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题名: 真菌-细菌联合修复芘和砷污染土壤研究
作者: 刘爽
学位类别: 博士
答辩日期: 2010
授予单位: 中国科学院研究生院
授予地点: 北京
导师: 朱永官
关键词: 细菌 ; bacteria ; 真菌 ; Gene-overexpression ; fugunas ; ; Arsenic ; ; pyrene ; arsM基因 ; arsM ; 基因超量表达
其他题名: Bioremedation of Pyrene-Arsenic Contaminated soils with Fungus and Bacteria
中文摘要:       复合污染研究已成为环境科学发展的重要方向之一,芘和砷是土壤环境中的重要污染物,开展两者复合污染研究,对于农业环境生态安全具有重要意义。本研究以耐砷(As)的芘降解细菌为研究目标,从长期遭受多环芳烃(PAHs)-As复合污染的土壤中,筛选得到三株能在As胁迫下降解芘的细菌。对这三株菌株进行了形态学和生理生化反应的研究。提取了16S rDNA,通过与同源菌株的比对,分别鉴定为两株细菌Bacillus idriensis 和Sphingomonas desiccabilis,一株真菌Fusarium solani.本论文以这三种菌株为主要研究对象,研究其对芘和砷污染的去除效能。研究结果如下: (1)通过研究两株细菌B.idriensis and S. desiccabilis 以及一株真菌F. solani利用芘作为唯一碳源和能源生长情况,可知这几株菌均具有一定的耐芘性能,在芘污染环境下均可生长,并且对芘具有降解能力。 (2)在芘(100 mg L-1)污染液体培养基中,几种处理均对芘有降解能力,实验测定结果显示,混合菌两株细菌和真菌共同降解芘(B+S+F)组合降解率最高,液体培养基中的芘残留仅剩3.97%。B+S+F混合菌在第4天降解能力就进入平台期,降解速率较快于其他几种菌处理。其中降解速率大小为B+S+F组合>B+S组合>S>F>B,说明混合菌降解芘效果要好于单独菌处理。 (3)将单独菌与混合菌其投入到人工处理的污染土壤中(芘 100mg l-1),观察土壤芘的降解残留值,单独菌株的不同处理中,细菌的降解时间要显著快于真菌,实验测定结果显示第63天混合菌(B+S)组合和(B+S+F)组合的降解能力都比较高,土壤的芘残留分别为仅剩16.56%和15.78%。说明应用微生物去除土壤芘的污染,混合菌降解芘效果要好于单独菌处理。应用混合菌(B+S+F)原位修复芘污染将更高效。 (4)利用红假单胞杆菌(R. palustris)中的arsM基因转导至细菌B. idriensis 和 S. desiccabilis中,重组体后的细菌,均能检测到arsM基因片段,并且具有一定的耐砷性,在48小时内重组体的细菌和野生型的细菌均能在As (Ⅲ) 20 μM处理条件下均能生长。 (5)利用细菌重组体的甲基化功能,将其应用到砷污染的培养基中,伴随着培养基中的砷总量减少,有气态砷产生。超量表达菌处理气态化砷的能力要高于野生菌处理10倍,超量表达arsM基因的细菌B. idriensis气态砷产生量最高,并且对溶液中砷污染的移除率可以达到41.87%;利用重组体的甲基化功能,将其应用到砷污染的土壤中培养30天,各处理土壤均可以检测到MMA,DMA有机砷的形态,并且有气态砷产生,大约有2.2 – 4.5% 的砷能够被微生物移除,有机砷占总砷的最高比率为13.87%; (6)在芘浓度为100 mg kg-1,As (Ⅲ) 20 μM的复合污染液体培养基中,超量表达的细菌与真菌处理均能生长并且产生气态砷。对培养基中的砷移除率可达到约84%的高效移除率,芘的残留率仅为8.05%。并且超量表达的单一菌和混合菌对芘均具有降解能力,几种不同微生物组合降解芘的处理,降解能力不同,分别为B+S+F处理>B+S 处理> S处理> B处理。 这些结果显示,采用经济、高效的微生物修复来修复芘和砷污染的水和土壤是行之有效的。
英文摘要:       Combined pollution has become one of the important directions of environmental science at present. In soil environment, pyrene and arsenic are often present together. Three strains were isolated and studied in this thesis.Morphological appearance. Gram-straining, other physiological and biochemical characteristics and 16S rDNA were taken to identify isolates. The three strains were named Bacillus idriensis and Sphingomonas desiccabilis,Fusarium solani (Mart.), the efficient single strain and consortium (B+S+F) on the remedation of pyrene and arsenic contanimated soil were evaluted in the laboratory, the major findings were obtained as follows: Three pure strains were isolated and purified with pyrene as the single carbon and energey source. They have different degradation capability. The ability of removal efficiencies of pyrene were B+S+F>B+S>S>F>B, after 9 days in the mineral salts medium under initial pyrene concentration of 100 mg L-1.The coculture (fungal- bacterial) was a kind of high efficient degradating strain towards pyrene, residue efficiencies was 3.97%. Three pure strains will have different pyrene degradation rates (100 mg L-1) to soil after a 63-day cultivation. Inoculation of fungal-bacterial cocultures in PAH-contaminated soil resulted in significantly improved degradation of pyrene, and the coculture (fungal- bacterial) can also reach a highest degradation rate. It is showed that pyrene of residue efficiencies was under 15.78% in soil. In Rhodopseudomonas palustris, an arsM gene, encoding bacterial and archaeal homologues of the mammalian Cyt19 AsIII S-adenosylmethionine methytransferase, is regulated by arsenicals. An expression of arsM was introduced into strains for the methylation of arsenicin this study. The expression of arsM in isolated Bacillus idriensis and Sphingomonas desiccabilis strains enhanced resistance to As. In the aqueous system, S. desiccabilis and B. idriensis with the expression of arsM methylated about 10-fold higherlevels of gasous As, respectively, than the wild type cells without the expression of arsM. The amount of volatilized As correlated with the disappearance of total As in solution. The decrease in AsIII was correlated with the appearance of two soluble products in the medium, DMAV, AsV and As in the head space of the reaction vial. In the soil system, about 2.2 - 4.5% of As was removed by biovolatilization during the 30 day inculation. This study demonstrated that arsenic could be removed through volatilization from the contaminated soil by the expressionof arsM in indigenous bacteria. Fungal-bacterial were cocultured, and three different single strain were culture after 9 days in the mineral salts medium under initial pyrene100 mg kg-1 and As (Ⅲ) 20 μM. The results showed that the co-culture had synergistic effects on the removal of pyrene and As. In the co-culture treatement group, 84% of As was removed and only 8.05% of pyrene remained. These results show that it is possible to use microorganisms as an inexpensive, efficient strategy for pyrene and arsenic bioremediation from contaminated water and soil.
内容类型: 学位论文
URI标识: http://ir.rcees.ac.cn/handle/311016/35063
Appears in Collections:中澳联合土壤环境研究室_学位论文

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Recommended Citation:
刘爽. 真菌-细菌联合修复芘和砷污染土壤研究[D]. 北京. 中国科学院研究生院. 2010.
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