Impacts of sampling design on estimates of microbial community diversity and composition in agricultural soils

Authors: CASTLE, SARAH - University Of Minnesota; Samac, Deborah - Debby; SADOWSKY, MICHAEL - University Of Minnesota; ROSEN, CARL - University Of Minnesota; GUKNECHT, JESSICA - University Of Minnesota; KINKEL, LINDA - University Of Minnesota

Submitted to: Microbial Ecology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/2/2019
Publication Date: 3/9/2019
Citation: Castle, S.C., Samac, D.A., Sadowsky, M.J., Rosen, C.J., Guknecht, J.L., Kinkel, L.L. 2019. Impacts of sampling design on estimates of microbial community diversity and composition in agricultural soils. Microbial Ecology. 78:1-11. https://doi.org/10.1007/s00248-019-01318-6.
DOI: https://doi.org/10.1007/s00248-019-01318-6

Interpretive Summary: The members of microbial communities in soil and associated with plant roots can be easily identified and inventoried using high throughput DNA sequencing methods. However, information on the number of samples and replicates that are needed to accurately represent the complete assemblage and the influence of sampling methods community composition has not been investigated previously. Microbial communities were identified in a variable number of individual soil samples as well as composite soil samples. Results indicated that either a single sample per plot or a composite sample was sufficient for distinguishing bacterial and fungal communities among the three agricultural sites. However, a composite sample reduced the experimental variability, which increased the statistical significance of differences in community composition among sites. Moreover, a single sample did not capture all of the diversity in community members compared to several samples or a single composite samples. Overall these findings indicate that collecting several replicated individual samples or creating a composite sample may be most suitable for microbial community studies in agricultural soils. Accurate and economical inventories of soil microbes will enable researchers and producers to develop approaches for achieving healthy soils for sustainable crop production.

Technical Abstract: Soil microbiota play important and diverse roles in agricultural crop nutrition and productivity. Yet, despite increasing efforts to characterize soil bacterial and fungal assemblages, it is challenging to disentangle the influences of sampling design on assessments of communities. Here we sought to determine whether composite samples - often analyzed as a low cost, low effort alternative to replicated individual samples - provide representative summary estimates of microbial communities. At three agricultural research sites planted with an oat cover crop and regionally reflective of soil properties across Minnesota, we conducted amplicon sequencing for soil bacterial and fungal communities (16SV4 and ITS2) of replicated individual soil samples or homogenized composite samples. We compared soil microbiota from within and among plots and then among agricultural sites using both sampling strategies. Results indicated that either a single sample per plot or a composite sample was sufficient for distinguishing bacterial and fungal communities among the three agricultural sites. However, compositing reduced compositional variability among plots from each site, which increased the statistical significance of differences in community composition among sites. Analysis of a single sample captured only a fraction of the number of OTUs, diversity, and compositional variability compared to analysis of several individual samples or a single composite sample. On average, analysis of two or three individual samples from within a single plot offered greater compositional coverage (i.e. greater number of OTUs) than a single composite. Overall these findings indicate that collecting several replicated individual samples or creating a composite sample may be most suitable for microbiome studies in agricultural soils.