阿特拉津降解菌T_3 AB_1的分离鉴定及土壤修复

【目的】从阿特拉津污染土壤分离高效降解菌株,进行分类学鉴定、降解特性及黑土修复能力初步研究,为阿特拉津污染土壤微生物修复提供新的菌株。【方法】通过形态特征、生理生化特征和16S rDNA序列分析方法进行菌株鉴定;通过培养时间、温度、pH值等环境因素的研究得出菌株的最佳降解条件;通过降解菌株接种于不同种类除草剂为唯一碳氮源培养基获得该菌株的降解谱;通过土壤接种和敏感作物盆栽生测试验验证菌株对阿特拉津污染土壤修复能力。【结果】本试验从黑龙江省讷河市长期施用阿特拉津的玉米田地中分离出一株能以阿特拉津为唯一碳氮源生长的细菌T3AB1,初步鉴定为节杆菌属(Arthrobacter sp.),该菌株在72 h内对500 mg/L阿特拉津(pH 8.0)的降解率高达99%,其降解能力较高的条件为pH7.0-8.0、25-30℃、摇培72-108 h,该菌株能够利用甲氧咪草烟、咪唑乙烟酸、氟磺胺草醚、氟乐灵、异噁草松为唯一碳氮源进行生长,处理168 h的降解率能够达到12.66%-40.54%,该菌株处理21 d能够显著恢复敏感作物水稻的各项生物量指标,且随着处理时间的延长,其对土壤的修复作用也会逐渐增强。【结论】从黑龙江省污染土壤中筛选得到的高效降解阿特拉津的节杆菌属近缘种T3AB1,土壤接种实验表明该菌株具有很好的土壤修复作用,可为阿特拉津生物修复的研究提供适宜菌种资源。     

阿特拉津降解菌株的分离和鉴定

从农药厂废水中分离到6株能以除草剂阿特拉津为唯一氮源生长的细菌,即假单胞菌（Pseudomonas spp,.)AD1,AD2和AD6,土壤杆菌（Agrobacterium sp.)AD4,黄单胞菌（Xanthomonas sp.)AD5,欧氏菌（Erwinia sp.)AD7,AD1菌株能使无机盐培养基中的0．3g/L阿特拉津在72h内降解99．9％,当以AD1,AD2,AD4,AD5,AD6和AD7菌株的总DNA为模板进行PCR扩增时,除AD2菌株以外,均得到了与献报道的假单胞菌ADP菌株的阿特拉津氯水解酶基因（atzA)同源的PCR产物。     

一株阿特拉津降解菌的分离鉴定及降解特性

从农药厂废水处理池的活性污泥中分离到一株阿特拉津降解菌X-4, 根据其生理生化特性和16S rRNA基因序列相似性分析, 将其初步鉴定为节杆菌属(Arthrobacter sp.)。该菌能以阿特拉津为唯一碳氮源生长, 42 h内对100 mg/L的阿特拉津降解效果为95.7%, 降解阿特拉津的最适温度为30 °C, pH为7.0。该菌对多种重金属离子都存在抗性, 显示了其在去除阿特拉津和重金属复合污染方面的应用潜力。对其降解基因的初步研究显示, 该菌含有trzN、atzB和atzC 3个阿特拉津降解相关基因。     

阿特拉津降解菌SA1的分离鉴定及其降解特性研究

为进行阿特拉津(AT)污染的生物修复,从AT降解混合菌群中,经长期的交替液体摇瓶培养和平板划线分离,筛选到一株能完全降解AT的菌株SA1。经生理生化特征及16S rDNA序列分析,将该菌鉴定为假单胞菌属(Pseudomonas sp.)。与已报道的AT降解菌Pseudomonas sp.ADP不同,SA1能以AT为唯一碳源、氮源和能源生长,培养基中添加铵盐不抑制SA1的降解功能,而添加葡萄糖时,累积的氰尿酸会被快速降解。SA1生长的最适温度为37℃,最适pH值为7.0。SA1的静息细胞在10℃~40℃或pH值4~11时均能高效降解AT,比ADP降解具有更广的pH和温度范围,表明SA1降解菌株具有广阔的应用前景。SA1中AT降解基因为保守的atzABCD,并含有IS1071的tnpA基因片段,传代过程中降解基因会以一定频率丢失。     

