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厌氧微生物在碳氮铁循环以及典型污染物迁移和转化中作用的机制研究
Alternative TitleTHE CONTRIBUTION OF ANAEROBIC MICROORGANISMS TO C/N/IRON CYCLING AND TYPICAL POLLUTANT TRANSFORMATION AND DISSEMINATION
周国伟
Subtype博士后
Thesis Advisor朱永官
2020-10-29
Degree Grantor中国科学院生态环境研究中心
Place of Conferral北京
Degree Name理学
Degree Discipline环境科学
Keyword厌氧微生物 、土壤动物肠道 、 硝酸盐 /铁 还原 、 砷还原和甲基化 、水平基 因转移 、 蛋白质降解 Anaerobic Microorganism, Gut Microbiota, Nitrate/iron(Iii) Reduction, Arsenic Reduction And Methylation, Plasmid Conjugation, Protein Degradation
Abstract

        厌氧微生物广泛的分布在多种生境中,包括土壤、土壤动物肠道以及沉积物。虽然厌氧微生物仅占环境微生物的很少一部分,但是他们对元素(碳、氮和铁)循环和环境污染物(如含砷化合物和抗生素抗性基因)迁移和转化有非常重要的作用。目前,关于厌氧微生物对这些过程贡献的研究仍然比较薄弱。本研究选择了几种富含厌氧微生物的环境样品(土壤、土壤动物肠道内容物和海洋沉积物);围绕反硝化、铁还原、砷酸钠的还原和甲基化以及抗生素抗性基因水平 转移等过程;结合生化分析、 16S rRNA高通量测序和 RNA-SIP等手段;旨在揭示厌氧细菌和古菌在 碳 /氮 /铁元素的循环、含砷化合物转化以及抗生素抗性基因的水平转移过程中的作用。主要研究结果如下:
      (1)与周围土壤相比,土壤蚯蚓肠道中铁还原、硝酸盐还原、乙酸盐氧化的速率都更高。无论是在土壤还是蚯蚓肠道,硝 酸盐还原均早于铁还原过程。对于铁还原过程而言,具有发酵功能的 Clostridium、 Bacillus以及 Desulfotomaculum 在蚯蚓肠道中占主导地位,而 Geobacter则是土壤中丰度最高的铁还原微生物。此外,蚯蚓肠道中的多个种属既具有铁还原功能也具备反硝化能力。
      (2)两种菌株 E. coli BL21和 BL21M均能成功地在秀丽隐杆线虫的肠道内定殖,并且能够介导线虫肠道中 As V)的还原和甲基化。由肠道微生物介导产生的 As V)和 DMAs会诱导线虫细胞内 VTG GST和 SOD水平的升高,这与线虫 的繁殖和氧化应激防御密切相关。毒性测试进一步表明,肠道中 E. coli BL21或 BL21M介导的砷转化显著的降低了暴露于 100 μM As V)的线虫的产卵率。
      (3) 秀丽隐杆线虫肠道中的微生物群落与周围土壤中的微生物群落的结构相差甚大,并且肠道中质粒的接合率比土壤中的高出很多。宿主的基因型和 年龄是影响线虫肠道微生物群落结构和质粒接合率的重要因素。肠道核心菌群主要是Enterobacteriaceae,包括 E. fergusonii、 Salmonella enterica subsp. enterica serovar Typhimurium、 Shigella flexneri和 Shigella sonnei,他们也是质粒发生接合的主要受体菌。
      (4)我们证实了三个门中的五个厌氧古细菌亚组具有蛋白质降解活性,这些亚组包括: SG8-5、 uncultured Thermoplasmata、 Methanomassiliicoccales  related、Bathy-15和 Loki-2b related。在全球范围内,这五个古细菌亚组广泛的分布在多种沉积物中,并显示出不同的蛋白质利用能力。其中 SG8-5可以在蛋白质 降解过程中同时同化有机碳和无机碳,而且能够灵活的清除蛋白质降解过程中产生的电子。
      总而言之,厌氧微生物通过有机酸氧化、铁还原、反硝化、蛋白质降解、抗生素抗性基因水平转移、砷酸钠的还原和甲基化等过程驱动着环境中 C、 N和 Fe等元素的循环。系列研究不仅加深了我们 对厌氧微生物生境多样性的认知,而且丰富了我们对厌氧微生物种类和功能多样性的理解。

