土壤管理措施及环境因素对土壤微生物多样性影响研究进展

本文综述了土壤管理措施及环境因素对土壤微生物多样性影响的研究进展,并介绍了土壤微生物多样性的研究方法,土壤微生物多样性包括微生物物种多样性、遗传多样性和生态多样性。传统上,土壤微生物群落的分析依赖于培养技术,但使用该技术只能培养和分离出一部分土壤微生物群落。现在国际上普遍使用Biolog分析、磷脂脂肪酸(PLFA)分析和核酸分析等多种现代技术研究和表征土壤微生物多样性。土壤微生物多样性受土壤管理措施和多种环境因素的影响。农药可能使土壤微生物多样性减少或改变其结构和功能;施有机肥有利于维持土壤微生物的多样性及活性;但在施用无机肥的影响上目前的报道有矛盾之处。农业土壤减少耕作可能增加微生物多样性和生物量;轮作可能比单一栽培耕作更有利于维持土壤微生物的多样性及活性。土壤微生物多样性也受土壤有机质、植被、季节变化等因素的影响,且通常遭受干旱、过度放牧、营养缺乏等的胁迫作用。     

植被对土壤微生物群落结构的影响

研究了不同土壤及覆盖其上的植被与土壤微生物群落结构和多样性的关系．植被使土壤中的微生物种类更丰富,群落多样性更高．表层土壤微生物群落中没有明显的优势种群,种间竞争作用较弱．并介绍了研究土壤微生物群落的分子生物学方法．     

转基因作物对土壤微生物多样性影响的研究方法

土壤微生物是土壤生态系统的一个重要组成部分,对土壤中的生物化学循环起着不可替代的驱动作用。转基因作物在生长过程中会不可避免地与土壤微生物发生交流,开展转基因作物对土壤微生物群落影响的研究对于科学评价转基因作物的潜在风险具有重要意义。随着现代生物技术的不断发展,土壤微生物多样性及其分析方法已经从传统的分离培养发展到从种群角度去研究整个土壤微生态系统内的微生物。但是由于土壤微生物的各种特性(如大部分不可培养、体积微小及群体效应等)和仪器设备检测性能的局限性,单一的研究方法会存在一些弊端,还需要结合其他的手段共同研究土壤生态系统中的微生物多样性。目前,人们对土壤微生物多样性的研究主要包括物种多样性、功能多样性、结构多样性及遗传多样性等4个方面,国内外关于转基因作物对土壤微生物多样性及其群落结构影响的研究方法很多,对常见的研究方法进行了总结,并提出了今后转基因作物对土壤微生物多样性及群落结构影响的研究策略。     

放射性污染对铀矿区土壤微生物群落结构的影响

为探究放射性及其伴生污染对铀矿区附近土壤微生物群落功能多样性的影响，采集退役铀矿冶周围不同辐射值的土壤为研究对象，通过测定其土壤理化性质、土壤可培养微生物计数及Biolog-ECO微孔板培养，分析土壤放射性污染程度及理化性质对土壤微生物群落功能多样性和数量的影响。结果表明，矿区放射性污染土壤中重金属含量普遍较高，尤其是铅已超出管控标准。随着辐射值的增加，土壤中可培养微生物数量显著下降，特别是放线菌和真菌的数量；微生物群落功能多样性指数、物种丰富度指数、优势度、均一性指数显著下降；对碳源利用也逐渐下降，尤其是对酚酸类和胺类碳源，利用率分别下降25.8%、29.7%。RDA分析发现放射性核素是该区域驱动土壤微生物群落代谢发生变化的主要因素。铀矿区放射性污染及伴随的重金属污染明显改变土壤微生物群落结构，抑制了土壤微生物群落的代谢活性，造成土壤微生物群落功能多样性下降。     

土壤微生物群落多样性解析法:从培养到非培养

土壤微生物群落多样性是土壤微生物生态学和环境科学的重点研究内容之一.传统的土壤微生物群落多样性解析技术是指纯培养分离法(平板分离和形态分析法以及群落水平生理学指纹法).后来,研究者们建立了多样性评价较为客观的生物标记法(磷脂脂肪酸法和呼吸醌指纹法).随着土壤基因组提取技术和基因片段扩增(PCR)技术的发展,大量的现代分子生物学技术不断地涌现并极大地推动了土壤微生物群落多样性的研究进程.这些技术主要包括:G+C％含量、DNA复性动力学、核酸杂交法(FISH和DNA芯片技术)、土壤宏基因组学以及DNA指纹图谱技术等.综述了这些技术的基本原理、比较了各种技术的优缺点并且介绍了他们在土壤微生物群落多样性研究中的应用,展望了这些技术的发展方向.     

