糖脂类生物表面活性剂的特性及其对白腐真菌降解蒽的强化作用

【目的】对铜绿假单胞菌(Pseudomonas aeruginosa)所产生物表面活性剂的稳定性进行分析,考察该生物表面活性剂对乳白耙齿菌F17(Irpex lacteus F17)降解蒽的强化作用。【方法】采用三氯甲烷萃取的方法从铜绿假单胞菌的发酵液中提取生物表面活性剂,采用表/界面张力仪测定该生物表面活性剂在不同条件下的表面张力值,对其进行稳定性研究。在乳白耙齿菌F17降解蒽的过程中加入适量的生物表面活性剂,测定蒽的降解率,探讨其对蒽生物降解的强化作用。【结果】铜绿假单胞菌所产生物表面活性剂的临界胶束浓度为40 mg/L,在15-150°C及pH 6.0-13.0范围内表现出优良的稳定性,对盐浓度的耐受性也很高。在蒽的生物降解过程中,生物表面活性剂能极大地促进蒽的降解,在生物表面活性剂浓度为50 mg/L时,第15天蒽的降解率达到了82.9%。生物表面活性剂在接种乳白耙齿菌F17前1天加入培养基中,能更好地促进蒽的降解。与化学表面活性剂相比,生物表面活性剂对蒽降解的强化作用更显著。【结论】该生物表面活性剂性能优良、稳定性好,能够显著强化乳白耙齿菌F17对蒽的降解,具有良好的应用前景。     

乳白耙齿菌F17对单一和复合多环芳烃的降解差异解析

[目的] 针对菲、蒽、荧蒽多环芳烃（PAHs）污染物,利用乳白耙齿菌F17,研究单一和复合PAHs污染物的生物降解规律。[方法] 采用气相色谱-质谱法（GC-MS）分析降解过程中PAHs的浓度,并采用准一级反应动力学模型对降解结果进行拟合。[结果] 对于单一PAHs,第15天时菲、蒽、荧蒽的降解率由高到低依次为菲（97.8%） > 蒽（89.3%） > 荧蒽（81.5%）。菲、蒽和荧蒽的降解过程具有准一级反应动力学特征,菲的生物降解速率最快,其次是蒽,荧蒽的降解速率最慢。与单一PAHs的降解相比,在复合PAHs的降解过程中,乳白耙齿菌F17的生长和锰过氧化物酶的合成均表现出不同的特征。此外,水溶性极可能是复合污染物降解的重要控制因子,三者水溶性为：菲 > 荧蒽 > 蒽。因此,在菲或荧蒽加入条件下,微生物能优先降解这些污染物,抑制了污染物蒽的降解;同时,蒽或菲的存在对荧蒽的降解也有抑制作用;然而外源加入水溶性较差的蒽和荧蒽,则对菲的生物降解无显著影响。[结论] 复合PAHs的生物降解主要表现为相互竞争的特点,通过GC-MS分析了PAHs的生物降解途径。     

一株蒽降解细菌的分离及降解特性研究

【目的】从盐碱土壤中筛选蒽降解菌株并分析其降解特性。【方法】采用极度稀释结果流式细胞检测法筛选分离纯化菌株,通过16S rRNA基因序列分析对菌株进行初步鉴定,采用气质联用仪(GC-MS)分析蒽的降解特性。【结果】从盐碱土壤中筛选出一株高效蒽降解菌株。经过16S rRNA基因序列分析,鉴定该菌株为Demequina salsinemorus BJ1。菌株可以利用蒽作为唯一碳源生长,降解率可达92%。在一定浓度范围内,随着蒽浓度的降低,细菌生长速率变快,降解率升高。添加外加碳源后,细菌生长速率明显变快,而对蒽降解率变低。对萃取中间代谢产物的质谱分析表明,降解蒽的中间代谢产物主要有9,10-anthracenedione (9,10-蒽醌)和Phthalic acid (邻苯二甲酸)等,说明它可能通过邻苯二甲酸途径降解蒽。【结论】筛选得到一株新的耐盐碱蒽降解菌,该菌降解效率高,对修复石油污染的土壤有一定的现实意义。     

