Hydrochemistry and microbial community characteristics and environmental response in different functional zones of a typical coal mine in Ordos
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摘要:
为探究煤矿井下不同功能区的微生物群落分布及其对水文地球化学特征的响应，以鄂尔多斯某矿为研究对象，在涉及矿井水的来源、形成、汇集和排放全过程的6个功能区中采集了24份水样，进行水化学组分检测和微生物16S rRNA基因高通量测序，并采用多元统计方法进行序列数据处理。研究结果表明：矿井水的水化学类型继承了其直接充水水源，均为高矿化度的SO4Na型，而特征污染物SO4 2−在煤巷和地表水池中浓度最高。不同功能区的微生物群落组成差异明显：属水平上检出的优势菌属包括能氧化硫化物的发硫菌属和硫氧化菌属，以及可降解有机物的新鞘氨醇杆菌属和短波单胞菌属等在煤巷、采空区中分布较高；好氧的Uliginosibacterium和具有极强吸附力且能降解有机物的不动杆菌属在岩巷中丰度最高；与氮循环有关的噬氢菌属和红细菌属在水仓和地表水中分布较高。微生物群落与水化学过程响应灵敏且关系密切；井工煤矿微生物群落分布不仅与C、N、Ca、Mg等营养元素相关，还与Fe、COD和SO4 2−等氧化还原敏感物质密切相关。煤巷和采空区是地下水污染防控的关键区域；开采扰动后，煤中伴生的低价硫化物在化学氧化和硫氧化菌的催化氧化作用下生成大量SO4 2−；然而，当工作面停采半年到3年后，在物理或化学吸附、沉淀作用（前期占主导），以及硫酸盐还原菌的还原作用下（后期占主导），SO4 2−降低了15%～34%，这说明采空区具有一定的自净能力。综上所述，研究成果可为矿井水污染防控的工程应用提供以下理论支撑：一方面可通过通氮气保持工作面的厌氧条件，从源头上减少SO4 2−的生成；另一方面可以筛选、培养硫酸盐还原菌和有机物降解菌，制作成生物材料投加到井下进行原位地下水污染修复。
Abstract:To explore the distribution of microbial communities in different functional zones of coal mine and their response to hydrogeochemical characteristics, a coal mine in Ordos was taken as the research object. 24 water samples were collected from six typical functional zones involved in the whole process of the mine water source, formation, collection and discharge. Hydrochemical components detection and high-throughput sequencing of microbial 16S rRNA genes were carried out. and multivariate statistical methods were used for sequence data processing. The results showed that the hydrochemical type of mine water was high salinity SO4-Na type, which directly inherited the supplied water-source, while the concentration of characteristic pollutant${\rm{SO}}_4^{2 - } $was highest in coal roadways and surface water pools. The compositions of microbial communities in different functional zones presented significant differences. The dominant bacterial genera detected at the genus level includedThiothrixand sulfur oxidizing bacteria that could oxidize sulfides (i.e.,ThiothrixandSulfuricurvum), as well as newSphingobacteriaandShortwave Monocmonasthat can degrade organic matter (i.e.,NovosphingobiumandBrevundimonas), while they distributed relatively high in coal tunnels and goafs. The abundances of aerobicUliginosibacteriumandAcinetobacterwith strong adsorption and organic degradation, was highest in rock roadways. Bacteria related to nitrogen cycle (i.e.,HydrogenophagaandRhodobacter) accounted for the higher proportion in water sumps and surface water. Microbial communities were sensitive and closely related to the hydro-chemical processes. The distribution of microbial communities in underground coal mine was not only related to nutrients such as C, N, Ca and Mg, but also closely interrelated to redox sensitive substances such as Fe, COD and${\rm{SO}}_4^{2 - } $. Coal roadways and goafs are the key zones for groundwater pollution prevention and control. After mining disturbance, the low valent sulfides associated with coal have generated a large amount of${\rm{SO}}_4^{2 - } $through the chemical oxidation and catalytic oxidation of sulfur oxidizing bacteria. However, it is worth noting that when the working pannel stopped for six months to three years, the characteristic pollutant${\rm{SO}}_4^{2 - } $was reduced by 15%-34% due to the physical or chemical adsorption, precipitation (dominated in the early stage), and the reduction of sulfate reducing bacteria (dominated in the later stage). This result indicated that the goaf had a certain degree of self-cleaning ability. In summary, the research results could provide theoretical supports for the engineering applications of mine water pollution prevention and control, which was reflected in the following two aspects: on the one hand, to reduce the generation of${\rm{SO}}_4^{2 - } $from the source by maintaining the anaerobic condition on the working pannel through nitrogen gas supply; on the other hand, after screening and cultivating the sulfate-reducing bacteria and organic matter degrading bacteria, they would be produced into bio materials, and added to the underground for in-situ remediation of groundwater pollution.
