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South-East Asia, a global biodiversity hotspot, is home to four species of otters: smooth-coated otter (Lutrogale perspicillata), Asian small-clawed otter (Aonyx cinereus; Figure 1), hairy-nosed otter (Lutra sumatrana), and Eurasian otter (Lutra lutra; Basnet et al., 2020). Among these four species, the smooth-coated otter and the Asian small-clawed otter are listed as Vulnerable, the hairy-nosed otter as Endangered, and the Eurasian otter as Near-Threatened in the IUCN's Red List assessment (Khoo et al., 2021; Roos et al., 2021; Sasaki et al., 2021; Wright et al., 2021). There is a dearth of information about the distribution and conservation status of these four species of otters in South-East Asia (Duplaix & Savage, 2018), where populations are threatened by rapid economic development, habitat loss, illegal wildlife trade, pollution and degradation of aquatic habitat, human-otter conflict, and climate change (Cianfrani et al., 2018; Duckworth & Hill, 2008; Foster-Turley, 1992). Increased pollution of aquatic habitats has impacted otter food resources and expanding linear infrastructure such as road networks is emerging as one of the major causes of otter mortality (Peterson & Schulte, 2016).
Enlarge this image.FIGURE 1. A group of the Asian small-clawed otter (Aonyx cinereus) in their tidal habitat of the Sundarban mangrove forest in Bangladesh.
Increasing threats to otter populations throughout the world have called for increased and focused efforts to monitor their status and distributions (Duplaix & Savage, 2018). However, due to their elusive habits and rarity in distribution, monitoring otters is not an easy task. A plethora of noninvasive survey techniques, which possess the advantages of being relatively inexpensive and replicable, exist to monitor small carnivores (Long et al., 2012). Sign surveys using footprints and spraints have been traditionally used for monitoring presence, population trends, and habitat use, specifically for the Eurasian otter and North American River otter (Lontra canadensis). Otter spraints are typically deposited at latrine sites located along the banks of rivers and wetlands. Fresh otter spraints are easily detected by their strong fishy odor. In the last two decades, advancements in camera trapping and noninvasive DNA analysis techniques have opened new avenues for otter monitoring. Camera traps at latrine sites have provided valuable data on species, group size, and social behavior (Ben-David et al., 1998; Green et al., 2015; Stevens & Serfass, 2008). Noninvasive molecular genetic sampling using spraints has been successfully employed in ecological and demographic studies of Eurasian and North American otters (Hájková et al., 2009; Jansman et al., 2001; Klütsch & Thomas, 2018; Sittenthaler et al., 2020). These methods increase the scope of population monitoring by providing data on otter presence and habitat use, including additional information such as individual identification by DNA analysis, which can be used to estimate population size and density.
Accurate species identification is vital for population monitoring. However, where the ranges of multiple species of otters overlap, as in South-East Asia, surveys using sign or camera traps are limited by uncertainty and difficulties in definitively assigning spraints, footprints, or photographs to species. Kruuk et al. (1993) and Kistner et al. (2022) provide detailed descriptions to distinguish among tracks, spraints, and spraint sites of three otter species (Lu. lutra, A. cinereus, and L. perspicillata), but these signs, specifically tracks, persist for a short period in tropical weather conditions, and their reliable interpretation depends largely on the consistency of the substrate. A majority of South Asian and South-East Asian countries have more than two otter species. Cambodia, Indonesia (Sumatra), Thailand, Myanmar, and Vietnam report four species. Thus, efficient and cost-effective methods to overcome the challenge of species identification are crucial for monitoring otter populations in these regions.
Although all four species of otters have been reported from Malaysia, only three species, the smooth-coated otter, Asian small-clawed otter, and hairy-nosed otter, have consistently confirmed distribution records. Smooth-coated otters and Asian small-clawed otters are the most common and occur through much of tropical Asia including Peninsular Malaysia (Abdul-Patah et al., 2020; Wright et al., 2021) and Malaysian Borneo (Forest Department Sarawak, 2020; Khoo et al., 2021). The hairy-nosed otter is the only species of otter endemic to South-East Asia, but is rare with sporadic records from its known range, including Malaysia (Sasaki et al., 2021). A recent study (Abdul-Patah et al., 2020) provided the most comprehensive information on the spatial distribution, associated habitats, and genetic diversity (mitochondrial) of the three species of otters in Peninsular Malaysia. This study confirmed otter species in 138 locations of Peninsular Malaysia, out of which hairy-nosed otters were only confirmed from seven sparsely distributed locations. Roadkills of the hairy-nosed otter have been reported near peat swamp forests in Perak, Pahang (Sebastian, 1995), and more recently in Northern Selangor, Malaysia (Tan, 2015; Woo, 2021). The presence of the Eurasian otter in Malaysia is highly uncertain and limited to just two records from the northern part of the peninsula (Miller, 1900; Mohd-Azlan & Sharma, 2006) and one from Borneo (Phillipps & Phillipps, 2016). All species of otter are “Totally Protected” based on the Wildlife Conservation Act 2010 in Peninsular Malaysia, and “Protected” in Malaysian Borneo by the Wildlife Conservation Enactment 1997 of the state of Sabah and the Wild Life Protection Ordinance 1998 of the state of Sarawak.
Species identification by DNA analysis of scat samples has relied mainly on the amplification of mitochondrial DNA sequences, such as the control and 16s rRNA regions (e.g., Mills et al., 2000) cytochrome b gene (e.g., Hsieh et al., 2001; Madisha et al., 2015) and mitochondrial D-loop region (Bozarth et al., 2010). DNA from noninvasive sources such as hair and feces usually contain only trace amounts of low-quality fragmented DNA, which degrades further with time, resulting in amplification failure in old decayed samples (Shih et al., 2017; Waits & Paetkau, 2005). Abdul-Patah et al. (2014, 2020) successfully identified otter species from spraints by sequencing a fragment of ~450-bp DNA, amplified from the mitochondrial D-loop region. They were successful in amplifying DNA from ~50% of the spraint samples (A.-P. Pazil, personal communication) to ascertain species. The amplification of short (<300 bp) fragments of mitochondrial DNA is reported to produce higher amplification success rates (Madisha et al., 2015; Shih et al., 2017). PCR amplification of mitochondrial fragments has been successfully combined with restriction-fragment-length polymorphisms (PCR-RFLP) analysis to distinguish among closely related taxa (e.g., Bidlack et al., 2007; Cossíos & Angers, 2006; Hansen & Jacobsen, 1999; Mukherjee et al., 2010). This approach has the advantages of being relatively rapid and cost-effective because amplification products can be separated by size, producing distinct species-specific band patterns and eliminating the need for sequencing. Scats deposited by other small carnivores may sometimes be misidentified as otter spraints. Erroneous identification of scats of American mink (Neovison vison), European mink (Mustela lutreola), and European polecat (Mustela putorius) as those of European otter is reported (Hansen & Jacobsen, 1999), but no such information is reported by studies on Asian otter species.
Otters are apex carnivores in aquatic habitats and thus crucial for regulating aquatic food webs and ecosystem-level trophic dynamics (Ben-David et al., 1998; Kruuk, 2006). Tracking the status of otter populations is therefore fundamental to ensure the stability of aquatic ecosystems in South and South-East Asia. Conservation monitoring programs for elusive species such as otters require significant effort to achieve sufficient sample sizes for reliable estimates. Higher amplification success rates and accurate species identification with noninvasive samples will greatly improve the quality of data needed for accurate mapping of otter species range, population distributions, estimation of occupancy, density, and population trends. Here, we develop and evaluate three PCR-RFLP (polymerase chain reaction—restriction-fragment-length polymorphism) protocols with the aim of designing an efficient and cost-effective noninvasive species identification method for three otter species found in Malaysia and other parts of South-East Asia. Further, we compare these PCR-RFLP protocols for their cost-effectiveness and their ability in successfully and reliably identifying these otter species.
MATERIALS AND METHODS Field sampling and reference sample collectionThis study is the preliminary part of a larger project to systematically survey and map the distribution of otters along the North Central Selangor Coast (Lat 3.8444° N, Long 101.8152° E to Lat 2.9762° N, Long 101.2811° E) on the west coast of Malaysia (Figure 2). Opportunistic records and surveys confirm that all three species of otter inhabit this region (Abdul-Patah et al., 2020; Woo, 2021). We collected 33 otter spraint samples from various locations in this region (Figure 2). We obtained reference samples (blood samples on FTA cards and/or tissue samples) of two species of otters (smooth-coated otter and Asian small-clawed otter), common palm civet (Paradoxurus hermaphroditus), and short-tailed mongoose (Herpestes brachyurus) from the Department of Wildlife and National Parks, Peninsular Malaysia (PERHILITAN; Table S1). The common palm civet and short-tailed mongoose are two small carnivores that occur sympatrically with otters in the North Central Selangor Coast. Their scats are similar in size to those of otters, although chances of scat misidentification are generally minimal due to morphological dissimilarities between scats and spraints and dietary differences among these species.
Enlarge this image.FIGURE 2. Map of the North Central Selangor Coast in Peninsular Malaysia, where 33 otter spraint samples were collected for our study. Insets A and B are magnified views of sampled sections of North Central Selangor Coast. The identification and assignment of these samples to otter species are based on PCR-RFLP results of this study.
We surveyed potential otter habitat to locate spraint sites and collected fresh spraints in 5-ml collection vials with absolute ethanol as preservative. These samples were stored at room temperature/−20°C before lab analysis.
