Southeastern Naturalist B.B. Gregory, J.O. Whitaker, Jr., and G.D. Hartman 2014 Vol. 13, No. 4 762 2014 SOUTHEASTERN NATURALIST 13(4):762–769 Diet of Rafinesque’s Big-eared Bat (Corynorhinus rafinesquii) in West-central Louisiana Beau B. Gregory¹,*, John O. Whitaker, Jr.², and Gregory D. Hartman³ Abstract - We investigated the diet of Corynorhinus rafinesquii (Rafinesque’s Big-eared Bat) in west-central Louisiana by examining fecal pellets collected from beneath 3 bridges used by these bats as day roosts. Fresh fecal material was found under the bridges during every month of the year. We detected 5 insect orders, including 5 families, in fecal pellets collected from 25 August 2005 to 5 January 2007. Lepidoptera represented 93.7% of the total volume and was the only order observed in 100% of our samples. Coleopterans, mostly Scarabaeidae, were the second most abundant food item and represented 5.8% of the total volume. Hemiptera, Diptera, and Hymenoptera together represented 0.4 % of the total volume. We observed Diptera, Hemiptera, Hymenoptera, and Coleoptera in fecal pellets collected under some, but not all bridges. No insect orders were observed that had not previously been reported as prey of Rafinesque’s Big-eared Bats. Our results were similar to those reported in studies conducted in Kentucky, North Carolina, and Florida, and we concluded that Rafinesque’s Big-eared Bats primarily prey upon lepidopterans in Louisiana. Introduction Corynorhinus rafinesquii (Lesson) (Rafinesque’s Big-eared Bat) occurs in much of the eastern US and is considered to be a moth specialist (Hurst and Lacki 1997, Johnson and Lacki 2013, Lacki and Dodd 2010, Whitaker et al. 2007). This bat currently is considered a species of concern in every state where it occurs (Bayless et al. 2011). There are no published studies on the diet of these bats within the western portion of their geographic range. Because life-history information can aid management efforts, we report foods detected in feces of Rafinesque’s Big-eared Bats in Louisiana, near the western extent of the species’ range. Study Area We conducted our study within a 500-km2 area of Vernon Parish in west-central Louisiana within the boundaries of Fort Polk Military Reservation and neighboring Vernon Unit of the Kisatchie National Forest, Leesville, LA (Fig. 1). The area was characterized by gently rolling hills dominated by managed stands of Pinus palustris Mill. (Longleaf Pine), Pinus taeda L. (Loblolly Pine), and Pinus elliottii Engelm. (Slash Pine). Deciduous trees including Quercus spp. (oaks), Nyssa spp. (black gums), and Taxodium distichum (L.) Rich. (Baldcypress) dominated riparian areas (Carrie et al. 2002, Lance et al. 2001). ¹Coastal and Nongame Resources Division, Louisiana Department of Wildlife and Fisheries, Baton Rouge, LA 70808. ²Department of Biology, Indiana State University, Terre Haute, IN 47809. ³Department of Biology, Gordon State College, Barnesville, GA 30204. *Corresponding author - bgregory@wlf.la.gov. Manuscript Editor: Renn Tumlison Southeastern Naturalist 763 B.B. Gregory, J.O. Whitaker, Jr., and G.D. Hartman 2014 Vol. 13, No. 4 We collected fecal pellets under 3 T-beam girder bridges hereafter referred to as bridges I, II, and III. Bridges I and II were on paved roads and bridge III, a gravel road. The distances between bridges I and II, I and III, and II and III were 12.5, 15, and 5 km, respectively. Riparian habitats within a 0.25-km radius of bridges I and II were undeveloped forest, and there was a wastewater treatment facility in a 10-ha clearing ~0.1 km from bridge III. Methods From 25 August 2005 to 5 January 2007, we