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Author: Willenbücher, Katharina × Type: Text ×

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    Impact of process temperature and organic loading rate on cellulolytic/hydrolytic biofilm microbiomes during biomethanation of ryegrass silage revealed by genome-centered metagenomics and metatranscriptomics
    (London : BioMed Central, 2020) Maus, Irena; Klocke, Michael; Derenkó, Jaqueline; Stolze, Yvonne; Beckstette, Michael; Jost, Carsten; Wibberg, Daniel; Blom, Jochen; Henke, Christian; Willenbücher, Katharina; Rumming, Madis; Rademacher, Antje; Pühler, Alfred; Sczyrba, Alexander; Schlüter, Andreas
    Background: Anaerobic digestion (AD) of protein-rich grass silage was performed in experimental two-stage two-phase biogas reactor systems at low vs. increased organic loading rates (OLRs) under mesophilic (37 °C) and thermophilic (55 °C) temperatures. To follow the adaptive response of the biomass-attached cellulolytic/hydrolytic biofilms at increasing ammonium/ammonia contents, genome-centered metagenomics and transcriptional profiling based on metagenome assembled genomes (MAGs) were conducted. Results: In total, 78 bacterial and archaeal MAGs representing the most abundant members of the communities, and featuring defined quality criteria were selected and characterized in detail. Determination of MAG abundances under the tested conditions by mapping of the obtained metagenome sequence reads to the MAGs revealed that MAG abundance profiles were mainly shaped by the temperature but also by the OLR. However, the OLR effect was more pronounced for the mesophilic systems as compared to the thermophilic ones. In contrast, metatranscriptome mapping to MAGs subsequently normalized to MAG abundances showed that under thermophilic conditions, MAGs respond to increased OLRs by shifting their transcriptional activities mainly without adjusting their proliferation rates. This is a clear difference compared to the behavior of the microbiome under mesophilic conditions. Here, the response to increased OLRs involved adjusting of proliferation rates and corresponding transcriptional activities. The analysis led to the identification of MAGs positively responding to increased OLRs. The most outstanding MAGs in this regard, obviously well adapted to higher OLRs and/or associated conditions, were assigned to the order Clostridiales (Acetivibrio sp.) for the mesophilic biofilm and the orders Bacteroidales (Prevotella sp. and an unknown species), Lachnospirales (Herbinix sp. and Kineothrix sp.) and Clostridiales (Clostridium sp.) for the thermophilic biofilm. Genome-based metabolic reconstruction and transcriptional profiling revealed that positively responding MAGs mainly are involved in hydrolysis of grass silage, acidogenesis and/or acetogenesis. Conclusions: An integrated-omics approach enabled the identification of new AD biofilm keystone species featuring outstanding performance under stress conditions such as increased OLRs. Genome-based knowledge on the metabolic potential and transcriptional activity of responsive microbiome members will contribute to the development of improved microbiological AD management strategies for biomethanation of renewable biomass. © 2020 The Author(s).
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    Degradation Kinetics of Lignocellulolytic Enzymes in a Biogas Reactor Using Quantitative Mass Spectrometry
    (Basel : MDPI, 2023) Küchler, Jan; Willenbücher, Katharina; Reiß, Elisabeth; Nuß, Lea; Conrady, Marius; Ramm, Patrice; Schimpf, Ulrike; Reichl, Udo; Szewzyk, Ulrich; Benndorf, Dirk
    The supplementation of lignocellulose-degrading enzymes can be used to enhance the performance of biogas production in industrial biogas plants. Since the structural stability of these enzyme preparations is essential for efficient application, reliable methods for the assessment of enzyme stability are crucial. Here, a mass-spectrometric-based assay was established to monitor the structural stability of enzymes, i.e., the structural integrity of these proteins, in anaerobic digestion (AD). The analysis of extracts of Lentinula edodes revealed the rapid degradation of lignocellulose-degrading enzymes, with an approximate half-life of 1.5 h. The observed low structural stability of lignocellulose-degrading enzymes in AD corresponded with previous results obtained for biogas content. The established workflow can be easily adapted for the monitoring of other enzyme formulations and provides a platform for evaluating the effects of enzyme additions in AD, together with a characterization of the biochemical methane potential used in order to determine the biodegradability of organic substrates.