dietary implications of intrapopulation variation in ... - Oxford Academic

Journal of Mammalogy, 89(5):1184–1190, 2008

DIETARY IMPLICATIONS OF INTRAPOPULATION VARIATION IN NITROGEN ISOTOPE COMPOSITION OF AN OLD WORLD FRUIT BAT L. GERARDO HERRERA M., CARMI KORINE,* THEODORE H. FLEMING,

AND

ZEEV ARAD

Estacioń de Biologıá de Chamela, Instituto de Biologıá, Universidad Nacional Autońoma de Me´xico, Apartado Postal 21, 48980 San Patricio, Jalisco, Me´xico (LGHM) Department of Biology, Technion-Israel Institute of Technology, Haifa 32000, Israel (CK, ZA) Department of Biology, University of Miami, Coral Gables, FL 33124, USA (THF) Present address of CK: Mitrani Department of Desert Ecology, Blaustein Institutes for Desert Research, Ben-Gurion University of the Negev, Midreshet Ben-Gurion 84990, Israel

We used nitrogen isotope analysis from pectoral muscle of the Egyptian fruit bat Rousettus aegyptiacus (Chiroptera: Pteropodidae) to determine intrapopulation variation in sources of dietary protein throughout the year in northern Israel. In Mediterranean climates, winter and summer are stable seasons, whereas spring and fall are transitional seasons. Number of species of fruit-bearing plants is higher during the transitional periods, and we therefore predicted that intrapopulation variation would be higher during spring than in winter and summer; we made no prediction for fall because sample size was small. We also reconstructed sources of dietary protein for each individual using nitrogen isotope ratios (d15N) to determine whether individuals foraged on the same sources of food within each season. Intrapopulation variation in d15N was significantly higher in spring (d15N range: 9.7–17.5&) compared to winter (8.8–11.1&) and summer (9.5–11.2&), suggesting that individuals during this period varied more in their use of protein sources. Dietary reconstruction revealed intrapopulation partitioning among the bats in the use of plant food items, and interspecific partitioning among plants in their dependence on dispersal by bats. Key words: dietary mixing models, dietary protein, intrapopulation variation, pteropodid bats, Rousettus aegyptiacus, stable-isotope analysis

Field studies show that diets of pteropodid bats (megachiropterans) include a large variety of fruits in addition to pollen and leaves (Banack 1998; Bumrunsgri et al. 2007; Korine et al. 1988, 1999; Nyhagen et al. 2005; Ritcher and Cumming 2006; Stier and Mildestein 2005), with a few reports of insect consumption (Barclay et al. 2006; Courts 1998). Although these studies offer detailed dietary information, their sampling approaches are more appropriate to describe feeding strategies at the population level, and they give little consideration to individual variation. Intrapopulation variation in resource use has been recognized for its potential ecological, evolutionary, and conservation implications (Bolnick et al. 2002, 2003). A population may be composed of individuals that feed on different subsets of items. In Carollia perspicillata (Phyllos-

* Correspondent: [email protected]

Ó 2008 American Society of Mammalogists www.mammalogy.org 1184

tomidae), for example, the diets of harem males differ significantly from those of bachelor males and females (Fleming 1988). Previous dietary field studies of pteropodid bats were based on the examination of day-roost droppings collected in traps that produced long-term population information, or of individual stomachs, feces, or pellets that rendered snapshot diet information. Snapshot information can infer longterm individual diets when individuals can be sampled repeatedly over relatively long periods (Estes et al. 2003), but this is not a viable option in most populations of freeranging bats. An alternative method for studying long-term intrapopulation feeding strategies is the use of stable isotope analysis (Bolnick et al. 2002). Its use is based on the existence of isotopic differences among groups of food items that are incorporated into animal tissues, allowing for quantification of their relative nutritional importance to an animal (Ehleringer et al. 1986; Hobson 1999; Lajtha and Michener 1994). Depending on the type of tissue examined, this method allows one to integrate individual feeding strategies at different timescales.