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图 1 研究区主要含水层与煤层赋存关系示意
Figure 1. Schematic of the relative positions of the main aquifers and coal seams in the study coal mine
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图 3 不同功能区水样的Piper和Durov图
Figure 3. Piper and durov diagrams of the water samples in different zones
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图 4 不同功能区水化学特征组分半箱线图
（GW—地下水；RR—岩巷水；CR—煤巷水；goaf—采空区；sump—水仓水；SW—地表水；方法检出限分别为pH：0；DO：0；ORP：−2000 mV；K++Na+：0.04 mg/L；Ca2+：0.01 mg/L；Mg2+：0.05 mg/L；Fe3+：0.03 mg/L；Fe2+：0.03 mg/L；NH4 +：0.01 mg/L；Sr：0.01 mg/L；Cl−：10 mg/L；SO4 2−：10 mg/L；NO3 −：0.08 mg/L；NO2 −：0.01 mg/L；TDS：20 mg/L；CO2：0.1 mg/L；H2SiO3：0.1 mg/L；COD：0.5 mg/L）
Figure 4. Half-box and whisker diagrams of the major hydrochemical components in the six zones
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图 5 某矿6个分区微生物的总物种、独特物种和共有物种数量在属水平上的分布（Venn图）
Figure 5. The numbers of total species, unique species and shared species of the microorganisms in the six zones of the research coal mine on genus level (Venn diagram)
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图 6 细菌门水平Circos图及Kruskal-Wallis H组间差异检验结果
Figure 6. Circos diagram and Kruskal-Wallis H test of distribution of microbial phyla in different zones
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图 7 细菌属水平上相对丰度柱形图及Kruskal-Wallis H组间差异检验结果
Figure 7. Relative abundance of the dominant lineages on genus levels and Kruskal-Wallis H test of distribution of microbial genera in different zones
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图 8 环境因子和微生物属分类群落基于Bray-Curtis距离的RDA分析
Figure 8. Bray-Curtis distance-based RDA of the relations between the selected environmental variables and microbial communities in water samples
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图 9 丰度前100的属物种与环境因子之间的相关关系网络图
（每个节点的大小与其连接度成正比;两个节点间连接线的粗细代表Spearman相关系数绝对值的大小。连接显示|r|>0.5和显著(p<0.05)的Spearman相关性）
Figure 9. Network of correlation between bacterial genera (top 100 most abundant species) and environmental variables
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图 10 地下水含水层水化学组成及微生物群落的垂直分布特征
Figure 10. Vertical distribution characteristics of microbial community and hydrochemical composition in groundwater aquifers
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图 11 采煤工作面从开采到关闭全过程中特征水化学指标变化特征
