正构烷烃降解菌的筛选及降解过程中的变化特征

发布时间：2018-02-05 03:20

  本文关键词： 降解菌筛选 正构烷烃 微生物修复 细菌组成　出处：《沈阳大学》2016年硕士论文　论文类型：学位论文

【摘要】：本研究利用辽河油田石油污染土壤筛选分离正十六烷降解菌,并对所分离菌株进行理化性质分析及分类鉴定,同时监测其对正十六烷的代谢活性。利用模拟土壤的泥浆体系的正十六烷降解实验验证功能菌株对土壤污染环境的适应性。通过辽河油田油井周边石油污染土壤的微生物及电动修复实验,观测污染土壤修复过程中目标降解菌及微生物整体群落的生长及代谢活性变化特征,由此明确目标降解菌在有机污染土壤修复中的应用价值。研究的主要结论包括:(1)以辽河油田石油污染土壤为菌源,以正十六烷为唯一碳源,筛选分离出3株具有较强的正十六烷代谢功能的降解菌,分别记为T1、T2和T5,依据菌落的生长形态及16s r DNA测序分析确定T1、T5为铜绿假单胞菌(Pseudomonas aeruginosa),T2为鞘氨醇菌(Spingopyxis terrae)。摇瓶培养的正交实验确定混合菌株在温度30℃,p H为7,C:N:P为100:10:2,盐度为1.5%的溶液体系中获得对正十六烷的最佳降解效果。(2)土壤模拟(泥浆)体系中的正十六烷降解实验表明,前述筛选所得正十六烷功能降解菌具有较好的正十六烷降解特性,微生物处理30天,正十六烷降解率达到29.3%。试验初期,T2(Spingopyxis terrae)细菌数量丰度较高,中期替换为T1与T5(Pseudomonas aeruginosa)的数量增多,各菌株数量的变化结合正十六烷降解率的变化特征,反映了多菌株组合应用所产生的共代谢优势。DGGE群落组成分析表明,各降解菌株在直流弱电场条件下均具有一定程度的电场耐受性,但适应速率存在差异。弱场强梯度(1.2 V·cm-1)刺激功能降解菌的代谢活性,电动-微生物处理30天,正十六烷降解率达到65.10%,较微生物处理降解率提高35.80%,表现出电动与微生物的协同降解作用。(3)辽河油田石油污染场地土壤修复实验表明,功能降解菌对辽河油田石油组分中饱和烷烃的降解作用效果明显,处理48天后降解率达到37.58%。直流弱电场的施用进一步提高正十六烷的降解能力,处理48天后降解率达到61.97%,较单独微生物处理降解率提高24.39%。土壤脱氢酶(DHA)活性分析表明,微生物组(BIO)及电动-微生物(EK-BIO)组中DHA活性远高于电动(EK)组,反映了功能降解菌具有较好的污染物代谢活性。相关性分析表明,施加功能降解菌的土壤脱氢酶活性与烷烃的降解速率呈现显著的正相关性。EK-BIO处理中脱氢酶的活性高于BIO组,验证了功能降解菌受弱电场刺激,其烷烃代谢能力得到提高。DGGE图谱分析表明,电动处理有效增强了土壤微生物群落多样性,同时提高功能降解菌对污染土壤环境的适应性。本研究分离得到的烷烃降解菌不仅适用于石油污染土壤的微生物修复处理,而且适用于电场条件下的微生物-电动联合修复技术的应用。本研究为石油污染土壤微生物修复过程的调控及新的联合方法的建立提供理论基础。
[Abstract]:In this study, n-hexadecane degrading bacteria were isolated from oil-contaminated soil in Liaohe Oilfield, and their physicochemical properties were analyzed and classified. At the same time, the metabolic activity of cetane was monitored. The adaptability of the functional strain to soil pollution was verified by the n-cetane degradation experiment of simulated mud system. Oil contaminated soil around the oil well of Liaohe Oilfield was used to verify the adaptability of the functional strain to soil pollution. Microbiological and electric repair experiments, The characteristics of growth and metabolic activity of the target degrading bacteria and microbial community during the remediation of contaminated soil were observed. The main conclusions of the study include: the petroleum contaminated soil in Liaohe Oilfield was used as the bacterial source and the hexadecane as the sole carbon source. Three strains of biodegradable bacteria with strong function of cetane metabolism were screened out. It was recorded as T _ 1 T _ 2 and T _ 5, respectively. According to the growth morphology of colony and 16s r DNA sequencing analysis, it was confirmed that T _ 1 T _ 5 was Pseudomonas aeruginosafida T _ 2 as sphingosine sphingosporium. The orthogonal experiment of shaking flask culture confirmed that the mixed strain was 7C / N: P = 100: 10: 2 at 30 鈩，

本文编号：1491969