土壤动物（线虫、蚯蚓）对污染土壤多环芳烃去除的影响
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摘要
随着工农业的迅速发展,大量的有机污染物进入土壤,严重影响了土壤环境质量及其生态功能,尤其是POPs、PAHs和PCBs有机污染在我国土壤中均有不同程度的输出。由于多环芳烃的毒性作用和土壤中过量存在严重影响和抑制作物的生长发育,降低农产品品质,通过食物链的富集传递作用,危害人体健康,故PAHs等污染问题已成为人类面临的十分严重的环境问题。开展对土壤有机污染的生物修复研究,对防治有机污染土壤修复具有重要意义。
土壤动物(线虫和蚯蚓)作为土壤生物的重要组成部分,与土壤微生物之间存在着密切的关系,并在生物修复过程中发挥着重要的作用,成为PAHs污染土壤生物修复的研究热点。然而,土壤动物(线虫和蚯蚓)在PAHs污染的生物修复体系(土壤动物-微生物-植物)中的作用及机理还不清楚。因此,本文以线虫和蚯蚓作为土壤动物的代表,菲和荧蒽作为PAHs污染物的代表,设置了一系列的室内培养试验,主要研究：(1)食细菌线虫与恶臭假单胞菌相互作用对砂培体系中菲去除的影响及外加碳源(葡萄糖)与否的影响；(2)食细菌线虫与恶臭假单胞菌相互作用对未灭菌土壤中菲去除的影响；(3)蚯蚓与黑麦草的相互作用对土壤中荧蒽去除的影响；(4)蚯蚓粘液对黑麦草生长及其对荧蒽去除的影响。以试图揭示线虫和蚯蚓在PAHs污染的生物修复体系(土壤动物-微生物-植物)中的作用及机理,为线虫和蚯蚓在PAHs污染的生物修复中的应用提供理论依据。
主要结果如下：
(1)通过悉生培养试验研究食细菌线虫与恶臭假单胞菌相互作用对砂培体系中细菌去除菲的影响。结果显示,与对照及只接种细菌的处理相比,接种线虫显著促进处理中菲的去除(P<0.05)；接种食细菌线虫处理还促进了C230酶活性和过氧化氢酶活性的增强。
(2)采用悉生培养试验,探讨了外加碳源(葡萄糖)对食细菌线虫和恶臭假单胞菌的相互作用去除砂培体系中菲的影响。结果表明,与对照相比,外加碳源葡萄糖和接种食细菌线虫处理在培养结束时菲的去除率分别提高了25.6%和36.6%；在外加葡萄糖条件下,食细菌线虫显著提高了培养体系中细菌数量以及C230酶活性。因此,葡萄糖和食细菌线虫可能通过刺激细菌数量和C230酶活性来促进培养体系中菲的去除。
(3)通过室内培养试验研究了食细菌线虫与恶臭假单胞菌相互作用对未灭菌土壤中菲去除的影响。结果表明,与对照相比,接种食细菌线虫处理能显著(P<0.05)促进土壤酶活性(过氧化氢酶和FDA水解酶)。但是接种线虫处理并未显著促进土壤中菲的去除,可能是由于复杂的土壤环境所致。
(4)通过室内土培试验研究了蚯蚓和黑麦草相互作用对土壤中荧蒽去除的影响。结果表明,各处理条件下土壤中荧蒽浓度均按照一阶指数方程递减,其中种植黑麦草同时接种蚯蚓的处理土壤荧蒽的半衰期(5.3天)最小,比对照减少了36.3%；在培养结束(第70天)时,蚯蚓和黑麦草处理显著促进土壤微生物生物量及过氧化物酶活性(P<0.01)；相关分析表明,微生物生物量和过氧化物酶活性与土壤中荧蒽的残留量均呈显著(P<0.01)的负相关关系。因此,蚯蚓促进土壤中荧蒽的去除与土壤微生物生物量和过氧化物酶活性密切相关。
(5)通过室内水培试验,探讨蚯蚓粘液对黑麦草生长及其对荧蒽去除的影响。结果表明,与只添加蚯蚓粘液和只种植黑麦草的处理相比,两者的交互作用对培养体系中荧蒽的去除率更高,尤其在培养第12天时最显著。蚯蚓粘液促进黑麦草根系生长以及超氧化物歧化酶(SOD)的活性(P<0.05)；蚯蚓粘液提高了荧蒽在黑麦草体内的富集量以及黑麦草对荧蒽的传导系数,这可能是蚯蚓促进土壤荧蒽去除的机理之一。
以上结果表明,(1)食细菌线虫能促进悉生培养体系中菲的去除；(2)外加碳源(葡萄糖)可有效的提高食细菌线虫对细菌数量的增多和C230酶活性的增强,进而有助于培养体系中菲的去除；(3)蚯蚓存在条件下,土壤中荧蒽的去除跟土壤中微生物生物量与过氧化物酶(POD)活性密切相关；(4)蚯蚓粘液可有效的促进微生物的生长繁殖、黑麦草根系生长及其超氧化物歧化酶(SOD)的活性,以及黑麦草对荧蒽富集量和传导系数的提高。
With the rapid development of industry and agriculture, there are amounts of organic contaminants enter into the soil, which seriously affect the soil environmental quality and ecological function. Especially the POPs, PAHs and PCBs, of which there are different levels of output in soil in our country. As the PAHs toxicity and its amounts in soil seriously inhibit the crops growth, it reduces the quality of agricultural product. Moreover, it is also harmful to human health through the food chain enrichment transmission function. The PAHs pollution problem becomes the most serious environmental problems. The development of bioremediation research is conducive to the remediation of organic contaminated soil.
