山西阳泉老窑水硫酸盐还原菌的分离鉴定及其还原功能驯化

Isolation and identification of sulfate-reducing bacteria in goaf water in Yangquan of Shanxi Province and domestication for the reduction

摘要:

【背景】含硫煤矿开采后，地表水/地下水回流至采空区形成酸性老窑水，含有高浓度重金属离子和硫酸盐，严重危害生态系统健康。利用微生物自身生长处理老窑水具有成本低、环境友好等特点，具有良好的应用前景。目前利用的硫酸盐还原菌大多只在适宜温度和中性pH条件下具有较高活性，在北方低温和酸性条件下难以发挥作用。【目的】本研究旨在从山西阳泉山底河流域的老窑水环境中分离硫酸盐还原菌，并调节温度和pH进行驯化，从而得到高效耐低温耐酸菌株，为北方老窑水微生物治理提供可用菌种资源。【方法】对山底河流域典型老窑水样品中的微生物进行富集培养，并筛选硫酸盐还原菌。通过革兰氏染色、扫描电镜对菌株形貌特性进行表征，利用16S rRNA基因序列比对进行菌种鉴定，探究其生长特性和硫酸盐还原性能。在此基础上降低温度和pH，对高效还原硫酸盐菌株进行驯化，探讨其在北方老窑水污染治理中的应用潜力。【结果】本研究筛选得到2株硫酸盐还原菌，命名为YQ-1和YQ-2，分别属于革兰氏阴性瘤胃解蛋白质菌属(Proteiniclasticum)和脱硫弧菌属(Desulfovibrio)。在30℃、pH 7.5条件下，YQ-1和YQ-2对1 100 mg/L硫酸根的还原效率分别为96.75%和75.48%。选取高效菌株YQ-1进行定向驯化，该菌株经低温驯化后，在15℃、pH 7.5条件下对硫酸根的去除率为91.49%，比驯化前提高了85.69%；进一步进行低pH驯化后，在15℃、pH 4.5条件下对硫酸根的去除率为37.21%，比驯化前提高了34.30%。【结论】通过驯化培养提高了硫酸盐还原菌YQ-1对低温和低pH环境的耐受性，同时也提高了其在低温和低pH条件下对硫酸根的还原效率，为北方老窑水治理提供了菌种资源和理论依据。

关键词:老窑水;硫酸盐还原菌;单菌株筛选;细菌驯化

Abstract:

[Background] Surface water/ground water flows back to the goaf of sulfur-containing coal mine, forming the acidic goaf water. The water, with high salinity and sulfate content, threatens ecosystem health. Microorganisms, which feature low cost and environmental protection, has promising prospect of application in the treatment of goaf water. However, the currently available sulfate-reducing bacteria (SRB) are only highly active under suitable temperature and neutral pH, and are intolerant to low temperature and acidic conditions in northern China. [Objective] To isolate SRB from goaf water in Shandi River Basin of Yangquan, Shanxi, and to domesticate them so that they can tolerate the low temperature and acidic condition and be used for the treatment of goaf water in northern China. [Methods] Microorganisms in goaf water sample from Shandi River Basin were enriched and SRB were isolated. Then the SRB were characterized by Gram staining and scanning electron microscopy and identified based on 16S rRNA sequence alignment. Their growth characteristics and sulfate-reducing capacity were also investigated. On this basis, temperature and pH were lowered to domesticate the efficient SRB, thereby exploring their potential in the treatment of goaf water. [Results] Two strains of SRB, YQ-1 and YQ-2, were isolated, belonging to the Gram-negative Proteiniclasticum and Desulfovibrio, respectively. At 30 ℃ and pH 7.5, YQ-1 and YQ-2 reduced 96.75% and 75.48% of the 1 100 mg/L sulfate, respectively. The highly efficient strain YQ-1 was selected for domestication at low temperature and pH. The sulfate removal rate of the strain was 91.49% at 15 ℃ and pH 7.5 after low-temperature domestication, 85.69% higher than that before domestication. The removal rate was up to 37.21% at 15 ℃ and pH 4.5 after low-pH domestication, which was 34.30% higher than that before domestication. [Conclusion] The tolerance of YQ-1 to low temperature and low pH was improved, and the efficiency of sulfate reduction was also enhanced after domestication. This work provided strain resources and theoretical basis for the treatment of goaf water in northern China.

Key words:goaf water;sulfate-reducing bacteria;single-strain screening;bacterial domestication