植物-微生物联合修复石油污染土壤的实验研究

筛选高效石油降解菌并考察菌株的石油降解能力,通过植物-微生物联合修复石油污染土壤室内实验,在修复过程中测定了土壤中细菌和固氮菌,碱解氮、速效磷和速效钾的含量变化,同时采用傅立叶变换离子回旋共振质谱(ESI FT-ICR MS)考察了植物-微生物联合修复效果。结果表明,菌株3#、4#的生长适应性较强,其混合菌的降解效果最好,将其混合菌液与植物进行植物-微生物联合修复不同浓度的石油污染土壤,经过150 d的温室降解,最高降解率达到73.47%。ESI FT-ICR MS分析结果表明,与空白组相比,植物组的O1、O2和N1类等化合物相对丰度都发生了显著变化,石油污染物得到一定程度的生物降解。     

鼠李糖脂与菌剂对原油污染土壤的联合修复

为研究RL强化MI修复原油污染土壤的效果,通过盆栽实验,测定了修复过程中微生物、脱氢酶、原油降解率和正构烷烃等变化。结果表明,提取RL具有良好表面和乳化活性。修复过程中微生物数目、脱氢酶和石油降解率具有相关性。RL+MI联合修复对原油的降解效果最显著,第14、21和28天的降解率分别达67.6%、78.6%和81.3%,较MI和RL处理分别提高了22%~24%和32%~38%;细菌、放线菌和霉菌最高生物量分别比CK提高约20、5.8和4.7倍;GC-MS示修复14 d后∑C21-/∑C22+(1.334)和Pr/Ph(1.152)均大于CK,说明降解早期具有高碳正构烷烃、奇数烷烃的降解优势以及对类异戊二烯烷烃的降解。     

固定化耐低温混合菌对焦化土壤多环芳烃的降解研究

研究了固定化耐低温真菌-细菌混合菌在低温环境下,对焦化厂污染土壤中的菲(Phe)和苯并[b]荧蒽(BbF)降解的动态变化,利用高通量测序技术分析了降解过程中微生物群落多样性变化。结果表明:在低温条件下固定化混合菌对土壤中Phe、BbF的去除率远高于游离混合菌与固定化单菌,在60d的降解周期下,固定化混合菌对土壤中Phe和BbF的降解率分别可达59.61%和45.24%。处理前,土壤中细菌与真菌初始Shannon多样性指数分别为2.79和0.33,细菌远高于真菌,土壤中土著微生物以细菌为主,高丰度的细菌抑制了真菌的生长代谢。处理后,细菌的Shannon多样性指数下降至1.33,真菌的Shannon多样性指数增加至1.01,Phe和BbF的降解与细菌多样性呈负相关,且细菌多样性的降低减少了其对真菌的抑制作用。对比分析了处理前后土壤中微生物群落组成的变化,结果表明:投加固定化混合菌后,固定化混合菌中的假单胞菌(Pseudomonas sp.)SDR4和高山被孢霉(Mortierella alpina)JDR7在低温下生长代谢良好,并成为降解过程中的优势菌,其物种相对丰度分别提高至79.84%与58.63%。固定化混合菌对低温环境有良好的耐性,固定化混合菌的投加提高了菌株对多环芳烃(PAHs)的生物利用有效性,改变了土壤中微生物群落的结构和丰度,可应用于低温土壤PAHs的原位修复。     

黑土微生物群落对副球菌属(Paracoccus sp.)QD15-1修复邻苯二甲酸二甲酯污染土壤的响应

采用Illumina MiSeq高通量测序法探究黑土微生物群落对副球菌属(Paracoccus sp.)QD15-1修复邻苯二甲酸二甲酯(DMP)污染土壤的响应。结果表明,副球菌属QD15-1在7d内可快速有效降解土壤中的DMP,平均降解率为94.7%,并能保持稳定;Shannon、Sobs指数显示,土壤中微生物多样性、丰富性受到DMP的影响;土壤中的优势门为放线菌门(Actinobacteria),占总菌门数的50%以上;鞘氨醇单胞菌属(Sphingomonas)是第一优势菌属,第二优势菌属为类诺卡氏菌属(Nocardioides)。     

