Degradation Of Aniline And Crude Oil By Rhodococcus Sp. DH-2 And Its Tolerance To Heavy Metals

Aniline,which is composed of a benzene ring and an amino group,is one of the most important amines and a ubiquitous pollutant.Because aniline often appears in the groundwater environment polluted by crude oil or in the area polluted by heavy metals,the biodegradation efficiency of aniline is reduced.Biodegradation is considered to be an effective,economical and simple process in aniline-contaminated environments.In this paper,a highly efficient aniline-degrading bacterium was isolated and screened from soil contaminated by aniline for a long time,and its metabolic pathway,degradation ability of aniline,degradation ability of crude oil,and ability to tolerate heavy metals were studied.In order to achieve the goal of bioremediation of aniline,crude oil and heavy metals,the multi polluted soil was simulated.The specific research work is as follows:(1)A strain DH-2 was isolated from the soil polluted by aniline for a long time.Through morphological observation and 16 S r RNA gene sequence analysis,it was identified as Rhodococcus.The growth curve of strain DH-2 with ethanol was done,and the optimal seed culture time was determined as 48 h.Under the condition of adding 500-1500 mg / L aniline and shaking culture at 30 ℃ and 200 rpm for 48 h,the degradation rate of aniline by strain DH-2 was more than 50%,indicating that strain DH-2 has a good application prospect.(2)Through single factor optimization experiment,the optimal degradation conditions of aniline by strain DH-2 were determined as follows: temperature 35 ℃,p H 8.0,Na Cl content less than 3%,degradation rate of 1000 mg / L aniline was more than 80% after 36 h.Under these conditions,strain DH-2 could completely degrade aniline at a concentration of 1000mg/ L in 48 h,and the degradation pathway of aniline by strain DH-2 was supposed to be catechol pathway by HPLC-MS analysis.(3)The effect of different crude oil addition amount on DH-2 crude oil degradation was studied.It was proved that strain DH-2 had good degradation effect on crude oil(mass fraction was 2%).Furthermore,the degradation efficiency of aniline and crude oil dual pollution was investigated.The results showed that strain DH-2 could completely degrade aniline in 2 days under the optimal culture condition of 1000 mg / L aniline and 2% crude oil dual pollution,and the degradation rate of crude oil reached 92.6% within 4 days;the expression of aniline and crude oil related degradation genes was detected by real-time fluorescence quantitative PCR,and strain DH-2 had a good degradation effect on aniline and crude oil.Moreover,strain DH-2preferentially used alkanes as carbon source for its own growth under dual pollution,and the presence of alkanes indirectly enhanced the degradation effect of strain DH-2 on aniline.(4)The tolerance of strain DH-2 to heavy metals was studied.The results showed that low concentrations of Pb(II)and Fe(II)could promote the degradation of aniline.The simulated soil bioremediation experiment showed that strain DH-2 had a good degradation effect on the double pollution of 1000 mg / L aniline and 2% crude oil in the presence of 20 mg / L Fe(Ⅱ)and Pb(Ⅱ).Aniline and crude oil were completely degraded after 3 days and 12 days,respectively.The results showed that strain DH-2 had a variety of catabolism abilities,and could tolerate certain concentrations of heavy metals while degrading aniline and crude oil.Low concentrations of Pb(II)and Fe(II)could promote the degradation of strain DH-2.So far,few studies have reported that aniline degrading bacteria can maintain degradation ability in the presence of a variety of pollution.In this paper,the effects of heavy metals on the bioremediation of aniline and crude oil in soil were also studied.The results show that DH-2strain has certain potential to bioremediation of soil under multiple stresses of aniline,heavy metals and crude oil.