Analysis of bacterial communities in heavy metal-contaminated soils at different levels of resolution

The impact of heavy metal contamination on soil bacterial communities was studied in soils amended for many years with sewage sludge contaminated with heavy metals to varying extents. At the broad level of resolution, DNA reassociation analysis indicated a dramatic decrease in bacterial diversity from 16000 bacterial genomes (g soil [wet wt]-1 in the non-contaminated soil to 6400 bacterial genomes (g soil [wet wt])-1 in soil with low metal amendments and only 2000 bacterial genomes (g soil [wet wt]-1 in soil with high metal amendments. No differences between bacterial communities of these soils, however, were displayed in the %G+C profiles analysed by thermal denaturation. At a coarse level of characterisation, in situ hybridisation analysing larger phylogenetic groups of bacteria revealed a general decrease in the percentage of cells detected with probes ARCH915, BET42a, GAM42a, SRB385, CF319a, LGCb and HGC69a with increasing metal amendment. Only cells detected with probe ALF1b increased significantly from 3.1 +/- 0.8% of the cells detected by the domain-specific probe EUB338 in the non-contaminated soil to 6.5 +/- 1.3% in soil with high metal amendments. These shifts in populations of larger phylogenetic groups were largely confirmed by dot blot analysis of 16S and 23S rDNA clone libraries from bacteria in soil with low metal and high metal amendments, respectively. For a fine-level characterisation, 72 clones of 16S rDNA libraries were identified by comparative sequence analysis. A few sequences could not be assigned to the major taxa described. Most of the sequences were assigned to the Gram-positive bacteria with a high DNA G+C content (45%) and the alpha-subdivision of Proteobacteria (24%). However, only minor differences were seen between bacterial communities from the low and high metal soils. In the soil with high metal amendment, more sequences clustered to the alpha-subdivision of Proteobacteria, while in the low metal soil, more sequences clustered to the Gram-positive bacteria with a high DNA G+C content.