Control of doppler shift compensation in the greater horseshoe bat,Rhinolophus ferrumequinum

Summary FlyingRhinolophus ferrumequinum lower the frequency of the constant frequency part (f_A) of the emitted sounds in order to compensate for Doppler shifts caused by the flight speed. The echo frequency (f_E) is kept constant within a frequency band of about 200 Hz, the center frequency of which is about 150 Hz above the average or resting frequency (f_R) emitted by roosting bats shortly before take off. For the compensation they use a feedback control system in which the emission frequency is changed to hold the echo frequency at a criterion value. This feedback system was demonstrated by experiments with bats flying in an experimental wind tunnel and in a He-O₂-micture. In the wind tunnelRhinolophus lowers the emission frequency in order to compensate for Doppler shifts which are caused by the ground speed flown by the bat. In the He-O₂-mixtureRhinolophus compensates for Doppler shifts which correspond to the different sound speeds in the gas mixture.I would like to thank D. R. Griffin for his generous support and stimulating criticism. I express my appreciation to the New York Zoological Society for the use of its facilities and to R. Brown for technical assistance. The work was supported by grant number GB 7155 from the National Science Foundation to the New York Zoological Society. I also thank J. D. Pye for his suggestions.     

The photoreceptor populations in the retina of the greater horseshoe bat Rhinolophus ferrumequinum

Recently, we reported the existence of AII "rod" amacrine cells in the retina of the greater horseshoe bat Rhinolophus ferrumequinum (Jeon et al., 2007). In order to enhance our understanding of bat vision, in the present study, we report on a quantitative analysis of cone and rod photoreceptors. The average cone density was 9,535 cells/mm2, giving a total number of cones of 33,538 cells/retina. The average rod density was 368,891 cells/mm2, giving a total number of rods of 1,303,517 cells. On average, the total populations of rods were 97.49%, and cones were 2.51% of all the photoreceptors. Rod: cone ratios ranged from 33.85:1 centrally to 42.26:1 peripherally, with a mean ratio of 38.96:1. The average regularity index of the cone mosaic in bat retina was 3.04. The present results confirm the greater horseshoe bat retina to be strongly rod-dominated. The rod-dominated retina, with the existence of AII cells discovered in our previous study, strongly suggests that the greater horseshoe bat retina has a functional scotopic property of vision. However, the existence of cone cells also suggests that the bat retina has a functional photopic property of vision.     

Studies on the vaginal plug of the Japanese greater horseshoe bat, Rhinolophus ferrumequinum nippon

The vaginal plug of the Japanese greater horseshoe bat is composed of an outer, thick, hard and homogeneous layer originating from the stratum disjunctum of the vaginal mucosa and a thin, soft and opaque central core containing accidentally trapped dead spermatozoa. The vaginal plug appeared to be expelled just before or after ovulation in spring. Immediately after copulation in autumn, uterine spermatozoa were dead but few leucocytes were present. As time passes after copulation, however, the infiltration of leucocytes into the uterus increased gradually, and extensive leucocytic phagocytosis of dead spermatozoa occurred during hibernation.     

Parentage, reproductive success and breeding behaviour in the greater horseshoe bat (Rhinolophus ferrumequinum)

Female greater horseshoe bats form maternity colonies each summer in order to give birth and raise young. During the mating period, females visit males occupying territorial sites, copulation takes place and sperm are stored until ovulation occurs, normally in April. Using microsatellite markers and a likelihood method of parentage analysis, we studied breeding behaviour and male reproductive success over a five-year period in a population of bats in south-west Britain. Paternity was assigned with 80% confidence to 44% of young born in five successive cohorts. While a small annual skew in male reproductive success was detected, the variance increased over five years due to the repeated success of a few individuals. Mating was polygynous, although some females gave birth to offspring sired by the same male in separate years. Such repeated partnerships probably result from fidelity for either mating sites or individuals or from sperm competition. Females mated with males born both within and outside their own natal colony; however, relatedness between parents was no less than the average recorded for male female pairs. Gene flow between colonies is likely to be primarily mediated by both female and male dispersal during the mating period rather than more permanent movements.     

