高效诱变菌与生物炭复合修复重金属污染土壤的研究
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摘要
生物强化技术作为土壤重金属污染的一种原位修复手段,近年来成为环境领域的研究热点。菌种及外来营养源的选择共同决定了修复技术的成败。本文以枯草芽孢杆菌经紫外诱变所获高耐镉菌株B38作为活性菌,以玉米秸秆和猪粪2种农业废弃物为原材料制备的2种生物炭(分别记为CBC和PBC)作为微生物载体,分别研究了上述材料对镉(Cd)、铬(Cr)、汞(Hg)和铅(Pb)4种典型重金属离子的吸附能力及环境条件对吸附的影响；利用盆栽实验确认了生物强化技术的修复效果；并通过傅里叶变换-红外光谱(FT-IR)、扫描电子显微镜(SEM)、X射线光电子能谱(XPS)、DNA扩增—变性梯度凝胶电泳(PCR-DGGE)、荧光标记技术等手段,对吸附剂形貌、表面元素组成及官能团变化、外源微生物和土著微生物生长情况等进行了系统的解析,为阐明载体—微生物强化固定土壤重金属机制提供了理论依据与技术手段。
主要研究结果包括：(1)诱变菌B38对3种重金属阳离子普遍表现出高亲和力,接触初期快速吸附于细胞壁表面,之后达平衡。阴离子Cr(VI)在死体菌株上的吸附行为与阳离子类似,而在活体菌株上吸附进行得非常缓慢,可能是因为B38受到铬胁迫影响生长与活性；整体上,死体菌株的吸附能力略高于活体菌株；活体菌株的吸附动力学符合指数方程,死体菌株对阳离子和阴离子的吸附曲线分别符合假一级动力学和假二级动力学模型；pH值对吸附影响显著,pH=7.0时吸附率最高；体系的温度几乎不影响吸附；离子强度对Hg的吸附几乎没有影响,对Cd、Cr存在一定影响,特别是对Pb影响较大,说明静电吸附在Hg的吸附过程中可以忽略,而Cd的静电吸附能要高于Pb；具有相同吸附点位的离子,在单一体系下具有最高吸附量的离子(Pb)在多元体系下对其它离子存在强抑制作用,而Cd与Hg、Pb与Hg离子分别在二元体系下吸附行为互不影响,因此吸附点位不同；B38细胞壁含有羟基、羰基、羧基、酰胺基等活性基团,在吸附过程中与金属离子形成络合物。(2)为探究B38对土壤重金属污染的修复效果,选择了一种工业废弃物诺沃肥作为外来营养源。盆栽实验研究发现生物强化技术对土壤重金属起到了固定作用,降低植物积累。但仅添加B38对植物生长促进作用不明显,单独施加诺沃肥反而起到抑制作用,B38和诺沃肥协同作用显著增加植物可食部分生物量。B38在外来营养源的辅助下活性最高。(3)选择二乙基三胺五乙酸(DTPA)、复合制剂Mehlich3(M3)和BCR1(Community Bureau of Reference推荐连续提取法第1步)3种化学提取法预测重金属的生物可利用性。研究发现DTPA对叶菜类植物的生物有效性预测效果最佳,M3和BCR1分别适用于根茎类和豆科植物。(4)生物炭对重金属的吸附研究发现CBC对Hg表现出了高亲和力,PBC对Pb亲和力更高；当生物炭与B38复合后,2种复合吸附剂均对Pb吸附能力更强；同种重金属离子,B38菌株与PBC复合的吸附能力要强于与CBC复合；2种生物炭的吸附行为符合Freundlich模型,2种复合吸附剂符合Langmuir-Langmuir模型；Hg和Pb离子在生物炭上发生拮抗作用,在复合吸附剂上的相互之间略有竞争,但作用不大,说明2种离子在生物炭上的吸附点位有所重叠,但B38的引入改变了生物炭的表面性状,使2种离子拥有各自的专性吸附点位。(5)SEM研究发现2种生物炭的微观形貌存在显著差异,CBC为筛板状多孔片层结构,微孔密集分布于炭层上；PBC为表面凹陷的多孔颗粒状结构,表面向内凹陷形成塌陷的空腔,可以为重金属离子提供更多吸附点位,为微生物的复合提供更大的空间。FT-IR光谱和XPS能谱研究发现,与B38复合使CBC表面官能团数量减少、多样性降低,却可增加PBC表面的官能团数量和种类；在吸附过程中,生物炭表面的羟基、羰基和羧基等含氧官能团、芳香环上的羰基和酚羟基、CBC纤维素上的脂肪族醚类(C-O-C)和醇羟基(-OH)参与了离子交换吸附作用；生物炭表面呋哺γ-CH以及吡啶β环等杂环化合物参与了阳离子-π键作用。(6)将诱变菌B38与载体CBC或PBC应用于生物强化处理。PBC复合B38后显著促进植株生长,CBC与B38复合对作物生长的影响不大；各处理均能不同程度的降低植物对重金属的积累和土壤中重金属有效态含量,其中以PBC和B38复合效果最佳,植物可食部分Cd和Pb含量均达到国家标准。(7)利用荧光标记示踪技术监测不同载体辅助下B38的生长情况。仅添加B38,菌体浓度不断上升,缓慢趋于稳定；CBC和诺沃肥辅助处理,菌体浓度迅速达到稳定状态；PBC的处理菌体浓度先迅速上升,之后呈现下降趋势。因此,B38对恶劣的环境有较高的适应性及发展潜力,在与土著微生物的竞争中处于优势,载体物质为土著微生物和外源微生物均能提供碳源、氮源等营养物质和栖息地。
本论文的研究表明：高耐镉菌株B38对多种重金属离子均具有较高的吸附能力；生物炭作为一种新型环境材料,不仅能够吸附重金属污染物,而且能够提高土壤肥力,改善土壤生境,为微生物的生长繁殖提供营养物质和栖息场所,特别是动物源的生物炭成为微生物的优良载体。微生物与生物炭协同固定土壤重金属污染,实现了对土壤重金属的原位修复,为农业生产废弃物的就地取材、二次利用,及生物强化技术修复土壤重金属复合污染提供了一个新的途径。
Bioaugmentation has been receiving much attention recently as an in-situ technology for the remediation of heavy metal polluted soils. The microorganism species and its carrier, used as the nutrition source for microorganism, are the two crucial factors affecting the bioaugmentation efficiency. In this thesis, the immobilization of heavy metals in soil by an active species of microorganism and several carriers was studied. The bacterium used in this study was a mutant species (B38) from Bacillus subtilis obtained by UV irradiation under high cadmium (Cd) concentration. Two types of biochar made of different biomass by pyrolysis were used as the carrier of B38mutant species. One was produced by pyrolyzing corn straw, designated as CBC. The other was produced by pyrolyzing pig manure, designated as PBC. The adsorption behavior of cadmium (Cd), chromium (Cr), mercury (Hg) and lead (Pb) by B38or biochar, and various factors affecting the adsorption, such as the contact time, amount of biomass, pH, temperature and ionic strength of the solution, were investigated. To evaluate the feasibility of bioaugmentation, the pot experiment was conducted. The Fourier