凋落物对土壤有机碳与微生物功能多样性的影响

森林凋落物是影响土壤微生物群落和有机碳含量的重要因素,但其作用的程度和机制尚不清楚,研究该问题对于分析森林生态系统碳循环和资源管理具有重要意义。研究凋落物去除与添加处理下土壤有机碳含量与土壤微生物对碳源利用的差异,明确凋落物去除与添加对土壤微生物群落代谢功能及其多样性的影响,探究不同处理下SOC含量变化的土壤微生物群落代谢机理。选取承德市雾灵山1405-1435 m海拔范围内核桃楸-蒙古栎混交林的表层土壤,采用室内培养结合Biolog-ECO方法,测定了培养第21天的土壤有机碳（soil organic carbon,SOC）含量及微生物群落的AWCD值、Shannon-Wiener多样性指数、Simpson优势度指数、McIntosh均匀度指数、Pielou丰富度指数,分析培养期内凋落物的不同处理下SOC含量与微生物功能多样性的变化特征。结果表明：1）不同凋落物处理对SOC含量与土壤微生物群落多样性具有显著影响（P<0.05）,DL > HL > NL > CK;2）不同凋落物处理下土壤微生物群落代谢活性和土壤微生物对碳源的利用程度具有显著差异（P<0.05）,碳水化合物类和氨基酸类是土壤微生物的主要碳源;3）不同处理的SOC含量与土壤微生物多样性具有正相关关系。双倍凋落物添加在短期内对土壤微生物多样性影响难以达到显著水平且在一定程度上对土壤微生物的代谢活性具有抑制作用,土壤微生物群落功能多样性对SOC含量具有重要影响。     

啶虫脒污染下土壤微生物多样性

避开传统的分离培养过程,采用现代分子生物学方法探讨了杀虫剂啶虫脒污染条件下旱地土壤微生物种群多样性.通过对不同培养时间、不同浓度啶虫脒污染下旱地土壤微生物进行DGGE基因多样性的分子指纹图谱分析,发现随着培养时间不同,各处理之间的土壤微生物基因多样性出现了一定的差异.但在整个试验过程中,正常田间使用浓度（0.5mg kg^-1干土）的啶虫脒对土壤微生物群落的影响不明显,DGGE图谱条带与对照没有明显差异,土壤微生物基因多样性没有明显下降,这说明在旱地中使用正常田间浓度的啶虫脒不会对微生物群落造成较大的影响,高浓度啶虫脒对土壤微生物群落基因多样性有一定的影响,但是影响时间不长.在培养第五周时,浓度为5 mg kg^-1干土的土样出现了特异性条带,为对照所没有,其他处理浓度染色暗淡.经序列比对分析,与来自土壤的Uncultured bacterium具有100﹪的相似率,可能为不可培养或未培养过的细菌种.     

PCR-DGGE和FAMEs技术在土壤微生物多态性研究中的应用

将PCR-DGGE和FAMEs等生物技术新方法用于土壤微生物分析可有效的克服传统培养方法的局限性,为土壤微生物多样性、群落结构及动态变化的研究提供了更全面、可靠的方法,受到了国内外研究者的重视.本文综述了PCR-DGGE和FAMEs技术的原理及其在土壤微生物多样性、群落结构及动态变化方面的应用效果,指出了实际应用存在的问题.     

农用化学品污染对土壤微生物群落DNA序列多样性影响研究

采用ＲＡＰＤ分子遗传标记技术研究了农用化学品不同使用环境下的４种土壤微生物群落ＤＮＡ序列多样性的变化。结果表明,４种土壤微生物群落ＤＮＡ序列在其丰富度、多样性指数、均匀度等方面均存在差异;农用化学品的使用会对土壤微生物群落在ＤＮＡ分子水平上的多样性产生影响;而冰同的农用化学品对土壤微生物群落ＤＮＡ序列多样性影响各不相同：化肥污染会引起某些土壤微生物的富集和一些微生物物种的丧失;农药杂会引起土壤微生     

农业土壤微生物基因与群落多样性研究进展

介绍了群落基因组多样性、结构多样性与功能多样性相互关系的研究方法 ,重点论述了近年来农业土壤微生物群落遗传、结构与功能多样性的研究进展。同时总结了耕作措施和养分管理对农业土壤微生物群落多样性的影响 ,提出微生物序列分析、比较基因组学和微生物芯片技术与传统研究技术结合将有助于对微生物群落结构与功能和生物与环境因素对土壤微生物群落影响的深刻理解     

