Kinetics of competitive inhibition and cometabolism in the biodegradation of benzene, toluene, and p-xylene by two Pseudomonas isolates

Two Pseudomonas species (designated strains B1 and X1) were isolated from an aerobic pilot-scale fluidized bed reactor treating groundwater containing benzene, toluene, and p-xylene (BTX). Strain B1 grew with benzene and toluene as the sole sources of carbon and energy, and it cometabolized p-xylene in the presence of toluene. Strain X1 grew on toluene and p-xylene, but not benzene. In single substrate experiments, the appearance of biomass lagged the consumption of growth substrates, suggesting that substrate uptake may not be growth-rate limiting for these substrates. Batch tests using paired substrates (BT, TX, or BX) revealed competitive inhibition and cometabolic degradation patterns. Competitive inhibition was modeled by adding a competitive inhibition term to the Monod expression. Cometabolic transformation of nongrowth substrate (p-xylene) by strain B1 was quantified by coupling xylene transformation to consumption of growth substrate (toluene) during growth and to loss of biomass during the decay phase. Coupling was achieved by defining two transformation capacity terms for the cometabolizing culture: one that relates consumption of growth substrate to the consumption of nongrowth substrate, and second that relates consumption of biomass to the consumption of nongrowth substrate. Cometabolism increased decay rates, and the observed yield for strain B1 decreased in the presence of p-xylene. (c) 1993 Wiley & Sons, Inc.     

罕山土壤微生物群落组成对植被类型的响应

选取分布在中国东北部地区的阔叶林-针叶林-亚高山草甸这一明显的植被垂直带谱来研究植被类型对土壤微生物群落组成的影响。选取5种植被类型-山杨(Populus davidiana)(1250—1300 m),山杨(P.davidiana)与白桦(Betula platyphylla)的混交林(1370—1550 m),白桦(B.platyphylla)(1550—1720 m),落叶松(Larix principis-rupprechtii)(1840—1890 m),亚高山草甸(1900—1951 m),采用磷脂脂肪酸(Phopholipid Fatty Acids,PLFAs)分析方法测定不同植被类型下的土壤微生物群落组成。分别采用主成分分析(Principal Components Analysis,PCA)以及冗余分析(Redundancy Analysis,RDA)来解释单种特征PLFAs的分异以及土壤理化指标与微生物PLFAs指标间的相关性。结果表明不同植被类型下土壤有机碳(SOC)对土壤微生物PLFAs总量,各类群(真菌(f)、细菌(b)、革兰氏阳性菌(G+)、革兰氏阴性菌(G-))生物量以及群落结构影响显著;土壤微生物PLFAs总量及各类群的生物量随土层加深总体上表现降低趋势,G+/G-和f/b分别随土层加深总体上表现升高趋势。不同植被类型下,阔叶混交林土壤PLFAs总量及各类群生物量总体上最高;针叶林比阔叶林下的f/b和G+/G-高;亚高山草甸下低的p H值对有机碳的可利用性有一定的抑制作用,导致f/b和G+/G-的值相对较高。总之,不同植被类型下SOC对土壤微生物群落组成的影响最为显著,而较低的p H对有机碳的可利用性有一定的抑制作用;真菌对植被类型的变化比细菌更敏感,而细菌更易受可利用性养分和p H变异的影响,这对预测不同林型下的土壤微生物群落组成有重要的启示作用。     

Quantification of toluene dioxygenase induction and kinetic modeling of TCE cometabolism by Pseudomonas putida TVA8.

As measured by the toluene-induced bioluminescent response of Pseudomonas putida TVA8 in batch experiments, toluene dioxygenase (Tod) enzyme activities are dependent on toluene concentration between 0 and 30 mg/L. To provide a measure of the Tod activity for use in Michaelis-Menten competitive-inhibition kinetics, a correlation between toluene concentration and induced Tod activity as measured by an induced bioluminescent response of P. putida TVA8 is presented as a nondimensional Tod activity parameter. A packed-bed, radial-flow bioreactor (RFB) using the bioreporter P. putida TVA8A serves as the model system for studying the effect of the enzyme activity parameter on model predictions of vapor-phase toluene oxidation and trichloroethylene (TCE) cometabolism. Mass balances were performed on a differential section of the RFB to describe the radial transport of vapor-phase toluene and TCE through a bulk gas phase and the concomitant biological reaction in a stationary biofilm phase. The finite-element Galerkin weak-statement formulation with first-order basis functions was used to find the optimum solution to the highly nonlinear, coupled equations. For this RFB system with toluene concentrations less than 1 mg/L in the bulk gas phase, the Tod activity parameter enables accurate predictions of steady-state TCE degradation rate (0.27 microg TCE/min).     

