Abundance, classification and genetic potential of Thaumarchaeota in metagenomes of European agricultural soils: a meta-analysis.

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[Formula: see text]) 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.

The novel oligopeptide utilizing species Anaeropeptidivorans aminofermentans M3/9T, its role in anaerobic digestion and occurrence as deduced from large-scale fragment recruitment analyses.

Research on biogas-producing microbial communities aims at elucidation of correlations and dependencies between the anaerobic digestion (AD) process and the corresponding microbiome composition in order to optimize the performance of the process and the biogas output. Previously, Lachnospiraceae species were frequently detected in mesophilic to moderately thermophilic biogas reactors. To analyze adaptive genome features of a representative Lachnospiraceae strain, Anaeropeptidivorans aminofermentans M3/9T was isolated from a mesophilic laboratory-scale biogas plant and its genome was sequenced and analyzed in detail. Strain M3/9T possesses a number of genes encoding enzymes for degradation of proteins, oligo- and dipeptides. Moreover, genes encoding enzymes participating in fermentation of amino acids released from peptide hydrolysis were also identified. Based on further findings obtained from metabolic pathway reconstruction, M3/9T was predicted to participate in acidogenesis within the AD process. To understand the genomic diversity between the biogas isolate M3/9T and closely related Anaerotignum type strains, genome sequence comparisons were performed. M3/9T harbors 1,693 strain-specific genes among others encoding different peptidases, a phosphotransferase system (PTS) for sugar uptake, but also proteins involved in extracellular solute binding and import, sporulation and flagellar biosynthesis. In order to determine the occurrence of M3/9T in other environments, large-scale fragment recruitments with the M3/9T genome as a template and publicly available metagenomes representing different environments was performed. The strain was detected in the intestine of mammals, being most abundant in goat feces, occasionally used as a substrate for biogas production.

Adaptation of a microbial community to demand-oriented biological methanation.

BACKGROUND: Biological conversion of the surplus of renewable electricity and carbon dioxide (CO2) from biogas plants to biomethane (CH4) could support energy storage and strengthen the power grid. Biological methanation (BM) is linked closely to the activity of biogas-producing Bacteria and methanogenic Archaea. During reactor operations, the microbiome is often subject to various changes, e.g., substrate limitation or pH-shifts, whereby the microorganisms are challenged to adapt to the new conditions. In this study, various process parameters including pH value, CH4 production rate, conversion yields and final gas composition were monitored for a hydrogenotrophic-adapted microbial community cultivated in a laboratory-scale BM reactor. To investigate the robustness of the BM process regarding power oscillations, the biogas microbiome was exposed to five hydrogen (H2)-feeding regimes lasting several days. RESULTS: Applying various "on-off" H2-feeding regimes, the CH4 production rate recovered quickly, demonstrating a significant resilience of the microbial community. Analyses of the taxonomic composition of the microbiome revealed a high abundance of the bacterial phyla Firmicutes, Bacteroidota and Thermotogota followed by hydrogenotrophic Archaea of the phylum Methanobacteriota. Homo-acetogenic and heterotrophic fermenting Bacteria formed a complex food web with methanogens. The abundance of the methanogenic Archaea roughly doubled during discontinuous H2-feeding, which was related mainly to an increase in acetoclastic Methanothrix species. Results also suggested that Bacteria feeding on methanogens could reduce overall CH4 production. On the other hand, using inactive biomass as a substrate could support the growth of methanogenic Archaea. During the BM process, the additional production of H2 by fermenting Bacteria seemed to support the maintenance of hydrogenotrophic methanogens at non-H2-feeding phases. Besides the elusive role of Methanothrix during the H2-feeding phases, acetate consumption and pH maintenance at the non-feeding phase can be assigned to this species. CONCLUSIONS: Taken together, the high adaptive potential of microbial communities contributes to the robustness of BM processes during discontinuous H2-feeding and supports the commercial use of BM processes for energy storage. Discontinuous feeding strategies could be used to enrich methanogenic Archaea during the establishment of a microbial community for BM. Both findings could contribute to design and improve BM processes from lab to pilot scale.

Effect of Growth Media on the Diversity of Neocallimastigomycetes from Non-Rumen Habitats.

