Multivariate Analyses of Burkholderia Species in Soil: Effect of Crop and Land Use History

The assessment of Burkholderia diversity in agricultural areas is important considering the potential use of this genus for agronomic and environmental applications. Therefore, the aim of this work was to ascertain how plant species and land use management drive the diversity of the genus Burkholderia. In a greenhouse experiment, different crops, i.e., maize, oat, barley, and grass, were planted in pots containing soils with different land use histories, i.e., maize monoculture, crop rotation, and permanent grassland, for three consecutive growth cycles. The diversity of Burkholderia spp. in the rhizosphere soil was assessed by genus-specific PCR-denaturing gradient gel electrophoresis (DGGE) and analyzed by canonical correspondence analysis (CCA). CCA ordination plots showed that previous land use was the main factor affecting the composition of the Burkholderia community. Although most variation in the Burkholderia community structure was observed between the permanent grassland and agricultural areas, differences between the crop rotation and maize monoculture groups were also observed. Plant species affected Burkholderia community structure to a lesser extent than did land use history. Similarities were observed between Burkholderia populations associated with maize and grass, on the one hand, and between those associated with barley and oat, on the other hand. Additionally, CCA ordination plots demonstrated that these two groups (maize/grass versus barley/oat) had a negative correlation. The identification of bands from the DGGE patterns demonstrated that the species correlated with the environmental variables were mainly affiliated with Burkholderia species that are commonly isolated from soil, in particular Burkholderia glathei, B. caledonica, B. hospita, and B. caribiensis.     

Rhizosphere microbial community and its response to plant species and soil history

The plant rhizosphere is a dynamic environment in which many parameters may influence the population structure, diversity and activity of the microbial community. Two important factors determining the structure of microbial community present in the vicinity of plant roots are plant species and soil type. In the present study we assessed the structure of microbial communities in response to four plant species (i.e. maize (Zea mays L.), oat (Avena sativa L.), barley (Hordeum vulgare L.) and commercial grass mix) planted in soil with different land use history (i.e. arable land under crop rotation, maize monoculture and permanent grassland). Both factors, plant species and land use history, showed clear effects on microbial community and diversity as determined by PCR-DGGE fingerprinting with universal and group-specific bacterial primers. Moreover, we explored the rhizosphere effect of these plant species on the abundance of bacterial antagonists of the potato pathogen Rhizoctonia solani AG3. The data showed that the abundance and taxonomic composition of antagonists differed clearly between the different plants. The highest percentages of antagonists were found in maize and grass rhizosphere. When antagonistic Pseudomonas populations were compared, the highest, abundance and diversity of antagonists were detected in barley and oat rhizospheres, as compared to maize and grass rhizosphere. The results obtained in our study demonstrate clearly that plant species and soil type are two important factors affecting the structure of total bacterial, Pseudomonas and Bacillus community.     

Effect of Agricultural Management Regime on Burkholderia Community Structure in Soil

The main objective of this study was to determine the Burkholderia community structure associated with areas under different agricultural management and to evaluate to which extent this community structure is affected by changes in agricultural management. Two fields with distinct soil history (arable land and permanent grassland) were exposed to three agricultural management regimes (crop rotation, maize monoculture, and grassland). By using a culture-independent approach, based on a Burkholderia-specific polymerase chain reaction–denaturing gradient gel electrophoresis system, it was possible to observe the conversion of Burkholderia communities typical for permanent grassland to those of arable land after four consecutive years. However, the time needed to achieve the reverse transition, i.e., converting the Burkholderia community associated with arable land to that of grassland, was beyond the duration of the field experiment. In addition, by applying principal response curves, the direction and extent of the conversion from grassland to arable land (maize monoculture and to crop rotation) were determined. Hence, the results suggested that agricultural practices, such as fertilization and tillage, were more effective in changing the Burkholderia community structure than agricultural management regime. To determine the effect of agricultural management on the Burkholderia population with biocontrol abilities, the culturable fraction of the Burkholderia community was assessed. The areas under permanent grassland and grassland converted to maize monoculture had the highest percentages of Burkholderia strains with antagonistic activity against Rhizoctonia solani AG-3, mainly Burkholderia pyrrocinia and Burkholderia sp. LMG 22929. The isolation frequency of antagonistic isolates from arable land was extremely low. Our results indicate that (changes in) agricultural management, mainly crop rotation, affect the frequency of isolation of antagonistic Burkholderia strains and that grassland represents a reservoir of Burkholderia species with great potential for agricultural applications.     

