海洋生态与环境科学重点实验室知识库

深海因其高压、黑暗以及光合作用来源的有机物匮乏等特征，长期被视为“生命荒漠”，随着海洋研究的发展，人们发现在深海中存在另一种特殊的生态系统，即深海化能合成生态系统。化能自养菌利用氧化环境中CH₄、H₂S等还原性的物质释放的化学能合成有机物，供养整个生态系统。深海Bathymodiolus属贻贝广泛存在于很多深海化能合成生态系统中，其中，Bathymodiolus platifrons贻贝是西太平洋很多热液和冷泉环境共有的优势种，基于其生态系统中的重要地位以及该系统中多种生物基因组大小的调查结果，选定该物种作为研究模型，采用多组学和生物信息学的研究方法，围绕B. platifrons（深海贝）及其亲缘关系较近的偏顶蛤Modiolus kurilensis（近海贝）开展了比较基因组和比较转录组的测序和分析工作，揭示了B. platifrons在基因组水平以及基因表达水平上对环境的适应性特征。其中，与化能自养菌的共生关系是其适应深海化能还原环境的一个关键因素，本研究也重点探讨了深海贝与其共生菌之间的相互作用关系。此外，深海生物的起源问题一直饱受争议，本研究基于线粒体基因组的测序结果，重建了已有数据的所有贻贝的系统发育树，以探讨深海贝的起源，同时比较了B. platifrons热液群体和冷泉群体，以探讨不同群体之间的连通性。获得的主要结论如下：

1. 基因组大小和特征

基于流式细胞术测定了所研究区域多种优势生物的基因组大小，选定深海B. platifrons作为研究模型开展基因组测序，流式细胞术测定基因组大小约为2.4 Gb,基因组survey评估结果约为2.2 Gb。

深海贝中转座子含量增加，发生了种特异的近期扩张，尤其是helitron元件比例较高。已有研究表明，helitron元件与基因组重组有关，这些过程会导致基因组发生变化，可能与深海贝适应深海环境相关。此外，贻贝的基因内含子数量和长度都大于其他软体生物，同时可变剪切数量也对应增加，这些可能是贻贝特有的基因组特征。

2. 适应性特征

在应对环境胁迫方面，几乎所有代表性的抗胁迫基因都在深海贝和近海贝中存在，表明同近海贝一样，深海贝也具有适应各种环境刺激的潜力。而相比于近海贝，深海贝最显著的特征是含有内共生菌。在基因组水平上，与内吞作用、糖基化以及通道蛋白相关的基因家族在深海贝中扩张或者受到正选择，可能是长期进化过程中，共生关系的存在对宿主的选择作用导致的。

尽管深海贝主要依赖于其共生菌提供营养物质来源，但是深海贝在长期进化过程中，依旧保留了其消化潜力，这种多元的营养关系，有助于深海贝适应多样的深海环境。

3. 深海贝起源

基于线粒体基因组开展的系统发育分析结果表明，所有的Bathymodiolus贻贝聚在一起，以M. kurilensis贻贝作为姐妹枝。深海Bathymodiolus贻贝较近期分化时间（37.4 Mya）以及其位于系统进化树远离根节点的末端节点的位置，都支持深海贝并不是古老物种，而是不晚于白垩纪中期的时候从近海向深海演化而来。因此，这些深海贝不属于“活化石”。

4. 深海贝-共生菌相互作用关系

宿主中免疫识别相关基因在整体上发生了收缩，有利于共生的建立和维持；宿主通过内吞作用主动获取共生菌，溶酶体活动及其调控过程在宿主的营养获取、共生菌稳态维持中发挥重要作用，相关基因受到了选择并高表达。对于深海贝，自吞噬、凋亡以及溶酶体活动等过程可能与共生菌的调控有关，然而具体的调控机制仍不明确，需要更系统地结合组学和共生菌侵染实验等进一步阐明。

5. 热液和冷泉不同群体的比较

热液区和冷泉区的深海贝基因组相似度很高，且在群体水平上，基因组大小和线粒体基因组分析的结果都显示，两个群体没有发生分化，相距甚远（> 980 km） 的南海冷泉区和冲绳海槽热液区的B. platifrons贻贝不同群体间具有较好的连通性。