植物混种原位修复多环芳烃污染农田土壤

通过比较实验前后土壤微生物主要类群数量、PAHs降解菌数量、土壤PAHs含量和植物不同部位PAHs含量变化,评价植物单种和混种野外原位修复多环芳烃(PAHs)污染农田土壤的效果。结果显示,150 d天生长期内,黑麦草/小麦混种及黑麦草/蚕豆混种修复效果最好,对土壤PAHs总量的降解率分别达到了59.4%和64.8%。同时,这2个混种处理土壤细菌、真菌和PAHs降解菌数量分别显著高于相应的小麦、蚕豆和黑麦草单种处理。植物不同部位PAHs含量高低次序为根部>茎叶≈籽粒。混种模式下,蚕豆和小麦不同部位PAHs含量比单种模式的不同程度降低,特别是籽粒部。植物混种模式不仅显著提高了土壤PAHs的降解率,还降低了农作物体内PAHs含量,实现了边生产边修复,在污染农田土壤修复领域有着广阔的应用前景。     

不同土地利用类型土壤微生物群落特征及其影响因素

土壤微生物群落结构特征能够提前反映土壤环境质量的变化,是土壤质量评价的重要因子。为弄清不同土地利用类型土壤微生物群落特征及其影响因素,以内蒙古呼和浩特赛罕区为研究区,从绿地、园林地、耕地以及荒废地4种不同土地利用类型土壤进行采样研究。研究结果表明:4种不同土地利用类型土壤中主要菌种为细菌、真菌以及放线菌,放线菌比例最大,其次是细菌,真菌比例最小。采样深度、温度、土地利用类型及含水率均影响微生物群落特征。随着采样深度的增加,4种土地利用类型土壤中微生物数量不断减少,0～10cm微生物总数最多。荒废地土壤微生物数量最多,因为荒废地土层中有机质含量高,为微生物的生长提供了营养物质。含水率过高或过低均不利于微生物生长,当处于最佳含水率时微生物数量最大。对于绿地和园林地而言,最佳含水率为10%～15%,耕地最佳含水率为15%～20%,荒废地最佳含水率为10%左右。     

采煤沉陷区耕地土壤微生物数量及酶活性的空间特征

通过野外调查和采样分析,研究了焦作矿区韩王矿沉陷区不同沉陷部位和不同深度耕地的土壤微生物数量及酶活性特征。研究结果表明:与对照区比较,沉陷区耕地土壤微生物数量、酶活性赋存特征及微生物类群组成比例发生了明显变化。沉陷区表层(0~5 cm)、上层(5~10 cm)、中层(10~20 cm)土壤微生物总数、细菌数量均明显减少,而下层(20~40 cm)土壤微生物总数、细菌、真菌、放线菌数量均明显升高。沉陷区表层、上层土壤细菌数量所占微生物总数比例分别降低了20.64%和13.17%,而放线菌数量所占比例分别升高了20.69%和12.66%。沉陷区表层土壤脲酶、蔗糖酶、磷酸酶、脱氢酶、过氧化氢酶和多酚氧化酶活性分别降低了1.5%、17.5%、22.0%、35.3%、20.4%和5.4%。不同沉陷部位的土壤微生物数量及部分酶活性指标空间异质性显著(p0.05)。沉陷区土壤真菌、放线菌数量、蔗糖酶、磷酸酶、脱氢酶、过氧化氢酶、多酚氧化酶活性较对照区均具有不同的垂向分布特征。表明采煤引发的地表沉陷使耕地土壤微生物数量及酶活性在水平和垂直方向上均发生了显著变化,而这些变化是导致沉陷区耕地退化、生产力降低的重要原因。     

