Cd-DDT复合污染土壤的植物与微生物联合修复及机理
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摘要
土壤-作物系统中呈现出“新老污染物共存、无机／有机复合污染”的复杂状况,特别是重金属和持久性有机污染物(POPs)造成的复合土壤污染已受到广泛关注。近年来不断爆发的人体、土-水环境污染事件,为污染土壤治理与修复提出了迫切的需求。本研究以重金属(Cd)-有机氯农药(DDT)复合污染农田土壤为研究对象,开展了复合污染土壤植物-微生物联合修复技术及机制研究,取得的主要研究结果如下：
1.研究21份不同基因型南瓜(Cucurbita pepo ssp)品种对复合污染土壤中Cd和DDs (DDT, DDD, DDE的总称)的吸收和积累能力的差异性。结果表明,不同南瓜品种对土壤DDs和Cd的积累及转运能力差异较大。南瓜果实、叶、茎、根DDs的平均含量分别为338.4-793.2、1619-1812、1273-3548、3396-12811ng g-1DW, Cd平均含量分别为0.26-1.12、0.49-2.25、1.04-4.84、1.61-7.72mg kg-1DW.南瓜地上部对Cd和DDs的转运系数(TF)平均值分别为0.79和0.60,富集系数(BAF)则分别为1.68和1.54。经盆栽实验验证,“特别选蜜本王”可以认定为Cd和DDs低积累型南瓜,在中轻度Cd和DDs (Cd≤1.50mg kg-1, DDs≤1.00mg kg-1)复合污染土壤上种植该品种亦可保障农产品的质量安全,而“日本红甜蜜”南瓜具有共富集Cd和DDs的潜力,适宜作为Cd-DDs复合污染土壤的植物修复材料。
2.研发复合污染土壤的生物修复技术是治理污染土壤的重要举措。在Cd-DDs复合污染土壤条件下,盆栽实验中土壤接种DDT-1增加了东南景天的根系生物量,采用东南景天与DDT-1联合修复技术,去除了土壤中32.1-40.3%Cd和33.9-37.6%DDs,随之进行了为期18个月的大田试验,土壤Cd和DDs污染水平分别降低了31.1%和53.6%,大田数据很好地验证了该修复方法的有效性和可行性。可见,应用东南景天与DDT降解菌联合修复技术,是修复Cd-DDs复合污染土壤的有效手段。
3.采用东南景天/小白菜(油冬儿)与DDT-1联合修复技术,增加了油冬儿的根生物量(P<0.05),但对其地上部的生物量无显著影响,增加了东南景天的根部和地上部的生物量(P<0.05)；应用该联合修复技术显著降低了油冬儿根部和地上部Cd含量,降幅分别高达48.0-51.5%和54.7-71.4%；显著降低了其根部总DDs含量,降幅高达37%。土壤Cd和DDs的去除率分别达到30-46%和36.8-42.7%。激光扫描共聚焦显微镜的观测的结果表明,DDT-1大量定殖在根际土壤及东南景天的根毛和根表处。对土壤微生物多样性进行T-RFLP分析,结果表明,在所有包含植物的处理中土壤细菌群落丰富度指数(S)、Shannon指数(H’)、均匀度指数(E)和Simpson指数(D’)明显高于无植物对照组。因此,应用该修复技术既可以修复中轻度复合污染土壤又可以提高小白菜的安全系数,是有应用前景的生物修复策略。
4.采用东南景天／南瓜与DDT-1联合修复技术,显著增加了南瓜根部和地上部的生物量(P<0.05),增幅分别为26.8和21.7%,但对东南景天根部和地上部的生物量均无显著影响。在低Cd-DDs浓度处理中,无明显降低两者根部及地上部的Cd含量,但在高Cd-DDs浓度处理中,降低了南瓜根部的Cd含量(P<0.05),降幅为33.0-40.6%。南瓜根部DDs含量无显著变化,地上部DDs含量则显著降低了38.2-44.5%；东南景天根部DDs含量显著增加了34.7-67.7%,地上部DDs含量显著降低了19.3-39.2%(P<0.05)。土壤Cd和DDs的去除率分别达到41.9-60.7%7.5-45.2%。研究了南瓜中Cd和DDs的微区分布特征,结果表明,南瓜根部Cd主要分布在表皮细胞壁和和维管束,而茎部Cd主要分布在厚壁组织(纤维)和基本组织的液泡；南瓜根部DDs主要分布在中柱和吸附在根毛表面,而茎部DDs主要分布在维管束、基本组织的液泡和腺毛体；激光扫描共聚焦显微镜观测结果表明,DDT-1菌株在东南景天与南瓜根部大量定殖。对土壤微生物多样性进行T-RFLP分析,结果表明,该修复技术对土壤细菌群落丰富度指数(S)、Shannon指数(H’)、均匀度指数(E)和Simpson指数(D’)有明显影响,土壤细菌群落多样性明显丰富。因此,应用该修复技术既可以修复中轻度复合污染土壤又可以提高南瓜的安全系数,是有应用前景的生物修复策略。
Environmental pollution of the soil-crop system is very complex due to the coexistence of old and new pollutants, as well as inorganic and organic compounds. Especially soil pollution caused by heavy metals and persistent organic pollutants (POPs) is widespread concerned. In recent years, the frequent outbreak of environmental pollution accidents due to soil-water pollution puts forward urgent demand for remediation and restoration of contaminated soil.
