were not

J Comp Physiol A (2014) 200:799–809

targeted when pulses were emitted, classified into three equal ranges: 0°–60°, 60°–120° and 120°–180°. For this analysis, data were taken from a total of 94 pulses that were emitted before the approach phase, including all long-duration pulses, from a total of 12 flight recordings for 4 bats (note that when the bats decided between two target moths, the approach phase began). The highest probability that a moth would be targeted by the bat occurred when θmoth was within 60°–120° relative to the bat’s pulse direction. These angles encompass 70°–100°, which were demonstrated to produce strong (both in frequency and amplitude) acoustical glints of the echoes (see Fig. 2b, c). Conversely, the bats tended not to aim at moths whose θmoth was between 120° and 180°, which produced small acoustical glints. These findings suggest that distinct acoustical glints appear to attract bats for prey selection.

Discussion Previous acoustical measurements in the laboratory have demonstrated that the wing movements of fluttering insects cause both frequency and amplitude modulations in the CF component of returning echoes (Schnitzler and Ostwald 1983; Schuller 1984; Kober and Schnitzler 1990). Furthermore, behavioral studies based on a two-alternative forcedchoice procedure have shown that CF-FM bats can discriminate the wing-beat rates of fluttering insects (von der Emde and Menne 1989) and also can recognize insect species using acoustical glints (von der Emde and Menne 1989). These findings strongly suggest that CF-FM bats use acoustical glints produced by insect fluttering as an important cue during foraging. In this study, bats in flight were tasked with choosing between two fluttering moths, and changes in the direction of the bat’s attention (pulse direction) were monitored while the bat approached the moths for capture. The results show that the bats were more attracted to moths that produced “glinting” echoes, i.e., acoustical glints with strong frequency and amplitude modulation. The amplitude and frequency of glints strongly depend on the incident angle of the sound. Angles at which the wing area of the fluttering moth was perpendicular to the emitted sound produced the strongest acoustical glints (increased by 6–11 dB compared with those from the back and front of the fluttering moth in Fig. 2b). To detect the presence of small prey and even identify or classify insects using ultrasound against high atmospheric attenuation in the air, bats use acoustical glints as a dominant cue during prey capture (Trappe and Schnitzler 1982; Schnitzler and Flieger 1983; Schnitzler and Ostwald 1983; von der Emde and Menne 1989; von der Emde and Schnitzler 1990; reviewed in Schnitzler and Denzinger 2011). Our results confirm these earlier findings using the Telemike recordings combined

807

with microphone arrays, and provide behavioral evidence that bats use information from acoustical glints during inflight prey selection for capture. For more reliable quantitative results, statistical analyses should be used to compare bat behavioral responses among various insect orientation. Because it is still difficult to manipulate insect orientation against bat flight direction using real moths, we will conduct further investigations using quasi-fluttering moths that will allow us to directly manipulate the extent of acoustical glints for data collection and statistical analysis. Not only such acoustical cues, but also the flight direction and/or relative positions of the prey to the bat appear to be potentially relevant features enabling bats to determine prey items during foraging. In the preliminary experiment for this study, when two moths were placed at different distances, i.e., 2.5 and 4 m, from the bat’s starting point, the bats were observed to approach the moth whose initial position was closer to the bat’s starting point (data not shown). However, when the initial target distances of the two moths were the same, such as in this study, the bats tended to select the moth that most likely produced more echoes containing stronger glints, especially during the consecutive emissions of long-duration pulses. The bat can repeatedly detect the fluttering period of target moths by increasing pulse duration, which provides the bats with detailed information for prey selection. To date, we have not found any evidence that small differences in the distance between the two moths relative to the bats curtail bat prey choice: thus, during foraging, the bats did not always direct their attention to the nearest moth. However, our microphone array recordings of wild bats during natural foraging suggest the possibility that the bats may conduct path planning for multiple prey capture (Fujioka et al., under review). When the sonar emissions of Japanese house bats (Pipistrellus abramus) were recorded with a 32-channel microphone array for measurement of their flight path and pulse direction in the field, the acoustically tracked flight paths of the bats suggested that during foraging, the bats sometimes follow complicated flight paths, which do not simply direct them toward the nearest prey. Thus, when bats consecutively completed two interception maneuvers to capture targets, they sometime aimed their sonar sounds toward the second target before completing the capture of the first target. This also indicates that the bat focus not only on immediate, close-at-hand targets but also on upcoming targets. It is well known that, during flight, each respiration cycle of bats is accompanied by a wing-beat cycle (Suthers et al. 1972; Lancaster et al. 1992, 1995). Since the bats adjust their pulse emission timing according to their distance from a target of interest, foraging behavior should be achieved through a collaboration of flight and echolocation. Therefore, various factors in addition to the simple distance

