Advertisement calls of four species of anurans from van long wetland nature reserve, Ninh Binh province, Viet Nam

151 HNUE JOURNAL OF SCIENCE DOI: 10.18173/2354-1059.2018-0040 Natural Sciences 2018, Volume 63, Issue 6, pp. 151-158 This paper is available online at ADVERTISEMENT CALLS OF FOUR SPECIES OF ANURANS FROM VAN LONG WETLAND NATURE RESERVE, NINH BINH PROVINCE, VIETNAM Le Trung Dung and Luong Mai Anh Hanoi National University of Education Abstract. We describe quantitatively the advertisement calls of four species of anurans from Van Long Wetland Nature Reserve (Microhyla heymonsi, M. mukhlesuri, M. pulchra, and Rhacophorus orlovi). This is the first study that describes calls from anurans of this area. Five acoustics parameters were measured, including call duration (s), call repetition rate (calls/s), inter–call interval(s), number of notes or pulses in a call, dominant frequency of call (kHz). An acoustic review of Vietnamese members of the genera showed that the dominant frequency of the calls to be is distinctly different in the species of investigated Microhyla and Rhacophorus species, it could be considered as static properties. Keywords: Anura, advertisement calls, Microhyla, Rhacophorus, Van Long Wetland Nature Reserve. 1. Introduction Advertisement calls are generally recognized as being the main premating reproductive isolating mechanism in anurans (Blair, 1958 [1]; Gerhardt, 1988 [2]; Ryan and Rand, 1993 [3]). The detailed description of anuran advertisement calls has proved to be a powerful tool for the determination of the taxonomic status of specimens collected in less explored areas. In order for the comparisons of calls to become a useful tool, a quantitative description of the characteristics of the advertisement calls of anuran species is needed. Van Long is the largest wetland nature reserve in northern Vietnam, covers about 3,000 hectares in Gia Vien district, Ninh Binh province. Currently, 17 species of anurans from three different families are known from Van Long (Le et al., 2015 [4]). In this paper we describe the calls of four species and compare the advertisement calls with previous studies, when possible. 2. Content 2.1. Material and methods Sampling: Field surveys (Figure 1) were conducted July 2015, September 2015 and March 2016 in Van Long Wetland Nature Reserve (Ninh Binh province). Specimens were collected between 19:00 and 24:00. After taking photograph specimens were preserved in 80% ethanol and subsequently transferred to 70% ethanol for permanent storage. Specimens were deposited in the Hanoi National University of Education. Received July 30, 2018. Revised August 24, 2018. Accepted August 30, 2018. Contact Le Trung Dung, e-mail address: letrungdung_sp@hnue.edu.vn Le Trung Dung and Luong Mai Anh 152 Figure 1. Map showing the survey sites in Van Long Wetland Nature Reserve, Ninh Binh province Morphological characters: Measurements were taken with a digital caliper to the nearest 0.1 mm. Abbreviations are as follows: a.s.l., above sea level; terminology of morphological characters followed Ohler et al. (2011) [5]. Recording methods: Advertisement calls were recorded with a Sony Recorder ICD-UX200F and a Parabolic microphone Telinga Pro-4PIP. Calls were recorded at a distance of approximately 0.2 m and ambient temperatures at the calling site were taken immediately after recording using an electronic thermal hygrometer Nakata NJ-2099-TH. Acoustic analysis: Calls were analyzed by using Raven Pro., version 1.3 (Cornell Laboratory of Ornithology, Ithaca, New York, U.S.A.) at resolution of 16 bit and frequency of 44.1 kHz. Spectrograms were performed with Hann window type, frame length of 1024 samples, and corresponding 3 dB filter bandwidth of 67.4 Hz; frame overlapped 50% with time–grid resolution of 512 samples and frequency-grid resolution of 46.9 