Pesticides Biodegradation Performance Of Microorganism And Its Remediation Application In Composite Pesticides Contaminated Soils

Pesticide pollution has become one of the most concerning environmental problems in recent Chinese environmental pollutions.In China,the increasing production and consumption of pesticides,the specific distributed argriculture,the irregular and abusive use of pesticides,etc,aggravate the pesticide pollutions in nowadays.Meanwhile,the production of pesticides and relocation of many pesticides industries have caused the mixture residues of different pesicides and intermediate synthesis organics,leading to the composite pesticide pollutions in soils.Screening some microorganisms with broad-spectrum pesticides degradation abilities have been considered as one of the most efficient solutions in the composite pesticide pollutions remediation in soil.This study aimed at isolating several broad-spectrum pesticides degradation microorganisms by normal domestication procedures,investigating their biodegradation characters and proposed biodegradation pathways,and discussing their impacts on the composite pesticides residues and the soil microorganisms.The main results were as follows:1.The reasearches on the isolation and identification of broad-spectrum pesticides degradation microorganisms.An organophosphate pesticide trichlorfon(TCF)was first utilized in sreening biodegradation microorganisms.By normal domesticating procedures,one bacterial strain PA F-3 and one fungus strain PA F-2 were isolated from pesticide-polluted soil and water samples.These two strains,which both degraded TCF as sole carbon,phosphorus and carbon-phosphorus sources,could also utilize 15 different kinds of pesticides on minimal salt medium(MSM)plates as sole carbon sources.These two strains were identified as Bacillus tequilensis and Aspergillus sydowii,respectively.2.The investigations on the TCF microbial degradation performance.The TCF degradation abilities of PA F-3 and PA F-2 were both impacted by combined environmental factors.Optimal TCF degradation conditions of PA F-3 and PA F-2 were determinded by Response Surface Methodology(RSM).The predicted maximum degradation for the PA F-3 were 88.7%at 28 ℃,125 initial TCF concentration and 4%inoculum amount.While the optimal conditions for the PA F-2 facilitated TCF degradation were as 28 ℃,pH 6.5 and 50 mg/L initial TCF concentration with predicted maximum degradation of 75.3%.Adding exogenous compound yeast extract could significantly imcrease the TCF degradation of PA F-3 and PA F-2,reaching the highest degradation rates of 98.0%and 100%within 5 days,respectively.Meanwhile,the first-order kinetic results revealed that the degradation time(DT50)in yeast-containing experiments were shortened to 1.82 and 0.9 days,respectively.3.The investigations on the TCF microbial degradation pathways.On the basis of the chemical structure and intermediate products,which were detected by gas chromatography-mass spectrometry(GC-MS),the TCF degradation pathways were proposed for the first time.Both PA F-3 and PA F-2 had the similar TCF degradation pathways.TCF was firstly hydrolyzed through the cleavage of the C-P bond to produce dimethyl hydrogen phosphite and chloral hydrate,and then degraded to the biological available phosphorus compounds and glucuronide to microorganisms.The fungus PA F-2 could also degrade the TCF hydrolyzation product dichlorvos(DDVP).Trimethyl orthophosphate and dichloromethane in the DDVP biodegradation pathways were proposed as primary metabolites through the cleavage of C=C double bond.Meanwhile,PA F-2 could also degrade TCF and DDVP to the new intermediate products trimethyl phosphonoacetate,dichloroacetic acid methyl ester and dimethyl ethyphosphonate in order to produce phosphorus compounds and glucuronide.4.The studies on the composite pesticedes biodegradation performance and pathways by PA F-3.PA F-3 and PA F-2 could utilize 15 different kinds of pesticides on minimal salt medium(MSM)plates as sole carbon sources,while bacterium strain PA F-3 had better broad-spectrum pesticide degradation abilities than fungus strain PA F-2.After domesticated by composite pesticides,PA F-3 exhibied better degradation abilities of o-chlorophenol and 2,4-dichlorophenol(2,4-DCP)than 2,4-dichlorophenoxy acetic acid(2,4-D)and 4-methyl-2-chlorophenoxyacetic acid(MCPA),reaching the highest degrading rates of 48.5%and 61.5%in MSM in 14 days,respectively.In MSM containing 5 different pesticides,PA F-3 could completely degrade 2,4-DCP and TCF,and reach the highest degrading rates of 96.1%,69.5%and 71.4%of beta-cypermethrin((3-CP),2,4-D and acetochlor in 41 days.By analyzing the intermediate products of composite pesticides by GC-MS,these results indicated that PA F-3 degraded chlorophenol pesticides to produce phenol through dechlorination and oxidation,and then to produce biological available alcohols and acids through ortho ring-cleavage pathways.The acetochlor biodegradation pathway in PA F-3 suggested that the important intermediate 6-ethyl-2-methylaniline was produced by cleavage of the ethoxymethyl and acylamino bonds,and subsequently degraded to phenol by denitrification.The β-CP biodegradation pathway in PA F-3 demonstrated that(3-CP was initially hydrolyzed to produce the major intermediate product 3-phenoxybenzaldehyde,and then oxidized,dechlorinated to the further metabolisms.5.The studies on the bio-remediation performance of PA F-3 in TCF contaminated soils.The addition of PA F-3 significantly improved the TCF degradation in soil,leading to the completely degradation of 50 mg/kg TCF in 6 days.By comparative analyzing of DGGE and band sequencing results,the dominate species Proteobacteria were greatly affected by TCF in soil without PA F-3 inoculation,while the changes in Actinobacteria and Verrucomicrobia were negligible.However,the inoculation of PA F-3 in soil tremendously increased the relative abundance of Bacillus of Firmicutes and decreased the relative abundance of Actinobacteria in 41 days.These results indicated that TCF slightly impacted the soil bacterial comminities,while the addition of Bacillus sp.strain PA F-3 changed the development of the dominate bacterial species to the degrading bacterial species and reduced the relative abundances of other soil bacterial comminities.By contrast,the relative abundances of fungus communities were improved by the addition of TCF degrading bacterium PA F-3.6.The investigations on the bio-remediation performance of PA F-3 in composite pesticides contaminated soils.In soils containing five composite pesticides,PA F-3 exhibited good broad-spectrum pesticides degradation abilities and completely degraded 50 mg/kg of 2,4-DCP,TCF and β-CP,and reached the highest degrading rates of 91.6%and 90.3%of 2,4-D and acetochlor in 41 days,respectively.Compared to non-inoculated groups,the average degradation rate(v),DT50 and degradation constant(k)in first-order kinetic models were significantly improved.The DGGE and bands sequencing results showed that in soils polluted by the 5 composite pesticides,the dominant bacterial communities were α-,β-andγ-Proteobacteria.However,the addition of PA F-3 greatly changed the development of the dominate bacterial species to the degrading bacterial species Bacillus and reduced the relative abundance of Proteobacteria comminities.The inoculation of the degrading bacterium increased the fungus communities to Ascmycota and Basidiomycota,and mitigated the influences of the composite pesticides to the fungus communities.