菲降解菌株GY2B的分离鉴定及其降解特性

对一株菲降解菌进行了鉴定并对其降解特性进行了研究.初步鉴定菌株GY2B为鞘氨醇单胞菌(Sphingomonassp.).菌株GY2B有较高的降解效率,当无机盐培养液中菲初始浓度为100mg/L时,48h内对菲降解率达到99.1%.添加100mg/L的葡萄糖和蛋白胨均可以促进菌株的生长和菲的降解,pH值为中性条件时对细胞的生长较为有利.GY2B菌株还能降解1-羟基-2-萘酸、2-萘酚、萘、水杨酸、邻苯二酚和苯酚等多种芳香化合物并利用其为碳源和能源生长繁殖.GY2B菌株对菲的降解可能通过水杨酸途径.     

Isolation of marine benzo[a]pyrene-degrading Ochrobactrum sp. BAP5 and proteins characterization

A bacterial strain BAP5 with a relatively high degradation ability of benzo[a]pyrene (BaP) was isolated from marine sediments of Xiamen Western Sea, China and identified as Ochrobactrum sp. according to 16S rRNA gene sequence as well as Biolog microbial identification system. Strain BAP5 could grow in mineral salt medium with 50 mg/L of BaP and degrade about 20% BaP after 30 d of incubation. Ochrobactrum sp. BAP5 was able to utilize other polycyclic aromatic hydrocarbons (PAHs) (such as phenanthrene, pyrene and fluoranthene) as the sole carbon source and energy source, suggesting its potential application in PAHs bioremediation. The profile of total soluble protein from Ochrobactrum sp. BAP5 was also investigated. Some over- and special-expressed proteins of strain BAP5 when incubated with the presence of BaP were detected by two-dimensional polyacrylamide gel electrophoresis, and found to be related with PAHs metabolism, DNA translation, and energy production based on peptide fingerprint analysis through matrix-assisted laser desorption/ionization-time of flight mass spectrometry.     

耐冷腐殖酸吸附态PAHs降解菌筛选及其降解特性

为了寻找自然环境中高效PAHs降解菌,并应用于北方寒冷地区PAHs污染土壤修复,从沈抚灌渠冻融土壤中筛选出一株以腐殖酸(HA)吸附态PAHs为碳源和能源且生长良好的耐冷菌株,结合其生理生化特征和16S rDNA序列比对,鉴定此菌株为假单胞菌(Pseudomonas sp.SDR4),并研究了其对冻融土壤中HA吸附态PAHs的降解性能.结果表明:Pseudomonas sp.SDR4对冻融土壤中Phe、Pyr和Ba P均具有一定的降解能力,42 d降解率分别为73.88%、64.88%和49.39%.加入HA能够提高其对Phe、Pyr和Ba P的降解率,42 d降解率分别提高15.9%、13.8%和8.6%;在降解初期加入HA可显著提高Phe、Pyr和Ba P的降解速率,第1周,降解速率分别提高37.8%、28.4%和39.1%,但在后期促进效果减弱.若要在寒冷地区选择Pseudomonas sp.SDR4作为固定化菌株,可添加HA提高其修复效果.本研究为北方寒冷地区PAHs污染土壤修复提供了新的固定化菌种.     

Biodegradation of polycyclic aromatic hydrocarbons in sediments from the Daliao River watershed, China