Anaerobic fungi (AF), belonging to the phylum Neocallimastigomycota, are a pivotal component of the digestive tract microbiome of various herbivorous animals. In the last decade, the diversity of AF has rapidly expanded due to the exploration of numerous (novel) habitats. Studies aiming at understanding the role of AF require robust and reliable isolation and cultivation techniques, many of which remained unchanged for decades. Using amplicon sequencing, we compared three different media: medium with rumen fluid (RF), depleted rumen fluid (DRF), and no rumen fluid (NRF) to enrich the AF from the feces of yak, as a rumen control; and Przewalski's horse, llama, guanaco, and elephant, as a non-rumen habitats. The results revealed the selective enrichment of Piromyces and Neocallimastix from the feces of elephant and llama, respectively, in the RF medium. Similarly, the enrichment culture in DRF medium explicitly manifested Piromyces-related sequences from elephant feces. Five new clades (MM1-5) were defined from llama, guanaco, yak, and elephant feces that could as well be enriched from llama and elephant samples using non-conventional DRF and NRF media. This study presents evidence for the selective enrichment of certain genera in medium with RF and DRF from rumen as well as from non-rumen samples. NRF medium is suggested for the isolation of AF from non-rumen environments.

Simultaneous Metabarcoding and Quantification of Neocallimastigomycetes from Environmental Samples: Insights into Community Composition and Novel Lineages.

Anaerobic fungi from the herbivore digestive tract (Neocallimastigomycetes) are primary lignocellulose modifiers and hold promise for biotechnological applications. Their molecular detection is currently difficult due to the non-specificity of published primer pairs, which impairs evolutionary and ecological research with environmental samples. We developed and validated a Neocallimastigomycetes-specific PCR primer pair targeting the D2 region of the ribosomal large subunit suitable for screening, quantifying, and sequencing. We evaluated this primer pair in silico on sequences from all known genera, in vitro with pure cultures covering 16 of the 20 known genera, and on environmental samples with highly diverse microbiomes. The amplified region allowed phylogenetic differentiation of all known genera and most species. The amplicon is about 350 bp long, suitable for short-read high-throughput sequencing as well as qPCR assays. Sequencing of herbivore fecal samples verified the specificity of the primer pair and recovered highly diverse and so far unknown anaerobic gut fungal taxa. As the chosen barcoding region can be easily aligned and is taxonomically informative, the sequences can be used for classification and phylogenetic inferences. Several new Neocallimastigomycetes clades were obtained, some of which represent putative novel lineages such as a clade from feces of the rodent Dolichotis patagonum (mara).

Phage Genome Diversity in a Biogas-Producing Microbiome Analyzed by Illumina and Nanopore GridION Sequencing.

The microbial biogas network is complex and intertwined, and therefore relatively stable in its overall functionality. However, if key functional groups of microorganisms are affected by biotic or abiotic factors, the entire efficacy may be impaired. Bacteriophages are hypothesized to alter the steering process of the microbial network. In this study, an enriched fraction of virus-like particles was extracted from a mesophilic biogas reactor and sequenced on the Illumina MiSeq and Nanopore GridION sequencing platforms. Metagenome data analysis resulted in identifying 375 metagenome-assembled viral genomes (MAVGs). Two-thirds of the classified sequences were only assigned to the superkingdom Viruses and the remaining third to the family Siphoviridae, followed by Myoviridae, Podoviridae, Tectiviridae, and Inoviridae. The metavirome showed a close relationship to the phage genomes that infect members of the classes Clostridia and Bacilli. Using publicly available biogas metagenomic data, a fragment recruitment approach showed the widespread distribution of the MAVGs studied in other biogas microbiomes. In particular, phage sequences from mesophilic microbiomes were highly similar to the phage sequences of this study. Accordingly, the virus particle enrichment approach and metavirome sequencing provided additional genome sequence information for novel virome members, thus expanding the current knowledge of viral genetic diversity in biogas reactors.

Variimorphobacter saccharofermentans gen. nov., sp. nov., a new member of the family Lachnospiraceae, isolated from a maize-fed biogas fermenter.