阿特拉津降解菌株DNS32的降解特性及分类鉴定与降解途径研究

【目的】研究阿特拉津降解菌株DNS32的菌种分类、降解特性及降解途径,丰富阿特拉津降解菌菌种资源。【方法】在长期施用阿特拉津的东北地区寒地黑土中筛选出一株以阿特拉津为唯一氮源生长的降解菌株DNS32,测定其基本降解特性,通过16S rRNA序列分析进行分类鉴定,并利用阿特拉津降解基因PCR扩增技术及降解产物生成量的测定,进一步揭示其降解途径。【结果】实验结果发现DNS32菌株具有较好的降解能力,且在相对较低温度下也具有一定的降解能力。16S rRNA序列分析结果表明DNS32与鲁氏不动杆菌(Acinetobacter lwoffii)16S rRNA序列同源性高达99%。成功地扩增降解基因trzN、atzB及atzC,实验结果表明DNS32遵循Arthrobacter aurescens TC1的降解模式,可将阿特拉津降解为氰尿酸,降解产物的生成量测定也证明了这一点。【结论】实验结果丰富了阿特拉津降解菌菌种资源,为不动杆菌属的阿特拉津降解菌研究提供了参考。     

阿特拉津降解菌株的分离和鉴定*

从农药厂废水中分离到６株能以除草剂阿特拉津为唯一氮源生长的细菌,即假单胞菌（Ｐｓｅｕ－ｄｏｍｏｎａｓ ｓｐｐ．）ＡＤ１、ＡＤ２和 ＡＤ６,土壤杆菌（Ａｇｒｏｂａｃｔｅｒｉｕｍ　ｓｐ．）ＡＤ４,黄单胞菌（Ｘａｎｔｈｏｍｏｎａｓ ｓｐ．）ＡＤＳ,欧文氏菌（Ｅｒｗｉｎｉａ ｓｐ．）ＡＤ７。ＡＤ１菌株能使无机盐培养基中的 ０．３ｇ／Ｌ阿特拉津在７２ｈ内降解９９,９％。当以ＡＤ１、ＡＤ２、ＡＤ４、ＡＤ５、ＡＤ６和ＡＤ７菌株的总ＤＮＡ为模板进行ＰＣＲ扩增时,除Ａ     

阿特拉津降解菌株AD111的分离鉴定及其降解特性

【背景】玉豆轮作过程中,玉米田中长残留除草剂阿特拉津易对下茬大豆作物产生不良影响。【目的】从黑龙江省安达市的农田土筛选一株能适应该土壤环境生长的阿特拉津降解菌并研究其降解特性。【方法】利用富集培养法,分离、筛选一株阿特拉津高效降解菌并结合外观形态、生理生化及16SrRNA基因序列测定对其进行鉴定,通过单一变量法设置不同的碳源、pH、温度和阿特拉津浓度,研究降解菌株最佳发酵及降解条件。【结果】得到一株在BSM-G中能够以阿特拉津为唯一氮源生长的高效阿特拉津降解菌AD111,鉴定为马德普拉塔无色小杆菌(Achromobacter marplatensis)。菌株AD111降解阿特拉津的最适温度为35℃,最适pH为8.0,最佳碳源为蔗糖,24 h内对浓度为50 mg/L的阿特拉津降解率达到99.7%,对300 mg/L的阿特拉津降解率达到81.9%。【结论】降解菌AD111具有较好的环境适应及阿特拉津降解能力,为解决黑龙江偏碱土壤中阿特拉津残留提供了良好的候选菌株。     

