Impacts of 3 years of elevated atmospheric CO2 on rhizosphere carbon flow and microbial community dynamics

Carbon (C) uptake by terrestrial ecosystems represents an important option for partially mitigating anthropogenic CO₂ emissions. Short‐term atmospheric elevated CO₂ exposure has been shown to create major shifts in C flow routes and diversity of the active soil‐borne microbial community. Long‐term increases in CO₂ have been hypothesized to have subtle effects due to the potential adaptation of soil microorganism to the increased flow of organic C. Here, we studied the effects of prolonged elevated atmospheric CO₂ exposure on microbial C flow and microbial communities in the rhizosphere. Carex arenaria (a nonmycorrhizal plant species) and Festuca rubra (a mycorrhizal plant species) were grown at defined atmospheric conditions differing in CO₂ concentration (350 and 700 ppm) for 3 years. During this period, C flow was assessed repeatedly (after 6 months, 1, 2, and 3 years) by ¹³C pulse‐chase experiments, and label was tracked through the rhizosphere bacterial, general fungal, and arbuscular mycorrhizal fungal (AMF) communities. Fatty acid biomarker analyses and RNA‐stable isotope probing (RNA‐SIP), in combination with real‐time PCR and PCR‐DGGE, were used to examine microbial community dynamics and abundance. Throughout the experiment the influence of elevated CO₂ was highly plant dependent, with the mycorrhizal plant exerting a greater influence on both bacterial and fungal communities. Biomarker data confirmed that rhizodeposited C was first processed by AMF and subsequently transferred to bacterial and fungal communities in the rhizosphere soil. Over the course of 3 years, elevated CO₂ caused a continuous increase in the ¹³C enrichment retained in AMF and an increasing delay in the transfer of C to the bacterial community. These results show that, not only do elevated atmospheric CO₂ conditions induce changes in rhizosphere C flow and dynamics but also continue to develop over multiple seasons, thereby affecting terrestrial ecosystems C utilization processes.     

Rhizodeposition shapes rhizosphere microbial community structure in organic soil

The aims of the study were to determine group specificity in microbial utilization of root-exudate compounds and whole rhizodeposition; quantify the proportions of carbon acquired by microbial groups from soil organic matter and rhizodeposition, respectively; and assess the importance of root-derived C as a driver of soil microbial community structure. Additions of 13C-labelled root-exudate compounds to organic soil and steady-state labelling of Lolium perenne, coupled to compound-specific isotope ratio mass spectrometry, were used to quantify group-specific microbial utilization of rhizodeposition. Microbial utilization of glucose and fumaric acid was widespread through the microbial community, but glycine was utilized by a narrower range of populations, as indicated by the enrichment of phospholipid fatty acid (PLFA) analysis fractions. In L. perenne rhizospheres, high rates of rhizodeposit utilization by microbial groups showed good correspondence with increased abundance of these groups in the rhizosphere. Although rhizodeposition was not the quantitatively dominant C source for microbes in L. perenne rhizospheres, relative utilization of this C source was an important driver of microbial group abundance in organic soil.     

Carbon allocation mediated by arbuscular mycorrhizal fungi alters the soil microbial community under various phosphorus levels

Studies have shown that arbuscular mycorrhizal fungi (AMF) can shape the rhizosphere microbial community of the host plant, but the underlying mechanisms are unclear. Here, we tested the hypotheses that AMF may affect the rhizosphere microbial community by mediating carbon (C) allocation of the host plant, and that this mediation may be modulated by the soil phosphorus (P) level. Using a split-root system, we conducted a microcosm experiment with three main effects (soil P level: 5 or 25 mg kg⁻¹; AMF: with or without inoculation; and spatial niche (i.e., rhizosphere, hyphosphere, and mycorrhizosphere). Host plant growth benefited from AMF under low soil-P conditions. ¹³CO₂ isotope labeling showed that AMF increased C allocation to the colonized root and AMF mycelia under low-P conditions, which promoted AMF growth. ¹³C-DNA-SIP and 16S rRNA sequencing further indicated that the enhanced C allocation from the host altered the soil microbial community. Our results suggest that AMF enhances the C allocation of the host plant below ground, which can shape microbial community composition. These AMF effects were greater with a low than with a high level of soil P.     