collected 39 samples of fecal pellets from Big-eared Bats; 13 samples from under each of the 3 bridges. All months of the year were represented by at least 1 sample from each of the 3 bridges; however, samples corresponding to a given month were not always collected on the same date. Months represented twice in our dataset were August 2005 and 2006 for bridges I and II, and September 2005 and 2006 for bridge III. The number of Rafinesque’s Big-eared Bats present during sampling events was variable; ranges = 1–9, 0–26, and 0–23 bats for bridges I, II, and III, respectively. Fecal pellets were collected with forceps, deposited in 7-ml glass vials, and stored at room temperature until we analyzed the samples. We assumed that fecal Figure 1. Location of study area. Southeastern Naturalist B.B. Gregory, J.O. Whitaker, Jr., and G.D. Hartman 2014 Vol. 13, No. 4 764 pellets adhered to the concrete roosting surface were the most fresh and collected them first; pellets located on the ground directly below roost sites were collected next. Eptesicus fuscus (Beauvois) (Big Brown Bat), Myotis austroriparius (Rhoads) (Southeastern Myotis), and Perimyotis subflavus (Cuvier) (Eastern Pipistrelle) also occurred at the roosts, which could complicate collection of fecal material. However, fecal pellets of Rafinesque’s Big-eared Bats typically are distinctively helical (Hurst and Lacki 1997), which made it possible to distinguish Rafinesque’s Big-eared Bat pellets from those of other species. All samples consisted of multiple fecal pellets. After we collected fecal pellets, we cleared the roosting surface and substrate at the collection site of any remaining feces and foreign debris to provide a clean surface for feces to accumulate in the future. Because collection sites were sometimes flooded by the creeks running under the bridges, use of fecal-pellet collection sheets was not practical. Substrate under the bridges predominately was loose sandy soil, allowing collection sites to be easily scraped clean using a piece of lumber. We covered fecal pellets with ethanol in a Petri dish and examined them using a 10–70-power zoom dissecting microscope (Olympus America, SZH, Melville, NY). We identified food items to the lowest taxonomic level possible and visually estimated the percent volume of each food item for each pellet. Insect remains had been chewed into pieces and thoroughly mixed; thus, it was not possible to measure their volume directly or by water displacement. Data were summarized as percent volume (sum of volumes in individual pellets for each item/sum of total volume x 100; see Whitaker 1988), which indicates the relative amount of each type of prey in a sample. Percent frequency was recorded as the percentage of samples in which a given food occurred. There were many particles that we could not identify; these were assumed to be in similar proportion to identified foods. All fecal pellet analysis was conducted by J.O. Whitaker, Jr. Results We found fresh fecal pellets during every month sampling was conducted, but they were more abundant during warmer months. We identified representatives of 5 orders of insects (Table 1). Lepidoptera occurred in 100% of the samples, represented 93.7% of the total volume, and was the only order detected in 19 of our 39 samples. The total volume attributable to Lepidoptera ranged from 88.9 to 99.2% in the combined samples from each of the 3 roost sites (Table 1). Coleopterans were the second most abundant food item and represented 5.7% of the total volume; the families Scarabaeidae and Carabidae represented 4.0% and 0.5% of the total volume, respectively. Hemiptera, Diptera, and