Manis Javanica Research Articles

Molecular identification of tick-borne Rickettsia, Anaplasma, Ehrlichia, Babesia, and Colpodella in confiscated Malayan pangolins.

The traditional concept of "tonic food" and demand for traditional Chinese medicine make pangolins the largest population of illegally smuggled mammals in the world. Illegal hunting and trade are not only responsible for the sharp decline in pangolin populations but also provide conditions for pathogenic transmission. In 2021, we rescued 21 confiscated unhealthy Malayan pangolins, none of which survived. This study aimed to investigate the reasons for their unexpected deaths and the potential pathogens that may be transmitted during smuggling. Physical examination found that more than 80% pangolins were parasitized with A. javanense ticks. Autopsy and pathological staining analysis revealed multiple organ damage in the deceased pangolins. Pathogens nucleic acid detection of 33 tick samples showed that the positive rate of Rickettsia spp., Anaplasma spp., Ehrlichia spp. Babesia spp., and Colpodella spp. were 90.91%, 6.06%, 6.06%, 15.15% and 18.18%, respectively. Furthermore, pangolin samples were positive for Rickettsia spp. (42.86%, 9/21), Ehrlichia sp. (4.76%, 1/21), and Babesia sp. (4.76%, 1/21). This study confirmed that spotted fever triggered by Rickettsia spp. from A. javanense might accelerate the most death of confiscated pangolins, while Ehrlichia sp., and Babesia sp. infection potentially accelerating a few deaths. Of note, A. javanense ticks carrying Colpodella spp. were detected for the first time in Malayan pangolins. However, whether Colpodella spp. are pathogenic to pangolins is unknown. Further research on the diagnosis, treatment, surveillance, and elimination of ticks and tick-borne diseases in humans, livestock, and wildlife should provide insight into wildlife conservation and zoonotic disease prevention.

ACE2-independent sarbecovirus cell entry can be supported by TMPRSS2-related enzymes and can reduce sensitivity to antibody-mediated neutralization.

The COVID-19 pandemic, caused by SARS-CoV-2, demonstrated that zoonotic transmission of animal sarbecoviruses threatens human health but the determinants of transmission are incompletely understood. Here, we show that most spike (S) proteins of horseshoe bat and Malayan pangolin sarbecoviruses employ ACE2 for entry, with human and raccoon dog ACE2 exhibiting broad receptor activity. The insertion of a multibasic cleavage site into the S proteins increased entry into human lung cells driven by most S proteins tested, suggesting that acquisition of a multibasic cleavage site might increase infectivity of diverse animal sarbecoviruses for the human respiratory tract. In contrast, two bat sarbecovirus S proteins drove cell entry in an ACE2-independent, trypsin-dependent fashion and several ACE2-dependent S proteins could switch to the ACE2-independent entry pathway when exposed to trypsin. Several TMPRSS2-related cellular proteases but not the insertion of a multibasic cleavage site into the S protein allowed for ACE2-independent entry in the absence of trypsin and may support viral spread in the respiratory tract. Finally, the pan-sarbecovirus antibody S2H97 enhanced cell entry driven by two S proteins and this effect was reversed by trypsin while trypsin protected entry driven by a third S protein from neutralization by S2H97. Similarly, plasma from quadruple vaccinated individuals neutralized entry driven by all S proteins studied, and availability of the ACE2-independent, trypsin-dependent pathway reduced neutralization sensitivity. In sum, our study reports a pathway for entry into human cells that is ACE2-independent, can be supported by TMPRSS2-related proteases and may be associated with antibody evasion.

Structural basis for human DPP4 receptor recognition by a pangolin MERS-like coronavirus.