Bacterial diversity in the rhizosphere of Proteaceae species

The Cape Floral Kingdom is an area of unique plant biodiversity in South Africa with exceptional concentrations of rare and endemic species and experiencing drastic habitat loss. Here we present the first molecular study of the microbial diversity associated with the rhizosphere soil of endemic plants of the Proteaceae family (Leucospermum truncatulum and Leucadendron xanthoconus). Genomic DNA was extracted from L. truncatulum rhizosphere soil, L. xanthoconus rhizosphere and non-rhizosphere soil and used as a template for the polymerase chain reaction (PCR) amplification of the 16S ribosomal RNA gene (rDNA). Construction and sequencing of 16S rDNA libraries revealed a high level of biodiversity and led to the identification of several novel bacterial phylotypes. The bacterial community profiles were compared by 16S rDNA denaturing gradient gel electrophoresis (DGGE). Cluster analysis and biodiversity indices revealed that the rhizosphere soil samples were more similar to each other than to non-rhizosphere soil and the rhizosphere soil contained a bacterial diversity that was richer and more equitable compared with non-rhizosphere soil. A Chloroflexus and an Azospirillum genospecies were restricted to the L. xanthoconus rhizosphere soil and Stenotrophomonas genospecies was identified in all rhizosphere soil samples but was not present in the non-rhizosphere soil. Taxon-specific nested PCR and DGGE-identified differences between the Proteaceae plant rhizosphere soil with a Frankia genospecies restricted the L. truncatulum rhizosphere. Archaea-specific rDNA PCR, DGGE and DNA sequencing revealed that Crenarcheote genospecies were excluded from the plant rhizosphere soil and only present in non-rhizosphere soil.     

Effects of site and plant species on rhizosphere community structure as revealed by molecular analysis of microbial guilds

The bacterial and fungal rhizosphere communities of strawberry (Fragaria ananassa Duch.) and oilseed rape (Brassica napus L.) were analysed using molecular fingerprints. We aimed to determine to what extent the structure of different microbial groups in the rhizosphere is influenced by plant species and sampling site. Total community DNA was extracted from bulk and rhizosphere soil taken from three sites in Germany in two consecutive years. Bacterial, fungal and group-specific (Alphaproteobacteria, Betaproteobacteria and Actinobacteria) primers were used to PCR-amplify 16S rRNA and 18S rRNA gene fragments from community DNA prior to denaturing gradient gel electrophoresis (DGGE) analysis. Bacterial fingerprints of soil DNA revealed a high number of equally abundant faint bands, while rhizosphere fingerprints displayed a higher proportion of dominant bands and reduced richness, suggesting selection of bacterial populations in this environment. Plant specificity was detected in the rhizosphere by bacterial and group-specific DGGE profiles. Different bulk soil community fingerprints were revealed for each sampling site. The plant species was a determinant factor in shaping similar actinobacterial communities in the strawberry rhizosphere from different sites in both years. Higher heterogeneity of DGGE profiles within soil and rhizosphere replicates was observed for the fungi. Plant-specific composition of fungal communities in the rhizosphere could also be detected, but not in all cases. Cloning and sequencing of 16S rRNA gene fragments obtained from dominant DGGE bands detected in the bacterial profiles of the Rostock site revealed that Streptomyces sp. and Rhizobium sp. were among the dominant ribotypes in the strawberry rhizosphere, while sequences from Arthrobacter sp. corresponded to dominant bands from oilseed rape bacterial fingerprints.     

番茄根际微生物种群动态变化及多样性

采用盆栽试验的方法对番茄根际主要微生物种群在不同生育期的动态变化进行了跟踪研究.结果表明,在番茄整个生育期内,可培养细菌数量在初花期和初果期时最多;放线菌数量从苗期到末期逐渐减少;真菌数量逐渐增多.番茄对细菌根际效应明显.DGGE图谱显示不同生育期番茄根际均具有较高的细菌多样性.根际细菌种类和数量在初花期发生较为显著的变化,初果期根际群落多样性指数(H)和物种丰度(S)值都达到最高,微生物最丰富,是筛选拮抗菌的较好时期.     

