Metagenomics has proven to be one of the most important research fields for microbial ecology during the last decade. Starting from 16S rRNA marker gene analysis for the characterization of community compositions to whole metagenome shotgun sequencing which additionally allows for functional analysis, metagenomics has been applied in a wide spectrum of research areas. The cost reduction paired with the increase in the amount of data due to the advent of next-generation sequencing led to a rapidly growing demand for bioinformatic software in metagenomics. By now, a large number of tools that can be used to analyze metagenomic datasets has been developed. The Bielefeld-Gießen center for microbial bioinformatics as part of the German Network for Bioinformatics Infrastructure bundles and imparts expert knowledge in the analysis of metagenomic datasets, especially in research on microbial communities involved in anaerobic digestion residing in biogas reactors. In this review, we give an overview of the field of metagenomics, introduce into important bioinformatic tools and possible workflows, accompanied by application examples of biogas surveys successfully conducted at the Center for Biotechnology of Bielefeld University.

To follow the hypothesis that agricultural management practices affect structure and function of the soil microbiome regarding soil health and plant-beneficial traits, high-throughput (HT) metagenome analyses were performed on Chernozem soil samples from a long-term field experiment designated LTE-1 carried out at Bernburg-Strenzfeld (Saxony-Anhalt, Germany). Metagenomic DNA was extracted from soil samples representing the following treatments: (i) plough tillage with standard nitrogen fertilization and use of fungicides and growth regulators, (ii) plough tillage with reduced nitrogen fertilization (50%), (iii) cultivator tillage with standard nitrogen fertilization and use of fungicides and growth regulators, and (iv) cultivator tillage with reduced nitrogen fertilization (50%). Bulk soil (BS), as well as root-affected soil (RS), were considered for all treatments in replicates. HT-sequencing of metagenomic DNA yielded approx. 100 Giga bases (Gb) of sequence information. Taxonomic profiling of soil communities revealed the presence of 70 phyla, whereby Proteobacteria, Actinobacteria, Bacteroidetes, Planctomycetes, Acidobacteria, Thaumarchaeota, Firmicutes, Verrucomicrobia and Chloroflexi feature abundances of more than 1%. Functional microbiome profiling uncovered, i.a., numerous potential plant-beneficial, plant-growth-promoting and biocontrol traits predicted to be involved in nutrient provision, phytohormone synthesis, antagonism against pathogens and signal molecule synthesis relevant in microbe–plant interaction. Neither taxonomic nor functional microbiome profiling based on single-read analyses revealed pronounced differences regarding the farming practices applied. Soil metagenome sequences were assembled and taxonomically binned. The ten most reliable and abundant Metagenomically Assembled Genomes (MAGs) were taxonomically classified and metabolically reconstructed. Importance of the phylum Thaumarchaeota for the analyzed microbiome is corroborated by the fact that the four corresponding MAGs were predicted to oxidize ammonia (nitrification), thus contributing to the cycling of nitrogen, and in addition are most probably able to fix carbon dioxide. Moreover, Thaumarchaeota and several bacterial MAGs also possess genes with predicted functions in plant–growth–promotion. Abundances of certain MAGs (species resolution level) responded to the tillage practice, whereas the factors compartment (BS vs. RS) and nitrogen fertilization only marginally shaped MAG abundance profiles. Hence, soil management regimes promoting plant-beneficial microbiome members are very likely advantageous for the respective agrosystem, its health and carbon sequestration and accordingly may enhance plant productivity. Since Chernozem soils are highly fertile, corresponding microbiome data represent a valuable reference resource for agronomy in general.

