Isolation And Screening Of Plant Endophytic Bacteria And Plant Bacteria Combined-remediation Of PAHs In Soil

Along with the global industrialization and the modernized development,coal and petroleum were widely applied within industrial production,transportation as well as living-life,so that pollution in soils by PAHs was day by day serious and had becomed a problem urgently solved.Microorganism plant combined-remediation of PAHs were green and cheap and moreover it has merits which microorganism or plant sole remediation did not have.In this experiment,that endophytic bacterium plant combined-remediation strengthening merits of microorganism and plant,has significantly theoretic and practical meanings.Thirty-six plant endophytic bacteria were isolated from water-earthnut and wild garlic in oil-contaminated soils in Nanjing.According to the potential of removing PAHs, promoting wheats growth and colonizing in wheat tissue interior,strain 7J2 was screened out for degrading phenanthrene and strain 12J1 was screened out for degrading pyrene through the plant growth promoting and colonization experiments.Based on the morphological and biochemical characteristics and 16SrDNA sequence analysis,strain 7J2 and strain 12J1 were both identified as Enterobacter spp.Characteristics of the strain 7J2,12J1 were investigated and the results showed that the strain 7J2,12J1 can dissolve calcium phosphate,and produce IAA,siderophore.The degradation rate of phenanthrene(30 mg·L^-1) by the strain 7J2 was 99.81%at 28℃on the rotary shaker at 150 rpm for 6 days.The degradation rate of pyrene(5 mg·L^-1) by the strain 12J1 was 83.75%at 28℃on the rotary shaker at 150 rpm for 7 days.The experiment of the strains influenced by environmental factors showed that the strain 7J2 and 12.11 can grow within 4-45℃,within pH 4.0-11.0 and the optimum was 20-37℃,pH7.2-9.0.The strains adapted easily to environment and were able to grow under salinity of 5%.The strains had different resistances to the heavy metal.Under Pb²⁺,Cr²⁺,Cu²⁺,Ni²⁺,Zn²⁺,Cd²⁺, lethal concentrations of the strain 7J2 were respectively 3000,10,5,10,1500,50 mg.L^-1, and those of the strain 12J1 respectively were 3000,10,10,20,2500,50 mg.L^-1.The strains had same resistances to antibiotic.The strain 7J2 and 12J1 could grow well in culture medium containing Km 10 mg.L^-1,Amp 100 mg.L^-1,Str 10 mg.L^-1,Tc 10 mg.L^-1,Rif 20 mg.L^-1,or Spe 10 mg.L^-1.The experiment of endophytic bacterium plant combined-remediation of PAHs in soils showed that,in soils containing phenanthrene 200 mg.L^-1,pyrene 100 mg.L^-1,or phenanthrene 200 mg.L^-1 and pyrene 100 mg.L^-1,the biomass of inoculated plant was obviously more than that of non-inoculated plant.The treatments of microorganism plant combined-remediation were higher than the sole remediation of plant or microorganism, and also higher than CK obviously.In treatments of plants,the concentrations of residual PAHs within inoculated plants were obviously more than those of non-inoculated plants. Moreover,in plants by solely polluted,the concentrations of residual PAHs were obviously more than in plants by compound pollution.In treatments of plants,sucrose enzyme, polyphenol oxidase,and urea enzyme were obviously higher than in treatments of no plants. In inoculated treatments,sucrose enzyme,polyphenol oxidase,and urea enzyme were obviously higher than in non-inoculated treatments.By selective culture mediums to separate and count,it showed the strain 7J2 and 12J1 could colonize well in the plant rhizosphere.Along with the experiment continuing,the quantities of bacterium,epiphyte in the soil increased gradually.Endophytic bacteria strain 7J2 and 12J1 survived in the wheat for long.After 60 days they still could be separated from wheat and their best survival time was within 25 days.They survived in the maize for long.After 30 days they still could be separated from maize and their best survival time was within 20 days.