Hz. For each acoustic signal, the following parameters were measured: call duration or call length(s): duration of time between beginning and end of a call; call repetition rate (calls/s): (total number of calls-1)/duration of time between beginning of the first call and end of the last call; inter-call interval(s): duration of time between end of the first call and beginning of the second call; number of notes per call: total number of notes in a call; number of pulses per note: total number of pulses within a note (in some cases); dominant frequency of call (kHz): the emphasized harmonic in the spectrum (after Duellman and Trueb, 1994 [6]; Cocroft and Ryan, 1995 [7]). The ten first notes of each call were removed according to research by Pröhl (2003) [8]. We calculated the coefficients of variation (CV=[SD/mean]×100%) of the acoustic features measured in calls. We employed the criteria of Gerhardt (1991) [9] and classified as static those acoustic properties with average within individual CV 12%, and CV from 5-12% was intermediate level of within–individual variation. Advertisement calls of four species of anurans from Van Long Wetland Nature Reserve 153 2.2. Results and discussions 2.2.1. Acoustic characters of four Anuran’s species from Van Long Wetland Nature Reserve Family MICROHYLIDAE Microhyla heymonsi Vogt, 1911/ Taiwan Rice Frog–mon General aspect. Morphological characters of the specimen from Van Long wetland nature reserve agreed well with the description of Hecht et al. (2013) [10]: Body small, vomerine teeth absent; tympanum hidden, fingers free webbing, with small discs; finger I shorter than II; toes webbed rudimentally. Dorsal and ventral skin smooth; supratympanic fold indistinct (Figure 2a). Figure 2. Dorsolateral views of: a, Microhyla heymonsi; b, M. mukhlesuri; c, M. pulchra Acoustic properties. The call emitted by one male (HNUE.VL.2015.139) were recorded at an air temperature of 28 o C and a relative humidity of 88%. The calls were composed of multinote, lasting for 0.529 - 1.421 s (0.836 ± 0.507 s, n = 3), call repetition rate 0.467 calls/s (n = 1), inter- call interval 1.862 - 2.058 s (1.96 ± 0.139 s, n = 2). Each note consisted of multipulse (7 - 12 pulse). The dominant frequency of the primary calls from 1.637 to 2.929 kHz (2.067 ± 0.746, n = 3) (Figure 3a, b). Variation. Rise time of calls (mean CV=60.57%), inter–call interval (mean CV=7.07%) and dominant frequency (mean CV=36.08%). Calling behavior. Calling males were found standing on the ground near stream, about 0.2–0.5 m from the stream. The smallest distance between two calling males was about 1–1.5 m. Remarks. Advertisement calls of the specimen agreed well with the description of Sheridan et al. (2010) [11], based on specimens collected in Taiwan, Thailand and Singapore. Advertisement calls of the specimen in Manchou, Taiwan has a dominant frequency of 3 kHz. The samples were collected from Singapore (2,406 ± 0,127 kHz) and Chiang Mai, Thailand (2.49 ± 0.066 kHz). Microhyla mukhlesuri Hasan, Islam, Kuramoto, Kurabayashi, and Sumida, 2014 / Mukhlesuri Rice Frog /muk–le–su–ri General aspect. Morphological characters of the specimen from Van Long wetland nature reserve agreed well with the description of Hasan et al. (2014) [12]: Body small and slightly elongated, vomerine teeth absent, tongue elliptical, tympanum hidden; fingers slender, free and tips not swollen, himdlimb long and stout. Skin smooth or warty, dark X-shaped mark on the dorsum. Ventral side is a slightly whitish with few very small fine speckles along the throat (Figure 2b). Acoustic properties. The calls emitted by one male (HNUE.VL.2015.105) were recorded at an air temperature of 29.7 o C and a relative humidity of 78%. The calls were composed of multinote, lasting for 0.085 - 0.28 s (0.221 ± 0.0444 s, n = 48), call repetition rate 1.702 