乳白耙齿菌F17好氧降解四溴双酚A的特性及机理研究

[目的] 为了探究乳白耙齿菌F17（Irpex lacteus F17）降解溴代阻燃剂的可能性,研究了该菌好氧降解四溴双酚A （Tetrabromobisphenol A,TBBPA）的特性以及影响降解的因素,并结合降解产物的分析,推测其降解途径。[方法] 采用高效液相色谱法测定TBBPA的浓度,并通过气相色谱-质谱联用仪分析降解过程的中间产物。[结果] I.lacteus F17可以通过共代谢的方式好氧降解TBBPA,最适共代谢基质是葡萄糖。在葡萄糖浓度为8 g/L、菌悬液接种量为5%、pH 5.0的优化条件下,当TBBPA初始浓度为20 mg/L时降解率可达85.5%,脱溴率为14.6%。对降解过程中锰过氧化物酶的研究发现TBBPA的降解率受到该酶活性的影响。通过气相色谱-质谱联用仪检测到7种中间产物。[结论] I.lacteus F17可以有效降解四溴双酚A,其降解机理主要包括脱溴、β-断裂、羟基化、去质子和氧化等过程。     

施氏假单胞菌YC-YH1的萘降解特性及产物分析

【目的】萘是一种重要的环境污染物,它在环境中的积累会对人类健康造成危害,生物降解是解决这一问题的有效方法。本实验室保存的施氏假单胞菌YC-YH1对萘具有较强的降解能力,在此基础上,研究和分析菌株对萘的降解特性、环境因素对萘降解率的影响以及代谢产物。【方法】本文首先采用单因素实验法研究pH、温度、接种量、萘初始浓度对萘降解率的影响;并在单因素实验结果的基础上,利用Design-Expert 8.0.5软件和Box-Behnken设计对pH、温度、接种量3个影响因素进行响应面优化分析,建立环境因素对萘降解率影响的优化模型。利用LC-MS检测萘降解过程中产生的重要代谢产物,从而推测菌株对萘的代谢途径。【结果】响应面分析结果表明,优化模型极显著(P<0.001),拟合度良好,预测结果可信度高。降解实验证明,在培养温度为32.4 °C、pH为7.10、接种量5.74% (体积比)的优化条件下培养3 d即可将浓度为100 mg/L的萘100%降解。LC-MS分析表明,菌株降解萘的过程中,能够被检测到的主要代谢产物有1,2-二羟基萘、水杨酸、邻苯二酚等。【结论】施氏假单胞菌YC-YH1对萘有高的降解效率,pH、温度、接种量3个因素对菌株的降解率有较大影响。利用响应面法优化菌株对萘的降解条件,能够提高YC-YH1菌株对萘的生物降解性能。初步推测菌株YC-YH1对萘的降解是通过水杨酸途径,萘首先被其代谢为1,2-二羟基萘,而后被转化为水杨酸和邻苯二酚,最后进入三羧酸循环被彻底降解。     

厌氧丁草胺降解菌BAD-20的分离鉴定及降解特性研究

【目的】分离并鉴定能够降解除草剂丁草胺的厌氧微生物菌株,研究其厌氧降解丁草胺的特性和代谢途径,为深入研究丁草胺厌氧降解机制提供依据。【方法】以丁草胺为碳源作为选择压力从水稻田土壤中富集驯化丁草胺降解菌,利用16S rRNA基因系统发育分析结合菌株培养特征对降解菌株进行初步鉴定,利用液相色谱-时间飞行质谱(LC-TOF-MS)检测菌株降解丁草胺的代谢产物。【结果】筛选到一株降解丁草胺的厌氧细菌,命名为BAD-20,初步鉴定为嗜蛋白质菌属(Proteiniphilum),菌株BAD-20降解丁草胺的最适条件为温度30–35℃、pH 7.5–8.0和0–0.5%NaCl,在有氧条件下该菌不能降解丁草胺。最适条件下,菌株BAD-20在10d降解90%的20mg/L丁草胺。菌株BAD-20还能降解甲草胺、乙草胺、丙草胺,降解效率从高到低依次为甲草胺乙草胺丙草胺丁草胺,对这些氯乙酰胺除草剂的降解动力学符合一级动力学方程。鉴定到2个丁草胺降解代谢产物,分别是N-(2,6-二乙基苯基)-N-(丁氧甲基)乙酰胺(DEPBMA)和N-(2,6-二乙基苯基)乙酰胺(DEPA),表明菌株BAD-20降解丁草胺的起始步骤为脱氯,随后脱去N-丁氧甲基。【结论】本研究富集分离到一株降解丁草胺的厌氧细菌嗜蛋白质菌属(Proteiniphilum) BAD-20,为深入研究丁草胺厌氧降解机制及研发含丁草胺废水厌氧生物处理技术提供依据。     

