Diet Composition of the Wild Stump-Tailed Macaque(Macaca arctoides) in Perlis State Park, Peninsular Malaysia, Using a Chloroplast tRNL DNA Metabarcoding Approach: A Preliminary Study

Understanding dietary diversity is a fundamental task in the study of stump-tailed macaque, Macaca arctoides in its natural habitat. However, direct feeding observation and morphological identification using fecal samples are not effective and nearly impossible to obtain in natural habitats because this species is sensitive to human presence. As ecological methods are challenging and time-consuming, DNA metabarcoding offers a more powerful assessment of the diet. We used a chloroplast tRNL DNA metabarcoding approach to identify the diversity of plants consumed by free-ranging M. arctoides in the Malaysia–Thailand border region located in Perlis State Park, Peninsular Malaysia. DNA was extracted from three fecal samples, and chloroplast tRNL DNA was amplified and sequenced using the Illumina MiniSeq platform. Sequences were analyzed using the CLC Genomic Workbench software. A total of 145 plant species from 46 families were successfully identified as being consumed by M. arctoides. The most abundant species were yellow saraca, Saraca thaipingensis (11.70%), common fig, Ficus carica (9.33%), aramata, Clathrotropis brachypetala (5.90%), sea fig, Ficus superba (5.44%), and envireira, Malmea dielsiana (1.70%). However, Clathrotropis and Malmea are not considered Malaysian trees because of limited data available from Malaysian plant DNA. Our study is the first to identify plant taxa up to the species level consumed by stump-tailed macaques based on a DNA metabarcoding approach. This result provides an important understanding on diet of wild M. arctoides that only reside in Perlis State Park, Malaysia

Haplotype analysis of long-tailed macaques in TNB Bukit Selambau, Peninsular Malaysia, leads to inferences of fissionfusion social structure

Long-tailed macaques have been roaming near Tenaga National Berhad Bukit Selambau Solar(TBSS) causing human–macaque conflicts. This study reveals the social organization and genetic variation of these macaques. Macaques’ groups were determined via direct observation and closed circuit television. Genomic DNA from 29 fecal samples were extracted and proceeded with amplification of the D-loop region of mitochondrial DNA. There are four main groups of primates at TBSS. Some members of the Kuil group and the Pekan group shared haplotype 1, forming a fission-fusion society. This finding can be used for TNB in mitigation plans involving human– macaque conflict

Metabarcoding data analysis revealed the plant dietary variation of long-tailed macaque Macaca fascicularis (Cercopithecidae, Cercopithecinae) living in disturbed habitats in Peninsular Malaysia

The long-tailed macaque (Macaca fascicularis) has a wide range in both Peninsular Malaysia and Borneo. Although the primates are especially vulnerable to habitat alterations, this primate lives in disturbed habitats due to human-induced land-use. Thus, this study presents a faecal metabarcoding approach to clarify the plant diet of long-tailed macaques from five locations in Peninsular Malaysia to represent fragmented forest, forest edge, island and recreational park habitats. We extracted genomic DNA from 53 long-tailmacaque faecal samples. We found 47 orders, 126 families, 609 genera and 818 species across these five localities. A total of 113 plant families were consumed by long-tailed macaques in Universiti Kebangsaan Malaysia, 61 in the Malaysia Genome and Vaccine Institute, 33 in Langkawi Island, 53 in Redang Island and 44 in the Cenderawasih Cave. Moraceae (33.24%) and Fabaceae (13.63%) were the most common families consumed by long-tailed macaques from the study localities. We found that habitat type impacted diet composition, indicating the flexibility of foraging activities. This research findings provide an understanding of plant dietary diversity and the adaptability of this macaque with the current alteration level that applies to long-tailed macaque conservation management interest in the future. Keywords Malaysian primates, Southeast Asia, DNA metabarcoding, trnL, next-generationsequencin

Molecular phylogeny confirms the subspecies delineation of the Malayan Siamang (Symphalangus syndactylus continentis) and the Sumatran Siamang (Symphalangus syndactylus syndactylus) based on the hypervariable region of mitochondrial DNA

Siamangs (Symphalangus syndactylus) are native to Peninsular Malaysia, Sumatra and southern Thailand and their taxonomical classification at subspecies level remains unclear. Morphologically, two subspecies were proposed as early as 1908 by Thomas namely Symphalangus s. syndactylus and Symphalangus s. continentis. Thus, this study aims to clarify the Siamang subspecies status, based on mtDNA D-loop sequences. Faecal samples were collected from wild Siamang populations at different localities in Peninsular Malaysia. A 600-bp sequence of the mitochondrial D-loop region was amplified from faecal DNA extracts and analysed along with GenBank sequences representing Symphalangus sp., Nomascus sp., Hylobates sp., Hoolock sp. and outgroups (Pongo pygmaeus, Macaca fascicularis and Papio papio). The molecular phylogenetic analysis in this study revealed two distinct clades formed by S. s. syndactylus and S. s. continentis which supports the previous morphological delineation of the existence of two subspecies. Biogeographical analysis indicated that the Sumatran population lineage was split from the Peninsular Malaysian population lineage and a diversification occurrred in the Pliocene era (~ 3.12 MYA) through southward expansion. This postulation was supported by the molecular clock, which illustrated that the Peninsular Malaysian population (~ 1.92 MYA) diverged earlier than the Sumatran population (~ 1.85 MYA). This is the first study to use a molecular approach to validate the subspecies statuses of S. s. syndactylus and S. s. continentis. This finding will be useful for conservation management, for example, during Siamang translocation and investigations into illegal pet trade and forensics involving Malayan and Sumatran Siamangs

