寒区水体中溶藻铜绿假单胞菌的分离和性质研究

我国北方,尤其是东北地区的一些水体由于富营养化导致的水华现象呈增加趋势,成为急需解决的水环境问题.溶藻菌可从生态调控的角度来控制水华的爆发,因此,在富营养化水体治理方面逐渐得到重视.本文从寒区水华爆发水体中分离获得了一株溶藻细菌JM1,系统发育分析表明,菌株JM1与铜绿假单胞菌的同源性最高.VITEK系统鉴定表明,菌株JM1与铜绿假单胞菌的相似度为98%.菌株JM1在不同的碳源和氮源培养基中可产生不同颜色和p H值的代谢产物,而以葡萄糖和蛋白胨分别为碳源和氮源时发酵液的溶藻效果明显好于其它培养基.菌株JM1的溶藻效果随藻细胞初始密度的降低而增强,随发酵液添加量的增加而增加;当菌体细胞的密度达到1011CFU·m L-1时,可单一抑制藻细胞的生长;当菌体细胞的密度大于105CFU·m L-1时,菌体细胞可促进发酵液的溶藻作用.发酵液作用于藻细胞6 h时可明显影响其恢复能力,当暴露时间达18 h时,藻细胞彻底丧失了恢复能力.菌株JM1发酵液在中性和偏碱性的藻液中的溶藻效果最强,并且经过酸化、碱化和高温处理后,仍然保持较高的溶藻活性.本研究可为基于微生物作用过程的水华控制提供优势的土著菌种及其应用上的技术支持.     

Microcystis aeruginosa/Pseudomonas pseudoalcaligenes interaction effects on off-flavors in algae/bacteria co-culture system under different temperatures

We conducted an experiment to study the interaction effects of Microcystis aeruginosa and Pseudomonas pseudoalcaligenes on off-flavors in an algae/bacteria co-culture system at three temperatures (24, 28 and 32°C). Gas chromatography–mass spectrometry was applied to measure off-flavor compounds dimethyl sulfide (DMS), dimethyl trisulfide (DMTS), 2-methylisoborneol, geosmin (GEO) and β-cyclocitral. During the lag phase of co-cultured M. aeruginosa (first 15 days), P. pseudoalcaligenes significantly increased the production of DMS, DMTS and β-cyclocitral at all three temperatures. In the exponential phase of co-cultured M. aeruginosa (after 15 days), M. aeruginosa became the main factor on off-flavors in the co-culture system, and β-cyclocitral turned to the highest off-flavor compound. These results also indicated that DMS, DMTS and β-cyclocitral were the main off-flavor compounds in our M. aeruginosa/P. pseudoalcaligenes co-culture system. Univariate analysis was applied to investigate the effects of M. aeruginosa and P. pseudoalcaligenes on the production of off-flavors. The results demonstrated that both M. aeruginosa and P. pseudoalcaligenes could increase the production of DMS and DMTS, while β-cyclocitral was mainly determined by M. aeruginosa. Our results also provide some insights into understanding the relationship between cyanobacteria and heterotrophic bacteria.     

Microcystis aeruginosa/Pseudomonas pseudoalcaligenes interaction effects on off-flavors in algae/bacteria co-culture system under different temperatures

We conducted an experiment to study the interaction effects of Microcystis aeruginosa and Pseudomonas pseudoalcaligenes on off-flavors in an algae/bacteria co-culture system at three temperatures(24, 28 and 32℃). Gas chromatography–mass spectrometry was applied to measure off-flavor compounds dimethyl sulfide(DMS), dimethyl trisulfide(DMTS),2-methylisoborneol, geosmin(GEO) and β-cyclocitral. During the lag phase of co-cultured M. aeruginosa(first 15 days), P. pseudoalcaligenes significantly increased the production of DMS, DMTS and β-cyclocitral at all three temperatures. In the exponential phase of co-cultured M. aeruginosa(after 15 days), M. aeruginosa became the main factor on off-flavors in the co-culture system, and β-cyclocitral turned to the highest off-flavor compound. These results also indicated that DMS, DMTS and β-cyclocitral were the main off-flavor compounds in our M. aeruginosa/P. pseudoalcaligenes co-culture system. Univariate analysis was applied to investigate the effects of M. aeruginosa and P. pseudoalcaligenes on the production of off-flavors. The results demonstrated that both M. aeruginosa and P. pseudoalcaligenes could increase the production of DMS and DMTS, while β-cyclocitral was mainly determined by M. aeruginosa. Our results also provide some insights into understanding the relationship between cyanobacteria and heterotrophic bacteria.     

