好氧同时硝化-反硝化菌的分离鉴定及系统发育分析

从土壤中分离到一株好氧同时硝化-反硝化菌,编号为SDN,该菌株革兰氏染色呈阳性,球状或杆状,菌落颜色橙红色.该菌株以硝酸钠为氮源时能进行好氧反硝化作用;以乙酸钠和硫酸铵分别为碳源和氮源能进行异养硝化作用,能以乙酰胺为唯一碳源和氮源进行生长.部分长度的16S rDNA序列分析表明,分离菌株SDN与Rhodococcusruber的16SrDNA序列具有99%相似性.并用PHYLIPS程序将该菌株与报道的相关微生物进行了系统发育分析.图4表1参13     

异养硝化-好氧反硝化菌Psracoccus pantotrophus ATCC 35512的研究进展

近年来发现了一类具有偶联异养硝化-好氧反硝化功能的细菌.Paracoccus pantotrophus ATCC 35512是其中最早发现的细菌.通过对该菌株16S rRNA序列和细胞色素c的氨基酸序列分析,确立其为一个新种Paracoccuspantotrophus,ATCC 35512被确定为模式株.该菌株的异养硝化-好氧反硝化偶联途径假说被提出,其反应途径中涉及的一系列酶和电子传递体也陆续被纯化研究.本文综述了对该菌株系统分类、形态和生理生化特征、硝化-反硝化偶联假说及氮循环酶系等的研究进展,并提出了进一步揭示异养硝化-好氧反硝化现象的研究方向.     

一种新的好氧反硝化菌筛选方法的建立及新菌株的发现

利用间歇曝气富集,氰化钾(KCN)选择培养基筛选好氧反硝化的细菌,通过形态学特征、生理生化反应及16SrDNA同源性比较对筛得菌株进行鉴定,并对其好氧反硝化相关基因napA进行扩增并测序比较.筛选到一株可以柠檬酸钠为碳源,硝酸钾为氮源,进行好氧反硝化的细菌.在溶解氧(DO)为(9.0±0.5)mg/L的培养基中,该菌株5 d内将硝态氮由282.0 mg L-1降解至149.2 mg L-1,其硝态氮去除率为46.47 ng mg-1min-1,同时亚硝态氮仅有少量的积累.经鉴定,初步判定它为假单胞菌属,命名为Pseudomonas sp.Y2-1-1.从其基因组中扩增出与好氧反硝化相关的周质硝酸盐还原酶(NAR)的亚基napA基因,并与已报道的napA基因进行Blast比较,发现具有较大差别.利用间歇曝气富集,氰化钾(KCN)选择培养基筛选好氧反硝化的细菌是非常有效的.初步认为Pseudomonas sp.Y2-1-1是一株新的好氧反硝化菌.图6表3参12     

异养硝化-好氧反硝化菌Paracoccus pantotrophus ATCC 35512的研究进展

近年来发现了一类具有偶联异养硝化-好氧反硝化功能的细菌．Paracoccus pantotrophus ATCC 35512是其中最早发现的细菌．通过对该菌株16S rRNA序列和细胞色素c的氨基酸序列分析,确立其为一个新种Paracoccus pantotrophus,ATCC 35512被确定为模式株．该菌株的异养硝化一好氧反硝化偶联途径假说被提出,其反应途径中涉及的一系列酶和电子传递体也陆续被纯化研究．本文综述了对该菌株系统分类、形态和生理生化特征、硝化-反硝化偶联假说及氮循环酶系等的研究进展,并提出了进一步揭示异养硝化-好氧反硝化现象的研究方向．表2参50     

一株碱性果胶酶高产细菌的分离、系统发育分析和产酶条件的初步优化

从腐烂的苹果表皮筛选到一株碱性果胶酶的高产菌株,命名为WSHB04-02.分离菌株为革兰氏阳性细菌,有芽孢,菌落颜色为乳白色.分离菌株WSHB04-02的16SrDNA全序列分析表明,该菌株与Bacillussubtilus具有99%的相似性,并与其他13株产碱性果胶酶菌株的16SrDNA结果进行同源性分析,并构建系统发育树.发现菌株WSHB04-02在不含果胶类物质与Mg2 的培养基上能高产碱性果胶酶,在优化的培养条件下,碱性果胶酶的酶活达到34U/mL,在国内还未见报道.图6表2参15     