In this research, the plant-microbe remediation of Cd and DDT was carried out under pot and field experiments to identify bioremediation strategy of Cd-DDs co-contaminated soil and to investigate the mechanism for enhancing bioremediation efficiency. Primary results were summarized as follows:
1. Cadmium (Cd) and dichlorodiphenyltrichloroethane (DDT) or its metabolite residues DDD/DDE (DDT, DDE, and DDD are collectively called DDs) are frequently detected in agricultural soils and agricultural products, posing a threat to human health. The objective of this study was to compare the ability of21genotypes of Cucurbita pepo ssp in mobilizing and uptake of Cd and DDs (p,p'-DDT, o,p-DDT, p,p'-DDD and p,p'-DDE) in the co-contaminated soil. The plant genotypes varied greatly in the uptake and accumulation of Cd and DDs, with mean concentrations of0.26-1.12、0.49-2.25、1.04-4.84and1.61-7.72mg kg-1DWfor Cd, and338.4-793.2,1619-1812,1273-3548and3396-12811ng g-1DW for DDs in leaf, stem and root, respectively. The TF and BAF values were0.79and1.68for Cd, and0.60and1.54for DDs, respectively. These results indicate that Cucurbita pepo cv."Tebiexuan mibenwang" has a low ability to absorb and accumulate Cd and DDs from the contaminated soils, but Cucurbita pepo cv."Riben Hongtianmi" has great potential for accumulating Cd and DDs from moderately co-contaminated soil (Cd≤1.50mg kg-1, DDs≤1.00mg kg-1)。
2. The development of an integrated strategy for the remediation of soil co-contaminated by heavy metals and persistent organic pollutants is a major research priority for the decontamination of soil slated for use in agricultural production. The objective of this study was to develop a bioremediation strategy for fields co-contaminated with cadmium (Cd), dichlorodiphenyltrichloroethane (DDT), and its metabolites1,1-dichloro-2,2-bis (4-chlorophenyl) ethylene (DDE) and1,1-dichloro-2,2-bis (4-chlorophenyl) ethane (DDD) using an identified Cd-hyperaccumulator plant Sedum alfredii (SA) and DDT-degrading microbes (DDT-1). Initially, inoculation with DDT-1was shown to increase SA root biomass in a pot experiment. When SA was applied together with DDT-1, the levels of Cd and DDs in the co-contaminated soil decreased by32.1-40.3%and33.9-37.6%, respectively, in a pot experiment over18months compared to3.25%and3.76%decreases in soil Cd and DDs, respectively, in unplanted, untreated controls. A subsequent field study (18-month duration) in which the levels of Cd and DDs decreased by31.1%and53.6%, respectively, confirmed the beneficial results of this approach. This study demonstrates that the integrated bioremediation strategy is effective for the remediation of Cd-DDs co-contaminated soils.
3. Co-cropping, a widely accepted agronomical practice in China for2000years, can increase total crop yields through increased resource use efficiency. In our present experiments, co-cropping of Sedum alfredii and B. campestris ssp chinensis associated with DDT-1increased the biomass and metal phytoextraction of S. alfredii (P＜0.05) and also enhanced the root growth of B. campestris ssp chinensis (P＜0.05), but had no significant effects on shoot biomass of B. campestris ssp chinensis. The combined remediation strategy decreased the Cd concentration by48.0-51.5%and54.7-71.4%in the roots and shoots of B. campestris ssp chinensis, respectively, and also corresponded to a decrease in root DDs concentration by37%. The removal efficiencies were30-46%for Cd and36.8-42.7%for DDs. Laser scanning confocal microscopy revealed that gfp-tagged DDT-1heavily colonized in rhizosphere soil, and on rhizoplane of Sedum alfredii. The soil diversity indices, such as richness index (S), evenness index (E), Shannon's diversity (H'), Simpson's index (D'), were higher in all of the planted treatments than in the unplanted control. Principal component analysis of bacterial T-RFLP data also revealed strong shifts in bacterial community composition with the planted treatments. The results of this study indicate that the co-cropping of Sedum alfredii and B. campestris ssp chinensis associated with DDT-1appears to be a promising approach for the bioremediation of soils co-contaminated by Cd and DDs, while simultaneously decreasing the pollutant concentration in edible part of B. campestris ssp Chinensis, and hence maintaining product safety and and reliability of this vegetable.