嗜油菌NY3对蜡质油污泥无害化处理

蜡质油污泥(WOS)成分复杂,含大量老化原油、蜡质油、沥青质及其他化学药剂,属危险性固体废弃物,大量排放已成为石化企业可持续发展的障碍.研究利用铜绿假单胞菌NY3处理高浓度WOS的条件,结果表明,WOS∶木屑(m/m)(d =0.30 mm) 7∶1并结合热熔分散为最佳油污泥预处理条件;液相降解体系含油量为24.5 g/L时,6d内,油泥中C20以下的正构烷烃完全被去除,C21～C34和C35～C38大分子量正构烷烃去除率分别在91％ ～99％和78％ ～ 89％之间.添加鼠李糖脂能明显提高长链烃和多环芳烃的降解率.6d内210 mg/L鼠李糖脂使C29 ～ C38的降解率提高17.27％ ～36.65％,芘、菲分别提高13.67％和16.12％.添加葡萄糖和Fe2+的条件下,NY3菌可将油污泥转化成为糖蛋白,产量达28 g/L.     

Pseudomonas sp.fz胞外分离物降解四溴双酚A

四溴双酚A(TBBPA)是全球消耗量最大的溴系阻燃剂。通过活性物质的定位、分离和TBBPA产物的分析等步骤,对Pseudomonas sp.fz胞外分离物降解TBBPA进行了研究。结果表明,存在于胞外的活性物质通过异丙基断裂和脱溴两条途径降解TBBPA。通过超滤,Sephadex G-10分离纯化得到了具有降解活性的小分子物质,分子量约为376~456 Da,初步鉴定为短肽类物质。这种小分子物质具有很好的热稳定性(30~80℃),在弱酸性条件下(2.0~5.0)活性较高,其活性还受金属离子、氧气和抑制剂(Na N3)的影响。此外,部分纯化的小分子物质在一定条件下能够产生羟基自由基(·OH)。     

恶臭假单胞菌降解苯酚的动力学研究

研究了恶臭假单胞菌（Pseudomonas putida LY1）利用苯酚作为唯一碳源和能源时的生长动力学特性。通过对该菌株在不同温度、pH和初始苯酚浓度下的生长和降解情况的研究,可知该菌株在温度为25℃左右、pH为中性和偏碱性的条件下具有更高的降解活性。细菌的生长动力学过程符合基质抑制型的Haldane方程,可利用该...     

聚合物驱采出水中聚丙烯酰胺的微生物联合降解作用研究

通过对2株细菌的培养降解实验研究聚丙烯酰胺(hydrolyzed polyacrylamide,HPAM)降解菌对水环境下聚丙烯酰胺的降解作用,讨论协同降解机理。2株降解聚丙烯酰胺的菌株假单胞菌CJ419、枯草芽孢杆菌FA16在初始30℃废水样品上培养,定期测量细菌生物量和HPAM降解率。培养30 d后CJ419和FA16对聚合物的降解率最大值分别达到30.4%和25%,而以1∶1比例的混合菌降解率最大值达到80.3%。对2株菌胞外各组分研究表明:混合菌降解HPAM的机理主要由胞外降解酶系水解聚合物侧链基团导致HPAM降解为小分子物质,同时生长过程中降解菌还会释放非蛋白还原性物质引发氧化反应共同参与HPAM降解。     