阿特拉津及其降解菌的使用对土壤微生物群落的影响

比较了阿特拉津及降解菌株BTAH1的使用对土壤微生物的影响.结果表明,在实验周期内阿特拉津对土壤微生物的代谢作用有较明显的刺激作用,与空白土壤(未施用阿特拉津和降解菌)相比,对照土壤(施用50mg·kg^-1土阿特拉津)呼吸强度显著增加,且土壤中的阿特拉津浓度对土壤NH₄⁺-N和NO₃^--N浓度的影响显著.降解菌BTAH1可在1周内降解土壤中98%以上的阿特拉津,从而使土壤呼吸强度有所下降,土壤中NH₄⁺-N和NO₃^--N的浓度基本与空白土壤持平,对微生物量C和微生物量N影响不显著;放线菌和真菌数量也基本与空白持平,细菌数量较高.对土壤细菌的16SrDNA文库的ARDRA分析发现,阿特拉津及其降解菌的使用对土壤细菌群落结构有一定程度的影响,阿特拉津的使用会降低细菌群落的多样性,而降解菌的使用会恢复土壤细菌的多样性.     

阿特拉津降解菌SYSA的分离筛选和鉴定

从长期施用阿特拉津的土壤中筛选到1株能够以阿特拉津为惟一碳源生长的菌株SYSA,经生理生化特性鉴定和16S rDNA序列分析,该菌为阴沟肠杆菌(Enterobacter cloacae).对SYSA菌的生物学特性研究表明,pH 7-8,30℃时,在以阿特拉津(20 mg/L)为惟一碳源的培养基上经146 h培养,降解率为87%.     

除草剂阿特拉津对土壤脲酶活性的影响

研究了阿特拉津对4种典型施肥处理的土壤脲酶活力的影响。结果表明,处理初期,低浓度阿特拉津对土壤脲酶有一定刺激作用,高浓度处理在整个试验过程中对脲酶有明显抑制作用,阿特拉津对不同肥力土壤中脲酶的影响有明显差异,对照土壤和NPK肥土壤中脲酶活力较低,脲酶受抑制明显,抑制率分别高达30．35％和28．89％;NPK+秸秆和NPK+有机肥土壤的脲酶活力高,脲酶抑制率低,最高抑制率分别为21．35％和16．86％,不同肥力土壤在整个处理过程中,脲酶抑制率均为先逐渐增大到最大值,然后又逐渐降低;高肥力土壤脲酶抑制率最大值出现的时间比低肥力土壤迟,表明高肥力土壤对阿特拉津有较强的耐受能力。     

节杆菌菌株LZP08X的分离鉴定及其降解苯酚特性

目的:从柳州市某钢铁公司焦化污水处理厂的活性污泥中,筛选分离出高效降解苯酚的菌株,对其进行分类鉴定和苯酚降解特性的研究。方法:采用苯酚为唯一碳源和能源的无机盐培养基,经过富集培养,平板分离,耐受性试验和苯酚降解能力测定。结果:获得一株能高效降解苯酚的菌株,命名为LZP08X,通过形态观察和生理生化特征分析,初步鉴定为节杆菌属(Ar-throbacter sp.)。该菌株降解苯酚的最适条件为:温度35℃,pH 9.0,装液量20%(v/v);其降解苯酚过程符合一级反应动力学方程,在苯酚初始浓度为400mg/L时,于12h内,降解率达99%,降解速率常数K值为0.39,半衰期为1.78h。结论:节杆菌属(Arthrobact-er sp.)菌株LZP08X苯酚降解能力较强,对该菌的继续研究,可使其在含酚工业废水处理的实际应用中起到重要作用。     

Isolation and characterization of Arthrobacter sp. strain MCM B-436, an atrazine-degrading bacterium, from rhizospheric soil