Laboratory analysisWe extracted DNA from approximately 0.25 mg of spraint samples via the alkaline lysis method using the GF-1 Soil Sample DNA Extraction Kit (Vivantis Technologies). DNA extraction procedure followed the manufacturer's instructions with a slight modification; the incubation time was increased to 5 min prior to DNA elution to maximize DNA yield. We also extracted DNA from reference samples of A. cinereus, L. perspicillata, and H. brachyurus from blood samples stored on the FTA cards and P. hermaphroditus from tissue samples using the GF-1 Tissue Blood Combi DNA Extraction Kit (Vivantis Technologies). The extracted DNA was stored at −20°C.
We amplified two fragment sizes of the mt D-loop region using two different sets of primers: (i) forward TanaD-F, 5′-ACCATCAGCACCCAAAGCTG-3′ (Masuki et al., 2008) and reverse TanaD-R, 5′-GGGCTGATTAGTCATTAGTCCATC-3′ (Masuki et al., 2008) primers for amplification of 450-bp fragment and (ii) forward TanaD-Mod-F (5′-CACCATGCCTCGAGAAACCA-3′) designed in this study and reverse TanaD-R primers for a shorter 200-bp fragment. Additionally, we designed another set of primers (forward OTR-ND4-F, 5′- GGCAACCAAACAGAACGCC-3′ and reverse OTR-ND4-R, 5′- GTTTAAGGAGTACGGCGGCAA-3′) to amplify a 300-bp fragment of the mt ND4 gene.
We designed the OTR-ND4 primer using Primer-Blast (Ye et al., 2012) along with the complete mitochondrial genomes of three otter species (A. cinereus: GenBank KY117535.1, L. perspicillata: GenBank KY117557.1, Lu. sumatrana: GenBank KY117556.1; Salleh et al., 2017). We used Primer-Blast to design the TanaD-Mod-F primer using 324 published sequences (Abdul-Patah et al., 2020) representing all three species (A. cinereus: GenBank OL588639.1–OL588743.1, L. perspicillata: GenBank OL588744.1–OL588954.1, Lu. sumatrana: GenBank OL588632.1–OL588638.1; Appendix 1). We used the OligoAnalyzer™ Tool (Integrated DNA Technologies) to check for secondary structures in these primers. Thereafter, we searched for appropriate sets of restriction enzymes using an interactive web-based program VIRS (Chen et al., 2009). This tool can identify and visualize restriction sites in multiple DNA sequences and suggests appropriate restriction enzymes for those sites. Our objective was to select those enzymes that would not be affected by CpG methylation and provide discernible band patterns (ideally less than three bands with sufficient difference among their size) for easy visualization on agarose gels. We also conducted in silico PCR and restriction digestion, followed by gel visualization in the SnapGene program (GSL Biotech LLC).
DNA amplifications of the targeted fragments were conducted in a final reaction volume of 25 μl consisting of 12.5 μl ExPrime Taq Premix (2X) (GENETBIO Inc.), 7.5 μl distilled water, 1.5 μl of 10 pmol/μl forward and reverse primers, and 2 μl extracted otter fecal DNA. PCR was carried out using T100® Thermal Cycler (Bio-Rad Laboratories) with the following parameters: initial denaturation at 94°C for 10 min followed by 45 cycles of denaturation at 94°C for 30 s, annealing at 56°C and 62°C for D-loop fragments and ND4 fragment, respectively, for 30 s, and extension at 72°C for 1 min, and a final extension at 72°C for 7 min. The amplicons were electrophoresed and visualized on a 2% agarose gel to determine specific amplifications. We included extraction negatives and PCR negatives in every batch of PCR performed.
First, we confirmed all reference samples, including one spraint sample of hairy-nosed otter collected from our study area that was used as a reference for all follow-up lab analysis. These samples were sequenced (Applied Biosystems™ 3730xl DNA Analyzer) for the mt D-loop and mt ND4 regions with the primers mentioned above. Forward and reverse sequences were checked, edited, and merged into contigs using ChromasPro2.0 (Technelysium Pty. Ltd.). Sequences obtained from this study were compared with reference taxonomic sequences deposited in the National Center for Biotechnology Information (NCBI) using the nucleotide Basic Local Alignment Search Tool (nBLAST) with default settings. Species-specific amplifications were confirmed if query sequences shared high similarity with any taxa from the Lutrinae subfamily. After confirming species identity, these reference samples were used as positive controls for subsequent PCR reactions of 33 spraint samples collected from various sites in the study region.
After confirming the amplification of PCR products, restriction digestions were conducted in a final reaction volume of 30 μl consisting of 10 μl PCR products, 1–2 μl restriction enzyme (Thermo Fisher Scientific), 2 μl 1X buffer (Thermo Fisher Scientific), and 16–17 μl distilled water. The digestion mixtures were incubated at 37°C (Hin1II, NdeI, and BsuRII) and 65°C (MseI and HpyCH4III) for 4 h, and digested products were separated via electrophoresis on a 2.5% agarose gel at 60 V for 2 h.
Cost analysisOur aim was to develop a cost-effective and accurate protocol that would be useful and financially feasible to otter researchers in Asia. Thus, we evaluated the cost of the reagents required for processing a batch of 100 samples to determine the most cost-effective approach for species identification of three species of otters using the PCR-RFLP method with three primer sets and a combination of suitable restriction enzymes. We also compared the cost of PCR-RFLP with sequencing costs for species identification. Reagents used in this study were commercially available and were obtained from local distributors in Malaysia. We estimated and converted the cost in U.S. Dollars (US$).
RESULTSSequencing confirmed that all reference samples were from their respective species. Reference samples for three otter species, including the spraint sample of one hairy-nosed otter and the common palm civet, amplified successfully with the three primer sets (TanaD, TanaD-Mod, and OTR-ND4) used in this study and were ascertained to their respective species by an nBLAST search (Figure 3a,e,g). We also found that the fragment size for the common palm civet reference sample was larger (~500 bp) than those of the otter species (~450 bp; Figure 3a). The short-tailed mongoose reference sample amplified only with the TanaD-Mod primer, providing an amplification product similar in size to the other four species (Figure 3g), but did not produce any distinct fragments in RFLP analysis (Figure 3h). RFLP analysis of the remaining four species with three sets of primers and five restriction enzymes provided species-specific band patterns (Figure 3b–d,f,h).
Enlarge this image.FIGURE 3. Gel electrophoresis results of successful amplification of Aonyx cinereus (AC), Lutrogale perspicillata (LP), Lutra sumatrana (LS), Paradoxurus hermaphroditus (PH), and Herpestes brachyurus (HB) reference samples using three primers sets (TanaD, OTR-ND4, and TanaD-Mod) and combinations of restriction enzymes for reference samples of A. cinereus, L. perspicillata, Lu. sumatrana, P. hermaphroditus, and H. brachyurus, respectively. The two negative wells (N1 and N2) refer to DNA extraction and PCR negatives, respectively. Panels (a–h) show results of PCR amplification and RFLP digestions of three primers and restriction enzymes.
Amplification rates of 33 samples differed among the three primer sets. Amplification success decreased with increasing PCR fragment size. The TanaD primer with the largest PCR product (~450 bp) amplified just 60.6% (n = 20) of DNA from spraints and produced spurious bands of various sizes for several samples. This primer set also produced inconsistent results for RFLP analysis with three different restriction enzymes MseI, NdeI, and HpyCH4III. Therefore, we discontinued the use of TanaD to analyze further samples using the PCR-RFLP method. Amplification rates were slightly better (63.6% of spraints) for the OTR-ND4 primer, which produced a band of ~300 bp. Moreover, OTR-ND4 provided the expected RFLP profile for correct species identification after RFLP analysis with BsuRI for all 21 spraint samples that successfully amplified. The TanaD-Mod primer, which yielded the smallest PCR product (~200 bp) of all three primer sets, amplified all 33 samples. Subsequent RFLP analysis with Hin1II restriction enzyme identified the species of otter in 32 (97%) of the samples.
We compared the costs of PCR-RFLP analysis for the three protocols. The OTR-ND4 primer and BsuRI enzyme pair was the least expensive, totaling 437 USD for DNA extraction, PCR amplification, and RFLP analysis of 100 samples (Table 1). The TanaD-Mod primer with Hin1II enzyme would cost 455 USD for analysis. However, if compared both for consistency and accuracy in species identification and costs, the TanaD-Mod and Hin1II pair emerges as the most cost-effective PCR-RFLP protocol (Table 2).
TABLE 1 Detailed cost breakdown of lab analysis.
Note: Costs were calculated for the analysis of 100 samples and reported in the USD. Various colors (orange, green and blue) indicate different stages of laboratory analysis.
*Agarose, TBE buffer, DNA stain, loading dye, and plastic consumables.
TABLE 2 Comparison of amplification success, PCR-RFLP accuracy in species identification, and cost comparison of various primer sets and Sanger sequencing used in our study.
| Primers | TanaD-Mod | OTR-ND4 | TanaD | Sequencing | ||
| Amplification success (%) | 100 | 63.63 | 60.6 | |||
| Restriction enzymes | Hin1II | BsuRI | MseI | NdeI | HpyCH4II | |
| PCR-RFLP accuracy (%) | 96.97 | 61.9 | 30 | – | – | |
| Cost, 100× (US$) | 455 | 437 | 453 | 442 | 470 | 937 |
Note: Cost per 100 samples includes chemical consumables for DNA extraction, PCR amplification, and RFLP analysis.
