Effects of Pinus sylvestris root growth and mycorrhizosphere development on bacterial carbon source utilization and hydrocarbon oxidation in forest and petroleum-contaminated soils

The hypothesis that Pinus sylvestris L. root and mycorrhizosphere development positively influences bacterial community-linked carbon source utilization, and drives a concomitant reduction in mineral oil levels in a petroleum hydrocarbon- (PHC-) contaminated soil was confirmed in a forest ecosystem-based phytoremediation simulation. Seedlings were grown for 9 months in large petri dish microcosms containing either forest humus or humus amended with cores of PHC-contaminated soil. Except for increased root biomass in the humus/PHC treatment, there were no other significant treatment-related differences in plant growth and needle C and N status. Total cell and culturable bacterial (CFU) densities significantly increased in both rhizospheres and mycorrhizospheres that actively developed in the humus and PHC-contaminated soil. Mycorrhizospheres (mycorrhizas and extramatrical mycelium) supported the highest numbers of bacteria. Multivariate analyses of bacterial community carbon source utilization profiles (Biolog GN microplate) from different rhizosphere, mycorrhizosphere, and bulk soil compartments, involving principal component and correspondence analysis, highlighted three main niche-related groupings. The respective clusters identified contained bacterial communities from (i) unplanted bulk soils, (ii) planted bulk PHC and rhizospheres in PHC-contaminated soils, and (iii) planted bulk humus and rhizosphere/mycorrhizosphere-influenced humus, and mycorrhizosphere-influenced PHC contaminated soil. Correspondence analysis allowed further identification of amino acid preferences and increased carboxylic/organic acid preferences in rhizosphere and mycorrhizosphere compartments. Decreased levels of mineral oil (non-polar hydrocarbons) were detected in the PHC-contaminated soil colonized by pine roots and mycorrhizal fungi. These data further support our view that mycorrhizosphere development and function plays a central role in controlling associated bacterial communities and their degradative activities in lignin-rich forest humus and PHC-contaminated soils.     

Protozoan communities around conifer roots colonized by ectomycorrhizal fungi

Protozoan communities around roots with different types of ectomycorrhizae were distinct. These protozoan communities differed both qualitatively and quantitatively with the host (Pinus ponderosa, Pseudotsuga menziesii, Picea sitchensis, Tsuga heterophylla and Abies grandis) and the ectomycorrhizal fungal species. Based on the species identified and the numbers of individuals of each species, six communities of protozoa were found associated with specific ectomycorrhizae. Previous researchers have shown that mycorrhizal colonization of roots alters the amounts and types of exudates produced by roots, which in turn alters the bacterial community present. Most likely, mycorrhizal colonization of roots influences the protozoan community around roots by controlling the bacterial community. However, the protozoan community may in turn influence the successional dynamics of ectomycorrhizal fungi on different host root systems by a variety of mechanisms. These mechanisms could include: (1) preying upon individuals and perhaps removing particular species of bacteria from the mycorrhizosphere; and (2) controlling nitrogen mineralization in the rhizosphere. Further work needs to be performed to determine the interaction between these quadrate (plant-bacteria-fungi-protozoa) associations.     

Effect of inoculation of a TOL plasmid containing mycorrhizosphere bacterium on development of Scots pine seedlings, their mycorrhizosphere and the microbial flora in m-toluate-amended soil

