Éditeur(s) : HAL CCSD Elsevier
Résumé : Extracting and purifying a representative fraction of bacteria from soil is necessary for the application of many techniques of microbial ecology. Here the influence of different soil types on the quantitative and qualitative recovery of bacteria by soil grinding and Nycodenz density gradient centrifugation was investigated. Three soils presenting contrasted physicochemical characteristics were used for this study. For each soil, the total (AODC: acridine orange direct count) and culturable (cfu: colony-forming units) bacterial densities were measured in three distinct fractions: (i) the primary soil, (ii) the soil pellet (soil remaining after centrifugation), and (iii) the extracted cells. The automated-ribosomal intergenic spacer analysis (A-RISA) was used to characterize the community structure directly from the DNA extracted from each fraction. The physicochemical characteristics of soils were found to influence both the efficiency of bacterial cell recovery and the representativeness of the extracted cells in term of community structures between the different fractions. Surprisingly, the most representative extracted cells were obtained from the soil exhibiting the lowest efficiency of cell recovery. Our results demonstrated that quantitative and qualitative cell recovery using Nycodenz density gradient centrifugation are not necessarily related and could be differentially biased according to soil type.
ISSN: 1164-5563

hal-01031904
https://hal.archives-ouvertes.fr/hal-01031904
DOI : 10.1016/j.ejsobi.2005.08.003

Éditeur(s) : HAL CCSD Wiley
Résumé : International audience
Three soil DNA extraction procedures (homemade protocols and commercial kit) varying in their practicability were applied to contrasting soils to evaluate their efficiency in recovering: (i) soil DNA and (ii) bacterial diversity estimated by 16S rDNA pyrosequencing. Significant differences in DNA yield were systematically observed between tested procedures. For certain soils, 10 times more DNA was recovered with one protocol than with the others. About 15 000 sequences of 16S rDNA were obtained for each sample which were clustered to draw rarefaction curves. These curves, as well as the PCA ordination of community composition based on OTU clustering, did not reveal any significant difference between procedures. Nevertheless, significant differences between procedures were highlighted by the taxonomic identification of sequences obtained at the phylum to genus levels. Depending on the soil, differences in the number of genera detected ranged from 1% to 26% between the most and least efficient procedures, mainly due to a poorer capacity to recover populations belonging to Actinobacteria, Firmicutes or Crenarchaeota. This study enabled us to rank the relative efficiencies of protocols for their recovery of soil molecular microbial biomass and bacterial diversity and to help choosing an appropriate soil DNA extraction procedure adapted to novel sequencing technologies.
ISSN: 1751-7915

hal-01546177
https://hal.archives-ouvertes.fr/hal-01546177
DOI : 10.1111/j.1751-7915.2011.00307.x