The production of biogas by anaerobic digestion (AD) of agricultural residues, organic wastes, animal excrements, municipal sludge, and energy crops has a firm place in sustainable energy production and bio-economy strategies. Focusing on the microbial community involved in biomass conversion offers the opportunity to control and engineer the biogas process with the objective to optimize its efficiency. Taxonomic profiling of biogas producing communities by means of high-throughput 16S rRNA gene amplicon sequencing provided high-resolution insights into bacterial and archaeal structures of AD assemblages and their linkages to fed substrates and process parameters. Commonly, the bacterial phyla Firmicutes and Bacteroidetes appeared to dominate biogas communities in varying abundances depending on the apparent process conditions. Regarding the community of methanogenic Archaea, their diversity was mainly affected by the nature and composition of the substrates, availability of nutrients and ammonium/ammonia contents, but not by the temperature. It also appeared that a high proportion of 16S rRNA sequences can only be classified on higher taxonomic ranks indicating that many community members and their participation in AD within functional networks are still unknown. Although cultivation-based approaches to isolate microorganisms from biogas fermentation samples yielded hundreds of novel species and strains, this approach intrinsically is limited to the cultivable fraction of the community. To obtain genome sequence information of non-cultivable biogas community members, metagenome sequencing including assembly and binning strategies was highly valuable. Corresponding research has led to the compilation of hundreds of metagenome-assembled genomes (MAGs) frequently representing novel taxa whose metabolism and lifestyle could be reconstructed based on nucleotide sequence information. In contrast to metagenome analyses revealing the genetic potential of microbial communities, metatranscriptome sequencing provided insights into the metabolically active community. Taking advantage of genome sequence information, transcriptional activities were evaluated considering the microorganism’s genetic background. Metaproteome studies uncovered enzyme profiles expressed by biogas community members. Enzymes involved in cellulose and hemicellulose decomposition and utilization of other complex biopolymers were identified. Future studies on biogas functional microbial networks will increasingly involve integrated multi-omics analyses evaluating metagenome, transcriptome, proteome, and metabolome datasets.Electronic supplementary materialThe online version of this article (10.1007/s00253-018-8976-7) contains supplementary material, which is available to authorized users.
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.
High precision of measurement of methane potential is important for the economic operation of biogas plants in the future. The biochemical methane potential (BMP) test based on the VDI 4630 protocol is the state-of-the-art method to determine the methane potential in Germany. The coefficient of variation (CV) of methane yield was >10% in several previous inter-laboratory tests. The aim of this work was to investigate the effects of inoculum and the digestion system on the measurement variability. Methane yield and methane percentage of five substrates were investigated in a Hohenheim biogas yield test (D-HBT) by using five inocula, which were used several times in inter- laboratory tests. The same substrates and inocula were also tested in other digestion systems. To control the quality of the inocula, the effect of adding trace elements (TE) and the microbial community was investigated. Adding TE had no influence for the selected, well- supplied inocula and the community composition depended on the source of the inocula. The CV of the specific methane yield was <4.8% by using different inocula in one D-HBT (D-HBT1) and <12.8% by using different digestion systems compared to D-HBT1. Incubation time between 7 and 14 days resulted in a deviation in CV of <4.8%.
BackgroundTo elucidate biogas microbial communities and processes, the application of high-throughput DNA analysis approaches is becoming increasingly important. Unfortunately, generated data can only partialy be interpreted rudimentary since databases lack reference sequences.ResultsNovel cellulolytic, hydrolytic, and acidogenic/acetogenic Bacteria as well as methanogenic Archaea originating from different anaerobic digestion communities were analyzed on the genomic level to assess their role in biomass decomposition and biogas production. Some of the analyzed bacterial strains were recently described as new species and even genera, namely Herbinix hemicellulosilytica T3/55T, Herbinix luporum SD1DT, Clostridium bornimense M2/40T, Proteiniphilum saccharofermentans M3/6T, Fermentimonas caenicola ING2-E5BT, and Petrimonas mucosa ING2-E5AT. High-throughput genome sequencing of 22 anaerobic digestion isolates enabled functional genome interpretation, metabolic reconstruction, and prediction of microbial traits regarding their abilities to utilize complex bio-polymers and to perform specific fermentation pathways. To determine the prevalence of the isolates included in this study in different biogas systems, corresponding metagenome fragment mappings were done. Methanoculleus bourgensis was found to be abundant in three mesophilic biogas plants studied and slightly less abundant in a thermophilic biogas plant, whereas Defluviitoga tunisiensis was only prominent in the thermophilic system. Moreover, several of the analyzed species were clearly detectable in the mesophilic biogas plants, but appeared to be only moderately abundant. Among the species for which genome sequence information was publicly available prior to this study, only the species Amphibacillus xylanus, Clostridium clariflavum, and Lactobacillus acidophilus are of importance for the biogas microbiomes analyzed, but did not reach the level of abundance as determined for M. bourgensis and D. tunisiensis.ConclusionsIsolation of key anaerobic digestion microorganisms and their functional interpretation was achieved by application of elaborated cultivation techniques and subsequent genome analyses. New isolates and their genome information extend the repository covering anaerobic digestion community members.Electronic supplementary materialThe online version of this article (10.1186/s13068-017-0947-1) contains supplementary material, which is available to authorized users.
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.
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