Sperm collection and characteristics analysis of the critically endangered Chinese pangolin (Manis pentadactyla).

The Chinese pangolin (Manis pentadactyla) is a critically endangered species. However, there is a paucity of research on the male reproductive gamete biology of this species. The present study was the first to systematically analyse the sperm characterization of the Chinese pangolin, including semen collection, sperm morphometry and ultrastructure. The semen of five male Chinese pangolins was successfully collected using the electroejaculation method. CASA (computer-assisted sperm analysis) was used to assess semen quality and take images for sperm morphometric analysis. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used for sperm ultrastructure observation. The results showed that the semen of the Chinese pangolin was yellow to pale yellow in colour, viscous, with a fishy odour, and a slightly alkaline pH of between 7.7 and 7.9. The head defects were the main sperm defects; there were 13 kinds of head defects counted in this study. The total sperm length, head length, head width and tail length were 67.62 ± 0.21 µm, 10.47 ± 0.06 µm, 1.33 ± 0.006 µm and 57.16 ± 0.20 µm, respectively. SEM observed that the spermatozoa had a rod-shaped head with a distinct apical ridge, which was different from most mammals and similar to that in avians and reptiles. Interestingly, TEM found that the acrosome membrane of the Chinese pangolin had a double membrane structure rather than a multiple bi-lamellar membrane structure as reported by the previous study. Collectively, this study contributes to the development of artificial breeding efforts and assisted reproductive techniques for the Chinese pangolin, as well as providing technical support for research on germplasm conservation of this species.

Measuring ecosystem services and ecological sensitivity for comprehensive conservation in Giant Panda National Park.

China announced the development of its first 5 national parks in 2021, the primary objective of which is to conserve the natural state and integrity of natural ecosystems. As such, ecosystem services and biodiversity levels are crucial assessment factors for the parks. For Giant Panda National Park (GPNP), we evaluated ecological sensitivity based on water and soil erosion and rocky desertification; ecosystem services based on headwater conservation, soil and water conservation, and biodiversity conservation; and presence of giant panda (Ailuropoda melanoleuca) and sympatric species (e.g., takin [Budorcas taxicolor], Asiatic black bear [Ursus thibetanus]) habitat suitability derived from niche modeling to identify the ecosystem status and assess ecological problems within the park. From our results, we proposed ecologically critical areas to target to meet the park's goals. The suitable habitat for pandas and sympatric species encompassed 62.98% of the park and occurred mainly in the Minshan Mountains. One quarter of the total area (25.67%) contained areas important for ecosystem services. Ecologically sensitive and extremely sensitive areas covered 88.78% of the park and were distributed mainly in Qionglaishan and Minshan Mountains. This coverage indicated that there was much habitat for pandas and sympatric species but that the ecosystems in GPNP are vulnerable. Therefore, ecologically critical areas encompassed all suitable habitats for all the species examined and areas important and extremely important to ecosystem service provision,ecologically sensitive and extremely sensitive areas, encompassed 15.17% of panda habitat, accounted for 16.37% of the GPNP area, and were distributed mainly in the Minshan Mountains. Our results indicated where conservation efforts should be focused in the park and that by identifying ecologically critical areas managers can provide targeted protection for wildlife habitat and ecosystems and effectively and efficiently protect the composite ecosystem. Additionally, our methods can be used to inform development of new national parks.