The Daliao River, as an important water system in Northeast China, was reported to be heavily polluted by polycyclic aromatic hydrocarbons (PAHs). Aerobic biodegradations of four selected PAHs (naphthalene, phenanthrene, fluorene and anthracene) alone or in their mixture in river sediments from the Daliao River water systems were studied in microcosm systems. E ects of additional carbon source, inorganic nitrogen and phosphorus, temperature variation on PAHs degradation were also investigated. Results showed that the degradation of phenanthrene in water alone system was faster than that in water-sediment combined system. Degradation of phenanthrene in sediment was enhanced by adding yeast extract and ammonium, but retarded by adding sodium acetate and not significantly influenced by adding phosphate. Although PAHs could also be biodegraded in sediment under low temperature (5°C), much lower degradation rate was observed. Sediments from the three main streams of the Daliao River water system (the Hun River, the Taizi River and the Daliao River) demonstrated di erent degradation capacities and patterns to four PAHs. Average removal rates (15 or 19 d) of naphthalene, phenanthrene, fluorene and anthracene by sediment were in the range of 0.062–0.087, 0.005–0.066, 0.008– 0.016 and 0–0.059 mg/(L d), respectively. As a result, naphthalene was most easily degraded compound, anthracene was the hardest one. In multiple PAHs systems, the interactions between PAHs influenced each PAH biodegradation.     

青顶拟多孔菌对单一和复合多环芳烃的降解特性

利用中国东北林区普遍存在白腐菌——青顶拟多孔菌,降解单一和复合多环芳烃,分别测定了菲、蒽、芘于11,22,33d的累积降解率.结果显示,对于单一多环芳烃,该菌种降解能力由强到弱依次为菲>蒽>芘,33d累积降解率依次为96.56%、94.76%和57.53%;对复合多环芳烃降解中,菲和芘的累积降解率分别为99.46%和61.09%.在复合多环芳烃的降解研究中发现,少量蒽的加入,刺激了菌种对菲和芘的降解,使菲和芘的降解率分别提高了2.9%和3.56%.由此提示,在研究降解高环、难降解多环芳烃时,可利用低环多环芳烃对菌种的刺激作用,在体系内形成高、低环多环芳烃的共代谢,以达到更加高效降解多环芳烃的目的.     

Isolation of marine benzo[a]pyrene-degrading Ochrobactrum sp. BAP5 and proteins characterization

A bacterial strain BAP5 with a relatively high degradation ability of benzo[a]pyrene (BaP) was isolated from marine sediments of Xiamen Western Sea, China and identified as Ochrobactrum sp. according to 16S rRNA gene sequence as well as Biolog microbial identification system. Strain BAP5 could grow in mineral salt medium with 50 mg/L of BaP and degrade about 20% BaP after 30 d of incubation. Ochrobactrum sp. BAP5 was able to utilize other polycyclic aromatic hydrocarbons (PAHs) (such as phenanthrene, pyrene and fluoranthene) as the sole carbon source and energy source, suggesting its potential application in PAHs bioremediation. The profile of total soluble protein from Ochrobactrum sp. BAP5 was also investigated. Some over- and special-expressed proteins of strain BAP5 when incubated with the presence of BaP were detected by two-dimensional polyacrylamide gel electrophoresis, and found to be related with PAHs metabolism, DNA translation, and energy production based on peptide fingerprint analysis through matrix-assisted laser desorption/ionization-time of flight mass spectrometry.     

Degradation of pyrene by immobilized microorganisms in saline-alkaline soil

Biodegradation of polycyclic aromatic hydrocarbons (PAHs) is very difficult in saline-alkaline soil due to the inhibition of microbial growth under saline-alkaline stress. The microorganisms that can most effectively degrade PAHs were screened by introducing microorganisms immobilized on farm byproducts and assessing the validity of the immobilizing technique for PAHs degradation in pyrene-contaminated saline-alkaline soil. Among the microorganisms examined, it was found that Mycobacterium sp. B2 is the best, and can degrade 82.2% and 83.2% of pyrene for free and immobilized cells after 30 days of incubation. The immobilization technique could increase the degradation of pyrene significantly, especially for fungi. The degradation of pyrene by the immobilized microorganisms Mucor sp. F2, fungal consortium MF and co-cultures of MB+MF was increased by 161.7% (P < 0.05), 60.1% (P < 0.05) and 59.6% (P < 0.05) after 30 days, respectively, when compared with free F2, MF and MB+MF. Scanning electron micrographs of the immobilized microstructure proved the positive effects of the immobilized microbial technique on pyrene remediation in saline-alkaline soil, as the interspace of the carrier material structure was relatively large, providing enough space for cell growth. Co-cultures of different bacterial and fungal species showed different abilities to degrade PAHs. The present study suggests that Mycobacterium sp. B2 can be employed for in situ bioremediation of PAHs in saline-alkaline soil, and immobilization of fungi on farm byproducts and nutrients as carriers will enhance fungus PAH-degradation ability in saline-alkaline soil.     

