一株高效絮凝剂产生菌的分离鉴定与絮凝特性研究

采用富集培养及平板分离方法从成都某污水处理厂活性污泥中筛选出一株具有絮凝活性的菌株SNUX-1,经形态特征及16S rRNA鉴定该菌株为约氏不动杆菌(Acinetobacter johnsonii).对该菌株絮凝活性分布及在不同生长条件下所产絮凝剂的絮凝率进行了研究,并对其絮凝剂成分进行了分析.结果表明,该菌株所产微生物絮凝剂为胞外产物,显色剂显色结果表明,该絮凝剂不含蛋白质成分,含有糖类成分;在以蔗糖为碳源、培养基初始pH值为8、以氯化铜为助凝剂的条件下培养12 h,该菌株所产微生物絮凝剂的絮凝率最高,达96.71％,可作为治理环境污染的菌种资源.     

一种新型抗性质粒的构建及其在核黄素生产菌中的运用

枯草芽孢杆菌Bacillus subtilis RF1是课题组前期通过基因工程改造得到的一株核黄素高产菌,为了进一步提高核黄素的产量,需要对该菌株进行进一步的基因工程改造.本研究首先将编码的链丝菌素乙酰基转移酶基因sat克隆到p MA5质粒上,构建具有诺瓦丝菌素Norseothricin(NTC)抗性的重组质粒p MA5-sat,实验证实该重组质粒能够用于B.subtilis RF1的抗性筛选.随后将核黄素合成相关的关键酶葡萄糖-6-磷酸脱氢酶编码基因zwf克隆到重组质粒p MA5-sat上,获得重组质粒p MA5-sat-zwf,并成功构建重组菌B.subtilis RF1/p MA5-sat-zwf.结果显示,重组菌胞内葡萄糖-6-磷酸脱氢酶活力比原始菌提高了近50倍,说明葡萄糖-6-磷酸脱氢酶在重组菌中成功过量表达;根据发酵特性分析,重组菌B.subtilis RF1/p MA5-sat-zwf最终核黄素产量达到12.01 g/L,比原始菌B.subtilis RF1提高了30.3%.综上,本研究构建的新型抗性质粒能够成功运用于核黄素生产菌枯草芽孢杆菌的基因工程改造.     

Cytochrome P450 BM3 of Bacillus megaterium—A possible endosulfan biotransforming gene

Computing chemistry was applied to understand biotransforrnation mechanism of an organochlorine pesticide, endosulfan. The stereo specific metabolic activity of human CYP-2B6 （cytochrome P450） on endosulfan has been well demonstrated. Sequence and structural similarity search revealed that the bacterium Bacillus megaterium encodes CYP-BM3, which is similar to CYP-2B6. The functional similarity was studied at organism level by batch-scale studies and it was proved that B. megaterium could metabolize endosulfan to endosulfan sulfate, as CYP-2B6 does in human system. The gene expression analyses also confirmed the possible role of CYP-BM3 in endosulfan metabolism. Thus, our results show that the protein structure based in-silico approach can help us to understand and identify microbes for remediation strategy development. To the best of our knowledge this is the first report which has extrapolated the bacterial gene for endosulfan biotransformation through in silico prediction approach for metabolic gene identification.     

一株羽毛角蛋白降解菌的分离鉴定及特性研究

采用富集培养的方法,从广西大学家禽养殖场废弃羽毛处土样中筛选获得一株高效降解羽毛角蛋白的菌株GZD-23。经形态学、生理生化特性和16S rDNA序列分析,鉴定该菌株为枯草芽孢杆菌(Bacillus subtilis)。环境因素影响和酶学特性实验结果表明,蔗糖与蛋白胨是菌株GZD-23降解羽毛的最佳外加碳氮源,pH 8.0、温度30℃、转速100 r/min和羽毛含量20 g/L为其降解羽毛的最适反应条件。菌株GZD-23合成的角蛋白酶最适反应温度为60℃,最适pH为7.5;金属离子Cu2+、Hg2+、Fe2+、Co2+、Zn2+、Mn2+以及SDS、EDTA和高浓度TritonX-100、异丙醇和β-巯基乙醇对其酶活有不同程度抑制作用;低浓度TritonX-100、异丙醇和β-巯基乙醇对其酶活具有一定促进作用。     

As(Ⅲ)和As(Ⅴ)胁迫对嗜酸性氧化亚铁硫杆菌铁行为的影响

以嗜酸性氧化亚铁硫杆菌为受体材料,通过分析培养过程中培养液pH、Fe2+氧化率和总铁(TFe)沉淀率研究不同浓度As(Ⅲ)和As(Ⅴ)胁迫对嗜酸性氧化亚铁硫杆菌铁行为的影响。结果表明:As(Ⅲ)胁迫显著降低反应体系的pH降幅、Fe2+氧化速率和TFe沉淀率;低浓度As(Ⅴ)的加入对反应体系的pH降幅、Fe2+氧化速率和TFe沉淀率没有明显影响;随着初始浓度的增大,pH降幅、Fe2+氧化速率和TFe沉淀率也逐渐降低。As(Ⅲ)对嗜酸性氧化亚铁硫杆菌铁行为的影响较As(V)更明显。同一胁迫水平下,该菌对As(Ⅲ)的响应更强烈。     