The purpose of this study was to evaluate the influence of introduced bacteria containing a contaminant degrading plasmid on the growth and survival of pine seedlings and mycorrhizosphere microbial flora in contaminated soil. The Pseudomonas fluorescens strain OS81, originally isolated from fungal hyphae in contaminated soil, was supplied with the TOL plasmid pWW0::Km (to generate OS81(pWW0::Km)) and inoculated in humus-soil microcosms with and without pine seedlings mycorrhized with Suillus bovinus. After 3 months of regular treatment with m-toluate (mTA) solutions, the introduced catabolic plasmid was found to be disseminated in the indigenous bacterial population of both mycorrhizosphere and soil uncolonized by the fungus. Transconjugants were represented by bacteria of the genera Pseudomonas and Burkholderia and their number correlated positively with the concentration of mTA applied. Indigenous mTA degrading bacteria with low similarity to Burkholderia species were also enriched in microcosms. They were mostly associated with mycorrhizal soil or fungal structures and virtually absent in microcosms without pines. The total number of Tol(+) bacteria was higher in mycorrhizospheric soil compared with bulk soil. Inoculation with P. fluorescens OS81(pWW0::Km) had a positive effect on the development of roots and fungus in contaminated soil. Both inoculation with the P. fluorescens OS81(pWW0::Km) and mTA contamination as well as the presence of mycorrhized pine roots and fungal hyphae had an effect on the microbial community structure of soil as measured by carbon source oxidation patterns. However, the impact of mTA on the microbial community was more prominent. The study indicates that an effect on plant and fungal development can be obtained by manipulating the mycorrhizosphere. Both introduction of the bacterium carrying the degradative plasmid and the plasmid itself are likely to have a positive effect not only on the organisms involved, but also on bioremediation of contaminated soil, a factor that was not directly monitored here.     

Recent 14C-labelled assimilate allocation to Scots pine seedling root and mycorrhizosphere compartments developed on reconstructed podzol humus,E- and B- mineral horizons

In northern boreal forests, podzolic soils prevail that comprise of a distinct upper organic humus/mor (O) horizon that is supported by underlying eluvial (E) and illuvial (B) mineral horizons. The dominant tree species, Scots pine (Pinus sylvestris L.), is known to be highly dependent on root symbiosis with ectomycorrhizal fungi that develop in constituent podzol horizons for growth in these nutrient limited soils. The aim of this microcosm-based study was a quantification of photosynthetically fixed ¹⁴C allocation, following standard pulse-feeding of 7-month-old Scots pine seedling shoots, to respective root and mycorrhizosphere compartments that developed in the reconstructed podzol (O, E and B) profile. Biomass of roots and mycorrhizas decreased with increasing soil depth but no soil origin, control forest vs. clear-cut area, related differences were observed. Similarly, no major soil origin- or podzol horizon-related differences in categorised ectomycorrhizal morphotypes and number of mycorrhizas, in relation to pooled root and mycorrhiza biomass, were detected. However, the total recovery of ¹⁴C-label was significantly higher in clear-cut soil microcosms compared to control counterparts. A significant finding was equivalent ¹⁴C-carbon allocation to roots and ectomycorrhizas in all three major, organic and mineral, podzol profile horizons studied. These carbon allocation data provide additional support for direct (or indirect) roles of roots and symbiotic mycorrhizal fungi in mineral weathering and biodegradation of organic ligands that are central for plant acquisition of growth limiting nutrients and the podzolization process in boreal forest ecosystems.     

Characterization of culturable bacterial populations associating with Pinus sylvestris--Suillus bovinus mycorrhizospheres

Bacterial isolations were carried out on Pinus sylvestris--Suillus bovinus mycorrhizospheres obtained directly from boreal pine forest. When samples were taken during dry weather, the numbers of bacterial colony-forming units were significantly higher in uncolonized short roots and external mycelia than in mycorrhizal roots and soil outside the mycorrhizosphere. In contrast, the colony-forming unit counts were similar in all hypogeous samples after rainy weather. Culturable bacteria were absent from most Suillus bovinus sporocarps. The bacteria isolated from all types of mycorr hizo sphere samples, i.e. short roots, mycorrhizal roots, and external mycelia, consisted primarily of Burkholderia spp., whereas most isolates from soil outside the mycorrhizosphere were identified as Paenibacillus spp. This study shows that mycorrhizal external mycelia can expand the habitat favourable for common rhizosphere bacteria into the soil far from the immediate rhizosphere. Some of these bacteria may help the trees with nitrogen acquisition, since potentially diazotrophic bacteria harbouring nitrogenase reductase (nifH) genes were isolated from mycorrhizal root tips.     

