中国科学院昆明动物研究所机构知识库

对于分布于同一区域的不同物种，由于进化历史和生活习性的不同，其遗传结构有可能存在着差异。通过比较谱系地理研究，可以重建在过去气候变化和地理隔离的影响下同域分布物种的演化，以预测环境因子如何对区域生物多样性空间分布、遗传结构、种群动态产生影响。喜马拉雅地区经历了大尺度气候变迁，特别是在更新世，并且地形因河流系统而产生巨大改变，进而形成了独特的景观、生态系统和生物区系。然而喜马拉雅地区地理独特性对动植物区系遗传结构所产生的影响至今仍未得到很好的说明。因此，本研究拟以中喜马拉雅地区（尼泊尔）同域分布的熊猴（Macaca assamensis McClelland 1840）和哈努曼叶猴（Semnopithecus entellus Dufresne 1797）为研究对象，探究其时空分布、遗传多样性、群体遗传结构、进化历史和系统发育及其区域河流系统及历史气候变化所产生的影响。中（尼泊尔）喜马拉雅地区流淌着3大水系，从东到西分别为东部克溪水系、中部甘达基水系和西部利河－马哈卡利水系。为解决前述的提到科学问题，我们在尼泊尔3大水系中沿9条主要支流的两岸进行了野外调查，调查范围基本覆盖了两个物种的分布范围，调查其种群数量与分布，采集相关群体的粪便样品（无损伤采样），其中采集到熊猴样品277份，涉及39个群体；采集到哈努曼叶猴样品87份，涉及18个群体。从粪便样品中提取出总DNA，扩增并测定线粒体（D-loop、CYTB、COI和16S rRNA）和Y染色体（TSPY和SRY）基因片段序列。野外调查结果表明：熊猴的分布区包含在哈努曼叶猴分布区之内，哈努曼叶猴作为普适者，其分布范围从热带低地塔莱直至海拔3800 m以上，而熊猴仅分布在西瓦利克、中山区及从喜马拉雅山脉山脚至海拔2650 m；两个物种的种群大小在海拔1500 m以下较大，但其在海拔梯度分布上呈现明显的差异，但是种群组成在河流系统间和海拔梯度上没有明显差异。另外，基于Maxent模型的生态位预测结果表明：在潜在温室气体排放的效应下，熊猴将经历分布区变换和适宜栖息地减少的气候变化影响。通过对尼泊尔地区熊猴种群遗传多样性的分析，结果检出54个单倍型，其线粒体D-loop基因表现出较低的核苷酸多样性（0.0075 ±SD 0.0001）和高单倍型多样性（0.965 ±SD 0.004）。贝叶斯聚类分析将39个群体样品分成了7个大类群，尽管缺少较高的支持率。空间上的隔离对熊猴的群体遗传结构有显著影响，而水系的隔离则影响不明显。对于同域分布的哈努曼叶猴，共检出35个单倍型，其线粒体基因表现出高的核苷酸多样性 (0.0298 ±SD 0.0009) 和单倍型多样性 (0.957 ±SD 0.015)。18个群体聚成6个类群，与河流隔离的空间分布相一致，相关类群间表现出了较高的遗传变异，揭示喜马拉雅地区河流对哈努曼叶猴遗传的影响。基于RAxML和BEAST进行系统发育分析和古分布重建的进化历史分析结果表明：中（尼泊尔）喜马拉雅地区两种同域分布灵长类动物的种群动态和空间扩散分布模式不一致。熊猴尼泊尔种群进化时间较短，经历了末次盛冰期小的奠基种群的扩张。相反，哈努曼种群进化时间较长，该种群自早更新世便生存于中喜马拉雅地区，在末次盛冰期种群数量有所下降，并于全新世开始恢复。更新世气候变化对熊猴的种群遗传和种群历史动态变化的影响更甚于河流阻隔的影响，而哈努曼叶猴受到的影响则刚好相反。在末次盛冰期，熊猴种群缩小到尼泊尔半山区，而现在海拔梯度方向上双向扩张，并在经向上向东部和纬向上向北部高海拔地区扩张。哈努曼叶猴在末次盛次期虽然向低海拔迁移，但其分布面积并没有太大改变，现今正在向高海拔单向扩张。基于线粒体和Y染色体多基因位点的分析，在猕猴属Sinica种组中群体的系统发育分析表明，熊猴尼泊尔种群与不丹种群、达旺猴（M. munzala）和白颊猕猴（M. leucogenys）构成了单系群，但与东部的不丹亚种 (M. assamensis assamensis) 构成并系。熊猴尼泊尔种群与其单系群最近共同祖先的分化约在1.62 Mya前，且是最早从Sinica种组中分化出来的种群。基因树的拓扑结构和分化时间估算结果均显著支持熊猴尼泊尔种群是一个独立分化的物种。而系统发育分析表明尼泊尔哈努曼叶猴3个支系与基于形态学进行的亚种划分（Semnopithecus entellus hector，S. e. ajax 和 S. e. schistaceus）和地理分布相吻合：，其最大似然分析和贝叶斯分析均表现出较高的节点支持率。综上，基于线粒体和核基因多基因联合数据集，熊猴和哈努曼叶猴的尼泊尔种群系统发育分析结果佐证了其形态学、地理分布和行为学证据。针对该两物种，尼泊尔中山区仍然是它们的最主要适宜生境，尽管其大部分区域并不在自然保护区范围内，因此对该段区域的保护亟待加强，而基于多算法的生态位模拟也需要被应用到评估气候变化对该两个物种将来生存与分布的影响。 

Riverine barriers and climatic fluctuations may account inequitably on genetic structuring among different taxa. Comparative phylogeography of co-distributed species allows reconstruction of evolutionary changes under the influence of past climatic fluctuations and contemporary geophysical barriers, and thus can be useful for predicting how these factors have genetic, demographic, and spatial influence on biodiversity. Himalayan region experienced large-scale climatic fluctuations, especially during the Pleistocene, and the landmass is fragmented by numerous rivers. However, the influence of such rivers and the past climatic fluxes at shaping the genetic variations of the terrestrial biota in Nepal Himalaya are not documented yet. Therefore, this study aimed to explore the spatial and temporal distribution pattern, genetic diversity, population genetic structure, evolutionary history and phylogeny of sympatric primates- Assam macaque (Macaca assamensis McClelland 1840) and Hanuman Langur (Semnopithecus entellus Dufresne 1797) in Nepal Himalaya. Field surveys were conducted along both sides of nine major tributaries; three from each of Koshi River system in eastern, Gandaki River system in central, and Karnali-Mahakali River system in western Nepal covering almost the entire distribution range of both the species; and detailed population censuses were carried out. A total of 277 fecal samples from 39 wild troops of Assam macaque and 87 fecal samples from 18 wild troops of Hanuman Langurs were collected non-invasively. Genomic DNA was extracted from fecal samples, and mitochondrial (D-loop, CYTB, COI & 16S rRNA) and Y-chromosomal DNA (TSPY & SRY) loci were amplified and sequenced. Coalescent based phylogeographical analyses were complemented with ecological niche modeling that introduced spatial-geographic perspectives to statistical phylogeography. Further, phylogenetic analyses were performed using maximum likelihood and Bayesian inference methods in RAxML and BEAST, respectively. This study revealed the distribution range of Assam macaque is included within that of the Hanuman Langur. Hanuman Langur is distributed from tropical lowland Tarai up to almost 3800 m asl (above sea level) as the habitat generalist, whereas, Assam macaque is confined to subtropical and temperate climates of Siwaliks, mid-hills and foot-hills of Himalaya up to 2650 m asl. Population sizes