Mammalian Biology 78 (2013) 94–103

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Mammalian Biology

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Original Investigation Natural history, physiology and energetic strategies of Asellia tridens (Chiroptera)

Eran Amichai ∗, Eran Levin, Noga Kronfeld-Schor, Uri Roll, Yoram Yom-Tov

Department of Zoology, Faculty of Life Sciences, Tel Aviv University, Tel Aviv, article info abstract

Article history: We used radio-telemetry, observations and physiological measurements to study the basic biology and Received 20 February 2012 energetic strategies of Asellia tridens in northern Israel from 2009 to 2010. Between late May and early Accepted 21 June 2012 November, the occupied abandoned man-made structures in this area. Parturition occurred between Available online 25 July 2012 late June and mid-July, and juveniles were independent by late August. A. tridens foraged near the roost in a vegetation-rich, cluttered background environment, catching insects flying close to vegetation. Its Keywords: diet was diverse, with Coleoptera, Heteroptera, Diptera and Lepidoptera being the main diet components. Chiroptera During summer, males and females differed in their foraging patterns and energetic strategies: Lactating Foraging pattern Energetic strategies females departed for more frequent foraging bouts than males, and maintained euthermy throughout Reproductive cycle the day, while males became torpid on a daily basis. Torpor © 2012 Deutsche Gesellschaft für Säugetierkunde. Published by Elsevier GmbH. All rights reserved.

Introduction and Hymenoptera (Feldman et al., 2000; Whitaker and Yom-Tov, 2002). However, its natural history – foraging behaviour, prey pref- Asellia tridens (Geoffroy, 1813; ) is a medium- erence, home range, reproduction cycle, and seasonal movement – sized (forearm length ca. 50 mm, body mass ca. 12–13 g, is still poorly known. (Jones et al., 1993)). In its family it is the species most adapted Although are generally considered to be euther- to arid environments and has the widest distribution, inhabit- mic, a large number of mammalian species use heterothermy ing Mediterranean to extremely arid habitats from North-western either for short period (daily) torpor or for long term hiberna- Africa through the Arabian Peninsula east to Pakistan (Nowak, tion. Heterothermy is common in temperate zones small mammals, 1994). In Israel it is the only Hipposiderid, where it is relatively including bats, and is usually associated with strong seasonality, common in the Jordan and Arava Valleys, (part of the Great Rift low winter temperatures and low food availability. Lately, how- Valley). In this area A. tridens is known to roost in colonies number- ever, it is becoming evident that heterothermy is not only present, ing several hundred to several thousand individuals (Mendelssohn but probably common, in tropical and sub-tropical species as well and Yom-Tov, 1999). It occupies maternity and summer roosts from (Geiser and Stawski, 2011; Levy et al., 2011a,b; Liu and Karasov, late spring (May) to mid autumn (November), but in most cases 2011; Lovegrove, 2000). its wintering locations are unknown (Mendelssohn and Yom-Tov, Male and female A. tridens arrive at the summer roosts in dif- 1999). Little work has been carried out on the biology of A. tri- ferent stages in their reproductive cycle, and are therefore subject dens. There is some information on its reproductive cycle: in it to different energetic demands and constraints (Gittleman and gives birth to a single pup around June, after a pregnancy period of Thompson, 1988; Kurta et al., 1989). We hypothesized that these 9–10 weeks, and lactation lasts about 40 days (Mendelssohn and differences between the sexes will result in different energetic Yom-Tov, 1999; Qumsiyeh, 1996). It produces echolocation calls strategies that will be evident in fat reserve accumulation and with a cf. component at 111–124 kHz lasting 3–10 ms that ends daily body temperature patterns, and that males, but not lactating with a terminal downward sweep of 19–21 kHz (Gustafson and females, will use daily torpor. Such differences have been described Schnitzler, 1979) and its flight morphology have been character- in species from temperate zones (Dietz and Kalko, 2006; Grinevitch ized as suitable for cluttered environments (Jones et al., 1993). Its et al., 1995), but it is unclear whether they exist in species with trop- guild association in the Dead Sea area is a cluttered environment bat ical or sub-tropical distribution, where ambient temperatures are and its diet in the area consists mostly of Coleoptera, Lepidoptera much higher. In this study we used temperature sensitive radio-transmitters to record skin temperatures and combine these data with exten- sive observations and measurements to present some data on ∗ Corresponding author. Tel.: +972 3 6409397; fax: +972 3 6409403. the ecophysiology and biology of A. tridens at the limit of its E-mail address: [email protected] (E. Amichai). distribution.

1616-5047/$ – see front matter © 2012 Deutsche Gesellschaft für Säugetierkunde. Published by Elsevier GmbH. All rights reserved. http://dx.doi.org/10.1016/j.mambio.2012.06.006 E. Amichai et al. / Mammalian Biology 78 (2013) 94–103 95

