Filters

2012 - 201918

More filters

2019 - 2019112020 - 20227
Reset filters

Settings

search.filters.applied.f.access: openAccess × Author: Klocke, Michael × End date: 2019 × Start date: 2011 × WGL Institute: ATB ×

Search Results

Now showing 1 - 10 of 18
  • Thumbnail Image
    Item
    DNA and RNA extraction and quantitative real-time PCR-based assays for biogas biocenoses in an interlaboratory comparison
    (Basel : MDPI, 2016) Lebuhn, Michael; Derenkó, Jaqueline; Rademacher, Antje; Helbig, Susanne; Munk, Bernhard; Pechtl, Alexander; Stolze, Yvonne; Prowe, Steffen; Schwarz, Wolfgang H.; Schlüter, Andreas; Liebl, Wolfgang; Klocke, Michael
    Five institutional partners participated in an interlaboratory comparison of nucleic acid extraction, RNA preservation and quantitative Real-Time PCR (qPCR) based assays for biogas biocenoses derived from different grass silage digesting laboratory and pilot scale fermenters. A kit format DNA extraction system based on physical and chemical lysis with excellent extraction efficiency yielded highly reproducible results among the partners and clearly outperformed a traditional CTAB/chloroform/isoamylalcohol based method. Analytical purpose, sample texture, consistency and upstream pretreatment steps determine the modifications that should be applied to achieve maximum efficiency in the trade-off between extract purity and nucleic acid recovery rate. RNA extraction was much more variable, and the destination of the extract determines the method to be used. RNA stabilization with quaternary ammonium salts was an as satisfactory approach as flash freezing in liquid N2. Due to co-eluted impurities, spectrophotometry proved to be of limited value for nucleic acid qualification and quantification in extracts obtained with the kit, and picoGreen® based quantification was more trustworthy. Absorbance at 230 nm can be extremely high in the presence of certain chaotropic guanidine salts, but guanidinium isothiocyanate does not affect (q)PCR. Absolute quantification by qPCR requires application of a reliable internal standard for which correct PCR efficiency and Y-intercept values are important and must be reported.
  • Thumbnail Image
    Item
    Development of a flow-fluorescence in situhybridization protocol for the analysis of microbial communities in anaerobic fermentation liquor
    (London : BioMed Central, 2013) Nettmann, Edith; Fröhling, Antje; Heeg, Kathrin; Klocke, Michael; Schlüter, Oliver; Mumme, Jan
    Background: The production of bio-methane from renewable raw material is of high interest because of the increasing scarcity of fossil fuels. The process of biomethanation is based on the inter- and intraspecific metabolic activity of a highly diverse and dynamic microbial community. The community structure of the microbial biocenosis varies between different biogas reactors and the knowledge about these microbial communities is still fragmentary. However, up to now no approaches are available allowing a fast and reliable access to the microbial community structure. Hence, the aim of this study was to originate a Flow-FISH protocol, namely a combination of flow cytometry and fluorescence in situ hybridization, for the analysis of the metabolically active microorganisms in biogas reactor samples. With respect to the heterogenic texture of biogas reactor samples and to collect all cells including those of cell aggregates and biofilms the development of a preceding purification procedure was indispensable. Results: Six different purification procedures with in total 29 modifications were tested. The optimized purification procedure combines the use of the detergent sodium hexametaphosphate with ultrasonic treatment and a final filtration step. By this treatment, the detachment of microbial cells from particles as well as the disbandment of cell aggregates was obtained at minimized cell loss. A Flow-FISH protocol was developed avoiding dehydration and minimizing centrifugation steps. In the exemplary application of this protocol on pure cultures as well as biogas reactor samples high hybridization rates were achieved for commonly established domain specific oligonucleotide probes enabling the specific detection of metabolically active bacteria and archaea. Cross hybridization and autofluorescence effects could be excluded by the use of a nonsense probe and negative controls, respectively. Conclusions: The approach described in this study enables for the first time the analysis of the metabolically active fraction of the microbial communities within biogas reactors by Flow-FISH.
  • Thumbnail Image
    Item
    Dynamic variation of the microbial community structure during the long-time mono-fermentation of maize and sugar beet silage
    (Milton Park : Taylor & Francis, 2015) Klang, Johanna; Theuerl, Susanne; Szewzyk, Ulrich; Huth, Markus; Tölle, Rainer; Klocke, Michael
    This study investigated the development of the microbial community during a long-term (337 days) anaerobic digestion of maize and sugar beet silage, two feedstocks that significantly differ in their chemical composition. For the characterization of the microbial dynamics, the community profiling method terminal restriction fragment length polymorphism (TRFLP) in combination with a cloning-sequencing approach was applied. Our results revealed a specific adaptation of the microbial community to the supplied feedstocks. Based on the high amount of complex compounds, the anaerobic conversion rate of maize silage was slightly lower compared with the sugar beet silage. It was demonstrated that members from the phylum Bacteroidetes are mainly involved in the degradation of low molecular weight substances such as sugar, ethanol and acetate, the main compounds of the sugar beet silage. It was further shown that species of the genus Methanosaeta are highly sensitive against sudden stress situations such as a strong decrease in the ammonium nitrogen (NH4 +-N) concentration or a drop of the pH value. In both cases, a functional compensation by members of the genera Methanoculleus and/or Methanosarcina was detected. However, the overall biomass conversion of both feedstocks proceeded efficiently as a steady state between acid production and consumption was recorded, which further resulted in an equal biogas yield.
