Joko Pamungkas

Effects of Pleistocene glaciations and rivers on the population structure of Bornean orangutans (Pongo pygmaeus)

Sundaland, a tropical hotspot of biodiversity comprising Borneo and Sumatra among other islands, the Malay Peninsula, and a shallow sea, has been subject to dramatic environmental processes. Thus, it presents an ideal opportunity to investigate the role of environmental mechanisms in shaping species distribution and diversity. We investigated the population structure and underlying mechanisms of an insular endemic, the Bornean orangutan (Pongo pygmaeus). Phylogenetic reconstructions based on mtDNA sequences from 211 wild orangutans covering the entire range of the species indicate an unexpectedly recent common ancestor of Bornean orangutans 176 ka (95% highest posterior density, 72–322 ka), pointing to a Pleistocene refugium. High mtDNA differentiation among populations and rare haplotype sharing is consistent with a pattern of strong female philopatry. This is corroborated by isolation by distance tests, which show a significant correlation between mtDNA divergence and distance and a strong effect of rivers as barriers for female movement. Both frequency-based and Bayesian clustering analyses using as many as 25 nuclear microsatellite loci revealed a significant separation among all populations, as well as a small degree of male-mediated gene flow. This study highlights the unique effects of environmental and biological features on the evolutionary history of Bornean orangutans, a highly endangered species particularly vulnerable to future climate and anthropogenic change as an insular endemic.

Sex-Biased Dispersal and Volcanic Activities Shaped Phylogeographic Patterns of Extant Orangutans (genus: Pongo)

The Southeast Asian Sunda archipelago harbors a rich biodiversity with a substantial proportion of endemic species. The evolutionary history of these species has been drastically influenced by environmental forces, such as fluctuating sea levels, climatic changes, and severe volcanic activities. Orangutans (genus: Pongo), the only Asian great apes, are well suited to study the relative impact of these forces due to their well-documented behavioral ecology, strict habitat requirements, and exceptionally slow life history. We investigated the phylogeographic patterns and evolutionary history of orangutans in the light of the complex geological and climatic history of the Sunda archipelago. Our study is based on the most extensive genetic sampling to date, covering the entire range of extant orangutan populations. Using data from three mitochondrial DNA (mtDNA) genes from 112 wild orangutans, we show that Sumatran orangutans, Pongo abelii, are paraphyletic with respect to Bornean orangutans (P. pygmaeus), the only other currently recognized species within this genus. The deepest split in the mtDNA phylogeny of orangutans occurs across the Toba caldera in northern Sumatra and, not as expected, between both islands. Until the recent past, the Toba region has experienced extensive volcanic activity, which has shaped the current phylogeographic patterns. Like their Bornean counterparts, Sumatran orangutans exhibit a strong, yet previously undocumented structuring into four geographical clusters. However, with 3.50 Ma, the Sumatran haplotypes have a much older coalescence than their Bornean counterparts (178 kya). In sharp contrast to the mtDNA data, 18 Y-chromosomal polymorphisms show a much more recent coalescence within Sumatra compared with Borneo. Moreover, the deep geographic structure evident in mtDNA is not reflected in the male population history, strongly suggesting male-biased dispersal. We conclude that volcanic activities have played an important role in the evolutionary history of orangutans and potentially of many other forest-dwelling Sundaland species. Furthermore, we demonstrate that a strong sex bias in dispersal can lead to conflicting patterns in uniparentally inherited markers even at a genus-wide scale, highlighting the need for a combined usage of maternally and paternally inherited marker systems in phylogenetic studies.

Elucidating geological and biological processes underlying the diversification of Sulawesi tarsiers

Because of their exceptionally long independent evolution, a range diminution of their Eocene relatives, and a remarkable subsequent diversification in Southeast Asia, tarsiers are of particular importance to evolutionary primatologists. Little is known, however, on the processes shaping the radiation of these small enigmatic primates—especially on the Indonesian island of Sulawesi, their center of endemism. Geological reconstructions and progress in applying DNA sequence information to divergence dating now provide us with the tools and background to comprehend tarsier dispersal. Here, we describe effects of plate-tectonic movements, Pleistocene sea level changes, and hybridization on the divergence of central Sulawesi tarsiers. We analyzed 12 microsatellites, the cytochrome b gene, the hypervariable region I of the mitochondrial control region, and the sex-determining region on the Y-chromosome from 144 specimens captured along a transect crossing a species boundary and a contact zone between 2 microplates. Based on these differentially inherited genetic markers, geographic information, and recordings of vocalizations, we demonstrate that the species boundary coincides with a tectonic suture. We estimate the most recent common ancestor of the 2 taxa to have lived 1.4 Mya, we describe asymmetrical introgressive hybridization, and we give evidence of unbiased dispersal in one species and male-biased dispersal in another species. This study exemplifies that the distribution of tarsier acoustic forms on Sulawesi is consistent with the allocation of genetic variability and that plate-tectonic and glacial events have left traceable marks in the biogeography of this islands unique fauna.