Material and methods Three additional roosts were monitored occasionally (4–5 times each in the season) to provide comparisons from different regions: Study area Timna Cave colony in the southern Arava desert (29◦45N, 34◦59E), Tel-Azaz’yat colony in the Golan Heights (33◦13N, 35◦40E) and The bats were studied in the Beit-She’an valley (32◦33N, Kochav-Ya’ir colony in the eastern Mediterranean region (32◦13N, 35◦34E). This area is a part of the Great African Rift system, com- 35◦00E). prising low flatlands on the eastern and western sides of the , about 11 km south of Lake Kineret. The Jordan River valley Observations is ca. 6 km wide, its floor is ca. 250 m below sea level, and it is enclosed on both sides by steep slopes. The climate is subtropi- Observations were carried out at least once a week during the cal, with dry and hot summers (mean daily maximal temperature summer (May through October) of 2009 and 2010. Number of indi- in July: 37.6 ◦C) and moderate winters (mean minimal tempera- viduals was estimated by counts on site and from photographs tures in February: 9.2 ◦C, mean maximal temperature in February: taken on site and examined later. Sex ratio estimate was possi- 19.2 ◦C, mean annual precipitation: 389.1 mm, Israel Meteorolog- ble due to the males’ easily visible penis. Behavioural observations ical Service). Water sources are abundant along the valley, with were carried out at least once every two weeks on emergence of numerous artificial (fishing ponds, reservoirs) and natural water the bats and during foraging, with the aid of bat detectors (D1000X, sources (springs), in addition to the Jordan River itself. Native PetterssonElektronik, Sweden). riparian vegetation is concentrated in streambeds and consists of Sudanian, Saharo-Sindian, Oriental and Mediterranean elements Measurements (Yom-Tov and Tchernov, 1988). A mixture of native and disturbed habitat vegetation exists on fishpond banks and along irrigation Bats were captured inside the roosts using hand nets. Bats were channels. Most of the valley area is used for agriculture, mainly not individually marked, but from 60 of the total 341 adults cap- grain fields and banana plantations west of the Jordan, and orchards tured a wing biopsy was taken for future genetic analysis, and an and palm tree plantations east of the Jordan. additional 20 were fitted with transmitters. Only two of those 80 Four roosts in the area were consistently monitored during bats have been recaptured, suggesting a negligible recapture rate. 2009–2010 (Fig. 1): three abandoned military bunkers on the slopes Captures were carried out during both seasons, at least once every (TY south, TY north and Keren) and an abandoned fortified building 10 days, often more frequently. Each time 3–8 individuals were cap- on the valley floor (Gesher). tured, measured and released back into the colony. Measurements were carried out outside the roost to minimize disturbance, so that usually less than a minute was spent inside the roost. Our obser- vations show that a few minutes after such disturbances the bats relax and resume normal behaviour. Each captured individual was weighed to an accuracy level of 0.1 g using digital scales (Micron, China), its forearm (FA) length measured to an accuracy level of 0.1 mm using a caliper (Mitutoyo, Japan), and its sex was deter- mined. Bats were classified as either adult or juvenile according to the degree of finger joint ossification (Kunz, 1988) and pelage col- oration (Dietz, 2007). Reproductive state was noted in males (testes developed/un-developed) and in females (pregnant/lactating/post lactating/not reproductive). Fat reserve levels were assessed visu- ally according to the size of the uropatagium fat store. A 7-level index was created, from 0 (empty fat stores) to 6 (completely full fat stores). This fat level index was compared to body fat percentages obtained from 23 individuals using a Dual Emission X-ray Absorp- tiometry (DEXA, PIXImus, General Electric, USA). This method scans a subject using X-ray beams of two different energies, enabling the determination of fat percentage according to absorption lev- els, as well as other information such as bone mineral density. Fat index was found to be highly correlated with body fat percentage (Spearman rank correlation, N = 23, R2 = 0.87, p < 0.05) and was sub- sequently used to represent fat reserve levels. Roost temperature was obtained from TY south, TY north and Keren to an accuracy level of 1 ◦C using ThermochroniButtons (Dallas Semiconductors, USA) set to record ambient temperature at 3 hour intervals through- out the season. iButtons were attached to cables hanging from the ceiling about 40 cm under the bats.

Radio-telemetry

Radio-telemetry studies were carried out for two periods during the 2010 season. Five individuals – three males and two pregnant females, were monitored during the first period (13/6/2010–5/7/2010), which was aimed at developing a suit- able working protocol for the bats and our study area, allowing Fig. 1. Study area: Beit-She’an Valley, North-eastern Israel. Different shades of gray the identification of suitable vantage points for radio-telemetry represent different altitudes: lighter gray equals higher altitude. tracking and probable flight directions. During the second period 96 E. Amichai et al. / Mammalian Biology 78 (2013) 94–103