污染土壤微生物群落结构多样性及功能多样性测定方法

土壤微生物在促进土壤质量和植物健康方面发挥着重要的作用,土壤微生物群落结构和组成的多样性及其变化在一定程度上反映了土壤质量.为了更好地了解土壤健康状况,非常有必要发展有效的方法来研究污染土壤微生物的多样性、分布以及行为等.回顾了近年来国内外污染土壤微生物群落结构多样性及功能多样性的测定方法,包括生物化学技术和分子生物学技术,现将它们的原理、优缺点、实用性及其发展动态作一阐述,同时指出结合这两种技术可为微生物群落分析提供一个更全面的、精确的方法.     

Fatty acid methyl ester (FAME) profiles as measures of soil microbial community structure

Analysis of fatty acid methyl ester (FAME) profiles extracted from soils is a rapid and inexpensive procedure that holds great promise in describing soil microbial community structure without traditional reliance on selective culturing, which seems to severely underestimate community diversity. Interpretation of FAME profiles from environmental samples can be difficult because many fatty acids are common to different microorganisms and many fatty acids are extracted from each soil sample. We used principal components (PCA) and cluster analyses to identify similarities and differences among soil microbial communities described using FAME profiles. We also used PCA to identify particular FAMEs that characterized soil sample clusters. Fatty acids that are found only or primarily in particular microbial taxa-marker fatty acids-were used in conjunction with these analyses. We found that the majority of 162 soil samples taken from a conventionally-tilled corn field had similar FAME profiles but that about 20% of samples seemed to have relatively low, and that about 10% had relatively high, bacterial:fungal ratios. Using semivariance analysis we identified 21:0 iso as a new marker fatty acid. Concurrent use of geostatistical and FAME analyses may be a powerful means of revealing other potential marker FAMEs. When microbial communities from the same samples were cultured on R2A agar and their FAME profiles analyzed, there were many differences between FAME profiles of soil and plated communities, indicating that profiles of FAMEs extracted from soil reveal portions of the microbial community not culturable on R2A. When subjected to PCA, however, a small number of plated communities were found to be distinct due to some of the same profile characteristics (high in 12:0 iso, 15:0 and 17:1 ante A) that identified soil community FAME profiles as distinct. Semivariance analysis indicated that spatial distributions of soil microbial populations are maintained in a portion of the microbial community that is selected on laboratory media. These similarities between whole soil and plated community FAME profiles suggest that plated communities are not solely the result of selection by the growth medium, but reflect the distribution, in situ, of the dominant, culturable soil microbial populations.     

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。     

Diversity and seasonal fluctuations of the dominant members of the bacterial soil community in a wheat field as determined by cultivation and molecular methods

There is a paucity of knowledge on microbial community diversity and naturally occurring seasonal variations in agricultural soil. For this purpose the soil microbial community of a wheat field on an experimental farm in The Netherlands was studied by using both cultivation-based and molecule-based methods. Samples were taken in the different seasons over a 1-year period. Fatty acid-based typing of bacterial isolates obtained via plating revealed a diverse community of mainly gram-positive bacteria, and only a few isolates appeared to belong to the Proteobacteria and green sulfur bacteria. Some genera, such as Micrococcus, Arthrobacter, and Corynebacterium were detected throughout the year, while Bacillus was found only in July. Isolate diversity was lowest in July, and the most abundant species, Arthrobacter oxydans, and members of the genus Pseudomonas were found in reduced numbers in July. Analysis by molecular techniques showed that diversity of cloned 16S ribosomal DNA (rDNA) sequences was greater than the diversity among cultured isolates. Moreover, based on analysis of 16S rDNA sequences, there was a more even distribution among five main divisions, Acidobacterium, Proteobacteria, Nitrospira, cyanobacteria, and green sulfur bacteria. No clones were found belonging to the gram-positive bacteria, which dominated the cultured isolates. Seasonal fluctuations were assessed by denaturing gradient gel electrophoresis. Statistical analysis of the banding patterns revealed significant differences between samples taken in different seasons. Cluster analysis of the patterns revealed that the bacterial community in July clearly differed from those in the other months. Although the molecule- and cultivation-based methods allowed the detection of different parts of the bacterial community, results from both methods indicated that the community present in July showed the largest difference from the communities of the other months. Efforts were made to use the sequence data for providing insight into more general ecological relationships. Based on the distribution of 16S rDNA sequences among the bacterial divisions found in this work and in literature, it is suggested that the ratio between the number of Proteobacteria and Acidobacterium organisms might be indicative of the trophic level of the soil.     