DISCUSSIONMonitoring small carnivores is a daunting task because of their elusive nature and sparse distribution. Where closely related species of small carnivores co-occur, signs such as tracks and scats are often not very useful to ascertain species. Remote cameras have improved the quality of small carnivore surveys, but have limitations where morphologically similar species such as Asian otters co-occur. There is an enormous lack of information about small carnivore distribution and ecology, with no robust and scientifically validated information on distribution ranges and population size for a number of species, owing to the lack of research studies (Glatston & Duplaix, 2020; Torres-Romero & Giordano, 2022). Without this baseline information, assessment of the impacts and extent of threats to these species and the effectiveness of conservation measures is difficult.
The primary goal of this study was to develop an accurate, easy-to-implement, cost-effective DNA analysis method for the identification of three Asian otter species. We describe a cost-effective PCR-RFLP method for identifying three otter species that has very high amplification success with noninvasive samples and is half the cost of Sanger sequencing for species identification. We also detected an inverse relationship between fragment size and amplification success rates for the three primers in this study, which has also been reported in previous studies (Broquet et al., 2007; Deguilloux et al., 2002, 2003; Shih et al., 2017). Previously, the TanaD primer was used for amplifying DNA from otter spraints followed by Sanger sequencing to identify species (Abdul-Patah et al., 2020). Besides the high costs and technical expertise required for sequencing analysis, amplification success was low (~50%) in the study due to the large fragment size. In our study, RFLP analysis of the amplified PCR product of TanaD primer produced inconsistent results; thus, we consider it unsuitable for species identification using PCR-RFLP on noninvasive samples. Amplification success of large fragments is expected to be low with noninvasive samples such as spraints in which DNA is likely to degrade rapidly in warm, humid tropical environments. We designed a new forward primer, TanaD-Mod-F, to amplify a shorter fragment of the TanaD primer with the expectation that the smaller fragment length of the PCR product would increase amplification success of spraint samples. As expected, this modified primer yielded a very high amplification rate (99.3%) with field-collected samples.
One of the strengths of our study is that we used a published database of 324 reference sequences for three otter species (Lu. sumatrana, A. cinereus, and L. perspicillata) to select suitable restriction enzymes that had unique restriction sites for all the haplotypes reported for these three species in Malaysia (Abdul-Patah et al., 2020). This approach increased the likelihood that the selected primer–enzyme pair would produce an accurate RFLP profile while also ensuring replicability by checking for the possibility of mutations at enzyme cleavage sites by screening these 313 reference sequences collected from various locations in Malaysia. The D-loop region of mitochondrial DNA is highly variable, and any PCR-RFLP assay designed for this region with a few reference sequences might have a risk of nonreplicability when applied to samples collected from other regions. To further address this issue, we tested the TanaD-Mod primer with 151 field-collected spraint samples and amplified 99.3% of these samples (Sunway University, unpublished data). Of these, 84.1% (n = 127) were identified as spraints deposited by otter species and 15.9% (n = 24) as those of common palm civet. We designed the OTR-ND4 primer for a second primer–enzyme combination targeting the mitochondrial ND4 region of the three otter species. Although this primer–enzyme pair had less amplification success than TanaD-Mod, it had moderately high accuracy and was the least expensive among all three PCR-RFLP protocols. We suggest the use of the OTR-ND4 primer in conjunction with the TanaD-Mod primer–enzyme pair as a complimentary marker where species identification needs to be reconfirmed (e.g., forensic analysis of confiscated samples).
It is prudent to mention here that hybridization between L. perspicillata and A. cinereus has been reported in the putative otter sample of L. perspicillata collected from Singapore (Moretti et al., 2017), but this study also mentions that hybridization events such as these are likely to occur when the population of one species is much smaller than the other (A. cinereus in this case). This calls for further investigation with nuclear markers to ascertain species identification, particularly if one of the co-occurring species is rare in the study region.
Cost is an important determinant of the feasibility of any monitoring program. Most genetic monitoring programs require technical knowledge, access to highly expensive lab equipment, and analyses whose costs are often prohibitive for most researchers in South and South-East Asia. These are important limiting factors for research and conservation work in this region, where local students and wildlife researchers work on shoestring budgets for their projects. The protocols we present in this study were developed specifically to reduce the costs and processing time of molecular genetic species identification. For otter researchers planning large-scale monitoring programs for otters in Asia, we anticipate that this protocol will improve efficiency and enhance data quality.
We caution that our PCR-RFLP approach is based on a reference database consisting largely of otter species from the Malay peninsula. In other regions, different RFLP profiles may result because of regional haplotype differences. Thus, we suggest for an initial pilot study to confirm species identity via sequencing as an essential prelude to surveys using the PCR-RFLP methods described here. Also, our PCR-FLP approach can be applied through the Malay peninsula and Borneo where Eurasian otter is not a resident. In areas where the Eurasian otter is sympatric with smooth-coated otters or small-clawed otters (i.e., ~11 countries in tropical Asia; Khoo et al., 2021; Roos et al., 2021; Wright et al., 2021), we recommend screening spraint samples using Hansen and Jacobsen's (1999) PCR-RFLP protocol for Eurasian otter species identification for further confirmation of their presence. We also recommend to develop and test other upcoming advanced technique such as e-DNA analysis with next-generation sequencing (NGS) for distribution surveys of Asian otter species. New advanced techniques such as NGS hold much promise for noninvasive genetic studies, but may not be available nor affordable for ecological monitoring in several otter range countries in South and South-East Asia in the near future. The methodologies presented here provide a more affordable alternative.
CONCLUSIONOtters are sentinels of freshwater ecosystems. They are also resilient species that respond well to habitat conservation and population protection. In places where habitats (rivers, wetlands, and marine coastlines) are restored, and where pollution is controlled, otters have shown remarkable recolonization potential (Duplaix & Savage, 2018). Many of the world's species of otter coexist with people, including in highly urbanized areas such as Singapore (Khoo & Lee, 2020; Theng & Sivasothi, 2016). Making an active effort for otter conservation in these regions is also linked with some of the specific Sustainable Development Goals (SDGs) such as SDG 6—clean water and sanitation, SDG 14—life below water, SDG 15—life on land, and SDG 13—climate action. Asian otters are in peril due to loss of habitat, trafficking for pelts and the pet trade, and from conflicts with aquaculture industries. Conservation measures for these species would require robust, replicable, and cost-effective monitoring methods. The PCR-RFLP methods described in this paper match these criteria and can accurately identify three species of Asian otters. These protocols also have important forensic applications such as the identification of seized otter pelts and roadkills. Accurate species identification is also an important step that should precede genetic identification of individuals, sex, relatedness, population genetics, and diet assessment by metabarcoding. The methods we describe here can be used to generate baseline spatial distribution maps, including studies of occupancy, species interaction and habitat use, genetic monitoring, and other valuable ecological information that will facilitate the conservation of otters in Asia.
AUTHOR CONTRIBUTIONSSandeep Sharma: Conceptualization (equal); data curation (equal); formal analysis (equal); investigation (equal); methodology (equal); project administration (equal); supervision (equal); validation (equal); visualization (equal); writing – original draft (equal); writing – review and editing (equal). Woo Chee-Yoong: Conceptualization (equal); data curation (equal); funding acquisition (equal); investigation (equal); methodology (equal); project administration (equal); writing – original draft (equal); writing – review and editing (equal). Adrian Kannan: Data curation (equal); formal analysis (equal); investigation (equal); methodology (equal); validation (equal); visualization (equal); writing – original draft (equal). Suganiya Rama Rao: Data curation (equal); formal analysis (equal); investigation (equal); methodology (equal); validation (equal); visualization (equal). Pazil Abdul-Patah: Resources (equal); supervision (equal); validation (equal). Shyamala Ratnayeke: Conceptualization (equal); funding acquisition (equal); investigation (equal); methodology (equal); project administration (equal); supervision (equal); validation (equal); visualization (equal); writing – original draft (equal); writing – review and editing (equal).
ACKNOWLEDGMENTSWe express our gratitude to the following who either contributed or supported this project. The late Balu Perumal, Head of MNS Conservation Division and the co-founder of the MNS Otter Conservation Project for supporting the research since its inception. The local field team (Azlie bin Aris, Shakh Muhammad Iszuan bin Shakh Manzar Ahmed Manzu, Hassan Bin Mat, Shakthi A/L Periasamy, Kahar bin Buntal, Rusli bin Salleh, Mohd Helmi bin Mohd Agus, Kamarul bin Abdullah, Haizan anak Kamarul Zaman, Lim Wei Hang, and Norsyila binti Ali) that helped during the field surveys and sample collection. The government agencies, Department of Wildlife and National Parks, Peninsular Malaysia (Permit no: W-00042-15-20 and W-00528-16-21) and Forestry Department of Peninsular Malaysia who granted the research permits and provided reference samples. Kuala Selangor Nature Park authority for granting permission for sampling in the park. Sunway Lagoon Wildlife Park for providing spraint samples and Wildlife Reserve Singapore for helping in tracing the origin of captive Asian small-clawed otter of Sunway Lagoon Wildlife Park. The funding agencies of this project—the Rufford Foundation, MSIG Holdings (Asia) Pte Ltd., Conservation International Asia-Pacific Ltd., The Conservation, Food & Health Foundation and Keidanren Nature Conservation Fund.