transform-infrared spectroscopy (FT-IR), scanning electron microscope (SEM), X-ray photoelectron spectroscopy (XPS), polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), and fluorescent labeling technique were applied to analyze the surface functional group, the morphology, the bulk organic elemental composition, and the growth of native microorganism and B38, respectively, so that the mechanisms of heavy metal immobilization using carried microorganisms could be elucidated. The main results of this thesis are as follows:
(1) B38had a higher affinity to heavy metal cations than the anion. Rapid metal binding occurred during the beginning of the adsorption of heavy metal cations by both living and nonliving B38biomass, and then the equilibrium was achieved. A similar behavior was found for heavy metal anion (Cr(Ⅵ)) adsorption by nonliving B38. But the adsorption of Cr(Ⅵ) by living B38was a slow process. This implied that the B38growth was inhibited by the high Cr(Ⅵ) concentrations. The nonliving biomass generally showed greater or similar adsorptive capacities as compared to the living biomass. The sorption kinetics followed the exponential equation for living biomass, and followed the pseudo-first-order and pseudo-second-order model for nonliving biomass for the metal cations and the metal anion, respectively. The equilibrium adsorptive quantity of the heavy metals on B38changed with solution pH. At pH7.0, both of the two sorbents presented the maximum adsorption capacities. However, the effect of the temperature of the adsorption was less significant. It seemed obvious that the adsorption of Hg was not related to ionic strength. But the adsorption quantity of Cd, Cr and Pb decreased with the increase of the ionic strength of solution. This implied that the electrostatic binding of the B38cell wall with the Hg ion was negligible compared to the covalent binding, but important for other ions. The results of binary and ternary sorption experiments indicated that the metals with the higher sorption capacity in the single-component systems showed greater inhibitory effects on the biosorption of other metal ions in the multiple-component systems, but the sorption sites of Hg and Cd or Pb are likely to be different. The FT-IR spectra showed the evidence that some of the functional groups, such as the hydroxyl, carbonyl, carboxyl, and primary amide groups, were involved in the interaction with the heavy metals ions by forming complexes on the cell wall of B38.(2) NovoGro, an industrial by-product, was used as an organic amendment to provide the nutrients necessary for B38growth. In the pot experiment, it was found that B38mutant combined with NovoGro exhibited a high efficiency for the remediation of heavy metal polluted soil. But B38employed alone could not promote the growth of plants. Meanwhile, soil amended with NovoGro alone inhibited plant growth slightly. However, the biomass yields of plants increased significantly when the soil was amended with both B38and NovoGro. Thus, the B38exhibited high activity when assisted by nutrient materials.(3) Extraction by diethylene triamine pentacetate acid (DTPA), Mehlich3(M3), and the first step of BCR (Community Bureau of Reference) sequential extraction method (BCR1) were studied to predict the bioavailability of heavy metal pollutants in soil. The results revealed that DTPA was a good extractant for the predicting of heavy metal bioavailability for leaf vegetables, and M3and BCR1showed good capacity of predicting heavy metal bioavailability for rhizome vegetables and for leguminous plants, respectively.