一株耐盐菌Halomonas sp. A20的分离及降解糖精钠废水的特性

【背景】糖精钠废水是一种难处理的高盐有机工业废水。【目的】为了提高糖精钠废水的生物降解效果,需要研究糖精钠废水降解菌的特性。【方法】采用纯培养技术从处理糖精钠废水的多级生物接触氧化系统内的活性污泥中分离筛选糖精钠废水降解菌,对分离菌株的形态特征、生理生化特性和16S rRNA基因序列进行分析,利用单因素实验和响应面法考察分离菌株降解糖精钠废水的最佳条件。【结果】筛选获得一株糖精钠废水降解菌A20,归属于盐单胞菌属(Halomonas),当糖精钠废水的盐分为5%,菌接种量为15%,pH值为8.0,温度为30°C时,菌株A20对糖精钠废水中的化学需氧量(chemical oxygen demand,CODcr)去除率在60%以上;通过响应面法优化,菌株A20降解糖精钠废水的最佳条件为：pH 8.0,温度为30.3°C,接种量为14.1%,其CODcr去除率为65.4%。【结论】分离到一株能高效降解糖精钠废水中有机物的耐盐菌Halomonas sp. A20,可为高盐、高浓度糖精钠废水的处理提供优良的微生物菌种资源。     

一株尼古丁降解菌株Arthrobacter sp.D4的分离鉴定及其降解代谢途径

【背景】烟草在生产和加工中会产生高浓度的尼古丁废弃物,对环境造成较大的污染。【目的】筛选降解尼古丁的微生物菌种并解析其降解尼古丁的代谢途径,理解微生物如何降解尼古丁。【方法】用常规分离筛选方法、结合形态学观察和分子鉴定手段分离和鉴定菌株类别,进而利用单因素试验方法,通过设置不同的尼古丁浓度、温度和pH确定菌株降解尼古丁的最适发酵条件和降解率,利用气相色谱-质谱联用技术检测菌株在尼古丁降解过程中的主要代谢产物。【结果】获得一株以尼古丁为唯一碳源和氮源的节杆菌属(Arthrobacter)菌株,编号为D4;该菌株降解尼古丁的最适温度和pH分别为30.0℃和7.0;在1 g/L的尼古丁浓度下具备较快的尼古丁降解速率,培养18 h时尼古丁降解率可达到90%以上;尼古丁浓度≥4 g/L时菌株生长受到明显抑制;与目前报道的节杆菌属降解途径不同,该菌株降解尼古丁过程中产生了新的终产物N-甲基吡咯烷酮、可替宁及中间产物麦斯明。【结论】本研究分离鉴定到一株具有较快尼古丁降解速率的节杆菌,该菌株很可能存在新的尼古丁降解途径。     

球形红细菌厌氧降解2,4-二硝基甲苯

【目的】研究不同环境条件对2,4-二硝基甲苯(2,4-DNT)生物降解的影响。【方法】采用光合细菌球形红细菌在温度为30 °C的光照培养箱中厌氧降解2,4-DNT,并用高效液相色谱仪测定其浓度。【结果】去除2,4-DNT的最佳条件是初始浓度40 mg/L、初始pH 7.0和接种量15%。另外,2,4-DNT在菌体延滞期被细胞吸收,然后在指数期作为碳源被降解。2,4-DNT的去除率在72 h达到98.8%。从液相色谱图中观察到有2种中间代谢产物,但在120 h内产物被逐渐降解。2,4-DNT的去除动力学符合一级速率模型。【结论】不同条件下2,4-DNT的去除率表明球形红细菌能有效降解2,4-DNT。     