Do Botanical Pesticides Alter the Structure of the Soil Microbial Community?

The effects of synthetic pesticides on the soil microbial community have been thoroughly investigated in the past mostly by culture-dependent methods and only few recent studies have used culture-independent approaches for this purpose. However, it should be noted that most of these studies have been conducted in microcosms where the soil microbial community is exposed to unrealistic concentrations of the pesticides, providing an unrealistic exposure scheme for soil microorganism. On the other hand, little is known regarding the potential impact of botanical pesticides on the soil microbial community. Therefore, a laboratory study and a field study were conducted to investigate the effects of synthetic (metham sodium [MS], sodium tetrathiocarbonate [SoTe], and fosthiazate) and botanical pesticides (azadirachtin, quillaja, and pulverized Melia azedarach fruits [PMF]) on the soil microbial community using phospholipid fatty acids (PLFA) analysis. Principal component analysis (PCA) on the results of the laboratory study indicated that the application of PMF resulted in significant changes in the soil microbial community. This was obvious by the proportional increase in the abundance of fatty acids 18:1ω9cis, 18:1ω9trans, which are common in gram-negative bacteria and saprotrophic fungi, and 18:2ω6,9, which is a fungal indicator. This response was attributed to the release of copious amounts of organic carbon and nutrients in the soil by the PMF. On the other hand, MS inhibited fungi and gram-negative bacteria, while fosthiazate and the botanical pesticides quillaja and azadirachtin did not impose significant changes in the soil microbial community. Similar results were obtained by the field study where application of the fumigants MS and SoTe significantly altered the structure of the soil microbial community with the former having a more prominent effect. Fosthiazate imposed mild changes in the soil microbial community, whereas quillaja and azadirachtin again did not show a significant effect. Overall, botanical pesticides, at their recommended dose, did not alter the structure of the soil microbial community compared to synthetic nonfumigant and fumigant pesticides which induced significant changes.     

PCR-DGGE技术在农田土壤微生物多样性研究中的应用

变性梯度凝胶电泳技术(DGGE)在微生物生态学领域有着广泛的应用。研究采用化学裂解法直接提取出不同农田土壤微生物基因组DNA,并以此基因组DNA为模板,选择特异性引物F357GC和R515对16S rRNA基因的V3区进行扩增,长约230bp的PCR产物经变性梯度凝胶电泳(DGGE)进行分离后,得到不同数目且分离效果较好的电泳条带。结果说明,DGGE能够对土壤样品中的不同微生物的16S rRNA基因的V3区的DNA扩增片断进行分离,为这些DNA片断的定性和鉴定提供了条件。与传统的平板培养方法相比,变性梯度凝胶电泳(DGGE)技术能够更精确的反映出土壤微生物多样性,它是一种有效的微生物多样性研究技术。     

Culturing captures members of the soil rare biosphere

The ecological significance of rare microorganisms within microbial communities remains an important, unanswered question. Microorganisms of extremely low abundance (the 'rare biosphere') are believed to be largely inaccessible and unknown. To understand the structure of complex environmental microbial communities, including the representation of rare and prevalent community members, we coupled traditional cultivation with pyrosequencing. We compared cultured and uncultured bacterial members of the same agricultural soil, including eight locations within one apple orchard and four time points. Our analysis revealed that soil bacteria captured by culturing were in very low abundance or absent in the culture-independent community, demonstrating unexpected accessibility of the rare biosphere by culturing.     

Effects of elevated atmospheric CO2 concentrations on soil microorganisms

Effects of elevated CO(2) on soil microorganisms are known to be mediated by various interactions with plants, for which such effects are relatively poorly documented. In this review, we summarize and synthesize results from studies assessing impacts of elevated CO(2) on soil ecosystems, focusing primarily on plants and a variety the of microbial processes. The processes considered include changes in microbial biomass of C and N, microbial number, respiration rates, organic matter decomposition, soil enzyme activities, microbial community composition, and functional groups of bacteria mediating trace gas emission such as methane and nitrous oxide. Elevated CO(2) in atmosphere may enhance certain microbial processes such as CH(4) emission from wetlands due to enhanced carbon supply from plants. However, responses of extracellular enzyme activities and microbial community structure are still controversy, because interferences with other factors such as the types of plants, nutrient availabilitial in soil, soil types, analysis methods, and types of CO(2) fumigation systems are not fully understood.