| With the increasing demand for oil,oil exploitation process is constantly due to accidents such as leakage,which causes carcinogenic,teratogenic,mutagenic and other toxic petroleum hydrocarbons to the living organisms into the underground water and soil environment.As the essential zone of petroleum hydrocarbon migration to aquifer,the degradation of petroleum hydrocarbon by soil microorganisms using oxygen and ion receptors is the key to natural attenuation and purification of petroleum hydrocarbon before entering aquifer.However,most of the oil fields are widely distributed in the seasonal frozen soil areas in the north of China.The special seasonal freeze-thaw process will change the activity of soil microorganisms and related environmental factors such as temperature and REDOX conditions,making the degradation process of microorganisms in the frozen soil areas more complicated.At present,there is a lack of research at home and abroad on the degradation process and influencing factors of soil microorganisms in vadonic zone under freeze-thaw action,especially the response mechanism of microbial degradation process in permafrost area to freeze-thaw action remains to be explored.Based on this,this paper relies on the National Natural Science Foundation project,taking the oil contaminated site in Songyuan City,Northeast China as a typical research area,and comprehensively analyzes the response rules and interaction relationships of environmental indicators,pollutant content,nutrients,electron receptors,and microbial community structure under seasonal freezing and thawing and indoor daily freezing and thawing through site experiments and daily scale freezing and thawing cycle experiments,The characteristics of microbial degradation of petroleum hydrocarbons during different freezing and thawing processes were analyzed,and the evolution of microbial community abundance and structure under seasonal freezing and thawing processes and daily scale freezing and thawing cycles was revealed.The effects of different conditions such as freezing and thawing temperature,frequency,and water content on microbial degradation mechanisms were emphatically studied.Finally,the degradation mechanism of petroleum hydrocarbons by microorganisms in the aerated zone of seasonal frozen soil during freezing and thawing periods was depicted,The enhanced mechanism of microbial degradation during freezing and thawing period was obtained through a comparative study of adding artificial reinforcement bacteria.Through the research of the thesis,the following main conclusions and understandings have been obtained:The microorganisms in the original soil of the site are mainly composed of Anaerobic bacteria such as Arthrobacter of Actinobacteria and GP4,GP6,and GP16 of Acidobacter,which have significant denitrification and relatively slow sulfate reduction.However,the effect of anaerobic pathway on petroleum hydrocarbon attenuation is not significant in the short term.The aerobic bacteria in the original soil have no significant abundance at the beginning,and after adding petroleum hydrocarbons at room temperature,they undergo significant aerobic degradation with the rapidly growing Proteobacteria of Lysobacteria and Acinetobacter as the main aerobic bacteria,with a degradation rate of about 0.01 mg/g/day.Moreover,the species of bacterial flora are continuously derived under normal temperature conditions,with a significant increase in diversity.Seasonal freezing and thawing conditions have transformed the dominant aerobic bacteria species in the original soil into Rhodococcus and Nocardia of the actinomycetes that adapt to low-temperature environments,while the types of anaerobic bacteria are not affected.Due to the high abundance of anaerobic bacteria,they have significant denitrification and sulfate reduction effects when not frozen in the early stage of freezing and thawing;the main freezing period inhibits the availability of ion receptors,mainly due to the aerobic degradation of aerobic bacteria that grew up in the early stage of freezing and thawing at temperatures ranging from-10-0 ℃,while long-term low temperatures below-10 ℃ can cause aerobic and anaerobic bacteria to stagnate their growth and lose their degradation ability;At the melting stage,aerobic degradation,denitrification,and sulfate reduction capabilities will gradually recover and improve.During the entire freezing and thawing period,the degradation rate of petroleum hydrocarbons is about 0.004mg/g/day.Seasonal freezing and thawing conditions inhibit the oxygen consumption,denitrification,and sulfate reduction of microorganisms,as well as the increase in microbial diversity in the original soil.Different freezing and thawing temperature conditions did not affect the type of native soil flora,mainly through affecting the growth and activity of microorganisms,the availability of electron receptors and nutrients during freezing and thawing,affecting the degradation activity of microorganisms.The low-temperature freezing process can significantly inhibit the growth of microorganisms and the utilization of oxygen and ion receptors,while the melting process can gradually restore their growth activity.Freezing at too low a temperature of-15 ℃ will cause the microorganisms in the aeration zone to lose all their degradation pathways.Under the daily freezing and thawing cycles of-15 ℃ freezing and 5 ℃ thawing,it is not possible to restore the growth activity and degradation ability of microorganisms in a short period of time,but when the thawing temperature increases to 15 ℃,the degradation pathways of microbial oxygen consumption,denitrification,and sulfate reduction will recover quickly.The artificially added degradation bacteria belong to the Acinetobacter genus of the Proteobacteria.After being added at room temperature,the aerobic degradation effect in the original soil was significantly improved,and the degradation amount increased by about 6 times during the field experiment period.At the same time,it severely inhibited other bacteria and anaerobic degradation pathways in the original soil,but the abundance of the artificially degraded bacteria gradually decreased with the rapid consumption of oxygen at room temperature.However,the freeze-thaw effect significantly inhibited the aerobic enhancement and sulfate reduction of artificial degradation bacteria,and the degradation ability during the freeze-thaw period only increased by two times.The strengthening mechanism of its original soil microbial population during the freeze-thaw period is reflected in advancing and improving its aerobic degradation ability during the early freeze-thaw period and the winter freeze-thaw period.Water content conditions have a small impact on the evolution of microbial flora under freezing and thawing,mainly due to water migration caused by freezing and thawing affecting the content of electron receptors and nutrients available to microorganisms,thereby affecting the degradation effect.The frequency of freeze-thaw cycles will promote the recovery of microbial degradation pathways.During the freeze-thaw cycle,the sulfate reduction ability of microorganisms first recovers,followed by denitrification ability,and aerobic degradation ability takes longer.At the same time,the temperature difference in the freezing and thawing cycle significantly affects the utilization of oxygen and ion receptors by microorganisms,as well as the degradation effect of petroleum hydrocarbons.Higher freezing and thawing intensities are more conducive to the aerobic or anaerobic degradation effect of microorganisms. |
