Table_4_Insights into ecological roles of uncultivated bacteria in Katase hot spring sediment from long-read metagenomics.XLSX

Diverse yet-uncultivated bacteria and archaea, i.e., microbial dark matter, are present in terrestrial hot spring environments. Numerous metagenome-assembled genomes (MAGs) of these uncultivated prokaryotes by short-read metagenomics have been reported so far, suggesting their metabolic potential. However, more reliable MAGs, i.e., circularized complete MAGs (cMAGs), have been rarely reported from hot spring environments. Here, we report 61 high-quality (HQ)-MAGs, including 14 cMAGs, of diverse uncultivated bacteria and archaea retrieved from hot spring sediment (52°C, pH 7.2) by highly accurate long-read sequencing using PacBio Sequel II. The HQ MAGs were affiliated with one archaeal and 13 bacterial phyla. Notably, nine of the 14 cMAGs were the first reported cMAGs for the family- to class-level clades that these cMAGs belonged to. The genome information suggests that the bacteria represented by MAGs play a significant role in the biogeochemical cycling of carbon, nitrogen, iron, and sulfur at this site. In particular, the genome analysis of six HQ MAGs including two cMAGs of Armatimonadota, of which members are frequently abundant in hot spring environments, predicts that they are aerobic, moderate thermophilic chemoorganoheterotrophs, and potentially oxidize and/or reduce iron. This prediction is consistent with the environmental conditions where they were detected. Our results expand the knowledge regarding the ecological potential of uncultivated bacteria in moderately-high-temperature environments.

陆地热泉环境中存在多样性未培养细菌与古菌，即微生物暗物质（microbial dark matter）。迄今已有诸多研究通过短读长宏基因组学技术，获取了这类未培养原核生物的宏基因组组装基因组（metagenome-assembled genomes, MAGs），并揭示了其代谢潜力。然而，从热泉环境中获得的更为可靠的环状完整宏基因组组装基因组（circularized complete MAGs, cMAGs）却鲜有报道。
本研究通过PacBio Sequel II平台的高精度长读长测序技术，从温度52℃、pH 7.2的热泉沉积物中获得了61条高质量（high-quality, HQ）MAGs，其中包含14条cMAGs，涵盖多样的未培养细菌与古菌。这批高质量MAGs隶属于1个古菌门与13个细菌门。值得注意的是，14条cMAGs中有9条所属的科至纲级演化支，其对应的cMAGs此前尚无报道。
基因组信息表明，这些由MAGs代表的微生物在该位点的碳、氮、铁、硫生物地球化学循环中发挥重要作用。尤为特别的是，对6条高质量MAGs（包含2条装甲菌门（Armatimonadota）的cMAGs，该门类成员在热泉环境中通常较为丰富）的基因组分析预测：这类微生物为需氧型中度嗜热化能有机异养菌，具备铁氧化和/或铁还原的潜在能力。该预测与它们被检出的环境条件相符。本研究结果拓展了我们对中高温环境中未培养细菌生态潜力的认知。