Altering the mineral composition of soil causes a shift in microbial community structure

This study tests the hypothesis that altering the mineral composition of soil influences microbial community structure in a nutrient-deficient soil. Microcosms were established by adding mica (M), basalt (B) and rock phosphate (P) to soil separately, and in combination (MBP), and by planting with Lolium rigidum, Trifolium subterraneum or by leaving unplanted. The effects of mineral and plant treatments on microbial community structure were assessed using automated ribosomal intergenic spacer analysis. Bacterial community structure was significantly affected by both mineral (global R=0.73 and P<0.001) and plant (global R=0.71 and P<0.001) treatments, as was the fungal community structure: mineral (global R=0.65 and P<0.001) and plant (global R=0.65 and P<0.001) treatments. All pairwise comparisons of bacterial and fungal communities between different mineral treatments and between different plant treatments were significantly different (P<0.05). This study has shown that mineral addition to soil microcosms resulted in substantial changes in both bacterial and fungal community structure, dependent on the type of mineral added and the plant species present. These results suggest that the mineral composition of soil may be an important factor influencing the microbial community structure in soil.     

Cometabolic degradation of trichloroethylene in a bubble column bioscrubber

A bubble column bioreactor was used as bioscrubber to carry out a feasibility study for the cometabolic degradation of trichloroethylene (TCE). Phenol was used as cosubstrate and inducer. The bioreactor was operated like a conventional chemostat with regard to the cosubstrate and low dilution rates were used to minimize the liquid outflow. TCE degradation measurements were carried out using superficial gas velocities between 0.47and 4.07 cm s(-1) and TCE gas phase loads between 0.07 and 0.40 mg L(-1) Depending on the superficial gas velocity used, degrees of conversion between 30% and 80% were obtained. A simplified reactor model using plug flow for the gas phase, mixed flow for the liquid phase, and pseudo first order reaction kinetics for the conversionof TCE was established. The model is able to give a reasonable approximation of the experimental data. TCE degradation at the used experimental conditions is mainly limited by reaction rate rather than by mass transfer rate. The model can be used to calculate the reactor volume and the biomass concentration for a required conversion. (c) 1995 John Wiley & Sons Inc.     

土壤微生物群落构建理论与时空演变特征

土壤微生物作为陆地生态系统的重要组成部分,直接或间接地参与几乎所有的土壤生态过程,在物质循环、能量转换以及污染物降解等过程中都发挥着重要作用。对土壤微生物时空演变规律及其形成机制的研究,不仅是微生物演变和进化的基础科学问题,也是预测微生物及其所介导的生态功能对环境条件变化响应、适应和反馈的理论依据。讨论了土壤微生物群落的定义、测度方法和指标,认为群落是联系动植物宏观生态学与微生物生态学的基础,群落构建机制是宏观和微观生态学都需要研究的核心科学问题;从生态学的群落构建理论出发,阐述了包括生态位理论/中性理论、过程理论和多样性-稳定性理论在土壤微生物时空演变研究中的应用,以及微生物群落在时间和空间上的分布特征及其尺度效应;确立了以微生物群落构建理论为基础、不同时空尺度下土壤微生物群落演变特征为主要内容的微生物演变研究的基本框架。     

