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Methods for Experiment 284 -

Sampling-Streptomyces pathogen suppressive activity in plant communities varying in diversity

We sampled soil microbiomes under the dominant influence of four perennial prairie plant species (two C4 grasses: Andropogon gerardii and Schizachyrium scoparium; two legumes: Lespedeza capitata and Lupinus perennis), with each species grown in communities planted to 1-, 4-, 8-, or 16-species mixtures (Tilman et al. 2001). The plots sampled are located in the Big Biodiversity field. There were 48 soil samples collected in total (4 plant species X 4 plant richness treatments X 3 replicates). Detailed methods on sample collection and processing have been reported elsewhere (Bakker-etal_SoilBiolBiochem_2013).

acge284 - Streptomyces pathogen suppressive activity in plant communities varying in diversity

Methods-Streptomyces pathogen suppressive activity in plant communities varying in diversity

Streptomyces antagonistic activity was assessed in vitro as an estimate of the potential for suppression of plant pathogens. Briefly, antagonistic activity was measured as the density (log colony-forming units [CFU] g-1) and proportion of colonies on a Streptomyces-selective medium that created visible zones of inhibition when overlaid with an indicator organism. We used three different Streptomyces isolates (Davelos-etal_MicrobialBiology_2004) as indicators, and analyzed mean values across indicator strains. The intensity of inhibition was assessed as the mean radius of inhibition zone sizes across inhibitory colonies for a given soil sample. Detailed methods, reporting and interpretation of these data have been presented elsewhere (Bakker-etal_SoilBiolBiochem_2013). Here, we link these microbial community functional data to detailed characterization of community composition and structure. Our microbiome sequence data gave particular attention to the Streptomyces, but included taxa across the domain Bacteria. We took this approach because we expect that interactions with phylogenetically distinct taxa may be important to the generation and maintenance of the inhibitory phenotypes that contribute to pathogen suppression by Streptomyces. Our dataset included amplicon sequencing from PCR reactions using sets of 16S rRNA gene primers that were either universal and non-selective ('bacteria' dataset), or selective for Streptomyces ('Streptomyces' dataset). Detailed descriptions of the generation and processing of these sequence data have been reported elsewhere (Bakker-etal_SoilBiolBiochem_2013). Both sequence datasets were processed similarly, and operational taxonomic units (OTUs) were defined using a 97% sequence similarity criterion. The sample of bacterial communities consisted of 476,573 high quality sequence reads, which were distributed across 26,153 OTUs belonging to 16 different phyla. The Streptomyces-targeted dataset consisted of 59,184 high quality sequence reads, which were distributed across 409 OTUs. The vast majority of these sequence reads (> 90%) belonged to the Streptomyces, although other taxa within the phylum Actinobacteria were also represented at low relative abundances.