(20/7/2010–6/8/2010) we monitored 15 individuals – six males and calculated. Since each pellet usually holds more than one kind of nine lactating females. Bats were fitted with temperature-sensitive prey, the sum of relative frequencies is greater than one. LB-2T transmitters (Holohil Systems, Canada) weighing 0.61 g, within the accepted range of 5–7% of body weight (Aldridge and Permits Brigham, 1988; Bontadina et al., 2002). The hair between the bats’ shoulders was clipped and transmitters were attached using med- All procedures were performed under permits from the Israel ical adhesive (Perma-Type Company, USA). Tracking was done by Nature and Parks Authority, permits 2009/32077 and 2010/37754. triangulation between two observers, each equipped with an R1000 mobile receiver (Communication Specialist, USA) and a three- Results element yagi antenna. Cross-bearings of foraging bats were taken as often as possible, synchronized using walkie-talkies (Uniden, Roost characterizations and seasonal movements China). Location-points (fixes) obtained from cross-bearings differ- ing by less than 30◦ were discarded. Bearings were rounded to the The majority of A. tridens roosts known in Israel, including all nearest 5◦ mark and we estimated our location-point error to be four of the present study, are abandoned man-made structures, no more than 60–70 m, based on signals received from bats inside in most cases military bunkers. During the period the roosts were the roost. The bats were assumed to be foraging if they were active occupied, the mean temperature inside was higher by ca. 2 ◦C than in a specific area (i.e. rapid changes in signal direction) for more the outside environment mean temperature. However, the daily than 5 min (Russo et al., 2002; Zahn et al., 2010); and to be com- temperature range was narrower than that of the environment, and muting if they moved from one activity site to another in direct the mean daily maximum was lower than the outside environment flight. There were no occurrences of immobile bats outside the (Table 1). Temperature inside the roost changed during the day, lag- roost which in other studies are associated with night roosting. In ging behind the temperature outside, with maximal temperature addition, two RX900 stationary receivers and data loggers (Televilt in the afternoon and minimal temperature in the early morning. International, Sweden) were used to record activity patterns and Except for a southern colony of about 100 individuals (both skin temperature. These were stationed inside the roosts (Keren male and female) in the Arava desert (Timna cave) that was popu- and TY north during the first period, Gesher and Keren during the lated throughout the year, A. tridens spent the winter in locations second period), and were programmed to scan the relevant trans- unknown to us. They occupied the Beit-She’an valley roosts starting mitter frequencies, allocating 20 s for each frequency. Depending in late May, in a gradual process that lasted until mid-July. The bats upon the number of active transmitters at any given time, the max- began leaving their summer roosts towards the end of September, imum interval between scans for each frequency was 2.5 min. An and by mid-November the roosts were completely empty (Table 2). individual was considered on a foraging bout if it was absent from the stationary data log for more than 20 min, but reappeared at Physical measurements and fat reserve accumulation some point during the same night. During the second period forag- ing was verified by location-points from the mobile receivers. Maps Adult forearm length (FA) was 51.49 ± 1.1 mm (mean ± SD, were generated using Arcview-GIS (Esri, USA), which was also used N = 334). Males FA were significantly larger than females’: male for range calculations by way of minimum convex polygons (MCP). FA was 51.79 ± 0.15 mm while female FA was 50.95 ± 1.1 mm Skin temperature (Tsk) was obtained using the transmitter (mean ± SD, N males = 213, N females = 121, Shapiro–Wilk test for pulse rate, as measured and logged by the stationary receivers in normality, w = 0.99 ♂, 0.98 ♀, p = 0.53 ♂, 0.1 ♀ T-test, t = 6.76, df = 332, the roosts. Pulse rates from each transmitter were entered into p < 0.001). Both sexes were lean upon arrival to their summer equations calculated from calibration curves provided by theman- roosts but gained weight significantly during the summer, and ufacturer in which the linear range contains the entire range of reached maximal weight at fall (September–October). On aver- ◦ bat body temperature (Tb) (20–40 C). We used Tsk as an approxi- age, males gained 3.74 g (33.9% of their initial weight) and females mation of Tb, as studies have shown that in small mammals Tsk is gained 2.23 g (19.7% of their initial weight). Mean maximal weight closely correlated to Tb (Daniel et al., 2010; Dausmann, 2005; Liu for males was 14.76 ± 1.3 g (mean ± SD, N = 30), significantly more and Karasov, 2011). than females: 13.05 ± 1.47 g (N = 32), with no significant difference between the sexes at the beginning of summer (early June). A sim- Diet analysis ilar pattern was observed for fat reserves: both sexes arrived at the summer roosts with depleted fat reserves and accumulated signifi- Faeces were collected from all four roosts, none of which hosted cant amounts of fat during the summer, males having accumulated other bat species at any time during the study. Plastic sheets significantly larger fat reserves than females (Fig. 2). (1 × 1 m) were spread beneath bat aggregations and fresh faeces were collected about every two weeks. After collection the sheets Reproductive cycle were cleaned and repositioned. Due to the uniformity of pellet contents within each collection we used a sample of six faecal Nearly all females (92.8%, N = 56) arrived at the summer roosts at pellets from each collection for analysis, following the method an advanced stage of pregnancy. Parturition was well synchronized described by Whitaker (1988), and insect remains were identified in the population: all births took place within three weeks, between to the lowest taxonomic level possible (usually Order). Because the end of June and mid-July. Females gave birth to a single pup, identification is based on cuticle remains, faecal analysis may lead born hairless, with closed eyes, and weighing ca. 2.5–3 g (up to 30% to under-representation of soft-bodied prey (such as spiders). How- of its mother’s weight). Similar to Rhinolophids, the mother pos- ever, in Lepidoptera this is not a problem, as moths are easily sesses pubic nipples from which the pup hangs. For the first 2–3 recognizable by their typical wing scales. Two parameters were days the pups remained constantly attached to the mother, even used to characterize A. tridens diet: (1) mean percentage volume: during foraging bouts. At 3–4 days the pups had grown hair and in each pellet the volume of the remains from each Order relative to opened their eyes, and were left hanging from the ceiling when the other Orders was visually estimated, and the mean for each Order in mothers foraged. At 3 weeks the pups were already volant but did each sample was calculated and (2) relative frequency: the num- not leave the roost. At this stage they did not use the pubic nipples ber of pellets in which remains from each Order was found was but hung head down from the mother’s shoulders. At 6–7 weeks counted, and its frequency relative to the rest of the pellets was the pups were independent and lactation ceased. E. Amichai et al. / Mammalian Biology 78 (2013) 94–103 97

Table 1 Environmental and roost temperatures.