三种种植方式对土壤微生物群落组成的影响

土壤微生物是表征土壤质量变化的敏感指标之一。借助长期定位试验, 采用磷脂脂肪酸分析方法研究了3种种植方式(玉米(Zea mays)连作、玉米非连作和撂荒)对土壤微生物群落组成的影响。结果表明, 在不同的种植方式下, 土壤微生物群落组成有明显的差异。玉米连作的土壤中总磷脂脂肪酸和细菌磷脂脂肪酸含量最低, 分别为33.12 nmol·g^-1和18.09 nmol·g^-1。非连作的土壤真菌磷脂脂肪酸和真菌/细菌分别为0.61 nmol·g^-1和3.06%, 显著低于撂荒和连作(p < 0.05), 非连作方式下, 革兰氏阳性细菌/革兰氏阴性细菌增大。撂荒土壤的总磷脂脂肪酸和细菌磷脂脂肪酸分别为42.98和24.68 nmol·g^-1, 高于耕作处理。 同时, 在撂荒方式下, 革兰氏阳性细菌和革兰氏阴性细菌的含量增加, 革兰氏阳性细菌/革兰氏阴性细菌降低。主成分分析结果表明: 耕作处理(玉米连作和非连作)分布第一主成分负方向上, 第一主成分得分系数分别为-2.48和-1.84; 撂荒分布第一主成分正方向上, 第一主成分得分系数为2.31, 与连作和非连作差异显著(p < 0.05)。冗余分析(RDA)表明: 土壤pH、总氮、有效磷和土壤>0.25 mm水稳性团聚体含量与磷脂脂肪酸呈正相关, 并且土壤pH和土壤>0.25 mm水稳性团聚体含量对土壤微生物群落的影响最大。     

改变碳输入对亚热带人工林土壤微生物生物量和群落组成的影响

通过在亚热带杉木(Cunninghamia lanceolata)和米老排(Mytilaria laosensis)人工林中设置互换凋落物、去除凋落物、去除凋落物+去除根系和对照处理来分析改变地上、地下碳输入对人工林土壤微生物生物量和群落组成的影响。结果显示,改变地上、地下碳输入对土壤微生物生物量碳、氮的影响因树种而异。在米老排林中,土壤微生物生物量不受碳源的限制。而在杉木林中,加入米老排凋落物、去除凋落物和去除凋落物+去除根系3种处理中土壤微生物生物量碳、氮具有明显增加的趋势。磷脂脂肪酸分析结果显示,杉木林中,添加高质量的米老排凋落物后,革兰氏阳性细菌、阴性细菌、丛枝菌根真菌、放线菌和真菌群落生物量分别显著增加了24%、24%、53%、25%、28%,革兰氏阴性细菌和丛枝菌根真菌的相对丰度均有显著增加。与对照相比,杉木林中去除凋落物后革兰氏阳性细菌、阴性细菌、丛枝菌根真菌、放线菌和真菌群落生物量分别显著增加了22%、29%、44%、25%、52%,真菌与细菌比值显著增加了21%。但是,去除凋落物+去除根系处理对两个树种人工林土壤微生物群落组成均无显著影响。米老排和杉木林土壤微生物生物量碳、氮的季节变化格局不同,土壤养分有效性可能是驱动土壤微生物生物量季节变化的主要因子。未来研究需要关注凋落物和根系在不同树种人工林中对土壤微生物群落的相对贡献。     

甲烷及三氯乙烯驯化对垃圾填埋场覆盖土细菌群落结构的影响

对典型垃圾填埋覆盖土进行CH₄原位富集和三氯乙烯(TCE)驯化,研究了其生物氧化能力和微生物群落结构变化.覆盖土CH₄氧化速率为0.20～0.87 μmol·g^-1 soil·h^-1,TCE降解速率为0.009～0.013 mg·L^-1·h^-1,其中山东垃圾填埋场覆盖土土样甲烷氧化活性高于广东、上海和重庆地区土样.通过Illumina MiSeq测序技术分析了α多样性和驯化前后微生物菌群结构变化规律.结果表明: 在所有被注释的操作分类单元聚类结果中,细菌OTUs分配为39个门,85个纲,562个属,富集驯化后变形杆菌门、拟杆菌门、绿弯菌门和酸杆菌门仍为各土样的优势菌群,所占比例之和高于77.4%;γ-变形杆菌纲、β--变形杆菌纲、α-变形杆菌纲、放线菌纲和酸杆菌纲所占比例之和高于26.5%.嗜甲基菌属、厌氧绳菌属、节杆菌属和假单胞菌属经TCE驯化后,其相对丰度呈增加趋势.表明在覆盖土氯代烃生物降解过程中,除了被广泛认可的甲烷氧化菌异养共代谢机制以外,还存在非甲烷共代谢机制和氯代烃自养降解机制.     