芴降解优势菌的筛选鉴定及降解特性研究

从某焦化厂活性污泥中分离筛选出一株能以芴为碳源和能源生长的细菌(命名为W-2),在形态学观察和生理生化试验基础上,利用16S rDNA序列分析及系统发育学分析的方法,鉴定菌株W-2为微嗜酸寡养单胞菌(Stenotrophomonas acidaminiphila).考察了菌株W-2在液体培养基体系内对芴的降解效果,结果表明,该菌株对芴具有良好的降解特性,在初始芴浓度为40mg/L,接种量10%(V/V),pH 7.0,温度30℃条件下,接种该菌11d后,芴的降解效率达到86.0%,说明该菌在芴污染控制方面具有良好的应用前景.     

一株菲高效降解菌的环境适应性研究

生物降解是多环芳烃从环境中去除的主要途径,菲是一种典型的三环芳烃。本研究考察了一株能高效降解多环芳烃菲的鞘氨醇单胞菌GY2B在含河沙环境及不同盐度的人工海水环境中的生长特性与降解菲的情况。结果表明：河沙的加入对菌株GY2B的生长及其高效降解菲的性能均无明显影响,65 h可将起始浓度为100 mg/L的菲降解99.5%以上;而经过驯化后在添加85%人工海水的条件下该菌也仍可正常生长并高效降解菲,66 h可将起始浓度为100 mg/L的菲几乎完全降解。本研究结果可为菌株GY2B在受多环芳烃污染的河滩、河口及近海海洋环境修复中的应用提供参考依据。     

咪唑乙烟酸降解菌S181的分离鉴定及其降解特性

为咪唑乙烟酸污染土壤的生物修复提供菌源,采用瓶富集培养法,从长期施用咪唑乙烟酸的土壤中筛选出1株对咪唑乙烟酸降解能力较强的放线菌,命名为S181.该菌株可利用咪唑乙烟酸为唯一氮源生长.经形态特征、生理生化特性及16S rRNA序列分析,初步鉴定该菌株为大宫链霉菌(Streptomyces omiyaensis).以S1...     

Potential bioremediation of mercury-contaminated substrate using filamentous fungi isolated from forest soil

The use of filamentous fungi in bioremediation of heavy metal contamination has been developed recently. This research aims to observe the capability of filamentous fungi isolated from forest soil for bioremediation of mercury contamination in a substrate. Six fungal strains were selected based on their capability to grow in 25 mg/L Hg2+-contaminated potato dextrose agar plates. Fungal strain KRP1 showed the highest ratio of growth diameter, 0.831, thus was chosen for further observation.Identification based on colony and cell morphology carried out by 18S rRNA analysis gave a 98%match to Aspergillus flavus strain KRP1. The fungal characteristics in mercury(Ⅱ) contamination such as range of optimum pH, optimum temperature and tolerance level were 5.5–7 and 25–35℃ and 100 mg/L respectively. The concentration of mercury in the media affected fungal growth during lag phases. The capability of the fungal strain to remove the mercury(Ⅱ) contaminant was evaluated in 100 mL sterile 10 mg/L Hg2+-contaminated potato dextrose broth media in 250 mL Erlenmeyer flasks inoculated with 108spore/mL fungal spore suspension and incubation at 30℃ for 7 days. The mercury(Ⅱ) utilization was observed for flasks shaken in a 130 r/min orbital shaker(shaken) and nonshaken flasks(static) treatments. Flasks containing contaminated media with no fungal spores were also provided as control. All treatments were done in triplicate. The strain was able to remove 97.50%and 98.73% mercury from shaken and static systems respectively. A. flavus strain KRP1 seems to have potential use in bioremediation of aqueous substrates containing mercury(Ⅱ) through a biosorption mechanism.     