Molecular phylogeny confirms the subspecies delineation of the Malayan Siamang (Symphalangus syndactylus continentis) and the Sumatran Siamang (Symphalangus syndactylus syndactylus) based on the hypervariable region of mitochondrial DNA

Siamangs (Symphalangus syndactylus) are native to Peninsular Malaysia, Sumatra and southern Thailand and their taxonomical classification at subspecies level remains unclear. Morphologically, two subspecies were proposed as early as 1908 by Thomas namely Symphalangus s. syndactylus and Symphalangus s. continentis. Thus, this study aims to clarify the Siamang subspecies status, based on mtDNA D-loop sequences. Faecal samples were collected from wild Siamang populations at different localities in Peninsular Malaysia. A 600-bp sequence of the mitochondrial D-loop region was amplified from faecal DNA extracts and analysed along with GenBank sequences representing Symphalangus sp., Nomascus sp., Hylobates sp., Hoolock sp. and outgroups (Pongo pygmaeus, Macaca fascicularis and Papio papio). The molecular phylogenetic analysis in this study revealed two distinct clades formed by S. s. syndactylus and S. s. continentis which supports the previous morphological delineation of the existence of two subspecies. Biogeographical analysis indicated that the Sumatran population lineage was split from the Peninsular Malaysian population lineage and a diversification occurrred in the Pliocene era (~ 3.12 MYA) through southward expansion. This postulation was supported by the molecular clock, which illustrated that the Peninsular Malaysian population (~ 1.92 MYA) diverged earlier than the Sumatran population (~ 1.85 MYA). This is the first study to use a molecular approach to validate the subspecies statuses of S. s. syndactylus and S. s. continentis. This finding will be useful for conservation management, for example, during Siamang translocation and investigations into illegal pet trade and forensics involving Malayan and Sumatran Siamangs

Metabarcoding data analysis revealed the plant dietary variation of long-tailed macaque Macaca fascicularis (Cercopithecidae, Cercopithecinae) living in disturbed habitats in Peninsular Malaysia

The long-tailed macaque (Macaca fascicularis) has a wide range in both Peninsular Malaysia and Borneo. Although the primates are especially vulnerable to habitat alterations, this primate lives in disturbed habitats due to human-induced land-use. Thus, this study presents a faecal metabarcoding approach to clarify the plant diet of long-tailed macaques from five locations in Peninsular Malaysia to represent fragmented forest, forest edge, island and recreational park habitats. We extracted genomic DNA from 53 long-tailed macaque faecal samples. We found 47 orders, 126 families, 609 genera and 818 species across these five localities. A total of 113 plant families were consumed by long-tailed macaques in Universiti Kebangsaan Malaysia, 61 in the Malaysia Genome and Vaccine Institute, 33 in Langkawi Island, 53 in Redang Island and 44 in the Cenderawasih Cave. Moraceae (33.24%) and Fabaceae (13.63%) were the most common families consumed by long-tailed macaques from the study localities. We found that habitat type impacted diet composition, indicating the flexibility of foraging activities. This research findings provide an understanding of plant dietary diversity and the adaptability of this macaque with the current alteration level that applies to long-tailed macaque conservation management interest in the future

Short Communication : Effectiveness of nuclear gene in species and subspecies determination of captive orangutans

Abdul-Manan MN, Mohd-Ridwan AR, Aifat NR, Osman NA, Abdul-Latiff MA, Dharmalingam S, Md-Zain BM. 2020. Short Communication: Effectiveness of nuclear gene in species and subspecies determination of captive orangutans. Biodiversitas 21: 3665- 3669. Genetic identification of captive orangutans is of paramount importance in providing a correct identity that is essential for captive management. Thus, the utility of nuclear DNA sequences was tested in this study to identify the genetic identity of captive orangutans at Bukit Merah Orang Utan Island. Out of 24 DNA samples that were successfully extracted, only 10 orangutan samples were successfully sequenced for the von Willebrand factor (vWF) gene. From the results, this gene was able to separate the genus Pongo at the species level. Distance and character analyses indicated that a clear separation between P. pygmaeus and P. abelii at the species level. However, the degree of separation at species level was indicated in tree topology with moderate bootstrap values. At the subspecies level of P. pygmaeus, this gene was unable to show a clear separation between three Bornean subspecies. All the subspecies were formed clade together with each other. The vMF gene is a good nuclear gene for the study of phylogenetic relationships of orangutans in captivity at the species level, but the genetic identification at subspecies level in the genus level remains unclear. We suggest that future studies should involve multiple independent nuclear markers to increase the probability of getting reliable results

Haplotype analysis of long-tailed macaques in TNB Bukit Selambau, Peninsular Malaysia, leads to inferences of fissionfusion social structure

Long-tailed macaques have been roaming near Tenaga National Berhad Bukit Selambau Solar (TBSS) causing human–macaque conflicts. This study reveals the social organization and genetic variation of these macaques. Macaques’ groups were determined via direct observation and closedcircuit television. Genomic DNA from 29 fecal samples were extracted and proceeded with amplification of the D-loop region of mitochondrial DNA. There are four main groups of primates at TBSS. Some members of the Kuil group and the Pekan group shared haplotype 1, forming a fission-fusion society. This finding can be used for TNB in mitigation plans involving human– macaque conflict