Mean daily temperature (◦C) Mean daily maximum (◦C) Mean daily minimum (◦C) Mean daily range (◦C)

Environment 28.9 ± 3.1 36.1 ± 3.6 23 ± 3.5 13 ± 2.9 TY south 31.3 ± 2 34.2 ± 2.5 30 ± 1.9 4.2 ± 1.4 TY north 31.4 ± 1.9 33.6 ± 2.6 30.3 ± 1.8 3.3 ± 1.3 Keren 30.4 ± 2.5 33.2 ± 3.2 28.9 ± 2.6 4.3 ± 1.5

Table 2 Roost occupation and leaving times.

Roost Start of occupation Estimated maximum Last bat leaves Start of Estimated maximum number Last bat leaves 2009 number of individuals 2009 the roost, 2009 occupation 2010 of individuals 2010 the roost, 2010

TY south Mid May 300 21.10–13 16.6 300 7.11 TY north Mid May 300 21.10–13 6.6 350 26.10–14 Keren Mid May 500 5.11–26.10 6.6–25.5 400 15.11 Gesher Unknown 300 13.10 19.5–8 500 7.11

All males arrived at the summer roosts with under-developed Foraging patterns (regressed) testes, and for the most part remained this way until they left for their winter roosts. Only in a few individuals (6%, Between 30 min to one hour before sunset bat activity inside N = 216) did the testes begin to develop towards the end of summer. the roost increased, and bats gradually began to move towards the openings. About 10 min after sunset several individuals left Sexual segregation the roost and performed a quick flight, circling the area around the opening before returning inside. After several of these ‘recon- A high degree of sexual segregation initially existed in A. tri- naissance flights’ true emergence began, with groups of 2–20 bats dens colonies: more than 90% of individuals at Gesher were female, leaving the roosts at high speed and low altitude (1–1.5 m), flying while more than 90% of the individuals at TY south, TY north and close to terrain features and vegetation. Keren were male; although interestingly, females in these three lat- 140 foraging bouts were recorded from seven male (5.9 ± 5.8 ter roosts bred successfully. This segregation weakened in October, bouts per individual (mean ± SD)) and five female (19.8 ± 13.9 when juveniles became independent, as juveniles and adult females bouts per individual (mean ± SD)) radio-tagged individuals. Our left Gesher and settled in the other roosts, and males settled in findings revealed that male A. tridens display a bi-modal foraging Gesher. By the end of summer there was no discernible sexual pattern (Fig. 3A), with most foraging activity taking place at the segregation. beginning of the night – 54% in the first hour after sunset, and a In other regions of Israel different forms of sexual segregation second, smaller, peak at the end of the night. This pattern was not occur: in the Timna Cave colony in the southern Arava desert no found in females: while there was still an activity peak in the first segregation was observed and the cave was populated throughout hour after sunset (36%), for the rest of the night activity showed a the year (B. Shalmon, pers. com.); while the Tel-Azaz’yat colony more uniform pattern (Fig. 3B). The difference in activity pattern in the Golan Heights and the Kochav-Ya’ir colony in the eastern between the sexes is statistically significant – Chi-square test on Mediterranean region were populated by males alone, and for a percentage of bouts per hour after sunset: 2 = 49.9, df = 9, p < 0.01. relatively short period (less than four months) at the height of We obtained 60 full nights of foraging data: 24 full nights from summer. five different males and 36 full nights from five different lactating

Fig. 2. Seasonal changes in fat reserve levels (A) and weight (B) in adult female and male A. tridens in Beit-She’an valley, Israel in 2010. Data are given as mean ± SD, * Indicates statistical significance. Differences between the seasons were significant – Fat index: for females (᭹): Mann-Whitney U-test, N spring = 7, N fall = 26, p < 0.001. For males (): T-test, N spring = 30, N fall = 31, t-value = 12.5, df = 59, p < 0.001. Fat index did not differ between the sexes in the spring: Mann–Whitney U-test, N ♀ =7,♂ = 30, U = 96.5, p = 0.72 but did differ in the fall: N ♀ = 26, ♂ = 30, U = 238, p = 0.01. Weight: for females (᭹): Mann–Whitney U-test, N spring = 7, N fall = 25, U = 18.5, p = 0.002. For males (): T-test, N spring = 30, N fall = 30, t-value = 12.8, df = 58, p < 0.001. Weight did not differ between the sexes in the spring: Mann–Whitney U-test, N ♀ =7,♂ = 30, U = 103, p = 0.95 but did differ in the fall: T-test, N ♀ = 25, ♂ = 30, t-value = −3.97, df = 53, p < 0.001. 98 E. Amichai et al. / Mammalian Biology 78 (2013) 94–103