Atrazine is one of the most environmentally prevalent s-triazine-ring herbicides. The widespread use of atrazine and its toxicity necessitates search for remediation technology. As atrazine is still used in India as a major herbicide, exploration of atrazine-degrading bacterial community is of immense importance. Considering lack of reports on well characterized atrazine-degrading bacterial cultures from India and wide diversity and density of microorganisms in rhizosphere, soil sample from rhizosphere of atrazine-resistant plant was studied. Arthrobacter sp. strain isolated in this investigation utilizes atrazine as the sole nitrogen source. In addition, the bacterium degrades other triazines such as ametryn, cyanizine, propazine and simazine. PCR analysis confirms the presence of atzBCD and triazine hydrolase (trzN) genes on chromosomal DNA. Sequencing of the trzN gene reveals high sequence similarity with trzN from Nocardioides sp. C190. An inducible and intracellular atrazine chlorohydrolase enzyme was isolated and partially purified from this isolate. This study confirms the presence of atrazine-degrading microbial population in Indian soils and could be used efficiently for remediation of contaminated soils. Presence of trzN gene indicates possible presence of bacterial community with more efficient and novel enzymatic capabilities. Comparison of enzyme and gene structure of this isolate with other geographically distinct atrazine-degrading strains will help us in the better understanding of gene transfer and evolution.     

Isolation of Fenitrothion-degrading Strain Burkholderia sp. FDS-1 and Cloning of mpd Gene

A short rod shaped, gram-negative bacterium strain Burkholderia sp. FDS-1 was isolated from the sludge of the wastewater treating system of an organophosphorus pesticides manufacturer. The isolate was capable of using fenitrothion as the sole carbon source for its growth. FDS-1 first hydrolyzed fenitrothion to 3-methyl-4-nitrophenol, which was further metabolized to nitrite and methylhydroquinone. The addition of other carbon source and omitting phosphorus source had little effect on the hydrolysis of fenitrothion. The gene encoding the organophosphorus hydrolytic enzyme was cloned and sequenced. The sequence was similar to mpd, a gene previously shown to encode a parathion-methyl-hydrolyzing enzyme in Plesiomonas sp. M6. The inoculation of strain FDS-1 (10⁶ cells g⁻¹) to soil treated with 100 mg fenitrothion emulsion kg⁻¹ resulted in a higher degradation rate than in noninoculated soils regardless of the soil sterilized or nonsterilized. These results highlight the potential of this bacterium to be used in the cleanup of contaminated pesticide waste in the environment. Zhonghui Zhang, Qing Hong: Both authors contributed equally to this work     

Arthrobacter sp. lipase immobilization for improvement in stability and enantioselectivity

Arthrobacter sp. lipase (ABL, MTCC no. 5125) is being recognized as an efficient enzyme for the resolution of drugs and their intermediates. The immobilization of ABL on various matrices for its enantioselectivity, stability, and reusability has been studied. Immobilization by covalent bonding on sepharose and silica afforded a maximum of 380 and 40 IU/g activity, respectively, whereas sol–gel entrapment provided a maximum of 150 IU/g activity in dry powder. The immobilized enzyme displayed excellent stability in the pH range of 4–10 and even at higher temperature, i.e., 50–60°C, compared to free enzyme, which is unstable under extreme conditions. The resolution of racemic auxiliaries like 1-phenyl ethanol and an intermediate of antidepressant drug fluoxetine, i.e., ethyl 3-hydroxy-3-phenylpropanoate alkyl acylates, provided exclusively R-(+) products (∼99% ee, E=646 and 473), compared to cell free extract/whole cells which gave a product with ∼96% ee (E=106 and 150). The repeated use (ten times) of covalently immobilized and entrapped ABL resulted in no loss in activity, thus demonstrating its prospects for commercial applications.     

1株拮抗真菌的生防细菌Xj11的分离与鉴定

从江西省井冈山自然保护区发病毛竹中分离到1株具有明显抑制植物病原真菌活性的生防细菌Xj11。其对小麦赤霉病菌、尖孢镰刀菌、甘蔗节菱孢菌、链格孢菌和炭角菌均有较强的抑制作用。通过形态特征和培养特征观察、生理生化实验及16S rDNA序列系统发育分析,初步鉴定该菌为唐菖蒲伯克霍尔德氏菌,命名为Burkholderia gladioli strain Xj11。     

高效氯氰菊酯降解菌JCN13的分离鉴定及降解特性研究

从生产高效氯氰菊酯的农药厂污水曝气池中,分离到一株能降解高效氯氰菊酯并以之为唯一碳源进行生长的细菌JCN13.经生理生化试验和16S rDNA分析,鉴定菌株JCN13为沙雷菌属(Serratia sp.).气相色谱检测,菌株JCN13在4 d内对100 mg/L高效氯氰菊酯的降解率为89%,8 d内基本降解完全.经气质联用检测,发现高效氯氰菊酯在被菌株JCN13降解的过程中存在异构体的转化.     