多环芳烃降解菌的筛选及其在焦化场地污染土壤修复中的应用

经过富集、分离和纯化,从沈阳某焦化厂多环芳烃(PAHs)污染土壤中获得7株菌株B1~B7。通过初步降解实验和血平板实验,发现B4、B5、B7在15d时对PAHs总降解率均高于40%,为高效PAHs降解菌,B2为高效表面活性剂产生菌。将B4、B5、B7分别与B2等质量混合后对PAHs进行降解,发现添加B2可提高PAHs总降解率,B4+B2对PAHs的总降解率最大,在9d时平均值达到45.9%。经形态观察和16SrRNA基因序列比对,鉴定B2和B4分别归为假单胞菌属(Pseudomonas sp.)和芽孢杆菌属(Bacillus sp.)。接种B4+B2进行微生物修复实验,结果表明,接种B4+B2对PAHs污染土壤的微生物修复有明显的强化作用,在60d时PAHs总降解率达到48.1%;接种B4+B2对中环(4、5环)PAHs降解率的提高尤为明显,7种中环PAHs的平均降解率比不接种菌株的对照组提高29.6百分点。     

生物强化修复石油污染土壤

筛选高效石油降解菌,考察菌株的降解性能及降解机理,进行花盆模拟高效外源菌强化修复石油污染土壤实验,在降解后期添加激活剂H2O2以及木屑来试图改善微生物的修复环境,减缓微生物的衰亡,并考察修复效果。结果表明,菌株L-1的降解效果较好,其对pH和温度有较大范围的适应性,能分泌较多的表面活性物质,细胞疏水性较强。将其应用于土壤修复中,经过50 d的修复,石油残留率达到50.6%左右,生物强化比自然修复残留率降低了8%左右。在第45天添加激活剂能有效改善修复效果,70 d时添加外源菌的土样最小石油残留率达到37.9%。     

含油污泥中石油降解菌的分离及其降解特性

从渤海油田含油污泥中分离出3株石油烃降解菌,通过16S rRNA基因序列鉴定RS1、RS2和RS3分别为棒状杆菌、短杆菌和假单胞菌。经单因素实验确定,3株细菌对石油的最适降解条件分别为37℃、盐度3%、pH 8;32℃、盐度1%、pH 8;42℃、盐度1%、pH 6。降解实验结果表明,3株细菌30 d内对含油污泥中总石油烃的降解率分别为39.69%、31.13%和53.29%,而混合菌的降解效果明显高于单一菌株,降解率为58.08%;不同菌株对原油中不同组分的降解能力不同,其中,RS1对饱和烃的降解率最高,达20.74%,RS3对芳香烃的降解率最高,达到8.08%;GC-MS分析表明,混合菌对nC12~n-C34等正构烷烃均有明显降解,且对萘、苊、屈和苯并[b]荧蒽等多环芳烃的降解能力较强。     

固定化降解菌Q5去除喹啉的性能研究

为了研究固定化微生物在土壤生物修复中的应用,以实验室筛选出来的高效降解菌 Q5 为生物活性物质,利用生物大分子仿生合成出的纳米多孔氧化硅为载体,通过表面吸附同定化方法将其固定,制备出固定化微生物.考察固定化微牛物初始 pH 值、温度、摇床转速和菌种的接种量对喹啉去除的影响,得到适宜的去除条件,在相同条件下比较固定化微生物与游离菌种对底物的去除情况,研究单一固定化菌种对不同浓度的喹啉的去除情况,考察固定化微生物的稳定性.实验结果表明,菌株 Q5 经固定化后,对喹啉的去除能力大大增强,在 500 mg/L 浓度下,40 h 固定化 Q5 对底物去除率达96.6%,远高于未固定化 Q5 的去除率 56.1%;对于高底物浓度,固定化微生物的去除效果明显,初始底物浓度为1 500 mg/L,反应 70 h 后去除率为 91.6%,且这种固定化微生物的重复使用性能良好.     

