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.     

一株焦化废水降酚菌的质粒初步研究

从处理焦化废水的活性污泥池中,分离得到了一株具有降酚能力的细菌, Ｊ Ａ． Ｊ Ａ 可以在含酚的 Ｈ Ｐ 复杂培养基中生长,也可以在以酚为唯一碳源的 Ａ Ｂ Ｐ 培养基中生长,且具有产黑色素的特点．用碱裂解提取质粒的方法发现它含有一个质粒,定名为ｐ Ｘ Ｈ１ ,并测定了这个质粒的大小约为２ ．６ ｋｂ ．用限制性内切酶 Ｂｇｌ Ｉ和 Ｒｓａ Ｉ进行双酶切,作出了初步的物理图谱．ｐ Ｘ Ｈ１ 经过２０ 次没有酚作为选择压传代培养后仍然能１００ ％ 保留,同时, Ｊ Ａ菌仍具有降酚能力,表明其具有很高的遗传稳定性     

萘降解质粒pND6的分离和鉴定

从工业废水中分离的假单胞菌 (Pseudomonassp .)ND6菌株 ,能以萘为唯一碳源生长 ,使无机盐培养基(MM)中 2g/L的萘在 48h内降解 98% .该菌株含有一个 115kb的大质粒pND6 .DNA杂交实验表明 ,pND6质粒含有与恶臭假单胞菌 (P .putida)G7菌株的NAH7质粒同源的萘降解基因 .图 3表 1参 14     

多环芳烃降解菌ZL5分离鉴定及其降解质粒

通过选择性富集培养，从辽河油田石油污染土壤中分离到一株多环芳烃(PAHs)降解菌ZL5．它能以菲和芘为唯一碳源生长，但是不能利用萘．16S rDNA核苷酸序列分析结果表明，ZL5属于变形细菌α亚类中的鞘氨醇单胞菌属．该菌株含有一个大小约为60kb的质粒．丝裂霉素C消除实验表明，随着质粒的丢失，菌株利用菲和芘的能力也丧失．用电转化和氯化铷转化法分别将菌株ZL5的质粒导人大肠杆菌JM109和DH5α中，随着质粒的获得，这些转化子获得了降解菲和芘的能力．本研究结果表明，鞘氨醇单胞菌ZL5降解PAHs的功能和质粒有关。     

红球菌Chr-9降解吡啶和苯酚的特性

研究了红球菌(Rhodococcus)Chr-9菌株在基础盐培养基中降解吡啶和苯酚的特性,分析了菌株降解苯酚和吡啶间的差异.结果表明,菌株Chr-9能够在72 h内将基础盐培养基中的吡啶(200 mg L-1)和苯酚(200 mg L-1)完全降解,同时利用吡啶和苯酚进行生长.菌株降解吡啶的最适温度为35℃,最适pH为8.0.菌株降解吡啶和苯酚的速度与底物的初始浓度呈负相关;在无其它氮源的基础盐培养基中,菌株能够利用吡啶和苯酚协同生长.图7参12     

费氏中华根瘤菌ntrBC基因突变株的构建及其特性

从费氏中华根瘤菌(Sinorhizobium fredii)HN01基因文库的重组质粒pGXHN300上获得ntrB、ntrC基因完整序列.通过自杀质粒pK18mob同源单交换的方法,分别获得了HN01菌株的ntrB和ntrC基因的突变株GXHNB、GXHNC.功能检测证明GXHNB、GXHNC能以硝酸盐为唯一氮源进行正常生长,但不能以铵源作为唯一氮源进行正常生长.图4表2参15     

Identification of two alkane hydroxylases involved in long-chain n-alkane degradation by Dietzia sp. DQ12-45-1b北大核心CSCD

