Fine-tuning DNA approach and multiple enzyme assay to assess soil microbial properties under different forest floors.

Even in the current era of high-throughput molecular techniques, the enormous heterogeneity of soil still renders the assessment of its microbiota as an arduous challenge. We aimed to assess and monitor the microbial properties of forest topsoils (0-15 cm depth) at a small scale (2.5 cm intervals) under different vegetation representatives (grass, branches, moss, litter) of alpine areas (1400 m a.s.l; Trentino, Italy) at south vs. north exposure. To assess the vertical distribution of soil microbiota (biomass and activity), we performed and linked the fine-tuning DNA approach (extracellular vs. intracellular vs. total soil DNA) and multiple enzyme assay (12 hydrolases covering the principal biogeochemical cycles). The fine-tuning DNA approach was proved to be a sensitive tool to quantitatively assess the vertical distribution of the soil microbiota, taking into account also the well-known method-result effect. Likewise, the comparative assessment of multiple enzymatic activities allowed for the accurate characterisation of the soils under different forest floors, overcoming the probable misinterpretation of single enzyme determination due to their specificity. Overall, the coupled DNA-enzyme approach is suggested to accurately assess soil microbiological properties as a function of soil type (humus forms, physical and chemical properties, soil depth), forest floor and exposure. Furthermore, the quantitative monitoring of extracellular DNA along the topsoils in comparison to intracellular and total soil DNA provided further evidences of possible movement of soil DNA by vertical advection with evolutionary implications in terms of genetic exchange by natural transformation. Our results are suggested to provide information about the heterogeneous distribution of the soil microbiota and the interlinked effects of biotic and abiotic factors at a small scale, which should be considered for the data interpretation and/or the set up of an experimental design/sampling strategy in soil microbial ecology studies.