固定化微生物修复石油污染土壤影响因素研究

针对石油污染土壤修复,利用实验室已筛选的高效石油降解单菌SM-3,以天然有机材料为载体,吸附法制备固定化微生物。将游离与固定化微生物应用于室内花盆模拟修复石油污染土壤,对C/N/P、微生物投加量、石油含量、氧化剂和表面活性剂设计5因素4水平正交实验,探讨不同修复时期各影响因素的重要性顺序,最佳条件下各菌株的修复效果。结果表明,不同微生物在不同降解时期,各影响因素的重要性会发生变化;经过21 d的修复,固定化单菌SM-3石油降解率为22.77%,修复过程中,接种量是最重要的影响因素,营养元素N、P投加影响较大,表面活性剂和氧化剂影响次之。     

Degradation of pentachlorophenol by pure and mixed cultures in two different soils

GOAL, SCOPE AND BACKGROUND: Pentachlorophenol (PCP) is the second highest volume pesticide used in the United States. It is a mutagenic compound whose exposure poses significant health effects, One of the most desirable, environmentally friendly treatment methods is bioremediation. For soil-based contamination, the effectiveness of bioremediation will also be affected by the presence of an active indigenous population, sorption of the contaminant onto the soil, and environmental parameters. METHODS: Two pure strains and their mixed culture were used to evaluate PCP biodegradation in two different field soils, Columbia (CO) and New Mexico (NM). Biostimulation of the indigenous microbes was evaluated by adding nutrients. The efficiency of adding bacteria strains (bioaugmentation) for degrading PCP was determined with Arthrobacter sp., Flavobacterium sp. and a 50:50 mixture of the two bacteria strains. RESULTS: In CO soil, only 24%, 12% and 25% of the initial PCP concentration were degraded by Flavobacterium sp., Arthrobacter sp. and mixed culture, respectively. Arthrobacter sp. was used in NM soil with two initial concentrations and achieved degradation efficiencies of 57% and 61% for 361 and 95 mg kg- concentrations, respectively. Discussion. Analysis via statistical methods showed that the bacteria had different efficiencies on PCP degradation in each soil. 2 CONCLUSIONS: All bacteria catalyzed a higher PCP degradation when present in NM soil. Second, Flavobacterium sp. degraded more PCP than Arthrobacter sp. in CO soil. The mixed culture achieved the highest degradation efficiency regardless of the initial concentration or soil origin. RECOMMENDATIONS AND PERSPECTIVES: The effect of the soil properties, such as the soil organic matter (SOM) on PCP biodegradation should be investigated. Future work can also investigate the effect of aging time on biodegradation.     

Degradation of di-butyl-phthalate by soil bacteria

Twelve Gram-positive phthalate ester degraders were isolated from soil. Using Biolog GP2 plates, eight of them were identified as belonging to the Corynebacterium-Mycobacterium-Nocardia group, while the remaining four were unidentifiable. When cultured in the presence of di-butyl-phthalate (DBP) in basal salts solution, five of these isolates accomplished more than 90% of DBP degradation within 48 h (fast group), three were placed in the medium group, and the remaining four were placed in the slow group which caused less than 30% of DBP degradation within the same period of time. A 420 bp DNA fragment was amplified from six isolates and none of them fell within the slow group. When compared with the large subunit of phthalate dioxygenase gene (phtA) of Arthrobacter keyseri, 83% and 91% similarities were evident in the nucleotide and amino acid sequences, respectively. However, no correlation between cell surface hydrophobicity and phthalate degradation ability was evident. Six surfactants (Brij 30, Brij 35, Tergitoltype NP-10, Triton N-101, Triton X-100 and SDS) were tested for their abilities to increase degradation rate. When added at the critical micellar concentration (CMC), they all displayed strong growth inhibition against the three bacteria tested, with Brij 30 been the least toxic to isolates G2 and G11, and Brij 35 had the least inhibitory effect for G1. When half the CMC of Brij 30 was incorporated into the basal salts, the inhibitory effect on DBP degradation remained. Soil helped to minimize surfactant toxicity of surfactant and increase the degradation potential of some of the test bacteria. When DBP-amended soil had been aged for three months, decreases in bioavailability were observed but the effect varied tremendously between different organisms. For isolates G1, G2, G5, G7 and G17 the aging effects were almost non-exist. The present study indicates that selection of a suitable degrader may minimize the undesired effect of aging on bioremediation process.     