Hymenoptera were found in 7 samples and together they accounted for 0.4% of the total volume. A small portion (0.1% total volume) of the material was unidentifiable insect remains. Among the 39 samples from the 3 bridges, the total volume attributable to Coleoptera ranged from 0.7 to 10.1%. We detected Coleoptera in 15 samples (43.6%), with the family Scarabaeidae representing from 40 to 47% of the total volume of 3 samples. Scarabaeidae and Diptera were detected only in samples from bridges I Southeastern Naturalist 765 B.B. Gregory, J.O. Whitaker, Jr., and G.D. Hartman 2014 Vol. 13, No. 4 and II, and we observed Hemiptera and Hymenoptera only in fecal pellets collected at bridge II. Discussion In west-central Louisiana, Rafinesque’s Big-eared Bats primarily consume lepidopterans. Our findings were similar to those reported in other studies where fecal pellets were analyzed near the northern extent of the species’ range in southeastern Kentucky (Hurst and Lacki 1997, Johnson and Lacki 2013) and in the southeastern extent of the range in central Florida (Whitaker et al. 2007). West-central Louisiana is near the southwestern edge of the known range of Rafinesque’s Big-eared Bat. Our findings further support earlier observations of a high percentage of moths in the diet of members of the genus Corynorhinus (Clark 1991; Hurst and Lacki 1997; Lacki and Dodd 2010; Ober and Hayes 2008; Sample and Whitmore 1993; Whitaker et al. 1977, 1981, 2007). Dietary percentage volumes reported in our study and the Kentucky and Florida investigations were similar (Table 2). However, though trace amounts of Trichoptera were recorded by Hurst and Lacki (1997), and small amounts of Trichoptera and Orthoptera were reported by Whitaker et al. (2007), we did not observe trichopteran or orthopteran remains in any fecal pellets we analyzed. Arachnids and vegetation also were present in the fecal pellets of Rafinesque’s Big-eared Bats from Florida, but these taxa were not detected in fecal pellets from Kentucky or Louisiana. Seasonal differences in sampling regime, regional variation in prey availability, and differences in the numbers of fecal pellets Table 1. Mean percentage volume and frequency of insects observed from fecal pellets of Rafinesque’s Big-eared Bats collected throughout the year at 3 bridge roosts in Louisiana. Some of the columns with values for volume in the table do not sum to exactly 100 percent due to rounding. All = All roosts combined. Bridge I Bridge II Bridge III All Taxon Volume Freq. Volume Freq. Volume Freq. Volume Freq. Lepidoptera 93.2 100.0 88.9 100.0 99.2 100.0 93.7 100.0 Coleoptera Unknown Coleoptera 2.0 23.1 1.1 30.8 0.5 15.4 1.2 23.1 Scarabaeidae 4.2 15.4 7.9 23.1 0.0 0.0 4.0 12.8 Carabidae 0.2 7.7 1.2 7.7 0.2 7.7 0.5 7.7 Hemiptera Unknown Hemiptera 0.0 0.0 0.5 15.4 0.0 0.0 0.2 5.1 Cercopidae 0.0 0.0 0.2 7.7 0.0 0.0 0.1 2.6 Diptera Unknown Diptera 0.2 15.4 0.0 0.0 0.0 0.0 0.1 5.1 Tipulidae 0.0 0.0 0.2 2.6 0.0 0.0 0.1 0.9 Hymenoptera Formicidae 0.0 0.0 0.1 2.6 0.0 0.0 0.03 0.9 Unidentified insect 0.2 7.7 0.0 0.00 0.2 7.7 0.1 5.1 Southeastern Naturalist B.B. Gregory, J.O. Whitaker, Jr., and G.D. Hartman 2014 Vol. 13, No. 4 766 analyzed may account for some of the differences among the findings reported in the 3 studies (Table 2). We observed Diptera, Scarabaeidae, Hemiptera, and Hymenoptera in fecal pellets from some but not all of our sampling sites. Hurst and Lacki (1997) reported a similar pattern for Homoptera, Hemiptera, Hymenoptera, and Trichoptera. The availability of moths (Dodd et al. 2008, Johnson and Lacki 2013) and other Table 2. Comparison of percentage