Influence of plant developmental stage on microbial community structure and activity in the rhizosphere of three field crops

Seasonal shifts in rhizosphere microbial populations were investigated to follow the influence of plant developmental stage. A field study of indigenous microbial rhizosphere communities was undertaken on pea (Pisum satvium var. quincy), wheat (Triticum aestivum var. pena wawa) and sugar beet (Beta vulgaris var. amythyst). Rhizosphere community diversity and substrate utilization patterns were followed throughout a growing season, by culturing, rRNA gene density gradient gel electrophoresis and BIOLOG. Culturable bacterial and fungal rhizosphere community densities were stable in pea and wheat rhizospheres, with dynamic shifts observed in the sugar beet rhizosphere. Successional shifts in bacterial and fungal diversity as plants mature demonstrated that different plants select and define their own functional rhizosphere communities. Assessment of metabolic activity and resource utilization by bacterial community-level physiological profiling demonstrated greater similarities between different plant species rhizosphere communities at the same than at different developmental stages. Marked temporal shifts in diversity and relative activity were observed in rhizosphere bacterial communities with developmental stage for all plant species studied. Shifts in the diversity of fungal and bacterial communities were more pronounced in maturing pea and sugar beet plants. This detailed study demonstrates that plant species select for specialized microbial communities that change in response to plant growth and plant inputs.     

Fluorescent pseudomonads in the rhizosphere of plants and their relation to root exudates

Fluorescent pseudomonads were present in chernozem soil not influenced by plant roots (10(3)-10(4) per g dry soil) in the rhizosphere soil of various plants (10(4)-10(5) per g soil) and on roots (10(3) to 10(7) per g fresh roots), depending on the species and age of the plant. Relative species representation of fluorescent pseudomonads changed on the roots and in the plant rhizosphere as compared with free soil. Pseudomonas fluorescens, representing 60-93% of the population of fluorescent pseudomonads predominated on the roots of all plants investigated. Somewhat different results were obtained in rhizosphere soil. Relatively higher numbers of P. fluorescens were detected in the rhizosphere soil of cucumber and maize, numbers in the rhizosphere soil of wheat were practically the same as in free soil and higher numbers of P. putida were found in the rhizosphere soil of barley. Almost all components contained in the root exudates of the plants studied, including beta-pyrazolylalanine from the root exudates of cucumbers were utilized as carbon and energy sources. Root exudates of wheat and maize were utilized by the strain P. putida K2 with an efficiency of 73-91%, depending on species and age of the plant.     

Burkholderia, a genus rich in plant-associated nitrogen fixers with wide environmental and geographic distribution

The genus Burkholderia comprises 19 species, including Burkholderia vietnamiensis which is the only known N(2)-fixing species of this bacterial genus. The first isolates of B. vietnamiensis were recovered from the rhizosphere of rice plants grown in a phytotron, but its existence in natural environments and its geographic distribution were not reported. In the present study, most N(2)-fixing isolates recovered from the environment of field-grown maize and coffee plants cultivated in widely separated regions of Mexico were phenotypically identified as B. cepacia using the API 20NE system. Nevertheless, a number of these isolates recovered from inside of maize roots, as well as from the rhizosphere and rhizoplane of maize and coffee plants, showed similar or identical features to those of B. vietnamiensis TVV75(T). These features include nitrogenase activity with 10 different carbon sources, identical or very similar nifHDK hybridization patterns, very similar protein electrophoregrams, identical amplified 16S rDNA restriction (ARDRA) profiles, and levels of DNA-DNA reassociation higher than 70% with total DNA from strain TVV75(T). Although the ability to fix N(2) is not reported to be a common feature among the known species of the genus Burkholderia, the results obtained show that many diazotrophic Burkholderia isolates analyzed showed phenotypic and genotypic features different from those of the known N(2)-fixing species B. vietnamiensis as well as from those of B. kururiensis, a bacterium identified in the present study as a diazotrophic species. DNA-DNA reassociation assays confirmed the existence of N(2)-fixing Burkholderia species different from B. vietnamiensis. In addition, this study shows the wide geographic distribution and substantial capability of N(2)-fixing Burkholderia spp. for colonizing diverse host plants in distantly separated environments.     