Other Abstract

       Anaerobic microorganisms are widely distributed in diverse environments, including soil, soil animal intestines and sediments. Although anaerobic microorganisms account for only a small part of environmental microbes, they play a very important role in element cycling and pollutant (such as arsenic compounds and antibiotic resistance genes) transformation. Currently, research on contribution of anaerobic microorganisms in these processes is still under characterized. Combination of biochemical analysis, 16S rRNA high-throughput sequencing with RNA-SIP, this study focused on denitrification, iron reduction, sodium arsenate reduction and methylation, and horizontal transfer of antibiotic resistance genes in versatile environments. The main findings are as follows:
      (1) Rates of iron reduction, nitrate reduction and acetate oxidation in the gut of soil earthworms were higher than those in the surrounding soil. Nitrate reduction occurred earlier than iron reduction no matter in the soil or earthworm gut. Clostridium, Bacillus and Desulfotomaculum were dominant iron(III)reducers in the gut of earthworms, while Geobacter was the most abundant iron-reducing microorganism in the soil. In addition, iron(III)-reducing microbial population shared several genera with denitrifers in the gut, revealing a close link between iron(III) reduction and denitrification in earthworm gut.
      (2) Strains of E. coli BL21 and BL21M successfully colonized the gut of Caenorhabditis elegans, and both of them can stimulate the reduction and methylation of As(V) in the worm gut. As (V) and DMAs produced by gut microbiota via arsenic transformation can induce the increase of VTG, GST and SOD levels in the nematode cells, which were closely associated with reproduction and the defense against oxidative stress in C. elegans. The toxicity test further showed that arsenic biotransformation mediated by E. coli BL21 or BL21M in the gut significantly impaired the reproduction of C. elegans exposed to 100 μM As(V).
      (3) The microbial community composition in the gut of C. elegans was significantly different from that in the surrounding soil. The conjugation rate of plasmids in the gut was much higher than that in soil. Genotype and age were important factors that affect gut microbial community structure and plasmid conjugation rate of C. elegans. The intestinal core flora was Enterobacteriaceae, including E. fergusonii, Salmonella enterica subsp. enterica serovar Typhimurium, Shigella flexneri and Shigella sonnei, which were also the main recipient bacteria for plasmid conjugation.
      (4) Five archaea subgroups (SG8-5, uncultured Thermoplasmata, Methanomassiliicoccales related, Bathy-15 and Loki-2b related) affiliated to three phyla have protein degradation activity. They were globally distributed for all these five subgroups, highlighting the importance of protein degradation by varied uncultivated archaea in coastal marine sediments. Especially, SG8-5 can simultaneously assimilate both organic and inorganic carbon during protein degradation. All these five archaeal subgroups displayed divergent capabilities of protein utilization, which were determined by the features of electron scavenging strategy and availability of electron acceptors in sediments.
      All in all, anaerobic microorganisms served important roles in driving the cycling of C, N and Fe in the environment via fatty acid oxidation, protein degradation, iron reduction, antibiotic resistance gene horizontal transfer, sodium arsenate reduction and methylation. This research deepened our understanding on the diversity of anaerobic microorganism and their functions.
 

Pages128
Document Type学位论文
Identifierhttp://ir.rcees.ac.cn/handle/311016/43716
Collection土壤环境科学实验室
Recommended Citation
GB/T 7714
周国伟. 厌氧微生物在碳氮铁循环以及典型污染物迁移和转化中作用的机制研究[D]. 北京. 中国科学院生态环境研究中心,2020.
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