DATA AVAILABILITY STATEMENTThe data that support the findings of this study are available from the corresponding author upon reasonable request.
| S. No. | GenBank accession number | Description |
| 1 | OL588743.1 | Aonyx cinereus isolate SG_NIPAH_AC2 D-loop, partial sequence; mitochondrial |
| 2 | OL588742.1 | Aonyx cinereus isolate PSR_PANJANG_AC_2 D-loop, partial sequence; mitochondrial |
| 3 | OL588741.1 | Aonyx cinereus isolate PSR_PANJANG_AC_1 D-loop, partial sequence; mitochondrial |
| 4 | OL588740.1 | Aonyx cinereus isolate SG_Panjang_AC_1 D-loop, partial sequence; mitochondrial |
| 5 | OL588739.1 | Aonyx cinereus isolate Bgn_Lalang_AC_1 D-loop, partial sequence; mitochondrial |
| 6 | OL588738.1 | Aonyx cinereus isolate Sg_Pelek_AC_2 D-loop, partial sequence; mitochondrial |
| 7 | OL588737.1 | Aonyx cinereus isolate Sg_Pelek_AC_1 D-loop, partial sequence; mitochondrial |
| 8 | OL588736.1 | Aonyx cinereus isolate Ulu_Yam_AC_1 D-loop, partial sequence; mitochondrial |
| 9 | OL588735.1 | Aonyx cinereus isolate Sg_Buluh_AC_1 D-loop, partial sequence; mitochondrial |
| 10 | OL588734.1 | Aonyx cinereus isolate Sg_Timun_AC_3 D-loop, partial sequence; mitochondrial |
| 11 | OL588733.1 | Aonyx cinereus isolate Sg_Timun_AC_2 D-loop, partial sequence; mitochondrial |
| 12 | OL588732.1 | Aonyx cinereus isolate Sg_Timun_AC_1 D-loop, partial sequence; mitochondrial |
| 13 | OL588731.1 | Aonyx cinereus isolate LUKUT_AC_1 D-loop, partial sequence; mitochondrial |
| 14 | OL588730.1 | Aonyx cinereus isolate KPilah_AC_2 D-loop, partial sequence; mitochondrial |
| 15 | OL588729.1 | Aonyx cinereus isolate KPilah_AC_1 D-loop, partial sequence; mitochondrial |
| 16 | OL588728.1 | Aonyx cinereus isolate SERTING_AC_3 D-loop, partial sequence; mitochondrial |
| 17 | OL588727.1 | Aonyx cinereus isolate SERTING_AC_2 D-loop, partial sequence; mitochondrial |
| 18 | OL588726.1 | Aonyx cinereus isolate SERTING_AC_1 D-loop, partial sequence; mitochondrial |
| 19 | OL588725.1 | Aonyx cinereus isolate Klinggi_AC_3 D-loop, partial sequence; mitochondrial |
| 20 | OL588724.1 | Aonyx cinereus isolate Klinggi_AC_2 D-loop, partial sequence; mitochondrial |
| 21 | OL588723.1 | Aonyx cinereus isolate Klinggi_AC_1 D-loop, partial sequence; mitochondrial |
| 22 | OL588722.1 | Aonyx cinereus isolate Durian_Tunggal_AC3 D-loop, partial sequence; mitochondrial |
| 23 | OL588721.1 | Aonyx cinereus isolate Durian_Tunggal_AC2 D-loop, partial sequence; mitochondrial |
| 24 | OL588720.1 | Aonyx cinereus isolate Durian_Tunggal_AC1 D-loop, partial sequence; mitochondrial |
| 25 | OL588719.1 | Aonyx cinereus isolate Perlok_AC_3(2) D-loop, partial sequence; mitochondrial |
| 26 | OL588718.1 | Aonyx cinereus isolate Perlok_AC_3 D-loop, partial sequence; mitochondrial |
| 27 | OL588717.1 | Aonyx cinereus isolate Perlok_AC_2 D-loop, partial sequence; mitochondrial |
| 28 | OL588716.1 | Aonyx cinereus isolate Perlok_AC_1 D-loop, partial sequence; mitochondrial |
| 29 | OL588715.1 | Aonyx cinereus isolate Tmn_Negara_AC_3 D-loop, partial sequence; mitochondrial |
| 30 | OL588714.1 | Aonyx cinereus isolate K_Rompin_AC_2 D-loop, partial sequence; mitochondrial |
| 31 | OL588713.1 | Aonyx cinereus isolate Ksalang_AC_1 D-loop, partial sequence; mitochondrial |
| 32 | OL588712.1 | Aonyx cinereus isolate KSalang_AC_3 D-loop, partial sequence; mitochondrial |
| 33 | OL588711.1 | Aonyx cinereus isolate KPerlis3_AC_3 D-loop, partial sequence; mitochondrial |
| 34 | OL588710.1 | Aonyx cinereus isolate KPerlis3_AC_2 D-loop, partial sequence; mitochondrial |
| 35 | OL588709.1 | Aonyx cinereus isolate KPerlis3_AC_1 D-loop, partial sequence; mitochondrial |
| 36 | OL588708.1 | Aonyx cinereus isolate KPerlis2_AC_2 D-loop, partial sequence; mitochondrial |
| 37 | OL588707.1 | Aonyx cinereus isolate KPerlis1_AC_2 D-loop, partial sequence; mitochondrial |
| 38 | OL588706.1 | Aonyx cinereus isolate KPerlis1_AC_1 D-loop, partial sequence; mitochondrial |
| 39 | OL588705.1 | Aonyx cinereus isolate Kuala_Muda_AC_4 D-loop, partial sequence; mitochondrial |
| 40 | OL588704.1 | Aonyx cinereus isolate Kuala_Muda_AC_2 D-loop, partial sequence; mitochondrial |
| 41 | OL588703.1 | Aonyx cinereus isolate Kuala_muda_AC_1 D-loop, partial sequence; mitochondrial |
| 42 | OL588702.1 | Aonyx cinereus isolate Kg_hj_jamal_AC_2 D-loop, partial sequence; mitochondrial |
| 43 | OL588701.1 | Aonyx cinereus isolate Kubang_Ps_AC_2 D-loop, partial sequence; mitochondrial |
| 44 | OL588700.1 | Aonyx cinereus isolate Kubang_Ps_AC_1 D-loop, partial sequence; mitochondrial |
| 45 | OL588699.1 | Aonyx cinereus isolate Penaga_AC_1 D-loop, partial sequence; mitochondrial |
| 46 | OL588698.1 | Aonyx cinereus isolate Penaga_AC_2 D-loop, partial sequence; mitochondrial |
| 47 | OL588697.1 | Aonyx cinereus isolate P_Remis2_AC_3 D-loop, partial sequence; mitochondrial |
| 48 | OL588696.1 | Aonyx cinereus isolate P_Remis2_AC_2 D-loop, partial sequence; mitochondrial |
| 49 | OL588695.1 | Aonyx cinereus isolate P_Remis2_AC_1 D-loop, partial sequence; mitochondrial |
| 50 | OL588694.1 | Aonyx cinereus isolate P_Remis_AC_1 D-loop, partial sequence; mitochondrial |
| 51 | OL588693.1 | Aonyx cinereus isolate KGULA_AC_7 D-loop, partial sequence; mitochondrial |
| 52 | OL588692.1 | Aonyx cinereus isolate ENDAU_AC_2 D-loop, partial sequence; mitochondrial |
| 53 | OL588691.1 | Aonyx cinereus isolate SEDILI_AC_2 D-loop, partial sequence; mitochondrial |
| 54 | OL588690.1 | Aonyx cinereus isolate SEDILI_AC_3 D-loop, partial sequence; mitochondrial |
| 55 | OL588689.1 | Aonyx cinereus isolate RENGIT_AC_1 D-loop, partial sequence; mitochondrial |
| 56 | OL588688.1 | Aonyx cinereus isolate TG_PIAI_AC_4 D-loop, partial sequence; mitochondrial |
| 57 | OL588687.1 | Aonyx cinereus isolate TG_PIAI_AC_1 D-loop, partial sequence; mitochondrial |
| 58 | OL588686.1 | Aonyx cinereus isolate TG_PIAI_AC_9 D-loop, partial sequence; mitochondrial |
| 59 | OL588685.1 | Aonyx cinereus isolate TG_PIAI_AC_11 D-loop, partial sequence; mitochondrial |
| 60 | OL588684.1 | Aonyx cinereus isolate TG_PIAI_AC_3 D-loop, partial sequence; mitochondrial |
| 61 | OL588683.1 | Aonyx cinereus isolate TG_PIAI_AC_2 D-loop, partial sequence; mitochondrial |
| 62 | OL588682.1 | Aonyx cinereus isolate TG_PIAI_AC_10 D-loop, partial sequence; mitochondrial |
| 63 | OL588681.1 | Aonyx cinereus isolate PONTIAN_AC_5 D-loop, partial sequence; mitochondrial |
| 64 | OL588680.1 | Aonyx cinereus isolate PONTIAN_AC_3 D-loop, partial sequence; mitochondrial |
| 65 | OL588679.1 | Aonyx cinereus isolate PONTIAN_AC_2 D-loop, partial sequence; mitochondrial |
| 66 | OL588678.1 | Aonyx cinereus isolate G_PATAH_AC_4 D-loop, partial sequence; mitochondrial |
| 67 | OL588677.1 | Aonyx cinereus isolate G_PATAH_AC_2 D-loop, partial sequence; mitochondrial |
| 68 | OL588676.1 | Aonyx cinereus isolate G_PATAH_AC_1 D-loop, partial sequence; mitochondrial |
| 69 | OL588675.1 | Aonyx cinereus isolate SG_BUAYA_AC2 D-loop, partial sequence; mitochondrial |
| 70 | OL588674.1 | Aonyx cinereus isolate SG_BUAYA_AC1 D-loop, partial sequence; mitochondrial |
| 71 | OL588673.1 | Aonyx cinereus isolate T_AGAS_AC_8 D-loop, partial sequence; mitochondrial |