Echolocation sounds and hearing of the greater Japanese horseshoe bat (Rhinolophus ferrumequinum nippon)

Summary The relationship between the orientation sounds and hearing sensitivity in the greater Japanese horseshoe bat,Rhinolophus ferrumequinum nippon was studied.An orientation pulse consisted of a constant frequency (CF) component followed by a short downward frequency-modulated (FM) component. Sometimes, an initial upward FM component preceded the CF component. Duration of pulses was about 30 ms and the CF of resting pulses (RF) averaged 65.5 kHz. The best frequency (BF) at the lowest threshold in audiograms as measured by the pinna reflex averaged 66.1 kHz. Audiograms showed remarkable sharp cut-offs on both sides near the BF. The frequency difference between the BF and the RF was about 0.6 kHz, and the RF was always below the BF. The values of RF and BF were characteristically different from those of the European subspecies,Rhinolophus ferrumequinum ferrumequinum.Abbreviations BF best frequency - CF constant frequency - FM frequency modulated - RF resting frequency     

Prolonged survival of the Graafian follicle and fertilization in the Japanese greater horseshoe bat, Rhinolophus ferrumequinum nippon

After the mating season of the Japanese greater horseshoe bat in mid- or late October, only the right ovary maintained a single Graafian follicle throughout hibernation until early April. During this time the ovum was in prophase of meiosis I (resting stage) with many large lipid droplets as a nutrient source. In synchrony with stigma formation, there was resumption of meiotic activity, separation of the cumulus oophorus from the granulosa layer and dispersion of the follicle cells just before ovulation in spring. The block to polyspermy seemed to reside in the zona pellucida, because no spermatozoa could be detected in the perivitelline space of the 6 fertilized ova examined, although a second spermatozoon was recognized in the zona pellucida of 3 ova.     

Phylogeography of the greater horseshoe bat, Rhinolophus ferrumequinum: contrasting results from mitochondrial and microsatellite data

Phylogeographical studies are typically based on haplotype data, occasionally on nuclear markers such as microsatellites, but rarely combine both. This is unfortunate because the use of markers with contrasting modes of inheritance and rates of evolution might provide a more accurate and comprehensive understanding of a species' history. Here we present a detailed study of the phylogeography of the greater horseshoe bat, Rhinolophus ferrumequinum , using 1098 bp of the mitochondrial ND2 gene from 45 localities from across its Palaearctic range to infer population history. In addition, we re-analysed a large microsatellite data set available for this species and compared the results of both markers to infer population relationships and the historical processes influencing them. We show that mtDNA, the most popular marker in phylogeography studies, yielded a misleading result, and would have led us to conclude erroneously that a single expansion had taken place in Europe. Only by combining the mitochondrial and microsatellite data sets are we able to reconstruct the species' history and show two colonization events in Europe, one before the Last Glacial Maximum (LGM) and one after it. Combining markers also revealed the importance of Asia Minor as an ancient refugium for this species and a source population for the expansion of the greater horseshoe bat into Europe before the LGM.     

Vocal communication in adult greater horseshoe bats, Rhinolophus ferrumequinum

Whereas echolocation in horseshoe bats is well studied, virtually nothing is known about characteristics and function of their communication calls. Therefore, the communication calls produced by a group of captive adult greater horseshoe bats were recorded during various social interactions in a free-flight facility. Analysis revealed that this species exhibited an amazingly rich repertoire of vocalizations varying in numerous spectro-temporal aspects. Calls were classified into 17 syllable types (ten simple syllables and seven composites). Syllables were combined into six types of simple phrases and four combination phrases. The majority of syllables had durations of more than 100 ms with multiple harmonics and fundamental frequencies usually above 20 kHz, although some of them were also audible to humans. Preliminary behavioral observations indicated that many calls were emitted during direct interaction with and in response to social calls from conspecifics without requiring physical contact. Some echolocation-like vocalizations also appeared to clearly serve a communication role. These results not only shed light upon a so far widely neglected aspect of horseshoe bat vocalizations, but also provide the basis for future studies on the neural control of the production of communicative vocalizations in contrast to the production of echolocation pulse sequences.     