calls/s (n = 1), inter–call interval 0.275 - 1.042 s (0.399 ± 0.149, n = 47). Each note consisted of multipulse (11 - 20 pulse). The dominant frequency of the primary calls from 2.842 to 2.929 kHz (2.891 ± 0.043, n = 48) (Figure 3c, d). Variation. Rise time of calls (mean CV = 20.13%), inter–call interval (mean CV = 37.31%) and dominant frequency (mean CV = 1.53%) were the most stereotyped properties. Le Trung Dung and Luong Mai Anh 154 Calling behavior. Calling males were found standing on the ground under the shrubs, about 1.5 - 2 m from the stream. The smallest distance between two calling males was about 0.5 - 1 m. Females are found on the ground next to the males. Microhyla pulchra (Hallowell, 1861)/ Guangdong Rice Frog General aspect. Morphological characters of the specimen from Van Long wetland nature reserve agreed well with the description of Taylor (1962) [13]: Body moderately large, tympanum hidden, vomerine teeth absent, tongue narrow, oval; forelimbs weak, first finger very much smaller than second, toes one–third to one–half webbed. Dorsal smooth, skin of chin finely wrinkled, venter nearly smooth (Figure 2c). Figure 3. Advertisement calls of Microhyla heymonsi (a, waveform and b, corresponding spectrogram); M. mukhlesuri (c, waveform and d, corresponding spectrogram); M. pulchra (e, waveform and f, corresponding spectrogram) Acoustic properties. The calls emitted by one male (HNUE.VL.2015.100) were recorded at an air temperature of 29.6 o C and a relative humidity of 79%. The calls were composed of multinote, lasting for 0.072 - 0.838 s (0.292 ± 0.186 s, n = 21), call repetition rate 0.634 - 0.667 calls/s (0.65 ± 0,023 call/s, n = 2), inter–call interval 0.652 - 2.51 s (1.371 ± 0.511, n = 19). Each note consisted of multipulse (38 - 80 pulse). The dominant frequency of the primary calls from 2.24 to 2.412 kHz (2.334 ± 0.046, n = 21) (Figure 3e, f). Advertisement calls of four species of anurans from Van Long Wetland Nature Reserve 155 Variation. Rise time of calls (mean CV = 63.82%), inter–call interval (mean CV = 37.25%) and dominant frequency (mean CV = 1.99%) were the most stereotyped properties. Calling behavior. Calling males were found standing on the ground near small puddle, about 1.5 - 3 m from the stream. Male were found around the puddle and spaced about 0.2 - 0.5 m. Females are found on the ground next to the males. Family RHACOPHORIDAE Rhacophorus orlovi Ziegler and Köhler, 2001/ Orlov's Treefrog / Ếch cây ooc–lop General aspect. Morphological characters of the specimen from Van Long wetland nature reserve agreed well with the description of Ziegler and Köhler (2001) [14]: Body moderately, tympanum distinct, vomerine teeth in oblique, tongue slightly elongated; each finger of the hand bearing a well developed; finger with webbing; tip of each toe bearing flattened disc. Skin on throat smooth, on abdomen and inner thighs coarsely granular, limbs without distinct dermal flaps and folds except for a weakly developed fold along outer edge of 4th finger and 5th toe (Figure 3a). Figure 3. Rhacophorus orlovi (a, Waveform and corresponding spectrogram; b, Dorsolateral view) Acoustic properties. The calls emitted by one male (HNUE.VL.2015.101) were recorded at an air temperature of 29.6 o C and a relative humidity of 79%. The calls were composed of singernote, lasting for 0.009 - 0.033 s (0.02 ± 0.005 s, n = 369), call repetition rate 3.8 calls/s (n=1), inter–call interval 0.2 - 0.524 s (0.224 ± 0.03, n = 368). Each note consisted of multipulse (46 - 49 pulse). The dominant frequency of the primary calls from 2.24 to 2.326 kHz (2.321 ± 0.02, n = 369) (Figure 3b). Advertisement calls of the specimen from Van Long agreed well with the description of Wildenhues et al. (2011) [15]. Variation. Rise time of calls (mean CV = 23.62%), inter–call interval (mean CV = 12.39%) and