炔草酯降解菌Sphingopyxis sp. WP68的分离鉴定与降解特性

【背景】炔草酯可以高效防除麦田恶性杂草,但炔草酯的生产和使用也对环境造成了破坏,对动物和人类健康造成了威胁。【目的】分离筛选炔草酯高效降解菌株,研究其降解特性,为炔草酯污染生物修复提供优良菌种资源。【方法】采集农药厂活性污泥样品,通过富集培养和含有炔草酯的LB培养基进行炔草酯降解菌株的分离,通过形态和生理生化特性以及16S rRNA基因序列分析确定其分类学地位,通过单因素试验从温度、pH、接种量和底物浓度等方面考察菌株对炔草酯的降解特性,并利用UPLC-MS分析降解产物。【结果】筛选出一株炔草酯高效降解菌株WP68,经鉴定为鞘氨醇盒菌(Sphingopyxis sp.),该菌株在37°C和pH值为8.0时,10 h内可将200 mg/L的炔草酯降解98.26%。利用UPLC-MS鉴定菌株WP68降解炔草酯的产物为炔草酸。确定了该菌株降解炔草酯的最适温度、pH值、接种量、底物浓度分别是37°C、8.0、5%、200mg/L。菌株WP68还能降解氰氟草酯和精喹禾灵。【结论】Sphingopyxis sp. WP68对炔草酯有较强的降解能力和较高耐受性,在炔草酯污染土壤修复中具有潜在的应用前景。     

Utilization of l-alanine as carbon and nitrogen source by Deusulfovibrio HL21

Desulfovibrio HL21 is unable to grow with amino acids as energy substrates. Alanine, serine, aspartate and to some extent glutamate were used as carbon and nitrogen sources in the presence of hydrogen as the energy substrate. Dense cell suspensions converted alanine stoichiometrically to acetate, NH ₄ ⁺ and presumably HCO ₃ ^- , but at a very low rate. Desulfovibrio HL21 cells grown with alanine as carbon and nitrogen source contained increased levels of NAD(P)-dependent l-alanine dehydrogenase as compared to cells grown with NH₄Cl as nitrogen source. Unfavourable kinetic properties of this alanine dehydrogenase, repression of the synthesis of the enzyme by NH ₄ ⁺ and a low rate of NADH oxidation all have a negative effect on the rate of degradation of alanine and may partly explain the inability of the strain to grow with alanine as an energy substrate.     

牛粪堆肥中纤维素高效降解菌的筛选与产酶条件优化

【背景】纤维素是一种有待开发利用的生物质资源,对于能源危机、环境污染问题的解决具有重要作用。【目的】从牛粪堆肥中分离出产纤维素酶的细菌,研究该菌株的纤维素降解能力。【方法】采用纤维素固体平板刚果红染色法进行初筛、液体发酵纤维素酶活测定法进行复筛。【结果】筛选获得一株具有高产纤维素酶活性的解淀粉芽孢杆菌(Bacillusamyloliquefaciens),命名为N5。单因素分析试验结果显示,菌株N5具有较好的pH、温度和盐度耐受性,正交优化试验结果表明,菌株N5产纤维素酶的最佳条件为：发酵初始pH 5.0,发酵时间96 h,发酵温度40℃。在此条件下,羧甲基纤维素(carboxymethyl cellulose, CMC)酶活为189.27 U/mL。此外,菌株N5能够在7 d内使水稻秸秆减重率达到19.35%。扫描电镜结果表明菌株N5能够有效促进水稻秸秆降解。【结论】菌株N5具有较高的纤维素酶活力,具有开发成高效好氧堆肥菌剂的潜质,这为固体废弃物中纤维素的生物转化提供了优质菌种资源。     

枯草芽胞杆菌降解毒死蜱特性研究

研究生物量、pH、毒死蜱浓度和温度对枯草芽胞杆菌3374菌株(编号为GU086422)在水溶液中降解毒死蜱特性,考察该菌株对白菜上毒死蜱残留的降解特性。结果表明,在毒死蜱质量浓度为240 mg/L、pH7.0、温度30℃的适宜条件下,枯草芽胞杆菌3374菌株对毒死蜱的降解率达到92.48%。该菌株能够有效提高白菜叶面上毒死蜱残留的降解速度,表明其在白菜上具有有效降解毒死蜱的能力,在无公害农产品生产中具有广阔的应用潜力。     

库特氏菌属细菌的研究进展

库特氏菌属归属于动性球菌科,属内细菌分布环境多样,在土壤、地表水、野生动物体表、昆虫肠道等处均有检出。近几年,研究者对库特氏菌属的关注度显著增加并且在多个领域发现了其新价值,包括良好的抗逆性和多重降解功能等,同时确定少数菌株具有致病性。目前,仍存在着对属内各降解机制研究均较浅显、从功能的发现到具体应用方案之间缺乏有效衔接等问题亟待解决。对库特氏菌属的建立、分类学特征、属内成员分布、致病性、功能研究与应用等方面进行介绍,以期为该属菌株的分离鉴定、资源开发以及实际应用提供参考。     