of both the species are skewed to the elevations below 1500 m asl and have significant difference in distribution along the elevational gradients. However, there was no significant difference in their average troop size and age-sex composition among the river systems and elevational zones. The ecological niche modeling and future projection based on the principle of niche conservatism revealed that Assam macaque will experience climate change induced range shift and decline of suitable habitat, consistently in all the potential greenhouse emission scenarios (Representative Concentration Pathways, RCPs) defined by International Panel on Climate Change.The Assam macaque population in Nepal defined 54 haplotypes with low nucleotide diversity (0.0075 ±SD 0.0001) but high haplotype diversity (0.965 ±SD 0.004) of mtDNA D-loop. Bayesian clustering analysis clustered the 39 sampled troops into seven clusters but lacked the strong statistical significance in support of it. Isolation by distance had a stronger influence on population genetic structuring of Assam macaque, whereas the riverine barrier effects were insignificant. Contrastingly, Hanuman Langurs defined 35 haplotypes with high nucleotide diversity (0.0298 ±SD 0.0009) and haplotype diversity (0.957 ±SD 0.015) from mtDNA D-loop. The troops were grouped into six major clusters corresponding to their spatial distribution isolated by the rivers, and significantly high genetic variations among those clusters conformed the barrier effects of snow-fed Himalayan rivers in genetic structuring of Hanuman Langur. The evolutionary history recovered from molecular analyses and paleodistribution reconstruction revealed differential patterns for the two species in Nepal Himalaya. Nepalese population of the Assam macaque has short evolutionary history that experienced demographic and spatial expansion from small founder population after last glacial maximum (LGM). Incongruously, Hanuman Langurs have longer evolutionary history, survived in Nepal Himalaya since early Pleistocene and had a small-scale population decrease during LGM that revived with the onset of Holocene. The population genetic structure and demographic history of Assam macaque were affected by Pleistocene climatic fluctuations than riverine barrier effects, whereas, the same were much affected by riverine barriers than the climatic variations in Hanuman Langurs. Assam macaques retreated mid-hills of central Nepal as refugial population during LGM and currently have elevational range expansion, longitudinally eastward and latitudinally northward towards higher elevations. Hanuman Langurs shifted to lower elevations during LGM without considerable loss of habitat and currently have unidirectional range expansion towards higher elevation. Phylogenetic inference of Sinica-group of macaques using multiple mtDNA and Y-chromosomal loci concluded that Nepal population of Assam macaque is paraphyletic to the eastern subspecies of Assam macaque (M. assamensis assamensis), whereas it formed monophyletic clade with M. munzala, M. leucogenys and conspecific populations from Bhutan. The most recent common ancestor of Nepal population of Assam macaque and rest of the taxa of its monophyletic clade occurred 1.62 mya; and the Nepal population of Sinica-group macaque was the first to diverge from that common ancestral stock. The tree topology and divergence time estimation provide strong support for the distinct species status of Nepal population of Assam macaque.The phylogenetic tree retrieved three clades among Hanuman Langurs within Nepal corresponding to the morphological division into three species/subspecies i.e., Semnopithecus entellus hector, S. e. ajax and S. e. schistaceus and corroborates with their geographical distribution. The nodal support values for the clades within ‘entellus’ species group was high in both maximum likelihood and Bayesian inference methods. Detail phylogenetic assessment of Nepal population of Assam macaque and Himalayan Langurs using multiple mitochondrial and nuclear DNA loci with morphological, distribution and behavioral data is warranted. For both the species, the areas in mid-hills of Nepal remained as the prominent suitable habitats which mostly fall outside the current protected areas, hence conservation efforts on mid-hills are recommended. Ecological niche modelling employing multiple algorithms should be used to assess the potential impacts of climate change on their future distribution.