| 72 | OL588672.1 | Aonyx cinereus isolate T_AGAS_AC_4 D-loop, partial sequence; mitochondrial |
| 73 | OL588671.1 | Aonyx cinereus isolate T_AGAS_AC_3 D-loop, partial sequence; mitochondrial |
| 74 | OL588670.1 | Aonyx cinereus isolate P_Karang_AC_6 D-loop, partial sequence; mitochondrial |
| 75 | OL588669.1 | Aonyx cinereus isolate P_Karang_AC_5 D-loop, partial sequence; mitochondrial |
| 76 | OL588668.1 | Aonyx cinereus isolate Pekan_Nanas_AC_1 D-loop, partial sequence; mitochondrial |
| 77 | OL588667.1 | Aonyx cinereus isolate Sg_Kemaman_AC_4 D-loop, partial sequence; mitochondrial |
| 78 | OL588666.1 | Aonyx cinereus isolate Sg_Kemaman_AC_2 D-loop, partial sequence; mitochondrial |
| 79 | OL588665.1 | Aonyx cinereus isolate Sg_Kemaman_AC_1 D-loop, partial sequence; mitochondrial |
| 80 | OL588664.1 | Aonyx cinereus isolate Penarik_AC_14 D-loop, partial sequence; mitochondrial |
| 81 | OL588663.1 | Aonyx cinereus isolate Penarik_AC_13 D-loop, partial sequence; mitochondrial |
| 82 | OL588662.1 | Aonyx cinereus isolate Penarik_AC_5 D-loop, partial sequence; mitochondrial |
| 83 | OL588661.1 | Aonyx cinereus isolate Penarik_AC_3 D-loop, partial sequence; mitochondrial |
| 84 | OL588660.1 | Aonyx cinereus isolate Penarik_AC_2 D-loop, partial sequence; mitochondrial |
| 85 | OL588659.1 | Aonyx cinereus isolate Sabak_AC_3 D-loop, partial sequence; mitochondrial |
| 86 | OL588658.1 | Aonyx cinereus isolate Sabak_AC_2 D-loop, partial sequence; mitochondrial |
| 87 | OL588657.1 | Aonyx cinereus isolate Sabak_AC_1 D-loop, partial sequence; mitochondrial |
| 88 | OL588656.1 | Aonyx cinereus isolate Kg_Padang_Slm_AC_14 D-loop, partial sequence; mitochondrial |
| 89 | OL588655.1 | Aonyx cinereus isolate Padang_salimAC_13 D-loop, partial sequence; mitochondrial |
| 90 | OL588654.1 | Aonyx cinereus isolate AC_Kg_Padang_Salim_12 D-loop, partial sequence; mitochondrial |
| 91 | OL588653.1 | Aonyx cinereus isolate KgPadang_Salim_AC_11 D-loop, partial sequence; mitochondrial |
| 92 | OL588652.1 | Aonyx cinereus isolate KgPadang_Salim_AC_10 D-loop, partial sequence; mitochondrial |
| 93 | OL588651.1 | Aonyx cinereus isolate Kg_Padang_Salim_AC_9 D-loop, partial sequence; mitochondrial |
| 94 | OL588650.1 | Aonyx cinereus isolate Kg_Padang_Salim_8 D-loop, partial sequence; mitochondrial |
| 95 | OL588649.1 | Aonyx cinereus isolate KgPadang_Salim_AC_37570_7 D-loop, partial sequence; mitochondrial |
| 96 | OL588648.1 | Aonyx cinereus isolate KgPadang_Salim_AC_37570_4 D-loop, partial sequence; mitochondrial |
| 97 | OL588647.1 | Aonyx cinereus isolate Kg_Padang_Salim_AC_3 D-loop, partial sequence; mitochondrial |
| 98 | OL588646.1 | Aonyx cinereus isolate Kg_Padang_Salim_AC_2 D-loop, partial sequence; mitochondrial |
| 99 | OL588645.1 | Aonyx cinereus isolate Kg_Padang_Salim_AC_1 D-loop, partial sequence; mitochondrial |
| 100 | OL588644.1 | Aonyx cinereus isolate Tumpat_AC_15 D-loop, partial sequence; mitochondrial |
| 101 | OL588643.1 | Aonyx cinereus isolate Tumpat_AC_14 D-loop, partial sequence; mitochondrial |
| 102 | OL588642.1 | Aonyx cinereus isolate Tumpat_AC_12 D-loop, partial sequence; mitochondrial |
| 103 | OL588641.1 | Aonyx cinereus isolate Tumpat_AC_303 D-loop, partial sequence; mitochondrial |
| 104 | OL588640.1 | Aonyx cinereus isolate Tumpat_AC_304 D-loop, partial sequence; mitochondrial |
| 105 | OL588639.1 | Aonyx cinereus isolate AC_Tumpat_505 D-loop, partial sequence; mitochondrial |
| 106 | OL588954.1 | Lutrogale perspicillata isolate KgPadang_salim_LP_5 D-loop, partial sequence; mitochondrial |
| 107 | OL588953.1 | Lutrogale perspicillata isolate KgPadang_salim_LP_6 D-loop, partial sequence; mitochondrial |
| 108 | OL588952.1 | Lutrogale perspicillata isolate Tg_Piandang2_LP_14 D-loop, partial sequence; mitochondrial |
| 109 | OL588951.1 | Lutrogale perspicillata isolate Tg_Piandang2_LP_12 D-loop, partial sequence; mitochondrial |
| 110 | OL588950.1 | Lutrogale perspicillata isolate Tg_Piandang2_LP_13 D-loop, partial sequence; mitochondrial |
| 111 | OL588949.1 | Lutrogale perspicillata isolate Sg_Besut_LP_6 D-loop, partial sequence; mitochondrial |
| 112 | OL588948.1 | Lutrogale perspicillata isolate T_Alam_LP_1 D-loop, partial sequence; mitochondrial |
| 113 | OL588947.1 | Lutrogale perspicillata isolate Paya_Indah1_LP_4 D-loop, partial sequence; mitochondrial |
| 114 | OL588946.1 | Lutrogale perspicillata isolate Paya_Indah1_LP_3 D-loop, partial sequence; mitochondrial |
| 115 | OL588945.1 | Lutrogale perspicillata isolate Bkt_Tagar_LP_2 D-loop, partial sequence; mitochondrial |
| 116 | OL588944.1 | Lutrogale perspicillata isolate Bkt_Tagar_LP_1 D-loop, partial sequence; mitochondrial |
| 117 | OL588943.1 | Lutrogale perspicillata isolate JUGRA_LP_2 D-loop, partial sequence; mitochondrial |
| 118 | OL588942.1 | Lutrogale perspicillata isolate JUGRA_LP_1 D-loop, partial sequence; mitochondrial |
| 119 | OL588941.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_17 D-loop, partial sequence; mitochondrial |
| 120 | OL588940.1 | Lutrogale perspicillata isolate Kelanang_LP-1 D-loop, partial sequence; mitochondrial |
| 121 | OL588939.1 | Lutrogale perspicillata isolate SG_MAS_LP-1 D-loop, partial sequence; mitochondrial |
| 122 | OL588938.1 | Lutrogale perspicillata isolate Paya_Indah2_LP_10 D-loop, partial sequence; mitochondrial |
| 123 | OL588937.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_13 D-loop, partial sequence; mitochondrial |
| 124 | OL588936.1 | Lutrogale perspicillata isolate Paya_Indah1_LP_6 D-loop, partial sequence; mitochondrial |
| 125 | OL588935.1 | Lutrogale perspicillata isolate Paya_Indah1_LP_7 D-loop, partial sequence; mitochondrial |
| 126 | OL588934.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_12 D-loop, partial sequence; mitochondrial |
| 127 | OL588933.1 | Lutrogale perspicillata isolate Paya_Indah1_LP_5 D-loop, partial sequence; mitochondrial |
| 128 | OL588932.1 | Lutrogale perspicillata isolate Paya_Indah1_LP_1 D-loop, partial sequence; mitochondrial |
| 129 | OL588931.1 | Lutrogale perspicillata isolate Paya_Indah1_LP_2 D-loop, partial sequence; mitochondrial |
| 130 | OL588930.1 | Lutrogale perspicillata isolate Sg_Paka_LP_1 D-loop, partial sequence; mitochondrial |
| 131 | OL588929.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_11 D-loop, partial sequence; mitochondrial |
| 132 | OL588928.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_19 D-loop, partial sequence; mitochondrial |
| 133 | OL588927.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_18 D-loop, partial sequence; mitochondrial |
| 134 | OL588926.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_15 D-loop, partial sequence; mitochondrial |
| 135 | OL588925.1 | Lutrogale perspicillata isolate Panchang_Bedana1_LP_5 D-loop, partial sequence; mitochondrial |
| 136 | OL588924.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_14 D-loop, partial sequence; mitochondrial |
| 137 | OL588923.1 | Lutrogale perspicillata isolate P_JAWA_LP_2 D-loop, partial sequence; mitochondrial |
| 138 | OL588922.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_10 D-loop, partial sequence; mitochondrial |