Links between plant and rhizoplane bacterial communities in grassland soils, characterized using molecular techniques

Molecular analysis of grassland rhizosphere soil has demonstrated complex and diverse bacterial communities, with resultant difficulties in detecting links between plant and bacterial communities. These studies have, however, analyzed "bulk" rhizosphere soil, rather than rhizoplane communities, which interact most closely with plants through utilization of root exudates. The aim of this study was to test the hypothesis that plant species was a major driver for bacterial rhizoplane community composition on individual plant roots. DNA extracted from individual roots was used to determine plant identity, by analysis of the plastid tRNA leucine (trnL) UAA gene intron, and plant-related bacterial communities. Bacterial communities were characterized by analysis of PCR-amplified 16S rRNA genes using two fingerprinting methods: terminal restriction fragment length polymorphisms (T-RFLP) and denaturing gradient gel electrophoresis (DGGE). Links between plant and bacterial rhizoplane communities could not be detected by visual examination of T-RFLP patterns or DGGE banding profiles. Statistical analysis of fingerprint patterns did not reveal a relationship between bacterial community composition and plant species but did demonstrate an influence of plant community composition. The data also indicated that topography and other, uncharacterized, environmental factors are important in driving bacterial community composition in grassland soils. T-RFLP had greater potential resolving power than DGGE, but findings from the two methods were not significantly different.     

Links between Plant and Rhizoplane Bacterial Communities in Grassland Soils, Characterized Using Molecular Techniques

Molecular analysis of grassland rhizosphere soil has demonstrated complex and diverse bacterial communities, with resultant difficulties in detecting links between plant and bacterial communities. These studies have, however, analyzed “bulk” rhizosphere soil, rather than rhizoplane communities, which interact most closely with plants through utilization of root exudates. The aim of this study was to test the hypothesis that plant species was a major driver for bacterial rhizoplane community composition on individual plant roots. DNA extracted from individual roots was used to determine plant identity, by analysis of the plastid tRNA leucine (trnL) UAA gene intron, and plant-related bacterial communities. Bacterial communities were characterized by analysis of PCR-amplified 16S rRNA genes using two fingerprinting methods: terminal restriction fragment length polymorphisms (T-RFLP) and denaturing gradient gel electrophoresis (DGGE). Links between plant and bacterial rhizoplane communities could not be detected by visual examination of T-RFLP patterns or DGGE banding profiles. Statistical analysis of fingerprint patterns did not reveal a relationship between bacterial community composition and plant species but did demonstrate an influence of plant community composition. The data also indicated that topography and other, uncharacterized, environmental factors are important in driving bacterial community composition in grassland soils. T-RFLP had greater potential resolving power than DGGE, but findings from the two methods were not significantly different.     

利用DGGE评价不同培养基回收番茄根际细菌类群的能力

用营养肉汤、YG、根系分泌物、土壤浸渍液4种培养基从番茄根际分离培养细菌,并结合变性梯度凝胶电泳(DGGE)技术,对4种培养基回收番茄根际细菌种群的能力进行了比较研究。结果表明,不同培养基和培养温度,回收到的细菌种群有一定差异;低营养浓度的YG培养基在较低的培养温度20℃下进行较长时间的培养,比高营养浓度营养肉汤培养基产生更多、更具代表性的细菌;以根系分泌物为基础的培养基从番茄根际回收到的优势菌群最多。该研究初步建立了用DGGE技术对不同培养基回收分离细菌种群能力进行评价的方法。     

Rhizosphere Microbial Community Structure in Relation to Root Location and Plant Iron Nutritional Status

Root exudate composition and quantity vary in relation to plant nutritional status, but the impact of the differences on rhizosphere microbial communities is not known. To examine this question, we performed an experiment with barley (Hordeum vulgare) plants under iron-limiting and iron-sufficient growth conditions. Plants were grown in an iron-limiting soil in root box microcosms. One-half of the plants were treated with foliar iron every day to inhibit phytosiderophore production and to alter root exudate composition. After 30 days, the bacterial communities associated with different root zones, including the primary root tips, nonelongating secondary root tips, sites of lateral root emergence, and older roots distal from the tip, were characterized by using 16S ribosomal DNA (rDNA) fingerprints generated by PCR-denaturing gradient gel electrophoresis (DGGE). Our results showed that the microbial communities associated with the different root locations produced many common 16S rDNA bands but that the communities could be distinguished by using correspondence analysis. Approximately 40% of the variation between communities could be attributed to plant iron nutritional status. A sequence analysis of clones generated from a single 16S rDNA band obtained at all of the root locations revealed that there were taxonomically different species in the same band, suggesting that the resolving power of DGGE for characterization of community structure at the species level is limited. Our results suggest that the bacterial communities in the rhizosphere are substantially different in different root zones and that a rhizosphere community may be altered by changes in root exudate composition caused by changes in plant iron nutritional status.     