Many researches have focused on the effects of soil fauna (nematodes and earthworms) on the bioremediation of polycyclic aromatic hydrocarbons (PAHs) contaminated soil because soil fauna were closely related to soil microorganisms and played key roles in bioremediating the contaminated soils. However, the inherent mechanisms of soil fauna in the bioremediation of the PAHs contaminated soils were still unclear. Therefore, we conducted a series of experiments to investigate the effects of soil fauna (nematodes and earthworms) on the bioremediation of PAHs contaminated soils. Our studies were mainly concentrated on:(1) Effects of bacterial-feeding nematodes and bacteria(Pseudomonas Putida) interactions, in the presence of glucose or not, on the degradation of phenanthrene in gnotobiotic system;(2) Effects of bacterial-feeding nematodes on phenanthrene removal from the contaminated soil;(3) Effects of earthworms and ryegrass interaction on the removal of fluoranthene from contaminated soil;(4) Effects of earthworm mucus on ryegrass growth and the removal of fluoranthene from hydroponic system.
The main results were shown as following:
(1) A gnotobiotic culture experiment was constructed to study the effects of bacterial-feeding nematodes and bacteria(Pseudomonas Putida) interactions on phenanthrene removal from the gnotobiotic system. Bacterial-feeding nematodes could promote the removal of phenanthrene from gnotobiotic system. Compared with the treatments of control and bacteria inoculated only, the inoculation of bacterial-feeding nematodes significantly accelerated the phenanthrene removal from gnotobiotic system (P <0.05). Bacterial-feeding nematodes also could promote the catechol2,3-dioxygenase (C23O) and catalase (CAT) activity.
(2) The effect of adding carbon (glucose) and bacterial-feeding nematodes on the phenanthrene removal was investigated in a gnotobiotic system. Compared with control, the removal of phenanthrene was increased by25.6%and36.6%in the presence of carbon (glucose) and bacterial-feeding nematodes, respectively. Nematodes could also significantly increase the bacterial abundance and the catechol2,3-dioxygenase (C23O) activity, especially when the glucose present. These results implied that nematodes might promote the removal of phenanthrene from medium via stimulating bacteria performance and catechol2,3-dioxygenase (C23O) activity.