Molecular characterization of methanogenic microbial communities for degrading various types of polycyclic aromatic hydrocarbon

Knowledge on methanogenic microbial communities associated with the degradation of polycyclic aromatic hydrocarbons (PAHs) is crucial to developing strategies for PAHs bioremediation. In this study, the linkage between the type of PAHs and microbial community structure was fully investigated through 16S rRNA gene sequencing on four PAH-degrading cultures. Putative degradation products were also detected. Our results indicated that naphthalene (Nap)/2-methylnaphthalene (2-Nap), phenanthrene (Phe) and anthracene (Ant) sculpted different microbial communities. Among them, Nap and 2-Nap selected for similar degrading bacteria (i.e., Alicycliphilus and Thauera) and methanogens (Methanomethylovorans and Methanobacterium). Nap and 2-Nap were probably activated via carboxylation, producing 2-naphthoic acid. In contrast, Phe and Ant shaped different bacterial and archaeal communities, with Arcobacter and Acinetobacter being Phe-degraders and Thiobacillus Ant-degrader. Methanogenic archaea Methanobacterium and Methanomethylovorans predominated Phe-degrading and Ant-degrading culture, respectively. These findings can improve our understanding of natural PAHs attenuation and provide some guidance for PAHs bioremediation in methanogenic environment.     

生物炭固定化多环芳烃高效降解菌剂的制备及稳定性

高效多环芳烃降解微生物在污染环境中存活并保持一定生物量是实现生物强化修复的前提.本研究通过优选生物炭,优化生物炭固定化多环芳烃高效降解菌Martelella sp.AD-3的制备条件以及评估生物炭固定化菌剂的稳定性,期望获得具有应用前景的生物材料.结果显示,稻壳生物炭比表面积及孔隙大、Zeta电位高、固定化菌剂去除效果好,选择其作为固定化AD-3菌的载体.电镜观察及固定化菌剂对菲的去除率表明,固定化培养基为3% LB,接种量为2.9×10⁸ CFU·mL^-1,固定2 d时,稻壳固定化菌剂负载AD-3量最多,对菲的去除速率可达8.08 mg·L^-1·h^-1.室温保存21 d后,稻壳生物炭固定化菌剂对菲的去除速率仍达到4.46 mg·L^-1·h^-1,表明稻壳生物炭固定化AD-3菌不仅保持了菌对菲的高效降解能力,而且延长了降解微生物的保存时间,这为多环芳烃污染土壤修复提供了良好的生物修复功能材料.     

Enhancing plant-microbe associated bioremediation of phenanthrene and pyrene contaminated soil by SDBS-Tween 80 mixed surfactants

The use of surfactants to enhance plant-microbe associated dissipation in soils contaminated with polycyclic aromatic hydrocarbons(PAHs) is a promising bioremediation technology. This comparative study was conducted on the effects of plant-microbe treatment on the removal of phenanthrene and pyrene from contaminated soil, in the presence of low concentration single anionic, nonionic and anionic-nonionic mixed surfactants. Sodium dodecyl benzene sulfonate(SDBS) and Tween 80 were chosen as representative anionic and nonionic surfactants, respectively. We found that mixed surfactants with concentrations less than 150 mg/kg were more effective in promoting plant-microbe associated bioremediation than the same amount of single surfactants. Only about(m/m) of mixed surfactants was needed to remove the same amount of phenanthrene and pyrene from either the planted or unplanted soils, when compared to Tween 80. Mixed surfactants( 150 mg/kg) better enhanced the degradation efficiency of phenanthrene and pyrene via microbe or plant-microbe routes in the soils. In the concentration range of 60–150 mg/kg, both ryegrass roots and shoots could accumulate 2–3 times the phenanthrene and pyrene with mixed surfactants than with Tween 80. These results may be explained by the lower sorption loss and reduced interfacial tension of mixed surfactants relative to Tween 80, which enhanced the bioavailability of PAHs in soil and the microbial degradation efficiency. The higher remediation efficiency of low dosage SDBS-Tween 80 mixed surfactants thus advanced the technology of surfactant-enhanced plant-microbe associated bioremediation.     

