21 THE STOMACH CONTENTS OF SOME ANURAN TADPOLES FROM THAILAND Benjawan Moonasa1, Prapaiporn Thongproh1, Ekachai Phetcharat2, Wassana Kingwongsa3, Pramote Ratree4, Prateep Duengkae5, Theerasak Somdee6, Yodchaiy Chuaynkern1 & Chantip Chuaynkern1,* ABSTRACT The stomach contents of 18 tadpole species in Thailand were studied. The tadpoles in this study consisted of one species of the family Bufonidae (Duttaphrynus melanostictus); one species of Dicroglossidae (Fejervarya limnocharis); four species of Megophryidae (Megophrys carinense, Megophrys sp., Leptobrachium smithi and Leptolalax sp.); six species of Microhylidae (Microhyla butleri, Microhyla berdmorei, Microhyla heymonsi, Micryletta inornata, Kaloula pulchra and Glyphoglossus molossus); three species of Ranidae (Clinotarsus penelope, Hylarana erythraea and Sylvirana nigrovittata); and three species of Rhacophoridae (Polypedates leucomystax, Rhacophorus rhodopus and Rhacophorus jarujini). Tadpole specimens were collected by net from the following five areas: Phu Khiao Wildlife Sanctuary in Chaiyaphum Province, Yoddom Wildlife Sanctuary in Ubon Ratchathani Province, Phu Kradueng National Park in Loei Province, Umphang Wildlife Sanctuary in Tak Province, and Nam Nao National Park in Phetchabun Province. Specimens were dissected in order to remove their stomachs. All stomach contents were kept in order to create semi-permanent slides, which were observed under the light microscope. The results of tadpole stomach contents showed eight groups of food items, which included microalgae (25 genera), protozoa (3 genera), fungi like-organisms, rotifers (3 genera, including amictic eggs and bdelloid rotifers), diatoms (16 genera), arthropods, nematodes, and plant parts. Keywords: Amphibian, diet, food items, stomach analysis, Thailand INTRODUCTION Food is an important welfare factor for all living organisms. In amphibians, caecilians generally feed on earthworms and other invertebrates, while anurans and salamanders feed almost exclusively on insects (Vitt & Caldwell, 2009). Naturally, there is large diversity of food items available in their habitats, but the amphibians do not eat all of these prey items. Depending upon amphibian species and their foraging habits, only subsets of these available foods are eaten. Environment factors, such as physical and biological factors (or abiotic and biotic factors), influence the diversity of amphibian diets. 1 Department of Biology, Faculty of Science, Khon Kaen University, Mueang Khon Kaen, Khon Kaen, 40002, Thailand 2 Na Yung-Nam Som National Park, Udon Thani, 41380, Thailand 3 Center for Study Natural and Wildlife, Nam Yuen, Ubon Ratchathani, 34260, Thailand 4 Protected Area Regional Office 14 (Ubon Ratchathani), Department of National Parks, Wildlife and Plant Conservation, Mueang Ubon Ratchathani, Ubon Ratchathani, 63000, Thailand 5 Department of Forest Biology, Faculty of Forestry, Kasetsart University, Chatuchak, Bangkok, 10900, Thailand 6 Department of Microbiology, Faculty of Science, Khon Kaen University, Mueang Khon Kaen, Khon Kaen, 40002, Thailand * Corresponding author: [email protected] วารสารสัตว์ป่าเมืองไทย ปีที่ 25 พ.ศ. 2561 Journal of Wildlife in Thailand Vol. 25, 2018 22 To determine what amphibians eat, researchers try to use various methods and analyses to obtain their results (e.g., Solé et al., 2005; Chuaynkern et al., 2009; Camera et al., 2014; Ponpituk et al., 2015; Goldberg et al., 2018). Based on the items that we presently know that amphibians eat, we are still far from fully understanding their complete diets because what is known is mainly based on the diets of adult (or juvenile) amphibians. In amphibian tadpoles, food is very important for development (Kupferberg, 1997; Alford, 1999), which influences the size and shape of the tadpoles. However, little information about their food items has been published. Unlike the adult amphibians, the tadpoles generally feed on fungi, viruses, bacteria, algae, protists, helminths, plant cells, and on other tadpoles (Kupferberg et al., 1994; Hoff et al., 1999; Goldberg et al., 2018). Moreover, cannibalism in anuran larvae are also reported in several species (see e.g., Heusser, 1970; Jungfer & Schiesari, 1995). Approximately 178 amphibian species are known in Thailand, which is 10% of the known global species (Frost, 2018; Phusaensri et al., 2018). Among them, only some anuran species have had their food items reported on in detail (Chuaynkern et al., 2009; Ponpituk et al., 2015). However, for the tadpoles of the amphibians, almost nothing is known about their food items. Therefore, this present work has reported on the prey items of 18 anuran tadpoles based upon specimens collected from five localities of Thailand. MATERIALS AND METHODS From the following five