1株贫营养好氧反硝化菌的分离鉴定及其脱氮特性

从水库底泥样品中,以硝酸盐为唯一氮源进行驯化、分离筛选出1株能在贫营养及好氧条件下进行高效反硝化的菌株PY8,经过电镜形态学观察、生理生化和16S rDNA序列分析,并基于16SrDNA序列结果,构建了该菌株的系统发育树,最终确定菌株PY8为根瘤菌Rhizobiumsp.。考察了初始pH值、温度、C/N、初始硝酸钠质量浓度、投菌量对菌株PY8硝酸盐还原活性的影响,以及该菌株的异养硝化性能。结果表明,在pH6.0～10.0,温度25～30℃,C/N1.0～9.0,初始硝酸钠质量浓度0.01～0.50g·L-1,投菌量1%～15%时,菌株PY8培养72h后的硝氮去除率可达到95%以上。另外,该菌株具有同时硝化-反硝化作用,在培养过程中氨氮去除率可达到58%左右。实验结果表明,菌株PY8在微污染水体生物脱氮领域中具有很大的应用潜力。     

DNA同源性分析最终确定新疆中华根瘤菌（Sinorhizobium xinjiangensis）为一独立种

针对S.xinjiangensis分类地位的争议，从新疆再次分离获得34株快生大豆根瘤菌，在16SrDNAPCR-RFLP分析和SDS全细胞蛋白电泳分群的基础上，进行了16SrDNA全序列相似性和DNA同源性分析.所测4个菌株和S.fredii模式菌株USDA205的16SrDNA全序列有很高的相似性.而DNA同源性分析说明新分离的菌株代表与原定的S.xinjiangensis为一个DNA同源群.其模式菌株CCBAU110与S.fredii的2株参比菌株USDA194、2048之间的DNA同源性分别为31.5%和28.7%.S.fredii的模式菌株USDA205与新分离的菌株代表及原定的S.xinjiangensis的模式菌株和2个参比菌株之间的DNA同源性为20.8%～39%，远低于70%.属于种水平上的差异.按照国际细菌分类委员会以DNA同源性≥70%且△Tm≤5℃作为定种的标准，S.xinjiangensis是Sinorhizobium属中不同于S.fredii的一个独立的种.表3参20     

一株聚丁二酸丁二醇酯降解菌的分离鉴定

以聚丁二酸丁二醇酯（PBS）为唯一碳源,通过滤纸片法对西安郊区土壤中可有效降解PBS聚合物的微生物进行分离,并成功获得一株降解效果最为明显的细菌菌株,其在30 d内可使PBS聚合物的数均分子量降低19.80%。采用生理生化实验以及16S rDNA序列对比分析对该菌株进行了鉴定。结果表明,该菌株的生理生化特征与铜绿假单胞菌属相似,且其16SrDNA序列与Pseudomonas aeruginosa 39016 contig 00492的核苷酸序列同源性达99%,结合生理、生化指标鉴定结果,进一步确定该菌株为铜绿假单胞菌（Pseudanonas aeruginosa）。该菌株的分离鉴定为进一步研究PBS聚合物的生物降解特性及机理等奠定了基础。     