Rhizosphere microbial community structure in relation to root location and plant iron nutritional status

Root exudate composition and quantity vary in relation to plant nutritional status, but the impact of the differences on rhizosphere microbial communities is not known. To examine this question, we performed an experiment with barley (Hordeum vulgare) plants under iron-limiting and iron-sufficient growth conditions. Plants were grown in an iron-limiting soil in root box microcosms. One-half of the plants were treated with foliar iron every day to inhibit phytosiderophore production and to alter root exudate composition. After 30 days, the bacterial communities associated with different root zones, including the primary root tips, nonelongating secondary root tips, sites of lateral root emergence, and older roots distal from the tip, were characterized by using 16S ribosomal DNA (rDNA) fingerprints generated by PCR-denaturing gradient gel electrophoresis (DGGE). Our results showed that the microbial communities associated with the different root locations produced many common 16S rDNA bands but that the communities could be distinguished by using correspondence analysis. Approximately 40% of the variation between communities could be attributed to plant iron nutritional status. A sequence analysis of clones generated from a single 16S rDNA band obtained at all of the root locations revealed that there were taxonomically different species in the same band, suggesting that the resolving power of DGGE for characterization of community structure at the species level is limited. Our results suggest that the bacterial communities in the rhizosphere are substantially different in different root zones and that a rhizosphere community may be altered by changes in root exudate composition caused by changes in plant iron nutritional status.     

典型农田土壤中丛枝菌根真菌-根际细菌互作及与氮磷利用的关系

在农业生态系统中,土壤微生物是土壤-作物系统养分循环的重要驱动力,其中丛枝菌根真菌(Arbuscularmycorrhizalfungi,AMF)能够促进作物对养分的吸收,适应逆境胁迫。【目的】进一步揭示AMF和根际细菌群落的跨界网络互作,挖掘与作物氮磷利用显著相关的关键微生物类群,揭示关键类群的生态网络特征。【方法】利用Illumina测序技术对3种典型农田旱地土壤(黑土、潮土和红壤)中AMF和根际细菌群落结构进行分析;构建互作网络并利用偏冗余分析、相关性分析探究了与氮磷利用相关的潜在关键类群。【结果】3种土壤中AMF与根际细菌均以正相互作用为主。不同土壤中AMF与根际细菌互作关系差异明显,在红壤中跨界互作最为密切,其中球囊霉属真菌(Glomus)与根际细菌中的放线菌(Actinobacteria)和变形菌(Proteobacteria)之间的交互作用最多。而在黑土中主要体现为根际细菌的界内互作。与氮磷利用率显著相关的关键微生物类群主要属于球囊霉属真菌、放线菌和α变形菌。【结论】典型旱地土壤中AMF与根际细菌的正相互关系对作物氮磷利用有潜在促进作用,关键类群在有机质和养分贫乏的红壤中可能起到更重要的作用。     

Resilience of the rhizosphere Pseudomonas and ammonia-oxidizing bacterial populations during phytoextraction of heavy metal polluted soil with poplar

We assessed the effects of phytoextraction on the dynamics of Pseudomonas spp. and ammonia-oxidizing bacterial populations in a heavy metal (HM) polluted soil. Hybrid poplars were grown in two-compartment root containers with a medium history (> 4 years) of HM pollution for 13 weeks. Bulk and poplar rhizosphere soils were analysed by denaturing gradient gel electrophoresis (DGGE) of Pseudomonas (sensu stricto) 16S rRNA and amoA gene fragments. DGGE patterns revealed that Pseudomonas and amoA-containing populations in the contaminated soils were markedly different from those in the uncontaminated soils. Pseudomonas and amoA profiles appeared to be stable over time in the bulk soils. In contrast, contaminated rhizosphere soils revealed a clear shift of populations with removal of HM becoming similar or at least shifted to the populations of the uncontaminated soils. The effect of phytoextraction was, however, not evident in the bulk samples, which still contained large amounts of HM. Cloning and sequencing of dominant DGGE bands revealed that Pseudomonas were phylogenetically related to the Pseudomonas fluorescens cluster and the amoA sequences to Nitrosospira spp. At the last sampling, major prominent band sequences from contaminated rhizosphere soils were identical to sequences obtained from uncontaminated rhizosphere soils, indicating that the populations were dominated by the same phylotypes. This study suggests that two taxonomically different populations are able to recover after the relief of HM stress by phytoextraction practices, whereas bulk microbial activities still remained depressed.     

