中国大陆王锦蛇的表型和遗传变异：检测局域适应

Testing adaptation of organisms to local environment is a central goal of evolutionary ecology. Widely distributed species inhabiting environmentally heterogeneous regions provide an excellent opportunity in this respect. The selective forces imposed by local environments and phenotypes under selection have been studied in a wide range of vertebrate taxa, but not including any snake species. In this study, we will use king ratsnakes (Elaphe carinata) collected from 23 populations covering the entire range of the species in mainland China to fill this gap. Specifically, we will address four main questions: (1) Do fitness-related (morphological, physiological, behavioral and life-history) phenotypes vary in response to local environment, thus varying among geographically distinct populations? (2) Is the spatial pattern of population genetic diversity related to local environment？(3) Does a given phenotype have an unique pattern of geographic variation, thus having an unique position in the continuum of cogradient-countergradient variation? (4) Is the differential expression of stress (temperature) resistance gene (HSPS) among populations heritable？We will conduct a series of field and laboratory experiments to collect data on female reproductive traits (maternal size, postpartum mass, egg-laying date, clutch size and egg size), egg traits other than size (embryonic stage at laying, incubation length and hatching success) and offspring traits (size, early growth and cognitive behavior) and then calculate Pst for each phenotype. We will use SNP marker to resolve phylogenetic relationships among populations and to calculate Fst. We will contrast Pst with Fst to examine whether a phenotype is selected and thus locally adapted. We will use phylogenetic ANOVA, phylogenetic ANCOVA or phylogenetic generalized least squares regression to determine associations of the observed patterns of phenotypic variation with the species' range in mainland China and local environment. We will perform an integrative analysis of environmental, genetic and phenotypic data to test four hypotheses: (1) phenotypes should be similar among populations with similar environmental conditions if these phenotypes are tightly correlated with local environment; (2) population genetic diversity should be higher in regions where habitat stability is higher if it varies geographically; (3) phenotypes should be more likely to be optimized in the peripheral regions of distribution if they vary along the continuum of cogradient-countergradient variation; and (4) the expression of HSPS should be adaptive in different populations climate regions if its differential expression among populations is heritable. Data generated from this study will allow us to better understand how snakes adapt to local environments in mainland China and, more importantly, they might provide insight into the underlying mechanisms for the formation of the spatial patterns of genetic and phenotypic variation of snakes in the region.

检测局域适应是进化生态学的核心任务之一。申请人拟研究中国大陆王锦蛇的遗传和表型变异，检测局域适应，通过假设检验的方法探讨4个具体科学问题：（1）适合度相关的形态、生理、行为和生活史表型是否与物种的地理分布和局域环境有关？（2）种群遗传多样性的空间格局是否与局域环境有关？（3）特定表型在不同种群是否呈现逆梯度顺梯度的连续性变异？（4）种群间温度抗逆基因在面临环境剧烈变化时表达量的变异是否具有遗传性？将在中国大陆23个种群采集王锦蛇，用SNP基因标记研究遗传多样性并计算Fst；其中6个种群用于测定抗逆基因的表达量，收集母体繁殖和幼体特征数据并计算Pst，比较Pst和Fst辨析适应性表型。将对环境、遗传、表型数据进行整合分析，阐明气候和植被等环境因子对地理分布、遗传多样性空间格局的影响，以及王锦蛇对局域环境的表型适应。本项目对揭示局域适应对大陆蛇类遗传多样性和表型空间格局的影响具有重要意义。

本文从遗传格局、生活史特征、生理适应和进化等方面阐述了王锦蛇的局域适应机制。分析王锦蛇线粒体和SNP数据证实王锦蛇个体主要可以聚为以太行山—巫山为分界线的东西两大支系。两个进化支的王锦蛇都在进化过程中都经历过种群扩张事件，东部群体的扩张过程显得更为渐进而西部群体的则显得更为突然。共筛选出24个局域适应相关的SNP位点。王锦蛇遗传格局受到距离和环境异质性的共同影响。以分布范围更大的中华蟾蜍为对象进一步验证东亚地区陆生变温脊椎动物的遗传格局，分析SNP数据显示，21个中华蟾蜍种群被划分成三个进化支，其遗传格局受到距离和环境异质性的共同影响。筛选出30个与局域适应相关的SNP位点。研究基因易损性发现全球变暖将会导致中华蟾蜍栖息地进一步片段化，并可能导致其东南种群灭绝。.不同种群王锦蛇的生活史特征具有显著差异，相同体长情况下舟山种群的母体产后体重最大，江山种群的最小，其余种群介于两者之间。产卵时间从六月底到八月初，种群间差异较显著。王锦蛇窝卵数和窝卵重均随母体体长的增大而增多，但不同种群繁殖输出无显著差异。比较王锦蛇和黑眉锦蛇怀卵母体和新生幼体的肠道微生物发现不同组肠道微生物的alpha多样性无显著差异。王锦蛇和黑眉锦蛇肠道微生物群组成相似。两种蛇的肠道菌群只有相对丰度存在差异，而初次摄食后幼蛇的肠道菌群组成变化迅速，且两种幼蛇肠道菌群之间存在差异。这说明饮食是影响了幼蛇的肠道微生物定植的主要因素。.从进化角度看，卵生和卵胎生蛇类胚胎中的卵黄分配发现卵生到卵胎生的进化并没有改变胚胎的卵黄分配机制。鳞片结构和形态的进化分析显示蛇腹鳞上的微皮纹反映了蛇类对其微生境的一种生态适应。有鳞类动物鳞片微皮纹特征和皮肤感受器的形态是对其所处环境多重压力的优化选择。比较陆生脊椎动物的生理和生态位进化发现：1）热生理耐受进化显著慢于冷耐受生理进化速率；2）最高温度生态位的变化慢于最低温度生态位的变化；3）对冷热的生理耐受性比相应的气候生态位变量进化得更慢。