The aim of this study was to identify genes involved in long-chain alkane degradation in Dietzia sp. DQ12-45-1b. Functional genes were annotated by genome analysis. Induction of alkane hydroxylase genes by C28 n-alkane was analyzed by using quantitative real-time PCR in wild-type Dietzia sp. DQ12-45-1b and its alkW1 gene knockout mutant strain M 5-5. From the genome of Dietzia sp. DQ12-45-1b, two homologues, G1 and G2 genes were annotated, which showed 50% amino acid sequence similarity with AlmA from Acinetobacter sp. DSM17874, and 48% amino acid sequence similarity with LadA from Geobacillus thermodenitrificans NG80-2, respectively. In addition, G1 showed 71% amino acid sequence similarity with G1a, and G2 showed 34% and 87% amino acid sequence similarities with G2a and G2ß, respectively, which were annotated from Dietzia sp. E1 genome. In addition, the alkW1 gene knockout strain M 5-5 could grow with C28 n-alkane as the sole carbon source, indicating the presence of potential long-chain alkane hydroxylase gene(s) other than alkW1 in Diezia sp. DQ12-45-1b. Accordingly, induction of G1 and G2 genes was observed when Dietzia ap. DQ12-45-1b and alkW1 knockout mutant strain M 5-5 grew with C28 n-alkane as sole carbon source. The results indicated that G1 and G2 genes are mostly responsible for the degradation of long-chain alkanes in Dietzia sp. DQ12-45-1b, which has unique multiple alkane hydroxylase systems.     

一株能产生聚乙烯醇降解酶的委内瑞拉链霉菌

以聚乙烯醇(PVA)为唯一碳源,从土壤中筛选到1株能产生胞外PVA降解酶的放线菌GY1.根据扩增出的该菌株的16SrDNA全序列在GenBank中的比较结果,结合生理生化实验、细胞化学成分及菌落形态分析,确定该菌为委内瑞拉链霉菌(Stretopmycesvenezuelae).GY1菌株以PVA为唯一碳源时,产生的PVA降解酶活性达到120UL-1,是以葡萄糖或可溶性淀粉为唯一碳源时的10倍.当在PVA培养基中添加1gL-1至3gL-1的葡萄糖时,该菌株细胞量和PVA降解酶总酶活随着葡萄糖添加量的增加而增加,最大细胞量是未添加葡萄糖时的2.4倍,最高总酶活是未添加葡萄糖时的2.6倍,而单位质量细胞产生PVA降解酶的能力提高不大.但当添加的葡萄糖浓度从3gL-1增至10gL-1时,总酶活随着细胞量的增加出现下降趋势,同时单位质量细胞产生PVA降解酶的能力也大大降低.在相同PVA聚合度下,高醇解度PVA比低醇解度PVA更有利于GY1菌株的生长及产生PVA降解酶.图5表1参16     

间苯二酚的微生物降解研究

从土壤中筛选的一株降解活性较高的细菌Ｔ１，当以间苯二酚为唯一碳源时，降解率可达９８％，研究表明，菌体接种量的大小对间苯二酚降解速度有一定的影响；降解酶为胞内酶，经鉴定，Ｔ１菌为黄单胞菌，其某些特性与已知菌种有差异，需进一步研究。     

苯甲酸类化合物的微生物降解研究

用富集培养法从工业污水中分离到5个能以苯甲酸为唯一碳源和能源而生长的细苗菌株：不动杆菌（Acinetobactersp.）BJ1,无色杆菌（Achromobactersp.）BY1,假单胞苗（Pseudomonasspp.）SJ1、SY1和SH1、测定了这5个菌株的底物特异性和抗菌素抗性其中BJ1菌株含有1个大质粒和2个小质位在最适培养条件下BJ1菌株对苯甲酸的降解率达98％以上．     