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GERARDO HERRERA M. ET AL.—VARIATION IN PROTEIN USE BY FRUIT BATS

For example, examination of liver, whole blood, and hair reflects the integrated diet of the animal over days to several weeks and months (Miroń et al. 2006; Tieszen et al. 1983). Intrapopulation variation in feeding strategies has been examined using the isotopic approach in several taxa, including gray wolves (Canis lupus—Urton and Hobson 2005), earthworms (Aporrectodea longa—Schmidt 1999), sea otters (Enhydra lutis—Estes et al. 2003), and blue tilapia fish (Oreochromis aureus—Gu et al. 1997). Stable isotope analysis has been used to study feeding habits of microchiropteran bats (Ceballos et al. 1997; Des Marais et al. 1980; Fleming et al. 1993; Herrera et al. 1998, 2001a, 2001b, 2002; Nassar et al. 2003; Voigt and Kelm 2006), but it has not been used previously in Old World fruit bats (Pteropodidae). We used nitrogen isotope analysis of pectoral muscle of the Egyptian fruit bat (Rousettus aegyptiacus) to determine intrapopulation variation in sources of protein throughout the year in the vicinity of Haifa, Israel. Protein is an essential nutrient for growth and reproduction in animals, and its availability influences the feeding ecology of organisms (White 1993). We tested the hypothesis that intrapopulation variation in d15N values would be higher during spring when fruit availability (number of species) increases compared to other times of the year. We further reconstructed the origin of dietary protein to determine whether individuals within the population foraged on the same dietary items, or if there were groups of individuals that favored subsets of available food sources.

MATERIALS AND METHODS Study site.— Samples of bats were collected in Rupin Cave, Haifa, Israel (338479N, 348599E), located on the northern slope of Mount Carmel, 3 km east of the Mediterranean shore. The cave in this urban area serves as a day-roost and is occupied year-round by thousands of Egyptian fruit bats (R. aegyptiacus). The Mediterranean climate at this site is characterized by cool, rainy winters and prolonged dry summers. The mean annual rainfall is approximately 600 mm, and the mean temperature is 288C in August and 138C in January (Amiran et al. 1970). Sample collection.— Adult male bats were captured with mist nets (Avinet, Dryden, New York) near the entrance of the cave on a seasonal basis in 1994 during the 2nd week of January (winter), April (spring), August (summer), and November (fall). Bats were brought to the laboratory, where 6 they were maintained in a temperature-controlled room (X SD; 228C 6 0.28C, 60% 6 3% relative humidity, 12D:12L). Bats were anesthetized using Pentothal (4.5 mg/kg; Sigma, Jerusalem, Israel) by intraperitoneal injection according to Holmes (1987) and Fowler (1993). Hair around the flight muscle was shaved, and a small cut was made to expose the muscle. A triangular-shaped piece of muscle tissue (;0.7 mm) was taken, the cut was carefully stitched, and an antibacteriogenic cream was applied. Bats were transferred to a cage in a temperature-controlled room (258C) where they recovered from the operation after 5–6 h. Bats reached full recovery and

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flight within 10 days after the operation, while being injected daily with antibiotics (Pen-Strep, 20/25 Vet; Eurovet Animal Health B.V., Bladel, Netherlands). Within 15 days, bats flew freely and were released at the entrance of Rupin Cave. The research was done under a permit issued by the Israel Nature and National Parks Protection Authority. We followed the guidelines of animal care and surgery regulations of all participating authors’ institutions. Procedures also met guidelines approved by the American Society of Mammalogists (Gannon et al. 2007). Even though the bats were released safely at their roost, we now recommend less-invasive protocols such as the use of small samples (;1 mg dry mass) of blood or wing membrane instead of muscle (Herrera et al. 2001a, 2001b, 2002; Voigt and Kelm 2006). Mass spectrometers at the time when samples were collected still relied on relatively large samples (;12–15 mg dry mass), thus restricting our selection to muscle tissue. The diet of Egyptian fruit bats was studied simultaneously during 2 years from 1993 to 1995 from feces collected in 2 other day-roosts located in the study area (Korine et al. 1999), and we used this information to identify potential food sources. The diet in these roosts consisted mainly of fruit from several species of plants, leaves from 1 plant species, and nectar and pollen from 1 species of plant. From the 14 species of plants reported by Korine et al. (1999), we considered 9 species for isotopic diet reconstruction. In winter, we considered fruits of Cerotonia siliqua, Diospyros kaki, Ficus microcarpa, and Melia azedarach, and leaves of F. religiosa; in spring fruits, of C. siliqua, F. microcarpa, F. sycomorus, Morus nigra, Eriobotrya japonica, and M. azedarach, and leaves of F. religiosa; in summer, fruits of Phoenix dactylifera, F. religiosa, F. microcarpa, F. sycomorus and M. nigra; and in fall, fruits of C. siliqua, D. kaki, P. dactylifera, F. religiosa, F. microcarpa, F. sycomorus, and M. azedarach, and leaves of F. religiosa. With the exception of F. carica (4–56% in feces) and F. rubiginosa (0–30% in feces) in summer and fall, most plants not included in the analysis were found in a very small proportion of the feces of the bats (0–12%). No insects were included in the isotopic analysis because this item was not found in bat droppings (Korine et al. 1999). For stable isotope analysis, samples from at least 6 individuals were collected from each of the 9 species of plants considered in our study. Stable-isotope analysis.— The nitrogen (N) and carbon (C) composition of pectoral muscle from individual bats was assessed using the methodology described by Sealy et al. (1987). We did not use C data to reconstruct protein origin because C can enter different metabolic pathways at the macromolecular level, including lipids, carbohydrates, and proteins (Hobson and Stirling 1997; Podlesak and McWilliams 2006). We present C stable isotope values because they may be of further use for other researchers. Twelve to fifteen milligrams of ground muscle samples were soaked in a 1 N HCl (J. T. Baker, Phillipsburg, New Jersey) solution for 4 h to dissolve bone carbonate and then combusted at 8008C in Vycor ampullae with 1 g of cupric oxide wire (Fisher Chemical, Pittsburgh, Pennsylvania), 1 g of copper (Aldrich, St. Louis, Missouri), and approximately 50 mg of