Fig. 3. Temporal foraging patterns – number and percentage of foraging bouts for each hour since sunset. (A) Males – N = 8, no. of nights = 24, no. of foraging bouts = 41. (B) Females – N = 6, no. of nights = 36, no. of foraging bouts = 99. females. A. tridens performed up to eight foraging bouts each night males roosting to the south of Gesher simply foraged in the closer (Table 3), and on average spent a total of 226.7 ± 232.7 min forag- of the two foraging sites. ing per night (mean ± SD). We compared male and female foraging We defined a circular area four times larger than the home range patterns using Poisson log-normal mixed effects model, with total (14.25 km2) and centred around the centre of the home range as foraging time per night, foraging bout length and number of forag- ‘Full area’ and divided it into 10 habitat types (Supplementary Fig. ing bouts per night as fixed factors and individuals as the random S1) to investigate A. tridens habitat preference. We compared the factors, using lme4 R package (Bates et al., 2011). Total foraging time proportions of different habitat types between ‘full area’ and ‘home and single bout length were not significantly different between the range’ to the proportion of time spent in each habitat type (Fig. 5). sexes (total foraging time: z = −0.907, p = 0.364; single bout length: While ‘open fields’, ‘natural mountainside’ and ‘orchards and plan- z = 0.746, p = 0.456); although males went on significantly fewer tations’ had the largest proportion in both full area and home range, (Z = −1.990, p = 0.0466) foraging bouts than females (Table 3). it is clear that they were not the preferred habitats by the bats, who An estimated minimum of 230 h of active radio-tracking preferred instead to forage in the much smaller ‘rich bank vegeta- resulted in 311 location points from 16 bats (six males, ten females). tion’ and ‘riparian vegetation’ habitats. While a significant amount Spatial foraging patterns are given for the Gesher roost area (Fig. 4), of time was spent in ‘open fields’, some of that time was spent com- which yielded the most results due to terrain favourable for radio- muting to, from and between foraging sites, and the amount of time telemetry (13 of the 20 bats fitted with transmitters were recorded spent foraging in open fields is probably smaller than appears in the foraging here). Due to a lack in high vantage points and the bats’ low chart. Other habitat types were mostly or completely avoided. flight, individuals foraging around the other roosts quickly disap- The bats used two main flight paths for commuting from and peared. Moreover, following the bats physically was not possible as to the roost, identified by both radio-telemetry and direct observa- large parts in the area are closed by the military. Consequently not tions: (a) ‘Western path’: formed by the gap between the fish-pond enough location points around those roosts were obtained for spa- banks on one side and the valley slopes on the other. (b) ‘East- tial analysis, except for bats from these roosts that foraged around ern path’: an irrigation draining ditch between fields (Fig. 4). Flight Gesher. In the Gesher roost area foraging took place within a rela- within the paths was fast, direct and close to the terrain. Both males tively small area. We defined ‘Home Range’ as the area containing and females used the same foraging sites, however since males usu- 90% of radio-telemetry location points (BÖRger et al., 2006). Home- ally arrived from different roosts, e.g. ‘Keren’ to the south, different range size was 3.57 km2 and at no point was it farther than 3 km flight paths were used. from the roost (Fig. 4). The bats left the roost and flew directly to foraging sites about 1.5–2 km from the roost, covering this distance Body temperature measurements in 3–5 min. The rest of the foraging bout was spent either at the foraging site or commuting between sites. Two foraging sites were Out of a total of 33,296 Tsk readings obtained during the study identified: a. ‘Western ponds’: fish-pond with vegetated banks and we isolated 63 occasions for which we had temperature data cov- the slopes of an adjacent artificial mound, 1.5 km from the roost. ering a 24 h period for an individual bat (hereafter: full day): 25 full Other fish-ponds lying within the home range had bare banks and days from four males (mean ± SD: 6.3 ± 4.3) and 38 full days from were not used by the bats for foraging; and b. ‘Tavor stream’: four lactating females (mean ± SD: 9.5 ± 3.1). Full days included riparian vegetation along a streambed, 2 km from the roost. Both foraging bouts, during which no temperature data were received. sites contained richer and denser vegetation than the surround- When Tsk was compared with roost temperature (Troost) it was evi- ing fields, and the bats were observed hunting while skimming dent that while lactating females maintained euthermia (around the vegetation, occasionally pursuing prey into more open air and ◦ 36 C), males did not, and instead allowed their Tb (represented immediately returning to skim the vegetation. While it appears as by Tsk) to fluctuate with Troost (Fig. 6). There was a strong corre- though there was sexual segregation in foraging sites’ with females lation between mean Troost at any given time and the mean male preferring ‘western ponds’ and males preferring ‘Tavor steam’, this Tsk at the same time (Spearman rank correlation: N = 134, p = 0.002, was probably a result of the sexual segregation in the roosts: some 2 R = 0.857, Fig. 7A). No such correlation existed between mean Troost

Table 3 Temporal foraging patterns.

Females (N = 5) Males (N = 5) Significance

Total foraging time per night (min) 242.8 ± 250.7 202.5 ± 201.7 NS (p = 0.36) Number of foraging bouts per night 2.7 ± 2.99 1.5 ± 0.64 p = 0.047 Length of single foraging bout (min) 88.7 ± 97.5 125.1 ± 126.1 NS (p = 0.45) E. Amichai et al. / Mammalian Biology 78 (2013) 94–103 99

Fig. 4. Home range and foraging sites of A. tridens around Gesher roost. Western flight path is depicted in dashed yellow line, eastern flight path with dashed red line.

2 and mean female Tsk (Spearman rank correlation, N = 192, p = 0.26,  (4) = 0.343, p = 0.987. Temporal variation in relative frequency: R2 = 0.426, Fig. 7B). Two daily peaks in body temperatures are dis- 2(4) = 1.529, p = 0.821). Additionally, a smaller number of faecal cernible in both sexes; one at the beginning of the night and one at pellets were collected from three colonies in other parts of Israel: the end. These peaks are associated with active re-warming before Tel Azaz’yat colony in the Golan Heights, Kochav-Ya’ir colony in foraging activity (Fig. 6). the Mediterranean region, and Timna colony in the Arava desert (Appendix A). Diet Discussion 125 faecal pellets collected from all four roosts throughout the summer of 2010 were analysed. Because individuals routinely Life history moved from one roost to another and sexual segregation was not complete, our diet analysis regards A. tridens in Beit-She’an valley Although no information exists regarding A. tridens location and as a single population with no further division into roosts or sexes time of mating in Israel, its reproductive cycle can be deduced (Fig. 8, Appendix A). There was temporal variation in the diet during from our findings: females arrived at the summer roost already the season (Fig. 9), but this variation was random and inconsistent pregnant, and males arrive with regressed testes that remained in and did not show a significant trend (temporal variation in mean this state throughout the summer and fall, suggesting spermato- percent volume: Friedman’s repeated measures ANOVA on ranks: genesis does not start before winter. Spermatogenesis is a process

