Improved zinc tolerance in Eucalyptus globulus inoculated with Glomus deserticola and Trametes versicolor or Coriolopsis rigida

The potential of interactions between saprophytic and arbuscular mycorrhizal (AM) fungi to improve Eucalyptus globulus grown in soil contaminated with Zn were investigated. The presence of 100 mg kg ⁻¹ Zn decreased the shoot and root dry weight of E. globulus colonized with Glomus deserticola less than in plants not colonized with AM. Zn also decreased the extent of root length colonization by AM and the AM fungus metabolic activity, measured as succinate dehydrogenase (SDH) activity of the fungal mycelium inside the E. globulus root. The saprophytic fungi Trametes versicolor and Coriolopsis rigida increased the shoot dry weight and the tolerance of E. globulus to Zn when these plants were AM-colonized. Both saprophytic fungi increased the percentage of AM root length colonization and elevated G. deserticola SDH activity in the presence of all Zn concentrations applied to the soil. In the presence of 500 and 1000 mg kg⁻¹ Zn, there were higher metal concentrations in roots and shoots of AM than in non-AM plants; furthermore, both saprophytic fungi increased Zn uptake by E. globulus colonized by G. deserticola. The higher root to shoot metal ratio observed in mycorrhizal E. globulus plants indicates that G. deserticola enhanced Zn uptake and accumulation in the root system, playing a filtering/sequestering role in the presence of Zn. However, saprophytic fungi did not increase the root to shoot Zn ratio in mycorrhizal E. globulus plants. The effect of the saprophytic fungi on the tolerance and the accumulation of Zn in E. globulus was mediated by its effect on the colonization and metabolic activity of the AM fungi.     

Zn accumulation and subcellular distribution in the Zn hyperaccumulator Sedurn alfredii Hance

Zn accumulation and subcellular distribution in leaves of the hyperaccumulating ecotype （HE） and non-hyperaccumulating ecotype （NHE） of Sedum alfredii Hance were studied using radiotracer and gradient centrifugation techniques. Leaf Zn accumulation in the HE of S. alfredii was 18.5-26.7 times greater than that in the NHE when the plants were grown at 1-500μmol Zn L-1. Leaf section uptake of 65Zn was highly dependent on external Zn levels. Greater 65Zn uptake in HE was noted only at external Zn levels 〉 100μmol L-1. Zinc subcellular distribution in the leaves of the two ecotypes of S. alfredii was： cell wall 〉 soluble fraction 〉 cell organelle. However, more Zn was distributed to the leaf cell wall and soluble fractions for HE than for NHE. In the leaf of HE, 91%-94% of the Zn was found in the cell walls and the soluble fraction and only 6%-9% Zn was distributed in the cell organelle fraction. For NHE, about 20%-26% Zn was recovered in the cell organelle fraction. In stems, Zn distribution to the ceil wail fraction was approximately two fold greater in the HE than that in the NHE. For the hyperaccumulating ecotype of S. alfredii, the cell wall and the vacuole played a very important role in Zn tolerance and hyperaccumulation.     

Zinc uptake kinetics in the low and high‐affinity systems of two contrasting rice genotypes

Rice (Oryza sativa L.) cultivars differ widely in their susceptibility to zinc (Zn) deficiency. The physiological basis of Zn efficiency (ZE) is not clearly understood. In this study, the effects of Zn‐sufficient and Zn‐deficient pretreatments on the time and concentration‐dependent uptake kinetics of Zn were examined at low (0–160 nM) and high Zn supply levels (0–80 μM) in two contrasting rice genotypes (Zn‐efficient IR36 and Zn‐inefficient IR26). The results show that ⁶⁵Zn²⁺ influx rate was over 10 times greater for the Zn‐deficient pretreatment plants than for the Zn‐sufficient pretreatment plants. At low Zn supply, significant higher ⁶⁵Zn²⁺ influx rates were found for the Zn‐efficient genotype than for the inefficient genotype, with a greater difference (over three‐fold) at Zn supply > 80 nM in the Zn‐deficient pretreatments. At high Zn supply levels, however, a difference (2.5‐fold) in ⁶⁵Zn²⁺ influx rate between the two genotypes was only noted in the Zn‐deficient pretreatments. Similarly, the ⁶⁵Zn²⁺ accumulation in the roots and shoots of Zn‐efficient IR36 pretreated with Zn‐deficiency were sharply increased with time and higher than that in the Zn‐inefficient IR26 with an over four‐fold difference at 2 h absorption time. However, with Zn‐deficient pretreatments, the Zn‐efficient genotype showed a higher shoot : root ⁶⁵Zn ratio at higher Zn supply. Remarkable differences in root and shoot ⁶⁵Zn²⁺ accumulation were noted between the two genotypes in the Zn‐deficiency pretreatment, especially at low Zn level (0.05 μM), with 2–3 times higher values for IR36 than for IR26 at an uptake time of 120 min. There appear to be two separate Zn transport systems mediating the low and high‐affinity Zn influx in the efficient genotype. The low‐affinity system showed apparent Michaelis–Menten rate constant (K_m) values ranging from 10 to 20 nM, while the high‐affinity uptake system showed apparent K_m values ranging from 6 to 20 μM. The V_max value was significantly elevated in IR36 and was 3–4‐fold greater for IR36 than for IR26 at low Zn levels, indicating that the number of root plasma membrane transporters in low‐affinity uptake systems play an important role for the Zn efficiency of rice.     

