DOI: 10.5281/zenodo.18755316
title: Database of soil fungi and bacteria under specific pesticide management

description: Database of soil fungi and bacteria under specific pesticide management

Contributors: Veronika Řezáčová, Ema Némethová, Oushadee A.J. Abeyawardana, Markéta Mayerová

Affiliation: Czech Agrifood Research Center, Drnovská 507/73, CZ-160 00 Prague 6, Czech Republic

Soil microorganisms play a central role in nutrient cycling, organic matter transformation and overall soil ecosystem functioning, yet their responses to agricultural chemical inputs remain insufficiently understood. While many studies have explored pesticide effects on individual microbial groups or enzymatic activities, less attention has been given to how broad-spectrum field applications of herbicides, insecticides and fungicides shape the taxonomic composition of complex soil microbial assemblages.

In this dataset, we present high-throughput DNA metabarcoding data of soil bacterial (16S) and fungal (ITS) communities collected from two field experiments subjected to controlled applications of herbicides and an insecticide. Soil samples were processed using Illumina sequencing, and the resulting reads were processed through basic bioinformatic steps (quality filtering and OTU assignment).

Database of soil fungi and bacteria contribute to the realization of the project´s goal to identify appropriate and inappropriate pesticides from the point of the view of their impact on the non-target organisms and soil characteristics.

Field I.

The field experiment was established in 2024 in the experimental area of the Czech Agrifood Research Center in Prague – Ruzyně (previously Crop Research Institute), (50.083986° N, 14.310269° E). The trial area was split into 20 randomised plots with 2 different herbicide treatments in 8 repetitions and control without herbicides in 4 repetitions. Herbicide treatments differed in the mode of action and target weeds (Table 1). Winter wheat was grown in the experimental field in 2024 and spring barley in 2025.

Table 1. Summary of the herbicides and active ingredients used in the trial. Classification Group by Herbicide Resistance Action Committee (HRAC).






Herbicide 



Dose



Formulation



Active ingredient



Content of a.i.



Mode

of action



Legacy

HRAC code





Agritox 50 SL



1.5 l/ha



EC



MCPA



500 g/l



4



O





Glean 75 PX (2024)



15 g/ha



WG



chlorsulfuron



750 g/kg



2



B





Tribenuron methyl (2025)



18.75 g/ha



WG



tribenuron methyl 



750 g/kg



2



B





 

The area of each plot was 100 m2 and the 10 by 10 m plots were separated from field boundaries and from each other by 2 m on all sides to eliminate interaction between plots. Herbicides were applied post emergency in spring (April 26, 2024; May 19, 2025) from the tillering crop stage to the beginning of stem elongation (BBCH 21–31) by the Agrio-Napa 12 sprayer. At the beginning of March, it was mineral fertilized with LAD (ammonium nitrate with dolomite - NH4NO3 + CaMg (CO3)2; 27 % N) at a dose 100 kg/ha. 

Mixed disturbed soil samples for microbiological and physicochemical analyses were taken from the 0-15cm upper soil layer in each replication before herbicide application (April 24, 2024), 14 days after herbicide application (May 9, 2024) and 7 weeks after herbicide application (June 14, 2024).  A total of 20 soil samples were collected at each sampling. The soil samples were subsequently dried and sieved through a 2 mm sieve, thus simultaneously homogenised. In 2025, soil sampling was carried out only once on June 3, 2025.

 

 

Field II

The field experiment was established in 2024 in the experimental area of the Czech Agrifood Research Center in Prague – Ruzyně (previously Crop Research Institute). The total area of the experiment is about 11 ha including the protective area around the entire experiment. The experimental area is divided into two halves, 120m wide and 300m long.  One half was treated with insecticide Decis forte (active ingredient deltamethrin) at a dose 62.5 ml/ha on the following dates: June 17, 2024, and June 3, 2025. The other half was without insecticide treatment. Both areas are further divided into other halves. One half was treated with herbicide Agritox (active ingredient MCPA) at a dose 1.5l/ha on the following dates: May 15, 2024, and May 24, 2025. The other was treated with hoeing only. We thus obtained 4 strips 60m wide with following treatment combinations: (A) herbicide + insecticide; (B) hoeing + insecticide; (C) hoeing; (D) herbicide. Spring wheat was grown in the experimental field in 2024 and winter triticale in 2025. It was fertilized with mineral nitrogen at a dose of 150 kg N/ha before sowing and with 39 kg N/ha (DAM 390 - ammonium nitrate with urea) in the tillering phenophase.

In the middle of each strip (i.e. treatment), 8 sampling sites were marked in a row, 20 m apart from each other. Mixed disturbed soil samples for microbiological and physicochemical analyses were taken from the 0-15cm upper soil layer at each sampling site 14 days after herbicide application (May 29, 2024, and May 6, 2025) and 14 days after insecticide application (June 20, 2024, and June 17, 2025). A total of 32 soil samples were collected at each sampling. Further sample processing was the same as for Field I.

DNA analyses

The procedure applies to both trials. 

DNA from soil was isolated by DNeasy PoweSoil DNA Isolation Kit (Qiagen) and DNA from roots was isolated by NucleoSpin Plant II Kit (Macherey-Nagel).

Extracted DNA was used for amplification of bacterial DNA in V4 region by unique tagged primer 515F and 806R. PCR amplification was performed by TP HS DNA-free 2x Master Mix (Top-Bio) under conditions  initial denaturation at 94C for 4 mins, followed by 32 cycles of denaturation at 94C for 45s, annealing at 50C for 30s and elongation at 72C for 70s, final elongation was at 72C for 10 mins.

In order to amplify fungal DNA we used unique tagged primer gITS7 and ITS4 for ITS2 and ITS4 regions. PCR amplification was performed by PPP Combi Master Mix (Top-Bio) under conditions initial denaturation at 94C for 5 mins, followed by 35 cycles of denaturation at 94C for 30s; annealing at 60C for 30s and elongation at 72C for 30s; final elongation was at 72C for 10 mins.

All samples were then purified with Gel/PCR DNA Fragments Extraction Kit (Geneaid) and concentration was measured by the Quantifluor ONE dsDNA system (Promega). 

DNA was then pooled together in equimolar concentration and sent for sequencing using pair-end 2x250 bp reads on the Illumina MiSeq platform (Illumina) at SeqMe. Bioinformatic analyses were performed using with SEED 2.1.2 pipeline. We assembled the paired-end reads, and removed the low-quality reads, short reads (350 bp for bacteria and 400 bp for fungi). We then clustered the sequences by applying a 97% similarity threshold. Chimeras were identified and excluded using USEARCH. Taxonomic assignment was performed by BLASTing representative OTU sequences against the SILVA 138.1 reference database, which we used to classify the operational taxonomic units (OTUs) into species and higher taxonomic units.

Funding: Development for this work is funded primarily by the Technology Agency of the Czech Republic, project SS07020100: “The impact of plant protection products on non-target biodiversity: soil microorganisms, invertebrates and wild plants”, and the Ministry of Agriculture of the Czech Republic, institutional support MZE-RO0426.

The database was approved in April 2026 by the Ministry of Agriculture of the Czech Republic.

 

Citation:

Řezáčová, V., Némethová, E., Abeyawardana, O.A.J & Mayerová M. (2026). Database of soil fungi and bacteria under specific pesticide management [Data set]. Zenodo. 10.5281/zenodo.18755317.