Functional analysis of the archaea, bacteria, and viruses from a halite endolithic microbial community

Halite endoliths (salt rocks) in the Atacama Desert represent one of the most extreme environments on Earth. We obtained a 9.6 Gb high quality shotgun metagenome from halite nodules collected in the Salar Grande area. Our metagenome is unique because it has high sequencing depth, low diversity, and representative members from all 3 domains of life and from viruses. Taxonomic assignment of the metagenomic reads confirmed that the community was dominated by members of the Archaea, including Nanohaloarchaea, and also comprised members of the Bacteria and Eukarya. The functional annotation of the metagenome revealed that the cyanobacterium Halothece was responsible for most of the CO2 fixed in the community and that a number of archaea and bacteria carried out photoheterotrophy, via light-driven proton pumps, increasing the energy budget of the community from light. Our analysis predicted that the alga identified in the community has one of the lowest protein isoelectric point (pI) distributions of any reported eukaryote. Its pI profile was similar to that of halophiles using a “salt-in” strategy. Our genome assembly produced a nanohaloarchaeon genome of 1.2 Mbp. We have named this new microorganism Candidatus Nanopetramus SG9. Analysis of its genome sequence revealed a photoheterotrophic life style, a low median isoelectric point for all its predicted proteins, suggesting a “salt-in” strategist, and the presence of a unique CRISPR/Cas system in its genome; one of the spacers from this unique CRISPR/Cas system matched a partial viral genome from the metagenome. We developed new methods to identify viral genomic content from metagenome data, resulting in the identification of over 30 complete or near complete viral or proviral genomes in the halite metagenome. These viral genomes were diverse in genome structure, gene content, host, and genome size. Putative hosts for these viruses included Halobacteriaceae, Nanohaloarchaea, and Cyanobacteria. Despite the dependence on deliquescence of the halite community for liquid water availability, this study revealed an ecosystem spanning three phylogenetic domains, containing a large diversity of viruses, and a predominant “salt-in” strategy to balance the high osmotic pressure of the environment.

阿塔卡马沙漠中的盐岩内生菌（Halite endoliths，即盐岩）是地球最为极端的环境之一。我们从大盐湖（Salar Grande）区域采集的盐结核中获取了一份9.6 Gb的高质量鸟枪宏基因组（shotgun metagenome）。本宏基因组具有独特性：其测序深度高、物种多样性低，且涵盖了生命所有三个域——古菌域（Archaea）、细菌域（Bacteria）、真核生物域（Eukarya）以及病毒的代表性类群。对宏基因组读段（metagenomic reads）的分类学注释（taxonomic assignment）证实，该群落以古菌成员为主，包括纳米嗜盐古菌（Nanohaloarchaea），同时也包含细菌与真核生物类群。宏基因组的功能注释（functional annotation）显示，蓝细菌（cyanobacterium）*Halothece*负责群落中绝大部分二氧化碳（CO₂）的固定；此外，多种古菌与细菌通过光驱动质子泵（light-driven proton pumps）进行光异养（photoheterotrophy），提升了群落的光能收支（energy budget）。我们的分析预测，群落中发现的藻类拥有所有已报道真核生物中最低的蛋白质等电点（protein isoelectric point, pI）分布之一，其等电点特征与采用"盐入"策略（salt-in strategy）的嗜盐微生物（halophiles）相似。我们的基因组组装（genome assembly）获得了一条1.2 Mbp的纳米嗜盐古菌基因组（nanohaloarchaeon genome），将这一新发现的微生物命名为候选纳米杯菌*Candidatus Nanopetramus SG9*。对其基因组序列的分析显示，该微生物具有光异养生活方式，其所有预测蛋白质的平均等电点较低，表明其属于"盐入"策略适应者，且基因组中携带一套独特的CRISPR/Cas系统（CRISPR/Cas system）；该系统的其中一段间隔序列（spacers）与宏基因组中的一段部分病毒基因组匹配。我们开发了全新的方法用于从宏基因组数据（metagenome data）中识别病毒基因组组分，最终在该盐岩宏基因组中鉴定出30余个完整或近完整的病毒或前病毒基因组（proviral genomes）。这些病毒基因组在基因组结构、基因含量、宿主范围与基因组大小上均具有多样性，其推定宿主包括盐杆菌科（Halobacteriaceae）、纳米嗜盐古菌与蓝细菌。尽管该盐岩群落依赖潮解（deliquescence）获取液态水（liquid water），本研究仍揭示了一个覆盖三个系统发育域（phylogenetic domains）的生态系统，其中包含大量多样的病毒类群，并以"盐入"策略作为主导机制以平衡环境的高渗透压（osmotic pressure）。