Mycorrhizal communities in relation to biomass production and nutrient use efficiency in two varieties of Douglas fir (Pseudotsuga menziesii var. menziesii and var. glauca) in different forest soils

To investigate the role of arbuscular and ectomycorrhizas for growth and nutrition, Douglas fir (Pseudotsuga menziesii), a species capable of establishing both types of symbioses, was used. Seedlings of two varieties of Douglas fir, var. menziesii (DFM) and var. glauca (DFG), differing in biomass production under non-mycorrhizal conditions, were planted in two soils of different nutrient status without preceding Douglas fir cultivation. After 2 years, the abundance of ectomycorrhizas was significantly higher on DFM than on DFG roots and higher in nutrient rich than in poor soil. Independent of the abundance of ectomycorrhiza colonization, roots of both Douglas fir varieties contained in most cases a minimum of six to nine different fungal species identified by ITS sequencing, which displayed both soil- and seed origin-specific patterns. Rhizopogon vinicolor was associated with DFM, whereas Cadophora finlandia, Sebacinaceous sp., Tricholoma sp. and a Tuber sp. were more frequently found on DFG roots. Ectendomycorrhizas were also identified and included under ectomycorrhiza for determination of nutrient relations. Net primary productivity and nitrogen-use efficiency were positively correlated with the degree of ectomycorrhiza colonization of Douglas fir seedlings suggesting that nitrogen accumulation did not keep pace with biomass production despite increased ectomycorrhizal colonization. Phosphorus-use efficiency was negatively correlated with the abundance of arbuscular mycorrhizas, suggesting higher efficiency of these fungi than of ectomycorrhizas for phosphorus supply to the host. DFM, which had higher inherited growth capacities than DFG, displayed higher ectomycorrhizal colonization than DFG. In our experimental design the physiology of the host tree and soil properties, but not the diversity of fungi present on roots, were decisive factors for formation and abundance of ectomycorrhiza.     

Rice (Oryza sativa L.) seedlings enriched with zinc or manganese: Their impacts on cadmium accumulation and expression of related genes

Cadmium (Cd) contamination in paddy soils means that the rice produced there may be unsafe for human consumption. A hydroponic study was conducted to enrich rice seedlings with zinc (Zn) or manganese (Mn), and the uptake and transport characteristics of Cd in these Zn- and Mn-rich seedlings were subsequently investigated using a greenhouse pot trial. The results showed that hydroponic cultivation in 10–50 μmol L^-1 Zn (ZnSO₄·7H₂O) or 50–250 μmol L^-1 Mn (MnSO₄·H₂O) for 30 d had no significant impact on rice growth, while the accumulation of Zn and Mn was 7.31–18.5 and 25.4–47.7 times higher, respectively, than in the control (no Zn or Mn addition). The accumulation of Cd in the Zn- and Mn-rich rice plants was 26.3%–38.6% and 34.4%–44.5% lower than that in the control, respectively, and the translocation factors of Cd from roots to shoots also decreased by 23.3%–41.3% and 25.3%–37.0%, respectively, after transplanting to Cd-contaminated soils. Furthermore, the relative expression levels of OsIRT1 (Oryza sativa iron-regulated transporter 1) were downregulated by 40.1%–59.3% and 16.0%–25.9%, respectively, in the Zn- and Mn-rich seedling roots. This downregulation may indicate a possible mechanism contributing to the reductions in Cd absorption. Field experiments confirmed that the Zn- and Mn-rich seedlings produced brown rice (unpolished rice grains) with significantly decreased concentrations of Cd (34.2%–44.4%). This study provides an innovative method for reducing the food safety risks from rice grown on slightly to moderately Cd-contaminated paddy soils.     

