Fecal metagenomics for the simultaneous assessment of diet, parasites, and population genetics of an understudied primate.

BACKGROUND: Rapid habitat loss and degradation are responsible for population decline in a growing number of species. Understanding the natural history of these species is important for designing conservation strategies, such as habitat enhancements or ex-situ conservation. The acquisition of observational data may be difficult for rare and declining species, but metagenomics and metabarcoding can provide novel kinds of information. Here we use these methods for analysing fecal samples from an endangered population of a colobine primate, the banded leaf monkey (Presbytis femoralis). RESULTS: We conducted metagenomics via shotgun sequencing on six fecal samples obtained from a remnant population of P. femoralis in a species-rich rainforest patch in Singapore. Shotgun sequencing and identification against a plant barcode reference database reveals a broad dietary profile consisting of at least 53 plant species from 33 families. The diet includes exotic plant species and is broadly consistent with > 2 years of observational data. Metagenomics identified 15 of the 24 plant genera for which there is observational data, but also revealed at least 36 additional species. DNA traces for the diet species were recovered and identifiable in the feces despite long digestion times and a large number of potential food plants within the rainforest habitat (>700 species). We also demonstrate that metagenomics provides greater taxonomic resolution of food plant species by utilizing multiple genetic markers as compared to single-marker metabarcoding. In addition, full mitochondrial genomes of P. femoralis individuals were reconstructed from fecal metagenomic shotgun reads, showing very low levels of genetic diversity in the focal population, and the presence of gut parasites could also be confirmed. Metagenomics thus allows for the simultaneous assessment of diet, population genetics and gut parasites based on fecal samples. CONCLUSIONS: Our study demonstrates that metagenomic shotgun sequencing of fecal samples can be successfully used to rapidly obtain natural history data for understudied species with a complex diet. We predict that metagenomics will become a routinely used tool in conservation biology once the cost per sample reduces to ~100 USD within the next few years.

Is Yuan in China's Three Gorges a Gibbon or a Langur?

Clarifying the scientific identity of ancient biological names in historical archives is essential to understand traditional knowledge and literary metaphors of animals in human culture. Adopting a cross-disciplinary (Primatology, Linguistics, Historiography, Historical Sociology) analysis, we developed a theoretical framework for studies of the scientific identity of Chinese primate traditional names (e.g., Yuan ) throughout history, and interpret the historical evolution of the understanding of the Chinese word Yuan. Presently, the Chinese generally understand Yuan to be a gibbon (or "ape" in a broader sense), but this statement has many contradictions with the understanding of the word in relevant historical discourse. We review and comment on key evidence to support the traditional understanding of Yuan as a gibbon (Hylobatidae) and clarify the historical and current thought concerning Yuan. We find that the referent of the word Yuan has changed from "François' langur (Trachypithecus francoisi) with long limbs" to the "long-armed ape or gibbon" known today through two major changes in the idea of Yuan. One transformation in the conceptualization of Yuan took place during the Tang-Song period, with the other beginning at the end of the nineteenth century and ending in the 1950s. An interaction between the conceptualization of animals and power (e.g., political opportunity; cultural movement toward learning western sciences in the semi-colonial era) played an important role in these two diachronic changes to the idea of Yuan. In contrast to the clear linear relation between a species and its Latin name, our study indicates that one traditional name can represent varying animal species in China. Our findings exemplify the implications of the sociocultural and linguistic basis for the species identification of primate names found in historical discourse for historical zoogeography, our understanding of the intricate cultural and religious connections between humans and primates, and efforts to decolonize primatology. Supplementary Information: The online version contains supplementary material available at 10.1007/s10764-022-00302-1.

Three decades of Philippine nonhuman primate studies: research gaps and opportunities for Philippine primatology.

