假单胞菌JX165及其完整细胞对硝基苯的好氧降解

利用Pseudomonas sp. JX165及其完整细胞考察了硝基苯的好氧降解特性,并对其降解机理进行了分析.结果表明,菌株JX165对硝基苯的好氧降解过程,首先还原产生2-氨基酚和苯胺,再经不同的氧化途径彻底降解;在高浓度-＞200mg/L-及高转速-＞200r/min-下,菌株JX165对硝基苯的降解过程会产生更难以降解的偶氮苯类化合物;经硝基苯诱导的JX165细胞中含有硝基苯、2-氨基酚、苯胺及邻苯二酚降解酶,且它们均为诱导酶,其动力学参数分别为K_m(硝基苯)=34.12mg/L,V_{max(硝基苯)}=3.01mg/(L.min);K_{m(2氨基酚)}=43.49mg/L,V_{max(2氨基酚)}s =2.56mg/-L-min-;K_{m(邻苯二酚)}51.03mg/L,_{Vmax(邻苯二酚)}=1.32mg/_(L-min);K_{m（苯胺）}=57.15mg/L,V_max(苯胺)=0.79mg/(L.min)     

Degradation of o-chloronitrobenzene as the sole carbon and nitrogen sources by Pseudomonas putida OCNB-1

A bacterial strain that utilized o-chloronitrobenzene(o-CNB) as the sole carbon,nitrogen and energy sources was isolated from an activated sludge collected from an industrial waste treatment plant. It was identified as Pseudomonas putida based on its morphology,physiological,and biochemical characteristics with an automatic biometrical system and the 16S rRNA sequence analysis. Microcosm study showed that the biodegradation of o-CNB was optimized at culture medium pH 8.0 and 32°C. At these conditions,the st...     

Isolation of a Pseudomonas Stutzeri strain that degrades 1,2,4-trichlorobenzene and characterization of its degradative plasmid

The genetic information encoding metabolic pathways for xenobiotic compounds in bacteria often resides on catabolic plasmids. The aim of the present work was to know the location of the genes for degrading 1,2,4-trichlorobenzen. In this paper a 1,2,4-trichlorobenzene-degrading strain THSL-1 was isolated from the soil of Tianjin Chemical Plant using 1,2,4-trichlorobenzene as the sole carbon source. The strain was identified as Pseudomonas stutzeri through morphologic survey and 16S rDNA sequence determination. A plasmid was discovered from strain THSL-1 by using the alkali lysis method. When the plasmid was transformed into E. coli. JM109 by the CaCl₂ method, the transformant could grow using 1,2,4-trichlorobenzene as the sole carbon source and had the degradation function of 1,2,4-trichlorobenzene. Therefore, it could be deemed that the plasmid carried the degradative genes of 1,2,4-trichlorobenzene. The average size of the plasmid was finally determined to be 40.2 Kb using selectively three kinds of restricted inscribed enzymes (HindIII, BamHI, and XholI) for single cutting and double cutting the plasmid pTHSL-1, respectively. __________ Translated from China Environmental Science, 2005, 25(4): 385–388 [译自: 中国环境科学]     

假单胞菌的氯儿茶酚1,2-双加氧酶基因的克隆和表达

 从土壤中分离得到1株降解2,4-二氯酚(2,4-DCP)能力较强的细菌菌株GT241-1,克隆了该菌株的3,5-二氯儿茶酚1,2-双加氧酶基因(dcpB),采用的克隆策略为:用Southern杂交对dcpB进行定位后,构建重组质粒,再用斑点杂交从重组质粒中筛选目的转化子.经序列测定得知dcpB亚克隆片段全长4303bp,其中dcpB基因编码区765bp.核苷酸和推测的氨基酸序列分析表明,dcpB与已在GenBank登记的相关基因有一定的差异.dcpB基因能够在大肠杆菌转化子中成功地表达有生物活性的酶.     

菲降解菌的特性及其降解酶纯化研究

研究考察了一株伯克霍尔德氏菌(Burkholderia sp.AFF)的基本特性,并对降解过程中的邻苯二酚2,3-双加氧酶进行了初步分离纯化。菌株AFF对链霉素和四环素具有抗性;当培养基中NaCl的浓度>1.5%时,菌株的生长降解能力受到抑制。菌株AFF生长与降解菲的最佳条件:温度30℃,pH=7.0,摇床转速为150r/min,接种量10%。菌株AFF的细胞粗提物具有邻苯二酚2,3-双加氧酶活性,用阴离子层析交换柱对其进行纯化,对收集到的具有酶活的组分进行SDS-PAGE分析,结果表明目的蛋白分子量在31kDa和43kDa之间。经过一步纯化后,比酶活提高33%,蛋白的回收率为27.6%。     