凋落物多样性及组成对凋落物分解和土壤微生物群落的影响

凋落物分解是生态系统营养物质循环的核心过程,而土壤微生物群落在凋落物分解过程中扮演着极其重要且不可替代的角色。随着生物多样性的丧失日益严峻,探讨凋落物多样性及组成对凋落物分解和土壤微生物群落的影响,不仅有助于了解凋落物分解的内在机制,而且可为退化草原生态系统的恢复提供参考。以内蒙古呼伦贝尔草原退化恢复群落中的草本植物为研究对象,依据植物多度、盖度、频度和物种的重要值及其在群落中的恢复程度筛选出排序前4的羊草（Leymus chinensis）、茵陈蒿（Artemisia capillaris）、麻花头（Serratula centauroides）、二裂委陵菜（Potentilla bifurca）的凋落物为实验材料,通过设置3种凋落物多样性水平（1,2,4）,包括11种凋落物组合（单物种凋落物共4种,两物种凋落物混合共6种,四物种凋落物混合共1种）,利用磷脂脂肪酸（PLFA）方法来研究分解60 d后凋落物多样性及组成对凋落物分解和土壤微生物群落的影响。结果表明：（1）凋落物物种多样性仅对C残余率具有显著影响,表现在两物种混合凋落物C残余率显著低于单物种凋落物,而凋落物组成对所观测的4个凋落物分解参数（质量、C、N残余率以及C/N）均具有显著影响;（2）凋落物物种多样性对细菌（B）含量具有显著影响,而凋落物组成对真菌（F）含量具有显著影响,两者对F/B以及微生物总量均无显著影响;（3）冗余分析结果表明凋落物组成与凋落物分解相关指标（凋落物质量、C、N残余率及C/N）和土壤微生物（真菌、细菌含量）的相关关系高于凋落物多样性。（4）进一步建立结构方程模型（Structural Equation Model,SEM）发现,凋落物初始C含量对凋落物质量、C、N残余率及C/N有显著正的直接影响;凋落物木质素含量对凋落物质量、C、N残余率有显著正的直接影响;凋落物初始N含量对N残余率有显著正的直接影响,而对C残余率及C/N有显著负的直接影响;凋落物初始C/N对凋落物质量、N残余率有显著正的直接影响,而对C/N有显著负的直接影响。此外,凋落物初始C、N、木质素含量及C/N均对真菌含量具有显著正影响,并可通过真菌对凋落物质量分解产生显著负的间接影响。以上结果表明该退化恢复区域优势种凋落物分解以初始C、木质素为主导,主要通过土壤真菌影响凋落物的分解进程,这将减缓凋落物的分解速率进而减慢草原生态系统的进程。这些结果为凋落物多样性及组成对自身分解和土壤微生物群落的影响提供了实验依据,也为进一步分析凋落物分解内在机制以及草原生态系统的恢复提供了数据参考。     

Orchard floor management practices that maintain vegetative or biomass groundcover stimulate soil microbial activity and alter soil microbial community composition

Groundcover management systems (GMS) are important in managing fruit-tree orchards because of their effects on soil conditions, nutrient availability, tree growth and yields. We employed a polyphasic approach, incorporating measures of soil microbial abundance, activity and community composition, to study the long-term effects of different GMS on biotic and abiotic factors in an orchard soil. Four GMS treatments – Pre-emergence residual herbicides (Pre-H), post-emergence herbicide (Post-H), mowed-sod (Grass), and hardwood bark mulch (Mulch) – were established in 2-m-wide strips within tree rows in an apple orchard in 1992, and have been maintained and monitored annually until the present. We have measured soil water and nutrient availability, tree growth, and yields annually from 1993 to 2003. Soil nematode numbers and trophic groups were evaluated in July and Oct. 2001, and Sept. 2003. Numbers of culturable bacteria and fungi, soil respiratory activity, eubacterial and fungal community composition were determined in May and Sept. 2003. The Pre-H treatment soil had the fewest culturable bacteria, while the Grass treatment had the largest population of culturable fungi. Soil nematode population size and diversity were also affected by GMS treatments; the Pre-H treatment had the lowest ratio of (bacteriovores + fungivores) to plant parasitic nematodes. Soil respiration rates were higher in the Mulch than in other treatments during a 40-day incubation period. Hierarchical cluster dendrograms of denaturing gradient gel electrophoresis (DGGE) fingerprints for eubacterial community 16S rRNA genes indicated that Post-H and Grass treatments clustered together and separately from the Pre-H and Mulch treatments, which were also grouped together. The influence of GMSs on the fungal community, as assessed by PCR-DGGE of the internal transcribed spacer (ITS) region, was not as pronounced as that observed for bacteria. Soil fungal community composition under the Mulch differed from that under other treatments. The effects of GMS on soil microbial community abundance, activity, and composition were associated with observed differences in soil organic matter inputs and turnover, nutrient availability, and apple tree growth and yields under the different GMS treatments.     