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.     

Impact of Ectomycorrhizosphere on the Functional Diversity of Soil Bacterial and Fungal Communities from a Forest Stand in Relation to Nutrient Mobilization Processes

The ectomycorrhizal symbiosis alters the physicochemical and biological conditions in the surrounding soil, thus creating a particular environment called ectomycorrhizosphere, which selects microbial communities suspected to play a role in gross production and nutrient cycling. To assess the ectomycorrhizosphere effect on the structure of microbial communities potentially involved in the mobilization of nutrients from the soil minerals in a poor-nutrient environment, we compared the functional diversity of soil and ectomycorrhizosphere bacterial communities in a forest stand. Two hundred and sixty-four bacterial strains and 107 fungal strains were isolated from the bulk soil of an oak (Quercus petraea) stand and from oak–Scleroderma citrinum ectomycorrhizosphere and ectomycorrhizae, in two soil organo-mineral horizons (0 to 3 cm and 5 to 10 cm). They were characterized using two in vitro tests related to their capacities to mobilize iron and phosphorus. We demonstrated that the oak–S. citrinum ectomycorrhizosphere significantly structures the culturable bacterial communities in the two soil horizons by selecting very efficient strains for phosphorus and iron mobilization. This effect was also observed on the diversity of the phosphate-solubilizing fungal communities in the lower soil horizon. A previous study already demonstrated that Laccaria bicolor–Douglas fir ectomycorrhizosphere structures the functional diversity of Pseudomonas fluorescens population in a forest nursery soil. Comparing to it, our work highlights the consistency of the mycorrhizosphere effect on the functional diversity of bacterial and fungal communities in relation to the mineral weathering process, no matter the fungal symbiont, the age and species of the host tree, or the environment (nursery vs forest). We also demonstrated that the intensity of phosphorus and iron mobilization by the ectomycorrhizosphere bacteria isolated from the lower soil horizon was significantly higher compared to that which was isolated from the upper horizon. This reveals for the first time a stratification of the functional diversity of the culturable soil bacterial communities as related to phosphorus and iron mobilization.     

Bacterial and fungal communities in bulk soil and rhizospheres of aluminum-tolerant and aluminum-sensitive maize (Zea mays L.) lines cultivated in unlimed and limed Cerrado soil

Liming of acidic soils can prevent aluminum toxicity and improve crop production. Some maize lines show aluminum (Al) tolerance, and exudation of organic acids by roots has been considered to represent an important mechanism involved in the tolerance. However, there is no information about the impact of liming on the structures of bacterial and fungal communities in Cerrado soil, nor if there are differences between the microbial communities from the rhizospheres of Al-tolerant and Al-sensitive maize lines. This study evaluated the effects of liming on the structure of bacterial and fungal communities in bulk soil and rhizospheres of Al-sensitive and Al-tolerant maize (Zea mays L.) lines cultivated in Cerrado soil by PCR-DGGE, 30 and 90 days after sowing. Bacterial fingerprints revealed that the bacterial communities from rhizospheres were more affected by aluminum stress in soil than by the maize line (Al-sensitive or Al-tolerant). Differences in bacterial communities were also observed over time (30 and 90 days after sowing), and these occurred mainly in the Actinobacteria. Conversely, fungal communities from the rhizosphere were weakly affected either by liming or by the rhizosphere, as observed from the DGGE profiles. Furthermore, only a few differences were observed in the DGGE profiles of the fungal populations during plant development when compared with bacterial communities. Cloning and sequencing of 16S rRNA gene fragments obtained from dominant DGGE bands detected in the bacterial profiles of the Cerrado bulk soil revealed that Actinomycetales and Rhizobiales were among the dominant ribotypes.     

