萼花臂尾轮虫应对河流氨氮污染的适应性机制研究

The increasing anthropogenic activities have led to release of massive pollutants, causing dramatic biodiversity losses and severe function impairment of river ecosystem. To illuminate the processes and trends, and the related mechanisms of biota development in polluted rivers is the principal key question for restoring and managing impaired river ecosystems. Specifically, clarifying adaptive mechanisms underlying responses of organisms to pollutants is an important foundation. In the present proposal, in perspectives of gene expression and its relationship with phenotypes, we will apply the methods of the population transcriptomics to investigate Brachionus calyciflorus in the Chaobai River, which is a typical polluted river with distinct ammonia nitrogen gradient. Firstly, with complete analysis of gene expressing and genotypes, we will study the gene expression differences of Brachionus calyciflorus in different ammonia nitrogen gradient and the relationship between gene expression and phenotypes. Then, the critical signaling pathways and genes involved in responses of Brachionus calyciflorus to ammonia-nitrogen will be identified based on differentially expressed genes (DEGs). Subsequently, the key adaptive phenotypes will be determined based on the gene expression-phenotype relationships and the identified critical genes. And finally, this proposal will elucidate the adaptive mechanism of Brachionus calyciflorus to ammonia nitrogen stress in perspectives of both gene expression and genotypes. This study will provide an important basis for revealing the adaptive mechanisms of invertebrate to water pollution and provide a theoretical basis for effectively predicting the biodiversity and biogeographical distribution in polluted river ecosystem.

日益频繁的人类活动带来大量污染物排放，导致河流水生生物多样性锐减，河流生态系统功能严重受损。解答污染环境下生物区系的演化过程、方向及其生态机制是恢复和管理受损河流生态系统亟需解决的首要科学问题。其中，明确解析生物体响应污染物胁迫的适应性机制是回答上述科学问题的基础。因此，本项目拟以具显著氨氮污染梯度的潮白河为研究体系，以萼花臂尾轮虫（Brachionus calyciflorus）为研究对象，以群体转录组学为主要技术手段，以基因表达和表型的关系为切入点，通过对不同氨氮梯度下基因表达差异的分析及基因表达水平变化与表型特征差异关系的建立，重点揭示参与氨氮胁迫响应的关键信号通路和基因及响应氨氮胁迫的关键适应性表型，最终阐明萼花臂尾轮虫应对氨氮污染胁迫的适应性机制。本研究将为揭示无脊椎动物响应水体污染的适应性机制提供重要研究基础，同时也为有效预测水体污染对生物多样性及生物地理分布提供理论基础。

河流生态系统已经成为受威胁程度最高的生态系统之一，其生物多样性锐减。水体污染对河流生物多样性减少的影响得到越来越多的关注。污染河流的生物效应机制解析是污染环境下生物区系演化过程、方向及其生态机制亟需解决的首要科学问题，这一问题的回答可以为恢复和管理受损河流生态系统提供科学指导。由此，为了分析河流污染的形态响应机制，本项目以具有显著环境梯度的河流为研究体系，以萼花臂尾轮虫为模式生物，通过野外样品水质因子与形态参数的分析，发现与体型大小相关的形态参数包括头部宽度（G），体宽（H），体长（I）和身体体积（V）为水体敏感形态学指标，与水体氨氮浓度增加密切相关，是关键适应性表型；进一步通过氨氮处理实验表明急性（24小时）刺激条件下随着氨氮浓度增加，萼花臂尾轮虫群体的敏感参数长度呈现显著增加，但是长期（7天）氨氮刺激实验条件下与体积相关的敏感参数在胁迫处理后逐渐恢复到原有水平。为了分析萼花臂尾轮虫对氨氮胁迫的基因表达响应机制，本研究在对不同氨氮浓度梯度处理的萼花臂尾轮虫进行转录组测序的基础上，通过转录组序列组装注释、基因表达水平定量分析、差异表达基因筛选共得到1640个差异基因，比较不同浓度处理的差异表达基因发现不同浓度氨氮处理条件下，其表达模式不同；通过对差异基因的功能分析和基因共表达网络分析发现glnS基因（glutamine-tRNA ligase）、Hsp20基因、Rplp2基因（60S acidic ribosomal protein P2-like isoform X2）、cyp524A1基因（cytochrome P450 family protein）、vwkA基因（protein serine/threonine kinase）为关键响应基因。通过对差异基因与氨氮浓度值之间的趋势分析，本项目研究发现了与氨氮浓度正相关的基因集profile 19（包括56个基因），负相关的基因集profile 0（包括2个基因），这些基因集与形态参数差异潜在相关。通过对基因集profile 19进行基因功能富集分析表明基因功能主要与离子转运、跨膜运输相关。