一株贫营养异养硝化-好氧反硝化菌的筛选及脱氮特性

为了探究并优化菌剂应用于微污染水源水体修复的机制和条件,主要针对水库沉积物内筛选出的贫营养好氧反硝化菌进行了菌种鉴定及脱氮特性研究,考察菌株在不同环境条件下的脱氮效果,明确了该菌株的最适宜生长条件,并基于水库水体中贫营养条件对菌株进行水源水库原水的驯化培养试验研究,以期实现该菌株对微污染水源水库原水中氮源污染物的脱除,为原位投菌技术实际工程应用提供理论依据。从微污染水源水库沉积物中驯化筛分出一株高效异养硝化-好氧反硝化菌A14,通过扫描电镜观察、生理生化特征、16S rRNA基因测序和Biolog GenⅢ鉴定,确定该菌株为革兰氏阴性短杆菌,鉴定为皮特不动杆菌（Acinetobacter pittii）。在好氧条件下,菌株细胞内表达反硝化功能基因napA,以NO3-为唯一氮源进行反硝化作用时,36 h时NO3-去除率为78.89%。以NH4+为唯一氮源时,48 h NH4+去除率为95.25%,TN去除率达80.42%,TOC去除率达98.30%,表明该菌株具有异养硝化-好氧反硝化特性。在改变环境条件过程中,该菌株在以乙酸钠为碳源,温度为30℃,C/N为12,pH为7,接种量为10%时,NO3-去除率最高为86.62%,并且在10℃下脱氮率达到40.18%。在水源水库原水脱氮实验中,接种处理TN去除率为50.95%,NO3-去除率为80.25%。结果表明,菌株A14在微污染水源水体菌剂脱氮修复中具有良好的应用潜力。     

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     

低温贫营养好氧反硝化细菌SY13的反硝化特性

从底泥中分离出1株低温贫营养好氧反硝化细菌SY13,经常规生理生化鉴定和16SrDNA测序,鉴定出细菌SY13属于Acinetobactersp．。考察了温度、pH、C／N比及接种量对菌株SY13硝酸盐还原活性的影响,初始硝酸盐浓度为15mg／L左右,温度为15℃时低温贫营养好氧反硝化细菌SY13的硝酸盐去除率为49．26％,在中性环境适应性较强,pH值为7．0时72h的硝氮去除率达到58．08％,随着C／N比不断增加,菌株SY13硝酸盐的去除效果逐渐增强,接种量为10％时,菌株SY13培养72h后的硝氮去除率可达到59．62％。     

好氧异养硝化菌Acinetobacter sp.YY-5的分离鉴定及脱氮机理

通过异养硝化培养基获得一株高效脱氮细菌,并通过形态学特征、生理生化反应及16S rDNA同源性比较对筛得菌株进行了鉴定;分别以NO3--N和NO2--N为唯一氮源,通过对脱氮过程中各种含氮代谢物的定量及对脱氮相关基因氨单加氧酶基因(amoA)、羟胺氧化酶基因(hao)、周质硝酸盐还原酶亚基基因(napA)的扩增及测序比较,对该菌株的生理途径及脱氮机理进行了研究.结果表明,高效脱氮细菌YY-5不能发生好氧反硝化,但能在3 d内将氨氮由95.23 mg/L降解至1.29 mg/L,降解率达妻98.6%,同时未发现亚硝酸盐氮、硝酸盐氮积累;对该菌主要代谢气体产物进行检测,发现CO2和N2明显增多,无N2O生成;经鉴定,初步判定该菌为不动杆菌属,命名为Acinetobacter sp.YY-5;从该菌基因组中均能扩增出amoA、hao、napA等基因,其中napA与hao基因与已报道的napA与hao基因进行Blaster较,发现具有较大差别.图6表3参15     

集约化蔬菜种植区地下水中反硝化细菌的分离鉴定

从研究我国典型集约化蔬菜种植区地下水中硝酸盐的来源和浓度人手,进行富集、培养、分离、纯化,筛选出一株具有反硝化作用的菌株,通过形态学、革兰氏染色结合16SrDNA序列同源性分析鉴定,其鉴定结果为农杆菌(Agrobacterium sp.).该研究为开展地下水硝酸盐污染的生物修复储备宝贵的菌种资源,为地下水中硝酸盐污染的原位微生物修复和相关污水的生物处理提供微牛物基础,对于经济、有效的解决地下水硝酸盐污染和水资源短缺的问题有着十分重要的意义.     

