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The Archaeorhizomycetes are recently discovered fungi with poorly resolved ecology. Even their abundance in soil fungal communities is currently disputed. Here we applied a PCR-independent, RNA-based metatranscriptomic approach to determine their abundance among fungi in eleven different soils across Europe. Using small subunit (SSU) ribosomal RNA transcripts as marker, we detected Archaeorhizomycetes in 17 out of 28 soil metatranscriptomes. They had average relative SSU rRNA abundance of 2.0% with a maximum of 9.4% among fungal SSU rRNAs. Network analysis revealed that they co-occur with arbuscular mycorrhizal Glomerales, which is in line with their previously suggested association with plant roots. Moreover, Archaeorhizomycetes ranked among the potential keystone taxa. This metatranscriptomic survey exemplifies the usage of non-targeted molecular approaches for the study of soil fungi. It provides PCR- and DNA-independent evidence for the low abundance of Archaeorhizomycetes in soil fungal communities, although they might be non-negligible players despite their low abundance.
Závěrečná zpráva o řešení grantu Agentury pro zdravotnický výzkum MZ ČR
nestr.
Along with individual's genetic predisposition, host microbiome is an important factor in etiopathogenesis of gastroesophageal reflux disease (GERD), increasing significantly risk of developing esophagitis (RE), Barrett's esophagus (BE), and esophageal adenocarcinoma (EAC). The project aim is to determine markers at the level of microbiome, host genes and their interactions for earlier diagnosis/prediction of BE and EAC The project includes analyses of (1) metagenome and metatranscriptome at several sites of gastrointestinal tract (GIT) in GERD patients, (2) host exome and transcriptome in esophageal tissue in patients with BE/EAC, and (3) genetic profile of GERD patients. Mutations in patient ́s exome/genome and gene expression levels in esophagus tissue will be correlated with microbiome and its functional activity in the GIT and clinical finding with in vivo endomicroscopy. Based on the obtained data, we will propose modification of the standard care regime for GERD patients to avoid progressive/irreversible changes reducing patients ́ life quality or even threating their lives.
Hostitelský mikrobiom spolu s genetickou predispozicí jedince jsou důležitými faktory v etiopatogenezi refluxní choroby jícnu (GERD), která významně zvyšuje riziko rozvoje ezofagitidy (RE), Barrettova jícnu (BE) a adenokarcinomu ezofagu (EAC). Cílem projektu je determinace markerů na úrovni mikrobiomu, genů hostitele a jejich interakcí pro časnější diagnostiku a predikci rozvoje BE a EAC. Projekt obsahuje 3 části: (1) analýzu metagenomu a metatranskriptomu v několika lokalitách gastrointestinálního traktu (GIT) u pacientů s GERD, (2) analýzu hostitelského exomu a transkriptomu ve tkáni jícnu u pacientů s BE/EAC, a (3) analýzu genetického profilu pacientů s GERD a jeho komplikacemi. Mutace v exomu/genomu pacienta a míra exprese genů ve tkáni jícnu budou korelovány s mikrobiomem a jeho funkční aktivitou v GITu a klinickým nálezem doplněným o in vivo endomikroskopii. Na základě získaných dat se pokusíme navrhnout modifikaci standardního režimu péče o pacienty s GERD s cílem předejít pokročilým a ireverzibilním změnám snižujícím kvalitu života pacientů, resp. ohrožujících je na životě.
- Klíčová slova
- sekvenování nové generace, mikrobiom, microbiome, Genetická predispozice, Genetic predisposition, In vivo diagnostika, refluxní choroba jícnu, Barrettův jícen, adenokarcinom jícnu, in vitro diagnostika, Barrett's esophagus, in vitro diagnostics, Next-generation sequencing, Esophageal adenocarcinoma, in vivo diagnostics, metatranskriptom, metatranscriptome, esophageal reflux disease,
- NLK Publikační typ
- závěrečné zprávy o řešení grantu AZV MZ ČR
Cimex lectularius, known as the common bed bug, is a widespread hematophagous human ectoparasite and urban pest that is not known to be a vector of any human infectious disease agents. However, few studies in the era of molecular biology have profiled the microorganisms harbored by field populations of bed bugs. The objective of this study was to examine the viruses present in a large sampling of common bed bugs and related bat bugs (Cimex pipistrelle). RNA sequencing was undertaken on an international sampling of > 500 field-collected bugs, and multiple workflows were used to assemble contigs and query these against reference nucleotide databases to identify viral genomes. Shuangao bed bug virus 2, an uncharacterized rhabdovirus previously discovered in Cimex hemipterus from China, was found in several bed bug pools from the USA and Europe, as well as in C. pipistrelle, suggesting that this virus is common among bed bug populations. In addition, Shuangao bed bug virus 1 was detected in a bed bug pool from China, and sequences matching Enterobacteria phage P7 were found in all bed bug pools, indicating the ubiquitous presence of phage-derived elements in the genome of the bed bug or its enterobacterial symbiont. However, viral diversity was low in bed bugs in our study, as no other viral genomes were detected with significant coverage. These results provide evidence against frequent virus infection in bed bugs. Nonetheless, our investigation had several important limitations, and additional studies should be conducted to better understand the prevalence and composition of viruses in bed bugs. Most notably, our study largely focused on insects from urban areas in industrialized nations, thus likely missing infrequent virus infections and those that could occur in rural or tropical environments or developing nations.