Role of dissimilatory fermentative iron-reducing bacteria in Fe uptake by common bean (Phaseolus vulgaris L.) plants grown in alkaline soil

Iron (Fe) is an essential element for plant growth and development. Some plant growth-promoting rhizobacteria can increase Fe uptake by plants through reduction of Fe(III) to Fe(II) at the root surface. The aim of this work was to identify novel bacterial strains with high Fe(III) reduction ability and to evaluate their role in plant Fe uptake. Four bacterial strains (UMCV1 to UMCV4) showing dissimilatory Fe-reducing activity were isolated from the rhizosphere of bean and maize plants and further identified by 16S rDNA amplification and sequence analysis. From these analyses, UMCV1 and UMCV2 isolates were identified as Bacillus megaterium and Arthrobacter spp., respectively, whereas UMCV3 and UMCV4 were identified as Stenotrophomonas maltophilia. All four isolates showed Fe reduction in a nonflooded soil and when associated with roots of bean plants grown in alkaline soil or in mineral medium. In addition, the bacterial isolates were able to stimulate plant growth in vitro and on a broad level, plants grown in inoculated soil were generally bigger and with higher Fe content than those grown in sterilized soil. These results indicate that bacterial species isolated from the rhizosphere of bean and maize plants contribute significantly to Fe uptake by plants likely through increased Fe(III) reduction in the rhizosphere.     

Utilization of different carbon sources by bacteria isolated from the roots of pine seedlings (Pinus silvestris L.) inoculated with root-free, rhizosphere and mycorrhizosphere soil

Studies were carried out on the utilization of different sugars (glucose, fructose, arabinose, xylose, sucrose) and organic acids (acetic, citric, fumaric, propionic, succinic) by fast and slow growing bacteria isolated from the roots of pine seedlings (P. silvestris L.) inoculated with root-free, rhizosphere and mycorrhizosphere soil from nursery and mature pine forest. Sucrose among sugars and propionic acid among organic acids were the less frequently utilized compounds. Sugars were better carbon sources than organic acids. Proportion of isolates utilizing respective sugars or organic acids was, in general, significantly higher among fast growing bacteria as compared with slow growing ones. No significant differences in number of strains assimilating the appropiate sugars depending on their original habitat or kind of soil were observed. Such differences were noted for utilization of some organic acids. 3-factor ANOVA confirmed that the growth speed of bacteria at the moment of their isolation had the strongest effect on utilization of the compounds studied.     

Endobacteria in some ectomycorrhiza of Scots pine (Pinus sylvestris)

The diversity of cultivable endobacteria associated with four different ectomycorrhizal morphotypes (Suillus flavidus, Suillus variegatus, Russula paludosa and Russula sp.) of Scots pine (Pinus sylvestris) was analysed by restriction fragment length polymorphism profiling of PCR-amplified rDNA intergenic spacer regions and by sequence analyses of 16S rRNA genes. Ectomycorrhizal root tip surface-sterilization methods were developed and assessed for their efficiencies. Bacterial communities from surface-sterilized ectomycorrhizal root tips were different from those of ectomycorrhizal root tips without surface-sterilization for all the morphotypes studied. Endobacteria belonging to the genera Pseudomonas, Burkholderia and Bacillus were isolated from more than one ectomycorrhizal morphotype, whereas species of Rahnella, Janthinobacterium and Rhodococcus were only isolated from the single morphotypes of S. variegatus, R. paludosa and Russula sp., respectively. Some of the isolated endobacteria utilized fungal sugars more readily than typical plant sugars in carbon utilization assays.     

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.     