volume and frequency of insects observed from fecal pellets of Rafinesque’s Big-eared Bats in Louisiana with results reported in Florida and Kentucky. Some of the columns with values for volume do not sum to exactly 100 percent due to rounding. Blank cell = not reported or not detected, * = this study, ** = Whitaker et al. 2007, *** = Hurst and Lacki 1997. Louisiana* Florida** Kentucky*** Mean Volume Freq. Volume Freq. Volume freq.A Taxon Lepidoptera 93.70 100.00 93.70 99.40 93.90 100.00 . Coleoptera (5.70) (43.60) (1.30) UnknownB 5.10 83.55 Unknown Coleoptera 1.20 23.10 0.80 2.30 Scarabaeidae 4.00 12.80 0.40 0.80 Carabidae 0.50 7.70 0.08 0.10 Circulionidae 0.02 0.20 Hemiptera (0.20) (5.10) (0.09) (0.50) 0.10 11.70 Unknown HemipteraC 0.20 5.10 0.09 0.50 Hemiptera (formerly Homoptera) (0.10) (2.60) (0.28) (0.60) 0.38 28.10 Cercopidae 0.10 2.60 Cicadellidae 0.08 0.50 Cicadidae 0.20 0.10 Diptera (0.20) (7.70) (2.96) UnknownB 0.23 16.90 Unknown Diptera 0.10 5.10 0.70 2.00 Calliphoridae 2.20 5.50 Tipulidae 0.10 2.60 0.03 0.10 Chironomidae 0.03 0.01 Hymenoptera (0.03) (0.90) (0.10) (0.30) 0.10 7.50 Formicidae 0.03 0.90 0.03 0.10 Ichneumonidae 0.07 0.30 Trichoptera 0.2 0.50 Trace 2.88 Orthoptera (0.04) (0.30) Gryllidae 0.04 0.30 Unidentified insect 0.10 5.10 0.80 2.10 0.23 13.38 Acarina Trace 0.20 Araneae 0.40 0.20 Vegetation 0.02 0.10 ATotal percentage frequency not reported, average of individual r oosts. BCannot be calculated from data provided. CMay include unidentifiable insects formerly included in the orde r Homoptera. Southeastern Naturalist 767 B.B. Gregory, J.O. Whitaker, Jr., and G.D. Hartman 2014 Vol. 13, No. 4 arthropods can vary with habitat, and Rafinesque’s Big-eared Bats forage in more than 1 type of habitat (Johnson and Lacki 2013). In our study, there appeared to be differences in the diet of bats roosting at different bridges; however, our samples were not all collected during the same day within a month, and we were unable to determine if the availability of prey varied across time and space. The remains of arthropods in the fecal pellets of bats usually can be identified only to the level of order or family. Moth-wing culls found beneath roosts of members of Corynorhinus have been used to identify lepidopteran prey of the bats at the level of genus or species, and to estimate the sizes of the prey (Burford and Lacki 1998, Hurst and Lacki 1997, Lacki and LaDeur 2001, Sample and Whitmore 1993, Whitaker et al. 2007). We encountered only 1 moth-wing cull during our study, and were not able to identify the species. In west-central Louisiana, at least some Rafinesque’s Big-eared Bats forage year-round. Bats in the genus Corynorhinus possess a suite of characteristics that allows them to be effective at gleaning (Lacki and Dodd 2010, and references therein), they are, however, capable of capturing moths during flight (Lacki and LaDeur 2001). Gleaning is a behavior that allows bats to forage later into the night and during cooler weather than is possible for those that forage primarily by aerial hawking (Barclay 1991, Swift 1998). We observed lesser amounts of guano at the roosting sites during cooler months; a pattern that could have been caused by reduced foraging activity or use of alternate roost sites during those times (Johnson et al. 2012, Jones 1977). Some moths in the families Noctuidae and Geometridae are known to fly during any month of the winter when air temperatures are at least 0 °C (Heinrich 1987, 1993), a behavioral capability that may result in reduced predation pressure on the moths by bats and birds (Heinrich 1993). Moths in these 2 families commonly are consumed