Characterization of depth-related microbial communities in lake sediment by denaturing gradient gel electrophoresis of amplified 16S rRNA fragments

The characterization of microbial communities of different depth sediment samples was examined by a culture-independent method and compared with physicochemical parameters, those are organic matter （OM）, total nitrogen （TN）, total phosphorus （TP）, pH and redox potential （Eh）. Total genomic DNA was extracted from samples derived from different depths. After they were amplified with the GC-341 f/907r primer sets of partial bacterial 16S rRNA genes, the products were separated by denaturing gradient gel electrophoresis （DGGE）. The profile of DGGE fingerprints of different depth sediment samples revealed that the community structure remained relatively stable along the entire 45 cm sediment core, however, principal-component analysis of DGGE patterns revealed that at greater sediment depths, successional shifts in community structure were evident. The principle coordinates analysis suggested that the bacterial communities along the sediment core could be separated into two groups, which were located 0-20 cm and 21-45 cm, respectively. The sequencing dominant bands demonstrated that the major phylogenetic groups identified by DGGE belonged to Bacillus, Bacterium, Brevibacillus, Exiguobacterium, γ-Proteobacterium, Acinetobacter sp. and some uncultured or unidentified bacteria. The results indicated the existence of highly diverse bacterial community in the lake sediment core.     

内蒙古河套灌区不同盐碱程度的土壤细菌群落多样性

采用变性梯度凝胶电泳(DGGE)技术对内蒙古河套灌区三种不同盐碱程度土壤(盐土,强度盐化土,轻度盐化土)不同深度(0~20cm和20~30cm)土壤细菌16S rDNA V3~V6可变区扩增片段进行分析,并对土壤理化性质进行了测定.结果表明:细菌群落多样性随土壤盐碱化程度的加深而减少(轻度盐化土 > 强度盐化土 > 盐土),随土壤深度的增加而降低(细菌群落多样性0~20cm土层大于20~30cm土层).细菌Shannon-Wiener指数在轻度盐化土中最大为3.36,在强度盐化土和盐土分别为3.05和2.49.不同盐碱程度土壤以细菌相似系数聚类,分为0~20cm层与20~30cm层两大族群,土壤细菌群落Shannon-Wiener指数在0~20cm层中(盐土为3.04,强度盐化土为3.29,轻度盐化土为3.36)均大于在20~30cm层(盐土为2.49,强度盐化土为3.05,轻度盐化土为3.14).相关性分析和典范对应分析表明,土壤w(EC)、pH值、w(SOC)、w(TP)是土壤细菌群落结构多样性的显著影响因素,不同盐碱程度土壤中细菌群落的Shannon-Wiener指数与土壤w(EC)(r=-0.542,P < 0.05)、pH(r=-0.526,P < 0.05)呈显著负相关,与土壤w(SOC)(r=0.700,P < 0.01)和w(TP)(r=0.805,P < 0.01)呈极显著正相关.w(EC)和pH对盐碱土壤细菌群落结构的影响力最大. 回收DGGE图谱中20个优势条带进行测序分析,结果显示,变形菌纲(α-变形菌纲、β-变形菌纲、γ-变形菌纲和δ-变形菌纲)是盐碱土壤的主要类群.     

农业废物好氧堆肥过程因子对细菌群落结构的影响

采用聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)技术研究了农业废物堆肥过程中细菌种群随时间的变化.同时,应用Quantity One2.0和Canoco4.5软件对获得的堆肥细菌种群数据与堆肥过程因子:环境温度、堆体温度、pH、含水率、水溶性有机碳(WSC)、C/N、水溶性氨氮(NH4+-N)和硝氮(NO3--N)进行冗余分析,并做出样点、种群与堆肥过程因子的二维排序图.结果表明,细菌群落(样点)以堆肥过程因子为梯度大体可划分为升温期(1～2d)、高温期(3～11d)、降温期(12～18d)和腐熟期(19～36d)4个阶段,每一阶段均有对应种群存在.不同的堆肥过程因子对细菌种群的影响大小依次为:NO3--N堆体温度WSCC/NNH4+-N含水率pH环境温度,其中,堆体温度、WSC、NO3--N、NH4+-N对细菌种群的影响极显著(p0.01),C/N、pH对细菌种群的影响显著(p0.05),含水率、环境温度对细菌种群的影响不显著.     