| 139 | OL588921.1 | Lutrogale perspicillata isolate Panchang_Bedana1_LP_7 D-loop, partial sequence; mitochondrial |
| 140 | OL588920.1 | Lutrogale perspicillata isolate Sg_Perepet_LP_4 D-loop, partial sequence; mitochondrial |
| 141 | OL588919.1 | Lutrogale perspicillata isolate Panchang_Bedana1_LP_1 D-loop, partial sequence; mitochondrial |
| 142 | OL588918.1 | Lutrogale perspicillata isolate Panchang_Bedana1_LP_3 D-loop, partial sequence; mitochondrial |
| 143 | OL588917.1 | Lutrogale perspicillata isolate Sg_Bewah_LP_3 D-loop, partial sequence; mitochondrial |
| 144 | OL588916.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_7 D-loop, partial sequence; mitochondrial |
| 145 | OL588915.1 | Lutrogale perspicillata isolate K_Kemaman_LP_1 D-loop, partial sequence; mitochondrial |
| 146 | OL588914.1 | Lutrogale perspicillata isolate Panchang_Bedana2_LP_4 D-loop, partial sequence; mitochondrial |
| 147 | OL588913.1 | Lutrogale perspicillata isolate Panchang_Bedana1_LP_8 D-loop, partial sequence; mitochondrial |
| 148 | OL588912.1 | Lutrogale perspicillata isolate Sg_Bewah_LP_4 D-loop, partial sequence; mitochondrial |
| 149 | OL588911.1 | Lutrogale perspicillata isolate Panchang_Bedana1_LP_4 D-loop, partial sequence; mitochondrial |
| 150 | OL588910.1 | Lutrogale perspicillata isolate P_JAWA_LP_3 D-loop, partial sequence; mitochondrial |
| 151 | OL588909.1 | Lutrogale perspicillata isolate Sg_Perepet_LP_3 D-loop, partial sequence; mitochondrial |
| 152 | OL588908.1 | Lutrogale perspicillata isolate P_JAWA_LP_4 D-loop, partial sequence; mitochondrial |
| 153 | OL588907.1 | Lutrogale perspicillata isolate Sg_Bewah_LP_2 D-loop, partial sequence; mitochondrial |
| 154 | OL588906.1 | Lutrogale perspicillata isolate Sg_Bewah_LP_1 D-loop, partial sequence; mitochondrial |
| 155 | OL588905.1 | Lutrogale perspicillata isolate K_Kemaman_LP_3 D-loop, partial sequence; mitochondrial |
| 156 | OL588904.1 | Lutrogale perspicillata isolate Sg_Perepet_LP_1 D-loop, partial sequence; mitochondrial |
| 157 | OL588903.1 | Lutrogale perspicillata isolate Sg_Besut_LP_5 D-loop, partial sequence; mitochondrial |
| 158 | OL588902.1 | Lutrogale perspicillata isolate K_Kemaman_LP_4 D-loop, partial sequence; mitochondrial |
| 159 | OL588901.1 | Lutrogale perspicillata isolate Sg_Cacing_LP_2 D-loop, partial sequence; mitochondrial |
| 160 | OL588900.1 | Lutrogale perspicillata isolate K_Kemaman_LP_2 D-loop, partial sequence; mitochondrial |
| 161 | OL588899.1 | Lutrogale perspicillata isolate Sg_Paka_LP_4 D-loop, partial sequence; mitochondrial |
| 162 | OL588898.1 | Lutrogale perspicillata isolate Sg_Paka_LP_3 D-loop, partial sequence; mitochondrial |
| 163 | OL588897.1 | Lutrogale perspicillata isolate Sg_Besut_LP_1 D-loop, partial sequence; mitochondrial |
| 164 | OL588896.1 | Lutrogale perspicillata isolate Sg_Besut_LP_3 D-loop, partial sequence; mitochondrial |
| 165 | OL588895.1 | Lutrogale perspicillata isolate Sg_Besut_LP_2 D-loop, partial sequence; mitochondrial |
| 166 | OL588894.1 | Lutrogale perspicillata isolate Penarik_LP_7 D-loop, partial sequence; mitochondrial |
| 167 | OL588893.1 | Lutrogale perspicillata isolate Penarik_LP_10 D-loop, partial sequence; mitochondrial |
| 168 | OL588892.1 | Lutrogale perspicillata isolate Penarik_LP_8 D-loop, partial sequence; mitochondrial |
| 169 | OL588891.1 | Lutrogale perspicillata isolate Penarik_LP_9 D-loop, partial sequence; mitochondrial |
| 170 | OL588890.1 | Lutrogale perspicillata isolate Penarik_LP_12 D-loop, partial sequence; mitochondrial |
| 171 | OL588889.1 | Lutrogale perspicillata isolate Penarik_LP_11 D-loop, partial sequence; mitochondrial |
| 172 | OL588888.1 | Lutrogale perspicillata isolate Penarik_LP_6 D-loop, partial sequence; mitochondrial |
| 173 | OL588887.1 | Lutrogale perspicillata isolate Taman_Negara7_LP_6 D-loop, partial sequence; mitochondrial |
| 174 | OL588886.1 | Lutrogale perspicillata isolate Taman_Negara9_LP_3 D-loop, partial sequence; mitochondrial |
| 175 | OL588885.1 | Lutrogale perspicillata isolate Sg_Mersing_LP_2 D-loop, partial sequence; mitochondrial |
| 176 | OL588884.1 | Lutrogale perspicillata isolate Taman_Negara8_LP_2 D-loop, partial sequence; mitochondrial |
| 177 | OL588883.1 | Lutrogale perspicillata isolate Taman_Negara6_LP_3 D-loop, partial sequence; mitochondrial |
| 178 | OL588882.1 | Lutrogale perspicillata isolate Sg_Mersing_LP_4 D-loop, partial sequence; mitochondrial |
| 179 | OL588881.1 | Lutrogale perspicillata isolate Taman_Negara8_LP_1 D-loop, partial sequence; mitochondrial |
| 180 | OL588880.1 | Lutrogale perspicillata isolate Sg_Mersing_LP_6 D-loop, partial sequence; mitochondrial |
| 181 | OL588879.1 | Lutrogale perspicillata isolate Sg_Mersing_LP_5 D-loop, partial sequence; mitochondrial |
| 182 | OL588878.1 | Lutrogale perspicillata isolate Taman_Negara1_LP_1 D-loop, partial sequence; mitochondrial |
| 183 | OL588877.1 | Lutrogale perspicillata isolate Sg_Mersing_LP_1 D-loop, partial sequence; mitochondrial |
| 184 | OL588876.1 | Lutrogale perspicillata isolate Sg_Mersing_LP_3 D-loop, partial sequence; mitochondrial |
| 185 | OL588875.1 | Lutrogale perspicillata isolate Tg_Piai_LP_8 D-loop, partial sequence; mitochondrial |
| 186 | OL588874.1 | Lutrogale perspicillata isolate Tg_Piai_LP_5 D-loop, partial sequence; mitochondrial |
| 187 | OL588873.1 | Lutrogale perspicillata isolate Tg_Piai_LP_7 D-loop, partial sequence; mitochondrial |
| 188 | OL588872.1 | Lutrogale perspicillata isolate Tg_Piai_LP_6 D-loop, partial sequence; mitochondrial |
| 189 | OL588871.1 | Lutrogale perspicillata isolate Taman_Negara7_LP_4 D-loop, partial sequence; mitochondrial |
| 190 | OL588870.1 | Lutrogale perspicillata isolate Taman_Negara2_LP_1 D-loop, partial sequence; mitochondrial |
| 191 | OL588869.1 | Lutrogale perspicillata isolate Taman_Negara7_LP_5 D-loop, partial sequence; mitochondrial |
| 192 | OL588868.1 | Lutrogale perspicillata isolate Taman_Negara5_LP_2 D-loop, partial sequence; mitochondrial |
| 193 | OL588867.1 | Lutrogale perspicillata isolate Taman_Negara4_LP_1 D-loop, partial sequence; mitochondrial |
| 194 | OL588866.1 | Lutrogale perspicillata isolate Taman_Negara2_LP_2 D-loop, partial sequence; mitochondrial |
| 195 | OL588865.1 | Lutrogale perspicillata isolate Balok_LP_1 D-loop, partial sequence; mitochondrial |
| 196 | OL588864.1 | Lutrogale perspicillata isolate Temerloh_LP_3 D-loop, partial sequence; mitochondrial |
| 197 | OL588863.1 | Lutrogale perspicillata isolate Temerloh_LP_1 D-loop, partial sequence; mitochondrial |
| 198 | OL588862.1 | Lutrogale perspicillata isolate Temerloh_LP_2 D-loop, partial sequence; mitochondrial |
| 199 | OL588861.1 | Lutrogale perspicillata isolate Tg_Piandang8_LP_2 D-loop, partial sequence; mitochondrial |
| 200 | OL588860.1 | Lutrogale perspicillata isolate Tg_Lumpur_LP_1 D-loop, partial sequence; mitochondrial |
| 201 | OL588859.1 | Lutrogale perspicillata isolate Tg_Piandang5_LP_9 D-loop, partial sequence; mitochondrial |
| 202 | OL588858.1 | Lutrogale perspicillata isolate Tg_Piandang9_LP_5 D-loop, partial sequence; mitochondrial |
| 203 | OL588857.1 | Lutrogale perspicillata isolate Tg_Piandang4_LP_7 D-loop, partial sequence; mitochondrial |