镁离子对氧化亚铁硫杆菌生物合成次生铁矿物的影响

通过摇瓶实验,在Mg2+分别为48,4.8mg/L,其他元素组成与9K液体培养基一致的体系中,采用氧化亚铁硫杆菌A.ferrooxidans催化合成次生铁矿物.考察了Mg2+含量对生物合成次生铁矿物体系pH值、氧化还原电位(ORP)、Fe2+氧化率、总Fe沉淀率、次生铁矿物矿相及矿物晶体尺寸的影响.结果表明,经过48h培养,Mg2+浓度为48,4.8mg/L生物成矿体系pH值分别从原来的2.50降低至2.30,2.19,ORP分别从初始259mV增加至269mV,276mV.两体系Fe2+氧化率培养至第48h均达到100%,然而两体系总Fe沉淀率及矿物形态及却不尽相同.Mg2+浓度为48mg/L生物成矿体系,总Fe沉淀率为23.7%,次生矿物紧密粘附于三角瓶底部.而Mg2+浓度为4.8mg/L生物成矿体系,总Fe沉淀率达到32.2%,次生矿物却均匀分散于溶液中.两体系合成次生铁矿物均为黄铁矾与施氏矿物共存的混合物,Mg2+含量4.8mg/L体系合成黄铁矾单个晶体长度(~1.60μm)约为Mg2+含量48mg/L体系的1.2倍.     

Isolation, identification and arsenic-resistance of Acidithiobacillus ferrooxidans HX3 producing schwertmannite

Schwertmannite, a ubiquitous mineral present in iron oxyhydroxides formed in iron- and sulfate-rich acid media, favors incorporation of some toxic anions in its structure. We reported an iron-oxidizing bacterial strain HX3 from a municipal sludge that facilitates the formation of pure schwertmannite in cultures. Ferrous iron oxidation by the isolated strain HX3 was optimum at an initial pH of 2.0-3.3 and temperature of 28-35°C. Pure schwertmannite was found through bacterial oxidation of ferrous iron at an initial pH 2.8and temperature 28°C. Following 16 S rDNA gene sequence analysis the bacterial strain HX3 was identified as Acidithiobacillus ferrooxidans. The arsenic-resistance A. ferrooxidans HX3 showed the potential of environmental application in arsenic removal from the As（Ⅲ）- and iron-rich acid sulfate waters directly by As（Ⅲ） adsorption or the formation of schwertmannite in the environment.     

Characterization of a salt-tolerant bacterium Bacillus sp. from a membrane bioreactor for saline wastewater treatment

High salt concentrations can cause plasmolysis and loss of activity of cells, but the salt-torlerant bacterium can endure the high salt concentrations in wastewater. In this research 7 salt-torlerant bacteria, which could survive in dry powder products and could degrade organic contaminants in saline wastewater, were isolated from a membrane bioreactor. The strain NY6 which showed the fastest growth rate, best property for organic matter degradation and could survive in dry powder more than 3 months was selected and characterized. It was classified as Bacillus aerius based on the analysis of the morphological and physiological properties as well as the 16 S rRNA sequence and Neigh borjoining tree. The strain NY6 could survive in the salinity up to 6% and the optimal growth salinity is2%; it belongs to a slightly halophilic bacterium. The capability of its dry powder products for COD removal was 800 mg COD/(g·day) in synthesized saline wastewater with salinity of 2%. According to salt-tolerant mechanism research, when the salinity was below 2%, the stain NY6 absorbed K+and Na+to maintain osmotic equilibrium, and when the salinity was above 2%, the NY6 kept its life by producing a large amount of spores.     

Evaluation of Bacillus sp. MZS10 for decolorizing Azure B dye and its decolorization mechanism

To evaluate decolorization and detoxification of Azure B dye by a newly isolated Bacillus sp. MZS10 strain, the cultivation medium and decolorization mechanism of the isolate were investigated. The decolorization was discovered to be dependent on cell density of the isolate and reached 93.55%(0.04 g/L) after 14 hr of cultivation in a 5 L stirred-tank fermenter at 2.0 g/L yeast extract and 6.0 g/L soluble starch and a small amount of mineral salts. The decolorization metabolites were identified with ultra performance liquid chromatography-tandem mass spectroscopy(UPLC-MS). A mechanism for decolorization of Azure B was proposed as follows: the C=N in Azure B was initially reduced to –NH by nicotinamide adenine dinucleotide phosphate(NADPH)-dependent quinone dehydrogenase, and then the –NH further combined with –OH derived from glucose to form a stable and colorless compound through a dehydration reaction. The phytotoxicity was evaluated for both Azure B and its related derivatives produced by Bacillus sp. MZS10 decolorization, indicating that the decolorization metabolites were less toxic than original dye. The decolorization efficiency and mechanism shown by Bacillus sp. MZS10 provided insight on its potential application for the bioremediation of the dye Azure B.     

两种生物农药对土壤蔗糖酶活性的影响

采集海南省儋州市国家热带农业高新技术示范园土样中农田土样,在分析土壤基本理化性质的基础上,用3,5-二硝基水杨酸比色法测定法.研究了不同模拟条件下苏云金杆菌、井冈霉素A对土壤蔗糖酶活性的影响.结果表明:不同浓度、pH条件下这两种生物农药对土壤蔗糖酶表现出不同程度的激活作用.在30 d的培养时间内,苏云金杆菌(bacillus thuringiensis)对土壤蔗糖酶活性影响的趋势是激活→抑制→恢复,而且波动范围较大;井岗霉素A(jinggangmycin A)对土壤蔗糖酶活性影响程度相对较小,总的趋势是激活→抑制.总体来看,施用这两种生物农药有利于提高土壤蔗糖酶的活性.