Termination of belowground C allocation by trees alters soil fungal and bacterial communities in a boreal forest

The introduction of photosynthates through plant roots is a major source of carbon (C) for soil microbial biota and shapes the composition of fungal and bacterial communities in the rhizosphere. Although the importance of this process, especially to ectomycorrhizal fungi, has been known for some time, the extent to which plant belowground C allocation controls the composition of the wider soil community is not understood. A tree-girdling experiment enabled studies of the relationship between plant C allocation and microbial community composition. Girdling involves cutting the phloem of trees to prevent photosynthates from entering the soil. Four years after girdling, fungal and bacterial communities were characterized using DNA-based profiles and cloning and sequencing. Data showed that girdling significantly altered fungal and bacterial communities compared with the control. The ratio of ectomycorrhizal to saprobic fungal sequences significantly decreased in girdled treatments, and this decline was found to correlate with the fungal phospholipid fatty acid biomarker 18:2ω6,9. Bacterial communities also varied in the abundance of the two dominant phyla Acidobacteria and Alphaproteobacteria . Concomitant changes in fungal and bacterial communities suggest linkages between these two groups and point toward plant belowground C allocation as a key determinant of microbial community composition.     

Rhizosphere disturbance influences fungal colonization and community development on dead fine roots

Little is known about the community dynamics of fungi on decomposing fine roots, despite the importance of fine roots as a source of carbon to detrital systems in forests. We examined fungal communities on dead roots in a sugar-maple dominated northern hardwood forest to test the hypothesis that community development is sensitive to rhizosphere disruption. We generated cohorts of dead fine roots in root windows and disturbed the rhizosphere microbial community in half of the windows by moving roots into sieved bulk soil. We sampled root fragments repeatedly over time and cultured fungi from these fragments to explore temporal patterns of fungal species composition. Disturbing the root rhizosphere prior to initiating decomposition changed the dominant fungal taxa, the distribution of dominant species within the community, and the temporal development in the culturable fungal community. Dominance in control roots shifted from Neonectria in early decay to Umbelopsis in later decay. Disturbance roots were more evenly dominated over time by Trichoderma, Neonectria, another species of Umbelopsis, and Pochonia. Our results suggest that species interactions are important in the ecology of fine root decay fungi, with the rhizosphere community of the living root influencing development of the decay community.     

16S rDNA fingerprinting of rhizosphere bacterial communities associated with healthy and Phytophthora infected avocado roots

Molecular techniques employing 16S rDNA profiles generated by PCR-DGGE were used to detect changes in bacterial community structures of the rhizosphere of avocado trees during infection by Phytophthora cinnamomi and during repeated bioaugmentation with a disease suppressive fluorescent pseudomonad. When the 16S rDNA profiles were analyzed by multivariate analysis procedures, distinct microbial communities were shown to occur on healthy and infected roots. Bacterial communities from healthy roots were represented by simple DNA banding profiles, suggestive of colonization by a few predominant species, and were approximately 80% similar in structure. In contrast, roots that were infected with Phytophthora, but which did not yet show visible symptoms of disease, were colonized by much more variable bacterial communities that had significantly different community structures from those of healthy roots. Root samples from trees receiving repeated applications of the disease suppressive bacterium Pseudomonas fluorescens st. 513 were free of Phytophthora infection, and had bacterial community structures that were similar to those of nontreated healthy roots. Sequence analysis of clones generated from four predominant bands cut from the DGGE gels revealed the presence of pseudomonads, as well as several previously unidentified bacteria. Differentiation of 16S rDNA profiles for healthy and infected roots suggests that rhizosphere bacterial community structure may serve as an integrative indicator of changes in chemical and biological conditions in the plant rhizosphere during the infection process.     