铜绿假单胞菌对铜和铅的吸附

研究了铜绿假单胞菌(Pseudomonas aeruginosa)对Cu和Pb的吸附特性.结果表明,相同条件下,该菌株对Cu~(2+)的吸附率低于Pb~(2+).对于单一重金属体系,吸附率均随时间的延长先上升后平稳变化,2 h达到稳定.吸附率随投菌量的增加先迅速增加,之后趋于平稳.对于Cu~(2+),投菌量为1 g·L~(-1)时吸附率达到稳定,而Pb~(2+)的吸附效果达到平稳时的投菌量为0.5 g·L~(-1).单位质量菌体对Cu~(2+)、Pb~(2+)的吸附量随投菌量的增加而下降.pH为3时,菌体的吸附效果较差,当pH为5~8时,2种重金属的吸附效果较高.对于活菌,Pb~(2+)对菌体吸附Cu~(2+)有抑制作用,而Cu~(2+)对菌体吸附Pb~(2+)的影响无明显规律.对于失活菌,P.aeruginosa吸附Pb~(2+)和Cu~(2+)的效果均随共存重金属浓度的增大而降低,但Cu~(2+)对Pb~(2+)的影响比Pb~(2+)对Cu~(2+)的影响更显著.扫描电镜观察发现,吸附后的菌体较吸附前聚集性更好.总体而言,P.aeruginosa能对水体中共存的Cu~(2+)和Pb~(2+)有较好的吸附效果.     

Pseudomonas sp.L19抗除草剂AHAS基因的克隆、表达及特性

乙酰羟酸合酶(AHAS)是磺酰脲类、咪唑啉酮类、三唑嘧啶类、磺酰胺类和嘧啶水杨酸类等乙酰羟酸合酶抑制剂类除草剂的作用靶标,获得能抗此类除草剂的AHAS突变基因资源具有非常重要的理论和应用价值.从长期使用甲磺隆的农田土壤中分离到1株对乙酰羟酸合酶抑制剂类除草剂有广谱抗性的菌株L19,根据表型特征、生理生化特性和16S rDNA序列系统发育分析,将其鉴定为假单孢菌属(Pseudomonas sp.).利用PCR技术从Pseudomonas sp.L19的总DNA中克隆AHAS基因,氨基酸序列比对结果表明,L19与对除草剂敏感菌株P.putida KT2440的AHAS调节亚基氨基酸序列完全相同,而催化亚基有4个氨基酸位点不同:[Val 29→Ala(L19→KT2440),Pro93→Ser,Val 345→Ala,Pro 484→Arg].分别将菌株L19与KT2440的AHAS催化亚基克隆到pET29a(+)的多克隆位点,构建了表达载体pET-L19AHASc和pET-KT2440AHASc,通过镍柱亲和层析纯化得到带有组氨酸标签的乙酰羟酸合酶.抗性试验结果表明菌株L19的乙酰羟酸合酶对四大类乙酰羟酸合酶抑制剂类除草剂的抗性均要强于KT2440的乙酰羟酸合酶,对甲磺隆、咪唑乙烟酸、唑嘧磺草胺和嘧草醚的抗性倍数分别达到53.6、9.3、8.2和9.5倍.菌株L19的乙酰羟酸合酶比活力、对ThDP和Mg2+的Kc值相应地比KT2440乙酰羟酸合酶的要低;而对底物丙酮酸钠的米氏常数Km值要比KT2440乙酰羟酸合酶的要高近1倍.     

Effects of methamidophos on the community structure,antagonism towards Rhizoctonia solani,and phlD diversity of soil Pseudomonas

A microcosm incubation study using an aquic brown soil from northeast China (a Cambisol in the UN Food and Agriculture Organization FAO Soil Taxonomy) was conducted to examine the effects of different concentrations (0, 50, 150, and 250 mg kg^?1) of methamidophos (O,S-dimethyl phosphoramidothioato) on Pseudomonas, one of the most important gram-negative bacteria in soil. Amplified ribosomal DNA restriction analysis (ARDRA) was performed to study the Pseudomonas community structure, an in vitro assay was made to test the antagonistic activity of isolated Pseudomonas strains against soil-borne Rhizoctonia solani, a major member of the pathogens highly related to soil-borne plant diseases, and special primer amplification and sequencing were performed to investigate the diversity of phlD, an essential gene in the biosynthesis of 2, 4-diacetylphloroglucinol (2, 4-DAPG), which has biocontrol activity in phlD ⁺isolates. With exposure to increasing methamidophos concentrations, the total number of soil Pseudomonas ARDRA patterns decreased significantly, but with less change in the same treatments over 1, 3, and 5 weeks of incubation. The number of isolated Pseudomonas strains with antagonistic activity against R. solani as well as the diversity and appearance frequency of the strains' phlD gene also decreased with increasing concentrations of methamidophos, especially at high methamidophos concentrations. Applying methamidophos could increase the risk of soil-borne plant diseases by decreasing the diversity of the soil Pseudomonas community and the amount of R. solani antagonists, particularly those with the phlD gene.