Medición de los servicios ambientales y la sensibilidad ecológica para una conservación integral en el Parque Nacional del Panda Gigante Resumen China anunció el crecimiento de sus primeros cinco parques nacionales en 2021, con el objetivo principal de conservar el estado natural y la integridad de los ecosistemas naturales. Para ello, los servicios ambientales y los niveles de biodiversidad son factores cruciales de evaluación para los parques. Para poder identificar el estado del ecosistema y evaluar los problemas ecológicos dentro del Parque Nacional del Panda Gigante (PNPG), analizamos la sensibilidad ecológica con base en la erosión del agua y del suelo y la desertificación rocosa; los servicios ambientales con base en el suministro de conservación del agua, del agua y del suelo y de la biodiversidad; y la idoneidad de hábitat del panda gigante (Ailuropoda melanoleuca) y de especies simpátricas (takín [Budorcas taxicolor], oso negro asiático [Ursus thibetanus]) derivada del modelo de nichos. A partir de nuestros resultados proponemos enfocarnos en áreas ecológicamente críticas para lograr los objetivos del parque. El hábitat idóneo para los pandas y las especies simpátricas englobó el 62.98% del parque y se ubicó principalmente en las montañas Minshan. Un cuarto del área total (25.67%) albergó áreas importantes para los servicios ambientales. Las áreas ecológicamente sensibles y extremadamente sensibles cubrieron el 88.78% del parque y se distribuyeron en las montañas Minshan y Qionglaishan. Esta cobertura indica que hay bastante hábitat para los pandas y las especies simpátricas pero que los ecosistemas en el PNPG son vulnerables. Por lo tanto, las áreas ecológicamente críticas englobaron todos los hábitats para todas las especies analizadas y todas las áreas importantes y extremadamente importantes para el suministro de servicios ambientales. Las áreas ecológicamente sensibles y extremadamente sensibles englobaron el 15.17% del hábitat del panda, representaron el 16.37% del área del PNPG y se localizaron principalmente en las montañas Minshan. Nuestros resultados indican en dónde se deben enfocar los esfuerzos de conservación dentro del parque y que, si identificamos las áreas ecológicamente críticas, los gestores pueden proporcionar una protección focalizada para el hábitat y los ecosistemas y así proteger efectiva y eficientemente el ecosistema compuesto. Además, nuestro método puede usarse para guiar el desarrollo de nuevos parques nacionales.

大熊猫国家公园能实现物种和生态完整性的多重保护 中国在2021年宣布设立首批5个国家公园, 保护自然生态系统的真实性和完整性是其优先目标, 而生态系统服务和生物多样性水平是关键的评估要素。大熊猫国家公园作为首批唯一以单一物种命名的国家公园, 通过评估其生态系统服务、生态敏感性, 同时结合大熊猫及同域分布物种(羚牛、亚洲黑熊)的栖息地适宜性, 以揭示其国家公园内生态系统的状态及其面临的问题, 明确其生态关键区以实现多重保护的目标。我们发现大熊猫国家公园包含了超过62.98%的大熊猫和同域物种的适宜栖息地, 主要分布在岷山山系;其次, 大熊猫国家公园包含了25.67%的生态系统服务重要区域和高达88.78%的生态敏感区域, 主要分布在岷山山系和邛崃山山系。这表明尽管大熊猫国家公园内包含了大熊猫及同域物种所需的大面积适宜栖息地, 但是其生态系统具有较强的脆弱性。若将同时包含大熊猫及同域物种的适宜栖息地、生态系统服务重要和极重要区、生态敏感和极敏感的区域定义为生态关键区, 其面积占比为16.37%, 覆盖了15.17%的大熊猫栖息地, 主要分布在岷山山系。因此, 基于生态关键区制定新的科学的、针对性的保护措施, 不仅可以更好的来保护野生动物栖息地和应对生态系统的威胁, 而且也有效且高效地保护多重生态系统。.

The mating system of Himalayan marmots as inferred by microsatellite markers.

The Himalayan marmot Marmota himalayana is widely distributed across the Qinghai-Tibetan Plateau and lives in social groups, yet the mating system of this highly social marmot species is unknown. In this study, the genetic mating system of Himalayan marmots was investigated using microsatellite markers to determine which mating strategies individuals employ. Results revealed that both monogamous and polygamous mating relationships occur in our study population, indicating that the genetic mating system of this marmot species is promiscuity. This study presents the first genetic evidence on the mating system for Himalayan marmots, yet indicates that further studies employing both a genetic and behavioral framework are needed to better understand the social structure and reproductive biology of this marmot species.

Fatty acid metabolization and insulin regulation prevent liver injury from lipid accumulation in Himalayan marmots.