Biodegradation of trichloroethylene and toluene by indigenous microbial populations in soil.

The biodegradation of trichloroethylene (TCE) and toluene, incubated separately and in combination, by indigenous microbial populations was measured in three unsaturated soils incubated under aerobic conditions. Sorption and desorption of TCE (0.1 to 10 micrograms ml-1) and toluene (1.0 to 20 micrograms ml-1) were measured in two soils and followed a reversible linear isotherm. At a concentration of 1 micrograms ml-1, TCE was not degraded in the absence of toluene in any of the soils. In combination, both 1 microgram of TCE ml-1 and 20 micrograms of toluene ml-1 were degraded simultaneously after a lag period of approximately 60 to 80 h, and the period of degradation lasted from 70 to 90 h. Usually 60 to 75% of the initial 1 microgram of TCE ml-1 was degraded, whereas 100% of the toluene disappeared. A second addition of 20 micrograms of toluene ml-1 to a flask with residual TCE resulted in another 10 to 20% removal of the chemical. Initial rates of degradation of toluene and TCE were similar at 32, 25, and 18 degrees C; however, the lag period increased with decreasing temperature. There was little difference in degradation of toluene and TCE at soil moisture contents of 16, 25, and 30%, whereas there was no detectable degradation at 5 and 2.5% moisture. The addition of phenol, but not benzoate, stimulated the degradation of TCE in Rindge and Yolo silt loam soils, methanol and ethylene slightly stimulated TCE degradation in Rindge soil, glucose had no effect in either soil, and dissolved organic carbon extracted from soil strongly sorbed TCE but did not affect its rate of biodegradation.     

Degradation of trichloroethylene by a growth-arrestedPseudomonas putida

A toluene-oxidizing strain ofPseudomonas mendocina KR1 containing toluene-4-mono-oxygenase (TMO) completely degrades TCE with the addition of toluene as a co-substrate in aerobic condition. In order to constructin situ bioremediation system for TCE degradation without any growth-stimulating nutrients or toxic inducers such as toluene, we used the carbon-starvation promoter ofPseudomonas putida MK1 (Kim, Y.et al., J. bacteriol., 1995). Upon entry into the stationary phase due to the deprivation of nutrients, this promoter is strongly induced without further cell growth. The TMO gene cluster (4.5 kb) was spliced downstream of the carbon starvation promoter ofPseudomona putida MK1, already cloned in pUC19. TMO under the carbon starvation promoter was not expressed inE. coli cells either in stationary phase or exponential phase. For TMO expression inPseudomonas strains,tmo and carbon starvation promoter region were recloned into a modified broad-host range vector pMMB67HES which was made from pMMB67HE (8.9 kb) by deletion oftac promoter andlacI ^q (about 1.5 kb). Indigo was produced by TMO under the carbon starvation promoter in aPseudomonas strain of post-exponential phase on M9 (0.2% glucose and 1mM indole) or LB. 18% of TCE was degraded in 14 hours after entering the stationary phase at the initial concentration of 6.6μ M in liquid phase.     

林下植物剔除对毛竹林土壤微生物群落结构的影响

本研究以中亚热带地区广泛分布的毛竹林为对象,采用随机区组实验设计,分析了林下植物剔除对毛竹林土壤微生物群落结构和土壤理化特性的影响,探讨林下植物对毛竹林土壤微生物群落结构的调控机制.结果 表明:林下植物剔除对土壤理化特性产生显著影响,主要表现为土壤全氮、硝态氮和有效磷含量增加,而土壤铵态氮、全磷含量及土壤pH值降低.此...     