Strain MD1T is an anaerobic, Gram-stain-negative bacterium isolated from a lab-scale biogas fermenter fed with maize silage. It has a rod-shaped morphology with peritrichously arranged appendages and forms long chains of cells and coccoid structures. The colonies of MD1T were white, circular, slightly convex and had a smooth rim. The isolate is mesophilic, displaying growth between 25 and 45 °C with an optimum at 40 °C. It grew at pH values of pH 6.7-8.2 (optimum, pH 7.1) and tolerated the addition of up to 1.5% (w/v) NaCl to the medium. The main cellular fatty acids of MD1T are C14:0 DMA and C16:0. Strain MD1T fermented xylose, arabinose, glucose, galactose, cellobiose, maltose, maltodextrin10, lactose starch, and xylan, producing mainly 2-propanol and acetic acid. The genome of the organism has a total length of 4163427 bp with a G+C content of 38.5 mol%. The two closest relatives to MD1T are Mobilitalea sibirica P3M-3T and Anaerotaenia torta FH052T with 96.44 or 95.8 % 16S rRNA gene sequence similarity and POCP values of 46.58 and 50.58%, respectively. As MD1T showed saccharolytic and xylanolytic properties, it may play an important role in the biogas fermentation process. Closely related variants of MD1T were also abundant in microbial communities involved in methanogenic fermentation. Based on morphological, phylogenetic and genomic data, the isolated strain can be considered as representing a novel genus in the family Lachnospiraceae, for which the name Variimorphobacter saccharofermentans gen. nov., sp. nov. (type strain MD1T=DSM 110715T=JCM 39125T) is proposed.

Identification of Beneficial Microbial Consortia and Bioactive Compounds with Potential as Plant Biostimulants for a Sustainable Agriculture.

A growing body of evidence demonstrates the potential of various microbes to enhance plant productivity in cropping systems although their successful field application may be impaired by several biotic and abiotic constraints. In the present work, we aimed at developing multifunctional synthetic microbial consortia to be used in combination with suitable bioactive compounds for improving crop yield and quality. Plant growth-promoting microorganisms (PGPMs) with different functional attributes were identified by a bottom-up approach. A comprehensive literature survey on PGPMs associated with maize, wheat, potato and tomato, and on commercial formulations, was conducted by examining peer-reviewed scientific publications and results from relevant European projects. Metagenome fragment recruitments on genomes of potential PGPMs represented in databases were also performed to help identify plant growth-promoting (PGP) strains. Following evidence of their ability to coexist, isolated PGPMs were synthetically assembled into three different microbial consortia. Additionally, the effects of bioactive compounds on the growth of individually PGPMs were tested in starvation conditions. The different combination products based on microbial and non-microbial biostimulants (BS) appear worth considering for greenhouse and open field trials to select those potentially adoptable in sustainable agriculture.

The Role of Petrimonas mucosa ING2-E5AT in Mesophilic Biogas Reactor Systems as Deduced from Multiomics Analyses.

Members of the genera Proteiniphilum and Petrimonas were speculated to represent indicators reflecting process instability within anaerobic digestion (AD) microbiomes. Therefore, Petrimonas mucosa ING2-E5AT was isolated from a biogas reactor sample and sequenced on the PacBio RSII and Illumina MiSeq sequencers. Phylogenetic classification positioned the strain ING2-E5AT in close proximity to Fermentimonas and Proteiniphilum species (family Dysgonomonadaceae). ING2-E5AT encodes a number of genes for glycosyl-hydrolyses (GH) which are organized in Polysaccharide Utilization Loci (PUL) comprising tandem susCD-like genes for a TonB-dependent outer-membrane transporter and a cell surface glycan-binding protein. Different GHs encoded in PUL are involved in pectin degradation, reflecting a pronounced specialization of the ING2-E5AT PUL systems regarding the decomposition of this polysaccharide. Genes encoding enzymes participating in amino acids fermentation were also identified. Fragment recruitments with the ING2-E5AT genome as a template and publicly available metagenomes of AD microbiomes revealed that Petrimonas species are present in 146 out of 257 datasets supporting their importance in AD microbiomes. Metatranscriptome analyses of AD microbiomes uncovered active sugar and amino acid fermentation pathways for Petrimonas species. Likewise, screening of metaproteome datasets demonstrated expression of the Petrimonas PUL-specific component SusC providing further evidence that PUL play a central role for the lifestyle of Petrimonas species.

Analyzing large scale genomic data on the cloud with Sparkhit.

Motivation: The increasing amount of next-generation sequencing data poses a fundamental challenge on large scale genomic analytics. Existing tools use different distributed computational platforms to scale-out bioinformatics workloads. However, the scalability of these tools is not efficient. Moreover, they have heavy run time overheads when pre-processing large amounts of data. To address these limitations, we have developed Sparkhit: a distributed bioinformatics framework built on top of the Apache Spark platform. Results: Sparkhit integrates a variety of analytical methods. It is implemented in the Spark extended MapReduce model. It runs 92-157 times faster than MetaSpark on metagenomic fragment recruitment and 18-32 times faster than Crossbow on data pre-processing. We analyzed 100 terabytes of data across four genomic projects in the cloud in 21 h, which includes the run times of cluster deployment and data downloading. Furthermore, our application on the entire Human Microbiome Project shotgun sequencing data was completed in 2 h, presenting an approach to easily associate large amounts of public datasets with reference data. Availability and implementation: Sparkhit is freely available at: https://rhinempi.github.io/sparkhit/. Contact: asczyrba@cebitec.uni-bielefeld.de. Supplementary information: Supplementary data are available at Bioinformatics online.

