Effects of fertilization with urban and agricultural organic wastes in a field trial – Prokaryotic diversity investigated by pyrosequencing

Abstract

The impact of different fertilizer treatments on prokaryotic diversity in a Danish urban waste field trial was investigated using tag-encoded amplicon pyrosequencing. The field trial was established in 2003 to investigate the application of urban organic waste as fertilizer in agriculture and to identify the effects on soil quality. The fertilizers (e.g. composted organic household waste, sewage sludge and human urine) contain a large amount of nutrients but possibly also undesirable toxic compounds that may influence the bacterial flora in the soil. A 561bp fragment of the 16S rRNA gene flanking the V4, V5 and V6 regions, was amplified from each soil sample, tagged and sequenced using pyrosequencing. The major classified bacterial phyla and proteobacterial classes for all treatments were Actinobacteria, Acidobacteria and Betaproteobacteria, while the Crenarchaeota was the most frequent phylum of Archaea. No major changes in the community composition due to different fertilizer treatments were found, demonstrating a high robustness of the soil microbiota. However, some differences were observed e.g. Cyanobacteria were most frequent in the unfertilized soil, in comparison to the soils treated with nitrogen containing fertilizers and Firmicutes had higher occurrence in the soil with the composted household waste compared to all other treatments. Additionally, we used quantitative PCR (qPCR) to quantify specific bacterial groups, and used these numbers to convert the relative abundances of all bacteria obtained by pyrosequencing, to the actual numbers present in one gram of soil. All treatments resulted in a total number of bacteria between 1.99×109 and 4.11×109 gram−1 soil.