  • Thumbnail Image
    Item
    Metagenome, metatranscriptome, and metaproteome approaches unraveled compositions and functional relationships of microbial communities residing in biogas plants
    (Berlin ; Heidelberg ; New York : Springer, 2018-4-30) Hassa, Julia; Maus, Irena; Off, Sandra; Pühler, Alfred; Scherer, Paul; Klocke, Michael; Schlüter, Andreas
    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. © 2018, The Author(s).
  • Thumbnail Image
    Item
    Complete Genome Sequence of a New Ruminococcaceae Bacterium Isolated from Anaerobic Biomass Hydrolysis
    (Washington, DC : American Soc. for Microbiology, 2018) Hahnke, Sarah; Abendroth, Christian; Langer, Thomas; Codoñer, Francisco M.; Ramm, Patrice; Porcar, Manuel; Luschnig, Olaf; Klocke, Michael
    A new Ruminococcaceae bacterium, strain HV4-5-B5C, participating in the anaerobic digestion of grass, was isolated from a mesophilic two-stage laboratoryscale leach bed biogas system. The draft annotated genome sequence presented in this study and 16S rRNA gene sequence analysis indicated the affiliation of HV4-5- B5C with the family Ruminococcaceae outside recently described genera. © 2018 Hahnke et al.
  • Thumbnail Image
    Item
    Impact of surface structure and feed gas composition on Bacillus subtilis endospore inactivation during direct plasma treatment
    (Lausanne : Frontiers Media, 2015) Hertwig, Christian; Steins, Veronika; Reineke, Kai; Rademacher, Antje; Klocke, Michael; Rauh, Cornelia; Schlüter, Oliver
    This study investigated the inactivation efficiency of cold atmospheric pressure plasma treatment on Bacillus subtilis endospores dependent on the used feed gas composition and on the surface, the endospores were attached on. Glass petri-dishes, glass beads, and peppercorns were inoculated with the same endospore density and treated with a radio frequency plasma jet. Generated reactive species were detected using optical emission spectroscopy. A quantitative polymerase chain reaction (qPCR) based ratio detection system was established to monitor the DNA damage during the plasma treatment. Argon + 0.135% vol. oxygen + 0.2% vol. nitrogen as feed gas emitted the highest amounts of UV-C photons and considerable amount of reactive oxygen and nitrogen species. Plasma generated with argon + 0.135% vol. oxygen was characterized by the highest emission of reactive oxygen species (ROS), whereas the UV-C emission was negligible. The use of pure argon showed a negligible emission of UV photons and atomic oxygen, however, the emission of vacuum (V)UV photons was assumed. Similar maximum inactivation results were achieved for the three feed gas compositions. The surface structure had a significant impact on the inactivation efficiency of the plasma treatment. The maximum inactivation achieved was between 2.4 and 2.8 log10 on glass petri-dishes and 3.9 to 4.6 log10 on glass beads. The treatment of peppercorns resulted in an inactivation lower than 1.0 log10. qPCR results showed a significant DNA damage for all gas compositions. Pure argon showed the highest results for the DNA damage ratio values, followed by argon + 0.135% vol. oxygen + 0.2% vol. nitrogen. In case of argon + 0.135% vol. oxygen the inactivation seems to be dominated by the action of ROS. These findings indicate the significant role of VUV and UV photons in the inactivation process of B. subtilis endospores.
  • Thumbnail Image
    Item
    Complete Genome Sequence of a New Firmicutes Species Isolated from Anaerobic Biomass Hydrolysis
    (Washington, DC : American Soc. for Microbiology, 2017) Abendroth, Christian; Hahnke, Sarah; Codoñer, Francisco M.; Klocke, Michael; Luschnig, Olaf; Porcar, Manuel
    A new Firmicutes isolate, strain HV4-6-A5C, was obtained from the hydrolysis stage of a mesophilic and anaerobic two-stage lab-scale leach-bed system for biomethanation of fresh grass. It is assumed that the bacterial isolate contributes to plant biomass degradation. Here, we report a draft annotated genome sequence of this organism. © 2017 Abendroth et al.
  • Thumbnail Image
    Item
    Prediction of the biogas production using GA and ACO input features selection method for ANN model
    (Amsterdam [u.a.] : Elsevier, 2019) Beltramo, Tanja; Klocke, Michael; Hitzmann, Bernd
    This paper presents a fast and reliable approach to analyze the biogas production process with respect to the biogas production rate. The experimental data used for the developed models included 15 process variables measured at an agricultural biogas plant in Germany. In this context, the concentration of volatile fatty acids, total solids, volatile solids acid detergent fibre, acid detergent lignin, neutral detergent fibre, ammonium nitrogen, hydraulic retention time, and organic loading rate were used. Artificial neural networks (ANN) were established to predict the biogas production rate. An ant colony optimization and genetic algorithms were implemented to perform the variable selection. They identified the significant process variables, reduced the model dimension and improved the prediction capacity of the ANN models. The best prediction of the biogas production rate was obtained with an error of prediction of 6.24% and a coefficient of determination of R2 = 0.9.
  • Thumbnail Image
    Item
  • Thumbnail Image
    Item