Kinetic analysis of the inhibitory effect of trichloroethylene (TCE) on nitrification in cometabolic degradation

In this study, the inhibitory effect of TCE on nitrification process was investigated with an enriched nitrifier culture. TCE was found to be a competitive inhibitor of ammonia oxidation and the inhibition constant (K _I ) was determined as 666–802 μg/l. The TCE affinity for the AMO enzyme was significantly higher than ammonium. The effect of TCE on ammonium utilization was evaluated with linearized plots of Monod equation (e.g., Lineweaver–Burk, Hanes–Woolf and Eadie–Hofstee plots) and non-linear least square regression (NLSR). No significant differences were found among these data evaluation methods in terms of kinetic parameters obtained.     

亚热带森林土壤微生物生物量及群落功能特征的城乡梯度变化

土壤微生物在土壤养分循环以及生物地球化学循环中起着重要作用,土壤微生物对环境变化响应灵敏.为研究城乡梯度环境变化对亚热带森林土壤微生物的影响,本研究选取合肥市大蜀山国家森林公园(城市林)、紫蓬山国家森林公园(远郊林)、六安市万佛山(乡村自然林)为样地,分析比较其微生物生物量碳(MBC)、微生物生物量氮(MBN)及微生物...     

Recovery of trichloroethylene removal efficiency through short-term toluene feeding in a biofilter enriched withPseudomonas putida F1

Trichloroethylene (TCE) is an environmental contaminant provoking genetic mutation and damages to liver and central nerve system even at low concentrations. A practical scheme is reported using toluene as a primary substrate to revitalize the biofilter column for an extended period of TCE degradation. The rate of trichloroethylene (TCE) degradation byPseudomonas putida F1 at 25°C decreased exponentially with time, without toluene feeding to a biofilter column (11 cm I.D.×95 cm height). The rate of decrease was 2.5 times faster at a TCE concentration of 970 μg/L compared to a TCE concentration of 110 μg/L. The TCE itself was not toxic to the cells, but the metabolic intermediates of the TCE degradation were apparently responsible for the decrease in the TCE degradation rate. A short-term (2 h) supply of toluene (2,200 μg/L) at an empty bed residence time (EBRT) of 6.4 min recovered the relative column activity by 43% when the TCE removal efficiency at the time of toluene feeding was 58%. The recovery of the TCE removal efficiency increased at higher incoming toluene concentrations and longer toluene supply durations according to the Monod type of kinetic expression. A longer duration (1.4∼2.4 times) of toluene supply increased the recovery of the TCE removal efficieny by 20% for the same toluene load.     

茂兰喀斯特森林小生境类型对土壤微生物群落组成的影响

喀斯特山区是我国典型的生态脆弱区,区内分布复杂多样的小生境类型(石洞、石缝、石沟、石槽、土面等)。土壤微生物是陆地生态系统最活跃的成分,推动着生态系统能量流动和物质循环。研究不同小生境类型对土壤微生物群落组成的影响,对于喀斯特山区森林土壤微生物特征的认识和生态保护具有积极的意义。以贵州茂兰喀斯特森林小生境为对象,对其土壤理化性质和微生物组成进行了比较分析,并对小生境类型进行了数量聚类。结果表明:6种小生境土壤中共检测到48种磷脂脂肪酸(PLFA),其中,标记土壤微生物的特征PLFA有18种;土壤微生物总量、细菌、真菌、放线菌、G~+、G~-的含量均表现为石槽石沟土面表层石洞石缝土面下层,同时,各微生物类型的含量在不同生境类型中表现为细菌放线菌真菌; G~+/G~-在六种生境中表现为石槽石沟土面表层石缝石洞土面下层;真菌/细菌变幅为0.14—0.16,具体表现为土面表层石缝石沟石槽石洞土面下层;相关性分析和多元回归树分析显示,不同小生境类型的土壤有机质是影响土壤微生物群落组成的首要因子。聚类结果表明,小生境可划分为石槽-石沟型、石洞-石缝型、土面表层型以及土面下层型4种生境类型。研究结果证实了不同小生境类型对微生物分布特征存在显著影响。