Soil DNA Samples Research Articles

Soil fungal community assembly in a primary successional glacier forefront ecosystem as inferred from rDNA sequence analyses.

• Soil fungal community assembly in nonvegetated areas on the forefront of a receding glacier was analyzed by cloning of the PCR-amplified partial small subunit (18S) of the ribosomal RNA genes (rDNA) from soil DNA samples. • Fungal sequences obtained from areas adjacent to the present glacier terminus (young substrate) represented three fungal phyla, whereas those obtained adjacent to the terminal moraine (old substrate) were distributed among Ascomycetes and Hymenomycetes. The cloned sequences from both substrates represented mainly filamentous ascomycetes or basidiomycetes with a likely affinity to Agaricales. Unexpected biotrophic fungi with affinities to Taphrinomycetes, Urediniomycetes (the rust fungi) and Ustilaginomycetes (the smut fungi) plus an unknown, likely chytridiomycetous group were detected exclusively in the young substrates. • These observations of biotrophic fungi are attributed to an aerially deposited, dormant spore bank, which may also be present in the older substrate but is masked by larger active mycelial biomass. • This study underlines the importance of stochastic events and airborne spore deposition in the assembly of early fungal communities.

Identification of the key genes of naphthalene catabolism in soil DNA

The key genes nahAc and xylE of the naphthalene catabolism of fluorescent Pseudomonas spp. in the total soil DNA samples were detected by the polymerase chain reaction (PCR) technique. The collection of fluorescent Pseudomonas spp. was screened for the occurrence of these genes. The results obtained show the possibility of using this approach in the goal-directed search for plasmid-containing naphthalene-degrading fluorescent pseudomonads in soil. The distribution of the naphthalene catabolism genes in soils contaminated with creosote and petroleum products was also studied.

Spatial distribution of the below-ground mycelia of an ectomycorrhizal fungus inferred from specific quantification of its DNA in soil samples

In natural forest ecosystems several ectomycorrhizal fungal species cohabit on host plant root systems. To evaluate the ecological and functional impact of each species, it is necessary to appreciate the distribution and abundance of its mycelia in the soil. We developed a competitive PCR (cPCR) method for the basidiomycete Hebeloma cylindrosporum that allows quantification of its DNA in complex DNA mixtures extracted directly from soil samples. The target sequence chosen for the cPCR analysis was a 533-bp fragment of the nuclear ribosomal intergenic spacer, amplified using two species-specific primers. The detection threshold of the cPCR protocol developed was 0.03 pg of genomic DNA. This method was applied to soil samples collected from beneath and at various distances from a group of fruit bodies in a Pinus pinaster forest stand. The results revealed that H. cylindrosporum below-ground biomass was concentrated directly underneath the fruit bodies or very close to them, while no DNA of this species could be detected in soil samples collected at more than 50 cm away. In the vicinity of fruit bodies, H. cylindrosporum soil DNA concentration varied considerably (between 10 and 0.07 ng g soil(-1)) and decreased sharply with increased distance from the fruit bodies. This work demonstrates the potential of competitive quantitative PCR for the study of the distribution, abundance and persistence of the mycelia of an ectomycorrhizal fungal species in soil.

Spatial distribution of the below-ground mycelia of an ectomycorrhizal fungus inferred from specific quantification of its DNA in soil samples

In natural forest ecosystems several ectomycorrhizal fungal species cohabit on host plant root systems. To evaluate the ecological and functional impact of each species, it is necessary to appreciate the distribution and abundance of its mycelia in the soil. We developed a competitive PCR (cPCR) method for the basidiomycete Hebeloma cylindrosporum that allows quantification of its DNA in complex DNA mixtures extracted directly from soil samples. The target sequence chosen for the cPCR analysis was a 533-bp fragment of the nuclear ribosomal intergenic spacer, amplified using two species-specific primers. The detection threshold of the cPCR protocol developed was 0.03 pg of genomic DNA. This method was applied to soil samples collected from beneath and at various distances from a group of fruit bodies in a Pinus pinaster forest stand. The results revealed that H. cylindrosporum below-ground biomass was concentrated directly underneath the fruit bodies or very close to them, while no DNA of this species could be detected in soil samples collected at more than 50 cm away. In the vicinity of fruit bodies, H. cylindrosporum soil DNA concentration varied considerably (between 10 and 0.07 ng g soil −1) and decreased sharply with increased distance from the fruit bodies. This work demonstrates the potential of competitive quantitative PCR for the study of the distribution, abundance and persistence of the mycelia of an ectomycorrhizal fungal species in soil.

Soil conservation for DNA preservation for bacterial molecular studies

The aim of this study was to select a method for preserving bacterial DNA in soil samples in cases where there are no possibilities of using freezing or cooling methods. To overcome this difficulty, we hypothesized that adding absolute ethanol to soil samples could be as successful to preserve bacteria as it is to preserve insect or tissue samples for molecular studies. In an attempt to test this assumption, we compared four conservation conditions. After inoculation of soil samples with Escherichia coli, they were either kept at 28 °C, stored in the cold (4 °C), dried at 60 °C, or treated with absolute ethanol. The relative effectiveness of the methods was evaluated by using both DNA recoveries and bacterial 16S rDNA amplification as criteria. Two kinds of soils, i.e. sandy clay and clayey soil, were used. Soil conservation was tested for seven time periods ranging from 2 d to 1 year after bacterial inoculation. Results showed that cold conservation or addition of absolute ethanol to the samples yielded similar DNA recoveries. For these treatments, successful amplifications are still obtained after one year of conservation. The ethanol conservation of soil samples appears to be the easiest method to preserve the bacterial DNA because of its reliability and field convenience.