多环芳烃降解菌的分离鉴定及其生理特性研究

从兰州石化污水处理厂的污泥中分离出一株能分别以萘、蒽为唯一碳源生长的细菌(BDP01). 通过菌株形态观察及16S rDNA序列比对, 确定该菌株属于假单胞菌属的绿脓杆菌(Pseudomonas aeruginosa). 菌株在LB培养基中48 h内呈对数生长, 48 h到72 h进入生长稳定期, 72 h后开始进入衰亡期;其最佳培养温度为30 ℃, 最适生长pH为中性或弱碱性, 该菌能在萘和蒽的质量浓度分别为100 mg·L^-1和50 mg·L^-1的无机盐培养基中良好生长, 与已报道的其他微生物比较, BDP01在较短时间内具有较强的降解多环芳烃能力, 1%的接种量15 d内可降解89.64%的萘和77.21%的蒽. 采用PCR和琼脂糖凝胶电泳技术检测到菌株基因组中有邻苯二酚2,3-双加氧酶 (C23O)基因. 测序结果与NCBI数据库发布的C23O基因序列相似度为92%. 该酶在20~65 ℃以邻苯二酚为底物时, 细胞裂解液中酶活力变化范围为13.10~540.86 U.     

一株硝基苯高效降解菌的筛选及其降解特性

自南京化工厂下水道底泥和废水处理系统曝气池活性污泥中驯化分离得到一株能快速降解硝基苯的菌株,初步鉴定其为不动杆菌属菌株。该菌株降解硝基苯的最适宜环境条件为温度25℃～35℃,pH7～8,振荡速率大于120r/m。在适宜环境条件下,该菌株能够在24h内全部降解初始浓度不超过400mg/L的硝基苯,该浓度范围内硝基苯的降解过程符合零级动力学特征;当硝基苯的初始浓度超过400mg/L时,降解菌的生长受到毒害作用,该浓度范围内硝基苯的降解在开始有一个明显的停滞期,降解过程不再符合零级或一级动力学特征。     

Simultaneous biodegradation of nitrogen-containing aromatic compounds in a sequencing batch bioreactor

Many nitrogen-containing aromatic compounds (NACs), such as nitrobenzene (NB), 4-nitrophenol (4-NP), aniline (AN), and 2,4-dinitrophenol (2,4-DNP), are environmentally hazardous, and their removal from contaminated water is one of the main challenges facing wastewater treatment plants. In this study, synthetic wastewater containing NB, 4-NP, 2,4-DNP, and AN at concentrations ranging from 50 to 180 mg/L was fed into a sequencing batch reactor (SBR). Analyses of the SBR system indicated that it simultaneously removed more than 99% of the NACs at loading rates of 0.36 kg NB/(m3·d), 0.3 kg 4-NP/(m3·d), 0.25 kg AN/(m3·d), and 0.1 kg 2,4-DNP/(m3·d). Bacterial groups of Bacteriodetes, Candidate division TM7, α-Proteobacteria, and β-Proteobacteria were dominant in the clone libraries of 16S rRNA genes retrieved from the microbial communities in the SBR system. "Cycle tests" designed to alter feeding and aeration parameters of the SBR system demonstrated that the resident microbial biome of the SBR system responded rapidly to changing conditions. Consumption of O2 was concomitant with the apparent mineralization of NACs. Aromatic ring-cleaving dioxygenase activities suggested that (1) AN and NB were degraded via catechol 2,3-dioxygenase; (2) 4-NP was degraded via 1,2,4-benzentriol 1,2-dioxygenase; and (3) 2,4-DNP was degraded via an unresolved pathway.     

Screening for a new Streptomyces strain capable of e cient keratin degradation

Keratinous wastes could be degraded by some microorganisms in nature. Native human foot skin （NHFS） was used as sole nitrogen source to screen microorganisms with keratin-degrading capability. From approximately 200 strains, a strain of Streptomyces sp. strain No. 16 was found to possess the strongest keratinolytic activity, and the total activity in the culture was 110 KU/ml with specific activity of 2870 KU/mg protein （KU： keratinase unit）. Substrate specificity test indicated that the crude keratinase could degrade keratin azure, human hair, cock feathers and collagen. The optimal pH of the crude keratinase ranged from 7.5 to 10 and the temperature ranged from 40℃ to 55℃. Metal chelating agent ethylenediamine tetraacetic acid obviously stimulated the keratinolytic activity but suppressed the proteolytic activity. To our knowledge, this is the first report on specific induction of keratinases by NHFS from an actinomycete. Moreover, excellent characteristics of its crude keratinase may lead to the potential application in waste treatment and recovery, poultry and leather industry, medicine, and cosmetic development.     