(4) The sorption experiment by biochars showed that CBC had a high affinity to Hg ions, while PBC had a high adsorption capacity for Pb ions. Two combined sorbents between B38and the two biochars had high adsorption capacities and preferences for Pb ions. For the same metal ion, the adsorption capacity of B38combined with PBC was higher than that combined with CBC. The experimental adsorption data by the two biochars were fitted well to the Freundlich isotherms, and the results by the two combined sorbents were fitted well to Langmuir-Langmuir isotherms. Hg ions competed for the same adsorption sites with Pb ions on the biochar surface. But it seemed that the addition of B38altered the physico-chemical properties of biochar surface. The binding sites of Hg ions and Pb ions on the two combined sorbents seemed to be different.(5) The SEM photographs showed significant differences in structures between CBC and PBC. The CBC materials looked like the sieve plates with polyporous structure, while the PBC materials were polyporus structure with lacunose surface. Thus, PBC could provide more adsorption activity sites for heavy metal binding and more room for microorganism growth. The FT-IR and XPS spectra of the two biochar before and after combined with B38indicated that the addition of B38decreased the amount and diversity of the CBC surface, while increased those of the PBC surface. The oxygen-containing functional groups, such as the hydroxyl, carbonyl, carboxyl groups, and the aliphatic ethers groups (C-O-C) in cellulose of CBC were involved in the ion exchange with heavy metal ions. The cation-π interactions involved binding between heavy metal ions and the y-CH group of furan or the β-ring of heterocyclic compounds on the surface of biochar.(6) CBC and PBC were tested as the B38mutant carrier in the remediation of heavy metal contaminated soils by bioaugmentation. Soil amended with B38and PBC increased the plant biomass yield, but soil amended with B38and CBC had no significant effect on the growth of plants. All the four amendments (two biochars and two combined materials of B38and the two biochars) could inhibit the bioaccumulation of heavy metal pollutants by plants, and reduce the bioavailability of heavy metal pollutants in soil. Soil amended with PBC and B38exhibited the maximum remediation efficiency among the four treatments, and the Cd and Pb concentrations in the edible part of plants were below the maximum levels of contaminants in foods.(7) The thesis for the first time used the fluorescent labeling technique for the investigation of the growth of a bioaugmented species in soil. In the soil amended with B38alone, the concentration of B38mutant increased slowly and sustainingly, then reached a steady state. In the soil amended with B38and CBC or NovoGro, the concentration of B38increased sharply, and then achieved the equilibrium state. However, in the soil amended with B38and PBC, the concentration of B38mutant increased first, then decreased. Thus, the B38mutant showed great resilience to the stressful environmental conditions, had great potential for the development, and exhibited a competitive superiority when competed with the native microorganisms. All the three carrier materials could provide nutrients for the growth of both native microorganisms and B38mutant.