Anaerobic degradation of 4‐chlorophenol under denitrifying conditions with nitrate as the only electron acceptor

A denitrifying bacterium was isolated from activated sludge after enrichment on 4‐chlorophenol under anoxic, denitrifying conditions, and the microbial degradation of 4‐chlorophenol by this bacterium, with nitrate as the only electron acceptor, was investigated. The bacterium was capable of degrading up to 5 mM 4‐chlorophenol with stoichiometric release of chloride under denitrifying conditions. Anaerobic degradation of 4‐chlorophenol was dependent on nitrate as an electron acceptor. Nitrate (10 mM) severely inhibited the growth by the toxic effect of the accumulated nitrite. The optimum nitrate concentration for cell growth and chlorophenol degradation was 6 mM.     

Promotive effect of pyridine on indole degradation by activated sludge under anoxic conditions

Batch experiments were carried out to investigate the promotive effect of pyridine on indole degradation under denitrifying conditions. The seed sludge was obtained from a local coal-coking wastewater treatment facility and was acclimated in the laboratory. Indole and pyridine were supplemented to the synthetic wastewater at different ratios. The optimum ratio of chemical oxygen demand (COD) to nitrate (C/N) was 8.4–8.9 for both denitrification and indole and pyridine degradation. At a temperature of 28°C and pH of 7.0–7.5, the nitrate reductase activity (NRA) was in the best state. The addition of pyridine could promote NRA and the degradation of indole. When the initial concentration of indole was 150 mg/L, the concentration ratio of indole to pyridine was in the range of 1–10. Under optimum C/N conditions, the degradation of indole could be described with pseudo-zero-order kinetics. There was no accumulation of nitrite during the reaction. When the concentration ratio of pyridine to indole was less than 0.25 with an increase in the pyridine proportion, there were more significant augment rates for NRA and the degradation of indole than the situation when the concentration ratio was more than 0.25.     

Toxic effect of sodium perfluorononyloxy-benzenesulfonate on Pseudomonas stutzeri in aerobic denitrification,cell structure and gene expression

• OBS inhibited the growth of P. stutzeri and destroyed its structure. • OBS was toxic to the aerobic denitrification process of P. stutzeri. • OBS induced the production of ROS in P. stutzeri. • OBS affected the expression of key genes involved in denitrification and SOD. The toxicities of sodium perfluorononyloxy-benzenesulfonate (OBS) to animals and plants are similar to those of perfluorooctane sulfonate. However, the mechanism of its toxicity to aerobic denitrifying bacteria is still unclear. In the present study, the ecotoxicity of OBS on an aerobic denitrifying strain, Pseudomonas stutzeri, was evaluated. The results showed that a dosage of OBS clearly affected the growth and aerobic denitrification of P. stutzeri. When compared with an unamended control, the degradation efficiency of the total nitrogen decreased by 30.13% during exposure to 200 mg/L of OBS, and the complete degradation time of nitrate-nitrogen was delayed by 4 h. The lactate dehydrogenase and superoxide dismutase produced by the bacteria increased with the concentration of OBS, and reactive oxygen species were also detected by confocal laser scanning microscope imaging. Transmission electron microscope imaging revealed chromosome deformation of the cells and damage to cell content; moreover, outer membrane vesicles were secreted from the bacteria, which was the important detoxification mechanism of P. stutzeri to OBS. Expression of the genes involved in aerobic nitrification and oxidative stress were also changed under OBS stress, which further confirmed the toxicity of OBS to P. stutzeri. This study reveals the environmental exposure risk of OBS from the perspective of microorganisms.     

A/O-MBR处理某经济区市政污水的效能

采用A/O-MBR法处理某经济区市政污水,对其进、出水的水质进行连续监测,考察系统稳定性和处理效果。结果表明,在水力停留时间12小时,污泥龄30天,回流比75%,好氧段DO浓度1.3～8.5 mg/L的条件下,系统对COD、氨氮、总氮、总磷的平均去除率分别为92.04%、93.66%、78.04%、91.06%。反硝化...     

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。