4. In our present experiments, co-cropping Sedum alfredii and Cucurbita pepo cv."Tebiexuan mibenwang" associated with DDT-1increased the root (26.8%) and shoot biomass (21.7%) of "Tebiexuan mibenwang"(P＜0.05), but had no significant effects on the root and shoot biomass of Sedum alfredii. In general, the combined remediation strategy did not significantly decrease the Cd concentrations in roots and shoots of both plants. However, for "Tebiexuan mibenwang", the bioremediation strategy decreased the shoot DDs concentration by38.2-44.5%(P＜0.05), but had no significant effects on root DDs concentration; for Sedum alfredii, increased and decreased DDs concentration by34.7-67.7%and38.2-44.5%in roots and shoots of Sedum alfredii, respectively. The removal efficiencies were41.9-60.7%for Cd and37.5-45.2%for DDs. In the present study, for roots of "Tebiexuan mibenwang", Cd was largely retained in the cell walls and vessel tissue, DDs was largely retained in central cylinder, and was adsorbed on root hairs, but for shoots, Cd was largely retained in the collenchyma tissue and vacuoles, DDs was largely retained in vessel tissue, vacuoles and glandular trichomes. Laser scanning confocal microscopy revealed that gfp-tagged DDT-1heavily colonized on rhizoplane of Sedum alfredii and "Tebiexuan mibenwang". The soil diversity indices, such as richness index (S), evenness index (E), Shannon's diversity (H'), Simpson's index (D'), were higher in the treatment than in the unplanted control. The results of this study indicate that the co-cropping of Sedum alfredii and "Tebiexuan mibenwang" associated with DDT-1appears to be a promising approach for the bioremediation of soils co-contaminated by Cd and DDs, and while decreasing the DDs concentration in shoot of "Tebiexuan mibenwang", and hence ensuring product safety of the pumpkin.
1.研究21份不同基因型南瓜(Cucurbita pepo ssp)品种对复合污染土壤中Cd和DDs (DDT, DDD, DDE的总称)的吸收和积累能力的差异性。结果表明,不同南瓜品种对土壤DDs和Cd的积累及转运能力差异较大。南瓜果实、叶、茎、根DDs的平均含量分别为338.4-793.2、1619-1812、1273-3548、3396-12811ng g-1DW, Cd平均含量分别为0.26-1.12、0.49-2.25、1.04-4.84、1.61-7.72mg kg-1DW.南瓜地上部对Cd和DDs的转运系数(TF)平均值分别为0.79和0.60,富集系数(BAF)则分别为1.68和1.54。经盆栽实验验证,“特别选蜜本王”可以认定为Cd和DDs低积累型南瓜,在中轻度Cd和DDs (Cd≤1.50mg kg-1, DDs≤1.00mg kg-1)复合污染土壤上种植该品种亦可保障农产品的质量安全,而“日本红甜蜜”南瓜具有共富集Cd和DDs的潜力,适宜作为Cd-DDs复合污染土壤的植物修复材料。
2.研发复合污染土壤的生物修复技术是治理污染土壤的重要举措。在Cd-DDs复合污染土壤条件下,盆栽实验中土壤接种DDT-1增加了东南景天的根系生物量,采用东南景天与DDT-1联合修复技术,去除了土壤中32.1-40.3%Cd和33.9-37.6%DDs,随之进行了为期18个月的大田试验,土壤Cd和DDs污染水平分别降低了31.1%和53.6%,大田数据很好地验证了该修复方法的有效性和可行性。可见,应用东南景天与DDT降解菌联合修复技术,是修复Cd-DDs复合污染土壤的有效手段。
3.采用东南景天/小白菜(油冬儿)与DDT-1联合修复技术,增加了油冬儿的根生物量(P<0.05),但对其地上部的生物量无显著影响,增加了东南景天的根部和地上部的生物量(P<0.05)；应用该联合修复技术显著降低了油冬儿根部和地上部Cd含量,降幅分别高达48.0-51.5%和54.7-71.4%；显著降低了其根部总DDs含量,降幅高达37%。土壤Cd和DDs的去除率分别达到30-46%和36.8-42.7%。激光扫描共聚焦显微镜的观测的结果表明,DDT-1大量定殖在根际土壤及东南景天的根毛和根表处。对土壤微生物多样性进行T-RFLP分析,结果表明,在所有包含植物的处理中土壤细菌群落丰富度指数(S)、Shannon指数(H’)、均匀度指数(E)和Simpson指数(D’)明显高于无植物对照组。因此,应用该修复技术既可以修复中轻度复合污染土壤又可以提高小白菜的安全系数,是有应用前景的生物修复策略。
4.采用东南景天／南瓜与DDT-1联合修复技术,显著增加了南瓜根部和地上部的生物量(P<0.05),增幅分别为26.8和21.7%,但对东南景天根部和地上部的生物量均无显著影响。在低Cd-DDs浓度处理中,无明显降低两者根部及地上部的Cd含量,但在高Cd-DDs浓度处理中,降低了南瓜根部的Cd含量(P<0.05),降幅为33.0-40.6%。南瓜根部DDs含量无显著变化,地上部DDs含量则显著降低了38.2-44.5%；东南景天根部DDs含量显著增加了34.7-67.7%,地上部DDs含量显著降低了19.3-39.2%(P<0.05)。土壤Cd和DDs的去除率分别达到41.9-60.7%7.5-45.2%。研究了南瓜中Cd和DDs的微区分布特征,结果表明,南瓜根部Cd主要分布在表皮细胞壁和和维管束,而茎部Cd主要分布在厚壁组织(纤维)和基本组织的液泡；南瓜根部DDs主要分布在中柱和吸附在根毛表面,而茎部DDs主要分布在维管束、基本组织的液泡和腺毛体；激光扫描共聚焦显微镜观测结果表明,DDT-1菌株在东南景天与南瓜根部大量定殖。对土壤微生物多样性进行T-RFLP分析,结果表明,该修复技术对土壤细菌群落丰富度指数(S)、Shannon指数(H’)、均匀度指数(E)和Simpson指数(D’)有明显影响,土壤细菌群落多样性明显丰富。因此,应用该修复技术既可以修复中轻度复合污染土壤又可以提高南瓜的安全系数,是有应用前景的生物修复策略。