The present study found that B38, as a high Cd-tolerant mutant strain, had high adsorption capacities to multiple heavy metals. Biochar could not only immobilize the heavy metal contaminants, but also improve and fertilize the soil, and provide necessary nutrient for the microorganism growth. Biochar, especially the one produced from animal wastes, showed to be a good carrier for microorganism. The synergism of microorganisms with biochar is an effective bioaugmentation techonology for the in-situ remediation of heavy metal contamination in soil. Meanwhile, it provided a new concept of the solid waste reuse in agriculture.
主要研究结果包括：(1)诱变菌B38对3种重金属阳离子普遍表现出高亲和力,接触初期快速吸附于细胞壁表面,之后达平衡。阴离子Cr(VI)在死体菌株上的吸附行为与阳离子类似,而在活体菌株上吸附进行得非常缓慢,可能是因为B38受到铬胁迫影响生长与活性；整体上,死体菌株的吸附能力略高于活体菌株；活体菌株的吸附动力学符合指数方程,死体菌株对阳离子和阴离子的吸附曲线分别符合假一级动力学和假二级动力学模型；pH值对吸附影响显著,pH=7.0时吸附率最高；体系的温度几乎不影响吸附；离子强度对Hg的吸附几乎没有影响,对Cd、Cr存在一定影响,特别是对Pb影响较大,说明静电吸附在Hg的吸附过程中可以忽略,而Cd的静电吸附能要高于Pb；具有相同吸附点位的离子,在单一体系下具有最高吸附量的离子(Pb)在多元体系下对其它离子存在强抑制作用,而Cd与Hg、Pb与Hg离子分别在二元体系下吸附行为互不影响,因此吸附点位不同；B38细胞壁含有羟基、羰基、羧基、酰胺基等活性基团,在吸附过程中与金属离子形成络合物。(2)为探究B38对土壤重金属污染的修复效果,选择了一种工业废弃物诺沃肥作为外来营养源。盆栽实验研究发现生物强化技术对土壤重金属起到了固定作用,降低植物积累。但仅添加B38对植物生长促进作用不明显,单独施加诺沃肥反而起到抑制作用,B38和诺沃肥协同作用显著增加植物可食部分生物量。B38在外来营养源的辅助下活性最高。(3)选择二乙基三胺五乙酸(DTPA)、复合制剂Mehlich3(M3)和BCR1(Community Bureau of Reference推荐连续提取法第1步)3种化学提取法预测重金属的生物可利用性。研究发现DTPA对叶菜类植物的生物有效性预测效果最佳,M3和BCR1分别适用于根茎类和豆科植物。(4)生物炭对重金属的吸附研究发现CBC对Hg表现出了高亲和力,PBC对Pb亲和力更高；当生物炭与B38复合后,2种复合吸附剂均对Pb吸附能力更强；同种重金属离子,B38菌株与PBC复合的吸附能力要强于与CBC复合；2种生物炭的吸附行为符合Freundlich模型,2种复合吸附剂符合Langmuir-Langmuir模型；Hg和Pb离子在生物炭上发生拮抗作用,在复合吸附剂上的相互之间略有竞争,但作用不大,说明2种离子在生物炭上的吸附点位有所重叠,但B38的引入改变了生物炭的表面性状,使2种离子拥有各自的专性吸附点位。(5)SEM研究发现2种生物炭的微观形貌存在显著差异,CBC为筛板状多孔片层结构,微孔密集分布于炭层上；PBC为表面凹陷的多孔颗粒状结构,表面向内凹陷形成塌陷的空腔,可以为重金属离子提供更多吸附点位,为微生物的复合提供更大的空间。FT-IR光谱和XPS能谱研究发现,与B38复合使CBC表面官能团数量减少、多样性降低,却可增加PBC表面的官能团数量和种类；在吸附过程中,生物炭表面的羟基、羰基和羧基等含氧官能团、芳香环上的羰基和酚羟基、CBC纤维素上的脂肪族醚类(C-O-C)和醇羟基(-OH)参与了离子交换吸附作用；生物炭表面呋哺γ-CH以及吡啶β环等杂环化合物参与了阳离子-π键作用。(6)将诱变菌B38与载体CBC或PBC应用于生物强化处理。PBC复合B38后显著促进植株生长,CBC与B38复合对作物生长的影响不大；各处理均能不同程度的降低植物对重金属的积累和土壤中重金属有效态含量,其中以PBC和B38复合效果最佳,植物可食部分Cd和Pb含量均达到国家标准。(7)利用荧光标记示踪技术监测不同载体辅助下B38的生长情况。仅添加B38,菌体浓度不断上升,缓慢趋于稳定；CBC和诺沃肥辅助处理,菌体浓度迅速达到稳定状态；PBC的处理菌体浓度先迅速上升,之后呈现下降趋势。因此,B38对恶劣的环境有较高的适应性及发展潜力,在与土著微生物的竞争中处于优势,载体物质为土著微生物和外源微生物均能提供碳源、氮源等营养物质和栖息地。
本论文的研究表明：高耐镉菌株B38对多种重金属离子均具有较高的吸附能力；生物炭作为一种新型环境材料,不仅能够吸附重金属污染物,而且能够提高土壤肥力,改善土壤生境,为微生物的生长繁殖提供营养物质和栖息场所,特别是动物源的生物炭成为微生物的优良载体。微生物与生物炭协同固定土壤重金属污染,实现了对土壤重金属的原位修复,为农业生产废弃物的就地取材、二次利用,及生物强化技术修复土壤重金属复合污染提供了一个新的途径。
Bioaugmentation has been receiving much attention recently as an in-situ technology for the remediation of heavy metal polluted soils. The microorganism species and its carrier, used as the nutrition source for microorganism, are the two crucial factors affecting the bioaugmentation efficiency. In this thesis, the immobilization of heavy metals in soil by an active species of microorganism and several carriers was studied. The bacterium used in this study was a mutant species (B38) from Bacillus subtilis obtained by UV irradiation under high cadmium (Cd) concentration. Two types of biochar made of different biomass by pyrolysis were used as the carrier of B38mutant species. One was produced by pyrolyzing corn straw, designated as CBC. The other was produced by pyrolyzing pig manure, designated as PBC. The adsorption behavior of cadmium (Cd), chromium (Cr), mercury (Hg) and lead (Pb) by B38or biochar, and various factors affecting the adsorption, such as the contact time, amount of biomass, pH, temperature and ionic strength of the solution, were investigated. To evaluate the feasibility of bioaugmentation, the pot experiment was conducted. The Fourier transform-infrared spectroscopy (FT-IR), scanning electron microscope (SEM), X-ray photoelectron spectroscopy (XPS), polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), and fluorescent labeling technique were applied to analyze the surface functional group, the morphology, the bulk organic elemental composition, and the growth of native microorganism and B38, respectively, so that the mechanisms of heavy metal immobilization using carried microorganisms could be elucidated. The main results of this thesis are as follows:
(1) B38had a higher affinity to heavy metal cations than the anion. Rapid metal binding occurred during the beginning of the adsorption of heavy metal cations by both living and nonliving B38biomass, and then the equilibrium was achieved. A similar behavior was found for heavy metal anion (Cr(Ⅵ)) adsorption by nonliving B38. But the adsorption of Cr(Ⅵ) by living B38was a slow process. This implied that the B38growth was inhibited by the high Cr(Ⅵ) concentrations. The nonliving biomass generally showed greater or similar adsorptive capacities as compared to the living biomass. The sorption kinetics followed the exponential equation for living biomass, and followed the pseudo-first-order and pseudo-second-order model for nonliving biomass for the metal cations and the metal anion, respectively. The equilibrium adsorptive quantity of the heavy metals on B38changed with solution pH. At pH7.0, both of the two sorbents presented the maximum adsorption capacities. However, the effect of the temperature of the adsorption was less significant. It seemed obvious that the adsorption of Hg was not related to ionic strength. But the adsorption quantity of Cd, Cr and Pb decreased with the increase of the ionic strength of solution. This implied that the electrostatic binding of the B38cell wall with the Hg ion was negligible compared to the covalent binding, but important for other ions. The results of binary and ternary sorption experiments indicated that the metals with the higher sorption capacity in the single-component systems showed greater inhibitory effects on the biosorption of other metal ions in the multiple-component systems, but the sorption sites of Hg and Cd or Pb are likely to be different. The FT-IR spectra showed the evidence that some of the functional groups, such as the hydroxyl, carbonyl, carboxyl, and primary amide groups, were involved in the interaction with the heavy metals ions by forming complexes on the cell wall of B38.(2) NovoGro, an industrial by-product, was used as an organic amendment to provide the nutrients necessary for B38growth. In the pot experiment, it was found that B38mutant combined with NovoGro exhibited a high efficiency for the remediation of heavy metal polluted soil. But B38employed alone could not promote the growth of plants. Meanwhile, soil amended with NovoGro alone inhibited plant growth slightly. However, the biomass yields of plants increased significantly when the soil was amended with both B38and NovoGro. Thus, the B38exhibited high activity when assisted by nutrient materials.(3) Extraction by diethylene triamine pentacetate acid (DTPA), Mehlich3(M3), and the first step of BCR (Community Bureau of Reference) sequential extraction method (BCR1) were studied to predict the bioavailability of heavy metal pollutants in soil. The results revealed that DTPA was a good extractant for the predicting of heavy metal bioavailability for leaf vegetables, and M3and BCR1showed good capacity of predicting heavy metal bioavailability for rhizome vegetables and for leguminous plants, respectively.