Molecular method to assess the diversity of Burkholderia species in environmental samples

In spite of the importance of many members of the genus Burkholderia in the soil microbial community, no direct method to assess the diversity of this genus has been developed so far. The aim of this work was the development of soil DNA-based PCR-denaturing gradient gel electrophoresis (DGGE), a powerful tool for studying the diversity of microbial communities, for detection and analysis of the Burkholderia diversity in soil samples. Primers specific for the genus Burkholderia were developed based on the 16S rRNA gene sequence and were evaluated in PCRs performed with genomic DNAs from Burkholderia and non-Burkholderia species as the templates. The primer system used exhibited good specificity and sensitivity for the majority of established species of the genus Burkholderia. DGGE analyses of the PCR products obtained showed that there were sufficient differences in migration behavior to distinguish the majority of the 14 Burkholderia species tested. Sequence analysis of amplicons generated with soil DNA exclusively revealed sequences affiliated with sequences of Burkholderia species, demonstrating that the PCR-DGGE method is suitable for studying the diversity of this genus in natural settings. A PCR-DGGE analysis of the Burkholderia communities in two grassland plots revealed differences in diversity mainly between bulk and rhizosphere soil samples; the communities in the latter samples produced more complex patterns.     

Long-term effects of crop management on Rhizobium leguminosarum biovar viciae populations

Little is known about factors that affect the indigenous populations of rhizobia in soils. We compared the abundance, diversity and genetic structure of Rhizobium leguminosarum biovar viciae populations in soils under different crop managements, i.e., wheat and maize monocultures, crop rotation, and permanent grassland. Rhizobial populations were sampled from nodules of pea- or vetch plants grown in soils collected at three geographically distant sites in France, each site comprising a plot under long-term maize monoculture. Molecular characterization of isolates was performed by PCR-restriction fragment length polymorphism of 16S-23S rDNA intergenic spacer as a neutral marker of the genomic background, and PCR-restriction fragment length 0polymorphism of a nodulation gene region, nodD, as a marker of the symbiotic function. The diversity, estimated by richness in types and Simpson's index, was consistently and remarkably lower in soils under maize monoculture than under the other soil managements at the three sites, except for the permanent grassland. The highest level of diversity was found under wheat monoculture. Nucleotide sequences of the main rDNA intergenic spacer types were determined and sequence analysis showed that the prevalent genotypes in the three maize fields were closely related. These results suggest that long-term maize monoculturing decreased the diversity of R. leguminosarum biovar viciae populations and favored a specific subgroup of genotypes, but the size of these populations was generally preserved. We also observed a shift in the distribution of the symbiotic genotypes within the populations under maize monoculture, but the diversity of the symbiotic genotypes was less affected than that of IGS types. The possible effect of such changes on biological nitrogen fixation remains unknown and this requires further investigation.     

A multivariate analysis of cropping effects on Irish ground beetle assemblages (Coleoptera: Carabidae) in mixed arable and grass farmland

The assemblage of carabid species trapped over a 3 yr period in eight separate but contiguous fields was analysed by Canonical Correspondence Analysis (CCA), Seven of the fields were subject to arable crop rotations involving a mixture of autumn and early spring‐sown cereals (wheat or barley), late spring‐sown crops (potatoes, sugar and fodder beet and maize) and short‐term uncultivated grass leys, The eighth field was an established ‘permanent’ grass pasture of at least 50 yrs standing, Pooled samples collected in the first half of the growing season (April‐June) showed clear evidence of soil cultivation effects on community structure, Systematic exclusion of samples and re‐analysis distinguished the fauna of firstly, old pasture samples, samples from ley pastures of increasing age and finally samples from fields with different times of soil cultivation, In the latter analysis, the main effects of soil cultivation were related to differences in ground cover over the winter and a direct effect of spring soil cultivation on autumn breeding populations, The ordination of pooled catches for the second half of the growing season (July‐September) could not be related to known year, field or crop cultivation variables, The underlying species ordination suggested that later in the summer the effect of crop cover on soil microclimate may mask cultivation effects by influencing the post‐emergence dispersal of autumn‐breeding populations and the reproductive success of spring‐breeders, The combination of earlier soil cultivation effects, and probably microclimatic influences later in the season, resulted in the strong distinction of whole season carabid catches from individual fields, It is concluded that the uniqueness of individual field histories may provide a mechanism to promote the co‐existence of ecologically similar species within the farmed landscape and enhance the abundance and bio‐diversity of species in the face of routine soil cultivation.     