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Diet reconstruction.— Dietary mixing models can be used to solve for a unique combination of source proportions when the number of sources exceeds the number of stable isotopes analyzed by no more than 1 (Phillips 2001). In our case, there was only 1 isotope and 5–7 sources, depending on the season. Given the wide variation in plant d15N values, we grouped plants into 2 subsets of species that differed in mean d15N values (Mann– 6 SE; Whitney U ¼ 2.5, P ¼ 0.01): 15N-depleted plants (X 15 d N ¼ 4.9 6 0.4&, range: 3.4–6.3&, n ¼ 6; Fig. 1) and 15 N-enriched plants (7.8 6 0.5&, range: 7–9.3&, n ¼ 4; Fig. 1) species. We used the following 2-end-point mixing model (Fleming 1995) to estimate the extent to which each bat used each subset of plants as food according to the equation: d15 Nmuscle ¼ Pp ðd15 N15Ndepletedplants þ aÞ þ ð1 Pp Þðd15 N15Nenrichedplants þ aÞ; FIG. 1.—Nitrogen (d15N, &) and carbon (d13C, &) stable isotope composition of the species of plants included in diet reconstruction of individual Egyptian fruit bats (Rousettus aegyptiacus) in Haifa, Israel. All values represent the analysis of a single sample obtained from the homogenous mixture of at least 6 individuals for each species. Plants were separated into 2 groups that differed in mean d15N values: 15 6 SE; d15N ¼ 4.9 6 0.4&) and 15N-enriched (7.8 6 N-depleted (X 0.5&) species. All samples are fruits, except where indicated otherwise (L ¼ leaves). Plants were selected according to their presence in a previous diet study of this bat (Korine et al. 1999) and considered for diet reconstruction.

silver foil. N and C were cryogenically purified from the combustion products in a vacuum system. Purified N and C were analyzed in a PRISM micromass spectrophotometer (VG Isogas Ltd., Manchester, United Kingdom). There is no information on the turnover rate of N in bat muscle tissue, but C analysis of rodent muscle reflects the diet of the animals in the previous 1–2 months (Tiezsen et al. 1983). Half-life of N in whole blood of a 10-g phyllostomid bat is ;25 days (Miroń et al. 2006), but turnover rate in pteropodid bats in protein-based tissues is likely to be slower because of their larger body mass and may also result from their low maintenance N requirements (Korine et al. 1996). We assumed that N analysis of pteropodid muscle reflects the integrated diet of the animal in the previous 6–8 weeks. Stable-isotope ratios of C and N were expressed in dnotation as parts per thousand deviations from the Peedee belemnite marine limestone (C) or air (N) according to the equation: dX ¼ ½ðRsample =Rstandard Þ 1 1; 000;

ð1Þ

where X is the element of interest and R is the corresponding ratio 15N:14N or 13C:12C. Standard error was 60.1& for both elements. A similar procedure was followed with plant samples, except that we used 20 mg of plant tissue per sample. Fruit pulp and leaves were oven-dried at 608C to constant mass. Ground material from at least 6 individuals was homogenously mixed to form a single sample for each species of plant.