细菌群落结构对水体富营养化的响应

于2005年2月在太湖不同营养水平湖区采集水样,寞接提取水样的总DNA,以细菌16SrDNA通用引物进行V3区PCR扩增,产物经DGGE(变性凝胶梯度电泳)分离后,获得水体细菌群落的16SrDNA指纹图谱;并运用FDC(表面荧光直接计数)方法对细菌丰度进行了测定.结果表明,水体中细菌的群落结构随着水体富营养化程度的改变产生明显变化,细菌数量呈现随水体营养水平增加而上升的趋势,营养水平较低的湖心区细菌数量仅为2.87×106 cells·mL-1,超富营养化的五里湖区水体中.细菌数量高达5.92×106cells·mL-1;细菌群落多样性随水体营养水平增加呈现显著的下降趋势,在超富营养化的1#、2#采样位点,细菌DGGE主要条带数量仅为23条,在沉水植物丰富区(贡湖湾)细菌DGGE主要条带数量达33条.PCA(主成分分析)的聚类结果表明,太湖水体中不同营养水平下的细菌群落多样性可大致归为3种类型,超富营养化的五里湖区水体细菌群落结构与梅粱湾及汞湖湾存在明显不同,可以大体归为超富营养类型、藻型湖区及草型湖区类型3种.此结果反映了水体富营养化过程中,由于营养物质的增加,促进了某些类群微生物的大量繁殖,水体中微生物的生物量显著增加;生态环境条件的恶化,造成了微生物多样性的下降,这可能导致原本稳定的水生态系统脆弱化.     

太湖不同湖区冬季沉积物细菌群落多样性

为了探究太湖不同营养水平湖区冬季沉积物细菌群落多样性,利用高通量测序对太湖水草区、湖心区和河口区共9个采样点表层沉积物细菌的16S rRNA基因进行序列测定.结果发现：不同湖区的物种丰富度和均匀度为水草区 > 湖心区 > 河口区;不同湖区优势细菌群落组成存在差异.其中,硝化螺旋菌门（Nitrospirae）和酸杆菌门（Acidobacteria）2类门水平优势细菌以及厌氧绳菌纲（Anaerolineae）和硝化螺菌纲（Nitrospira）2类纲水平优势细菌在不同湖区之间差异显著.冬季太湖沉积物细菌优势类群为绿弯菌门（Chloroflexi）、变形菌门（Proteobacteria）、蓝藻门（Cyanobacteria）和硝化螺旋菌门（Nitrospirae）;在属分类水平上,主要优势类群为unidentified_Chloroplast和微囊藻属（Microcystis）.Cyanobacteria和Microcystis在所有采样点均有检出,平均丰度均以湖心区最高.对沉积物细菌群落与环境因子的关系进行冗余分析,结果表明：营养盐水平、水温和pH值均能影响沉积物细菌群落结构,NO₃^--N和水温是Microcystis的主要影响因子.     

PCR-DGGE技术解析生物制氢反应器微生物多样性

为了揭示发酵法生物制氢反应器厌氧活性污泥的微生物种群多样性 ,从运行不同时期取厌氧活性污泥 ,通过细胞裂解直接提取活性污泥的基因组DNA .以细菌 16SrRNA基因通用引物F338GC/R5 34进行V3高变异区域PCR扩增 ,长约 200bp的PCR产物经变性梯度凝胶电泳 (DGGE)分离后 ,获得微生物群落的特征DNA指纹图谱 .研究表明 ,不同时期的厌氧活性污泥中存在共同种属和各自的特异种属 ,群落结构和优势种群数量具有时序动态性 ,微生物多样性呈现出协同变化的特征 .微生物多样性由强化到减弱 ,群落结构之间的相似性逐渐升高 ,演替速度由快速到缓慢 .优势种群经历了动态演替过程 ,最终形成特定种群构成的顶级群落 .     