(3) A soil microcosm incubation experiment was constructed to study the effect of bacteria-feeding nematodes, which feed phenanthrene-degrading bacteria (Pseudomonas Putida), on phenanthrene removal from the unsterilied soil. Compared with control, the inoculation of bacteria-feeding nematodes could increase the soil enzyme activities (Catalase and Hydrolysis rate of fluorescein diacetate)(P<0.05). But inoculation of bacteria-feeding nematodes could not significantly increase the removal of phenanthrene from contaminated soils. It was contributed to the complex environment in soil.
(4) The effects of earthworms and ryegrass interaction on the removal of fluoranthene from contaminated soil were investigated in a70-day microcosm experiment. During the incubation time, the changes of fluoranthene in soil among all four treatments were fitted with the first-order kinetic model. The half-life (T1/2) of the fluoranthene in soil with earthworms and ryegrass together was the smallest among all treatments and36.3%smaller than that of control. Earthworms significantly increased microbial biomass and polyphenol oxidase activity (POD) in the presence of ryegrass at the final sampling (P<0.01). Microbial biomass and polyphenol oxidase activity (POD) were significantly (P<0.05) and negatively related to the residual fluoranthene concentration, respectively. The results implied that earthworms might promote the removal of fluoranthene from soil via stimulating microbial biomass and POD activity.
(5) A hydroponic culture study was carried out to study the effects of earthworm mucus on ryegrass growth and the removal of fluoranthene from hydroponic system. Results indicated that compared with treatments with only amending earthworm mucus or growing ryegrass, the interaction between earthworm mucus and ryegrass was more significantly on the removal of fluoranthene, especially on the12th day. Earthworm mucus significantly increased ryegrass root growth and superoxide dismutase (SOD) activity (P<0.05). The earthworm mucus also promoted the concentration and translocation factor (TF) of fluoranthene in plant.
In conclusion, these results demonstrated:1) Bacteria-feeding nematodes promoted the removal of phenanthrene from the contaminated gnotobiotic culture;2) Adding the carbon (glucose) could promote the stimulation effects of the bacteria-feeding nematodes on the bacteria abundance and C23O activity, and be helpful on the removal of phenanthrene from the gnotobiotic system;3) In the presence of earthworm, the fluoranthene removal from contaminated soil were closely related to the increase of soil microbial biomass and polyphenol oxidizes (POD) activity;4) Earthworm mucus significantly increased the bacterial growth and reproduction, and ryegrass root growth and superoxide dismutase (SOD) activity. Earthworm mucus also promoted the concentration and translocation factor (TF) of fluoranthene in plant.
