The role of fungal denitrifiers in the nitrous oxide emission from soils with organic residues amendment

Abstract

Different biologic processes are responsible for the nitrous oxide (N2O) production from soils. So, decipher the main players is of paramount importance due to global warming, since N2O is a potent greenhouse gas. It is well known that the combined application of organic residues with inorganic nitrogen (N) usually increase N2O emissions and change the structure of the soil microbiome. The use of organic vinasse, a by-product of ethanol production, applied in the sugarcane fields is no different. The present project aims to investigated the role of the fungal denitrifier community on the N2O production in soils amended with straw, concentrated vinasse (CV) plus N fertilizer in two different seasons, rainy (RS) and dry (DS) season. Fungal denitrifiers play an important role in N2O production, with a contribution from 17 to 89%, but we have very limited knowledge about who they are and what they are doing in the soil and their distribution, consequently their functions in the ecosystem. The limitation has been decreasing with the recently designed fungal nirK and P450nor PCR primers. However, there are still problems with the specificity of the primers. In a previews study, we found a positive correlation of qPCR nirK-fungi with N2O in sugarcane soils, however, we must have in mind that the literature primers for fungal denitrifiers are degenerated and also amplify bacterial nirK. During the post-doctorate (post-doctorate Project - FAPESP 2018/20698-6) we found that, on average, only 32% of the nirK-fungal amplified with the literature primer were actually nirK from fungi, 68% of the total relative abundance were from bacteria (results not published). So, we have biased results that need to be improved. Based on this, the specific goal of this project is to design a nirK-fungal primers to soils with sugarcane, consequently decipher the role of fungal denitrifier in the production of N2O in tropical soils. The field experiments were conducted in Brazil and N2O emissions were determined. In parallel, the air and soil temperatures were evaluated and soil samples were taken for soil moisture, concentrations of N-NO3- and N-NH4+ (field studies in Brazil) and for nucleic acids (DNA) extractions (to be done in the Netherland). After DNA extractions, the bacterial and archaeal amoA gene, bacterial nirK, bacterial nirS and nosZ, and total bacteria (16S rRNA) and fungi (18S rRNA) will be quantified by real-time PCR. At NIOO, we are going to employ a deep nirK-fungal amplicon sequencing approach to design specific nirK-fungal primers for our samples and after that quantify and determine their specificity. These results can be used as an input tool to determine sustainable management practices for organic residues. IAC group members have strong expertise in soil sciences, sugarcane nitrogen nutrition (N-fixing bacteria and fertilizer), bioenergy wastes recycling, soil fertility, bioenergy crop management, plant nutrition, and greenhouse gases emissions, whereas the NIOO group members have strong expertise in soil ecology, microbial ecology, ecological genomics, molecular biology, metagenomics, metatranscriptomics, and bioinformatics. Therefore, the exchange of researchers is fundamental not only for the success of the project but also give them a thorough education opportunity, given the complementarity of both country's research teams. (AU)