ð2Þ

where Pp is the proportion of assimilated N derived from N-depleted plants, 1 Pp is the proportion of assimilated N derived from 15N-enriched plants, and a is the diet–tissue isotopic discrimination factor appropriate for a plant diet. We interpreted these values qualitatively as individuals that derived protein predominantly from 15N-depleted plants (Pp ¼ 0.6–1), predominantly from 15N-enriched plants (Pp ¼ 0–0.4), or equally from both sets of plants (Pp ¼ 0.39–0.59). We also estimated the importance of each species of food sources using the Moore–Penrose pseudoinverse matrix (HallAspland et al. 2005). There are 2 methodological alternatives when the number of food sources exceeds the number of elements by more than 1: the Moore–Penrose and the IsoSource methods (Phillips and Gregg 2003). In contrast to IsoSource, which produces a distribution of all feasible combinations that adequately predict the observed consumer isotopic signature, the Moore–Penrose matrix produces a specific solution to the contribution of each food source (HallAspland et al. 2005). We constructed Moore–Penrose pseudoinverse matrices (Dþ) for each season using d15N values of food sources. We then obtained relative fractions for the use of each food source by multiplying Dþ by the d15N values of individual bats. We also used the IsoSource method but obtained equivocal results for most individuals, even after combining food sources with similar isotopic values. Therefore, we do not present the results of this method. Diet reconstruction requires that source d15N values were corrected by an appropriate tissue-diet discrimination factor before the analysis. Because no estimates of 15N isotopic discrimination are available for pteropodid bats, we used the 6 SE ¼ 3.6 6 0.4&) from 2 previous average value (X independent studies with a species of nectarivorous phylllostomid bat. We averaged the value obtained from bats fed 1.3% N (3.1& and 4.4&—Miroń et al. 2006; Voigt and Matt 2004) and 2.2% N (3.3&—Miroń et al. 2006) diets. In the 3 diets, the N source was of plant origin, and plants had N contents similar to the fruit eaten by pteropodid bats in this study. Data analysis.— We tested the prediction that intrapopulation variation in the use of protein sources would be higher in spring than in summer and winter using a square rank test of 15

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homogeneity of d15N variance (except fall because of a small sample size—StatSoft, Inc. 1995). We used a Kruskal–Wallis test to test for significant intrapopulation variation in use of protein sources within each season (except fall) using values derived from Moore–Penrose analysis. P , 0.05 was chosen as the lowest acceptable level of significance.

TABLE 1.—Nitrogen (d15N, &) and carbon (d13C, &) stable isotope composition of individual Egyptian fruit bats (Rousettus aegyptiacus) in northern Israel. Individuals are numbered in each season for further identification in Table 2 and Fig. 2. A dash (—) indicates a missing sample. Season Winter

RESULTS Bat stable isotope composition.— There was intrapopulation variation in bat d15N values in each of the seasons examined (with the exception of fall, in which only 2 samples were analyzed; Table 1). In winter, the difference between extreme d15N values was 2.3&, in spring it was 7.8&, and in summer it was 1.7&. Accordingly, there were significant differences in variance of bat d15N values among seasons (square rank test; v2 ¼ 13.9, d.f. ¼ 2, P , 0.001); variance was higher in spring (10.9) than in summer (0.54; P , 0.001) and winter (0.79; P , 0.001) but it did not differ between winter and summer (P ¼ 0.8). In particular, the d15N values of 3 individuals in spring were exceptionally high compared to the rest of the bats and to the highest plant d15N value. Diet reconstruction.— Diet reconstruction suggests that individuals used different sets of plants within the population (Fig. 2). In winter, 2 individuals (4 and 5) relied predominantly (Pp ¼ 0.65–0.85) on 15N-depleted plants (fruits of C. siliqua and D. kaki and leaves of F. religiosa), 2 individuals (1 and 3) used predominantly (Pp ¼ 0.28–0.40) 15N-enriched fruits (M. azedarach and F. microcarpa), and 1 individual (2) used both sets of plants equally (Pp ¼ 0.57). In spring, 3 individuals (2, 3, and 4) relied predominantly (Pp ¼ 0–0.13) on 15N-enriched fruits (M. azedarach, E. japonica, F. sycomorus, and F. microcarpa), 1 individual (1) used both sets of plants equally (Pp ¼ 0.65), and 3 individuals (5, 6, and 7) had d15N values that fell well outside the range of values of the food sources considered in the study, suggesting that they were feeding on other items. In summer, 4 individuals (1, 3, 5, and 6) relied predominantly (Pp ¼ 0–0.05) on 15N-enriched fruits (F. sycomorus and F. microcarpa), 1 individual (7) relied predominantly (Pp ¼ 0.69) on 15N-depleted fruits (M. nigra, P. dactylifera, and F. religiosa), and 2 (2 and 4) individuals used both subsets of plants to the same extent (Pp ¼ 0.52). In fall, both individuals examined relied predominantly (Pp ¼ 0.03–0.23) on 15N-enriched fruits (M. azedarach, F. sycomorus, and F. microcarpa). Interindividual variation in the use of protein sources did not differ significantly between individual bats during winter (P . 0.96) and summer (P . 0.82) but differed significantly during spring (Kruskal–Wallis, H ¼ 32.92, d.f. ¼ 6, P , 0.001).