Possible role of mother-daughter vocal interactions on the development of species-specific song in gibbons.

Mother-infant vocal interactions play a crucial role in the development of human language. However, comparatively little is known about the maternal role during vocal development in nonhuman primates. Here, we report the first evidence of mother-daughter vocal interactions contributing to vocal development in gibbons, a singing and monogamous ape species. Gibbons are well known for their species-specific duets sung between mates, yet little is known about the role of intergenerational duets in gibbon song development. We observed singing interactions between free-ranging mothers and their sub-adult daughters prior to emigration. Daughters sang simultaneously with their mothers at different rates. First, we observed significant acoustic variation between daughters. Co-singing rates between mother and daughter were negatively correlated with the temporal precision of the song’s synchronization. In addition, songs of daughters who co-sang less with their mothers were acoustically more similar to the maternal song than any other adult female’s song. All variables have been reported to be influenced by social relationships of pairs. Therefore those correlations would be mediated by mother-daughter social relationship, which would be modifiable in daughter’s development. Here we hypothesized that daughters who co-sing less often, well-synchronize, and converge acoustically with the maternal acoustic pattern would be at a more advanced stage of social independence in sub-adult females prior to emigration. Second, we observed acoustic matching between mothers and daughters when co-singing, suggesting short-term vocal flexibility. Third, we found that mothers adjusted songs to a more stereotyped pattern when co-singing than when singing alone. This vocal adjustment was stronger for mothers with daughters who co-sang less. These results indicate the presence of socially mediated vocal flexibility in gibbon sub-adults and adults, and that mother-daughter co-singing interactions may enhance vocal development. More comparative work, notably longitudinal and experimental, is now needed to clarify maternal roles during song development.

Marked Population Structure and Recent Migration in the Critically Endangered Sumatran Orangutan (Pongo abelii)

A multitude of factors influence how natural populations are genetically structured, including dispersal barriers, inhomogeneous habitats, and social organization. Such population subdivision is of special concern in endangered species, as it may lead to reduced adaptive potential and inbreeding in local subpopulations, thus increasing the risk of future extinctions. With only 6600 animals left in the wild, Sumatran orangutans (Pongo abelii) are among the most endangered, but also most enigmatic, great ape species. In order to infer the fine-scale population structure and connectivity of Sumatran orangutans, we analyzed the most comprehensive set of samples to date, including mitochondrial hyper-variable region I haplotypes for 123 individuals and genotypes of 27 autosomal microsatellite markers for 109 individuals. For both mitochondrial and autosomal markers, we found a pronounced population structure, caused by major rivers, mountain ridges, and the Toba caldera. We found that genetic diversity and corresponding long-term effective population size estimates vary strongly among sampling regions for mitochondrial DNA, but show remarkable similarity for autosomal markers, hinting at male-driven long-distance gene flow. In support of this, we identified several individuals that were most likely sired by males originating from other genetic clusters. Our results highlight the effect of natural barriers in shaping the genetic structure of great ape populations, but also point toward important dispersal corridors on northern Sumatra that allow for genetic exchange.

Call cultures in orang-utans?

Background Several studies suggested great ape cultures, arguing that human cumulative culture presumably evolved from such a foundation. These focused on conspicuous behaviours, and showed rich geographic variation, which could not be attributed to known ecological or genetic differences. Although geographic variation within call types (accents) has previously been reported for orang-utans and other primate species, we examine geographic variation in the presence/absence of discrete call types (dialects). Because orang-utans have been shown to have geographic variation that is not completely explicable by genetic or ecological factors we hypothesized that this will be similar in the call domain and predict that discrete call type variation between populations will be found. Methodology/Principal Findings We examined long-term behavioural data from five orang-utan populations and collected fecal samples for genetic analyses. We show that there is geographic variation in the presence of discrete types of calls. In exactly the same behavioural context (nest building and infant retrieval), individuals in different wild populations customarily emit either qualitatively different calls or calls in some but not in others. By comparing patterns in call-type and genetic similarity, we suggest that the observed variation is not likely to be explained by genetic or ecological differences. Conclusion/Significance These results are consistent with the potential presence of ‘call cultures’ and suggest that wild orang-utans possess the ability to invent arbitrary calls, which spread through social learning. These findings differ substantially from those that have been reported for primates before. First, the results reported here are on dialect and not on accent. Second, this study presents cases of production learning whereas most primate studies on vocal learning were cases of contextual learning. We conclude with speculating on how these findings might assist in bridging the gap between vocal communication in non-human primates and human speech.