Environmental pollution of the soil-crop system is very complex due to the coexistence of old and new pollutants, as well as inorganic and organic compounds. Especially soil pollution caused by heavy metals and persistent organic pollutants (POPs) is widespread concerned. In recent years, the frequent outbreak of environmental pollution accidents due to soil-water pollution puts forward urgent demand for remediation and restoration of contaminated soil.
In this research, the plant-microbe remediation of Cd and DDT was carried out under pot and field experiments to identify bioremediation strategy of Cd-DDs co-contaminated soil and to investigate the mechanism for enhancing bioremediation efficiency. Primary results were summarized as follows:
1. Cadmium (Cd) and dichlorodiphenyltrichloroethane (DDT) or its metabolite residues DDD/DDE (DDT, DDE, and DDD are collectively called DDs) are frequently detected in agricultural soils and agricultural products, posing a threat to human health. The objective of this study was to compare the ability of21genotypes of Cucurbita pepo ssp in mobilizing and uptake of Cd and DDs (p,p'-DDT, o,p-DDT, p,p'-DDD and p,p'-DDE) in the co-contaminated soil. The plant genotypes varied greatly in the uptake and accumulation of Cd and DDs, with mean concentrations of0.26-1.12、0.49-2.25、1.04-4.84and1.61-7.72mg kg-1DWfor Cd, and338.4-793.2,1619-1812,1273-3548and3396-12811ng g-1DW for DDs in leaf, stem and root, respectively. The TF and BAF values were0.79and1.68for Cd, and0.60and1.54for DDs, respectively. These results indicate that Cucurbita pepo cv."Tebiexuan mibenwang" has a low ability to absorb and accumulate Cd and DDs from the contaminated soils, but Cucurbita pepo cv."Riben Hongtianmi" has great potential for accumulating Cd and DDs from moderately co-contaminated soil (Cd≤1.50mg kg-1, DDs≤1.00mg kg-1)。
2. The development of an integrated strategy for the remediation of soil co-contaminated by heavy metals and persistent organic pollutants is a major research priority for the decontamination of soil slated for use in agricultural production. The objective of this study was to develop a bioremediation strategy for fields co-contaminated with cadmium (Cd), dichlorodiphenyltrichloroethane (DDT), and its metabolites1,1-dichloro-2,2-bis (4-chlorophenyl) ethylene (DDE) and1,1-dichloro-2,2-bis (4-chlorophenyl) ethane (DDD) using an identified Cd-hyperaccumulator plant Sedum alfredii (SA) and DDT-degrading microbes (DDT-1). Initially, inoculation with DDT-1was shown to increase SA root biomass in a pot experiment. When SA was applied together with DDT-1, the levels of Cd and DDs in the co-contaminated soil decreased by32.1-40.3%and33.9-37.6%, respectively, in a pot experiment over18months compared to3.25%and3.76%decreases in soil Cd and DDs, respectively, in unplanted, untreated controls. A subsequent field study (18-month duration) in which the levels of Cd and DDs decreased by31.1%and53.6%, respectively, confirmed the beneficial results of this approach. This study demonstrates that the integrated bioremediation strategy is effective for the remediation of Cd-DDs co-contaminated soils.