dominant frequency (mean CV = 0.86%) were the most stereotyped properties. Le Trung Dung and Luong Mai Anh 156 Calling behavior. Calling males were found standing on small vines near small puddle, about 0.5 - 1.2 m from the ground. The smallest distance between two calling males was about 5 - 10 m. 2.2.2. Variation of acoustic in the general Microhyla and Rhacophorus in Vietnam Genus Microhyla Tschudi, 1838 is small–bodied frogs, currently comprising 42 species. It’s inhabit is land with varying heights in the South, Southeast and East Asia ở (Frost, 2018) [15]. Vietnam is center of this genus with 12 species in 2009 and so far 16 species (Poyarkov et al., 2014) [17], of which eight species have known advertisement call (Heyer, 1971) [18]. Comparisons species of this study and species previously described in the Mycrohyla genus: M. berdmorei has the lowest dominant frequency of 1.5 - 1.8 kHz and high in M. negeriola 3 - 5,5 kHz (Das & Haas, 2010) [19] and less than 3.85 - 5.05 kHz in M. petrigena (Dehling, 2010) [20]. The frequency overlap occurs only in M. fissipe, M. berdmorei (Sheridan et al., 2010) [11], M. ornata (Kuramoto & Joshy, 2006) [21], M. orientalis (Matsui, 2013) (see Figure 4) [22]. Figure 4. Oscillate dominat frequency of the Microhyla species in Vietnam Figure 5. Oscillate dominat frequency of the Rhacophorus species in Vietnam Genus Rhacophorus Kuhl and Van Hasselt, 1822 are high numbers of extinct species, currently comprising 92 species. Vietnam is center of this genus with 16 species in 2009 so far 23 species (Frost, 2018) [16], of which seven species have known advertisement call (Tran, 2013) [23]. Comparisons species of this study and previously described acoustic species in the Rhacophorus genus (see Figure 5) show that the vocal cries of the dominant species are multinote: lowest in R. annamensis 0.86 - 1.94 kHz and highest in R. orlovi 1.599 - 3.975 kHz. The cries of the species Advertisement calls of four species of anurans from Van Long Wetland Nature Reserve 157 are overlapping but not overlapping. The dominant frequency overlap is focused only on some species such as R. bimaculatus (Heyer, 1971) [18], R. rhodopus, R. calcaneus (Tran, 2013) [23], R. dennysi, R. kio (Ziegler, 2002) [24]. The most prevalent in R. annamensis in Tran's study (2013) [23] was four types of vocalizations and the dominant frequency of cries for each individual recorded, which could explain that R. annamensis is a species rather than a single species. 3. Conclusion Acoustic characters of four spcies (Microhyla heymonsi, M. mukhlesuri, M. pulchra, and Rhacophorus orlovi) were described from Van Long Wetland Nature Reserve, Ninh Binh Province. This is the first study that describes calls from anurans of this area. Five acoustics parameters were measured, including call duration (s), call repetition rate (calls/s), inter–call interval(s), number of notes or pulses in a call, dominant frequency of call (kHz). An acoustic review of Vietnamese members of the genera Microhyla and Rhacophorus showed that the dominant frequency of the calls to be is distinctly different in the species, it could be considered as static properties. Acknowledgements. We are grateful to the directorates of the Forest Protection Department of Ninh Binh Province, and Van Long Wetland Nature Reserve for support of our field work and issuing relevant permits. D. T. Le thanks A.M. Luong, D.T. Pham (Hanoi) and staff of Van Long Wetland Nature Reserve for their assistance in the field. We thank D.T. Pham and H. T. Ngo (Hanoi) for laboratory assistance. This research was partially funded by the Hanoi National University of Education (Grant No. SPHN16-19TT). REFERENCES [1] Blair, W.F., 1958. Mating call in the speciation of anuran amphibians. American Naturalist 92, pp. 27-51.
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