Influence of Forest Trees on the Distribution of Mineral Weathering-Associated Bacterial Communities of the Scleroderma citrinum Mycorrhizosphere

In acidic forest soils, availability of inorganic nutrients is a tree-growth-limiting factor. A hypothesis to explain sustainable forest development proposes that tree roots select soil microbes involved in central biogeochemical processes, such as mineral weathering, that may contribute to nutrient mobilization and tree nutrition. Here we showed, by combining soil analyses with cultivation-dependent analyses of the culturable bacterial communities associated with the widespread mycorrhizal fungus Scleroderma citrinum, a significant enrichment of bacterial isolates with efficient mineral weathering potentials around the oak and beech mycorrhizal roots compared to bulk soil. Such a difference did not exist in the rhizosphere of Norway spruce. The mineral weathering ability of the bacterial isolates was assessed using a microplaque assay that measures the pH and the amount of iron released from biotite. Using this microplate assay, we demonstrated that the bacterial isolates harboring the most efficient mineral weathering potential belonged to the Burkholderia genus. Notably, previous work revealed that oak and beech harbored very similar pHs in the 5- to 10-cm horizon in both rhizosphere and bulk soil environments. In the spruce rhizosphere, in contrast, the pH was significantly lower than that in bulk soil. Because the production of protons is one of the main mechanisms responsible for mineral weathering, our results suggest that certain tree species have developed indirect strategies for mineral weathering in nutrient-poor soils, which lie in the selection of bacterial communities with efficient mineral weathering potentials.The mobilization of nutrients via the biotic and abiotic weathering of soil minerals is crucial to satisfying plant nutritional needs (2, 17), especially in acidic forest soils, which are mainly nonfertilized and nutrient poor. Besides the physicochemical weathering reactions, evidence is presently accumulating which indicates that certain soil bacterial strains increase mineral weathering and improve tree nutrition (5, 9, 32, 39-41).By way of their root exudates, plants alter the structure and activity of microbial communities (6, 25, 51) and selectively favor certain ones that are potentially beneficial to them (15, 16, 21, 45, 46). A hypothesis for sustainable forest development proposes that tree roots select from the soil efficient mineral weathering bacterial communities that may contribute to nutrient mobilization and tree growth (20). In this manner, recent studies (10, 46) have revealed that the oak-Scleroderma citrinum ectomycorrhizal symbiosis selects bacterial communities that are more efficient in mineral weathering than those of the surrounding soil, suggesting that the mycorrhizal symbiosis has an indirect effect on plant nutrition through its selective pressure on the functional diversity of the mycorrhizosphere bacterial communities.Distinct impacts of the tree species on the soil bacterial community structure have been previously reported (23, 38), suggesting that the composition and activity of soil bacterial communities depend on tree physiology and notably on its impact on the soil physicochemical properties and nutrient cycling (24, 26, 37). However, no study has ever addressed the question of the impact of tree species on the structure of forest soil bacterial communities involved in mineral weathering. This question regarding the impact of tree species on the functional diversity of the bacterial communities remains a major issue in forestry, especially in the context of today''s climate change, which will give rise to a shift in the spatial distribution of forest tree species.To appreciate the effect of tree species on mycorrhizosphere bacterial communities, we focused on a single but ubiquitous mycorrhizal fungus, S. citrinum, which forms mycorrhizae with different tree species. Since no functional genes have been identified to date, a cultivation-dependent analysis was developed in this study. A total of 155 bacterial isolates were randomly chosen among a collection of 400 bacterial isolates from the soil-Scleroderma citrinum mycorrhiza interface (ectomycorrhizosphere), the extramatrical mycelium (hyphosphere), and the surrounding soil (bulk soil) in 28-year-old stands of oak (Quercus sessiliflora Smith), beech (Fagus sylvatica L.), and Norway spruce (Picea abies Karst.). The mineral weathering potential of each bacterial isolate was evaluated by way of an in vitro microplate assay, putting in interaction a calibrated bacterial suspension and the biotite, a mineral widespread in soils (46). The bacterial isolates were genotypically characterized by amplifying and sequencing a portion of the 16S rRNA gene. Their mineral weathering efficiencies and the functional structure of the bacterial communities were compared with the physicochemical characteristics of the surrounding soil.