一株耐酒精纤维素分解菌的分离鉴定及生长条件研究

木薯酒精糟因其低营养高含水量而难以利用和处理,目前对新鲜木薯酒精糟主要采用机械脱水及干燥的方法,但此类方法的经济成本和能耗都较高,很难形成产业化。该实验在木薯酒精糟中通过用刚果红培养基初筛和杜博氏培养基复筛得到一株耐酒精的纤维素分解菌,编为13号。经16S rRNA序列鉴定与链霉菌同源性达99%。其最适生长pH值为6.5,最适生长温度为40℃。对酒精有一定的耐受性,能在酒精浓度为6%的环境中生长,并且在酒精浓度为1%时对菌株生长影响不大。同时该菌能在40℃的温度条件下保持良好的生长态势,对适应废物堆肥发酵后期熟化产生有益次生代谢产物有利。所以利用此菌耐酒精、产纤维素酶且较耐高温等特性,对于处理含酒精的纤维素废物具有较好的应用前景。     

多氯联苯降解菌Pseudomonas sp. DN2的分离鉴定及其降解特性研究

从长期受PCBs污染的土壤中经富集培养筛选分离到1株能以联苯为唯一碳源和能源生长的革兰氏阴性细菌DN2.经形态观察、16S rDNA序列分析和系统发育分析初步鉴定为Pseudomonas sp..对其联苯双加氧酶的bphA1基因核心区和2,3-二羟基联苯-1,2-双加氧酶活力进行了分析测定,结果表明该菌具有降解联苯/PCBs的相关基因存在.利用GC-MS测试该菌的静息细胞降解PCBs的能力表明,该菌株可以在3 d内显著而高效地降解Aroclor 1242总量的67%,其对Aroclor 1242中各单体同系物的降解率在30%～90%之间.这对PCBs污染场地的原位生物修复具有实际应用意义.     

Characterization and phylogenetic analysis of a phenanthrene-degrading strain isolated from oil-contaminated soil

Bacterium strain EVA17 was isolated from an oil-contaminated soil, and identified as Sphingomonas sp.based on analysis of 16S rDNA sequence, cellular fatty acid composition and physiological-chemical tests. The salicylate hydroxylase and catechol 2, 3-dioxygenase (C23O) were detected in cell-free lysates, suggesting a pathway for phenanthrene catabolism via salicylate and catechol. Alignment showed that both of the C23O and GST genes of the strain EVA17 had high similarity with homologues of strains from genus Sphingomonas. The phylogenetic analysis based on 16S rDNA and C23O gene sequence indicated that EVA17 should be classified into genus Sphingomonas, although the two phylogenetic trees were slightly different from each other. The results of co-amplification and sequence determination indicated that GST gene should be located upstream of the C230 gene.     

Isolation and preliminary characterization of a 3-chlorobenzoate degrading bacteria

A study was conducted to compare the diversity of 2-, 3-, and 4-chlorobenzoate degraders in two pristine soils and one contaminated sewage sludge. These samples contained strikingly different populations of mono-chlorobenzoate degraders. Although fewer cultures were isolated in the uncontaminated soils than contaminated one, the ability of microbial populations to mineralize chlorobenzoate was widespread. The 3- and 4-chlorobenzoate degraders were more diverse than the 2-chlorobenzoate degraders. One of the strains isolated from the sewage sludge was obtained. Based on its phenotype, chemotaxonomic properties and 16S rRNA gene, the organism S-7 was classified as Rhodococcus erythropolis. The strain can grow at temperature from 4 to 37℃. It can utilize several （halo）aromatic compounds. Moreover, strain S-7 can grow and use 3-chlorobenzoate as sole carbon source in a temperatures range of 10-30℃ with stoichiometric release of chloride ions. The psychrotolerant ability was significant for bioremediation in low temperature regions. Catechol and chlorocatechol 1,2-dioxygenase activities were present in cell free extracts of the strain, but no （chloro）catechol 2,3- dioxygenase activities was detected. Spectral conversion assays with extracts from R. erythropolis S-7 showed accumulation of a compound with a similar UV spectrum as chloro-cis,cis-muconate from 3-chlorobenzoate. On the basis of these results, we proposed that S-7 degraded 3-chlorobenzoate through the modified ortho-cleave pathway.     