硝基苯降解菌的分离鉴定及降解特性

从化工厂污水处理池污泥中分离到一株能高效降解硝基苯的菌株XY-1,通过形态观察、生理生化特征和16SrDNA序列同源性分析,将该菌株鉴定为假单胞菌属(Pseudomonas sp.).该菌株能以硝基苯为唯一碳源、氮源和能源生长,硝基苯初始浓度为200 mg/L时,20 h降解率可达97%.该菌在温度25～35℃、pH 7.0～9.0范围内均能高效降解硝基苯,并且对对氯硝基苯、对氯苯胺也有良好的降解效果.测序分析表明,克隆到了该菌中的硝基苯还原酶基因,推测该菌的降解途径是硝基苯部分还原途径.图6参19     

乐果降解菌LGX1的筛选及其降解特性研究

通过富集培养,从连续施用农药乐果的土壤中分离得到一株具有较强降解有机磷农药乐果能力的细菌菌株LGX1,通过菌落形态观察及细菌的16SrDNA测序,对其进行了鉴定,同时初步研究了其降解性能。结果表明：该菌株为蜡状芽孢杆菌（Bacillus cereus）,在接种量为20%,乐果初始质量浓度为100mg·L-1,外加碳源葡萄糖质量分数为2%,温度为35℃,初始pH值为4时该菌株对乐果的降解能力最强;并且能在以辛硫磷、毒死蜱和三唑磷为唯一碳源的基础盐培养基中生长,且均比在以乐果为唯一碳源的基础盐培养基中生长的OD600值要高,初步推断菌株LGX1对有机磷农药的降解有一定的广谱性。     

1,2,4-三氯苯对3种海洋微藻的毒性效应

1,2,4-三氯苯(1,2种,4-TCB)是环境中普遍存在的持久性有机污染物之一,而1,2,4-TCB对藻类的毒性作用报道很少。以1,2,4-TCB处理后,3种海洋微藻(金藻Isochrysis Zhanjiangensis、角毛藻Platymonas Subcordiformis和扁藻Chaetoceros miielleri)的生长状况、蛋白质质量分数和叶绿素质量分数的改变来检测1,2,4-TCB对海洋微藻的毒性效应。结果表明:1,2,4-TCB对3种海洋微藻的生长均有一定的抑制作用,该效应表现出一定的浓度和时间依赖性;1,2,4-TCB处理4d后,3种海洋微藻细胞蛋白质质量分数和叶绿素质量分数下降,呈现一定的浓度—效应关系,这表明1,2,4-TCB对3种海洋微藻产生毒害效应,其作用机制可能与藻类光合作用功能降低和蛋白质功能受损有关。1,2,4-TCB在一定程度上降低了藻类饵料的利用价值,而且对食物链下游生物具有潜在的危害性。     

一株嗜碱产甲烷八叠球菌的分离鉴定与系统发育分析

在15℃条件下用产甲烷菌培养基对采自四川省红原县的牦牛粪进行富集培养,采用Hungate厌氧操作技术从富集培养物中分离得到一株在8~45℃范围生长、最适生长pH为8.5的嗜碱产甲烷菌T13.该菌株革兰氏染色阳性,细胞聚集体,在液体培养基中为肉眼可见的颗粒状物,在固体培养基上菌落为淡黄色桑葚状;可利用甲醇、乙酸盐和甲胺作为唯一碳源生长;对氯霉素和庆大霉素敏感;生长pH范围为6.5~9.5;最适NaCl浓度为0~0.15 mol L-1;最适生长温度为30℃.形态和生理生化特征以及16S rDNA序列分析表明菌株T13为梅氏产甲烷八叠球菌(Methanosarcina mazei).由于该菌最适生长pH为8.5,所以初步认为菌株T13是一株梅氏产甲烷八叠球菌的新菌株.     