Biodegradation of polystyrene-graft-starch copolymers in three different types of soil

Materials based on polystyrene and starch copolymers are used in food packaging, water pollution treatment, and textile industry, and their biodegradability is a desired characteristic. In order to examine the degradation patterns of modified, biodegradable derivates of polystyrene, which may keep its excellent technical features but be more environmentally friendly at the same time, polystyrene-graft-starch biomaterials obtained by emulsion polymerization in the presence of new type of initiator/activator pair (potassium persulfate/different amines) were subjected to 6-month biodegradation by burial method in three different types of commercially available soils: soil rich in humus and soil for cactus and orchid growing. Biodegradation was monitored by mass decrease, and the highest degradation rate was achieved in soil for cactus growing (81.30 %). Statistical analysis proved that microorganisms in different soil samples have different ability of biodegradation, and there is a significant negative correlation between the share of polystyrene in copolymer and degree of biodegradation. Grafting of polystyrene on starch on one hand prevents complete degradation of starch that is present (with maximal percentage of degraded starch ranging from 55 to 93 %), while on the other hand there is an upper limit of share of polystyrene in the copolymer (ranging from 37 to 77 %) that is preventing biodegradation of degradable part of copolymers.     

Experimental study on effect of anion surfactant on degradation rate of aldicarb in soil

Degradation kinetics of aldicarb [2-methyl-2-(methylthio) propionaldehyde O-(methyl carbamoyl) oxime] in surface and subsurface soil containing different levels of sodium dodecylbenzenesulfonate (SDBS) were determined to understand complex effect of SDBS on aldicarb degradation process. The results showed that degradation curves of aldicarb in soil can be described with first order kinetics formula and the degradation rate constant. k (d(-1)), in surface soil was larger than that in subsurface soil. SDBS can accelerate the degradation of aldicarb in soil and there was a good linear relationship between degradation rate constant and the logarithm of SDBS concentration. The possible reasons were that SDBS could change pH value of soil, have solubilization effect on aldicarb, and be used as carbon source of microorganisms. But SDBS had a larger promotion to the degradation of aldicarb in surface than in subsurface soil. When SDBS concentration was 1000 mg/kg of dried soil the first order degradation rate constant of aldicarb could be increased by 56.6 percent in surface soil and by 27.6 percent in subsurface soil, respectively.     

Phytoremediation of petroleum hydrocarbons in tropical coastal soils II. microbial response to plant roots and contaminant

GOAL, SCOPE AND BACKGROUND: The goal of this study was to understand the interaction between plants and microorganisms during petroleum-hydrocarbon bioremediation in Pacific Islands coastal soils. Total bacteria and hydrocarbon-degrading microorganisms population dyanamics were examined in the rhizospheres of tropical trees and shrubs, which were evaluated for their phytoremediation potential in a greenhouse experiment. The respective and combined effects of plant roots and diesel contaminant on the microbial populations were determined in relation to diesel fuel depletion. An increase in the grading populations size of the hydrocarbon-degrading populations of microbes, elicited by rhizodeposition, is generally regarded as conducive to an enhanced degradation of petroleum hydrocarbon pollutants in vegetated soil. METHODS: The soil was a coastal sandy loam (pH 7.8) which was artificially contaminated with 10 g of No. 2 diesel fuel/kg soil or left uncontaminated. The pots were irrigated with fertilizer and 1% NaCl. The enumerations were carried out in the contaminated and uncontaminated rhizospheres of three trees, kiawe (Prosopis pallida), milo (Thespesia populnea), and kou (Cordia subcordata) and three shrubs, beach naupaka (Scaevola sericea), false sandalwood (Myoporum sandwicense), and oleander (Nerium oleander). Unplanted control soils were included in the experiment. Total bacteria and phenanthrene-degrading bacteria were enumerated on plates. Diesel- and pristane-degrading microorganisms were enumerated by the most-probable-number technique in tissue-culture plates. RESULTS AND DISCUSSION: All four types of microorganisms responded to the rhizosphere of the 6 plants in uncontaminated soil and to the diesel contaminant in unplanted soil. In contaminated rhizospheres, no effect of the plant on the hydrocarbon-degrader numbers was visible. Total bacteria responded more to the plant roots than to the contaminant. The phenanthrene-degrading bacteria and pristane-degrading microorganisms were more influenced by the contaminant than by the plants. The diesel-degrading microorganisms were equally stimulated by the plants and the contaminant. The numbers of hydrocarbon degraders were similar in the contaminated rhizospheres of the three effective plants (kiawe, kou, and milo) and in those of the three ineffective shrubs. CONCLUSION: The results suggest the quality of the rhizodeposition is plant-dependent and governs the type of diesel-degrader populations that will be enhanced by a given plant. RECOMMENDATIONS AND OUTLOOK: In the proposed phytoremediation-benefit model plant roots maintain high levels of hydrocaron degraders in uncontaminated soil. When the root enters a contaminated zone of soil, those hydrocarbon degraders that prefer the contaminant would switch to the contaminant as a carbon source, effectively removing the hydrocarbons. If the root exudates and the contaminant are equally attractive to the hydrocarbon degraders, the contaminant degradaton would be less effective.     