Fat storage and weight gain are dominant traits for hibernating mammals. However, excessive fat accumulation may cause liver damage. Here, we explore the lipid accumulation and metabolic processes of the Himalayan marmot (Marmota himalayana), a hibernating rodent species. We find that the unsaturated fatty acid (UFA) content in food was consistent with a large increase in the body mass of Himalayan marmots. Metagenomic analysis shows that Firmicutes Bacterium CAG:110 plays a synergistic role by synthesizing UFAs, which is demonstrated by fecal transplantation experiments, indicating that the gut microbiome promotes fat storage in Himalayan marmots for hibernation. Microscopic examination results indicate that the risk of fatty liver appears at maximum weight; however, liver function is not affected. Upregulations of UFA catabolism and insulin-like growth factor binding protein genes provide an entry point for avoiding liver injury.

A Fine-Grained Image Classification Approach for Dog Feces Using MC-SCMNet under Complex Backgrounds.

In a natural environment, factors such as weathering and sun exposure will degrade the characteristics of dog feces; disturbances such as decaying wood and dirt are likely to make false detections; the recognition distinctions between different kinds of feces are slight. To address these issues, this paper proposes a fine-grained image classification approach for dog feces using MC-SCMNet under complex backgrounds. First, a multi-scale attention down-sampling module (MADM) is proposed. It carefully retrieves tiny feces feature information. Second, a coordinate location attention mechanism (CLAM) is proposed. It inhibits the entry of disturbance information into the network's feature layer. Then, an SCM-Block containing MADM and CLAM is proposed. We utilized the block to construct a new backbone network to increase the efficiency of fecal feature fusion in dogs. Throughout the network, we decrease the number of parameters using depthwise separable convolution (DSC). In conclusion, MC-SCMNet outperforms all other models in terms of accuracy. On our self-built DFML dataset, it achieves an average identification accuracy of 88.27% and an F1 value of 88.91%. The results of the experiments demonstrate that it is more appropriate for dog fecal identification and maintains stable results even in complex backgrounds, which may be applied to dog gastrointestinal health checks.

Vocal signals with different social or non-social contexts in two wild rodent species (Mus caroli and Rattus losea).

The ultrasonic vocalizations (USVs) of rodents play a substantial role in the communication and interaction between individuals; exhibit a high degree of complexity; and are influenced by a multitude of developmental, environmental, and phylogenetic factors. The functions of USVs are mainly studied in laboratory mice or rats. However, the behavioral relevance of USVs in wild rodents is poorly studied. In this work, we systematically investigated the vocal repertoire of the wild mouse Mus caroli and wild rat Rattus losea in multiple social or non-social contexts, e.g., pup-isolation, juvenile-play, paired opposite-sex encounter, female-female interaction, social-exploring, or foot-shock treatment. Unlike the laboratory mice, M. caroli, whose USVs were recorded during pup-isolation and courtship behaviors, did not produce any vocal sounds during juvenile-play and female-female interactions. R. losea, similar to laboratory rats, emitted USVs in all test situations. We found higher peak frequencies of USVs in both these two wild rodent species than in laboratory animals. Moreover, the parameters and structures of USVs varied significantly across different social or non-social contexts even within each species, confirming the context-sensitivity and complexity of vocal signals in rodents. We also noted a striking difference in call types between these two species: no downward type occurred in M. caroli, but no upward type occurred in R. losea, thereby highlighting the interspecific difference of vocal signals among rodents. Thus, the present study presents behavioral foundations of the vocalization context in wild mice and wild rats, and contributes to revealing the behavioral significance of widely used USVs in rodents.

The Relationship Between Gut Microbiome and Bile Acids in Primates With Diverse Diets.