Effect of organic and inorganic nitrogenous compounds on RDX degradation and cytochrome P-450 expression in <Emphasis Type="Italic">Rhodococcus</Emphasis> strain YH1

We hypothesized that biodegradation of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX)—a widely used explosive contaminating soil and groundwater—by Rhodococcus strain YH1 is controlled by the presence of external nitrogen sources. This strain is capable of degrading RDX while using it as sole nitrogen source under aerobic conditions. Both inorganic and organic nitrogen sources were found to have a profound impact on RDX-biodegradation activity. This effect was tested in growing and resting cells of strain YH1. Nitrate and nitrite delayed the onset of RDX degradation by strain YH1, while ammonium inhibited it almost completely. In addition, 2,4,6-trinitrotoluene (TNT) inhibited RDX degradation and growth of strain YH1. On the other hand, tetrahydrophthalamide did not influence biodegradation or growth. Growth on RDX induced the expression of a cytochrome P-450 enzyme that is suggested to be involved in the first step in the aerobic pathway of RDX degradation, as identified by SDS-PAGE analysis. Ammonium and nitrite strongly repressed cytochrome P-450 expression. Our findings suggest that effective RDX bioremediation by strain YH1 requires the design of a treatment scheme that includes initial removal of ammonium, nitrite, nitrate and TNT before RDX degradation can take place.     

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     

Purification and properties of pyrophosphatase of Acinetobacter johnsonii 210A and its involvement in the degradation of polyphosphate

Inorganic pyrophosphatase (E.C. 3.6.1.1) of Acinetobacter johnsonii210A was purified 200-fold to apparent homogeneity. The enzyme catalyzedthe hydrolysis of inorganic pyrophosphate and triphosphate to orthophosphate.No activity was observed with other polyphosphates and a wide variety oforganic phosphate esters. The molecular mass of the enzyme was estimatedto be 141 kDa by gelfiltration. Sodium dodecyl sulfate-polyacrylamide gelelectrophoresis indicated a subunit composition of six identical polypeptideswith a molecular mass of 23 kDa. The cation Mg² was required foractivity, the activity with Mn², Co² and Zn² was 48, 48 and 182% of the activity observed with Mg², respectively. The enzyme was heat-stable and inhibited by fluoride and iodoacetamide. The analysis of the kinetic properties of the enzyme revealed an apparent K_m for pyrophosphate of 0.26 mM. In A. johnsonii 210A, pyrophosphatase may be involved in the degradation of high-molecular polyphosphates under anaerobic conditions: (i) it catalyses the further hydrolysis of pyrophosphate and triphosphate formed from high-molecular weight polyphosphates by the action of exopolyphosphatase, and (ii) it abolishes the inhibition of polyphosphate: AMP phosphotransferase-mediated degradation by pyrophosphate and triphosphate.     

Propylphenols are metabolites in the anaerobic biodegradation of propylbenzene under iron-reducing conditions

The metabolism of monoaromatic hydrocarbons by an iron-reducing bacterial enrichment culture originating from diesel-contaminated groundwater was examined using d₇-propylbenzene as a model hydrocarbon. Sequence analysis of the 16S rDNA gene showed that the dominant part (10 of 10 clones) of the enrichment culture consisted of a bacterium closely related to clones found in benzene-contaminated groundwater and to the iron-reducing -proteobacterium, Rhodoferax ferrireducens (similarity values were 99.5% and 98.3%, respectively). In degradation studies conducted over 18 weeks, d₇-propylphenols were detected by gas chromatography–mass spectrometry (GC/MS) as intra-cellular metabolites concomitant with cell growth in the cultures. The amount of propylphenols increased during the exponential growth phase, and by the end of this phase 4 × 10^–14 moles of ferric iron were reduced and 3 × 10^–15 moles propylphenol produced for every cell formed. During the stationary growth phase the cell density was approximately 10⁷ ml^–1, with significantly correlated amounts of propylphenols. Succinate derivates of propylbenzene or phenylpropanol previously shown to be the initial metabolites in the anaerobic degradation of alkylbenzenes could not be identified. This study is the first to report that oxidation of propylbenzene to propylphenols can initiate anaerobic propylbenzene degradation and that iron-reducing bacteria are responsible for this process. In addition, the study shows the importance of taking account of the metabolites adhering to solid phases when determining the extent of biodegradation, so as not to underestimate the extent of the process.