氯代酰胺类除草剂降解菌的分离及降解性能

从生产乙草胺的农药厂废水生物处理池活性污泥中分离到一株氯代酰胺类除草剂降解细菌,命名为Y3B-1.根据表型特征、生理生化特性和16S rDNA序列系统发育分析,将其鉴定为副球菌属(Paracoccus sp.).研究了菌株Y3B-1在不同条件下对多种氯代酰胺类除草剂的降解性能.结果表明:菌株Y3B-1能以乙草胺为碳源生长,并能降解乙草胺、丁草胺和丙草胺,3 d对这3种氯代酰胺类除草剂的降解率分别达到86.7%、65.5%和69.1%,不能降解异丙甲草胺.该菌降解乙草胺的最适温度为30℃,最适pH为7.0,对乙草胺的降解效果与接种量成正相关,对较低浓度的乙草胺有很好的降解效果,过高的起始浓度抑制其对乙草胺的降解,外加营养如酵母膏和土壤悬液则显著促进其对乙草胺的降解.图7参23     

一株降解除草剂膦化麦黄桐(PPT)菌的筛选与鉴定

从喷洒有除草剂的土壤中分离到一株能分解膦化麦黄桐(PPT)的细菌.该菌在以PPT为唯一碳源的培养基上生长,能利用PPT的最高浓度为2. 7g/L.采用常规生理生化鉴定方法,并结合16SrDNA序列分析法对该菌进行鉴定.结果表明,该菌与生癌肠杆菌(Enterobactercancerogenus)序列相似性为99. 3%,在细菌分类学上属于肠杆菌.将它命名为Enterobactersp. PPT. 图4表2参7     

丁草胺高效降解细菌的分离

通过瓶培养法富集培养,从肥东县一块单晚稻田土中分离出一株丁草胺高效降解细菌ＷＹ３０６,经鉴定,该菌为节杆菌属菌Ａｒｔｈｒｏｂａｃｔｅｒ ｓｐ．ＷＹ３０６．Ａｒｔｈｒｏｂａｃｔｅｒ ｓｐ．ＷＹ３０６降解丁草胺的影响因素研究表明：丁草胺降解半衰期与初始菌量近似成反比;当丁草胺添加浓度为５、９ｍｇ／Ｌ时,其降解半衰期分别为０．９７ｈ和１．８６ｈ,随浓度的增大而增大,而当丁草胺添加浓度为０．８ｍｇ／Ｌ或其     

氯代苯及氯代酚类优先污染物好氧生物降解动力学的研究

利用测定微生物呼吸耗氧量的方法,对氯代苯及氯代酚类中的７种优先污染物,用经它们分别驯化的活性污泥对其好氧生物降解动力学进行了研究。     

有机污染物生物降解途径控制反应的预测与优势菌选择模型的建立

本文把有机污染物生物降解途径的理论测和量化剖析相结合确定生物降解途径中的控制反应,给出控制反ＱＳＢＲ的概念模式。在此基础上上建立了Ｐｓｅｕｄｏｍｏｎａｓｓｐ．典型生物降解途径以应的ＱＳＢＲ模型,利用建立的模型指导１,２－１,４－二氯苯和１,２,４－三氯苯生物处理优势菌的选择。     

Biodegradation of paclobutrazol by a microbial consortium isolated from industrially contaminated sediment

Biodegradability of the plant growth retardant paclobutrazol by a microbial consortium in which Pseudomonas was the predominant strain was investigated in batch culture. The consortium which had been isolated from an industrially contaminated sediment was proven to be useful for the treatment of effluents containing paclobutrazol. Paclobutrazol was degraded by the pure isolated strain of Pseudomonas sp. as well as the microbial consortium. Paclobutrazol was utilized as the sole source of carbon and energy. Sixty percent of the paclobutrazol was degraded by the microbial consortium from an initial concentration of 54 mg L^?1 within 48 h and more than 98% of an initial concentration of 3.4 mg L^?1 was degraded within 36 h. The optimum temperature and pH were determined to be 30°C and 7.0, respectively. A pure strain of a bacterium, isolated from the enrichment culture was identified as Pseudomonas sp. The microbial consortium was tolerant of high pH and could degrade paclobutrazol faster than the pure strain. The degradation rate of this plant growth regulator in an aerobic environment was greater than that under anaerobic conditions.