| Biochar is increasingly promoted as a beneficial soil conditioner. Polycyclic aromatic hydrocarbons(PAHs), typical organic pollutants, are mainly originated from the incomplete combustion of organic materials in a variety of agricultural, domestic and industrial processes. Bioremediation is an emerging cleanup technology for PAHs-contaminated soil, which is a low-cost and environmental-friendly alternative for decontamination of the polluted soil. However, PAHs-contaminated soil is often suffered from nutrient and organic matter deficiency, which seriously limit the microbial degradation capacity in soil. In this study, biochars produced from rice straw and dairy manure at 350 and 500 ℃ by oxygen limited pyrolysis were used to investigate the influence of biochars on soil indigenous microorganisms. Microbial functional genes and their quantitative analysis of microbial diversity were also investigated by terminal restriction polymorphism analysis(T-RFLP) and fluorescence quantitative PCR(qPCR) technology. Then, taking persistent organic pollutants polycyclic aromatic hydrocarbons(PAHs) as the research object, microbial immobilization technique was used to fix the PAHs degrading bacteria on biochar. The remediation of 16 PAHs in soil combined pollution was carried out. And the potential mechanism for enhancing remedation of PAH contamination soil was explored. This study has important theoretical and practical significance for the development of contaminated soil in situ bioremediation technology and clarified the remediation mechanism and potential ecological effects of biochar immobilized degrading bacteria in PAHs contamination soil.The main conclusions are as follows:1. Functional groups carbonyl(C=O), carboxyl(-COOH), hydroxyl(-OH), ether(-O-) aromatic C=N bond and N-H3 bond existed on the surface of biochar-derived dairy manure, rice husk and rice straw. Except C element, biochar contains N, P, Ca, Na, Mg, K, and Mn, Zn, Cu and so on. All biochars examined exhibited alkaline. And pH value of biochar-derived rice straw RS500 was maximum(up to 10.50). The content of C and ash in biochar produced at 500 ℃ was more than those at 350℃. Under the same temperature pyrolysis conditions, the C/N in biochar-derived rice husk and rice straw were more than those in the biochar-derived dairy manure.2. Biochar increased significantly soil invertase activity and inhibited actinomycetes activity. The influence of biochars on activity of soil phosphatase, urease, number of bacteria and fungi, emissions of CO2 and N2 O depend on pyrolysis temperature and application rate. 4% of biochar amendment to soil stimulated significantly bacterial growth. Stimulation intensity on bacteria was greater by biochar at 500 ℃ pyrolysis temperature than those at 350 ℃. The effect of biochar on soil N2 O emission showed a typical hormesis characteristic.3. Biochar induced significantly soil bacterial community change. In biochar-amended soil, five groups were formed and nirS gene copy number increased by 67.22~70.18%. However, microbial community diversity, amoA gene copy number were closely related to the biochar pyrolysis temperature, feed stock and application rate. Among them, at 4% of application rate was, 350 ℃ low temperature pyrolysis biochar increased significantly soil microbial diversity. Except biochar DM350 and RH350, amoA gene copy number reduced by 93~98.10% compared with control. Gene copy number of bacterial 16 S rDNA and PAH-RHDα GP increased significantly in soil with biochar. The stimulation intensity was greater in soil with biochar produced at lower temperature treatments than those at the higher temperature. The maximum of PAH-RHDα GP gene copy number is biochar derived rice straw RS350(up to1.73 × 107 copies g-1 dry soil).4. In soil with PAHs combined pollution, biochar induced amoA gene copy number decrease. The lowest one was in soil with biochar-derived rice straw RS500. In contrast, the number of PAH-RHDα GP and nirS genes increased significantly. The inhibition rate of amoA gene copy number was from 23.53 to 85.13% in contaminated soil with biochar. The maximum proportional increase of PAH-RHDα GP gene copy number was induced by biochar-derived rice straw RS350. Its copy number reached 1.73 × 107 copies g-1 dry soil. Under the same treatment condition, the number of nirS gene copy in soil with biochar pyrolysed at 500℃ is more than those at 350 ℃. And the maximium increase was in soil with biochar-derived rice straw RS350, which was 2.9 times over that in the control soil. The copy number in control without biochar reached 6.45 × 107 copies g-1 dry soil.5. One PAHs degrading bacteria was obtained from soil contaminated with PAHs and heavy metal by enrichment culture methods. The results from 16 S rDNA molecular identification showed that this degrading bacterium belong to Cupriavidus and is named B35.6. Biochar produced at higher pyrolysis temperature(500℃) has greater absorption and degradation capacity to PAHs than those at lower pyrolysis temperature(350℃). At the same pyrolysis temperature, the adsorption capacity of B35 agents immobilized on biochar-derived rice husk and straw were greater than dairy manure. The order of adsorption capacity of B35 agents immobilized on biochar for PAHs were: rice husk RH350> rice straw RS350> dairy manure RM350> husk RH500> dairy manure DM500> rice straw RS500. The highest capacity degradation for PAHs was B35 agents immobilized biochar from rice husk RH350. Its degradation rate was 31.34%.7. There was significant difference in adsorption and degradation capacity of immobilized bacterial B35 agent for PAHs of various ring. Except for biochar-derived rice straw RS500, the rate of adsorption by biochar immobilized B35 agents for 4-ring PAHs was more than 2 and 3- ring PAHs. The average absorption rate was 44.98% and 24.50%, respectively. The effect of pyrolysis temperature on adsorption capapcity of biochar immobilized B35 agents for 5~6-ring PAHs were greater than those for 2 ~4-rings PAHs. The degradation rate of 2~4-ring PAHs on 350℃ pyrolysis biochar immobilized B35 agents was more than those on 500℃ pyrolysis biochar immobilization agents. The average degradation rate of 2~3-rings, 4-rings and 5~6-ring PAHs were 15.03%, 25.08% and 10.08%, respectively. Their maximum degradation rate reached 23.33%, 36.50% and 22.75%, respectively.8. The adsorption and degradation of PAHs on biochar immobilized B35 agents in medium were similar to those in contaminated soil. The rate of adsorption and degradation were higher on low temperature 350℃ biochar immobilized agents than those on biochar from high temperature 500℃. And the rate of adsorption for 2~4-ring PAHs was more than those for 5~6-ring PAHs. The biochar-derived rice straw RS350 had the maximium adsorption rate(22.80%).9. The mechanisms behind PAHs bioremediation by immobilized B35 agents with biochar as carrier maybe the synergy between bioaugmentation and biostimulation in PAH-contaminated soil. The order of bioremediation capacity of PAHs by immobilized B35 agents with biochar as carrier in PAH-contaminated soil were: rice straw RS350> husk RH350> dairy manure RM350> husk RH500> dairy manure DM500> rice straw RS500. |
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