Effects of agronomical measures on the microbial diversity of soils as related to the suppression of soil-borne plant pathogens

The diversity of soil microbial communities can be key to the capacity of soils tosuppress soil-borne plant diseases. As agricultural practice, as well as directedagronomical measures, are known to be able to affect soil microbial diversity, it isplausible that the soil microflora can be geared towards a greater suppressivity ofsoil-borne diseases as a result of the selection of suitable soil management regimes.In the context of a programme aimed at investigating the microbial diversity of soilsunder different agricultural regimes, including permanent grassland versus arableland under agricultural rotation, we assessed how soil microbial diversity is affectedin relation to the suppression of the soil-borne potato pathogen Rhizoctoniasolani AG3. The diversity in the microbial communities over about a growingseason was described by using cultivation-based – plating on different media – and cultivation-independent – soil DNA-based PCR followed by denaturing gradient gel electrophoresis (DGGE) community fingerprinting – methods. The results showed great diversity in the soil microbiota at both the culturable and cultivation-independent detection levels. Using cultivation methods, various differences between treatments with respect to sizes of bacterial and fungal populations were detected, with highest population sizes generally found in rhizospheres. In addition, the evenness of eco-physiologically differing bacterial types was higher in grassland than in arable land under rotation. At the cultivation-independent level, clear differences in the diversities of several microbial groups between permanent grassland and arable land under rotation were apparent. Bio-assays that assessed the growth of R. solani AG3 hyphae through soil indicated a greater growth suppression in grassland than in arable land soils. Similarly, an experiment performed in the glasshouse showed clear differences in both microbial diversities and suppressiveness of R. solani growth in soil, depending on the presence of either maizeor oats as the crop. The significance of these findings for designing soil managementstrategies is discussed.     

藏北高原草地群落的数量分类与排序

采用TWINSPAN数量分类和DCA、CCA排序的方法,对藏北高原草地29个样点进行统计分析。结果显示:(1)TWINSPAN数量分类将藏北高寒草地群落划分成10种类型。(2)样点DCA排序第一轴基本反映了水分环境梯度,第二轴基本反映了热量梯度。(3)TWINSPAN分类所划分的各群落在DCA排序图上都有各自的分布范围和界限,说明DCA排序能较好的反应各优势群落与其环境资源之间的关系。(4)样点CCA排序表明,影响群落分布的首要环境因子是水分因子(年均降水量)和空间因子(经度),其次是热量因子(年均温度),CCA排序进一步阐明了群落分布决定于水分和温度等环境因子,并间接验证了TWINSPAN的分类结果。(5)物种CCA排序和TWINSPAN分类结果表明:植物群落中物种的分布格局与植物群落类型的分布格局存在一定的相似性。     

青海湖鸟岛地区草地植物群落特征的研究

青海湖鸟岛地区草地群落共出现种子植物 6 3种 ,隶属 4 3属 ,19科 ,以北温带成分为主 ;从湖滨到典型地带性植被 ,物种多样性指数变化可以分为两大类型 ;随着高度增加 ,物种多样性呈现倒U型变化 ,沿着水平梯度物种多样性基本上逐渐增加。可以分为禾草、豆科固氮植物、非禾本科草类和半灌木功能组 ,物种水平与功能组水平的多样性、均匀度和丰富度相关系数分别为 0 2 79,0 4 36 ,0 0 19,均没有达到显著相关 ,功能组水平的多样性指数具有与物种水平不同的性质     

Communities of arbuscular mycorrhizal fungi in arable soils are not necessarily low in diversity

Communities of arbuscular mycorrhizal fungi (AMF) in five agricultural field sites of different management intensities were studied. Variable regions of the ribosomal RNA genes were used to detect and identify AMF directly within colonized roots. Roots from a continuous maize monoculture showed low AMF diversity, in agreement with previous reports on molecular diversity of AMF in agricultural soils. In contrast, a substantially higher diversity of AMF was found throughout the long term 'DOK' field experiment, where organic and conventional agricultural practices have been compared side by side since 1978. In this experiment, a 7-year crop rotation is performed under lower levels of inorganic fertilizer input and chemical pest control. These results are in good agreement with analyses of the spore community previously conducted in these field sites. In a third site, an organically managed leek field with soil of very high phosphate content reflecting the highly intensive conventional field history and intensive tillage, we detected a low-diversity community comparable to the maize monoculture. In addition to fungi from Glomus group A, which have previously been reported to dominate arable soils, we regularly found members of the genera Scutellospora, Paraglomus and Acaulospora. The genus Acaulospora was shown to occur more frequently early in the growing season, suggesting that the life history strategy of AMF may influence the active community at a given time. These data show that the diversity of AMF is not always low in arable soils. Furthermore, low-input agriculture involving crop rotation may provide better conditions to preserve AMF diversity, by preventing the selection for the few AMF taxa tolerating high nutrient levels.     