The deep-sea has been considered as a biological desert for marine animals due to its harsh conditions including complete darkness, low temperature and oligotrophic conditions. However, with more and more investigation of deap-sea, it is found that there are numerous flourished ecosystems scattered all around the deap-sea and called as deap-sea chemosythetic ecosystems, which are powered by chemosynthetic bacteria. Over past decades, the topic about how these species adapt to such “hash” habitats has drawn continuous attentions, but remains as an open question. Genome sequencing is a powerful method to study none-model species, and the knowledge of genome size is an important prerequisite. Mussels of subfamily Bathymodiolinae have been found to be one of the dominant species in diverse chemosynthetic ecosystems, among them, Bathymodiolus platifrons has been found to be common in the West-pacfic Ocean. Based on the genome size investigation of a wide range of invertebrates in sampling area, and considering the ecological importance, we choose B. platifrons as model species to study the adaptation to deep-sea. In order to identify the linaege-specific adaptive characteristics of B. platifrons, we conducted genome sequencing and RNA-seq of the mussel in a comparative frame with its costal relative Modiolus kurilensis. Compared with coastal mussels, one of the most extraordinary traits of Bathymodiolus mussels is their endosymbiosis. Based on the multi-omic analysis, we also disscussed the interaction relationship of this symbiotic system. Besides, there are debates about the origin of deep-sea life all these years. We also explored the phylogenetic interactions of mussels based on mitochondrial genomes sequencing and analysis, trying to understand the evolutionary transmission of the taxa. Meanwhile, we compared 20 complete mitogenomes of B. platifrons from a vent and a seep to explore the population connectivity. The main results are as follows:

1. Genome size and features

The genome size of B. platifrons was about 2.4 Gb and 2.2 Gb estimated by flow cytometry and genome survey respectively. Compared with other genomes of Lophotrochozoa species, the transposon elements were expanded in B. platifrons, especially the helitron elements, which could induce changes in genome, so it might play a role in the genomic rearrangement during the evolution.

In addition, the numbers and length of introns in three mussels were higher than those in other mollusks, with the increasing of numbers of alternative splicing, these might be the special features shared in mussels.

2. Adaptions

Both the deep-sea Bathymodiolus mussels and costal mussels possess diverse gene families related to stress responses, there were no specific or significantly expanded families in Bathymodiolus mussels, indicating that all mussels might possess the potential to cope with diverse stresses, but prefering different gene members of the same gene family in different species.

Compared with coastal relatives, the symbiotic relationship in deep-sea mussels are one of the most obvious and important difference. From the genomic insight, many gene families related to endocytosis, transporters, glycosylation were expanded or positively selected in deep-sea mussels, which indicated that the symbiotic relationship might have be a selective driver for the evolution of B. platifrons.

In deep-sea chemosynthetic ecosystems, many symbiotic invertebrates rely on the chemoautotrophic bacteria for nutrients, with the reduction of their digestive systems. We compared the number of digestion-related genes and expression pattern of different tissues between B. platifrons with other mollusks, the results indicated that B. platifrons mussels still have the potential of digestion, which might be good for the mussels to live in more diverse habitats and adapt to the unstable conditions.

3. The origin of deep-sea mussels

Combining evidence of the shallow divergence (37.4 Mya), the tip positioning in the phylogenetic trees as well as published fossil records, a recent invasion no older than the mid-cretaceous of Bathymodiolus spp. from shallow water to the deep-sea chemosynthetic ecosystem was supported. Hence, these deep-sea mussels are not living fossils.

4. Interactions between B. platifrons and its symbionts

Based on the comparative transcriptomic analysis, we identified several candidate pattern recognition receptors (PRRs) such as TLRs, C1qDC proteins, which might play a role in host-symbiont recognition. However, the results of statistics of representative PRR numbers using either genomic data or RNA-seq data all indicated that PRRs seemed to be globally reduced in B. platifrons, which might facilitate the infection and maintenance of their symbionts. Phagocytosis or endocytosis might be the routes for their entrance, after that, the symbionts were restricted in host-dedrived membrane and dwelled in host cell. Our results highlighted the lysosomal activities in deep-sea mussels, indicating that nutrients might be absorbed from the symbionts by cellular digestion. Meanwhile, diverse transporters were also identified to be positively selected or more highly expressed in the deep-sea mussels, which could provide more effective exchange of nutrients and intermediate metabolites.

To maintain a sustainale symbiotic relationship between Bathymodiolus mussels and their symbionts, related physiological processes such as lysosome activity, autophagy and apoptosis are need to be regulated coordinately, but the mechanisms remains unclear.

5. Population connectivity

According to the genome sequencing and analysis, the two genomes of B. platifrons from the studied hydrothermal vent and cold seep were highly similar. Besides, even with our hypervariable molecular marker, no structured genetic differentiation can be detected for B. platifrons from the seep and vent. The two populations were well connected, possibly by a strong Kuroshio Current during prolonged planktonic larvae stages.