土壤动物(线虫和蚯蚓)作为土壤生物的重要组成部分,与土壤微生物之间存在着密切的关系,并在生物修复过程中发挥着重要的作用,成为PAHs污染土壤生物修复的研究热点。然而,土壤动物(线虫和蚯蚓)在PAHs污染的生物修复体系(土壤动物-微生物-植物)中的作用及机理还不清楚。因此,本文以线虫和蚯蚓作为土壤动物的代表,菲和荧蒽作为PAHs污染物的代表,设置了一系列的室内培养试验,主要研究：(1)食细菌线虫与恶臭假单胞菌相互作用对砂培体系中菲去除的影响及外加碳源(葡萄糖)与否的影响；(2)食细菌线虫与恶臭假单胞菌相互作用对未灭菌土壤中菲去除的影响；(3)蚯蚓与黑麦草的相互作用对土壤中荧蒽去除的影响；(4)蚯蚓粘液对黑麦草生长及其对荧蒽去除的影响。以试图揭示线虫和蚯蚓在PAHs污染的生物修复体系(土壤动物-微生物-植物)中的作用及机理,为线虫和蚯蚓在PAHs污染的生物修复中的应用提供理论依据。
主要结果如下：
(1)通过悉生培养试验研究食细菌线虫与恶臭假单胞菌相互作用对砂培体系中细菌去除菲的影响。结果显示,与对照及只接种细菌的处理相比,接种线虫显著促进处理中菲的去除(P<0.05)；接种食细菌线虫处理还促进了C230酶活性和过氧化氢酶活性的增强。
(2)采用悉生培养试验,探讨了外加碳源(葡萄糖)对食细菌线虫和恶臭假单胞菌的相互作用去除砂培体系中菲的影响。结果表明,与对照相比,外加碳源葡萄糖和接种食细菌线虫处理在培养结束时菲的去除率分别提高了25.6%和36.6%；在外加葡萄糖条件下,食细菌线虫显著提高了培养体系中细菌数量以及C230酶活性。因此,葡萄糖和食细菌线虫可能通过刺激细菌数量和C230酶活性来促进培养体系中菲的去除。
(3)通过室内培养试验研究了食细菌线虫与恶臭假单胞菌相互作用对未灭菌土壤中菲去除的影响。结果表明,与对照相比,接种食细菌线虫处理能显著(P<0.05)促进土壤酶活性(过氧化氢酶和FDA水解酶)。但是接种线虫处理并未显著促进土壤中菲的去除,可能是由于复杂的土壤环境所致。
(4)通过室内土培试验研究了蚯蚓和黑麦草相互作用对土壤中荧蒽去除的影响。结果表明,各处理条件下土壤中荧蒽浓度均按照一阶指数方程递减,其中种植黑麦草同时接种蚯蚓的处理土壤荧蒽的半衰期(5.3天)最小,比对照减少了36.3%；在培养结束(第70天)时,蚯蚓和黑麦草处理显著促进土壤微生物生物量及过氧化物酶活性(P<0.01)；相关分析表明,微生物生物量和过氧化物酶活性与土壤中荧蒽的残留量均呈显著(P<0.01)的负相关关系。因此,蚯蚓促进土壤中荧蒽的去除与土壤微生物生物量和过氧化物酶活性密切相关。
(5)通过室内水培试验,探讨蚯蚓粘液对黑麦草生长及其对荧蒽去除的影响。结果表明,与只添加蚯蚓粘液和只种植黑麦草的处理相比,两者的交互作用对培养体系中荧蒽的去除率更高,尤其在培养第12天时最显著。蚯蚓粘液促进黑麦草根系生长以及超氧化物歧化酶(SOD)的活性(P<0.05)；蚯蚓粘液提高了荧蒽在黑麦草体内的富集量以及黑麦草对荧蒽的传导系数,这可能是蚯蚓促进土壤荧蒽去除的机理之一。
以上结果表明,(1)食细菌线虫能促进悉生培养体系中菲的去除；(2)外加碳源(葡萄糖)可有效的提高食细菌线虫对细菌数量的增多和C230酶活性的增强,进而有助于培养体系中菲的去除；(3)蚯蚓存在条件下,土壤中荧蒽的去除跟土壤中微生物生物量与过氧化物酶(POD)活性密切相关；(4)蚯蚓粘液可有效的促进微生物的生长繁殖、黑麦草根系生长及其超氧化物歧化酶(SOD)的活性,以及黑麦草对荧蒽富集量和传导系数的提高。
With the rapid development of industry and agriculture, there are amounts of organic contaminants enter into the soil, which seriously affect the soil environmental quality and ecological function. Especially the POPs, PAHs and PCBs, of which there are different levels of output in soil in our country. As the PAHs toxicity and its amounts in soil seriously inhibit the crops growth, it reduces the quality of agricultural product. Moreover, it is also harmful to human health through the food chain enrichment transmission function. The PAHs pollution problem becomes the most serious environmental problems. The development of bioremediation research is conducive to the remediation of organic contaminated soil.
Many researches have focused on the effects of soil fauna (nematodes and earthworms) on the bioremediation of polycyclic aromatic hydrocarbons (PAHs) contaminated soil because soil fauna were closely related to soil microorganisms and played key roles in bioremediating the contaminated soils. However, the inherent mechanisms of soil fauna in the bioremediation of the PAHs contaminated soils were still unclear. Therefore, we conducted a series of experiments to investigate the effects of soil fauna (nematodes and earthworms) on the bioremediation of PAHs contaminated soils. Our studies were mainly concentrated on:(1) Effects of bacterial-feeding nematodes and bacteria(Pseudomonas Putida) interactions, in the presence of glucose or not, on the degradation of phenanthrene in gnotobiotic system;(2) Effects of bacterial-feeding nematodes on phenanthrene removal from the contaminated soil;(3) Effects of earthworms and ryegrass interaction on the removal of fluoranthene from contaminated soil;(4) Effects of earthworm mucus on ryegrass growth and the removal of fluoranthene from hydroponic system.