辣椒青枯病罹病与健康植株根际土壤微生物群落多样性研究

系统研究和分析辣椒青枯病常发地发病与健康植株土壤微生物群落结构特征,为辣椒青枯病的绿色防治提供理论依据.基于16SrDNA基因高通量测序技术,对辣椒青枯病发病和健康植株根际土壤微生物群落结构和组成进行分析,同时采用biologyeco平板培养技术研究其土壤微生物群落代谢多样性和功能多样性的特征.结果表明,辣椒青枯病发病和健康植株根际土壤微生物群落组成之间存在显著差异,辣椒青枯病发病土壤的OTU为4566个,辣椒青枯病健康土壤的OTU为4167个.依据OTU所属细菌物种信息对土壤细菌群落结构进行分析,变形菌门在发病和健康土壤中均为优势细菌类群,其次为放线菌门类群.其中健康植株根际土壤中芽单胞菌门(Gemmatimonadetes)、装甲菌门(Armatimonadetes)的相对丰度比发病植株的分别高出了4.37,3.87倍,而发病植株根际土壤中厚壁菌门(Firmicutes)的相对丰度比健康植株的高出了3.87倍.辣椒青枯病发病土壤和健康土壤的土壤微生物代谢多样性也存在显著差异,同时,健康土壤中其微生物群落代谢得到显著增强,特别是对酚类化合物的利用显著增多,对辣椒抗病性存在显著的影响.研究表明,辣椒青枯病发病和健康植株根际土壤微生物群落组成和结构之间存在显著差异,并且健康土壤的微生物群落对酚类化合物的利用显著增强.     

Species-specific effects of polyploidisation and plant traits of Centaurea maculosa and Senecio inaequidens on rhizosphere microorganisms

Invasive plant species represent a threat to terrestrial ecosystems, but their effects on the soil biota and the mechanisms involved are not yet well understood. Many invasive species have undergone polyploidisation, leading to the coexistence of various cytotypes in the native range, whereas, in most cases, only one cytotype is present in the introduced range. Since genetic variation within a species can modify soil rhizosphere communities, we studied the effects of different cytotypes and ranges (native diploid, native tetraploid and introduced tetraploid) of Centaurea maculosa and Senecio inaequidens on microbial biomass carbon, rhizosphere total DNA content and bacterial communities of a standard soil in relation to plant functional traits. There was no overall significant difference in microbial biomass between cytotypes. The variation of rhizosphere total DNA content and bacterial community structure according to cytotype was species specific. The rhizosphere DNA content of S. inaequidens decreased with polyploidisation in the native range but did not vary for C. maculosa. In contrast, the bacterial community structure of C. maculosa was affected by polyploidisation and its diversity increased, whereas there was no significant change for S. inaequidens. Traits of S. inaequidens were correlated to the rhizosphere biota. Bacterial diversity and total DNA content were positively correlated with resource allocation to belowground growth and late flowering, whereas microbial biomass carbon was negatively correlated to investment in reproduction. There were no correlations between traits of the cytotypes of C. maculosa and corresponding rhizosphere soil biota. This study shows that polyploidisation may affect rhizosphere bacterial community composition, but that effects vary among plant species. Such changes may contribute to the success of invasive polyploid genotypes in the introduced range.     

Analysis of Bacterial Community in the Ginseng Soil Using Denaturing Gradient Gel Electrophoresis (DGGE)

The objective of this work was to determine the shifts in the PCR-DGGE profiles of bacterial communities associated with the rhizosphere soil of ginseng at varying age levels. Differences in the dominance of intense DNA bands in the DGGE profile was observed over the age of the plants indicating the fluctuation in the microbial community structure. The bacterial orders of actinomycetales of Actinobacteria and Spingomonadales and Rhizobiales of α-Proteobacteria were predominant in the ginseng soil.     