Within streptophyte green algae Zygnematophyceae are the sister group to the land plants that inherited several traits conferring stress protection. Zygnema sp., a mat-forming alga thriving in extreme habitats, was collected from a field site in Svalbard, where the bottom layers are protected by the top layers. The two layers were investigated by a metatranscriptomic approach and GC-MS-based metabolite profiling. In the top layer, 6569 genes were significantly upregulated and 149 were downregulated. Upregulated genes coded for components of the photosynthetic apparatus, chlorophyll synthesis, early light-inducible proteins, cell wall and carbohydrate metabolism, including starch-degrading enzymes. An increase in maltose in the top layer and degraded starch grains at the ultrastructural levels corroborated these findings. Genes involved in amino acid, redox metabolism and DNA repair were upregulated. A total of 29 differentially accumulated metabolites (out of 173 identified ones) confirmed higher metabolic turnover in the top layer. For several of these metabolites, differential accumulation matched the transcriptional changes of enzymes involved in associated pathways. In summary, the findings support the hypothesis that in a Zygnema mat the top layer shields the bottom layers from abiotic stress factors such as excessive irradiation.
- MeSH
- Chlorophyta genetika metabolismus MeSH
- ekosystém MeSH
- fotosyntéza genetika MeSH
- fyziologický stres MeSH
- metabolom MeSH
- Streptophyta genetika metabolismus MeSH
- transkriptom MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Arktida MeSH
- Svalbard MeSH
BACKGROUND: The planetary sulfur cycle is a complex web of chemical reactions that can be microbial-mediated or can occur spontaneously in the environment, depending on the temperature and pH. Inorganic sulfur compounds can serve as energy sources for specialized prokaryotes and are important substrates for microbial growth in general. Here, we investigate dissimilatory sulfur cycling in the brine and sediments of a southwestern Siberian soda lake characterized by an extremely high pH and salinity, combining meta-omics analyses of its uniquely adapted highly diverse prokaryote communities with biogeochemical profiling to identify key microbial players and expand our understanding of sulfur cycling under haloalkaline conditions. RESULTS: Peak microbial activity was found in the top 4 cm of the sediments, a layer with a steep drop in oxygen concentration and redox potential. The majority of sulfur was present as sulfate or iron sulfide. Thiosulfate was readily oxidized by microbes in the presence of oxygen, but oxidation was partially inhibited by light. We obtained 1032 metagenome-assembled genomes, including novel population genomes of characterized colorless sulfur-oxidizing bacteria (SOB), anoxygenic purple sulfur bacteria, heterotrophic SOB, and highly active lithoautotrophic sulfate reducers. Surprisingly, we discovered the potential for nitrogen fixation in a new genus of colorless SOB, carbon fixation in a new species of phototrophic Gemmatimonadetes, and elemental sulfur/sulfite reduction in the "Candidatus Woesearchaeota." Polysulfide/thiosulfate and tetrathionate reductases were actively transcribed by various (facultative) anaerobes. CONCLUSIONS: The recovery of over 200 genomes that encoded enzymes capable of catalyzing key reactions in the inorganic sulfur cycle indicates complete cycling between sulfate and sulfide at moderately hypersaline and extreme alkaline conditions. Our results suggest that more taxonomic groups are involved in sulfur dissimilation than previously assumed.
- MeSH
- Archaea klasifikace genetika metabolismus MeSH
- Bacteria klasifikace genetika metabolismus MeSH
- fylogeneze MeSH
- jezera chemie mikrobiologie MeSH
- koncentrace vodíkových iontů MeSH
- metagenom MeSH
- oxidace-redukce MeSH
- salinita MeSH
- síra analýza metabolismus MeSH
- soli chemie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Sibiř MeSH
Lignocellulose from plant biomass represents an abundant and renewable source for the production of environmentally friendly chemicals and biofuels. However, its recalcitrant nature entails the use of complex biochemical reactions that are still challenging. Since the degradation of lignocellulose is the current bottleneck of the conversion processes, the search for novel enzymes and microbial strains for degrading plant biomass is of high importance. Currently, bacteria are in the spotlight as promising candidates for novel conversion strategies due to their wide functional diversity and versatility. Here, we review the lines of evidence that show the high potential of bacterial strains from soil for biomass conversion ranging from strain characterization to metagenome and metatranscriptome analysis. Substantial and diverse fractions of soil bacteria are able to decompose the major lignocellulose components. To do that, bacteria evolved structurally variable and often highly complex lignocellulolytic systems composed of enzymes as well as proteins involved in efficient substrate binding. Both as individual components or in combination, bacterial enzymes, and accessory proteins appear to be promising tools in the biotechnological valorization of lignocellulose.