Restored meadow harvestman communities (Opiliones) in the Bílé Karpaty Protected Landscape Area, Czech Republic

Harvestman communities inhabiting plots treated differently for grassland restoration were investigated at the Vyzkum site near Malá Vrbka village (Bílé Karpaty Protected Landscape Area). Harvestman were sampled by pitfall trapping from 1999 to 2003 on plots sown with native haymeadow seed mixture, also on plots where narrow strips of regional seed mixtures were sown either within a matrix of commercial grass mixture or within vegetation cover in natural regeneration state and on plots in a natural regeneration state. Additionally, harvestman were collected in a field under permanent crop rotation and in a neighbouring xerothermic deciduous forest. In total, 5,086 individuals of harvestman representing 15 species from three families were obtained. Phalangium opilio was dominant (78%) and P. opilio, Rilaena triangularis and Zachaeus crista were the most frequent species. The results confirmed colonisation and subsequent development of harvestman communities on meadows in various state of restoration, including plots with spontaneous plant succession. Nevertheless, biotope character and successive formation of plant cover evidently influenced the structure of harvestman communities. The highest number of taxa (12) was recorded on plots with natural regeneration; the lowest one (9) was recorded in the field with permanent crop rotation. The highest values of diversity and equitability indices of harvestman communities were found in neighbouring forest habitats representing possible sources of harvestman migration.     

云雾山天然草地物种分布与环境因子的关系

应用数量生态学中的梯度分析方法研究了云雾山天然草地群落类型、物种分布以及与环境因子的关系.研究结果表明,作为重要的人为管理措施,封育通过改善草地的水分、养分等环境条件,显著影响了云雾山草地的物种分布和群落类型.在封育草地,植被类型为本氏针茅群落,未封育草地为铁杆蒿群落.本氏针茅、百里香、大针茅、直茎点地梅、阿尔泰狗哇花等中旱生植物在封育草地中占优势,铁杆蒿、硬质早熟禾、星毛委菱菜等在退化草地中占优势.除封育措施以外,20～100cm 的土壤含水量对物种分布影响显著.同时,养分作为一个重要的环境因素对物种的分布也具有重要影响,特别是速效钾和NO-3-N.海拔、坡位、坡向等地形因素对植物分布的影响不明显.     

Ground beetle species (Carabidae,Coleoptera) activity and richness in relation to crop type,fertility management and crop protection in a farm management comparison trial

Ground beetle activity and species richness was monitored using pitfall traps in a plot trial system on a farm in northern England where the effects of organic and conventional fertility and crop protection management were separated within different crop types between 2005 and 2008. As well as analyses on species activity the beetles were split into small, medium‐sized and large groups, and into groups of herbivores and specific Collembola feeders. Crop type had significant effects on the activity of the 20 most abundant species and all groups, generally with most in beans and winter barley and least in vegetables and spring barley. Most significant reactions to crop protection and fertility management were in cereals and grass/clover. Activity of small species was highest in conventionally crop‐protected cereals but not in vegetables, with more medium‐sized and herbivorous species in organic plots, but there was little influence of crop protection management on large and Collembola feeding species. However, large species were significantly more active in organically fertilised cereals and grass/clover, but not in vegetables, and there were more Collembola feeders in conventional cereals but not in grass/clover. Small species were more abundant in conventionally fertilised grass/clover but there were more in organic cereals and vegetables. These inconsistent activity reactions to management were also observed with individual species but most preferred organically managed plots. There were few significant crop protection : fertility management interactions. Species richness was also significantly affected by crop type and where management had an influence, more species were found in organically managed plots. Constrained ordination emphasised that ground beetle activity was influenced more by crops than by management. Given the diverse nature of organic crop rotations, crop type should be considered a major influence in any environmental manipulation aimed at increasing ground beetle activity for provision of ecosystem services.     