by Rafinesque’s Big-eared Bats (Johnson and Lacki 2013). Foraging activity of Rafinesque’s Big-eared Bats in Kentucky was positively associated with distribution and availability of moths (Johnson and Lacki 2013), and bats may consume individuals of some moth species disproportionately to the moth species’ abundance (Lacki and LaDeur 2001). Because arthropod remains in fecal pellets of bats usually can be identified only to order or family, it has been difficult for researchers to rigorously test some hypotheses about the diet of Rafinesque’s Big-eared and other bats. DNA recovered from bat feces has been used to successfully identify prey items, often to the species level (Bohmann et al. 2011; Clare et al. 2009, 2011; Dodd et al. 2012). Use of visual analyses coupled with DNA-based techniques will help researchers better identify remains of prey items in fecal pellets, and allow testing of hypotheses about diet of Rafinesque’s Big-eared Bats as it relates to habitat, season, and the distribution and abundance of prey. Because Rafinesque’s Big-eared Bat is considered a species of concern throughout its range, it is important to obtain detailed knowledge of diet from different geographic locations. More data are needed to understand diet in relation to season, locality, and habitat type, and to know if any beneficial or pest lepidopteran species of economic importance are being consumed. Forest and Southeastern Naturalist B.B. Gregory, J.O. Whitaker, Jr., and G.D. Hartman 2014 Vol. 13, No. 4 768 land-management practices potentially affect the suitability of a habitat for roosting and foraging by bats (Hayes and Loeb 2007, Johnson and Lacki 2013). Knowledge of the diet of Rafinesque’s Big-eared Bat at a finer taxonomic scale is important for the development of land-management practices suitable to sustain populations of these animals. Acknowledgments We thank the Conservation Branch at Fort Polk Military Reservation, Vernon Parish, LA, for funding part of this study, and Fort Polk biologists for assistance in data collection. M.J. Bender critically reviewed the manuscript and provided numerous suggestions that helped to improve it. Literature Cited Barclay, R.M.R. 1991. Population structure of temperate zone insectivorous bats in relation to foraging behavior and energy demand. Journal of Animal Ecology 60:165–178. Bayless, M.L., M.K. Clark, R.C. Stark, B.S. Douglas, and S.M. Ginger. 2011. Distribution and status of eastern big-eared bats (Corynorhinus spp.). Pp. 13−25, In S.C. Loeb, M.J. Lacki, and D.A. Miller (Eds.). Conservation and Management of Eastern Big-eared Bats: A Symposium. US Department of Agriculture, Forest Service, Southern Research Station, Asheville, NC. General Technical Report SRS-145. 157 pp. Bohmann, K., A. Monadjem, C.L. Noer, M. Rasmussen, M.R.K. Zeale, E. Clare, G. Jones, E. Willerslev, and M.T.P. Gilbert. 2011. Molecular diet-analysis of two African freetailed bats (Molossidae) using high throughput sequencing. PLoS ONE 6(6):e21441. doi:10.1371/journal.pone.0021441 Burford, L.S., and M.J. Lacki. 1998. Moths consumed by Corynorhinus townsendii virginianus in eastern Kentucky. American Midland Naturalist 139:141–146. Carrie, N.R., R.O. Wagner, K.R. Moore, J.C. Sparks, E.L. Keith, and C.A. Melder. 2002. Winter abundance of and habitat use by Henslow’s Sparrows in Louisiana. Wilson Bulletin 114:221–226. Clare, E.L., E.E. Fraser, H.E. Braid, M.B. Fenton, and P.D.N. Hebert. 2009. Species on the menu of a generalist predator, the Eastern Red Bat (Lasiurus borealis): Using a molecular approach to detect arthropod prey. Molecular Ecology 18:2532–2542. Clare, E.L., B.R. Barber, B.W. Sweeney, P.D.N. Hebert, and M.B. Fenton. 