A checklist of amphibians at Lubuk Semilang Recreational Park, Langkawi Island, Kedah, Peninsular Malaysia

A checklist of amphibians at Lubuk Semilang Recreational Park, Langkawi Island, Kedah, Peninsular Malaysia. A study on amphibian fauna at the lowland dipterocarp forest of Lubuk Semilang Recreational Park, Langkawi, Kedah, Peninsular Malaysia was conducted between December 2018 and June 2019. We recorded 18 species of amphibians from 14 genera and 6 families. These include seven new records for Lubuk Semilang, which are Fejervarya limnocharis, Limnonectes hascheanus, Megophrys aceras, Kaloula pulchra, Microhyla butleri, Pulchrana laterimaculata, and Polypedates discantus. We also recorded a new anuran record for Langkawi Archipelago, Pulchrana laterimaculata, and two endemic species of Malaysia, Leptobrachium smithi and Limnonectes macrognathus. Results from this survey indicate the amphibian species of the Langkawi Archipelago has increased from 28 to 29 species. Data published through GBIF (Doi:10.15470/f0diow)


Introduction
Lubuk Semilang Recreational Park is located at the base of Gunung Raya Forest Reserve, Langkawi, Kedah, Peninsular Malaysia. This forest covers 4,930 hectares and was declared a permanent forest in 1941 (Kedah Forestry Department, 2014). Lubuk Semilang Recreational Park is located beside Kampung Buku Malaysia and is approximately eight kilometres from the nearest town, Kuah. The stream at Lubuk Semilang, Sungai Korok, originates from the highest mountain of Langkawi, Gunung Raya (881 m a.s.l.). It flows into Sungai Kok, then into a larger stream, Sungai Melaka, before draining into the Straits of Malacca. In the Lubuk Semilang area, Sungai Korok is approximately two km long and it has a two-tier waterfall that is very popular for recreational and outdoor activities.
At Lubuk Semilang, only a small area had been explored and studied by previous researchers. Many areas around Lubuk Semilang, such as small streams, wetlands, swamps, seasonal pools, ditches and upper parts of Sungai Korok, remain unexplored. The present research was thus carried out to document amphibian species that inhabit the lowland dipterocarp forest of Lubuk Semilang.

Material and methods
This research was conducted at the lowland dipterocarp forest of Lubuk Semilang Recreational Park,'' E, elevation < 100 m a.s.l.) ( fig. 1) Kedah, Peninsular Malaysia, between December 2018 and June 2019. Twenty-five consecutive visits were made. Samplings of amphibian species were performed at eight different habitat types: the lower, middle and upper part of Sungai Korok, small streams beside Mardi and Kg. Buku, marsh wetland, slow-moving stream, and road trail ( fig. 2). Table 1 shows the characteristics of the habitat for each locality (see also fig. 3-6).
Tadpoles and egg clutches were collected between 10:00 and 13:00 h using a scoop net, and they were then placed in plastic containers. Amphibian individuals were captured between 20:00 and 23:00 h by means of a Visual Encounter Survey (VES), by hand or a scoop net, and with the help of headlights and flashlights. Two to three field members were involved in the surveys. Each live specimen was photographed in situ using a compact camera (Nikon COOLPIX L840) before being collected for identification.
Measurements taken for each individual were: weight (WT), tibia length (TL) (length of the outer surface of the flexed knee to the heel/tibiotarsal inflection), snout-vent length (SVL) (length from the tip of snout to posterior margin of the vent), and head width (HW) (the widest part of the head or angle of the jaws) (Watters et al., 2016). Measurements were made using electronic scales and a caliper.
We recorded habitat characteristics, including the type of vegetation, river width, water current flow, and canopy-covering. Microhabitat characteristics, which include horizontal position, vertical position and substrates, were also determined (Heyer et al., 1994). Horizontal position refers to the shortest distance of the species from any water bodies (streams or ponds). Vertical position refers to subsurface on the exposed soil surface (on a rock or on the plant), while substrate refers to the surface area that the individual sits on at the time of capture (rock, vegetation, log, mineral soil or dead leaf) (Zainudin, 2009;Zainudin et al., 2017). We also recorded abiotic conditions such as humidity, temperature, and weather using AccuWeather application (android).
Selected anuran species were fixed in 10 % formalin and stored in 7 % ethanol. Specimens were tagged and deposited at the School of Pharmaceutical Sciences in Universiti Sains Malaysia (USM) for future reference. Tissue samples (from thigh muscles) of the preserved specimens were also collected, stored in 95 % ethanol, and deposited at the same location for future molecular analysis. Samples were collected and sampled in accordance with legal regulations.