Predominant <Emphasis Type="Italic">Bacillus</Emphasis> spp. in Agricultural Soil under Different Management Regimes Detected via PCR-DGGE

A PCR system for studying the diversity of species of Bacillus and related taxa directly from soil was developed. For this purpose, a specific 24-bp forward primer located around position 110 of the 16S ribosomal RNA gene was designed and combined with a reverse bacterial primer located at the end of the gene. The specificity of this PCR system for bacilli and related taxons was confirmed on the basis of tests with diverse strains as well as with soil DNA. Analysis of a soil DNA derived clone library showed that the amplified fragments affiliated exclusively with sequences of gram-positive bacteria, with up to 95% of the sequences originating from putative Bacillus species. In particular, sequences affiliated to those of B. mycoides, B. pumilus, B. megaterium, B. thuringiensis, and B. firmus, as well as to related taxa such as Paenibacillus, were obtained. A minority, i.e., less than 6%, of the clones affiliated with other gram-positive bacteria, such as Arthrobacter spp., Frankia spp., and uncultured gram-positives. The amplified fragments were used as templates for a second PCR using bacterial 16S rDNA primers, yielding PCR products of about 410 bp, which were separated by denaturing gradient gel electrophoresis (DGGE). Amplicons indicating Bacillus spp. were found in the gel between 45% and roughly 60% denaturant, whereas those representing other, high-G+C% bacteria, were localized in gel regions with denaturant concentrations exceeding about 60%, thus allowing the distinction between these two groups of sequences. We applied this system to compare the group-specific diversity in bacterial communities in an agricultural soil under different regimes, i.e., permanent grassland, grassland recently turned to arable land, and arable land under agricultural rotation. Differences in the Bacillus-related community structures between the treatments were clearly detected. Higher diversities, as judged by Shannon–Weaver indices calculated on the basis of the molecular profiles, were consistently observed in the permanent grassland and the grassland turned into arable land, as compared to the arable land.     

Crop monoculture rather than agriculture reduces the spatial turnover of soil bacterial communities at a regional scale

The goal of this study was to investigate the spatial turnover of soil bacterial communities in response to environmental changes introduced by the practices of soybean monoculture or crop rotations, relative to grassland soils. Amplicon sequencing of the 16S rRNA gene was used to analyse bacterial diversity in producer fields through three successive cropping cycles within one and a half years, across a regional scale of the Argentinean Pampas. Unlike local diversity, which was not significantly affected by land use type, agricultural management had a strong influence on β‐diversity patterns. Distributions of pairwise distances between all soils samples under soybean monoculture had significantly lower β‐diversity and narrower breadth compared with distributions of pairwise distances between soils managed with crop rotation. Interestingly, good agricultural practices had similar degree of β‐diversity as natural grasslands. The higher phylogenetic relatedness of bacterial communities in soils under monoculture across the region was likely determined by the observed loss of endemic species, and affected mostly to phyla with low regional diversity, such as Acidobacteria, Verrucomicrobia and the candidates phyla SPAM and WS3. These results suggest that the implementation of good agricultural practices, including crop rotation, may be critical for the long‐term conservation of soil biodiversity.     

Characteristics of a Pythium arrhenomanes with High Frequency in Barley Soils

Soils from two long-term crop rotation experiments were examined for incidence of root pathogens with a test tube method, where a great number (hundreds) of small portions (15–68g) of soil were biotested. There was a 4–5 times higher frequency of a root-infecting Pythium sp. In barley monoculture soil when compared to crop rotation soil, where winter turnip rape was the preceding crop. In pathogenicity tests the isolated pathogen caused severe root rot on barley, wheat and rye, but did not affect growth of oats, maize, peas and winter rape. In all essential morphological characters it resembles P. arrhenomanes and we classify it as belonging to this species.     