DISCUSSION Previous studies on feeding habitats of pteropodids have demonstrated that some populations have large dietary breadths (Banack 1998; Bhat 1994; Bumrunsgri et al. 2007; Korine et al. 1999; Nelson et al. 2000; Nyhagen et al. 2005; Ritcher and

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Individual

d15N

d13C

1 2 3 4 5 6

10.6 9.9 11.1 8.8 9.6 — 10.0 6 0.4

22.7 24 23.1 24.5 24.6 23.7 23.7 6 0.3

1 2 3 4 5 6 7

9.7 11.5 11.0 11.3 16.2 17.5 17.1 13.4 6 1.2

23.9 23.8 24.7 24.2 23.7 23.6 24.6 24.1 6 0.1

1 2 3 4 5 6 7

11.2 9.9 11.2 9.9 11.1 11.0 9.5 10.5 6 0.3

23.8 19.5 22.9 23.0 23.6 — 23.3 22.6 6 0.6

1 2 3 4 5

10.8 11.5 — — —

24.5 23.9 23.7 24.1 23.1 23.8 6 0.5

6 SE X Spring

6 SE X Summer

6 SE X Fall

6 SE X

Cumming 2006; Stier and Mildestein 2005). Although methodological constraints in these studies do not allow examining long-term intrapopulation variation in diet, in some cases intrapopulation variation was suspected. For example, individuals of Pteropus tonganus and P. samoensis focus on preferred resources but also opportunistically include other items while foraging (Banack 1998). We used stable isotope analysis as a methodological alternative to study long-term individual feeding strategies in bats. We found that d15N values varied substantially in Egyptian fruit bats (8.8–17.5&). In support of our hypothesis, intrapopulation variance in d15N values was higher in spring than in summer and winter, suggesting that individuals during this period varied more in their use of protein sources. Most of the d15N range of values (8.8–11.5&) was in parallel with the range found in plants (3.4–9.3&). However, there were 3 bats that showed highly enriched d15N values (16.2–17.5&), suggesting that we probably missed dietary items with exceptionally high d15N values. Dietary reconstruction suggests that individuals within the bat population we studied do not have the same long-term dietary habits. Instead, there seem to be groups of individuals

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FIG. 2.—Origin of dietary nitrogen in Egyptian fruit bats throughout the year in Haifa, Israel. A 2-end-point mixing model (Fleming 1995) was used to estimate the relative contribution of d15N-depleted and d15N-enriched plants. A Moore–Penrose pseudoinverse matrix (Hall-Aspland et al. 2005) was used to estimate the relative contribution of each species of plant that could be used as food by the bats (Korine et al. 1999). All plants are fruits except Ficus religiosa in winter and spring (leaves), and in fall (leaves and fruits).

that used different subsets of food sources throughout the year: 2 groups of bats in winter, 3 groups in spring, and 3 groups in summer. In addition, our results also may indicate individual differences in the use of foraging areas because different plant species might have different isotopic composition at the landscape scale. It is important to point that we only examined adult males but our approach also could be used in the future to evaluate intrapopulation differences related to age, sex, or reproductive status of the bats. Although limited by small sample sizes, this study revealed long-term intrapopulation variation in the origin of dietary protein not attainable by conventional methods regularly used to study bats. Diet reconstruction is sensitive to variations in trophic fractionation (Gaye-Siessegger et al. 2004), and one should be cautious when interpreting it. For example, when we used the lowest (3.1&—Voigt and Matt 2004) and highest (4.4&—Miroń et al. 2006) 15N isotopic discrimination values reported for bats, the estimates of relative assimilated protein from 15N-depleted and 15N-enriched plants differed from the values obtained using the average discrimination factor (Table 2). However, the qualitative interpretation of our results was not affected in most cases. For example, when the highest discrimination value was used, 1 individual (individual 1 in winter) changed from predominantly feeding on 15N-enriched plants to predominantly feeding on 15N-depleted plants, and 6 individuals changed from feeding equally on both sets of plants to feeding predominantly on a particular set (or vice versa;

individuals 2 and 3 in winter, individual 1 in spring, individuals 2 and 4 in summer, and individual 1 in fall). In the remaining 14 individuals the qualitative interpretation did not change. The main message of our study is that there is long-term intrapopulation variation in the use of plant food items in Egyptian fruit bats. Not all bats are eating the same set of plant species. One consequence of this is that different subsets of plant species may be dispersed by different groups of bats rather than by the population as a whole. In other words, there seems to be intrapopulation partitioning among Egyptian fruit bats in the use of plant food items, and interspecific partitioning among plants in their dependence on dispersal by bats. To our knowledge, this is the 1st study to use stable-isotope analysis to reconstruct dietary patterns in pteropodid bats, and we hope that it will stimulate the use of this approach in future studies. In contrast to previous dietary studies of pteropodid bats that determined the diet of an entire population, our approach showed that individuals may differ in their use of food items, presumably equally available for the whole population. For some individuals it was not possible to reconstruct diet, which suggests that they were feeding on items that were highly enriched in 15N. One possibility points toward the consumption of insects because their tissues should be more enriched in this isotope than plants. Accordingly, hypothetical d15N values of insects feeding on the plant material with the average d15N value found in our study (6.1&) would be 9.7& (if we used a 3.6& enrichment factor). The