The Philippines is considered a megadiverse country. However, there are few published studies on its nonhuman primate (NHP) species (Carlito syrichta, Macaca fascicularis, and Nycticebus menagensis). Understanding the nature and extent of existing NHP research in the Philippines would help us to determine research gaps and opportunities. Thus, studies on NHPs of Philippine origin were retrieved from online databases including Google Scholar, ResearchGate, Primate Lit, the Integrated Taxonomic Information System (ITIS), and relevant authors. Of a total of 107 studies on Philippine NHP from 1989 to 2019, most focused on Philippine long-tailed macaque (52.78%), and Philippine tarsier (42.59%), with far fewer on Philippine slow loris (4.63%). Fewer studies were based on fieldwork (26.17%); more were based on research on captive animals or that undertaken in the laboratory (34.58%), or used only existing specimens or data (33.64%). Studies on wild NHPs were mostly conducted in Bohol Island. In terms of the type of research, studies on diseases (38.60%) were the most prevalent for macaques; ecological studies (23.91%) for tarsiers; evolutionary, taxonomic, and systematic studies (40%), and ecological and natural history studies (40%) for lorises. Philippine researchers and collaborations with foreign researchers contributed fewer studies (43.93%) than foreign-only researchers (56.07%). In conclusion, although research on Philippine NHPs is increasing, there is a bias with regards to the species studied, the type of research, and study location. Conservation-driven studies are also lacking. These gaps offer opportunities for further research to identify threats to the survival of NHPs in the Philippines, and for the development of plans and effective strategies for their conservation.

Faecal DNA to the rescue: Shotgun sequencing of non-invasive samples reveals two subspecies of Southeast Asian primates to be Critically Endangered species.

A significant number of Southeast Asian mammal species described in the 19th and 20th century were subsequently synonymized and are now considered subspecies. Many are affected by rapid habitat loss which creates an urgent need to re-assess the conservation status based on species boundaries established with molecular data. However, such data are lacking and difficult to obtain for many populations and subspecies. We document via a literature survey and empirical study how shotgun sequencing of faecal DNA is a still underutilized but powerful tool for accelerating such evaluations. We obtain 11 mitochondrial genomes for three subspecies in the langur genus Presbytis through shotgun sequencing of faecal DNA (P. femoralis femoralis, P. f. percura, P. siamensis cf. cana). The genomes support the resurrection of all three subspecies to species based on multiple species delimitation algorithms (PTP, ABGD, Objective Clustering) applied to a dataset covering 40 species and 43 subspecies of Asian colobines. For two of the newly recognized species (P. femoralis, P. percura), the results lead to an immediate change in IUCN status to Critically Endangered due to small population sizes and fragmented habitats. We conclude that faecal DNA should be more widely used for clarifying species boundaries in endangered mammals.

Current conservation status of Germain's langur (Trachypithecus germaini) in Vietnam.

Following the split of the silvered langurs of Indochina into two species based on molecular and phenotypic data, there is a need to reevaluate their distribution and update their conservation status. Here, we report the distribution and assess the population size of Germain's langur (Trachypithecus germaini) within its known range across Vietnam. We confirmed this species at six of seven survey sites in different habitats within three provinces in the Mekong Delta Region, including semi-evergreen forest at the Seven Mountains of An Giang Province, mangrove forest in Ngoc Hien and Nam Can Districts and Melaleuca forest in U Minh Ha National Park of Ca Mau Province, and limestone forest at Kien Luong Karst Area and semi-evergreen and evergreen forests at Phu Quoc National Park of Kien Giang Province. We found no evidence of this species in Mui Ca Mau National Park, Ca Mau Province where it was previously reported. We conservatively estimate that the total population of Germain's langurs in Vietnam consists of 362-406 individuals, with the largest population found in the Kien Luong Karst Area. Hunting and habitat loss are severely impacting Germain's langur, resulting in the extirpation of the population in Mui Ca Mau National Park and small, isolated populations in the Seven Mountains and Ngoc Hien and Nam Can Districts. However, the ability of this species to inhabit a wide range of forest types, and its increasing population sizes in Phu Quoc National Park and Kien Luong Karst Area, provide signs of hope that continued conservation actions may help in its long-term survival.

No evidence for mitochondrial genetic variability in the largest population of critically endangered Tonkin snub-nosed monkeys in Vietnam.

The Tonkin snub-nosed monkey (Rhinopithecus avunculus) with a global population of <250 is listed as critically endangered. It is endemic to northeastern Vietnam and was feared extinct until its rediscovery in 1989. The largest single population of R. avunculus consists of 125-130 individuals in an area of forest called Khau Ca in Ha Giang Province. We used non-invasively collected fecal samples to establish the amount of genetic diversity in this population based on mitochondrial information. We amplified and sequenced a 467- to 650-bp section of the hypervariable region I (HVI) of the mitochondrial D-loop for 201 samples and reconstructed the full mitochondrial genomes for five samples based on metagenomic data. All 201 HVI sequences were identical and no variability was found in the five mitochondrial genomes. Our results highlight the immediate need for a comprehensive assessment of the genetic diversity of all populations of R. avunculus based on mitochondrial and nuclear markers. The latter need to be developed for this species.