Anaerobic biodegradation of biphenyl in various paddy soils and river sediment

The anaerobic degradation of biphenyl was investigated in four uncontaminated Japanese paddy soils and one river sediment sample contaminated with benzene and chlorinated aliphatics. Two of the paddy soils and the sediment were capable of degrading biphenyl anaerobically without any additional medium or electron acceptors. The half-lives of biphenyl biodegradation in the three samples were 212 d in the Kuridashi soil, 327 d in the Kamajima soil, and 429 d in the river sediment. The Kuridashi soil metabolized 1+/-0.3% of [U-14C]-biphenyl into CO2 and 5+/-2% into water-soluble metabolites after 45 d of incubation. Submerged conditions, which result in lower nitrate and iron oxide contents, and neutral pH, appeared to be the common properties among the samples that influenced their degradation capacities. The addition of 10mM sulfate and 20mM Fe(III) as electron acceptors did not enhance the biphenyl degradation rate, whereas 10mM nitrate completely inhibited biphenyl degradation. The addition of different electron donors (lactate, acetate, or pyruvate) slightly slowed the degradation. Molybdate (an inhibitor of sulfate-reducing bacteria) had an inhibitory effect on biphenyl biodegradation, but bromoethanesulfonic acid (an inhibitor of methanogens) did not. Most biphenyl degradation was observed when only water was added, with no other electron acceptors or donors. These results suggest that sulfate-reducing bacteria and fermentative microbial populations play important roles in anaerobic biphenyl biodegradation in paddy soil.     

强化土著微生物处理硝基苯和苯胺污染地下水

以硝基苯、苯胺为主要污染物的污染地下水为研究对象,加入激活剂(乳糖、Na2HPO4、乳糖+Na2HPO4、乙醇、牛肉膏、蛋白胨)激活土著微生物,并考察其对土著微生物生长及硝基苯、苯胺降解效果的影响。加入激活剂3d后测各个水样的脱氢酶活性,对培养9d后的水样进行气相色谱/质谱(GC/MS)分析。结果表明,加入乳糖的水样中,其微生物相对增长率达157.2%,硝基苯、苯胺的相对去除率分别为14.90%和0.79%;加入Na2HPO4和乙醇的水样中,其微生物增长和硝基苯、苯胺降解情况均没有明显变化;加入乳糖+Na2HPO4的水样中,微生物相对增长率达180.3%,硝基苯、苯胺的相对去除率分别为24.20%和1.21%;加入牛肉膏的水样中,微生物的相对增长率为830.7%,硝基苯、苯胺的相对去除率分别为99.99%和99.67%;加入蛋白胨的水样中,其微生物相对增长率为686.0%,硝基苯、苯胺的相对去除率分别为99.33%和58.94%。GC/MS分析结果表明,加入激活剂后对氯苯胺、1-甲基-4-硝基苯等其他有机物的降解率均有提高。由此可见,通过激活土著微生物修复有机物污染地下水是可行的。