The main results were shown as following:
(1) A gnotobiotic culture experiment was constructed to study the effects of bacterial-feeding nematodes and bacteria(Pseudomonas Putida) interactions on phenanthrene removal from the gnotobiotic system. Bacterial-feeding nematodes could promote the removal of phenanthrene from gnotobiotic system. Compared with the treatments of control and bacteria inoculated only, the inoculation of bacterial-feeding nematodes significantly accelerated the phenanthrene removal from gnotobiotic system (P <0.05). Bacterial-feeding nematodes also could promote the catechol2,3-dioxygenase (C23O) and catalase (CAT) activity.
(2) The effect of adding carbon (glucose) and bacterial-feeding nematodes on the phenanthrene removal was investigated in a gnotobiotic system. Compared with control, the removal of phenanthrene was increased by25.6%and36.6%in the presence of carbon (glucose) and bacterial-feeding nematodes, respectively. Nematodes could also significantly increase the bacterial abundance and the catechol2,3-dioxygenase (C23O) activity, especially when the glucose present. These results implied that nematodes might promote the removal of phenanthrene from medium via stimulating bacteria performance and catechol2,3-dioxygenase (C23O) activity.
(3) A soil microcosm incubation experiment was constructed to study the effect of bacteria-feeding nematodes, which feed phenanthrene-degrading bacteria (Pseudomonas Putida), on phenanthrene removal from the unsterilied soil. Compared with control, the inoculation of bacteria-feeding nematodes could increase the soil enzyme activities (Catalase and Hydrolysis rate of fluorescein diacetate)(P<0.05). But inoculation of bacteria-feeding nematodes could not significantly increase the removal of phenanthrene from contaminated soils. It was contributed to the complex environment in soil.
(4) The effects of earthworms and ryegrass interaction on the removal of fluoranthene from contaminated soil were investigated in a70-day microcosm experiment. During the incubation time, the changes of fluoranthene in soil among all four treatments were fitted with the first-order kinetic model. The half-life (T1/2) of the fluoranthene in soil with earthworms and ryegrass together was the smallest among all treatments and36.3%smaller than that of control. Earthworms significantly increased microbial biomass and polyphenol oxidase activity (POD) in the presence of ryegrass at the final sampling (P<0.01). Microbial biomass and polyphenol oxidase activity (POD) were significantly (P<0.05) and negatively related to the residual fluoranthene concentration, respectively. The results implied that earthworms might promote the removal of fluoranthene from soil via stimulating microbial biomass and POD activity.
(5) A hydroponic culture study was carried out to study the effects of earthworm mucus on ryegrass growth and the removal of fluoranthene from hydroponic system. Results indicated that compared with treatments with only amending earthworm mucus or growing ryegrass, the interaction between earthworm mucus and ryegrass was more significantly on the removal of fluoranthene, especially on the12th day. Earthworm mucus significantly increased ryegrass root growth and superoxide dismutase (SOD) activity (P<0.05). The earthworm mucus also promoted the concentration and translocation factor (TF) of fluoranthene in plant.
In conclusion, these results demonstrated:1) Bacteria-feeding nematodes promoted the removal of phenanthrene from the contaminated gnotobiotic culture;2) Adding the carbon (glucose) could promote the stimulation effects of the bacteria-feeding nematodes on the bacteria abundance and C23O activity, and be helpful on the removal of phenanthrene from the gnotobiotic system;3) In the presence of earthworm, the fluoranthene removal from contaminated soil were closely related to the increase of soil microbial biomass and polyphenol oxidizes (POD) activity;4) Earthworm mucus significantly increased the bacterial growth and reproduction, and ryegrass root growth and superoxide dismutase (SOD) activity. Earthworm mucus also promoted the concentration and translocation factor (TF) of fluoranthene in plant.
引文
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