Soils of coniferous forest ecosystems are important for the global carbon cycle, and the identification of active microbial decomposers is essential for understanding organic matter transformation in these ecosystems. By the independent analysis of DNA and RNA, whole communities of bacteria and fungi and its active members were compared in topsoil of a Picea abies forest during a period of organic matter decomposition. Fungi quantitatively dominate the microbial community in the litter horizon, while the organic horizon shows comparable amount of fungal and bacterial biomasses. Active microbial populations obtained by RNA analysis exhibit similar diversity as DNA-derived populations, but significantly differ in the composition of microbial taxa. Several highly active taxa, especially fungal ones, show low abundance or even absence in the DNA pool. Bacteria and especially fungi are often distinctly associated with a particular soil horizon. Fungal communities are less even than bacterial ones and show higher relative abundances of dominant species. While dominant bacterial species are distributed across the studied ecosystem, distribution of dominant fungi is often spatially restricted as they are only recovered at some locations. The sequences of cbhI gene encoding for cellobiohydrolase (exocellulase), an essential enzyme for cellulose decomposition, were compared in soil metagenome and metatranscriptome and assigned to their producers. Litter horizon exhibits higher diversity and higher proportion of expressed sequences than organic horizon. Cellulose decomposition is mediated by highly diverse fungal populations largely distinct between soil horizons. The results indicate that low-abundance species make an important contribution to decomposition processes in soils.
- MeSH
- Bacteria klasifikace enzymologie genetika MeSH
- biodiverzita MeSH
- celulosa-1,4-beta-cellobiosidasa genetika MeSH
- celulosa metabolismus MeSH
- ekosystém MeSH
- fylogeneze MeSH
- houby klasifikace enzymologie genetika MeSH
- metagenom MeSH
- půdní mikrobiologie MeSH
- RNA ribozomální 16S genetika MeSH
- smrk fyziologie MeSH
- stromy mikrobiologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Stable isotope probing (SIP) provides the opportunity to label decomposer microorganisms that build their biomass on a specific substrate. In combination with high-throughput sequencing, SIP allows for the identification of fungal community members involved in a particular decomposition process. Further information can be gained through gene-targeted metagenomics and metatranscriptomics, opening the possibility to describe the pool of genes catalyzing specific decomposition reactions in situ and to identify the diversity of genes that are expressed. When combined with gene descriptions of fungal isolates from the same environment, specific biochemical reactions involved in decomposition can be linked to individual fungal taxa. Here we describe the use of these methods to explore the cellulolytic fungal community in forest litter and soil.
Molecular methods for the analysis of biomolecules have undergone rapid technological development in the last decade. The advent of next-generation sequencing methods and improvements in instrumental resolution enabled the analysis of complex transcriptome, proteome and metabolome data, as well as a detailed annotation of microbial genomes. The mechanisms of decomposition by model fungi have been described in unprecedented detail by the combination of genome sequencing, transcriptomics and proteomics. The increasing number of available genomes for fungi and bacteria shows that the genetic potential for decomposition of organic matter is widespread among taxonomically diverse microbial taxa, while expression studies document the importance of the regulation of expression in decomposition efficiency. Importantly, high-throughput methods of nucleic acid analysis used for the analysis of metagenomes and metatranscriptomes indicate the high diversity of decomposer communities in natural habitats and their taxonomic composition. Today, the metaproteomics of natural habitats is of interest. In combination with advanced analytical techniques to explore the products of decomposition and the accumulation of information on the genomes of environmentally relevant microorganisms, advanced methods in microbial ecophysiology should increase our understanding of the complex processes of organic matter transformation.
16S rRNA amplicon sequencing or, more recently, metatranscriptomic analysis are currently the only preferred methods for microbial profiling of samples containing a predominant ratio of human to bacterial DNA. However, due to the off-target amplification of human DNA, current protocols are inadequate for bioptic samples. Here we present an efficient, reliable, and affordable method for the bacteriome analysis of clinical samples human DNA content predominates. We determined the microbiota profile in a total of 40 human biopsies of the esophagus, stomach, and duodenum using 16S rRNA amplicon sequencing with the widely used 515F-806R (V4) primers targeting the V4 region, 68F-338R primers and a modified set of 68F-338R (V1-V2M) primers targeting the V1-V2 region. With the V4 primers, on average 70% of amplicon sequence variants (ASV) mapped to the human genome. On the other hand, this off-target amplification was absent when using the V1-V2M primers. Moreover, the V1-V2M primers provided significantly higher taxonomic richness and reproducibility of analysis compared to the V4 primers. We conclude that the V1-V2M 16S rRNA sequencing method is reliable, cost-effective, and applicable for low-bacterial abundant human samples in medical research.
- MeSH
- biopsie MeSH
- gastrointestinální trakt MeSH
- geny rRNA MeSH
- lidé MeSH
- mikrobiota * genetika MeSH
- reprodukovatelnost výsledků MeSH
- RNA ribozomální 16S genetika MeSH
- sekvenční analýza DNA metody MeSH
- vysoce účinné nukleotidové sekvenování metody MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