花生根系钙素吸收特性研究

运用电子探针、45Ca示踪及水培方法研究花生根系钙素吸收特性,结果表明花生根伸长区与成熟区均可吸钙,二者吸钙量分别占吸钙总量的28.9％～34.7％和71.1％～65.3％。在供钙浓度为0～0.5nmol/L范围,根系钙素吸收符合Michaelis-Menten酶动力学模型。此时其完全受代谢控制,完整根Km值为0.0549mmol/L、Fmax为3.81nmol/(cmh);断离根Km为0.0534mmol/L、Fmax为3.79nmol/(cm.h)。此后随供钙浓度增加,其受非代谢因素影响越大。供钙浓度为2.0mmol/L时,代谢因素对吸钙总量的贡献为63.2％,而非代谢因素的贡献为36.8％。     

Specific flavonoids as interconnecting signals in the tripartite symbiosis formed by arbuscular mycorrhizal fungi, Bradyrhizobium japonicum (Kirchner) Jordan and soybean (Glycine max (L.) Merr.)

Many legume plants benefit from the tripartite symbiosis of arbuscular mycorrhizal fungi (AMF) and rhizobia. Beneficial effects for the plant have been assumed to rely on increased P supply through the mycorrhizas. Recently, we demonstrated that P does not regulate the establishment of the tripartite symbiosis. Flavonoids appear to play a role as early signals for both rhizobia and AMF. Four soybean lines known to express different concentrations of the isoflavones genistein, daidzein and glycitein in the seed were used to test three hypotheses: (i) The establishment of the tripartite symbiosis is not dependent of a nutrient mediated effect; (ii) There is a positive correlation between seed isoflavone concentrations of different soybean lines and the progress of the tripartite symbiosis; (iii) Specific flavonoids control the establishment of the tripartite symbiosis in that a change in flavonoid root accumulation resulting from the development of one microbial partner can stimulate colonization of soybean roots by the other. Disturbed versus undisturbed soil treatments were produced to vary the potential for indigenous AMF colonization of soybean. In contrast, the potential for Bradyrhizobium was kept identical in both soil disturbance treatments. The uptake of P and Zn and the concentration of flavonoids in mycorrhizal soybean roots at 10 d after emergence were analysed either separately of Bradyrhizobium or in context of the tripartite symbiosis. Zinc nutrition did not differ between AMF treatments which supports the first hypothesis. The concentration of daidzein was at least four times greater in the root than in the seed reaching 3958±249 μg g⁻¹ dry across soybean lines. Coumestrol, which was absent in the seed, was synthesized to reach 2154±64 μg g⁻¹ dry. Conversely, the concentration of genistein was approximately three times smaller in the root that in the seed (301±15 μg g⁻¹ dry), while glycitein and formononetin were never detected. The establishment of the tripartite symbiosis was identical across soybean lines which does not support the second hypothesis. Concentrations of flavonoids were significantly greater in roots under disturbed soil, for which both symbioses were not as developed as in plants from undisturbed soil. This clearly supports the third hypothesis. This research provides the first data linking the function of different flavonoids to the establishment of the tripartite symbiosis, and suggests that these compounds are produced and released into the rhizosphere as a function of the colonization process.     

Phosphorus uptake by mycorrhizal and nonmycorrhizal roots of soybean

Kinetics of phosphorus absorption were investigated with mycorrhizal and nonmycorrhizal soybean (Glycine max Merr.) plants which were grown in a greenhouse and growth chamber and with nutrient solutions. Mycorrhizal plants depleted phosphorus from a dilute (<30uM) nutrient solution three times faster and had lower apparent efflux of phosphorus than the uninfected plants. The affinity (K_m) of roots for phosphorus from the solution was not increased by mycorrhizal infection. The results indicate that enhanced phosphorus absorption by mycorrhizal roots of soybean is due to an increase in the number of uptake sites per unit area of root (^v max).     