Monitoring impact of mefenacet treatment on soil microbial communities by PCR-DGGE fingerprinting and conventional testing procedures

The effect of acetanilide herbicide mefenacet on soil microbial communities was studied using paddy soil samples with different short-term treatments. The culturable bacteria （plate counts）, dehydrogenase activity and changes in community structure （denaturing gradient gel electrophoresis （DGGE） analysis） were used for biological community assessments. Mefenacet was a significant stimulus to cultural aerobic bacteria and dehydrogenase activity while Sphingobacterium multivorum Y1, a bacterium efficiently degrading the mefenacet, only induced the increasing colony-forming unit （CFU） of bacteria but little effect on dehydrogenase activity during the whole experiment. The degree of similarity between the 16S rDNA profiles of the communities was quantified by numerically analyzing the DGGE band patterns. Similarity dendrograms showed that the microbial community structures of the mefenacet-treated and non-treated soils were not significantly different. But supplement of S. multivorum Y1 could increase the diversity of the microbial community in the mefenacet-polluted paddy soil. This work is a new attempt to apply the S. multivorum Y1 for remediation of the mefenacet-polluted environments.     

Denaturing gradient gel electrophoresis fingerprinting of soil bacteria in the vicinity of the Chinese Great Wall Station, King George Island, Antarctica

Bacterial diversity was investigated in soil samples collected from 13 sites around the Great Wall Station, Fildes Peninsula, King George Island, Antarctica, using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes. The classes α-, β-, and γ-Proteobacteria, as well as the phylum Actinobacteria, were found to be the dominant bacteria in the soils around the Great Wall Station. Although the selected samples were not contaminated by oil, a relationship between soil parameters, microbial biodiversity, and human impact was still seen. Sample sites in human impacted areas showed lower bacterial biodiversity (average H′= 2.65) when compared to nonimpacted sites (average H′= 3.05). There was no statistically significant correlation between soil bacterial diversity and total organic carbon (TOC), total nitrogen, or total phosphorus contents of the soil. Canonical correlation analysis showed that TOC content was the most important factor determining bacterial community profiles among the measured soil parameters. In conclusion, microbial biodiversity and community characteristics within relatively small scales (1.5 km) were determined as a function of local environment parameters and anthropogenic impact.     

Effect of methamidophos on soil fungi community in microcosms by plate count, DGGE and clone library analysis

Methamidophos was widely used a pesticide in northern China. The potential influences of methamidophos on soil fungal community in black soil were assessed by plate count, 28S rDNA-PCR-DGGE, and clone library analysis. Three methamidophos levels (50, 150, and 250 mg/kg) were tested in soil microcosms. Results from plate count during a 60-d microcosm experiment showed that high concentrations of methamidophos (250 mg/kg) could significantly stimulate fungal populations. DCGE (denaturing gradient gel electrophoresis) fingerprinting patterns showed a significant difference between the responses of culturable and total fungi communities under the stress of methamidophos. Shannon diversity indices calculated from DGGE profiles indicated that culturable fungi in all microcosms with methamidophos treatment increased after 1 week of incubation. However, the diversity indices of total fungi decreased in the first week, as compared to the stimulation of culturable fungi. At the 8th week, however, all the microcosms treated by methamidophos were similar to the control microcosms in community structure as suggested by the Shannon diversity indices for both culturable and total fungi. In contrast, after 1 week the fungal structure of culturable and unculturable both were disturbed to different extent under the stresses of methamidophos by clustering analysis. Clone sequencing analysis indicated the stimulation of pathogenic and unculturable fungal populations by methamidophos treatment, suggetsing potential risks of plant disease outbreak.     

Bacterial community composition and abundance in leachate of semi-aerobic and anaerobic landfills

Different municipal solid waste landfill methods and landfill ages had crucial impacts on bacterial abundance and composition in leachate.     

底泥中红霉素耐受菌群的多样性分析

为了正确评估抗生素残留的环境风险,了解长期受低浓度红霉素污染条件下,水体底泥中红霉素耐受菌群结构的变化特点,采用传统的微生物培养法和分子生物学方法对长期受低浓度红霉素污染的水体底泥中红霉素耐受菌群结构的多样性进行了分析.结果表明:采用传统的培养方法,从底泥中可分离筛选出3株对红霉素有耐受能力的菌株,根据其形态学特征及16S rDNA序列分析,初步鉴定为赖氨酸芽孢杆菌(Lysinibacillussp.)、土壤芽孢杆菌(Solibacillus silvestris)以及蜡样芽孢杆菌(Bacillus cereus),其中敏感菌蜡样芽孢杆菌经低浓度红霉素诱导对红霉素表现出一定的耐药性.进一步构建底泥耐药菌群16S rDNA克隆文库发现,底泥中红霉素耐受菌群可分为三大类群,其中未获培养的细菌(Uncultured bacterium)在整个文库中比例最大,占65.06%,其次为芽孢杆菌纲(Bacilli)和梭菌纲(Clostridia),分别占文库比例33.73%和1.20%.文库中可培养的耐药菌优势类群为芽孢杆菌纲,与传统培养方法得到的结果相一致.上述研究表明,在长期受低浓度红霉素污染的环境中,红霉素耐药菌的存在具有一定的广泛性和潜在性,将可能威胁人类健康和生态系统,应当重视其生态风险评价与管理.     