Duttaphrynus melanostictus (Schneider, 1799)
This common Sunda toad was observed in the lower part of Sungai Korok and along small streams beside Kg. Buku and Mardi. They inhabit streams that have a high anthropogenic effect and usually perch on the bare soil of the sandy bank. On some nights, the calls of this toad can be heard from the lower part of Sungai Korok. This species can be identified by its stout body, blunt fingers, large round warts on its back, smaller warts on the side, and elongated parotoid glands (Berry, 1975).

Phrynoidis asper (Gravenhorst, 1829)
19USM This giant river toad can be spotted perched on large rocks, at midstream, and along the rocky bank at all parts of Sungai Korok, and also at a small, swift-flowing stream beside Mardi. During the daytime, they can be seen resting in cracks underneath large rocks. Sometimes they were found partially submerged underwater, beside the large rocks. The three specimens were collected at site 2 on 01/01/2019. They were identified by their large size, stout body, distinct tympanum, first finger slightly longer than second, large round warts on the back, and the roundish or subtriangular shape of prominent parotid glands (Berry, 1975). Limnonectes macrognathus --

Fejervarya limnocharis (Gravenhorst, 1829)
This common grass frog was found at all sites of Lubuk Semilang Recreational Park except at the upper part of Sungai Korok and on the road trail. At every visit, they were found sitting on sand/soil along the stream banks and marsh wetlands. This species was identified by its grey to brownish colour with darker spots or patches, dark cross-bars on limbs, dark bars on lips, white ventral surface, distinct tympanum, slightly pointed snout, undilated blunt fingers, and pointed toes with undilated tips (Berry, 1975).

Limnonectes blythii (Boulenger, 1920)
Blyth's river frog was frequently sighted at sites 1, 2, and 3 of Sungai Korok. Most were found sitting on medium-sized rocks midstream, on sand/soil of stream banks, or partially submerged in shallow streams. This species was recognized by its large size, head longer than broad, distinct tympanum, full webbing on toes and whitish-yellow ventral surface (Berry, 1975). Three specimens of hill forest frog were collected on 28/12/2018 and 04/01/2019. All three individuals were collected at site 8, hiding among leaf litter, on the road trail, 6 m away from a permanent river. This species was observed abundantly along a stream beside the road  Grismer et al. (2006); C, Daicus et al. (2005); CR, current research; -, not recorded; x, recorded.

Number of species (21) 7 4 7 18
trail and along Sungai Korok. It was identified by its small size, colour mostly pale brown, dark crossbar visible between the eyes, 'ᴧ' shaped mark between the shoulders, a blackish streak on each side of the head, distinct tympanum, short fingers and fingertips are dilated into small disks (Berry, 1975). Some orange and yellowish colour individuals were also seen. Two large-headed river frogs were caught on the 24/01/2019. The first specimen was caught hiding among leaf litter, on the rocky bank at site 7 (slow-moving stream). The second specimen was captured while hiding under a large rock on a rocky bank at the same location. On 29/06/2019, the calls of this species could be heard at the same location. This frog was identified by its distinctive large broad head and visible, large tympanum for male, smaller head and smaller tympanum in females, dark bars on lips, irregular dark marking on dorsum, limbs with rounded tubercles, blunt fingers, first finger slightly longer than second, and smooth dorsum with longitudinal skin fold (Norhayati et al., 2009;Smith, 1930). One Marten's oriental puddle frog was collected on 28/12/2018, sitting on the surface of leaf litter near a shallow stream beside Mardi (1-2 m width, 0.1-0.3 m depth). Another two individuals were caught partially submerged in watery mud and on the mud bank of marsh wetland on 20/01/2019. This frog was identified by its small, broad, oval-shaped head, hidden tympanum, brownish grey colouring on dorsal skin with dark round freckles and yellowish-white ventral skin (Norhayati, 2017).