Middle East respiratory syndrome coronavirus (MERS-CoV) and the pangolin MERS-like coronavirus MjHKU4r-CoV-1 employ dipeptidyl peptidase 4 (DPP4) as an entry receptor. MjHKU4r-CoV-1 could infect transgenic mice expressing human DPP4. To understand the mechanism of MjHKU4r-CoV-1 entry into cells, we determined the crystal structures of the receptor binding domain (RBD) of MjHKU4r-CoV-1 spike protein bound to human DPP4 (hDPP4) and Malayan pangolin DPP4 (MjDPP4), respectively. The overall hDPP4-binding mode of MjHKU4r-CoV-1 RBD is similar to that of MERS-CoV RBD. MjHKU4r-CoV-1 RBD shows higher binding affinity to hDPP4 compared to the bat MERS-like coronavirus Ty-BatCoV-HKU4. Via swapping residues between MjHKU4r-CoV-1 RBD and Ty-BatCoV-HKU4 RBD, we identified critical determinants on MjHKU4r-CoV-1 that are responsible for virus usage of hDPP4. Our study suggests that MjHKU4r-CoV-1 is more adapted to the human receptor compared to the bat HKU4 coronavirus and highlights the potential of virus emergence into the human population.

Georeferencing Sunda pangolin Manis javanica records in Singapore

Abstract In Singapore, the Critically Endangered Sunda pangolin Manis javanica is threatened by habitat loss and fragmentation, and road traffic collisions. To mitigate these threats, an understanding of its spatiotemporal distribution is needed, as identified in the National Conservation Strategy and Action Plan for the species. However, Sunda pangolin occurrence data are held in multiple separate databases, are typically collected using non-standardized methods, and often lack accurate location details. To compile a complete georeferenced database of Sunda pangolin records in Singapore, we consolidated occurrence data from heterogeneous databases and mainstream and social media, and converted locality descriptions into geographical coordinates. We demonstrate the use of this database to analyse data on rescued pangolins and those killed on roads, to aid conservation efforts in Singapore, and describe other potential applications. We georeferenced 482 records of pangolin sightings, rescues and roadkill for 1996–2021, finding an increase in all three over the study period. Roadkill and rescues occurred mostly in central and western Singapore, close to forested areas, and were predominantly of subadults and adult males. The data can be used to inform threat mitigation strategies, post-rescue release plans and further research. The database has already been used in practice, contributing to environmental impact assessments and conservation recommendations. Overall, this georeferenced database demonstrates the value of citizen science and collating wildlife data from multiple sources, and the methods used can be applied to other taxa to aid conservation strategies.

Proteomics and Microbiota Conjoint Analysis in the Nasal Mucus: Revelation of Differences in Immunological Function in Manis javanica and Manis pentadactyla.

All eight pangolin species, especially captive Manis pentadactyla, are critically endangered and susceptible to various pathogenic microorganisms, causing mass mortality. They are involved in the complement system, iron transport system, and inflammatory factors. M. pentadactyla exhibited a higher abundance of opportunistic pathogens, Moraxella, which potentially evaded complement-mediated immune response by reducing C5 levels and counteracting detrimental effects through transferrin neutralization. In addition, we found that the major structure of C5a, an important inflammatory factor, was lacking in M. javanica. In brief, this study revealed the differences in immune factors and microbiome between M. javanica and M. pentadactyla, thus providing a theoretical basis for subsequent immunotherapy.

Detection of tick-borne pathogens in the pangolin tick, Amblyomma javanense, from Vietnam and Laos, including a novel species of Trypanosoma

Two species of Southeast Asian pangolins (the Chinese pangolin, Manis pentadactyla and the Malayan or Sunda pangolin, Manis javanica) are critically endangered species. Therefore, knowledge on their parasitic infections is very important, especially considering ticks that can transmit which pathogens. In this study, 32 pangolin ticks (Amblyomma javanense), that were collected in Vietnam and Laos, were analyzed with molecular methods for the presence of tick-borne pathogens. Two members of the family Anaplasmataceae were shown to be present in 14 pangolin ticks, i.e., Candidatus Anaplasma pangolinii and an Ehrlichia sp. In three ticks, a single Rickettsia genotype was also detected, and in seven ticks four 18S rRNA sequence variants of a Babesia sp. Most importantly, a novel protozoan agent, tentatively called here Trypanosoma sp. “PAT14” was detected in one A. javanense nymph. These results imply the first molecular finding of any species of Anaplasma, Ehrlichia and Babesia in pangolin ticks from Vietnam and Laos. On the other hand, detection of a new tick-associated Trypanosoma sp. in A. javanense from Southeast Asia is not only important from a taxonomic point of view, but it is also the first finding of any trypanosomes in the genus Amblyomma in Eurasia and adds pangolins to the potential placental mammalian hosts of any trypanosomes.