Primates have evolved a variety of feeding habits and intestinal physiological structure. Gut microbiome act as metabolic organs in many biological processes and play a vital role in adaptation to dietary niches. Gut microbiome also convert primary bile acids (BAs) to secondary. BAs profile and gut microbiome are together influenced by diets and play a significant role in nutrient absorption. The regulation between gut microbiome and BAs metabolism is bidirectional although the relationship in primates consuming diverse diets is still unclear. Here, we investigated gut microbiome structures, fecal BAs profile, and their relationship in primates preferring three distinct diets. We found that gut microbiome communities are well differentiated among dietary groups. Folivorous primates had higher Firmicutes abundance and lower Prevotella to Bacaeroides ratios, possibly related to fiber consumption. Frugivorous primates are colonized predominantly by Prevotella and Bacteroides, pointing to an increased adaptation to high-sugar and simple carbohydrate diets. Likewise, BA profiles differ according to diet in a manner predictable from the known effects of BAs on metabolism. Folivorous primates have high conjugated bile acid levels and low unconjugated to conjugated BA ratios, consistent with their fiber-rich leaf-eating diet. Much of the differentiation in secondary and unconjugated BAs is associated with microbiome composition shifts and individual bile acid concentrations are correlated with the abundance of distinct bacterial taxonomic groups. Omnivores have higher concentrations of secondary BAs, mainly lithocholic acid (LCA). These levels are significantly positively correlated with the presence of Clostrida species, showing that the digestion requirements of omnivores are different from plant-eating primates. In conclusion, gut microbiome and BAs can respond to changes in diet and are associated with nutrient component consumption in each diet primate group. Our study is the first to demonstrate BA profile differentiation among primates preferring diverse diets. BAs thus appear to work with gut microbiome to help primates adapt to their diet.

Epidemiology and Genomic Characterization of Two Novel SARS-Related Coronaviruses in Horseshoe Bats from Guangdong, China.

Severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) and SARS-CoV-2, the causative agents of SARS, which broke out in 2003, and coronavirus disease 2019 (COVID-2019), which broke out in 2019, probably originated in Rhinolophus sinicus and R. affinis, respectively. Rhinolophus bats are important hosts for coronaviruses. Many SARS-related coronaviruses (SARSr-CoVs) have been detected in bats from different areas of China; however, the diversity of bat SARSr-CoVs is increasing, and their transmission mechanisms have attracted much attention. Here, we report the findings of SARSr-CoVs in R. sinicus and R. affinis from South China from 2008 to 2021. The full-length genome sequences of the two novel SARSr-CoVs obtained from Guangdong shared 83 to 88% and 71 to 72% nucleotide identities with human SARS-CoV and SARS-CoV-2, respectively, while sharing high similarity with human SARS-CoV in hypervariable open reading frame 8 (ORF8). Significant recombination occurred between the two novel SARSr-CoVs. Phylogenetic analysis showed that the two novel bat SARSr-CoVs from Guangdong were more distant than the bat SARSr-CoVs from Yunnan to human SARS-CoV. We found that transmission in bats contributes more to virus diversity than time. Although our results of the sequence analysis of the receptor-binding motif (RBM) and the expression pattern of angiotensin-converting enzyme 2 (ACE2) inferred that these viruses could not directly infect humans, risks still exist after some unpredictable mutations. Thus, this study increased our understanding of the genetic diversity and transmission of SARSr-CoVs carried by bats in the field. IMPORTANCE Severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2 probably originated from the SARS-related coronaviruses (SARSr-CoVs) carried by Rhinolophus bats from Yunnan, China. Systematic investigations of the reservoir hosts carrying SARSr-CoVs in Guangdong and the reservoir distribution and transmission are urgently needed to prevent future outbreaks. Here, we detected SARSr-CoV in Rhinolophus bat samples from Guangdong in 2009 and 2021 and found that the transmission of SARSr-CoV from different host populations contributes more to increased virus diversity than time. Bat SARSr-CoVs in Guangdong had genetic diversity, and Guangdong was also the hot spot for SARSr-CoVs. We once again prove that R. sinicus plays an important role in the maintenance of the SARS-CoVs. Besides, the SARSr-CoVs are mainly transmitted through the intestines in bats, and these SARSr-CoVs found in Guangdong could not use human ACE2 (hACE2), but whether they can pass through intermediate hosts or directly infect humans requires further research. Our findings demonstrate the ability of SARSr-CoVs to spread across species.

Captivity Influences the Gut Microbiome of Rhinopithecus roxellana.