十字花科作物根肿病对根际土壤微生物群落的影响

为探究根肿病对十字花科作物根际土壤微生物多样性的影响,以罹病大白菜和健康株根际土壤为研究对象,采用高通量测序技术对2组样本的细菌16S rDNA和真菌ITS基因进行序列测定,分析了样本间的微生物群落结构和组成差异,同时测定根际土壤理化性质,探讨根肿病、土壤微生物群落、土壤环境因子三者的相关性。研究表明:1)患病植株根际土壤pH和总磷、总钾、碱解氮、速效钾含量显著低于正常植株根际土,而交换性钙含量明显增加。2)根肿病的发生降低了根际土壤中细菌种群的丰富度和多样性程度,但对根际土壤中的真菌α-多样性无明显影响。3)变形菌门、拟杆菌门、放线菌门等是所测土壤样本的主要优势细菌种群,其中患病植株根际土壤中拟杆菌门丰度显著高于健康植株根际土壤,放线菌门丰度则显著降低(P0.05)。优势细菌纲为γ-变形菌纲、拟杆菌纲、α-变形菌纲、放线菌纲、酸杆菌纲等,2组土壤样本间多种优势细菌纲相对丰度差异显著。4)根际土壤优势真菌类群为子囊菌门、被孢霉门、担子菌门和壶菌门,其相对丰度在患病和健康株根际土壤样本中均有明显差异。主要真菌纲为散囊菌纲、被孢霉纲、锤舌菌纲等,并且土壤样本间的多种优势真菌纲相对丰度存在显著性差异。5)主坐标分析结果表明病株根际土壤与健康株根际土壤细菌和真菌群落结构差异明显,冗余分析结果显示速效钾和交换性钙是根际土壤微生物群落变化的主要影响因素。研究结果为揭示根肿病发生的根际微生态机制以及研发根肿病综合防控技术提供理论支撑。     

Bulk and rhizosphere soil bacterial communities studied by denaturing gradient gel electrophoresis: plant-dependent enrichment and seasonal shifts revealed

The bacterial rhizosphere communities of three host plants of the pathogenic fungus Verticillium dahliae, field-grown strawberry (Fragaria ananassa Duch.), oilseed rape (Brassica napus L.), and potato (Solanum tuberosum L.), were analyzed. We aimed to determine the degree to which the rhizosphere effect is plant dependent and whether this effect would be increased by growing the same crops in two consecutive years. Rhizosphere or soil samples were taken five times over the vegetation periods. To allow a cultivation-independent analysis, total community DNA was extracted from the microbial pellet recovered from root or soil samples. 16S rDNA fragments amplified by PCR from soil or rhizosphere bacterium DNA were analyzed by denaturing gradient gel electrophoresis (DGGE). The DGGE fingerprints showed plant-dependent shifts in the relative abundance of bacterial populations in the rhizosphere which became more pronounced in the second year. DGGE patterns of oilseed rape and potato rhizosphere communities were more similar to each other than to the strawberry patterns. In both years seasonal shifts in the abundance and composition of the bacterial rhizosphere populations were observed. Independent of the plant species, the patterns of the first sampling times for both years were characterized by the absence of some of the bands which became dominant at the following sampling times. Bacillus megaterium and Arthrobacter sp. were found as predominant populations in bulk soils. Sequencing of dominant bands excised from the rhizosphere patterns revealed that 6 out of 10 bands resembled gram-positive bacteria. Nocardia populations were identified as strawberry-specific bands.     

Effect of phenylurea herbicides on soil microbial communities estimated by analysis of 16S rRNA gene fingerprints and community-level physiological profiles

The effect of three phenyl urea herbicides (diuron, linuron, and chlorotoluron) on soil microbial communities was studied by using soil samples with a 10-year history of treatment. Denaturing gradient gel electrophoresis (DGGE) was used for the analysis of 16S rRNA genes (16S rDNA). The degree of similarity between the 16S rDNA profiles of the communities was quantified by numerically analysing the DGGE band patterns. Similarity dendrograms showed that the microbial community structures of the herbicide-treated and nontreated soils were significantly different. Moreover, the bacterial diversity seemed to decrease in soils treated with urea herbicides, and sequence determination of several DGGE fragments showed that the most affected species in the soils treated with diuron and linuron belonged to an uncultivated bacterial group. As well as the 16S rDNA fingerprints, the substrate utilization patterns of the microbial communities were compared. Principal-component analysis performed on BIOLOG data showed that the functional abilities of the soil microbial communities were altered by the application of the herbicides. In addition, enrichment cultures of the different soils in medium with the urea herbicides as the sole carbon and nitrogen source showed that there was no difference between treated and nontreated soil in the rate of transformation of diuron and chlorotoluron but that there was a strong difference in the case of linuron. In the enrichment cultures with linuron-treated soil, linuron disappeared completely after 1 week whereas no significant transformation was observed in cultures inoculated with nontreated soil even after 4 weeks. In conclusion, this study showed that both the structure and metabolic potential of soil microbial communities were clearly affected by a long-term application of urea herbicides.