Conservation, trade and (lack of) management of Sunda pangolins in Bali and Lombok

Context Limited information is available on the occurrence, conservation and management of Sunda pangolins (Manis javanica) on Bali and Lombok, Indonesia. Some have even questioned their presence here. Aims We present an overview of the current knowledge of pangolins on Bali and Lombok. Methods Information was drawn from camera traps (2023), animal market surveys (1994–2024), and review of occurrence data (museum specimens, iNaturalist, literature), rescues and releases, and seizure data. Key results We document the presence of wild pangolins from 11 sites in Bali (1994–2023) and 5 sites in Lombok (1939–2022). We document 8 rescues from Bali (2015–2023) but none from Lombok; there have been 11 releases of single pangolins in the forest near the Luhur Besi Kalung temple in Bali (2016–2023). We did not record, or find any records of, pangolins offered for sale in four animal markets in Bali (1994–2024) and Lombok (2018–2024), but there was one seizure of 15 live pangolins in Bali in 2009, and, unusually, limited evidence of trade from Lombok and Bali. Conclusions There is a paucity of data of pangolins on Lombok, and on Bali the species appears to occur scattered throughout the island, with much of the contemporary information being derived from accidental encounters or rescues and active management of pangolins is largely absent. Implications Compared to other parts of Indonesia, the conservation status of Sunda pangolins on Bali and Lombok is less bleak, and despite their small size, these two islands on the easternmost edge of the species’ range may hold important populations.

Mapping the distribution of the Sunda pangolin (Manis javanica) within natural forest in Sabah, Malaysian Borneo

Pangolins are the most trafficked mammals in the world and are severely threatened by poaching the loss, degradation, and fragmentation of habitat. In Malaysian Borneo, conservation initiatives for the Sunda pangolin (Manis javanica) are hindered by a paucity of data on their distribution and population size. Using MaxEnt niche modelling and consolidated species location data, we projected the distribution of Sunda pangolins in Sabah. Additionally, we assessed the accessibility of their forest habitats to humans to understand potential threats. Our model indicated that, as of 2015, approximately half of Sabah’s land area (39,530 km²) is suitable for pangolins, with 43% in protected forests, 38% in production forests, and 19% outside of these areas. Alarmingly, our data suggest that nearly all (91%) of these suitable habitats are relatively easily accessible to poachers. Our findings provide a state-level baseline understanding of Sunda pangolin distribution and assess potential threats in Sabah. These can inform short- and long-term conservation management plans for pangolin to safeguard this critically endangered species.

Characterization of a pangolin SARS-CoV-2-related virus isolate that uses the human ACE2 receptor.

Various SARS-CoV-2-related coronaviruses have been increasingly identified in pangolins, showing a potential threat to humans. Here we report the infectivity and pathogenicity of the SARS-CoV-2-related virus, PCoV-GX/P2V, which was isolated from a Malayan pangolin (Manis javanica). PCoV-GX/P2V could grow in human hepatoma, colorectal adenocarcinoma cells, and human primary nasal epithelial cells. It replicated more efficiently in cells expressing human angiotensin-converting enzyme 2 (hACE2) as SARS-CoV-2 did. After intranasal inoculation to the hACE2-transgenic mice, PCoV-GX/P2V not only replicated in nasal turbinate and lungs, but also caused interstitial pneumonia, characterized by infiltration of mixed inflammatory cells and multifocal alveolar hemorrhage. Existing population immunity established by SARS-CoV-2 infection and vaccination may not protect people from PCoV-GX/P2V infection. These findings further verify the hACE2 utility of PCoV-GX/P2V by in vivo experiments using authentic viruses and highlight the importance for intensive surveillance to prevent possible cross-species transmission.