Determinants of echolocation call frequency variation in the Formosan lesser horseshoe bat (Rhinolophus monoceros)

The origin and maintenance of intraspecific variation in vocal signals is important for population divergence and speciation. Where vocalizations are transmitted by vertical cultural inheritance, similarity will reflect co-ancestry, and thus vocal divergence should reflect genetic structure. Horseshoe bats are characterized by echolocation calls dominated by a constant frequency component that is partly determined by maternal imprinting. Although previous studies showed that constant frequency calls are also influenced by some non-genetic factors, it is not known how frequency relates to genetic structure. To test this, we related constant frequency variation to genetic and non-genetic variables in the Formosan lesser horseshoe bat (Rhinolophus monoceros). Recordings of bats from across Taiwan revealed that females called at higher frequencies than males; however, we found no effect of environmental or morphological factors on call frequency. By comparison, variation showed clear population structure, with frequencies lower in the centre and east, and higher in the north and south. Within these regions, frequency divergence was directional and correlated with geographical distance, suggesting that call frequencies are subject to cultural drift. However, microsatellite clustering analysis showed that broad differences in constant frequency among populations corresponded to discontinuities in allele frequencies resulting from vicariant events. Our results provide evidence that the processes shaping genetic subdivision have concomitant consequences for divergence in echolocation call frequency.     

The complete mitochondrial genome of the greater horseshoe bat subspecies, Rhinolophus ferrumequinum korai (Chiroptera: Rhinolophidae)

The total length of the mitogenome of Rhinolophus ferrumequinum korai is 16,839?bp with a total base composition of 31.8% A, 25.4% T, 28.7% C, and 14.0% G. The mitogenome consists of 13 protein-coding genes, 2 rRNA (12S and 16S RNA) genes, 22 tRNA genes, and 1 control region.     

Distribution of oxytocin and vasopressin neurons in the diencephalon of the Japanese horseshoe bat, Rhinolophus ferrumequinum. An immunohistochemical study.

The distribution of oxytocin (OXT) and vasopressin (VP) neurons in the diencephalon of the hibernating Japanese horseshoe bat, Rhinolophus ferrumequinum, was immunohistochemically investigated by the avidin-biotin complex method. Magnocellular OXT and VP neurons were localized mainly in the paraventricular nucleus and the supraoptic nucleus. In addition to these main nuclei, both kinds of magnocellular neurons were also found in the periventricular nucleus, perifornical area and lateral hypothalamic area. Extensively distributed parvocellular neurons containing only VP were observed in the rostral and middle portions of the suprachiasmatic nucleus. The size of OXT and VP magnocellular neurons was almost equal in the paraventricular and ventromedial supraoptic nuclei, whereas VP neurons were significantly larger than OXT neurons in the dorsolateral supraoptic nucleus. The OXT and VP cells in the ventral supraoptic nucleus showed a distinctive elliptical shape. Both OXT and VP fibers were distributed in the lateral habenular nucleus, stria medullaris thalami, lateral preoptic area, stria terminalis, and medial and supracapsular part of the bed nucleus of the stria terminalis. Moreover, OXT fibers were found in the substantia nigra, and VP fibers were noted in the nucleus reunions and the paraventricular nucleus of the thalamus.     