3. Co-cropping, a widely accepted agronomical practice in China for2000years, can increase total crop yields through increased resource use efficiency. In our present experiments, co-cropping of Sedum alfredii and B. campestris ssp chinensis associated with DDT-1increased the biomass and metal phytoextraction of S. alfredii (P＜0.05) and also enhanced the root growth of B. campestris ssp chinensis (P＜0.05), but had no significant effects on shoot biomass of B. campestris ssp chinensis. The combined remediation strategy decreased the Cd concentration by48.0-51.5%and54.7-71.4%in the roots and shoots of B. campestris ssp chinensis, respectively, and also corresponded to a decrease in root DDs concentration by37%. The removal efficiencies were30-46%for Cd and36.8-42.7%for DDs. Laser scanning confocal microscopy revealed that gfp-tagged DDT-1heavily colonized in rhizosphere soil, and on rhizoplane of Sedum alfredii. The soil diversity indices, such as richness index (S), evenness index (E), Shannon's diversity (H'), Simpson's index (D'), were higher in all of the planted treatments than in the unplanted control. Principal component analysis of bacterial T-RFLP data also revealed strong shifts in bacterial community composition with the planted treatments. The results of this study indicate that the co-cropping of Sedum alfredii and B. campestris ssp chinensis associated with DDT-1appears to be a promising approach for the bioremediation of soils co-contaminated by Cd and DDs, while simultaneously decreasing the pollutant concentration in edible part of B. campestris ssp Chinensis, and hence maintaining product safety and and reliability of this vegetable.
4. In our present experiments, co-cropping Sedum alfredii and Cucurbita pepo cv."Tebiexuan mibenwang" associated with DDT-1increased the root (26.8%) and shoot biomass (21.7%) of "Tebiexuan mibenwang"(P＜0.05), but had no significant effects on the root and shoot biomass of Sedum alfredii. In general, the combined remediation strategy did not significantly decrease the Cd concentrations in roots and shoots of both plants. However, for "Tebiexuan mibenwang", the bioremediation strategy decreased the shoot DDs concentration by38.2-44.5%(P＜0.05), but had no significant effects on root DDs concentration; for Sedum alfredii, increased and decreased DDs concentration by34.7-67.7%and38.2-44.5%in roots and shoots of Sedum alfredii, respectively. The removal efficiencies were41.9-60.7%for Cd and37.5-45.2%for DDs. In the present study, for roots of "Tebiexuan mibenwang", Cd was largely retained in the cell walls and vessel tissue, DDs was largely retained in central cylinder, and was adsorbed on root hairs, but for shoots, Cd was largely retained in the collenchyma tissue and vacuoles, DDs was largely retained in vessel tissue, vacuoles and glandular trichomes. Laser scanning confocal microscopy revealed that gfp-tagged DDT-1heavily colonized on rhizoplane of Sedum alfredii and "Tebiexuan mibenwang". The soil diversity indices, such as richness index (S), evenness index (E), Shannon's diversity (H'), Simpson's index (D'), were higher in the treatment than in the unplanted control. The results of this study indicate that the co-cropping of Sedum alfredii and "Tebiexuan mibenwang" associated with DDT-1appears to be a promising approach for the bioremediation of soils co-contaminated by Cd and DDs, and while decreasing the DDs concentration in shoot of "Tebiexuan mibenwang", and hence ensuring product safety of the pumpkin.
引文
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