Detection of a novel Babesia sp. in Amblyomma javanense, an ectoparasite of Sunda pangolins

BackgroundBabesia is a protozoal, tick-borne parasite that can cause life-threatening disease in humans, wildlife and domestic animals worldwide. However, in Southeast Asia, little is known about the prevalence and diversity of Babesia species present in wildlife and the tick vectors responsible for its transmission. Recently, a novel Babesia species was reported in confiscated Sunda pangolins (Manis javanica) in Thailand. To investigate the presence of this parasite in Singapore, we conducted a molecular survey of Babesia spp. in free-roaming Sunda pangolins and their main ectoparasite, the Amblyomma javanense tick.MethodsTicks and tissue samples were opportunistically collected from live and dead Sunda pangolins and screened using a PCR assay targeting the 18S rRNA gene of Babesia spp. DNA barcoding of the cytochrome oxidase subunit I (COI) mitochondrial gene was used to confirm the species of ticks that were Babesia positive.ResultsA total of 296 ticks and 40 tissue samples were obtained from 21 Sunda pangolins throughout the 1-year study period. Babesia DNA was detected in five A. javanense ticks (minimum infection rate = 1.7%) and in nine different pangolins (52.9%) located across the country. Phylogenetic analysis revealed that the Babesia 18S sequences obtained from these samples grouped into a single monophyletic clade together with those derived from Sunda pangolins in Thailand and that this evolutionarily distinct species is basal to the Babesia sensu stricto clade, which encompasses a range of Babesia species that infect both domestic and wildlife vertebrate hosts.ConclusionsThis is the first report documenting the detection of a Babesia species in A. javanense ticks, the main ectoparasite of Sunda pangolins. While our results showed that A. javanense can carry this novel Babesia sp., additional confirmatory studies are required to demonstrate vector competency. Further studies are also necessary to investigate the role of other transmission pathways given the low infection rate of ticks in relation to the high infection rate of Sunda pangolins. Although it appears that this novel Babesia sp. is of little to no pathogenicity to Sunda pangolins, its potential to cause disease in other animals or humans cannot be ruled out.Graphical abstract

Pangolin HKU4-related coronaviruses found in greater bamboo bats from southern China

Coronavirus (CoV) spillover originating from game animals, particularly pangolins, is currently a significant concern. Meanwhile, vigilance is urgently needed for coronaviruses carried by bats, which are known as natural reservoirs of many coronaviruses. In this study, we collected 729 anal swabs of 20 different bat species from nine locations in Yunnan and Guangdong provinces, southern China, in 2016 and 2017, and described the molecular characteristics and genetic diversity of alphacoronaviruses (αCoVs) and betacoronaviruses (βCoVs) found in these bats. Using RT-PCR, we identified 58 (8.0%) bat CoVs in nine bat species from six locations. Furthermore, using the Illumina platform, we obtained two representative full-length genomes of the bat CoVs, namely TyRo-CoV-162275 and TyRo-CoV-162269. Sequence analysis showed that TyRo-CoV-162275 shared the highest identity with Malayan pangolin (Manis javanica) HKU4-related coronaviruses (MjHKU4r-CoVs) from Guangxi Province, whereas TyRo-CoV-162269 was closely related to HKU33-CoV discovered in a greater bamboo bat (Tylonycteris robustula) from Guizhou Province. Notably, TyRo-CoV-162275 has a putative furin protease cleavage site in its S protein and is likely to utilize human dipeptidyl peptidase-4 (hDPP4) as a cell-entry receptor, similar to MERS-CoV. To the best of our knowledge, this is the first report of a bat HKU4r-CoV strain containing a furin protease cleavage site. These findings expand our understanding of coronavirus geographic and host distributions.

Visualisation and detection of latent DNA deposited by pangolin scales onto plastic packaging materials