复合菌系BYND-8的种群组成及其对沼气产量的影响

为了明确1组在中温下(30℃)高效分解木质纤维素的复合菌系的菌群组成.研究复合菌系预处理秸秆对沼气发酵的影响,利用平板分离法和变性梯度凝胶电泳(DGGE)法研究了中温木质纤维素降解复合菌系BYND-8的菌种组成多样性,通过添加该复合系菌液到以牛粪为原料的沼气发酵体系,研究了添加秸秆降解液对沼气产量的影响.利用平板法分离...     

应用变性梯度凝胶电泳研究厦门西海域超微型真核浮游生物多样性

应用变性梯度凝胶电泳(DGGE)方法对厦门西海域超微型真核浮游生物的遗传多样性进行周年变化特征研究分析.结果表明,厦门西海域典型测站的超微型真核浮游生物群落具有丰富的多样性组成;厦门西海域超微型真核浮游生物群落遗传结构和主要类群组成季节性变化显著:不同典型测站之间的超微型真核浮游生物群落遗传结构的周年变化特征相似;不同典型测站之间的超微型真核浮游生物群落的类群组成也很相似,由绿藻、Stramenopiles、定鞭金藻、甲藻、未确定类群(unidentified groups)和其他类群组成.     

利用PCR-DGGE技术指导高温油藏中功能微生物的分离

采用PCR-DGGE技术对高温油藏的微生物群落进行了多样性分析,并将得到的信息用于指导油藏微生物的分离.本研究通过PCR-DGGE技术对油藏水样中DNA的16S rDNA V3、V8、V9 3个高可变区的扩增产物进行了比较,确定采用可得到更多微生物多样性信息的V9区引物进行PCR扩增,优势条带序列分析表明,在高温油藏中存在的微生物与GenBank数据库中α,β,y-变形杆菌和芽孢杆菌的序列相似性最高.利用多元细菌培养技术,以序列信息为指导,采用富集培养、直接培养和特殊培养的方法,从水样中分离出5株高温菌(而传统分离方法只能获得3株),其中3株高温解烃菌分别属于Bacillus属、Geobacillus属和Petrobacter属,它们能够在55℃以上兼性厌氧条件良好生长,对原油的降解率分别为56.5%、70.01%和31.87%,对原油的降粘率分别为40%、54.55%和29.09%,使原油的凝固点分别降低3.7、5.2和3.1℃.因此,序列指导和改变培养条件是分离更多有效采油微生物的改进方法,这3株高温菌的对原油的作用效果证明其具备提高石油采收率的潜力.     

太湖湖滨带水向辐射带水生植物多样性及生境因子分析

水生植物是湖滨带水向辐射带的重要组成部分。基于2021年春季和夏季太湖湖滨带水向辐射带内水生植物调查结果,采用相关性分析和典范对应分析等方法,对水生植物分布及相应生境因子进行分析,探究其演变特征。结果表明：1）2次调查共记录太湖湖滨带水向辐射带内水生植物12种,分别属于7科9属,其中浮叶植物3种,沉水植物9种;2）相关性分析结果表明,春季水生植物群落的Alpha多样性指数与生境因子相关性不显著,夏季水生植物群落的Alpha多样性指数与水体溶解氧浓度呈极显著正相关,与透明度、透明度/水深（透明度和水深的比值）呈显著正相关,Shannon-Wiener指数、Margelef指数与总氮浓度呈显著负相关;3）典范对应分析结果表明,水体溶解氧浓度对调查结果的解释值最大,随后依次为透明度/水深以及水深。10个生境因子对水生植物分布的总解释值较低;4）与历史数据相比,本次调查结果的水生植物多样性降低且优势物种单一化,表明太湖湖滨带水向辐射带内水生植物群落结构和多样性水平受人类活动和环境因子影响较大。水生植物自然恢复周期较长,人为调控其群落结构和多样性是当前推动太湖水生植物恢复的关键。

