Study On The Isolation, Identification And Degradation Characteristics Of O-Dichlorobenzene-degrading Bacteria

In recent years, Microbial degradation of environmental pollutants has become the most promising treatment of environmental pollution.Microbial treatment was a method of low cost, high efficiency, no secondary pollution and easy operation. Therefore, screened the efficient degradation bacteria, studyed the degradation characteristics and its related enzymes, and applied into the environment had an important theoretical and practical significance.The mehtod was to gradually raise the condensation degree of o-dichlorobenzene for bacterium step by step. The result showed that the o-dichlorobenzene degrading abilities were greatly promoted from 40 mg/L to 200 mg/L. Three highly effective o-dichlorobenzene-degrading strains named DXL-1, DXL-2, DXL-3 through enrichiment culture were isolated from the stimulant constructed wetland reeds rhizosphere soil. Selected the highest degrading rate strain DXL-1 for further study. The strain was studied on the strain morphological characteristics and physiological and biochemical identification, the measurement of growth curve, resistant and optimal degradation conditions. At the same time, a preliminary study on the characterization of strain DXL-1 crude enzyme.The results showed that, a strain that had a capability to degrade o-dichlorobenzene was isolated from the reeds wetland. The strain could use o-dichlorobenzene as the sole carbon source in minimal medium. It was primarily identified as Alishewanella aestuarii based on morphological, physio-biochemical characteristics and 16S rDNA analysis of sequence. The strain was a facultative anaerobe, gram positive, straight rods, with peritrichous flaglla, catalase positive. Resistance of this strain to antibiotic is detected. The result showed this strain has resistance to streptomycin, but others.Through the determination of various factors including different carbon sources, nitrogen source, temperature, pH value, liquid volume in flask of degrading bacteria growth, Biological characteristics reaserch showed that, the strain DXL-1 could use glucose yeast extract as its optimal carbon and use yeast extract as its nitrogen source. Organic nitrogen source was better than inorganic nitrogen source. The strain DXL-1 could growth well with 25℃～35℃, pH 7.0-7.5. The factors influencing the strain's o-dichlorobenzene degrading ability, such as pH, temperature, initial o-dichlorobenzene concentration, inoculation volume were studied. The optimal conditions for the degrading o-dichlorobenzene were chosen by orthogonal experiment. The results showed that the optimum conditions were as follows:the initial o-dichlorobenzene concentration was 120 mg/L, temperature was 32℃, pH was 7.0, the inoculation volume was 10%, reacted 4 days, under which the biodegrading rate reached to 85.52%.Preliminary study on the strain DXL-1 crude enzyme showed that, the degradation of o-dichlorobenzene was catalyzed by catechol 1,2-dioxygenase.The optimal conditions for the enzymatic reaction were pH 8.0 and 30℃. The enzyme activities were relatively stable under pH 7.0-8.5. The effects of heavy metal ions on enzyme activities were not neglectable. Irons, such as Fe3+, Ba2+ and Mn2+ had a remarkalbe promotion effects on enzyme activities and Cu2+, Zn2+, Fe2+ were just on the contray.Comparison of degradation of a single strain and mixtuer strain, the mixtuer bacterial strains has a more advantaged performance on degrading o-dichlorobenzene than what pure bacterial strains do in this experiment. Co-degradation existed between the strains.