We report on the detection and visualisation of latent DNA from pangolin scales deposited onto a plastic packaging material through the use of a nucleic acid staining dye. This latent DNA deposited by pangolin scales was subsequently isolated and analysed using DNA barcoding method. Pangolins are the most illegally traded mammalian species due to the demand for their scales and meat. The demand for their scales were mostly fuelled by its use in traditional medicines. The scales are usually packed into bags and transported globally via sea routes. This is the first report detailing the detection of trace latent DNA from processed wildlife products, on surfaces of bags that they were packaged in. Prior to this report, it was not known if the dried pangolin scales contained transferable quantities of biological material for DNA analyses. To address this, scales were removed from a roadkill Sunda pangolin (Manis javanica), processed by drying and packaged into one of five plastic bags. The presence of pangolin latent DNA was detected on the surface of the plastic bags and visualised using Diamond™ nucleic acid dye. Swabs were then used to recover the stained biological material from various locations in the five bags. The DNA was isolated and quantified using a newly designed quantitative PCR (qPCR) specific to M. javanica to amplify a fragment of the mitochondrial DNA cytochrome b gene. There was a positive correlation between the number of stained particles and DNA quantity, and a greater number of stained particles were found at the bottom of the bag than were found at the top. Conventional PCR targeting part of the cyt b gene amplified a product from all 30 samples taken from the bags and in all cases, sequence data generated matched that of the Sunda pangolin, as expected. All negative controls yielded no results. The method described here is the very first use of a nucleic acid staining dye to detect latent DNA from a mammalian species, other than humans, and highlights the opportunity for further use of Diamond™ nucleic acid dye in wildlife forensic science. It is anticipated that this method will be invaluable in retrieving latent DNA deposited by wildlife products from the environment in which they were contained, to determine the presence of these illegal wildlife products even when previously hidden, inaccessible, or no longer present physically. Further research is required to understand if the use on non-human mammalian wildlife species is feasible.

The Effects of Anthropogenic Disturbances on the Spatiotemporal Patterns of Medium-Large Mammals in Tropical Volcanic Landscapes.

A comprehensive understanding of the consequences of human interactions with mammals is a critical factor in supporting and conserving species in landscapes dominated by humans, which are increasingly threatened. This study aimed to identify the spatial and temporal interactions between humans and mammals. A non-parametric statistical approach with kernel density was used to detect human-mammal temporal interactions. The species interaction factor (SIF) was applied to calculate the spatial overlap based on the two-species occupancy detection model. The activity patterns of medium mammals were nocturnal, diurnal, and cathemeral. The human-medium mammal pairs with SIF values that were <1 and statistically significant included the human-long-tailed macaque (Macaca fascicularis) pair, the human-leopard cat (Prionailurus bengalensis) pair, and the human-barking deer (Muntiacus muntjac) pair. Based on their SIF values and the high overlap in their activity times, the human-macaque pairings had a high risk of conflict. Barking deer and leopard cats displayed a coexistence with humans via time-sharing activities. Due to temporal niche variations with human activities, the existence of nocturnal mammals was relatively uninterrupted. This study showed that most mammals are able to adapt spatially and temporally to various human activities. Nonetheless, efforts to mitigate human-wildlife conflict must be maintained, particularly in the case of severely endangered species, such as the Sunda pangolin.

Genome sequence analysis of Malayan pangolin (Manis javanica) forensic samples reveals the presence of Paraburkholderia fungorum sequences.

The Malayan pangolin (Manis javanica) is a placental mammal and is listed as Critically Endangered on the IUCN Red List of Threatened Species. Most previous attempts to breed pangolins in captivity have met with little success because of dietary issues, infections, and other complications, although a previous study reported breeding pangolins in captivity to the third generation. In our previous pangolin genome sequencing data analysis, we obtained a considerable amount of bacterial DNA from a pregnant female Malayan pangolin (named "UM3"), which was likely infected by Paraburkholderia fungorum-an agent of biodegradation and bioremediation in agriculture. Here, we further confirmed and characterized this bacterial species using PCR, histological staining, whole-genome sequencing, and bioinformatics approaches. PCR assays with in-house designed primer sets and 16S universal primers showed clear positive bands in the cerebrum, cerebellum, lung, and blood of UM3 suggesting that UM3 might have developed septicaemia. Histological staining showed the presence of Gram-negative rod-shaped bacteria in the pangolin brain and lungs, indicating the colonization of the bacteria in these two organs. In addition, PCR screening of UM3's fetal tissues revealed the presence of P. fungorum in the gastrocnemius muscle, but not in other tissues that we examined. We also sequenced and reconstructed the genome of pangolin P. fungorum, which has a genome size of 7.7 Mbps. Our study is the first to present detailed evidence of the presence of P. fungorum in a pangolin and her fetus (although preliminary results were presented in our previous article). Here, we raise the concern that P. fungorum may potentially infect humans, especially YOPI (young, old, pregnant, and immunocompromised) people. Therefore, caution should be exercised when using this bacterial species as biodegradation or bioremediation agents in agriculture.