(4) The sorption experiment by biochars showed that CBC had a high affinity to Hg ions, while PBC had a high adsorption capacity for Pb ions. Two combined sorbents between B38and the two biochars had high adsorption capacities and preferences for Pb ions. For the same metal ion, the adsorption capacity of B38combined with PBC was higher than that combined with CBC. The experimental adsorption data by the two biochars were fitted well to the Freundlich isotherms, and the results by the two combined sorbents were fitted well to Langmuir-Langmuir isotherms. Hg ions competed for the same adsorption sites with Pb ions on the biochar surface. But it seemed that the addition of B38altered the physico-chemical properties of biochar surface. The binding sites of Hg ions and Pb ions on the two combined sorbents seemed to be different.(5) The SEM photographs showed significant differences in structures between CBC and PBC. The CBC materials looked like the sieve plates with polyporous structure, while the PBC materials were polyporus structure with lacunose surface. Thus, PBC could provide more adsorption activity sites for heavy metal binding and more room for microorganism growth. The FT-IR and XPS spectra of the two biochar before and after combined with B38indicated that the addition of B38decreased the amount and diversity of the CBC surface, while increased those of the PBC surface. The oxygen-containing functional groups, such as the hydroxyl, carbonyl, carboxyl groups, and the aliphatic ethers groups (C-O-C) in cellulose of CBC were involved in the ion exchange with heavy metal ions. The cation-π interactions involved binding between heavy metal ions and the y-CH group of furan or the β-ring of heterocyclic compounds on the surface of biochar.(6) CBC and PBC were tested as the B38mutant carrier in the remediation of heavy metal contaminated soils by bioaugmentation. Soil amended with B38and PBC increased the plant biomass yield, but soil amended with B38and CBC had no significant effect on the growth of plants. All the four amendments (two biochars and two combined materials of B38and the two biochars) could inhibit the bioaccumulation of heavy metal pollutants by plants, and reduce the bioavailability of heavy metal pollutants in soil. Soil amended with PBC and B38exhibited the maximum remediation efficiency among the four treatments, and the Cd and Pb concentrations in the edible part of plants were below the maximum levels of contaminants in foods.(7) The thesis for the first time used the fluorescent labeling technique for the investigation of the growth of a bioaugmented species in soil. In the soil amended with B38alone, the concentration of B38mutant increased slowly and sustainingly, then reached a steady state. In the soil amended with B38and CBC or NovoGro, the concentration of B38increased sharply, and then achieved the equilibrium state. However, in the soil amended with B38and PBC, the concentration of B38mutant increased first, then decreased. Thus, the B38mutant showed great resilience to the stressful environmental conditions, had great potential for the development, and exhibited a competitive superiority when competed with the native microorganisms. All the three carrier materials could provide nutrients for the growth of both native microorganisms and B38mutant.
The present study found that B38, as a high Cd-tolerant mutant strain, had high adsorption capacities to multiple heavy metals. Biochar could not only immobilize the heavy metal contaminants, but also improve and fertilize the soil, and provide necessary nutrient for the microorganism growth. Biochar, especially the one produced from animal wastes, showed to be a good carrier for microorganism. The synergism of microorganisms with biochar is an effective bioaugmentation techonology for the in-situ remediation of heavy metal contamination in soil. Meanwhile, it provided a new concept of the solid waste reuse in agriculture.
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