Use of atrazine sensitive leguminous plants as biological indicators to evaluate the atrazine degradation efficiency of a bacterial inoculum

Atrazine sensitive leguminous plants were grown in a soil spiked with atrazine and augmented with an atrazine-degrading bacterium, Arthrobacter sp. strain MCM B-436, to ascertain its degradative efficiency. Germination and survival of plants was correlated with atrazine removal from soil. This experiment was carried out at laboratory as well as field level, showing consistent results. This bioindicator approach serves as an efficient measure for atrazine removal and could be easily adapted to determine atrazine degradation efficiency of other microbial strains.     

1株金黄杆菌分离、鉴定及其胞外蛋白酶活性的初步研究

从无指盘臭蛙(Odorrana grahami)皮肤表面分离到1株具有胞外蛋白水解酶活性的菌株,命名为CW-E2T。初步研究发现CW-E2T系革兰阴性,菌体短杆状,无鞭毛,菌落呈圆形,边缘光滑并相互粘连,表面突起,产生黄色素,有金黄色光泽细菌。通过形态学、生理生化和16SrRNA基因序列分析,确定CW-E2T为金黄杆菌属内的一个未定种。在此基础上对该菌株蛋白水解酶进行了初步研究,确定菌株CW-E2T胞外蛋白酶为金属蛋白酶。以上对CW-E2T细菌分类学研究和胞外蛋白酶的初步研究,为该菌株的大量培养以及胞外蛋白酶的进一步研究提供理论依据。     

Isolation and partial characterization of plasmid DNA from Arthrobacter oxidans

A method for the extraction of the high molecular weight plasmid AO 1 from the gram-positive soil bacterium Arthrobacter oxidans is presented.Following digestion of this DNA with the restriction endonucleases Accl, Bam HI, Eco RI and Hind III, an average molecular mass of 157.8 kb was estimated. This value is in good agreement with the 160 kb size determined previously by electron microscopy (Brandsch et al. 1982).Using the same method, no plasmid DNA was found in strains of the genus Arthrobacter which do not degrade nicotine, e.g., A. albidus, A. globiformis and A. auricans.Abbreviations EDTA ethylenediaminetetraacetic acid - Kb kilobasepairs - SDS sodium dodecyl sulfate - Tris Tris-(hydroxymethyl)-aminomethan     

Mineralization of diuron [3-(3,4-dichlorophenyl)-1, 1-dimethylurea] by co-immobilized <Emphasis Type=Italic>Arthrobacter</Emphasis> sp. and <Emphasis Type=Italic>Delftia acidovorans</Emphasis>

Mineralization of diuron has not been previously demonstrated despite the availability of some bacteria to degrade diuron into 3,4-dichloroaniline (3,4-DCA) and others that can mineralize 3,4-DCA. A bacterial co-culture of Arthrobacter sp. N4 and Delftia acidovorans W34, which respectively degraded diuron (20 mg l⁻¹) to 3,4-DCA and mineralized 3,4-DCA, were able to mineralize diuron. Total diuron mineralization (20 mg l⁻¹) was achieved with free cells in co-culture. When the bacteria were immobilized (either one bacteria or both), the degradation rate was higher. Best results were obtained with free Arthrobacter sp. N4 cells co-cultivated with immobilized cells of D. acidovorans W34 (mineralization of diuron in 96 h, i.e., 0.21 mg l⁻¹ h⁻¹ vs. 0.06 mg l⁻¹ h⁻¹ with free cells in co-culture).