Fast Physically Correct Refocusing for Sparse Light Fields Using Block-Based Multi-Rate View Interpolation.

Digital refocusing has a tradeoff between complexity and quality when using sparsely sampled light fields for low-storage applications. In this paper, we propose a fast physically correct refocusing algorithm to address this issue in a twofold way. First, view interpolation is adopted to provide photorealistic quality at infocus-defocus hybrid boundaries. Regarding its conventional high complexity, we devised a fast line-scan method specifically for refocusing, and its 1D kernel can be 30× faster than the benchmark View Synthesis Reference Software (VSRS)-1D-Fast. Second, we propose a block-based multi-rate processing flow for accelerating purely infocused or defocused regions, and a further 3- 34× speedup can be achieved for high-resolution images. All candidate blocks of variable sizes can interpolate different numbers of rendered views and perform refocusing in different subsampled layers. To avoid visible aliasing and block artifacts, we determine these parameters and the simulated aperture filter through a localized filter response analysis using defocus blur statistics. The final quadtree block partitions are then optimized in terms of computation time. Extensive experimental results are provided to show superior refocusing quality and fast computation speed. In particular, the run time is comparable with the conventional single-image blurring, which causes serious boundary artifacts.

Methamphetamine induces alterations in the long non-coding RNAs expression profile in the nucleus accumbens of the mouse.

BACKGROUND: Repeated exposure to addictive drugs elicits long-lasting cellular and molecular changes. It has been reported that the aberrant expression of long non-coding RNAs (lncRNAs) is involved in cocaine and heroin addiction, yet the expression profile of lncRNAs and their potential effects on methamphetamine (METH)-induced locomotor sensitization are largely unknown. RESULTS: Using high-throughput strand-specific complementary DNA sequencing technology (ssRNA-seq), here we examined the alterations in the lncRNAs expression profile in the nucleus accumbens (NAc) of METH-sensitized mice. We found that the expression levels of 6246 known lncRNAs (6215 down-regulated, 31 up-regulated) and 8442 novel lncRNA candidates (8408 down-regulated, 34 up-regulated) were significantly altered in the METH-sensitized mice. Based on characterizations of the genomic contexts of the lncRNAs, we further showed that there were 5139 differentially expressed lncRNAs acted via cis mechanisms, including sense intronic (4295 down-regulated and one up-regulated), overlapping (25 down-regulated and one up-regulated), natural antisense transcripts (NATs, 148 down-regulated and eight up-regulated), long intergenic non-coding RNAs (lincRNAs, 582 down-regulated and five up-regulated), and bidirectional (72 down-regulated and two up-regulated). Moreover, using the program RNAplex, we identified 3994 differentially expressed lncRNAs acted via trans mechanisms. Gene ontology (GO) and KEGG pathway enrichment analyses revealed that the predicted cis- and trans- associated genes were significantly enriched during neuronal development, neuronal plasticity, learning and memory, and reward and addiction. CONCLUSIONS: Taken together, our results suggest that METH can elicit global changes in lncRNA expressions in the NAc of sensitized mice that might be involved in METH-induced locomotor sensitization and addiction.

[Detection of Angiostrongylus cantonensis larvae with a high resolution melt PCR assay].

OBJECTIVE: To develop a detection method of the third-stage larvae of Angiostrongylus cantonensis by real-time PCR and high-resolution melt curve analysis. METHODS: A pair of specific primers was designed based on the internal transcribed spacer 1 (ITS1) region of the nuclear ribosomal DNA of A. cantonensis. The third-stage larvae of A. cantonensis were detected by real-time PCR and high-resolution melt curve analysis. The specificity of the method was analyzed by testing DNAs of A. cantonensis, Clonorchis sinensis, and Gnathostoma spinigerum. The genomic DNA were extracted from 1 to 10 third-stage larvae of A. cantonensis, respectively, and used to identify the sensitivity of the method. RESULTS: This method could specifically detect A. cantonensis and the detection limit reached to one larva. No amplification curve and melt curve were found in C. sinensis and G. spinigerum. CONCLUSION: Real-time PCR and high-resolution melt curve analysis show good specificity and sensitivity for detecting the third-stage larvae of A. cantonensis.