Ex situ (captivity in zoos) is regarded as an important form of conservation for endangered animals. Many studies have compared differences in the gut microbiome between captive and wild animals, but few have explained those differences at the functional level due to the limited amount of 16S rRNA data. Here, we compared the gut microbiome of captive and wild Rhinopithecus roxellana, whose high degree of dietary specificity makes it a good subject to observe the effects of the captive environment on their gut microbiome, by performing a metagenome-wide association study (MWAS). The Chao1 index was significantly higher in the captive R. roxellana cohort than in the wild cohort, and the Shannon index of captive R. roxellana was higher than that of the wild cohort but the difference was not significant. A significantly increased ratio of Prevotella/Bacteroides, which revealed an increased ability to digest simple carbohydrates, was found in the captive cohort. A significant decrease in the abundance of Firmicutes and enrichment of genes related to the pentose phosphate pathway were noted in the captive cohort, indicating a decreased ability of captive monkeys to digest fiber. Additionally, genes required for glutamate biosynthesis were also significantly more abundant in the captive cohort than in the wild cohort. These changes in the gut microbiome correspond to changes in the composition of the diet in captive animals, which has more simple carbohydrates and less crude fiber and protein than the diet of the wild animals. In addition, more unique bacteria in captive R. roxellana were involved in antibiotic resistance (Acinetobacter) and diarrhea (Desulfovibrio piger), and in the prevention of diarrhea (Phascolarctobacterium succinatutens) caused by Clostridioides difficile. Accordingly, our data reveal the cause-and-effect relationships between changes in the exact dietary composition and changes in the gut microbiome on both the structural and functional levels by comparing of captive and wild R. roxellana.

Investment in science can mitigate the negative impacts of land use on declining primate populations.

Changes in land use and the conversion of natural forests to agricultural fields and cattle pastures are threatening the survival of many species of wild animals, including nonhuman primates. Given its almost 1.4 billion people, China faces a difficult challenge in balancing economic development, human well-being, environmental protection, and animal conservation. We examined the effects of poverty, anthropogenic land use (cropland and pasture/grazing), human population growth, government investment in science and public attention to primates during the period from the 1980s to 2015 on primate population persistence in China. We analyzed these data using generalized mixed-effects models, structural equation models (SEM) and random forests (a machine learning technique). We found that 16 of 21 (76%) primate species in China, for which data are available, have experienced a population decline over the past 35 years. Factors contributing most to primate population decline included human poverty and the conversion of natural habitat to cropland. In contrast, the five species of primates that were characterized by recent population increases were the subjects of substantial government research funding and their remaining distribution occurs principally in protected areas (PAs). We argue that increased funding for research, the establishment and expansion of PAs, a national policy focused on reducing poverty, and educational programs designed to inform and encourage local people to participate in scientific investigation and wildlife protection, can mitigate the negative impacts of historical patterns of land conversion on primate population survival in China.

Effects of human activity on the habitat utilization of Himalayan marmot (Marmota himalayana) in Zoige wetland.

Human activity is increasingly and persistently disturbing nature and wild animals. Affected wildlife adopts multiple strategies to deal with different human influences. To explore the effect of human activity on habitat utilization of Himalayan marmot (Marmota himalayana), habitat utilization patterns of three neighboring marmot populations in habitats affected differently by human activities were recorded and compared. We found that (a) distance between reproductive burrows (a represent of reproductive pairs) becomes shorter under the influence of human activities, and more burrows were dug as temporary shelters, resulting in shorter distance between those shelters and shorter distance flee to those shelters and, consequently, shorter flight initiation distance when threatened. More burrows that are closer to the disturbed habitats improve the ability to escape from threats. (b) Reproductive burrow site selection of the species is determined by the availability of mounds in the habitat, and breeding pairs selectively build reproductive (also the hibernation) burrows on mounds, potentially to improve surveillance when basking and the drainage of burrows. Human activities generally drive breeding pairs away from the road to dig their reproductive burrows likely to reduce disturbance from vehicles. However, even heavy human activity exerts no pressure on the distance of reproductive burrows from the road or the mound volume of the high disturbance population, potentially because mounds are the best burrowing site to reproduce and hibernate in the habitat. Marmots deal with disturbance by digging more burrows in the habitat to flee more effectively and building reproductive burrows on mounds to gain better vigilance and drainage efficiency.

Convergent Evolution of Himalayan Marmot with Some High-Altitude Animals through ND3 Protein.