| 204 | OL588856.1 | Lutrogale perspicillata isolate Kampar2_LP_3 D-loop, partial sequence; mitochondrial |
| 205 | OL588855.1 | Lutrogale perspicillata isolate Kampar2_LP_9 D-loop, partial sequence; mitochondrial |
| 206 | OL588854.1 | Lutrogale perspicillata isolate Kampar2_LP_5 D-loop, partial sequence; mitochondrial |
| 207 | OL588853.1 | Lutrogale perspicillata isolate Kuala_Gula2_LP_4 D-loop, partial sequence; mitochondrial |
| 208 | OL588852.1 | Lutrogale perspicillata isolate Kampar2_LP_4 D-loop, partial sequence; mitochondrial |
| 209 | OL588851.1 | Lutrogale perspicillata isolate Kampar2_LP_2 D-loop, partial sequence; mitochondrial |
| 210 | OL588850.1 | Lutrogale perspicillata isolate Kampar2_LP_12 D-loop, partial sequence; mitochondrial |
| 211 | OL588849.1 | Lutrogale perspicillata isolate Kampar2_LP_11 D-loop, partial sequence; mitochondrial |
| 212 | OL588848.1 | Lutrogale perspicillata isolate Kuala_Gula2_LP_9 D-loop, partial sequence; mitochondrial |
| 213 | OL588847.1 | Lutrogale perspicillata isolate Kampar2_LP_10 D-loop, partial sequence; mitochondrial |
| 214 | OL588846.1 | Lutrogale perspicillata isolate Kampar2_LP_8 D-loop, partial sequence; mitochondrial |
| 215 | OL588845.1 | Lutrogale perspicillata isolate Kampar2_LP_6 D-loop, partial sequence; mitochondrial |
| 216 | OL588844.1 | Lutrogale perspicillata isolate Kampar2_LP_7 D-loop, partial sequence; mitochondrial |
| 217 | OL588843.1 | Lutrogale perspicillata isolate Kampar2_LP_1 D-loop, partial sequence; mitochondrial |
| 218 | OL588842.1 | Lutrogale perspicillata isolate Kampar1_LP_7 D-loop, partial sequence; mitochondrial |
| 219 | OL588841.1 | Lutrogale perspicillata isolate Kampar1_LP_6 D-loop, partial sequence; mitochondrial |
| 220 | OL588840.1 | Lutrogale perspicillata isolate Kampar1_LP_5 D-loop, partial sequence; mitochondrial |
| 221 | OL588839.1 | Lutrogale perspicillata isolate Kampar1_LP_2 D-loop, partial sequence; mitochondrial |
| 222 | OL588838.1 | Lutrogale perspicillata isolate Kampar1_LP_3 D-loop, partial sequence; mitochondrial |
| 223 | OL588837.1 | Lutrogale perspicillata isolate Kuala_Gula1_LP_16 D-loop, partial sequence; mitochondrial |
| 224 | OL588836.1 | Lutrogale perspicillata isolate Kuala_Gula1_LP_19 D-loop, partial sequence; mitochondrial |
| 225 | OL588835.1 | Lutrogale perspicillata isolate Kampar1_LP_4 D-loop, partial sequence; mitochondrial |
| 226 | OL588834.1 | Lutrogale perspicillata isolate Tg_Piandang8_LP_3 D-loop, partial sequence; mitochondrial |
| 227 | OL588833.1 | Lutrogale perspicillata isolate Kampar1_LP_1 D-loop, partial sequence; mitochondrial |
| 228 | OL588832.1 | Lutrogale perspicillata isolate Kuala_Gula2_LP_2 D-loop, partial sequence; mitochondrial |
| 229 | OL588831.1 | Lutrogale perspicillata isolate Tanjung_Piandang3_LP_5 D-loop, partial sequence; mitochondrial |
| 230 | OL588830.1 | Lutrogale perspicillata isolate Tg_Piandang8_LP_1 D-loop, partial sequence; mitochondrial |
| 231 | OL588829.1 | Lutrogale perspicillata isolate Tg_Piandang9_LP_6 D-loop, partial sequence; mitochondrial |
| 232 | OL588828.1 | Lutrogale perspicillata isolate Kuala_Gula1_LP_15 D-loop, partial sequence; mitochondrial |
| 233 | OL588827.1 | Lutrogale perspicillata isolate Tg_Piandang9_LP_7 D-loop, partial sequence; mitochondrial |
| 234 | OL588826.1 | Lutrogale perspicillata isolate Tg_Piandang7_LP_14 D-loop, partial sequence; mitochondrial |
| 235 | OL588825.1 | Lutrogale perspicillata isolate Tg_Piandang9_LP_4 D-loop, partial sequence; mitochondrial |
| 236 | OL588824.1 | Lutrogale perspicillata isolate Tg_Piandang6_LP_13 D-loop, partial sequence; mitochondrial |
| 237 | OL588823.1 | Lutrogale perspicillata isolate Tanjung_Piandang3_LP_1 D-loop, partial sequence; mitochondrial |
| 238 | OL588822.1 | Lutrogale perspicillata isolate Tg_Piandang6_LP_11 D-loop, partial sequence; mitochondrial |
| 239 | OL588821.1 | Lutrogale perspicillata isolate T_Alam_LP_2 D-loop, partial sequence; mitochondrial |
| 240 | OL588820.1 | Lutrogale perspicillata isolate JUGRA_LP_3 D-loop, partial sequence; mitochondrial |
| 241 | OL588819.1 | Lutrogale perspicillata isolate Tg_Piandang5_LP_8 D-loop, partial sequence; mitochondrial |
| 242 | OL588818.1 | Lutrogale perspicillata isolate Panchang_Bedana1_LP_6 D-loop, partial sequence; mitochondrial |
| 243 | OL588817.1 | Lutrogale perspicillata isolate Tanjung_Piandang3_LP_6 D-loop, partial sequence; mitochondrial |
| 244 | OL588816.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_21 D-loop, partial sequence; mitochondrial |
| 245 | OL588815.1 | Lutrogale perspicillata isolate Tanjung_Piandang3_LP_4 D-loop, partial sequence; mitochondrial |
| 246 | OL588814.1 | Lutrogale perspicillata isolate Tanjung_Piandang3_LP_2 D-loop, partial sequence; mitochondrial |
| 247 | OL588813.1 | Lutrogale perspicillata isolate Tanjung_Piandang3_LP_3 D-loop, partial sequence; mitochondrial |
| 248 | OL588812.1 | Lutrogale perspicillata isolate Tg_Piandang2_LP_5 D-loop, partial sequence; mitochondrial |
| 249 | OL588811.1 | Lutrogale perspicillata isolate T_Alam_LP_3 D-loop, partial sequence; mitochondrial |
| 250 | OL588810.1 | Lutrogale perspicillata isolate Kelanang_LP-2 D-loop, partial sequence; mitochondrial |
| 251 | OL588809.1 | Lutrogale perspicillata isolate Tg_Piandang2_LP_8 D-loop, partial sequence; mitochondrial |
| 252 | OL588808.1 | Lutrogale perspicillata isolate Tg_Piandang2_LP_11 D-loop, partial sequence; mitochondrial |
| 253 | OL588807.1 | Lutrogale perspicillata isolate Paya_Indah2_LP_8 D-loop, partial sequence; mitochondrial |
| 254 | OL588806.1 | Lutrogale perspicillata isolate Panchang_Bedana3_LP_9 D-loop, partial sequence; mitochondrial |
| 255 | OL588805.1 | Lutrogale perspicillata isolate Paya_Indah2_LP_9 D-loop, partial sequence; mitochondrial |
| 256 | OL588804.1 | Lutrogale perspicillata isolate Sg_Dinding_LP_2 D-loop, partial sequence; mitochondrial |
| 257 | OL588803.1 | Lutrogale perspicillata isolate M_Nawar2_LP_5 D-loop, partial sequence; mitochondrial |
| 258 | OL588802.1 | Lutrogale perspicillata isolate Pergau_2_LP_3 D-loop, partial sequence; mitochondrial |
| 259 | OL588801.1 | Lutrogale perspicillata isolate P_Lalang_LP_3 D-loop, partial sequence; mitochondrial |
| 260 | OL588800.1 | Lutrogale perspicillata isolate P_Lalang_LP_1 D-loop, partial sequence; mitochondrial |
| 261 | OL588799.1 | Lutrogale perspicillata isolate Cenderoh_2_LP_2 D-loop, partial sequence; mitochondrial |
| 262 | OL588798.1 | Lutrogale perspicillata isolate P_Lalang_LP_2 D-loop, partial sequence; mitochondrial |
| 263 | OL588797.1 | Lutrogale perspicillata isolate Tumpat_LP_1 D-loop, partial sequence; mitochondrial |
| 264 | OL588796.1 | Lutrogale perspicillata isolate K_Salang_LP_2 D-loop, partial sequence; mitochondrial |
| 265 | OL588795.1 | Lutrogale perspicillata isolate M_Nawar1_LP_2 D-loop, partial sequence; mitochondrial |
| 266 | OL588794.1 | Lutrogale perspicillata isolate Kuala_Besar_LP_3 D-loop, partial sequence; mitochondrial |
| 267 | OL588793.1 | Lutrogale perspicillata isolate Kuala_Besar_LP_1 D-loop, partial sequence; mitochondrial |
| 268 | OL588792.1 | Lutrogale perspicillata isolate Sg_Dinding_LP_3 D-loop, partial sequence; mitochondrial |