Effects of arbuscular mycorrhizal fungi on zinc uptake, translocation and accumulation in winter wheat during whole plant growth stages

Although arbuscular mycorrhizal fungi (AMF) could play important roles in zinc (Zn) uptake in host plants, the effects of AMF on Zn uptake and transport in winter wheat during the whole growth stages remain unclear. A pot experiment was conducted to investigate the effects of Funneliformis mosseae (Fm) and Claroideoglomus etunicatum (Ce) on Zn absorption, transport, and accumulation in winter wheat growing in soils spiked with different Zn levels (0, 2.5, and 25 mg kg^-1). The results showed that there was a significant correlation between mycorrhizal colonization rate and Zn absorption efficiency in winter wheat roots during the post-anthesis period, but there was no significant correlation during the pre-anthesis period. Arbuscular mycorrhizal fungi significantly increased Zn concentrations (0.56-1.58 times) in wheat grains under 0 mg kg^-1 Zn level, but decreased Zn concentrations in wheat grains under 25 mg kg^-1 Zn level. Additionally, at the filling and maturity stages, AMF increased Zn absorption rate and the contribution of root Zn uptake to grain Zn by 3-14 and 0.36-0.64 times, respectively, under 0 mg kg^-1 Zn level and 0.21-1.02 and 0.27-0.37 times, respectively, under 2.5 mg kg^-1 Zn level. However, AMF decreased root Zn absorption rate (0.32-0.61 times) and increased the contribution of Zn remobilization in vegetative tissues to grain Zn (1.69-2.01 times) under 25 mg kg^-1 Zn level. This study would complement the mechanisms and effects of AMF on Zn absorption and transport in winter wheat and provide a potential method for the application of AMF to enrich wheat grain Zn.     

Gradient differences in soil metal solubility and uptake by shoots and roots of wheat (T. aestivum)

The aim of this research was to identify and quantify gaps in currents methods and models for predicting the plant availability of selected nutrient and contaminant metals (Cu, Ni, Zn, Cd) in soil. This study investigated relationships between the relative solubility of Cu, Ni, Zn, and Cd determined by six extraction methods with short-term uptake by shoots and roots of wheat (Triticum aestivum). For Cu, Ni, and Cd, relationships between solubility and plant uptake were found to be different for shoots and roots, with Cu and Ni solubility being more closely correlated with root uptake compared with shoot uptake. Correlation coefficients for Cd concentrations in shoot and root tissue for all six solubility methods were poor (r ²?<?0.5), while corresponding results for Zn explained more than 50 % of shoot variation but less than 50 % of root variation. Soil Cu solubility explained up to 85 % of variation in root uptake compared with 42–55 % for shoot uptake. These results clearly demonstrated that purely chemical and passive diffusion mechanisms were inadequate predictors of Cd uptake by shoots and roots, together with Cu uptake by shoots. Thus further attempts at refining soil metal bioavailability assays based solely on chemical extraction without consideration of plant responses are unlikely to improve prediction of plant uptake.     

Zinc and Lead Accumulation by Two Contrasting Ecotypes of Sedum alfredii Hance at Different Zinc/Lead Complex Levels

The old mined ecotype (OME) was more tolerant to high zinc (Zn) and lead (Pb) levels than the nonmined ecotype (NME) of Sedum alfredii H. The greatest stem values of Zn and Pb of the OME at Zn/Pb of 2.0/0.1 mM reached 19.9 and 0.460 g kg^?1 respectively, which was 15 times greater for Zn but similar for Pb as compared with the MNE. The greatest leaf values of Zn and Pb for the OME were about 67 and 400 times more than for the NME. At Zn/Pb level of 1.0/0.2 mM, the OME shoots achieved the maximum Zn and Pb accumulation. No antagonistic effect of Zn and Pb was noted in the OME grown below tolerable metal levels, and Zn addition could enhance Pb uptake by the roots. The extraordinary ability to tolerate and accumulate both Zn and Pb by the mined ecotype of Sedum alfredii H. could be valuable for phytoremediation of multimetal‐contaminated soils.     

Effect of vesicular-arbuscular mycorrhizae and zinc application on dry matter and zinc uptake of greengram (Vigna radiata L. Wilczek)

Summary In a greenhouse study we examined the effects of vesicular-arbuscular mycorrhizae (VAM) inoculation, using Glomus macrocarpum and of Zn application on dry matter production and Zn uptake by greengram in two mollisols. The VAM inoculation significantly increased the dry weight of different plant parts and the Zn uptake in both soils. Inoculated plants showed a greater response to the application of Zn at 2.5 and 5.0 mg kg^-1 soil in a Zn deficient clay loam soil. The inoculated plants also absorbed — more water than the uninoculated plants. Mass flow and diffusion were the principal processes by which Zn reached the plant roots; mass flow was particularly important in the absence of VAM in a sandy soil fertilized with higher Zn doses (5 and 10 mg kg^-1 soil). The greater supply of Zn to inoculated roots was attributed to an apparent diffusion process rather than to mass flow of Zn.     