The Himalayan marmot (Marmota himalayana) mainly lives on the Qinghai-Tibet Plateau and it adopts multiple strategies to adapt to high-altitude environments. According to the principle of convergent evolution as expressed in genes and traits, the Himalayan marmot might display similar changes to other local species at the molecular level. In this study, we obtained high-quality sequences of the CYTB gene, CYTB protein, ND3 gene, and ND3 protein of representative species (n = 20) from NCBI, and divided them into the marmot group (n = 11), the plateau group (n = 8), and the Himalayan marmot (n = 1). To explore whether plateau species have convergent evolution on the microscale level, we built a phylogenetic tree, calculated genetic distance, and analyzed the conservation and space structure of Himalayan marmot ND3 protein. The marmot group and Himalayan marmots were in the same branch of the phylogenetic tree for the CYTB gene and CYTB protein, and mean genetic distance was 0.106 and 0.055, respectively, which was significantly lower than the plateau group. However, the plateau group and the Himalayan marmot were in the same branch of the phylogenetic tree, and the genetic distance was only 10% of the marmot group for the ND3 protein, except Marmota flaviventris. In addition, some sites of the ND3 amino acid sequence of Himalayan marmots were conserved from the plateau group, but not the marmot group. This could lead to different structures and functional diversifications. These findings indicate that Himalayan marmots have adapted to the plateau environment partly through convergent evolution of the ND3 protein with other plateau animals, however, this protein is not the only strategy to adapt to high altitudes, as there may have other methods to adapt to this environment.

Endozoochorous seed dispersal by golden snub-nosed monkeys in a temperate forest.

Seed dispersal is essential for plant recruitment and the maintenance of biodiversity. Colobine monkeys are primarily folivorous, but they also consume fruits and are often assumed to be seed predators. Although they are known to be epizoochorous seed dispersers, their role as endozoochorous seed dispersers needs reassessment. We examined potential endozoochory in golden snub-nosed monkeys (Rhinopithecus roxellana) at Dalongtan in Shennongjia National Park, central China, by assessing potential germination of ingested seeds (n = 1806, 9 species) from fecal samples. Intact seeds were in almost all fecal samples (ranging from 5-130 seeds), and ingested seeds were from small seeded species (seed width <4.5 mm). The 2 most abundant species were Actinidia arguta (73%) and Rosa caudata (15%). The fruits of A. arguta were unripe when ingested (i.e. effective seed predation) and the ingested seeds did not germinate in the trials. Therefore, ingestion of unripe seeds does not lead to effective seed dispersal. However, germination rates of defecated R. caudata (9%) were greater than control seeds (6% and 0%), demonstrating potential endozoochorous seed dispersal. Thus, colobine monkeys do indeed disperse mainly small-seeded from multi-seeded fruits through potential endozoochory and this process enhances the recruitment of seedlings.

Burrowing and Anti-Predator Requirements Determine the Microhabitat Selection of Himalayan Marmot in Zoige Wetland.

To satisfy their requirements for food and safety, animals need certain habitats to live. Marmots generally select habitats with certain elevation, land surface temperature, soil and vegetation type, and certain mountain slope and aspect; however, what habitats are needed at relatively smaller scales are poorly known. The Himalayan marmot (Marmota himalayana) is distributed mainly on the Qinghai-Tibet Plateau, a region exhibiting diversified topographic features, and the Zoige wetland in the northeast part of the plateau is also the home of the Himalayan marmot. The region is famous for its plateau peat bog, and the suitable habitats for Himalayan marmots are patchily distributed in the wetland. To investigate what kinds of patches are preferred by the marmot in this wetland ecosystem, we measured and compared the soil and vegetation characteristics of used and unused patches. We found that unlike factors governing the habitat selection at macroscales, patches characterized by flat ground and low soil moisture content, with medium vegetation standing height and low vegetation density, are selected in the Zoige wetland. Patches of this kind are selected to meet the marmots' requirements for burrow construction and predator avoidance in such a wetland ecosystem. Together with previous studies on habitat selection of the marmot species at macroscales, we showed that to explore how the animals survive in an environment, it is important to conduct the analysis at multiple scales.