褐煤强化产甲烷菌群的群落分析及条件优化

Community structure analysis of methanogenic flora and optimization for bioaugmentation methane generation from lignite

【目的】以白音华褐煤为底物,利用从我国多地煤矿及污水处理厌氧罐中富集-混合-驯化得到的高效混合菌群进行产气,分析其群落组成并优化产气条件。【方法】采用Miseq高通量测序分析混合菌群结构,通过Plackett-Burman(P-B)和Box-Behnken(B-B)试验对褐煤产气影响因素和条件进行筛选和优化。【结果】本源和外源微生物样本混合样品(HN+MD+WT)经驯化后菌群产气效率最高。该样品菌群中细菌群落多样性丰富,以变形菌门的脱硫弧菌属Desulfovibrio(15.07%)、拟杆菌门的屠场杆状菌属Macellibacteroides(14.6%)、厚壁菌门的梭菌属Clostridiaceae(9.77%)、互营菌门的脱硫代硫酸盐弧菌属Dethiosulfovibrio(8.76%)以及热袍菌门Oceanotoga属(8.66%)为主。古菌全部为广古菌门,其群落多样性则较为单一,其中甲烷卵圆形菌属Methanocalculus(80.28%)占据绝对优势。Plackett-Burman(P-B)试验结果表明温度、CoCl_2添加量和NiCl_2添加量是影响褐煤产气的关键因素;Box-Behnken(B-B)试验结果表明最优产气条件为:温度36°C,CoCl_2添加量0.17 g/L,NiCl_2添加量0.02 g/L,最优条件下褐煤累计产甲烷量(周期20 d)达到159.33μmol/g。【结论】经过驯化可以得到高效的产气菌群,优化培养条件可使产气效率明显提高。

[Objective] The mixed methanogenic flora with high methane production from Baiyinhua lignite was obtained from several coalmines and an anaerobic polluted water processing tank by enrichment, mixture and domestication. The community structure was analyzed and its methane production condition from lignite was optimized. [Methods] The community structure was analyzed by Miseq high-throughput sequencing. Plackett-Burman and Box-Behnken tests were conducted to screen and optimize the main factors influencing the efficiency of methane production. [Results] The mixed flora （HN＋MD＋WT） combined with indigenous and exogenous microbial culture produced the highest methane after domestication. The diversity of bacterial community was abundant, in which the dominant portion was Desulfovibrio （belong to proteobacteria, 15.07%）, Macellibacteroides （belong to bacteroidetes, 14.6%）, Clostridiaceae （belong to Firmicutes, 9.77%）, Dethiosulfovibrio （belong to Synergistetes, 8.76%） and Oceanotoga （belong to thermotogae, 8.66%）. All the archaea were assigned to Euryarchaeota, and its diversity was single comparatively. Methanocalculus （80.28%） was the most dominant genus. Plackett-Burman test results indicated temperature, concentrations of COCl2 and NiCl2 were the most important influencing factors. Box-Behnken test results showed that the optimal conditions were as follows： temperature 36 ℃, COCl2 concentration 0.17 g/L, NiCl2 concentration 0.02 g/L, under which the methane accumulation is up to 159.3 μmol/g （20 d）. ]Conclusion] A mixed flora with high methane production could be obtained by domestication. The productivity of methane could be enhanced significantly by optimizing the culture condition.