2011. Eating local: Influences of habitat on the diet of Little Brown Bats (Myotis lucifugus). Molecular Ecology 20:1772–1780. Clark, M.K. 1991. Foraging ecology of Rafinesque’s Big-eared Bat, Plecotus rafinesquii, in North Carolina. Bat Research News 32:68. Dodd, L.E., M.J. Lacki, and L.K. Rieske. 2008. Variation in moth occurrence and implications for foraging habitat in Ozark Big-eared Bats. Forest Ecology and Management 255:3866–3872. Dodd, L.E., E.G. Chapman, J.D. Harwood, M.J. Lacki, and L.K. Rieske. 2012. Identification of prey of Myotis septentrionalis using DNA-based techniques. Journal of Mammalogy 93:1119–1128. Hayes, J.P., and S.C. Loeb. 2007. The influence of forest management on bats in North America. Pp. 207–236 In M.J. Lacki, J.P. Hayes, and A. Kurta (Eds.). Bats in Forests: Conservation and Management. Johns Hopkins University Press, Baltimore, MD. 329 pp. Southeastern Naturalist 769 B.B. Gregory, J.O. Whitaker, Jr., and G.D. Hartman 2014 Vol. 13, No. 4 Heinrich, B. 1987. Thermoregulation by winter-flying endothermic moths. The Journal of Experimental Biology 127:313–332. Heinrich, B. 1993. The Hot-blooded Insects: Strategies and Mechanisms of Thermoregulation. Harvard University Press, Cambridge, MA. 607 pp. Hurst, T.E., and M.J. Lacki. 1997. Food habits of Rafinesque’s Big-eared Bat in southeastern Kentucky. Journal of Mammalogy 78:525–528. Johnson, J.S., and M.J. Lacki. 2013. Habitat associations of Rafinesque’s Big-eared Bats (Corynorhinus rafinesquii) and their lepidopteran prey in bottomland hardwood forests. Canadian Journal of Zoology 91:94–101. Johnson, J.S., M.J. Lacki, S.C. Thomas, and J.F. Grider. 2012. Frequent arousals from winter torpor in Rafinesque’s Big-eared Bat (Corynorhinus rafinesquii). PLoS ONE 7(11):e49754. doi:10.1371/journal.pone.0049754. Jones, C. 1977. Plecotus rafinesquii. Mammalian Species 69:1–4. Lacki, M.J., and L.E. Dodd. 2010. Diet and foraging behavior of Corynorhinus in eastern North America). Pp. 39−52, In S.C. Loeb, M.J. Lacki, and D.A. Miller (Eds.). Conservation and Management of Eastern Big-eared Bats: A Symposium. US Department of Agriculture, Forest Service, Southern Research Station, Asheville, NC. General Technical Report SRS-145. 157 pp. Lacki, M.J., and K.M. LaDeur. 2001. Seasonal use of lepidopteran prey by Rafinesque’s Big-eared Bats (Corynorhinus rafinesquii). American Midland Naturalist 145:213–217. Lance, R.F., B.T. Hardcastle, A. Talley, and P.L. Leberg. 2001. Day-roost selection by Rafinesque’s Big-eared Bat (Corynorhinus rafinesquii) in Louisiana forests. Journal of Mammalogy 82:166–172. Ober, H.K., and J.P. Hayes. 2008. Prey selection by bats in forests of western Oregon. Journal of Mammalogy 89:1191–1200. Sample, B.E., and R.C. Whitmore. 1993. Food habits of the endangered Virginia Big-eared Bat in West Virginia. Journal of Mammalogy 74:428–435. Swift, S.M. 1998. Long-eared Bats. Cambridge University Press, Cambridge, UK. 182 pp. Whitaker, J.O., Jr. 1988. Food-habits analysis of insectivorous bats. Pp. 171–189 In T.H. Kunz (Ed.). Ecological and Behavioral Methods for the Study of Bats. Smithsonian Institution Press, Washington, DC. 533 pp. Whitaker, J.O., Jr., C. Maser, and L.E. Keller. 1977. Food habits of bats of western Oregon. Northwest Science 51:46–55. Whitaker, J.O., Jr., C. Maser, and S.P. Cross. 1981. Food habits of eastern Oregon bats, based on stomach and scat analyses. Northwest Science 55:281–292. Whitaker, J.O., Jr., M. Brooks, L. Scott, L.S. Finn, and C.L. Smith. 2007. Food habits of Rafinesque’s Big-eared Bat from Florida. Florida Scientist 70:202–206.