The genus Pseudomonas comprises many known plant-associated microbes with plant growth promotion and disease suppression properties. Genome-based studies allow the prediction of the underlying mechanisms using genome mining tools and the analysis of the genes unique for a strain by implementing comparative genomics. Here, we provide the genome sequence of the strain Pseudomonas brassicacearum 3Re2-7, formerly known as P. trivialis and P. reactans, elucidate its revised taxonomic classification, experimentally verify the gene predictions by transcriptome sequencing, describe its genetic biocontrol potential and contextualize it to other known Pseudomonas biocontrol agents. The P. brassicacearum 3Re2-7 genome comprises a circular chromosome with a size of 6,738,544 bp and a GC-content of 60.83%. 6267 genes were annotated, of which 6113 were shown to be transcribed in rich medium and/or in the presence of Rhizoctonia solani. Genome mining identified genes related to biocontrol traits such as secondary metabolite and siderophore biosynthesis, plant growth promotion, inorganic phosphate solubilization, biosynthesis of lipo- and exopolysaccharides, exoproteases, volatiles and detoxification. Core genome analysis revealed, that the 3Re2-7 genome exhibits a high collinearity with the representative genome for the species, P. brassicacearum subsp. brassicacearum NFM421. Comparative genomics allowed the identification of 105 specific genes and revealed gene clusters that might encode specialized biocontrol mechanisms of strain 3Re2-7. Moreover, we captured the transcriptome of P. brassicacearum 3Re2-7, confirming the transcription of the predicted biocontrol-related genes. The gene clusters coding for 2,4-diacetylphloroglucinol (phlABCDEFGH) and hydrogen cyanide (hcnABC) were shown to be highly transcribed. Further genes predicted to encode putative alginate production enzymes, a pyrroloquinoline quinone precursor peptide PqqA and a matrixin family metalloprotease were also found to be highly transcribed. With this study, we provide a basis to further characterize the mechanisms for biocontrol in Pseudomonas species, towards a sustainable and safe application of P. brassicacearum biocontrol agents.

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Background For a sustainable production of food, research on agricultural soil microbial communities is inevitable. Due to its immense complexity, soil is still some kind of black box. Soil study designs for identifying microbiome members of relevance have various scopes and focus on particular environmental factors. To identify common features of soil microbiomes, data from multiple studies should be compiled and processed. Taxonomic compositions and functional capabilities of microbial communities associated with soils and plants have been identified and characterized in the past few decades. From a fertile Loess–Chernozem-type soil located in Germany, metagenomically assembled genomes (MAGs) classified as members of the phylum Thaumarchaeota/Thermoproteota were obtained. These possibly represent keystone agricultural soil community members encoding functions of relevance for soil fertility and plant health. Their importance for the analyzed microbiomes is corroborated by the fact that they were predicted to contribute to the cycling of nitrogen, feature the genetic potential to fix carbon dioxide and possess genes with predicted functions in plant-growth-promotion (PGP). To expand the knowledge on soil community members belonging to the phylum Thaumarchaeota, we conducted a meta-analysis integrating primary studies on European agricultural soil microbiomes. Results Taxonomic classification of the selected soil metagenomes revealed the shared agricultural soil core microbiome of European soils from 19 locations. Metadata reporting was heterogeneous between the different studies. According to the available metadata, we separated the data into 68 treatments. The phylum Thaumarchaeota is part of the core microbiome and represents a major constituent of the archaeal subcommunities in all European agricultural soils. At a higher taxonomic resolution, 2074 genera constituted the core microbiome. We observed that viral genera strongly contribute to variation in taxonomic profiles. By binning of metagenomically assembled contigs, Thaumarchaeota MAGs could be recovered from several European soil metagenomes. Notably, many of them were classified as members of the family Nitrososphaeraceae, highlighting the importance of this family for agricultural soils. The specific Loess-Chernozem Thaumarchaeota MAGs were most abundant in their original soil, but also seem to be of importance in other agricultural soil microbial communities. Metabolic reconstruction of Switzerland_1_MAG_2 revealed its genetic potential i.a. regarding carbon dioxide (CO$$_2$$ 2 ) fixation, ammonia oxidation, exopolysaccharide production and a beneficial effect on plant growth. Similar genetic features were also present in other reconstructed MAGs. Three Nitrososphaeraceae MAGs are all most likely members of a so far unknown genus. Conclusions On a broad view, European agricultural soil microbiomes are similarly structured. Differences in community structure were observable, although analysis was complicated by heterogeneity in metadata recording. Our study highlights the need for standardized metadata reporting and the benefits of networking open data. Future soil sequencing studies should also consider high sequencing depths in order to enable reconstruction of genome bins. Intriguingly, the family Nitrososphaeraceae commonly seems to be of importance in agricultural microbiomes.