Analysis of broad-scale differences in microbial community composition of two pristine forest soils

Broad-scale differences in soil microbial community composition were analyz ed in two contrasting soils using DNA reassociation and % G+C profiles for analysis on the community-level, and filter- and whole cell hybridization t echniques for a coarse-level characterization of larger phylogenetic groups of bacteria. Reassociation analysis of DNA from bacterial fractions extrac ted from the organic soil Seim and the mineral soil Hau revealed similar co mplexity of the communities with 5700 and 4900 different bacterial genomes (g soil [dry wt])(-1), respectively. Thermal denaturation studies showed wi de % G+C distributions in DNA from bacteria of both soils. Differences in t he median % G+C with 55 to 61% for the bacterial community in soil Seim and 61 to 66% for that in soil Hall indicated a higher proportion of bacteria with a high DNA G+C content in soil Hau. In situ hybridization with fluores cent (Cy3-labeled) probes targeting larger phylogenetic groups showed minor differences between both soils, and between direct detection of bacteria i n dispersed soil slurries and in bacterial fractions extracted from soils t hough about 90% of the total bacteria were lost during extraction. In dispe rsed slurries of both soils, only probes ALF1b, SRB385, and PLA46 hybridize d to cells accounting for more than 1% of the DAPI-stained cells, while num bers obtained after hybridization with probes ARCH915, BET42a, GAM42a, HGC6 9a, and CF319a were below the detection limit set at <1%. These results wer e confirmed by in situ hybridization with horseradish peroxidase (HRP)-labe led probes and subsequent Cy3-tyramide signal amplification. In contrast, d ot blot hybridization with probe HGC69a indicated significant amounts of Gr am-positive bacteria with a high DNA G+C content in both soils. These could subsequently be visualized in non-dispersed soil slurries by in situ hybri dization with HRP-labeled probe HGC69a and Cy3-tyramide signal amplificatio n. Filamentous Gram-positive bacteria with a high DNA G+C content, likely a ctinomycetes, which are present in soil Hau in significant numbers are obvi ously destroyed by procedures used for soil dispersion.