Figure 11. Characteristics of hydrochemical composition changes in coal pannel from mining to closure
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图 12 采煤工作面从开采到关闭全过程中微生物群落变化特征
Figure 12. Characteristics of microbial community changes in coal pannel from mining to closure
表 1 6种功能分区24个取样点详细信息
Table 1 Details of 24 sampling sites in 6 zones
功能分区 样品编号 采样点位置 地下水GW M4 直罗组中部含水层 M7 直罗组底部含水层混约10%延安组水 M23 白垩系志丹群含水层 煤巷水CR M2 3102工作面回采中 M3 3102工作面进风措施巷道积水 M18 3-1煤南翼大巷巷道积水 M21 3105工作面回采中 M22 2-1煤集中巷L2联巷 水仓水sump M9 2-2中煤水仓（2煤巷道和采空区水） M11 2号水仓（3煤采空区水） M19 中央水仓（收集部分M9与M11水仓水
以及3煤巷道水）岩巷水RR M10 副井管子道冻结管淋水 M17 3-1煤南翼大巷巷道淋水，墙壁上挂白色结晶物 M20 井底车场岩巷淋水 采空水goaf M1 3102工作面2020年采空水 M5 2201工作面回撤通道2020年采空水 M8 3101工作面2018年采空水 M12 3103工作面回撤通道2019年采空水 M16 3104工作面回撤通道2021年3月采空水 地表水SW M24 2号水池矿井水排水口② M25 2号水池设备清洗废水排水口① M26 2号水池排水口正南水池边界 M27 1号水池南边界 M28 3号水池北边界 注：不同功能区简介：地下水为原生地下水含水层；岩巷是指开挖断面岩石面积大于80%的巷道，主要用于通风、运输设备和物料等；煤巷是指开挖断面煤的面积大于80%的巷道包括掘进工作面，主要用于采煤和运输煤炭；采空区是当采矿停止后，将建一堵墙来封闭形成的采矿“空洞”，被封闭的空间；水仓用于暂存井下所收集的矿井水；地表暂存水池是中央水仓的矿井水经抽排至井上水处理厂预处理后，排入3个地表水池暂存待深度处理。 
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表 2 不同功能区中地下水和矿井水微生物群落Alpha多样性指数
Table 2 Alpha diversity indexes of microbial community in water of different zones
位置分区 样品编号 序列 OTUs 覆盖率/% 群落丰富度 群落多样性指数 ACE Chao 1 Shannon Simpson 地下水GW M4 28542 153 99.73 269.46 232.80 2.28 0.20 M7 27072 127 99.82 166.63 166.06 2.28 0.19 M23 28374 161 99.78 248.04 231.50 2.58 0.16 岩巷水RR M10 28214 136 99.74 205.21 228.81 1.44 0.42 M17 21334 803 98.89 1035.25 1067.10 4.57 0.04 M20 28306 250 99.51 458.73 363.96 1.25 0.58 煤巷水CR M2 22183 434 99.42 567.18 567.98 4.10 0.04 M3 24298 599 99.13 785.39 772.79 3.22 0.19 M18 27376 303 99.49 534.29 489.39 3.14 0.10 M21 27087 572 99.81 588.03 589.73 4.75 0.04 M22 27741 177 99.74 240.46 225.13 2.02 0.27 采空水goaf M1 26277 320 99.56 407.21 402.27 2.60 0.22 M5 25235 508 99.03 926.01 792.28 2.53 0.29 M8 23872 502 99.17 847.03 717.79 3.10 0.15 M12 24495 554 99.97 1031.50 930.41 3.55 0.08 M16 25636 449 99.24 655.71 637.61 3.35 0.08 水仓水sump M9 23622 638 98.88 1242.34 995.00 4.24 0.04 M11 27031 666 98.70 1032.62 1013.51 3.39 0.09 M19 25999 337 99.40 652.45 537.03 3.43 0.07 地表水SW M24 27197 389 99.29 769.42 587.68 3.56 0.06 M25 27769 433 99.66 476.17 468.04 3.47 0.09 M26 27231 562 99.19 739.28 766.25 4.42 0.03 M27 25973 217 99.73 277.84 272.03 2.30 0.29 M28 27314 379 99.42 529.15 496.32 3.43 0.08
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