Fig. 5. Habitat preference – the proportion of different habitat types in the full area and home range, and the proportion of time spent in each habitat relative to its size. 100 E. Amichai et al. / Mammalian Biology 78 (2013) 94–103

Fig. 6. (A) Mean Skin temperature of male and female A. tridens and roost temperature in Beit-She’an colonies, 2010. Data are given as mean ± SD. Darkened area on the time axis represents night-time. The gap in the data around 19:30 is due to the very well synchronized emergence of all bats in all nights for the night’s first foraging bout. (B) and (C) Daily plot of skin temperature and activity of a single male (B) and female (C). Each actogram row represents one day and is made of separate columns, the height of which represents skin temperature. Gaps in the data represent absence from roost (foraging bout). Short gaps during day-time are a result of the bat momentarily moving out of reception inside the roost. White and black bars at the top represent the light schedule. Actograms were generated using the Clocklab program (Actimetrics Wilmette, IL). lasting about four months (Gustafson and Shemesh, 1976), so mat- stores (Fig. 2) which reveal significant energy reserve storage before ing cannot occur prior to middle to late March. In Iraq, gestation in winter, significantly reduced activity during winter among bats that this species lasts 9–10 weeks (Mendelssohn and Yom-Tov, 1999; remained in the Rift Valley (Feldman, 1998), and lack of foraging Qumsiyeh, 1996). Since parturition starts in mid-late June, mating activity during winter in the Timna year-round colony (B. Shalmon, cannot occur later than April. Hence, mating probably occurs during pers. com.). April, and such phenomena as delayed embryonic development or temporal separation between spermatogenesis and mating, which Foraging and diet were reported for other species (Hosken et al., 1998; Meenakumari and Krishna, 2005), apparently do not occur in A. tridens. The observed foraging strategy was in accordance with expec- Wintering locations of A. tridens in Israel are mostly unknown. tations from wing morphology and echolocation characteristics However, several phenomena suggest that A. tridens hibernate dur- (Norberg and Rayner, 1987): A. tridens searches for flying prey ing the winter, rather than simply migrate to winter activity areas. while skimming dense vegetation, with its low wing loading and These phenomena include seasonal changes in body mass and fat aspect ratio (Jones et al., 1993) enabling high manoeuvrability. In

Fig. 7. Correlation between mean roost temperature at a given time and mean skin temperature of males (A) and females (B). E. Amichai et al. / Mammalian Biology 78 (2013) 94–103 101

Fig. 8. Mean percent volume (left) and relative frequency (right) of insect orders in A. tridens faecal pellets collected in four roosts in Beit-She’an valley, Israel. addition, its high frequency CF-FM echolocation enables it to dis- that while A. tridens specializes in its foraging strategy, it is not a tinguish insect wing-beat against the cluttered background. All selective feeder; but, rather, it hunts whatever prey is available at summer colonies of A. tridens were located in the vicinity of water its foraging site. sources, but we never observed this bat drinking. Foraging sites were relatively close to the roost, reducing energetic costs during Energetic strategies and the differences between the sexes commuting flights to foraging habitats and water sources. Inter- estingly, in the , A. tridens only inhabits man-made Our hypothesis regarding different energetic strategies between structures close to the river, and was never found in other structures the sexes was confirmed: while in the roost, males let their T drop that are located more than a few hundred meters from the water. b close to ambient temperature (Fig. 6). Torpor in subtropical bats The use of protected linear landscape elements as flight paths may was reported for several different bat families (reviewed by Geiser serve as orientation guidelines and may also reduce predation risks and Stawski (2011)), including Hipposideridae (Baudinette et al., and provide shelter from the wind (Verboom and Huitema, 1997). 2000; Kulzer, 1965; Kulzer et al., 1970; Liu and Karasov, 2011). Male foraging displayed a bi-modal activity pattern similar to Pregnant and lactating females did not enter heterothermy but that documented in other insectivorous bat species, and corre- instead maintained euthermy. A similar difference between the sponding to the activity patterns of flying insects (Esbérard and sexes has been documented in temperate-zone species (e.g. Eptesi- Bergallo, 2010; Rydell et al., 1996; Taylor and Oneill, 1988). Forag- cusfuscus (Grinevitch et al., 1995) and Myotisdaubentonii (Dietz and ing bouts of females were shorter and more frequent than those of Kalko, 2006)), and the females’ euthermy may be explained by males, possibly related to the fact that all our radio-tagged females the cost of heterothermy for females: reduced metabolic rate may were lactating, and needed to minimize their time outside the roost slow embryonic development, lengthen pregnancy and lactation in order to care for their young. However, overall foraging time of periods, and shorten the time available for females to accumulate females was not shorter than that of males, possibly since nutri- fat reserves before winter and hibernation. tional and water demands on the mothers are high (Grinevitch et al., The males’ ability to enter daily torpor reduces energy con- 1995). sumption (Geiser, 2004) and results in the males accumulating Coleoptera and Heteroptera were found to be important com- significantly larger fat reserves by the end of summer, which may ponents in A. tridens’ diet, as was reported previously for other be necessary for courtship and mating towards the end of winter. areas of Israel (Feldman et al., 2000; Whitaker and Yom-Tov, 2002). Coleoptera were also found to be abundant in the diet of A. tridens in West Africa (Jones et al., 1993). Other diet components, however, Conservation varied between the areas, with Hymenoptera being significant in the Dead Sea area, Lepidoptera in the Arava and the Golan Heights, We recommend that conservation plans for A. tridens in the area and Diptera in the Beit-She’an valley, where several times a week should center on two issues: during the summer there is a mass emergence of Chironomidae Roosts: Almost all A. tridens populations in Israel depend upon (order: Diptera) from the local fish-ponds (Appendix A). The preva- abandoned man-made structures with stable micro-climate con- lence of two large and abundant insect orders in A. tridens diet, ditions for roosts. These structures, be they bunkers, shelters or coupled with regional variation in other diet components, suggests other buildings, should be preserved and should be protected