DIAGNOSTIC IMAGING AND OROTRACHEAL INTUBATION IN CLINICALLY HEALTHY WILD SUNDA PANGOLINS (MANIS JAVANICA) IN SINGAPORE.

Pangolins are amongst the most overexploited species in the world and all eight species of pangolins are threatened with extinction. These animals are rare in zoological collections and often suffer high mortality rates in captivity. Maintaining healthy populations in captivity has become more important with the declining populations in the wild, but knowledge of veterinary care of these animals is limited. Interpreting radiography and ultrasonography images in a patient can be challenging without knowledge of normal findings. The Wildlife Healthcare and Research Center (WHRC) at Mandai Wildlife Reserve (MWR) admits an average of 25 Sunda pangolins (Manis javanica) annually, which is a Critically Endangered species native to Singapore. All the pangolins are triaged on admission and anesthetized for a health assessment before release into the wild. Endotracheal intubation using an otoscope and stylet is a novel technique in the species that has been developed and is commonly performed with these pangolins. A retrospective study was done on 20 clinically healthy wild pangolins to determine normal ultrasonography and radiography findings in the species. Notable findings include the presence of radio-opaque particles in the stomachs of all pangolins, the presence of free fluid cranial to the left kidney (13/20), a spleen with multifocal hypoechoic regions (6/20), and open epiphyseal plates of long bones even in large individuals weighing as heavy as 6.8 kg. Ultrasonographic images and measurements of kidney, spleen, and adrenal gland sizes as well as intestinal, gallbladder, and urinary bladder wall thickness were also described. These diagnostic imaging findings can advance the veterinary care of captive and wild pangolins.

The knowledge, attitude and practice of local Sabahan in Malaysia on Sunda pangolin

The IUCN Red List of Threatened Species listed Sunda pangolin (Manis javanica) as critically endangered and remained among the most traded mammals and other pangolin species. In Malaysia, various conservation works on this species have been conducted these past years through government and NGO works. Nevertheless, their population is still declining despite all the efforts. To date, little to nothing is known about the local perception regarding the Sunda pangolin conservation. Therefore, through the Knowledge, Attitude, and Practice (KAP) approach, this study evaluates Sabahan’s awareness level regarding the Sunda pangolin conservation. A total of 109 respondents were interviewed from all across Malaysia to investigate their perception of Sunda Pangolin conservation. The KAP index score shows that respondents’ attitude toward Sunda Pangolin conservation is the highest (93.34%), followed by Knowledge (81.15%) and Practice (79.63%), of which all three fall within good KAP score. They also agreed that spreading awareness and being proactive in supporting conservation work helps to prevent this species from extinction. Interestingly, the correlation analysis using Spearman rho’s coefficient shows a positive relationship between KA and education but not for Practice. Generally, Malaysians had a positive understanding of the Sunda pangolin’s current status and the legal information regarding this species. This research may help improve species conservation strategies, particularly in Sabah, Malaysia. There is an urgent need to improve the Sunda pangolin conservation program in the future. Factors that influence Practice needs to be identified and scrutinized. Public awareness is not the only factor that needs to be considered for improvement in conservation works. More importantly, the people who work in the conservation field.

Presence of SARS-CoV-2-like coronaviruses in bats from east coast Malaysia.