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TABLE 2.—Ratio of assimilated nitrogen derived from 15N-depleted plants (Pp) in individuals of the Egyptian fruit bat using different blood–diet N discrimination factors (diet-consumer—Miroń et al. 2006; Voigt and Matt 2004). Pp values were qualitatively interpreted as nitrogen derived predominantly from 15N-depleted plants (Pp ¼ 0.6–1), predominantly from 15N-enriched plants (Pp ¼ 0–0.4), or equally from both sets of plants (Pp ¼ 0.39–0.59). Negative values indicate that it was not possible to trace the origin of assimilated nitrogen. Diet-consumer (&) Season

Individual

3.6

4.4

3.1

Winter

1 2 3 4 5

0.40 0.57 0.27 0.85 0.65

0.60 0.77 0.48 1.00 0.85

0.27 0.45 0.15 0.72 0.52

Spring

1 2 3 4 5 6 7

0.56 0 0.13 0.03 1.60 2.03 1.90

0.83 0.23 0.40 0.30 1.33 1.76 1.63

0.40 0.20 0 0.13 1.76 2.20 2.06

Summer

1 2 3 4 5 6 7

0 0.52 0 0.52 0 0.40 0.69

0.28 0.84 0.28 0.84 0.32 0.35 1.00

0 0.28 0.28 0.28 0.23 0.19 0.45

Fall

1 2

0.23 0.03

0.47 0.25

0.08 0.26

d15N value of bats feeding exclusively on these insects (13.3&) would still be far below the highest bat values found in our study. A 2nd possibility could be that these bats fed on exceptionally 15N-enriched plants. For example, commercially grown fruits may be enriched in 15N because agricultural soils have higher d15N values than forest soils due to higher rates of ammonia volatilization (Nadelhoffer and Fry 1994) and biological activity (Mariotti et al. 1981). Moreover, long-term use of nitrogen fertilizers increases the d15N value of soil N (Ho¨gber 1990). Differences in soil N stable isotope composition between cultivated and forest biomes are ultimately reflected in the composition of the food chain that they support. Egyptian fruit bats are known to include commercially grown fruits in their diet (Moran and Keidar 1993). However, the d15N values reported in this study for D. kaki and E. japonica are not particularly enriched in 15N, although it is likely that cultivated fruits exhibit geographic isotopic variation reflecting differences in local agricultural practices.

ACKNOWLEDGMENTS The Technion-Institute of Technology supported this study. We thank A. A. Nelson, L. Sternberg, and S. Goldenberg for technical assistance in the laboratory, and S. A. Hall-Aspland for help with the Moore–Penrose inverse matrix model. Two anonymous reviewers and C. Voigt helped improve the manuscript.