Fig. 9. Temporal variation in mean percent volume (A) and relative frequency (B) of insect orders in A. tridens faecal pellets collected in four roosts in Beit-She’an valley, Israel. 102 E. Amichai et al. / Mammalian Biology 78 (2013) 94–103 from excessive human interference, as roost availability may be an References important limiting factor for bat populations (Fenton et al., 2001). Foraging sites: Conserving riparian vegetation and native vegeta- Aldridge, H.D.J.N., Brigham, R.M., 1988. Load carrying and maneuverability in an insectivorous bat: a test of the 5% “Rule” of radio-telemetry. J. . 69, tion areas is of great importance. Even though citrus orchards, palm 379–382. groves and banana plantations exist in the area, sometimes within Bates, D., Maechler, M., Bolker, B., 2011. lme4: Linearmixed-effects Mod- A. tridens home range, we rarely recorded them foraging there (only els Using S4 Classes. R-package Version 0.999375-42. http://CRAN.R- project.org/package=lme4. 10 out of 311 location points), pointing to these bats’ preference Baudinette, R.V., Churchill, S.K., Christian, K.A., Nelson, J.E., Hudson, P.J., 2000. for natural and less monoculture vegetation. Fish ponds provide an Energy, water balance and the roost microenvironment in three Australian cave- abundant food source at specific times, and can also provide good dwelling bats (Microchiroptera). J. Comp. Physiol. B 170, 439–446. foraging sites if their banks are covered with vegetation. Though we Bontadina, F., Schofield, H., Naef-Daenzer, B., 2002. Radio-tracking reveals that lesser horseshoe bats (Rhinolophus ) forage in woodland. J. Zool. 258, have no direct observation of A. tridens drinking, in captivity they 281–290. drink when water is offered (E. Amichai, pers. observation), and so BÖRger, L., Franconi, N., De Michele, G., Gantz, A., Meschi, F., Manica, A., Lovari, S., fish ponds may be important in this respect as well. Coulson, T.I.M., 2006. Effects of sampling regime on the mean and variance of home range size estimates. J. Anim. Ecol. 75, 1393–1405. Daniel, S., Korine, C., Pinshow, B., 2010. The use of torpor in reproductive female Hemprich’s long-eared bats (Otonycteris hemprichii). Physiol. Biochem. Zool. 83, Future research 142–148. Dausmann, K.H., 2005. Measuring body temperature in the field – evaluation of Several aspects of A. tridens biology and ecology that arise from external vs. implanted transmitters in a small mammal. J. Therm. Biol. 30, this study are not completely understood and may warrant more 195–202. Dietz, C., 2007. Aspects of Ecomorphology in the Five European Horseshoe bats specific research. The reason for sexual segregation, for instance, (Chiroptera: Rhinolophidae) in the Area of Sympatry, Biology. University of is unclear, as both sexes roost in similar conditions and use the Tubingen, Tubingen. same foraging sites. Metabolic measurements will help elucidate Dietz, M., Kalko, E., 2006. Seasonal changes in daily torpor patterns of free-ranging female and male Daubenton’s bats (Myotis daubentonii). J. Comp. Physiol. B 176, the different physiological demands on males and females, and the 223–231. different thermoregulatory strategies employed by the different Esbérard, C., Bergallo, H., 2010. Foraging activity of the free-tailed bat Molos- sexes. While the advantages of daily torpor and seasonal hiberna- sus molossus (Chiroptera; Molossidae) in Southeastern Brazil. Braz. J. Biol. 70, 1011–1014. tion are well studied, the mechanisms that enable them in warm Feldman, R., 1998. The Insectivorous Bats (Microchiroptera) of the Dead Sea Area: and hot environments are less clear and A. tridens offers a good Food Habits and Habitat Use, Zoology. Tel-Aviv University, Tel-Aviv. model for such study. Feldman, R., Whitaker, J.O.J., Yom-Tov, Y., 2000. Dietary composition and habitat use in a desert insectivorous bat community in Israel. Acta Chiropterologica 2, 15–22. Acknowledgements Fenton, M.B., Bernard, E., Bouchard, S., Hollis, L., Johnston, D.S., Lausen, C.L., Ratcliffe, J.M., Riskin, D.K., Taylor, J.R., Zigouris, J., 2001. The bat fauna of Lamanai, Belize: roosts and trophic roles. J. Trop. Ecol. 17, 511–524. We are grateful to Dr. Amit Dolev of the Society for the Protec- Geiser, F., 2004. Metabolic rate and body temperature reduction during hibernation tion of Nature in Israel for use of equipment, good advice and help and daily torpor. Annu. Rev. Physiol. 