13

808

between the bat and prey, likely affect the flight path planning of bats during foraging, including physical restrictions on flight and echolocation controls and the amount of energy required. In our present study, the limitations of the flight space [8 (L) × 3 (W) × 2 (H)] may have restricted the path planning of the bats. As a result, rather than mutual interactions between the bat and the moths regarding flight paths and distance, acoustical features of the glints encoded in the echoes appeared to form a simple dominant cue for prey selection during the laboratory recordings. Goniocraspidum pryeri which have ears are observed to show evasive flight, during which they suddenly change their flight direction away from a bat, combined with an increase in flight speed when a bat approaches within approximately 1 m (Matsuta et al. 2013). Some additional evasive behaviors have been described in other moth species (reviewed in Miller and Surlykke 2001), such as stopping flight to reduce the chances of being caught by a bat (Fenton and Fullard 1979), and producing trains of ultrasonic clicks to jam bat sonar (Corcoran et al. 2009). These adaptations by the moths have been studied in the context of selective pressure from bats (Surlykke and Filskov 1999; Miller and Surlykke 2001). When the bats emit sounds directed toward the back of a fluttering moth, the amplitude and frequency glints of the returning echo are the minimal because of the small reflective surface area (Fig. 2b). Therefore, flying with their backs to approaching bats reduced the acoustic delectability of the moths. It is also interesting to see whether the moths intentionally plan “acoustical-stealth” flight paths during evasive action. Acknowledgments  We thank Dr. Takuma Takanashi and Dr. Ryo Nakano for their valuable support. We also thank Nobutaka Urano for assistance in capturing bats in the field. This work was partly supported by a Grant-in-Aid for Young Scientists (A) (Grant No. 70449510) from the Japan Society for the Promotion of Science (JSPS). These experiments complied with the Principles of Animal Care, publication no. 86-23, revised 1985, of the National Institutes of Health, and with current Japanese laws. All experiments were approved by the Animal Experiment Committee at Doshisha University.

References Corcoran AJ, Barber JR, Conner WE (2009) Tiger moth jams bat sonar. Science 325:325–327 Fenton MB, Fullard JH (1979) The influence of moth hearing on bat echolocation strategies. J Comp Physiol A Neuroethol Sens Neural Behav Physiol 132:77–86 Fujioka E, Aihara I, Watanabe S, Hosokawa Y, Kasai R, Hiryu S, Simmons JA, Riquimaroux H, Watanabe Y Rapid shifts of sonar attention to multiple prey items by foraging echolocating bats (under review) Hiryu S, Katsura K, Lin LK, Riquimaroux H, Watanabe Y (2005) Doppler-shift compensation in the Taiwanese leaf-nosed bat (Hipposideros terasensis) recorded with a telemetry microphone system during flight. J Acoust Soc Am 118:3927–3933