耐盐石油烃降解菌的筛选鉴定及其特性研究

为得到高效耐盐石油烃降解菌,从黄河三角洲石油污染盐渍化土壤中分离出39株细菌,经液体培养初筛和土壤培养复筛实验,得到1株高效耐盐石油烃降解菌BM38.通过形态特征、生理生化特征和16S rDNA序列分析,确定该菌为恶臭假单胞菌（Pseudomonas putida）.通过液体培养实验,研究了BM38的耐盐和产生物表面活性剂特性以及对不同烃的利用能力.结果表明,在含0.5%～6.0%NaCl液体培养基中BM38生长良好,属中度耐盐菌.在高盐环境下BM38具有较强的分解石油烃能力,其中在含1.0%NaCl液体培养基中,降解7 d后,原油降解率达到73.5%;在含盐量0.22%和0.61%土壤中添加BM38,降解40 d后,土壤总石油烃降解率达到40%以上.BM38能产生一种生物乳化剂,盐浓度对这种乳化剂的乳化能力影响较大,当NaCl浓度增加到1.0%,乳化值（EI24）开始迅速降低,但在NaCl浓度为2.0%时,EI24仍达到61.0%.BM38能够利用环己烷、甲苯、异辛烷、菲和正十六烷为唯一碳源生长,其中对正构烷烃和芳烃具有较强的利用能力.     

过硫酸钠对我国典型土壤中多环芳烃氧化降解效果的影响

过硫酸钠是污染土壤化学氧化修复技术中应用较为广泛的氧化剂.为研究过硫酸钠对不同土壤中PAHs（polycyclic aromatic hydrocarbons,多环芳烃）的修复效果,以我国多种典型土壤（黑土、潮土、黄土、紫色土、褐土、砖红壤）为试验样本,以萘、菲、蒽、芘、苯并[a]芘5种PAHs为目标污染物,分析活化过硫酸钠对人为老化的降解率;此外,通过对氧化前后土壤pH、w（有机碳）等土壤性质变化的比较和分析,探讨氧化修复过程对土壤性质的影响.结果表明:当活化过硫酸钠用量为0.8 mmol/g、温度为25℃时,PAHs污染土壤中萘、菲、蒽、芘、苯并[a]芘的降解率最高,分别为87.82%、79.68%、87.93%、83.40%、94.31%.随着温度的升高,PAHs降解率逐渐升高,当温度达到25℃时,PAHs的降解率（85.69%）达到最高,随后随着温度的继续升高,总PAHs的降解率没有明显增加;随着pH的升高,PAHs的降解率逐渐升高,当pH达到6~7时,PAHs降解率维持在一个较高水平;随后随着pH的继续升高,总PAHs的降解率逐渐降低.随着温度以及pH的变化,5种PAHs的降解率与总PAHs的降解率变化趋势一致. w（有机碳）越低,PAHs环数越高,PAHs降解率越高;高环（5~6环）、中环（4环）、低环（2~3环）PAHs降解率与总PAHs降解率变化趋势一致.此外,过硫酸钠氧化修复后土壤结构遭到一定程度的破坏,土壤的pH、w（有机碳）和土壤肥力会有不同程度的下降,对土壤的再次利用有较大影响.研究显示,过硫酸钠可有效氧化降解不同性质土壤中PAHs,在氧化修复PAHs污染土壤方面具有较好的应用前景.      