| 269 | OL588791.1 | Lutrogale perspicillata isolate Tg_Batu_LP_5 D-loop, partial sequence; mitochondrial |
| 270 | OL588790.1 | Lutrogale perspicillata isolate Tg_Batu_LP_4 D-loop, partial sequence; mitochondrial |
| 271 | OL588789.1 | Lutrogale perspicillata isolate Tg_Batu_LP_3 D-loop, partial sequence; mitochondrial |
| 272 | OL588788.1 | Lutrogale perspicillata isolate T_MUDA_LP_3 D-loop, partial sequence; mitochondrial |
| 273 | OL588787.1 | Lutrogale perspicillata isolate T_MUDA_LP_2 D-loop, partial sequence; mitochondrial |
| 274 | OL588786.1 | Lutrogale perspicillata isolate BELUM1_LP_2 D-loop, partial sequence; mitochondrial |
| 275 | OL588785.1 | Lutrogale perspicillata isolate BELUM2_LP_1 D-loop, partial sequence; mitochondrial |
| 276 | OL588784.1 | Lutrogale perspicillata isolate Merlimau_LP_2 D-loop, partial sequence; mitochondrial |
| 277 | OL588783.1 | Lutrogale perspicillata isolate YAN_LP_1 D-loop, partial sequence; mitochondrial |
| 278 | OL588782.1 | Lutrogale perspicillata isolate Merlimau_LP_1 D-loop, partial sequence; mitochondrial |
| 279 | OL588781.1 | Lutrogale perspicillata isolate M_Nawar2_LP_8 D-loop, partial sequence; mitochondrial |
| 280 | OL588780.1 | Lutrogale perspicillata isolate ALAI_LP_4 D-loop, partial sequence; mitochondrial |
| 281 | OL588779.1 | Lutrogale perspicillata isolate T_MUDA_LP_4 D-loop, partial sequence; mitochondrial |
| 282 | OL588778.1 | Lutrogale perspicillata isolate BELUM1_LP_1 D-loop, partial sequence; mitochondrial |
| 283 | OL588777.1 | Lutrogale perspicillata isolate Tlk_Kampi_LP_3 D-loop, partial sequence; mitochondrial |
| 284 | OL588776.1 | Lutrogale perspicillata isolate BELUM2_LP_2 D-loop, partial sequence; mitochondrial |
| 285 | OL588775.1 | Lutrogale perspicillata isolate Tlk_Kampi_LP_2 D-loop, partial sequence; mitochondrial |
| 286 | OL588774.1 | Lutrogale perspicillata isolate Cenderoh_3_LP_3 D-loop, partial sequence; mitochondrial |
| 287 | OL588773.1 | Lutrogale perspicillata isolate Bayram_LP_1 D-loop, partial sequence; mitochondrial |
| 288 | OL588772.1 | Lutrogale perspicillata isolate Bayram_LP_2 D-loop, partial sequence; mitochondrial |
| 289 | OL588771.1 | Lutrogale perspicillata isolate ALAI_LP_3 D-loop, partial sequence; mitochondrial |
| 290 | OL588770.1 | Lutrogale perspicillata isolate ALAI_LP_1 D-loop, partial sequence; mitochondrial |
| 291 | OL588769.1 | Lutrogale perspicillata isolate Sg_Dinding_LP_4 D-loop, partial sequence; mitochondrial |
| 292 | OL588768.1 | Lutrogale perspicillata isolate M_Nawar2_LP_6 D-loop, partial sequence; mitochondrial |
| 293 | OL588767.1 | Lutrogale perspicillata isolate TB_MERAH_LP_4 D-loop, partial sequence; mitochondrial |
| 294 | OL588766.1 | Lutrogale perspicillata isolate Cenderoh_2_LP_3 D-loop, partial sequence; mitochondrial |
| 295 | OL588765.1 | Lutrogale perspicillata isolate BIKAM_LP_1 D-loop, partial sequence; mitochondrial |
| 296 | OL588764.1 | Lutrogale perspicillata isolate Klinggi_NS_LP_2 D-loop, partial sequence; mitochondrial |
| 297 | OL588763.1 | Lutrogale perspicillata isolate Cenderoh_2_LP_1 D-loop, partial sequence; mitochondrial |
| 298 | OL588762.1 | Lutrogale perspicillata isolate M_Nawar2_LP_7 D-loop, partial sequence; mitochondrial |
| 299 | OL588761.1 | Lutrogale perspicillata isolate Kuala_Gula2_LP_5 D-loop, partial sequence; mitochondrial |
| 300 | OL588760.1 | Lutrogale perspicillata isolate Pergau_2_LP_2 D-loop, partial sequence; mitochondrial |
| 301 | OL588759.1 | Lutrogale perspicillata isolate BELUM2_LP_3 D-loop, partial sequence; mitochondrial |
| 302 | OL588758.1 | Lutrogale perspicillata isolate YAN_LP_2 D-loop, partial sequence; mitochondrial |
| 303 | OL588757.1 | Lutrogale perspicillata isolate Tg_Batu_LP_6 D-loop, partial sequence; mitochondrial |
| 304 | OL588756.1 | Lutrogale perspicillata isolate K_Salang_LP_1 D-loop, partial sequence; mitochondrial |
| 305 | OL588755.1 | Lutrogale perspicillata isolate Klinggi_NS_LP_1 D-loop, partial sequence; mitochondrial |
| 306 | OL588754.1 | Lutrogale perspicillata isolate Tg_Piandang5_LP_10 D-loop, partial sequence; mitochondrial |
| 307 | OL588753.1 | Lutrogale perspicillata isolate ALAI_LP_2 D-loop, partial sequence; mitochondrial |
| 308 | OL588752.1 | Lutrogale perspicillata isolate Temengor2_LP_1 D-loop, partial sequence; mitochondrial |
| 309 | OL588751.1 | Lutrogale perspicillata isolate M_Nawar1_LP_1 D-loop, partial sequence; mitochondrial |
| 310 | OL588750.1 | Lutrogale perspicillata isolate Bkt_Malut_LP_1 D-loop, partial sequence; mitochondrial |
| 311 | OL588749.1 | Lutrogale perspicillata isolate Pokok_Sena_LP_2 D-loop, partial sequence; mitochondrial |
| 312 | OL588748.1 | Lutrogale perspicillata isolate Bkt_Malut_LP_2 D-loop, partial sequence; mitochondrial |
| 313 | OL588747.1 | Lutrogale perspicillata isolate Pokok_Sena_LP_3 D-loop, partial sequence; mitochondrial |
| 314 | OL588746.1 | Lutrogale perspicillata isolate Pokok_Sena_LP_1 D-loop, partial sequence; mitochondrial |
| 315 | OL588745.1 | Lutrogale perspicillata isolate Temengor2_LP_2 D-loop, partial sequence; mitochondrial |
| 316 | OL588744.1 | Lutrogale perspicillata isolate Temengor5_LP_1 D-loop, partial sequence; mitochondrial |
| 317 | OL588638.1 | Lutra sumatrana isolate Tumpat_LS D-loop, partial sequence; mitochondrial |
| 318 | OL588637.1 | Lutra sumatrana isolate Tasik_Bera_LS D-loop, partial sequence; mitochondrial |
| 319 | OL588636.1 | Lutra sumatrana isolate Pontian_LS D-loop, partial sequence; mitochondrial |
| 320 | OL588635.1 | Lutra sumatrana isolate Pekan_LS D-loop, partial sequence; mitochondrial |
| 321 | OL588634.1 | Lutra sumatrana isolate Paya_Indah_LS D-loop, partial sequence; mitochondrial |
| 322 | OL588633.1 | Lutra sumatrana isolate Muazzam_LS D-loop, partial sequence; mitochondrial |
| 323 | OL588632.1 | Lutra sumatrana isolate Gopeng_LS D-loop, partial sequence; mitochondrial |
| 324 | OL588632.1 | Lutra sumatrana isolate Gopeng_LS D-loop, partial sequence; mitochondrial |
| Complete mitochondrial genome | ||
| S.No. | GenBank accession number | Description |
| 1 | KY117556.1 | Lutra sumatrana isolate 49 mitochondrion, complete genome |
| 2 | KY117557.1 | Lutrogale perspicillata isolate 21mitochondrion, complete genome |
| 3 | KY117535.1 | Aonyx cinerea isolate 16 mitochondrion, complete genome |
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Details
; Chee-Yoong, Woo 2 ; Kannan, Adrian 3 ; Suganiya Rama Rao 3 ; Abdul-Patah, Pazil 4 ; Ratnayeke, Shyamala 5 1 German Centre for Integrative Biodiversity Research (iDiv) Halle-Jena-Leipzig, Leipzig, Germany; Institute of Biology, Martin Luther University Halle-Wittenberg, Halle, Germany
2 Department of Biological Sciences, Sunway University, Selangor Darul Ehsan, Malaysia; Malaysian Nature Society, Kuala Lumpur, Malaysia
3 Department of Biological Sciences, Sunway University, Selangor Darul Ehsan, Malaysia
4 Department of Wildlife and National Parks (PERHILITAN), Peninsular Malaysia, Kuala Lumpur, Malaysia
5 Department of Biological Sciences, Sunway University, Selangor Darul Ehsan, Malaysia; Skidmore College, New York, Saratoga Springs, USA