13

J Comp Physiol A (2014) 200:799–809 Hiryu S, Shiori Y, Hosokawa T, Riquimaroux H, Watanabe Y (2008) On-board telemetry of emitted sounds from free-flying bats: compensation for velocity and distance stabilizes echo frequency and amplitude. J Comp Physiol A 194:841–851 Kober R, Schnitzler H-U (1990) Information in sonar echoes of fluttering insects available for echolocating bats. J Acoust Soc Am 87:882–896 Lancaster WC, Keating AW, Henson OW Jr (1992) Ultrasonic vocalizations of flying bats monitored by radiotelemetry. J Exp Biol 173:43–58 Lancaster WC, Henson OW Jr, Keating AW (1995) Respiratory muscle activity in relation to vocalization in flying bats. J Exp Biol 198:175–191 Link A, Marimuthu G, Neuweiler G (1986) Movement as a specific stimulus for prey catching behaviour in rhinolophid and hipposiderid bats. J Comp Physiol A 159:403–413 Mantani S, Hiryu S, Fujioka E, Matsuta N, Riquimaroux H, Watanabe Y (2012) Echolocation behavior of the Japanese horseshoe bat in pursuit of fluttering prey. J Comp Physiol A Neuroethol Sens Neural Behav Physiol 198:741–751 Matsuta N, Hiryu S, Fujioka E, Yamada Y, Riquimaroux H, Watanabe Y (2013) Adaptive beam-width control of echolocation sounds by CF-FM bats, Rhinolophus ferrumequinum nippon, during preycapture flight. J Exp Biol 216:1210–1218 Miller LA, Surlykke A (2001) How some insects detect and avoid being eaten by bats: tactics and countertactics of prey and predator. Bioscience 51:570–581 Neuweiler G, Metzner W, Heilmann U, Rubsamen R, Eckrich M, Costa HH (1987) Foraging behavior and echolocation in the rufous horseshoe bat (Rhinolophus rouxi) of Sri Lanka. Behav Ecol Sociobiol 20:53–67 Novick A (1963) Pulse duration in the echolocation of insects by the bats, Pteronotus. Ergebnisse Biol 26:21–26 Sano A (2006) Impact of predation by a cave-dwelling bat, Rhinolophus ferrumequinum, on the diapausing population of a troglophilic moth, Goniocraspidum preyeri. Ecol Res 21:321–324 Schnitzler HU, Denzinger A (2011) Auditory fovea and Doppler shift compensation: adaptations for flutter detection in echolocating bats using CF-FM signals. J Comp Physiol A 197:541–559 Schnitzler H-U, Flieger E (1983) Detection of oscillating target movements by echolocation. J Comp Physiol A 153:385–391 Schnitzler HU, Kalko EKV (2001) Echolocation by insect-eating bats. Bioscience 51:557–569 Schnitzler HU, Ostwald J (1983) Adaptations for the detection of fluttering insects by echolocation in Horseshoe bats. In: Ewert JP, Capranica RR, Ingle DJ (eds) Advances in vertebrate neuroethology. Plenum Press, New York, pp 801–827 Schnitzler HU, Hackbath H, Heilmann U, Herbert H (1985) Echolocation behavior of rufous horseshoe bats hunting for insects in the flycatcher-style. J Comp Physiol A 157:39–46 Schuller G (1984) Natural ultrasonic echoes from wing beating insects are encoded by collicular neurons in the CF-FM bat, Rhinolophus ferrumequinum. J Comp Physiol A 155:121–128 Surlykke A, Filskov M (1999) Auditory relationships to size in noctuid moths: bigger is better. Naturwissenschaften 86:238–241 Suthers RA, Thomas SP, Suthers BJ (1972) Respiration, wing-beat and ultrasonic pulse emission in an echo-locating bat. J Exp Biol 56:37–48 Tian B, Schnitzler HU (1997) Echolocation signals of the greater horseshoe bat (Rhinolophus ferrumequinum) in transfer flight and during landing. J Acoust Soc Am 101:2347–2364 Trappe M, Schnitzler HU (1982) Doppler-shift compensation in insect-catching horseshoe bats. Naturwissenschaften 69:193–194 Vogler B, Neuweiler G (1983) Echolocation in the noctule (Nyctalus noctula) and horseshoe bat (Rhinolophus ferrumequinum). J Comp Physiol A 152:421–432

J Comp Physiol A (2014) 200:799–809 von der Emde G, Menne D (1989) Discrimination of insect wingbeatfrequencies by the bat Rhinolophus ferrumequinum. J Comp Physiol A 164:663–671

809 von der Emde G, Schnitzler HU (1990) Classification of insects by echolocating greater horseshoe bats. J Comp Physiol A 167:423–430

13

Prey pursuit strategy of Japanese horseshoe bats during an in-flight target-selection task.

The prey pursuit behavior of Japanese horseshoe bats (Rhinolophus ferrumequinum nippon) was investigated by tasking bats during flight with choosing b...
1MB Sizes 0 Downloads 3 Views