低温硝基苯降解菌的筛选及降解特性研究

从被硝基苯污染的某河流底泥中分离到能在低温下生长并能以硝基苯为唯一碳源的7株细菌,其中菌株NB1在温度从2.5～35 ℃范围内时都可以生长并矿化20 mg/L的硝基苯,最适宜的生长温度为25 ℃左右;当培养温度为5 ℃时,该菌株在pH为6～9范围内可以快速降解20 mg/L硝基苯,偏碱性的条件比酸性条件更适合其生长;不超过100 mg/L的硝基苯可以被该细菌完全降解.通过生理生化反应特性、菌体形态以及16S rDNA序列测定结果,确定NB1为恶臭假单胞菌(Pseudomonas putida).不同温度条件,特别是低温下该菌株对硝基苯的快速降解特性为低温环境硝基苯污染的生物修复提供了可能.     

咪唑乙烟酸降解菌的分离、鉴定及其降解特性研究

从生产咪唑乙烟酸化工厂排污口的污泥和长期施用咪唑乙烟酸的混合土壤中分离到1株能降解咪唑乙烟酸的细菌.该菌株在72h内对500mS/L的咪唑乙烟酸降解率达到90%以上. pU为5时,500mS/L的咪唑乙烟酸72h内可全部降解,而pU 8和pH9条件下,72h咪唑乙烟酸的降解率仅为50%左右,酸性条件比碱性条件更适合降解菌的生长.25℃和30℃条件下,降解菌对咪唑乙烟酸的降解效率较高.25℃, pH 5是降解菌对咪唑乙烟酸降解的最佳条件.从形态特征、生理生化特性及 16S rRNA序列分析鉴定该菌株属于产碱菌属.     

Martelella sp. AD-3强化活性污泥耐盐降解菲效能

基于嗜盐菌Martelella sp. AD-3优配市政活性污泥搭建耐盐高效降解菲的生物反应器,在3.0 %盐度,进水菲浓度20mg/L的运行条件下,菲去除率高达97 %.批次实验证明,盐度为3.0 %,pH值为7.5~8.5,底物浓度为20~200mg/L是菲降解的最佳环境条件,此时污泥比活性高于1.0mg/(gVSS×h).各共存底物的受试浓度下,酵母提取物和苯酚促进菲降解,镉和氰化物则抑制该过程,乙酸钠、铜、铬对该过程没有明显影响.16S rRNA基因高通量测序结果表明AD-3菌在反应器内具有长期稳定性,其相对丰度维持在1.5 %.Shinella、Microbacterium和Brevundimonas是反应器中的优势菌,其相对丰度分别为20.7 %,15.1 %和11.9 %.RT-qPCR结果显示接种AD-3菌后,编码PAHs双加氧酶RHDa的功能基因差异倍数从2.1提升至11.7.     

真菌对污染旱地红壤中苯并[a]芘共代谢降解研究

在恒温和恒定转速培养条件下,模拟生物泥浆反应器法,选择从石油污染土壤中分离出来的青霉菌、黑曲霉、白腐真菌等3种真菌,在添加不同浓度菲和邻苯二甲酸作为共存底物情况下,研究其对旱地红壤中苯并[a]芘(B[a]P)的共代谢降解.结果表明,未灭菌土壤对B[a]P有降解能力,当土壤中添加菲时,提高了B[a]P在土壤中的降解率,100 mg.kg-1浓度菲处理的降解率显著高于200 mg.kg-1浓度菲处理,邻苯二甲酸对B[a]P降解影响不大.灭菌土壤中B[a]P几乎没有降解,添加共代谢底物后土壤中B[a]P降解率变化不明显.添加菲及邻苯二甲酸均促进了青霉菌对B[a]P的降解,提高其降解率,其中添加菲的效果更明显.与灭菌土壤相比,接种黑曲霉提高了B[a]P的降解率,但是添加菲与邻苯二甲酸却均抑制了黑曲霉菌对B[a]P的降解.白腐真菌对旱地红壤中B[a]P的降解能力较差,但当菲或邻苯二甲酸存在时能显著提高白腐真菌对B[a]P的降解,且菲比邻苯二甲酸效果好.