Changes in Soil Microbial Biomass and Residual Indices as Ecological Indicators of Land Use Change in Temperate Permanent Grassland

The relationship between microbial biomass, residues and their contribution to microbial turnover is important to understand ecosystem C storage. The effects of permanent grassland (100 % ryegrass—PG), conversion to modified grassland (mixture of grass and clover—MG) or maize monoculture (MM) on the dynamics of soil organic C (SOC), microbial biomass, fungal ergosterol and microbial residues (bacterial muramic acid and fungal glucosamine) were investigated. Cattle slurry was applied to quantify the effects of fertilisation on microbial residues and functional diversity of microbial community across land use types. Slurry application significantly increased the stocks of microbial biomass C and S and especially led to a shift in microbial residues towards bacterial tissue. The MM treatment decreased the stocks of SOC, microbial biomass C, N and S and microbial residues compared with the PG and MG treatments at 0–40 cm depth. The MM treatment led to a greater accumulation of saprotrophic fungi, as indicated by the higher ergosterol-to-microbial biomass C ratio and lower microbial biomass C/S ratio compared with the grassland treatments. The absence of a white clover population in the PG treatment caused a greater accumulation of fungal residues (presumably arbuscular mycorrhizal fungi (AMF), which do not contain ergosterol but glucosamine), as indicated by the significantly higher fungal C-to-bacterial C ratio and lower ergosterol-to-microbial biomass C ratio compared with the MG treatment. In addition to these microbial biomass and residual indices, the community level physiological profiles (CLPP) demonstrated distinct differences between the PG and MG treatments, suggesting the potential of these measurements to act as an integrative indicator of soil functioning.     

保护性耕作对陇东黄土高原轮作田杂草的影响

以陇东黄土高原旱地连续12年保护性耕作为基础,通过田间取样调查,研究了4种耕作措施下冬小麦-箭筈豌豆-玉米轮作田中杂草的组成和群落特征.结果表明： 不同作物生长期的杂草种类、密度和群落特征各异.免耕使冬小麦生长期杂草密度显著增大,轮作箭筈豌豆后免耕处理的杂草密度变小;秸秆覆盖显著降低了玉米生长期的杂草密度,玉米生长期免耕+秸秆覆盖处理的杂草密度最低.冬小麦和箭筈豌豆轮作前后杂草的群落多样性特征完全相反,从冬小麦生长期到箭筈豌豆生长期,免耕处理杂草群落的多样性指数先高于常规耕作处理,后期低于常规耕作处理;玉米生长期免耕处理的多样性指数均大于其他3种处理.因此,免耕和秸秆覆盖在一些作物生长期能抑制杂草的发生和分布,3种作物轮作对杂草防治具有明显作用.     

Assessment of differences in ascomycete communities in the rhizosphere of field-grown wheat and potato

To assess effects of plant crop species on rhizosphere ascomycete communities in the field, we compared a wheat monoculture and an alternating crop rotation of wheat and potato. Rhizosphere soil samples were taken at different time points during the growing season in four consecutive years (1999-2002). An ascomycete-specific primer pair (ITS5-ITS4A) was used to amplify internal transcribed spacer (ITS) sequences from total DNA extracts from rhizosphere soil. Amplified DNA was analyzed by denaturing gradient gel electrophoresis (DGGE). Individual bands from DGGE gels were sequenced and compared with known sequences from public databases. DGGE gels representing the ascomycete communities of the continuous wheat and the rotation site were compared and related to ascomycetes identified from the field. The effect of crop rotation exceeded that of the spatial heterogeneity in the field, which was evident after the first year. Significant differences between the ascomycete communities from the rhizospheres of wheat in monoculture and one year after a potato crop were found, indicating a long-term effect of potato. Sequencing of bands excised from the DGGE gels revealed the presence of ascomycetes that are common in agricultural soils.     

Molecular Method To Assess the Diversity of Burkholderia Species in Environmental Samples

In spite of the importance of many members of the genus Burkholderia in the soil microbial community, no direct method to assess the diversity of this genus has been developed so far. The aim of this work was the development of soil DNA-based PCR-denaturing gradient gel electrophoresis (DGGE), a powerful tool for studying the diversity of microbial communities, for detection and analysis of the Burkholderia diversity in soil samples. Primers specific for the genus Burkholderia were developed based on the 16S rRNA gene sequence and were evaluated in PCRs performed with genomic DNAs from Burkholderia and non-Burkholderia species as the templates. The primer system used exhibited good specificity and sensitivity for the majority of established species of the genus Burkholderia. DGGE analyses of the PCR products obtained showed that there were sufficient differences in migration behavior to distinguish the majority of the 14 Burkholderia species tested. Sequence analysis of amplicons generated with soil DNA exclusively revealed sequences affiliated with sequences of Burkholderia species, demonstrating that the PCR-DGGE method is suitable for studying the diversity of this genus in natural settings. A PCR-DGGE analysis of the Burkholderia communities in two grassland plots revealed differences in diversity mainly between bulk and rhizosphere soil samples; the communities in the latter samples produced more complex patterns.