Microbial Community Structure and Function in Groundwater of Abandoned Coal Mine and Its Response to Environment
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摘要: 为揭示微生物在煤矿开采区地下水环境中的空间分布特征及受控因素，为煤矿开采区地下水污染生态修复奠定基础，以河南新密某煤矿开采区地下水微生物为研究对象，利用16S rRNA高通量测序技术，分析了饮用水井和废弃矿井地下水微生物群落的结构与功能及其对环境的响应.研究表明在属水平上饮用水井主要以不动杆菌属（Acinetobacter）、草地土杆状菌属（Chthonobacter）和黄杆菌属（Flavobacterium）为主，而废弃矿井中则以短波单胞菌属（Brevundimonas）和甲基红色杆菌属（Methylorubrum）为主.两组微生物在多种合成酶、脱氢酶以及转运系统ATP结合蛋白等方面的潜在功能上存在显著差异.分子生态网络显示，矿井水微生物不仅联系更为紧密，且种间多为正相关关系，推测物种可能通过合作来应对极端寡营养环境.该区域地下水微生物群落结构主要受溶解性总固体和硝酸盐氮含量的影响，营养条件是制约当地地下水微生物群落结构与功能的先决因素.Abstract: To reveal the spatial distribution characteristics and controlled factors of microorganisms in the groundwater of the coal mining area, and to lay the foundation for the ecological restoration of groundwater pollution in the coal mining area, the structure and function of microbial communities in drinking water wells and abandoned mine wells, and their responses to the local environment were analyzed by using 16S rRNA high-throughput sequencing technology in a coal mining area in Xinmi, Henan Province, China. The results show that at the genus level, drinking water wells were mainly composed of Acinetobacter, Chthonobacter and Flavobacterium, while Brevundimonas and Methylorubrum were dominant in abandoned mine wells. The significant functional differences between the two groups were in multiple synthases, dehydrogenases and ATP-binding proteins of the transporter system. Molecular ecological network analysis shows that microorganisms in mine water were not only more closely linked but also mostly positively correlated, and species might respond to the extreme oligotrophic environment by cooperation. The groundwater microbial communities in the region were mainly influenced by total dissolved solids and nitrate nitrogen content, indicating that nutrient conditions were the prerequisite factors governing the structure and function of local microbial communities.
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图 1 研究区地理位置与采样点分布
Fig. 1. Geographical location of study area and distribution of sampling points
图 2 XMCHJ组和KJ组微生物多样性分析
a. α多样性指数箱线图；b. 两组样本ASV水平韦恩图；c. 样本分组PCoA分析
Fig. 2. Microbial diversity analysis of XMCHJ group and KJ group
图 3 XMCHJ组和KJ组在门水平和属水平上的物种组成
Fig. 3. Microbial community composition at the phylum and genus level of XMCHJ group and KJ group
图 4 LEFSe指示物种分析
柱状图颜色代表各自的组别，长短代表LDA score，即不同组间显著差异物种的影响程度
Fig. 4. LEFSe indicator species analysis
图 5 XMCHJ组和KJ组微生物群落的潜在功能比较
a. 功能分类的PLS-DA分析图；b. 差异功能基因分类柱形图，左侧为不同功能分类在两个样本中的丰度比例，中间图为95%置信区间内，功能丰度的差异比例，右侧为FDR值，FDR值小于0.05，表示差异显著；c. 差异基因聚类热图
Fig. 5. Comparison of potential functions of XMCHJ group and KJ group
图 6 XMCHJ组和KJ组的生态网络分析
Fig. 6. Interaction network analysis of XMCHJ group and KJ group
图 7 微生物群落与环境因子的冗余分析（RDA）
Fig. 7. Redundancy analysis (RDA) of microbial communities and environmental factors
表 1 不同取样点地下水样本的理化参数
Table 1. Physicochemical parameters of groundwater from different sampling points
ID 总硬度（mg/L） 浑浊度（NTU） 溶解性总固体（mg/L） 高锰酸盐指数（mg/L） 硝酸盐氮（mg/L） 硫酸盐（mg/L） 偏硅酸（mg/L） 碳酸氢根（mg/L） 砷（10-4 mg/L） 镁（mg/L） 钙（mg/L） 钠（mg/L） 钾（mg/L） 锌（mg/L） XMCHJ01 349 0 468 0.71 3.40 64.0 16.5 408 11 25.9 79 36.8 0.22 0.083 XMCHJ04 415 0.001 490 0.56 8.23 116.0 16.0 297 4 25.6 99 32.5 0.86 0 XMCHJ14 489 9.8 463 0.68 5.09 41.4 14.7 424 14 48.2 81.4 13.3 0.2 0 XMCHJ85 350 0.6 448 0.64 3.47 59.1 16.2 326 0 27.6 73.8 49.7 1.02 0.221 XMCHJ86 810 1.4 1 080 0.71 54.10 193.0 24.4 439 9 33.2 292 35 0.08 0 KJ027 511 348 721 0.46 14.10 77.6 27.9 481 6 27.1 189 20.3 0 0 KJ089 462 29 529 0.60 9.34 257.0 26.0 229 51 14.0 129 26.6 9.43 1 KJ100 612 2.8 869 0.51 12.10 232.0 21.5 484 5 35.6 226 34.3 0.73 0.106 KJ103 223 1.0 492 0.55 1.12 37.6 21.8 428 8 17.4 44.7 124 4.44 0.160 
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表 2 组间分子生态网络拓扑结构参数
Table 2. Parameters of the molecular ecological network topology between the two groups
分组 节点数 边数 平均度 平均加权度 模块化系数 平均聚类系数 正相关系数 负相关系数 XMCHJ 290 4 620 31.86 13.49 1.16 0.63 67.68% 32.32% KJ 170 3 494 41.11 50.75 0.52 0.77 100% 0
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