Parentage-based pedigree reconstruction reveals female matrilineal clusters and male-biased dispersal in nongregarious Asian great apes, the Bornean orang-utans (Pongo pygmaeus)

Philopatry and sex‐biased dispersal have a strong influence on population genetic structure, so the study of species dispersal patterns and evolutionary mechanisms shaping them are of great interest. Particularly nongregarious mammalian species present an underexplored field of study: despite their lower levels of sociality compared to group‐living species, interactions among individuals do occur, providing opportunities for cryptic kin selection. Among the least gregarious primates are orang‐utans (genus: Pongo), in which preferential associations among females have nevertheless been observed, but for which the presence of kin structures was so far unresolved because of the equivocal results of previous genetic studies. To clarify relatedness and dispersal patterns in orang‐utans, we examined the largest longitudinal set of individuals with combined genetic, spatial and behavioural data. We found that males had significantly higher mitochondrial DNA (mtDNA) variation and more unique haplotypes, thus underscoring their different maternal ancestries compared to females. Moreover, pedigree reconstruction based on 24 highly polymorphic microsatellite markers and mtDNA haplotypes demonstrated the presence of three matrilineal clusters of generally highly related females with substantially overlapping ranges. In orang‐utans and possibly other nongregarious species, comparing average biparental relatedness (r) of males and females to infer sex‐biased dispersal is extremely problematic. This is because the opportunistic sampling regime frequently employed in nongregarious species, combined with overlapping space use of distinct matrilineal clusters, leads to a strong downward bias when mtDNA lineage membership is ignored. Thus, in nongregarious species, correct inferences of dispersal can only be achieved by combining several genetic approaches with detailed spatial information.

The role of traditional beliefs in conservation of herpetofauna in Banten, Indonesia

Social taboos have been increasingly recognized for their role in determining human behaviour. Such informal institutions may also, in some instances, guide practices that serve as effective conservation measures. Here we present a case in Banten, Indonesia, where a local taboo has discouraged the collection of two herpetofaunal species, the water monitor lizard Varanus salvator and the reticulated python Python reticulatus , on Tinjil Island, an undeveloped island off the coast of Java. The taboo is not observed in the nearby mainland villages of Muara Dua and Cisiih, where the two species may be harvested for skin or meat, and where the water monitor may also be killed as a pest. Water monitors and reticulated pythons figure prominently in the international reptile leather trade, with skins produced from Indonesias wild populations representing the highest percentage of total global exports of both lizard and snake skins. The site-specific taboo documented here provides a strong deterrent to collection of these species in a location where they could be subject to illicit harvest as populations in nearby mainland areas decrease. Preliminary evidence also suggests that belief in forest guardian spirits may extend protection to other wildlife species on Tinjil Island.

HPLC-based metabolomics to identify cytotoxic compounds from Plectranthus amboinicus (Lour.) Spreng against human breast cancer MCF-7Cells

The objective of this study was to identify the active compounds in Plectranthus amboinicus (Lour.) Spreng which play a role to inhibit viability of breast cancer MCF-7 cells using HPLC-based metabolomics approach. Five fractions of the plant extract were observed including ethanol, hexane, chloroform, ethyl acetate and water fraction. There were 45 HPLC chromatograms resulted from 5 fractions with 3 replications and 3 wavelengths detection. The chromatograms were compared to the data of IC50 from MTT assay of each fraction against human breast cancer MCF-7 cells using metabolomics. The OPLS analysis result promptly pointed towards a chloroform fraction at retention time of 40.16-41.28min that has the greatest contribution to the cytotoxic activity. The data of mass spectra indicated that an abietane diterpene namely 7-acetoxy-6-hydroxyroyleanone was the main compound that contributed to the cytotoxic activity. This metabolomics application method can be used as a quick preliminary guideline to uncover the most dominant compound related to the bioactivity.

Therapeutic Vaccine Against Primate Papillomavirus Infections of the Cervix

Currently available prophylactic vaccines have no therapeutic efficacy for preexisting human papillomavirus (HPVs) infections, do not target all oncogenic HPVs and are insufficient to eliminate the burden of HPV induced cancer. We aim to develop an alternative HPV vaccine which is broadly effective and capable of clearing preexisting infection. In an initial attempt to develop a broadly reactive therapeutic vaccine, we designed a putative papillomavirus (PV) ancestor antigen (circulating sequence derived antigenic sequences E1E2—CDSE1E2) based on the conserved E1 and E2 protein sequences from existing oncogenic HPV strains. This antigen was found to be as related to circulating oncogenic Macaca fascicularis papillomaviruses (MfPVs) as to oncogenic HPVs. The CDSE1E2 antigen was fused to a T-cell adjuvant and encoded in chimpanzee 3 and 63 adenoviral vectors. We first showed that the combination of these 2 vaccines induced long-lasting potent CDSE1E2 specific T cell responses in outbred mice. This prime-boost regimen was then tested in female macaques naturally infected with MfPVs. All immunized animals (16/16) responded to the vaccine antigen but with reduced cross-reactivity against existing PVs. Preexisting MfPV infections did not prime vaccine inducible immune responses. Importantly, immunized oncogenic MfPV type 3 (MfPV3) infected animals that responded toward MfPV3 were able to diminish cervical MfPV3 DNA content. Although insufficient breadth was achieved, our results suggest that a relevant level of E1E2 specific T cell immunity is achievable and might be sufficient for the elimination of PV infection. Importantly, naturally infected macaques, offer a relevant model for testing vaccines aimed at eliminating mucosal PV infections.