Most of the public health importance coronaviruses, such as Severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV) and SARS-CoV-2 are likely originated from bats and spread to humans through intermediate hosts; civet cats, dromedary camel and Malayan pangolin, respectively. SARS-CoV-2-like coronaviruses were detected in Thailand, which is neighbouring with Kelantan in East Coast Malaysia. To date, there is no report on the presence of public health concerns (SARS-CoV, SARS-CoV-2 and MERS-CoV) coronaviruses in bats from Malaysia. This study was aimed to elucidate the presence of these coronaviruses in bat samples from East Coast, Malaysia. A total of hundred seventy oropharyngeal swab samples were collected from three states of East Coast Malaysia. Reverse Transcription-Polymerase Chain Reaction (RT-PCR) was conducted based on partial 3' Untranslated region (3'UTR) or ORF10 gene and the products were sequenced. The sequences were compared with all coronavirus sequences from the National Center for Biotechnology Information-GenBank (NCBI-GenBank) using NCBI-Basic Local Alignment Search Tool (NCBI-BLAST) software. A phylogenetic tree was constructed to determine the genetic relationship among the detected coronaviruses with the reference coronaviruses from the NCBI-GenBank. Our results showed that SARSCoV-2-like viruses were present in 3% (5/170) of the bats from East Coast Malaysia that have 98-99% sequence identities and are genetically related to SARS-CoV-2 from humans. This finding indicates the presence of SARS-CoV-2-like viruses in bats from East Coast Malaysia that may become a public health concern in the future.

Haplotype diversity of Pangolin (Manis javanica Desmarest, 1822) based on the COI gene

The Pangolin (Manis javanica) is a unique scaly mammal with the IUCN status as a critically endangered species. In Indonesia, M. javanica is distributed in Java, Sumatra, and Kalimantan. The population of M. javanica has been declined due to the high poaching. M. javanica has been reported present in some locations in West Sumatra however, there is no biological data, especially molecular data. Therefore, it is necessary to conduct the haplotype diversity study of M. javanica using the CO1 gene. Hair samples of M. javanica were collected during the release of M. javanica in the Biology Education and Research Forest (HPPB) of Andalas University. The result of the study was obtained 601 bp of the CO1 gene for analysis. Two haplogroups were obtained from five sequences in West Sumatra and 15 Genbank sequences. The value of haplotype diversity was 0.995. The value of nucleotide diversity (π) in the haplogroup ranged from 0.042 to 0.047. The value of nucleotide diversity (π) between haplogroups ranged from 0.0148 to 0.0161. Overall, the values indicate the low genetic diversity of pangolin (M. javanica). Therefore, it is necessary to maintain and increase the genetic diversity of M. javanica within haplogroups.

Genomic analysis reveals a cryptic pangolin species

Eight extant species of pangolins are currently recognized. Recent studies found that two mitochondrial haplotypes identified in confiscations in Hong Kong could not be assigned to any known pangolin species, implying the existence of a species. Here, we report that two additional mitochondrial haplotypes identified in independent confiscations from Yunnan align with the putative species haplotypes supporting the existence of this mysterious species/population. To verify the new species scenario we performed a comprehensive analysis of scale characteristics and 138 whole genomes representing all recognized pangolin species and the cryptic new species, 98 of which were generated here. Our morphometric results clearly attributed this cryptic species to Asian pangolins (Manis sp.) and the genomic data provide robust and compelling evidence that it is a pangolin species distinct from those recognized previously, which separated from the Philippine pangolin and Malayan pangolin over 5 Mya. Our study provides a solid genomic basis for its formal recognition as the ninth pangolin species or the fifth Asian one, supporting a new taxonomic classification of pangolins. The effects of glacial climate changes and recent anthropogenic activities driven by illegal trade are inferred to have caused its population decline with the genomic signatures showing low genetic diversity, a high level of inbreeding, and high genetic load. Our finding greatly expands current knowledge of pangolin diversity and evolution and has vital implications for conservation efforts to prevent the extinction of this enigmatic and endangered species from the wild.

Home ranges and activity patterns of Sunda pangolins Manis javanica (Pholidota: Manidae) in Vietnam

This study provides an update on the movement ecology of the Sunda pangolin (Manis javanica) in three distinct ecoregions of Vietnam. Using radio telemetry, we tracked 12 pangolins rescued from the illegal wildlife trade and translocated to protected areas. We found that the average home range of translocated Sunda pangolins was 1.58 km2 and did not vary significantly between sexes, mass, or habitat. However, there was a marked decrease in range size with decreasing size of suitable forest space and terrain, or localized abundance of resources in some locations. Pangolins were most active from 03:00 to 04:00. The study highlights the need for further research to fully understand the ecology of this unique and enigmatic species.