The influence of vesicular-arbuscular mycorrhizae on uptake of 90Sr from soil by soybeans

The Stanford-DeMent technique was used in a study of the influence of the vesicular-arbuscular (VA) mycorrhizal fungus Endogone mosseae on ⁹⁰Sr uptake by soybean. Thirteen-day-old mycorrhizal soybean plants absorbed significantly more ⁹⁰Sr than control plants after 1, 3 or 7 days contact with ⁹⁰Sr amended sterilized or nonsterilized soil. The same positive influence of Endogone mosseae on Sr absorption was observed in a second study which allowed for ⁹⁰Sr uptake concurrent with mycorrhizal infection and development. In the Stanford-DeMent study, soil sterilization exerted a short (1 day) negative influence on the uptake of ⁹⁰Sr by mycorrhizal roots. In the second study, mycorrhizal roots absorbed more ⁹⁰Sr from sterilized soil than from unsterilized soil while the reverse occurred with the control plants. Infected plants in both studies showed an early decrease in dry matter yield.     

Kinetics of zinc uptake by mycorrhizal (VAM) and non-mycorrhizal corn (Zea mays L.) roots

Summary The kinetics of Zn absorption were studied in mycorrhizal (Glomus macrocarpum) and non-mycorrhizal roots of corn (Zea mays L.) at pH 6.0 at Zn concentrations of 75 mol to 1.07 mol m^-3. Five concentration-dependent phases of Zn absorption were recognized; phase 0 (1.5–4.0 mmol m^-3) was linear but the other four phases (4.0 mmol to 1.07 mol m^-3) obeyed Michaelis-Menten kinetics. At low concentrations (less than 4 mmol m^-3), sigmoidal kinetics of Zn absorption were observed. The absorption of Zn by mycorrhizal maize was greater at low concentrations but decreased at higher levels. This appeared to be a result of a higher maximal uptake rate in phase 1 and lower K _m values in the subsequent phases. Kinetic models yielding continuous isotherms could not account for the observed multiphasic pattern.Research paper no. 6820 through the Director, Experiment Station, G.B. Pant University of Agriculture and Technology, Pantnagar 263 145, UP, India     

Arbuscular mycorrhizae affect the N and C economy of nodulated Phaseolus vulgaris (L.) during NH4 nutrition

In the symbiosis between nodulated legume roots and arbuscular mycorrhizal (AM) fungi, the C and N economy can be influenced by the source of N-supply from either AM-derived NH₄⁺ uptake or nodule-derived biological nitrogen fixation (BNF). This relationship was investigated in terms of NH₄⁺ supply and BNF by the two symbionts. Nodulated Phaseolus vulgaris seedlings with and without AM, were hydroponically grown with either 0 N or 1 mM NH₄⁺ supply. Plants were harvested at 30 days after emergence and measurements were taken for biomass, N₂ fixation, photosynthesis, CO₂ and O₂ root respiration, calculated C and N economy. AM roots had higher NH₄⁺ uptake and this was associated with the suppression of BNF and nodule growth. The higher NH₄⁺ uptake in AM roots occurred with lower root maintenance respiration, compared to when N was derived from BNF. There was also an increase in the below-ground sink strength of NH₄⁺ fed AM roots compared to NH₄⁺ fed non-AM roots, as evidenced by the increases in root CO₂ and O₂ respiration and photosynthetic stimulation. These results indicate that although the AM root had higher total below-ground respiratory costs during NH₄⁺ nutrition, there were lower respiratory C costs associated with N derived from AM symbionts in comparison to N from BNF.     

