Process Optimization And Mechanism Of Aerobic Cometabolic Biodegradation Of Trichloroethylene

Trichloroethene(TCE)which can lead to carcinogenesis,teratogenesis and mutagenesis effect is an important organic solvents and chemical raw materials.Leaks,improper disposal and direct emissions of TCE bring gross pollution in water,soil and air.As one of the important waste gas anthropogenic source,landfill also generated a lot of chlorinated hydrocarbon contaminants due to anaerobic degradation of origincs.Landfill is rich in a large number of function microorganisms in the long-term stress greenhouse gases of methane,carbon dioxide and volatile chlorinated hydrocarbons.So chlorinated hydrocarbon contaminants can be degraded by mixed bacteria with biological characteristics of high tolerance and mutual nutrition in the case of the presence of pollution sources through a variety of ways,such as co-metabolism and direct oxidation.It is considered to be an effective way to remove TCE and other chlorinated hydrocarbons pollutants.Here,in order to understand the population structure of function microorganisms in landfill,and to achieve the efficient methane emission reduction and TCE biodegradation.in this thesis,TCE co-metabolic biodegradation was studid by pure and mixed microorganism which were isolated from landfill cover soil under aerobic conditions.The main contents included the screening of function microbial,functional gene cluster sequence analysis and other chlorinated hydrocarbons biodegradation studies.The results are as follows:1)Capacity of methane oxidation in several typically landfill cover soils was studied,the results show the degradation rate of methane reached 99.8% when its initial volume concentration was 14% afterr 150 hours in cover soil of Chongqing which was higher than others.The optimum pH was 6.0-8.8 and maximum methane oxidation rate occurred at pH=7.02.In addation,adding NMS could enchance the capaciby of methane oxidation by the microorganism in landfill cover soils.2)A novel strain of methanotroph was isolated.The phylogenetic analysis indicated that the strain belongs to a species of Methylocystis,named as Methylocystis sp.JTC3.The degradation rate of TCE reached 93.79% when its initial concentration was 15.64 ?mol/L after 5 days,and TCE could promote the oxidation rate of methane when TCE initial concentration was from 12.55 ?mol/L to 20.76 ?mol/L.We obtained the pmoCAB gene cluster of 3 227 bp including pmoC gene of 771 bp,pmoA gene of 759 bp,pmoB gene of 1 260 bp and two noncoding sequences in the middle by semi-nested PCR,T-A cloning and sequencing.The theoretical molecular weight of their corresponding gamma,beta and alpha subunit were 29.1 kDa,28.6 kDa and 45.6 kDa separately analsised using ExPASy tool.3)Mixed microbial consortium named SWA1 and can use methane as a carbon source was isolated from landfill.SWA1 could be cultured continuously and stably.Non-methane-soluble carbon source could enriched related microbial community in mixed microbial consortium.TCE(14.06 ?mol/L)could promote the growth of SWA1.The growing ability and methane oxidation capacity of SWA1 could be improved by adding copper.4)The operating conditions of TCE biodegradation by SWA1 were optimized.The biodegradation of TCE was increased with TCE concentration(14.06-110.23 ?mol/L),and the maximum degradation efficiency was 87.79% in 110.23 ?mol/L TCE.In addition,the biodegradation rate of TCE can be strengthed by adding copper ion.There were two peak of TCE degradation rate,when copper ion concentration was 0.03 ?mol/L and 5 ?mol/L,the maximum degradation rate was 95.75% and 84.75%,respectively.5)In this paper,we analyzed the ability of transcript expression of genes encoding key enzymes by RT-qPCR and T-A cloning sequencing and the community structure in mixed microbial consortium by high throughput sequencing.The result of RT-qPCR showed that pMMO was thecritical enzyme for TCE degradation process.The maximum of pmoA gene and mmoX gene transcriptional expression abundance in 0.03 ?mol/L copper concentration and LmpH gene transcriptional expression abundance was increased with increasing copper concentration.The result of T-A cloning indicated that compared with control group without TCE,the abundance of methanotrophic bacteria was reduce and the abundance of non-methane oxidizing bacteria species increased in the presence TCE suggesting that the community sturcture of non-dominant and dominant microorganisms was change constantly.The result of High-throughput sequencing showed that Methylocystaceae which can co-metabolic degrade TCE were the dominant microorganisms,other microorganisms,such as Lactococcus and Bacillus,with the function of direct oxidation TCE were also included in SWA1 and the microbial diversity decreased significantly along with increasing of copper ion concentration.TCE degradation mechanism were different between low concentration of copper ion range(0-0.75 mol/L)and high concentration range(1-15 mol/L).TCE degradation mechanism were main pMMO,sMMO co-metabolic degradation of TCE and TCE direct degradation in low concentration range.Simultaneously in the high concentration range of copper ion,co-metabolism of non methane oxidizing bacteria having phenol hydroxylase also played a key role for TCE degradation.