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LITERATURE CITED AMIRAN, D. H. K., J. ELSTER, M. GILEAD, N. ROSEMAM, N. KADMON, AND U. PARAN. 1970. Atlas of Israel. Elsevier, Jerusalem, Israel. BANACK, S. A. 1998. Diet selection and resource use by flying foxes (genus Pteropus). Ecology 79:1949–1967. BARCLAY, R. M., L. E. BARCLAY, AND D. S. JACOBS. 2006. Deliberate insectivory by the fruit bat Rousettus aegyptiacus. Acta Chiropterologica 8:549–553. BHAT, H. R. 1994. Observations on the food and feeding behavior of Cynopterus sphinx Vahl (Chiroptera, Pteropodidae) at Pune, India. Mammalia 58:363–370. BOLNICK, D. I., R. SVANBACK, J. A. FORDYCE, L. H. YANG, AND J. M. DAVIS. 2003. The ecology of individuals: incidence and importance of individual specialization. American Naturalist 161:1–28. BOLNICK, D. I., L. H. YANG, J. A. FORDYCE, J. M. DAVIS, AND R. SVANBACK. 2002. Measuring individual-level resource specialization. Ecology 83:2936–2941. BUMRUNSGRI, S., W. LEELAPAIBUL, AND P. A. RACEY. 2007. Resource partitioning in sympatric Cynopterus bats in lowland tropical rainforest, Thailand. Biotropica 39:241–248. CEBALLOS, G., T. H. FLEMING, C. CHA´VEZ, AND J. NASSAR. 1997. Population dynamics of Leptonycteris curasoae (Chiroptera: Phyllostomidae) in Jalisco, Mexico. Journal of Mammalogy 78:1220–1230. COURTS, S. E. 1998. Dietary strategies of Old World fruit bats (Megachiroptera: Pteropodidae). How do they obtain sufficient protein? Mammalogy Review 28:185–194. DES MARAIS, D. J., J. M. MITCHELL, W. G. MEINSCHEIN, AND J. M. HAYES. 1980. The carbon isotope biogeochemistry of the individual hydrocarbons in bat guano and the ecology of insectivorous bats in the region of Carlsbad, New Mexico. Geochimica et Cosmochimica Acta 44:2075–2086. EHLERINGER, J. R., P. W. RUNDEL, AND K. A. NAGY. 1986. Stable isotopes in physiological ecology and food web research. Trends in Ecology and Evolution 1:42–45. ESTES, J. A., M. L. RIEDMAN, M. M. STAEDLER, M. T. TINKER, AND B. E. LYON. 2003. Individual variation in prey selection by sea otters: patterns, causes and implications. Journal of Animal Ecology 72:144–155. FLEMING, T. H. 1988. The short-tailed fruit bat. University of Chicago Press, Chicago, Illinois. FLEMING, T. H. 1995. The use of stable isotopes to study the diets of plant-visiting bats. Pp. 99–110 in Ecology, evolution and behaviour of bats (P. A. Racey and S. M. Swift, eds.). Zoological Society of London, Claredon Press, Oxford, United Kingdom. FLEMING, T. H., R. A. NUN˜EZ, AND L. S. L. STERNBERG. 1993. Seasonal changes in the diets of migrant and non-migrant nectarivorous bats as revealed by carbon stable isotope analysis. Oecologia 94:72–75. FOWLER, M. E. 1993. Zoo and wild animal medicine: current therapy 3. W. B. Saunders & Co., Philadelphia, Pennsylvania. GANNON, W. L., R. S. SIKES, AND THE ANIMAL CARE AND USE COMMITTEE OF THE AMERICAN SOCIETY OF MAMMALOGISTS. 2007. Guidelines of the American Society of Mammalogists for the use of wild mammals in research. Journal of Mammalogy 88:809–823. GAYE-SIESSEGGER, J., U. FOCKEN, S. MUETZEL, H. ABEL, AND K. BECKER. 2004. Feeding level and individual metabolic rate affect d13C and d15N values in carp: implications for food web studies. Oecologia 138:175–183. GU, B., C. L. SCHELSKE, AND M. V. HOYER. 1997. Feeding diversity in blue tilapia: evidence from stable-isotope analysis. Ecology 78:2263–2266.

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HALL-ASPLAND, S. A., A. P. HALL, AND T. L. ROGERS. 2005. A new approach to the solution of the linear mixing model for a single isotope: application to the case of an opportunistic predator. Oecologia 143:143–147. HERRERA, L. G., B. ALTUBE, W. DÍAZ, E. GUTIERREZ, K. A. HOBSON, AND V. SAŃCHEZ-CORDERO. 2002. Sources of assimilated protein in five species of New World frugivorous bats. Oecologia 133:280–287. HERRERA, L. G., T. H. FLEMING, AND L. S. STERNBERG. 1998. Trophic relationships in a neotropical bat community: a preliminary study using carbon and nitrogen isotopic signatures. Journal of Tropical Ecology 39:23–29. HERRERA, L. G., K. A. HOBSON, D. ESTRADA, A. MANZO, G. MEŃDEZ, ´ NCHEZ-CORDERO. 2001a. The role of fruits and insects in AND V. SA the nutrition of frugivorous bats: evaluating the use of stable isotope models. Biotropica 33:520–528. HERRERA, L. G., K. A. HOBSON, L. MIROŃ, N. RAMI´REZ, G. MEŃDEZ, ´ NCHEZ-CORDERO. 2001b. Sources of protein in two species AND V. SA of phytophagous bats in a seasonal dry forest: evidence from stableisotope analysis. Journal of Mammalogy 82:352–361. HOBSON, K. A. 1999. Tracing origins and migration of wildlife using stable isotopes: a review. Oecologia 120:314–326. HOBSON, K. A., AND I. STIRLING. 1997. Terrestrial foraging by polar bears during the ice free period in western Hudson Bay: metabolic pathways and limitations of the stable isotope approach. Marine Mammal Science 13:359–367. HO¨GBER, P. 1990. Forest losing large quantities of nitrogen has elevated 15N:14N ratios. Oecologia 84:229–231. HOLMES, D. 1987. Clinical laboratory animal medicine: an introduction. Iowa State University Press, Ames. KORINE, C., Z. ARAD, AND A. ARIELI. 1996. Nitrogen and energy balance of the fruit-bat, Rousettus aegyptiacus, on natural fruit diets. Physiological Zoology 69:618–634. KORINE, C., I. IZHAKI, AND Z. ARAD. 1998. Comparison of fruit syndromes between the Egyptian fruit-bat (Rousettus aegyptiacus) and birds in east Mediterranean habitats. Acta Oecologia 19:147–153. KORINE, C., I. IZHAKI, AND Z. ARAD. 1999. Is the Egyptian fruit bat Rousettus aegyptiacus a pest in Israel? An analysis of the bat’s diet and implications for its conservation. Biological Conservation 88:301–306. LAJTHA, K., AND R. H. MICHENER. 1994. Stable isotopes in ecology and environmental science. Blackwell Scientific Publications, Oxford, United Kingdom. MARIOTTI, A., ET AL. 1981. Experimental determination of nitrogen kinetic isotope fractionations: some principles; illustration for the denitrification and nitrification processes. Plant and Soil 62:413–430. MIROŃ, L. L. M., L. G. HERRERA M., N. RAMI´REZ P., AND K. A. HOBSON. 2006. Effect of diet quality on carbon and nitrogen turnover and isotopic discrimination in blood of a New World nectarivorous bat. Journal of Experimental Biology 209:541–548. MORAN, S., AND H. KEIDAR. 1993. Checklist of vertebrate damage to agriculture in Israel. Crop Protection 12:171–182. NADELHOFFER, K. J., AND B. FRY. 1994. Nitrogen isotope studies in forest ecosystems. Pp. 22–44 in Stable isotopes in ecology and