The invasion of pea roots, pisum sativum l., by soil microorganisms, Acanthamoeba palestinensis (reich) and pseudomonas SP

Pea roots (Pisum sativum L. cultivar Tall Sugar White) were inoculated with Pseudomonas sp., isolated from the roots of Timothy grass (Phleum pratensis L.). Microscopic examination showed that the bacterium had invaded many epidermal and outer cortical cells of the root. The presence of the soil amoeba, Acanthamoeba palestinensis (Reich), which readily ingests Pseudomonas sp., did not alter the root damage. The amoebae were also found in the epidermis and outer cortex of pea roots. No amoebae were found inside pea roots when Pseudomonas sp. was absent. Bacterial invasion also occurred in pea roots grown in garden soil previously sterilized by γ-irradiation and inoculated with Pseudomonas sp., but not in the same soil inoculated and unsterilized. Timothy grass roots were similarly uninfected in inoculated unsterilized garden soil.     

The uptake and translocation of phosphate by mycelial strands of pine mycorrhizas

Uptake and rapid translocation of ³²P-orthophosphate to Pinus radiata mycorrhizas from soil by mycelial strands of Rhizopogon luteolus was demonstrated. In greenhouse material, translocation occurred from soil for at least 12 mm and some 30–80 per cent of phosphate absorbed from 5 × 10^?6M as KH₂PO₄ was translocated. In the field, translocation occurred for 12 cm. Uptake by excised mycelial strands was metabolically mediated. Translocation occurred more rapidly when the untreated ends of strands were placed in an osmoticum: polarity in translocation was also observed. It was concluded that uptake and translocation by mycelial strands (as distinct from individual hyphae) provide an effective method for mycorrhizal exploitation of large inter-root soil volumes and assist the plants in competition for nutrients.Large differences occurred between strains of the same species in mycelial strand growth in soil. Mycelial strands of R. luteolus grew through the test soil at 1.3-2.9 mm/day and along P. radiata roots at 1.7 mm/day at 25°C day and 16°C night temperatures.     

Effect of elevated concentrations of Cd and Zn in soil on spring wheat yield and the metal contents of the plants

The effect of increasing concentrations of Cd and Zn in a sandy soil on spring wheat (Triticum vulgare L.) yields and the metal contents of the plants was examined in a pot experiment to establish critical levels of these metals in soil. The metals were added (individually and jointly) to the soil as sulfates in the following doses (in μg g^?1, dry wt.): Cd — 2, 3, 5,10, 15, 25, and 50; Zn ?200, 300, 500, 1000, 1500, 2500, and 5000. Cadmium added to soil did not affect yields of wheat. The Zn dose of 1000 μg g^?1 strongly reduced crop yields; at 1500 μg g^? Zn dose wheat did not produce grain. The metal contents of wheat increased with increasing concentrations of Cd and Zn in soil up to 10.3 and 1587 μ g^? of Cd and Zn in straw, respectively. The concentrations of both metals were higher in straw than in grain by factors of 3–7 and 1.5–2 for Zn and Cd, respectively. The relationships between Cd and Zn contents of the plants and soils were best expressed by exponential equations. High concentrations of Zn in soils (1042 and 1542 μg g^?1) enhanced uptake of Cd by plants. The tested threshold concentrations of the metals in soils (3 μg g^?1 for Cd and 200–300 μg g^?1 for Zn) are safe for Zn but are too high for Cd in terms of protecting plants from excessive metal uptake. The critical Cd content of sandy soil should not exceed 1.5 μg g^?.     

Characterisation of endobacterial communities in ectomycorrhizas by DNA- and RNA-based molecular methods

The diversity of endobacteria associated with ectomycorrhizas of Suillus variegatus and Tomentellopsis submollis, in two Corsican pine (Pinus nigra) stands was analysed by cultivation-dependent and cultivation-independent molecular methods. Denaturing gradient gel electrophoresis (DGGE) analysis revealed the cultivable endobacterial communities associated with S. variegatus were similar within the same stand. The most abundant cultivable bacterial species belonged to the genera Pseudomonas and Burkholderia. Cultivation-independent molecular analysis indicated that the structure of the endobacterial communities in ectomycorrhizas was consistent across all samples regardless of ECM fungal species or the pine stand from which the samples were collected. However, comparison between rDNA- and rRNA-derived DGGE gels showed that metabolically active endobacterial species were not always detected in rDNA-based profiles. Clone libraries constructed from rRNA molecules indicated that Pseudomonas and Burkholderia spp. were metabolically active bacteria. As some of the most abundant cultivable bacteria, including Bacillus/Paenibacillus spp., were not detected in cultivation-independent DGGE profiles, a combination of cultivation-dependent and -independent approaches provided a more complete assessment of the diversity of endobacteria associated with ectomycorrhizas.