硝基苯好氧降解菌筛选及其降解特性

针对化学试剂厂和制药厂废水中的硝基苯,经菌源筛选与长期好氧驯化,分离到1株能有效降解硝基苯的菌株Bacillus subilis(枯草芽孢杆菌)。系统研究了其生长条件及降解硝基苯的特性,适宜生长条件: pH为5.0～8.0,温度为30～40℃,NH4Cl浓度为100～200 mg/L,在降解硝基苯的过程中有苯胺生成,这为含低浓度硝基苯废水的生物处理提供了新的途径,即不需要厌氧工艺而直接用好氧技术就可将废水完全无害化。      

Isolation and characteristics of a novel biphenyl-degrading bacterial strain, Dyella ginsengisoli LA-4

A novel biphenyl-degrading bacterial strain LA-4 was isolated from activated sludge. It was identified as Dyella ginsengisoli according to phylogenetic similarity of 16S rRNA gene sequence. This isolate could utilize biphenyl as sole source of carbon and energy, which degraded over 95 mg/L biphenyl within 36 h. The major metabolites formed from biphenyl, such as 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid (HOPDA) and benzoic acid, were identified by LC-MS. The crude cell extract of strain LA-4 exhibited the activity of 2,3-dihydroxybiphenyl 1,2-dioxygenase (2,3-DHBD) and the kinetic parameters were Km= 26.48 μmol/L and Vmax= 8.12 μmol/mg protein. A conserved region of the biphenyl dioxygenase gene bphA1 of strain LA-4 was amplified by PCR and confirmed by DNA sequencing.     

强抗镉蜡状芽孢杆菌的分离鉴定及其抗性机理

从北京市远郊东三岔地区废弃的铅锌尾矿中筛选到1株强抗镉的细菌.通过细菌形态学观察、扫描电镜、X射线衍射能谱分析、甲基乙酰甲醇试验与酪氨酸水解试验等特征生理生化试验、菌株“G＋C mol%”值以及16S rDNA扩增与序列测定,鉴定此菌株为蜡状芽孢杆菌（Bacillus cereus）.分离菌株可以在镉浓度为1?000 mg/L的固体培养基平板上生长,表明菌株具有强抗镉的能力.进一步分析了菌株在添加Cd²⁺与未添加Cd²⁺的液体培养基中的生长曲线,并通过质粒消除试验证明该菌株的抗镉性质与抗性质粒的存在有关.     

一株苯酚降解菌的分离鉴定及响应面法优化其固定化

从太原市焦化厂废水活性污泥中分离、筛选出一株苯酚降解细菌,经生理生化反应和16S rRNA鉴定,该菌株为Diaphorobacter属细菌,命名为PD-07.代谢机制研究表明,苯酚可诱导该菌合成邻苯二酚2,3-加氧酶降解苯酚.为了提高该菌株对苯酚的降解率,以海藻酸钙为材料,对该菌株进行包埋固定化研究.首先采用Plackett–Burman实验设计筛选出影响固定化菌株苯酚降解率的关键因素,然后采用最陡爬坡实验逼近最大苯酚降解率响应区域.最后用Box–Behnken实验设计及响应面回归分析,应用二次方程对实验数据进行拟合得,拟合曲线与实验实测值相关性良好,最佳条件为海藻酸钠浓度3.83%(m/V)、CaCl2 0.3mol/L、菌胶比1:26.73、固定化时间2h、摇床转速180r/min、培养温度30℃、初始pH值7.2、液固比4.86:1,在此条件下苯酚降解率可达96.89%.     

一株菲降解细菌的分离及其特性

利用平板熏蒸和双氧加合酶活性测定的方法 ,从大庆油田水体中筛选出 1株对菲具有降解活性的细菌 ,Phx1.在LB培养基中 ,能将浓度为 10 0 μg/mL的菲在 2 4h内完全降解 .与无机盐培养基相比 ,LB和牛肉膏蛋白胨培养基能促进此菌对菲的降解 ,尤其能促进对中间产物的进一步转化 .经质粒消除证明 ,Phx1具有 2个质粒 ,且较大的 1个对菲的降解是必须的 .通过对其形态、生理生化特性 ,16SrDNA序列分析 ,鉴定Phx1为土壤杆菌属细菌 ,其 16SrDNA序列与Agrobacterium tumefaciensUP 3、Agrobacterium albertimagni的相似性最高 ,分别达到 97 8%和 96 6 % ,但对向日葵的致瘤实验为阴性 .     

两株假单胞菌降解酚类化合物的特性

从焦化废水中分离得到2株可降解对氯酚的假单胞菌(Pseudomonas)CO-1和CO-44.其最适降解温度为35~40℃,最适pH值为6.0~8.0.菌株降解对氯酚的速度与对氯酚初始浓度呈负相关.2株细菌均能较快地降解苯酚和甲酚,其中CO-1还能够降解1-萘酚和萘.在添加对氯酚的焦化废水中,CO-1和CO-44能够在42h内将苯酚、甲酚和对氯酚完全降解.从2株细菌中均检测到了苯酚羟化酶基因,分别从菌株CO-1和CO-44中检测到邻苯二酚1,2-双加氧酶基因和邻苯二酚2,3-双加氧酶基因.