localities in Thailand (Figure 1), the tadpole specimens were collected from nature by using dip-nets: Phu Kradueng National Park (Loei Province), Yoddom Wildlife Sanctuary (Ubon Ratchathani Province), Phu Khiao Wildlife Sanctuary (Chaiyaphum Province), Umphang Wildlife Sanctuary (Tak Province), and Nam Nao National Park (Phetchabun Province). The tadpoles were euthanized using 70% ethyl alcohol and were later preserved in a mixture solution of 10% formaldehyde and 70% ethyl alcohol at a ratio of 1:1 (followed Inthara et al., 2009). These tadpoles (Table 1) were catalogued and deposited in the Khon Kaen University Vertebrates Collection (KKUC) in Khon Kaen Province in northeastern Thailand. The identification of tadpoles was made by comparing the samples to relevant taxonomic works (Inthara, 2000; Grosjean et al., 2003, 2015; Taksintum, 2003; Meewattana, 2005; Stuart et al., 2006; Danisawat et al., 2010; Aran et al., 2012; Kaewtongkum et al., 2014a,b; Chuaynkern et al., 2017; Thongproh et al., 2018). The developmental stages of the tadpole specimens were determined according to the work of Gosner (1960). In order to investigate the stomach contents, tadpoles were selected, which were between Gosner’s stages of 31 to 39. Preparation of semi-permanent slides was modified from the protocol proposed by Somnark et al. (2011). The gut was excised by opening the tadpoles’ bellies using a surgical scissors. Later, the gut was opened, and all of the stomach contents were removed and placed into a Petri dish to which 70 % ethyl alcohol was added to preserve them. Then using a dropper was used to collect the solution of stomach contents, one drop was placed on the slide along with one drop of glycerin. Finally, the slide was covered with a cover slip and was sealed with nail polish at the edges of the cover slip. The stomach contents found on the semi-permanent slide were identified under a compound microscope (Zeiss) at 10x and 40x. Photographs were taken of all วารสารสัตว์ป่าเมืองไทย ปีที่ 25 พ.ศ. 2561 Journal of Wildlife in Thailand Vol. 25, 2018 23 the food items using a compound microscope (Zeiss). In order to identify these prey items, the photographs were compared with the standard literature review, and for some of the unidentified items, specialists were consulted. Figure 1 Map showing collected sites of larval specimens. 1=Umphang Wildlife Sanctuary (Tak Province), 2=Phu Kradueng National Park (Loei Province); 3=Nam Nao National Park (Phechabun Province); 4=Phu Khiao Wildlife Sanctuary (Chaiyaphum Province); 5=Yoddom Wildlife Sanctuary (Ubon Ratchathani Province). วารสารสัตว์ป่าเมืองไทย ปีที่ 25 พ.ศ. 2561 Journal of Wildlife in Thailand Vol. 25, 2018 24 Table 1 A list of tadpole specimens used in the present work. Map of localities was presented in Figure 1. Scientific names followed Frost (2018). Abbreviation: KD=Phu Kradueng NP, KKU=Khon Kaen Unviversity, NP=National Park, PK=Phu Khiao WS, PU=Protected Unit, WS=Wildlife Sanctuary, YC=Yodchaiy Chuaynkern, YD=Yoddom WS. Family/Species Number Stage No. of Locality Habitat specimens Family Bufonidae Duttaphrynus melanosctitus PK 007 31 3 Phu Khiao WS., “Temporary pond”, Chaiyaphum, Thung Kra Mung northeastern Thailand Family Dicroglossidae Fejervarya limnocharis PK 016 (R2) 31 3 Phu Khiao WS., “Temporary pond”, Chaiyaphum, Near Mor Chang Tai northeastern Thailand Family Megophryidae Megophrys carinense YC 122 36 1 Umphang WS., “Permanent forest steram”, Tak, western Thailand Huay Umphang Kee, Pa Do Tha PU. Megophrys sp. YC 157 36 1 Umphang WS., “Permanent forest stream”, Tak, western Thailand Tee Lor Zoo PU Leptobrachium smithi PK 009 (R2) 31,35,36 3 Phu Khiao WS., “Permanent forest stream”, Chaiyaphum, Huay Pong Hi northeastern Thailand Leptobrachium smithi YC 00814 37 1 Umphang WS., “Permanent forest stream”, Tak, western Thailand Huay Sae Dor Soup Gro Leptolalax sp. YC 00819 36 1 Umphang WS., “Permanent forest stream”, Tak, western Thailand Huay Tee Bue La Gro Family Microhylidae Glyphoglossus molossus PK 025 (R2) 34,35 3 Phu Khiao WS., “Temporary pond”, Chaiyaphum, near the Palace northeastern Thailand Kaloula pulchra KD 052 31,32,33 3 Phu Kradueng NP., “Temporary pond”, Loei, northeastern Lan Phra Kaew Thailand Microhyla berdmorei YC 00721 37 3 Umphang WS., “Permanent forest stream”, Tak, western Thailand Huay Tee Pha Do Microhyla berdmorei PK 006 33,34,35 3 Phu Khiao WS., “Temporary forest stream”, Chaiyaphum, Huay Kam Yan northeastern Thailand Microhyla butleri PK 020 (R2) 34,36,37 3 Phu Khiao WS., “Temporary pond”, Chaiyaphum, Thailand near Mor Chang Tai Microhyla butleri YD 0275 38 3 Yoddom WS., “Temporary pond”, Ubon Rachathani, Huay Chan Daeng PU northeastern Thailand Microhyla heymonsi PK 024 (R2) 31,32,33 3 Phu Khiao WS., “Temporary pond”, Chaiyaphum, Nong