     

太湖不同营养水平湖区表层沉积物的砷分布特征及其生态风险

以太湖不同营养水平湖区为研究对象,采用改进的砷(As)形态连续提取法对表层沉积物中As的化学形态进行分析研究,探讨了沉积物中总砷(TAs)和As形态的分布特征及其与沉积物中营养盐和总有机碳(TOC)的相关性,并利用潜在生态风险评价(Eir)和风险指数编码法(RAC)评估了各湖区沉积物中As的生态风险水平.结果表明,各湖区表层沉积物中TAs的平均含量约为14.23~16.59 mg·kg~(-1),其中,竺山湾的TAs平均含量相对最高.As形态表现出明显的空间分布特征,其中,北部富营养湖区(竺山湾、梅梁湾、贡湖湾)中的有效态As(非专性吸附态和专性吸附态)与潜在有效态As(无定形氧化铁结合态、晶体形氧化铁结合态、有机结合态)的含量与百分比均高于中营养水平的南太湖,而北部湖区的残渣态As含量则低于南太湖.Pearson相关分析结果显示,除晶体形氧化铁结合态As和残渣态As外,沉积物总氮(TN)、总磷(TP)和TOC与其他As形态均存在显著的正相关关系.潜在生态风险评价结果表明,各湖区沉积物TAs均处于低风险;而RAC评价结果表明,各湖区沉积物的有效态As基本处于中等风险水平,且北部湖区的RAC指数均明显高于南太湖.     

Methanogenic community dynamics in anaerobic co-digestion of fruit and vegetable waste and food waste

A lab-scale continuously-stirred tank reactor (CSTR), used for anaerobic co-digestion of fruit and vegetable waste (FVW) and food waste (FW) at different mixture ratios, was operated for 178 days at the organic loading rate of 3 kg VS (volatile solids)/(m³.day). The dynamics of the Archaeal community and the correlations between environmental variables and methanogenic community structure were analyzed by polymerase chain reactions - denaturing gradient gel electrophoresis (PCR-DGGE) and redundancy analysis (RDA), respectively. PCR-DGGE results demonstrated that the mixture ratio of FVW to FW altered the community composition of Archaea. As the FVW/FW ratio increased, Methanoculleus, Methanosaeta and Methanosarcina became the predominant methanogens in the community. Redundancy analysis results indicated that the shift of the methanogenic community was significantly correlated with the composition of acidogenic products and methane production yield. Different mixture ratios of substrates led to different compositions of intermediate metabolites, which may affect the methanogenic community. These results suggested that the analysis of microbial communities could be used to diagnose anaerobic processes.     

进水氨氮浓度对强化生物除磷(EBPR)系统除磷特性及微生物群落结构的影响

以富集聚磷菌的活性污泥为基础,研究了不同进水氨氮浓度对强化生物除磷(EBPR)系统的除磷效果、微生物胞内聚合物(PHA)代谢过程及微生物群落结构的影响.结果表明,SBR反应器运行6 d后,进水氨氮浓度为1、5、10、15和20 mg·L-1的系统除磷效率最终分别稳定在0、4%、80%、98%和65%左右.试验前后各系统...     

淀山湖浮游植物数量消长及其与环境因子的关系

根据2004年1月～2006年12月逐月测得的淀山湖浮游植物数量,以及水温、风力、透明度、氮磷营养盐、高锰酸盐指数等环境数据,运用Pearson相关分析和典范对应分析（CCA）对淀山湖浮游植物数量消长与环境因子的关系进行了探讨,以期为有效防治淀山湖藻类水华提供科学依据.相关分析表明,在已处于中-富营养及以上水平的淀山湖...     

MBR和CAS工艺污泥在贫营养培养条件下的微生物群落结构研究

采用聚合酶链式-变性梯度凝胶电泳（PCR-DGGE）技术,研究了膜生物反应器（MBR）和传统活性污泥工艺（CAS）反应器中微生物在贫营养条件下的总细菌群落结构.结果表明,在培养过程中,污泥的微生物种群经历了一个比较明显的变化过程,且以CAS污泥微生物种群的变化更为明显,演替过程中既有原始优势种群的消亡,又有新的优势种群...