Vol. 89, No. 5

environmental science (K. Lajtha and R. H. Michener, eds.). Blackwell Scientific Publications, Oxford, United Kingdom. NASSAR, J. M., H. BECK, L. S. L. STERNBERG, AND T. H. FLEMING. 2003. Dependence on cacti and agaves in nectar-feeding bats from Venezuelan arid zones. Journal of Mammalogy 84:106–116. NELSON, S. L., M. A. MILLER, E. J. HESKE, AND G. C. FAHEY, JR. 2000. Nutritional consequences of a change in diet from native to agricultural fruits for the Samoan fruit bat. Ecography 23:393–401. NYHAGEN, D. F., S. D. TURNBULL, J. M. OLESEN, AND C. G JONES. 2005. An investigation into the role of the Mauritian flying fox, Pteropus niger, in forest regeneration. Biological Conservation 122:491–497. PHILLIPS, D. L. 2001. Mixing models in analyses of diet using multiple stable isotopes: a critique. Oecologia 127:166–170. PHILLIPS, D. L., AND J. W. GREGG. 2003. Source partitioning using stable isotopes: coping with too many sources. Oecologia 127:166– 170. PODLESAK, D. W., AND S. R. MCWILLIAMS. 2006. Metabolic routing of dietary nutrients in birds: effects of diet quality and macronutrient composition revealed using stable isotopes. Physiological Biochemistry and Zoology 79:534–549. RITCHER, H. V., AND G. S. CUMMING. 2006. Food availability and annual migration of the straw-colored fruit bat (Eidolon helvum). Journal of Zoology (London) 268:35–44. SCHMIDT, O. 1999. Intrapopulation variation in carbon and nitrogen stable isotope ratios in the earthworm Aporrectodea longa. Ecological Research 14:317–328. SEALY, J. C., N. J. VAN DER MERWE, J. A. L. THORP, AND J. L. LANHAM. 1987. Nitrogen isotopic ecology in southern Africa: implications for environmental and dietary tracing. Geochimica et Cosmochimica Acta 51:2707–2717. STATSOFT, INC. 1995. STATISTICA for Windows. StatSoft, Inc., Tulsa, Oklahoma. STIER, S. C., AND T. L. MILDESTEIN. 2005. Dietary habits of the world’s largest bats: the Philippine flying foxes, Acerodon jubatus and Pteropus vampyrus lanensis. Journal of Mammalogy 86:719–728. TIESZEN, L. L., T. W. BOUTTON, K. G. TESDAHL, AND N. A. SLADE. 1983. Fractionation and turnover of stable carbon isotopes in animal tissues: implications for d13C analysis of diet. Oecologia 57:32–37. URTON, E. J. M., AND K. A. HOBSON. 2005. Intrapopulation variation in gray wolf isotope profiles: implications for the ecology of individuals. Oecologia 145:316–325. VOIGT, C. C., AND D. H. KELM. 2006. Host preference of the common vampire bat (Desmodus rotundus; Chiroptera) assessed by stable isotopes. Journal of Mammalogy 87:1–6. VOIGT, C. C., AND F. MATT. 2004. Nitrogen stress causes unpredictable enrichments of 15N in two bat species. Journal of Experimental Biology 207:1741–1748. WHITE, R. C. T. 1993. The inadequate environment. Nitrogen and the abundance of animals. Springer-Verlag, Berlin, Germany.

Submitted 17 August 2007. Accepted 19 March 2008. Associate Editor was Christian C. Voigt.