Laryngeal nerve activity during pulse emission in the CF-FM bat,Rhinolophus ferrumequinum

Summary The activity of the external (motor) branch of the superior laryngeal nerve (SLN), innervating the cricothyroid muscle, was recorded in the greater horseshoe bat,Rhinolophus ferrumequinum. The bats were induced to change the frequency of the constant frequency (CF) component of their echolocation signals by presenting artificial signals for which they Doppler shift compensated. The data show that the SLN discharge rate and the frequency of the emitted CF are correlated in a linear manner.Abbreviations SLN Superior laryngeal nerve - RLN Recurrent laryngeal nerve - DCS Doppler compensation system - CF Constant frequency - FM Frequency modulation Supported by grants of the Deutsche Forschungsgemeinschaft (DFG), Az.: Schu 390/1, /2 and SFB 45We are indebted to Dipl.-Ing. H. Zöller for providing the computer programs. We want to thank H. Hahn and A. Polotzek for technical help.     

Ontogenesis of the echolocation system in the rufous horseshoe bat,Rhinolophus rouxi (Audition and vocalization in early postnatal development)

1. The development of vocalization and hearing was studied in Sri Lankan horseshoe bats (Rhinolophus rouxi) during the first postnatal month. The young bats were caught in a nursing colony of rhinolophids in which birth took place within a two week period. 2. The new-born bats emitted isolation calls through the mouth. At the beginning these calls consisted of pure tones with frequencies below 10 kHz (Fig. 1). During the first postnatal week the call frequency increased to about 15 kHz, and the fundamental was augmented by two to four harmonics. No evoked potentials to pure tone stimuli could be elicited in the inferior colliculus of this age group, i.e., auditory processing at the midbrain level was not demonstrable. 3. Evoked potentials were first recorded in the second week, broadly tuned to 15-45 kHz, with a maximum sensitivity between 15-25 kHz. In the course of the second week, however, higher frequencies up to 60 kHz became progressively incorporated into the audiogram (Fig. 3). The fundamental frequency of the multiharmonic isolation calls, emitted strictly through the mouth, increased to about 20 kHz. 4. In the bats' third postnatal week an increased hearing sensitivity (auditory filter) emerged, sharply tuned at frequencies between 57 and 60 kHz (Fig. 4e). The same individuals were also the first to emit long constant frequency echolocation calls through the nostrils (Fig. 4c). The energy of the calls was arranged in harmonic frequency bands with the second harmonic exactly tuned to the auditory filter. These young bats continued to emit isolation calls through the mouth, which were, however, not harmonically related to the echolocation calls (Fig. 4b, d). 5. During the fourth week, both the auditory filter and the matched echolocation pulses (the second harmonic) shifted towards higher frequencies (Fig. 5). During the fifth week the fundamental frequency of the calls was progressively attenuated, and both the second harmonic of the pulses and the auditory filter reached the frequency range typical for adult bats of 73-78 kHz (Fig. 6). 6. The development of audition and vocalization is discussed with regard to possible interactions of both subsystems, and their incorporation into the active orientation system of echolocation.     

Birth timing and population changes in greater horseshoe bat colonies (Rhinolophus ferrumequinum) are synchronized by climatic temperature

We counted the births of greater horseshoe bats born at three small breeding colonies in southwest Wales and south-west England at the extreme edge of the species distribution over nine summers (1984–1993). Overall birth timing was almost identical at the three sites (mean of mean birth dates = 13–14 July), but varied widely from year to year. Mean birth timing was synchronized at the three sites in a given year. Early births followed warm springs; a significant negative regression of mean birth date on mean April plus May temperature was evident. A rise of 2^oC accelerated mean birth date by about 18 days. Population levels fell at all three sites following the very late mean birth date of 28 July recorded in 1986, after an extremely cold spring and summer. Recovery of populations in all sites followed a series of warm springs and early mean birth dates, but was hampered by birth sex ratios favouring males for several years. These findings confirm climate, through its effect upon birth timing, and possibly also on sex ratios, as a major factor controlling bat populations, and hence the distribution of bat species.