Family Megophryidae (2 species)
Leptobrachium smithi (Matsui, Nabhitabhata and Panha, 1999) ( fig. 12) 19USM/LA/LS01: TL = 16 mm, SVL = 40 mm, HW = 19 mm Temperature = 30 ºC; relative humidity = 54 %. An adult male Smith's litter frog was caught while calling under a large rock on a rocky bank, at the lower part of Sungai Korok on 28/01/2019. Their heavy-bodied tadpoles were also found grazing on the sandy bottom at the middle part of Sungai Korok and a small stream beside a road trail. This frog was identified by its wide head, wider than the body, large eyes, proportionally long arms, short legs, minimal webbing, and intense banded hind legs (Norhayati, 2017). On 01, 04/01/2019, an adult Perak spade-foot toad, was collected in the middle part of Sungai Korok and a juvenile was collected on a road trail. The adult was captured hiding under a large rock on a rocky bank. The juvenile was caught among leaf litter on the road, at least 50 m from the nearest stream. Their tadpoles were collected hiding among leaf litter sediment in a shallow slow-moving stream, beside the road trail (2-3 m width, 0.3 m depth). The frog was identified by its hidden tympanum, a triangular mark between the eyes, short dermal projection on top of eyelids, and limbs with dark crossbars (Norhayati et al., 2009).
Only some of the species mentioned in the dataset published in GBIF are discussed in the text. Species mentioned in the text were less common (except for P. asper) and represent all the samples in the dataset. Additional data on these species is given because this information was lacking. Samples of Pulchrana laterimaculata (six juvenile voucher specimens) are mentioned because represent a new record of amphibian species within Langkawi Archipelago.
In Lubuk Semilang, accessibility to the area has increased significantly over time, enabling detection of more species than in previous studies. This study was conducted in both dry and wet seasons for a longer period, minimizing bias concerning detections of amphibian species in Lubuk Semilang. During a gap in research of over ten years, habitat alteration and heterogeneity significantly increased detection of some amphibian species and absence of others. These alterations and heterogeneity were mainly due to changes in-stream structure, weather patterns (especially precipitation), and human recreational activities that may provide niches, microhabitats, continuous foraging, and especially breeding habitat for these amphibians (McCaffery, 2010;Nur Amalina et al., 2017). We consider the absence of Occidozyga lima is due to misidentification with Occidozyga martensii. Several Occidozyga collected in Lubuk Semilang were morphologically similar to Occidozyga martensii, rather than to the Occidozyga lima specimen (LRCUKM-0179) from Datai that were deposited in Langkawi Research Centre, Universiti Kebangsaan Malaysia (UKM). This showed that Occidozyga lima can be found in Langkawi, but not in Lubuk Semilang. For a long time, Polypedates discantus was known as Polypedates leucomystax. It was only recently separated from Polypedates leucomystax complex species and named Polypedates discantus in a study by Rujirawan et al. (2013). From molecular studies, Polypedates discantus has uncorrected pairwise distances of 6.61-7.16 % compared to its closest relative, Polypedates leucomystax. Morphological characteristics of Polypedates discantus are head skin that has not co-ossified with the skull, indistinct or absent white dots on the ventral part of the thigh skin, presence of round tubercle on the tibiotarsal articulation, and mildly visible 'X' marking on its back (Rujirawan et al., 2013). These morphological structures were similar to Polypedates found in Lubuk Semilang.
Pulchrana laterimaculata is a new record for the Langkawi Archipelago. Several Pulchrana laterimaculata juveniles and sub-adult individuals were found around marsh wetland/ swamp, showing this species emerges from the nearby forest to use the swampy areas as their breeding site (Shahriza et al., 2011a(Shahriza et al., , 2011bShahriza and Ibrahim, 2014). This finding has increased the number of amphibian species in the Langkawi Archipelago from 28 to 29 species.
The most common frog found in Lubuk Semilang, Fejervarya limnocharis, was identified in open areas at all the study sites except in the most disturbed sites (the upper part of Sungai Korok and forest trails), showing it is widely spread in both disturbed and less disturbed areas.