66, 239–274. Geiser, F., Stawski, C., 2011. Hibernation and torpor in tropical and subtropical bats in the field; to Dr. Benny Shalmon of the National Parks Author- in relation to energetics, extinctions, and the evolution of endothermy. Integr. ity for information and data collection from the Timna colony; to Comp. Biol. 51, 337–348. Dr. Leonid Freidman of the Department of Zoology, Tel Aviv Uni- Gittleman, J.L., Thompson, S.D., 1988. Energy allocation in mammalian reproduction. Am. Zool. 28, 863–875. versity for help with insect remains identification; to Hila Yaffe Grinevitch, L., Holroyd, S.L., Barclay, R.M.R., 1995. Sex differences in the use of daily from Neve-Ur, Shmulik Landau of Tel Aviv University and torpor and foraging time by big brown bats (Eptesicus fuscus) during the repro- OferAmichai for assistance with radio-telemetry, to Ofir Levy of ductive season. J. Zool. 235, 301–309. Gustafson, A.W., Shemesh, M., 1976. Changes in plasma testosterone levels during the Department of Zoology, Tel Aviv University, for assistance the annual reproductive cycle of the hibernating bat, Myotis lucifugus lucifu- with statistics, and to two anonymous reviewers for their help- gus with a survey of plasma testosterone levels in adult male vertebrates. Biol. ful comments. This research was supported by The Israel Science Reprod. 15, 9–24. Foundation (Grant No. 232/08). UR is supported by the Adams Fel- Gustafson, Y., Schnitzler, H.U., 1979. Echolocation and obstacle avoidance in the Hipposiderid bat Asellia tridens. J. Comp. Physiol. 131, 161–167. lowship Program of the Israel Academy of Sciences and Humanities. Hosken, D.J., Blackberry, M.A., Stewart, T.B., Stucki, A.F., 1998. The male repro- ductive cycle of three species of Australian vespertilionid bat. J. Zool. 245, 261–270. Appendix A. Diet composition Jones, G., Morton, M., Hughes, P.M., Budden, R.M., 1993. Echolocation, flight mor- phology and foraging strategies of some West African Hipposiderid bats. J. Zool. 230, 385–400. Order Beit-She’an valley Tel Azaz’yat Kochav-Ya’ir Timna Kulzer, E., 1965. TemperaturregulationbeiFledermausen (Chiroptera) ausver- schiedenKlimazonen. Z. Vergl. Physiol. 50, 1–34. Coleoptera 27.9 ± 22.9 23.9 ± 20.5 53.5 ± 23.6 22.5 ± 24.3 Kulzer, E., Nelsen, J.E., McKean, J.L., Ohres, P., 1970. Untersuchungen uber die Tem- Heteroptera 23.3 ± 22 21.2 ± 18.5 41.5 ± 24.8 19.6 ± 23.2 peraturregulation australischer Fledermause (Microchiroptera). Z. Vergl. Physiol. ± ± ± Hemiptera 1.5 29.1 2.3 7.1 0 11.7 12.5 69, 426–451. ± ± Hymenoptera 2 7.3 0.8 1.7 7.1 0.8 Kunz, T.H., 1988. Ecological and Behavioral Methods for the Study of Bats. Smith- ± ± ± Lepidoptera 8.3 23.3 25.8 21.1 0.7 25.4 25.1 sonian Institute, Washington, DC. Diptera 23.2 ± 25.1 9.2 ± 14.1 0.8 ± 2.5 3.3 ± 5.8 Kurta, A., Bell, G.P., Nagy, K.A., Kunz, T.H., 1989. Energetics of pregnancy and lactation Other/unknown 13.9 ± 16.2 16.9 ± 18.3 1.8 ± 5.9 16.7 ± 26.4 in free ranging little brown bats (Myotis lucifugus). Physiol. Zool. 62, 804–818. Mean percent volume (mean ± SD) of different insect orders in Levy, O., Dayan, T., Kronfeld-Schor, N., 2011a. Adaptive thermoregulation in golden spiny mice: the influence of season and food availability on body temperature. A. tridens faeces from four different areas in Israel. Dictyoptera, Physiol. Biochem. Zool. 84, 175–184. Orthoptera, Neuroptera and Embioptera were also identified but Levy, O., Dayan, T., Kronfeld-Schor, N., 2011b. Interspecific competition and torpor highly uncommon and were classified together with unidentified in golden spiny mice: two sides of the energy-acquisition coin. Integr. Comp. Biol. 51, 441–448. items. Liu, J.N., Karasov, W.H., 2011. Hibernation in warm hibernacula by free-ranging For- mosan leaf-nosed bats, Hipposiderosterasensis, in subtropical Taiwan. J. Comp. Physiol. B 181, 125–135. Appendix B. Supplementary data Lovegrove, B.G., 2000. Daily heterothermy in mammals: coping with unpre- dictable environments. In: Heldmaier, G., Klingenspor, M. (Eds.), Life in the Supplementary data associated with this article can be Cold: Eleventh International Hibernation Symposium. Springer-Verlag, Berlin, Jungholz, Austria, pp. 29–40. found, in the online version, at http://dx.doi.org/10.1016/j. Meenakumari, K.J., Krishna, A., 2005. Delayed embryonic development in the Indian mambio.2012.06.006. short-nosed fruit bat. Cynopterus Sphinx. Zool. 108, 131–140. E. Amichai et al. / Mammalian Biology 78 (2013) 94–103 103

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