Several frog species, including Hylarana erythraea, Kaloula pulchra, Microhyla butleri, Fejervarya limnocharis, and Duttaphrynus melanostictus, are commensal species (Shahriza et al., 2011a(Shahriza et al., , 2011bShahriza and Ibrahim, 2012, 2014, 2017. These species were often encountered at forest edges and disturbed areas with anthropogenic activities (Ibrahim et al., 2001;Tuen and Das, 2005;Norsham et al., 2005;Shahriza and Ibrahim, 2017) but rarely seen in forests (Inger and Stuebing, 1990). In Lubuk Semilang, these species can be found at sites 1, 4, 5 and 6 which are likely to be disturbed areas with anthropogenic activities. There are many activities here, including recreational activity and nature-based tourism. Alterations of local habitats provide various niches and microhabitats especially for the opportunists or generalist species (Nur Amalina et al., 2017). Pristine forest species or specialist species were replaced by species that adapted to secondary forest or generalist species (D'Cruze and Kumar, 2011). This explains why commensal species have high occurrences in Lubuk Semilang Recreational Park, especially at sites 1, 5 and 6.
The physical structure of streams determines different riparian frog species (Inger, 1969).
Limnonectes blythii, Phrynoidis asper, and Chalcorana labialis are riparian species and inhabit river banks (Berry, 1975;Inger and Stuebing, 1989;Shahriza et al., 2011aShahriza et al., , 2011b. These frog species were sighted through the sampling period in and along Sungai Korok. The presence of clean water species such as Odorrana hosii and Chalcorana labialis indicates that sites 2 and 3 were still undisturbed and pristine. According to Ibrahim et al. (2012Ibrahim et al. ( , 2013, is also considered a semi-disturbed species. In this study, Chalcorana labialis can be found in both disturbed and undisturbed areas. Odorrana hosii can be seen perched on rock boulders, either at midstream or rock banks of the middle and upper part of Sungai Korok. Both sites have moderate to fast stream currents, a suitable environment for this species (Ibrahim et al., 2008;Grismer, 2011;Shahriza and Ibrahim, 2012). Sites 4, 5, 6 and 7 in Lubuk Semilang were considered ideal as breeding grounds for amphibians. The shallow slow-moving stream in site 5 was considered an active breeding ground for Chalcorana labialis due to the presence of egg clutches, tadpoles and several gravid females of this species. Juveniles of Pulchrana laterimaculata and Pulchrana glandulosa were present at site 6, indicating this area is the breeding ground for both species. The shallow slow-moving stream in site 7 was a breeding ground for Microhyla berdmorei, Leptobrachium smithi, Megophrys aceras, and Limnonectes macrognathus. Several egg clutches and an amplexus pair of Microhyla berdmorei were found there. Additionally, tadpoles of Microhyla berdmorei, Leptobrachium smithi, and Megophrys aceras were also sighted there. Mating calls of Microhyla berdmorei, Pulchrana glandulosa, Occidozyga martensii and Limnonectes macrognathus were heard at the same site.
According to Barnett (2005), wetlands, slow backwater off-channel streams, shallow ponds, and seasonal pools with a water depth of less than 0.5 meters become suitable breeding grounds for amphibians. Amphibians prefer shallow water to breed, generally due to the presence of fewer invertebrates, absence of fish predators, and water that is warmer and thus suitable for egg and larval development (Barnett, 2005). These types of environments can be found at sites 4, 5, 6 and 7 of Lubuk Semilang.
Forest frog species that inhabit the lowland dipterocarp forest of Lubuk Semilang were Limnonectes hascheanus, Leptobrachium smithi, Megophrys aceras, Microhyla berdmorei, and Limnonectes macrognathus. The presence of these species indicates some parts of Lubuk Semilang are still undisturbed or have a minimal disturbance. The presence of two Langkawi endemic frogs, Leptobrachium smithi and Limnonectes macrognathus, shows that Lubuk Semilang is an important site for amphibian species and should be fully protected. The survey on amphibian species in these areas is ongoing and many potential areas are yet to be explored.