Biology subjects

Gemeinhardt, K.

Publications and source records attributed to Gemeinhardt, K..

4 recordsLinked to original sources

bioRxiv2025

MMonitor: Software for Real-Time Monitoring of Microbial Communities Using Long Reads

Real-time monitoring of microbial communities offers valuable insights into microbial dynamics across diverse environments. However, many existing metagenome analysis tools require advanced computational expertise and are not designed for monitoring. We present MMonitor, an open source software platform for the real-time analysis and visualization of metagenomic Oxford Nanopore Technologies (ONT) sequencing data. MMonitor includes two components: a desktop application for running bioinformatics pipelines through a graphical user interface (GUI) or command-line interface (CLI), and a web-based dashboard for interactive result inspection. The dashboard provides taxonomic composition over time, quality scores, diversity indices, and taxonomy-metadata correlations. Integrated pipelines enable automated de-novo assembly and reconstruction of metagenome-assembled genomes (MAGs). To validate MMonitor, we tracked human gut microbial populations in three bioreactors using 16S rRNA gene sequencing, and applied it to whole-genome sequencing (WGS) data to generate high-quality annotated MAGs and reveal functional insights. We also compare MMonitor to other software for real-time metagenomic analysis, highlighting the strengths and limitations of each tool for this use case. By performing automated time-series analyzes, sample management, and updating of reference databases, MMonitor addresses current limitations and supports dynamic microbiome research in various fields. IMPORTANCEMetagenome monitoring is essential for understanding and managing microbial communities in dynamic environments. Rapid and accurate analysis of these communities allows for timely interventions in biologically active environments, for example, in industrial biotechnology, environmental surveillance, and medical diagnostics. Metagenome monitoring tools should have real-time capabilities, intuitive interfaces, convenient setup and distribution, and suitable visual representations for effective monitoring. MMonitor addresses these challenges by offering a platform that combines real-time analysis of nanopore sequencing data with customizable pipelines and advanced visualization tools. It is designed to be accessible to researchers of all levels of computational skill. It supports efficient taxonomic and functional analyses based on popular metagenome software and automates taxonomic profiling, genome assembly, binning, and annotation. The dashboard helps to interpret data and present results. MMonitor is designed to track metagenomes and we hope that it can impact microbiome research and applications in health, biotechnology, agricultural, and environmental sciences.

bioinformatics↗

bioRxiv2025

Genetic modification of Clostridium kluyveri for heterologous n-butanol and n-hexanol production

The mesophilic microbe Clostridium kluyveri serves as the most commonly used model microbe to elucidate the physiology and biochemistry of ethanol-based chain elongation via reverse {beta}-oxidation. In this pathway, ethanol and acetate are converted into short- and medium-chain carboxylates. However, to date, no genetic system has been published in a peer-reviewed publication. Here, we report the development of versatile genetic tools for C. kluyveri, utilizing the pMTL Clostridia shuttle vector system and thiamphenicol as a selective marker. We identified the native restriction-modification system of C. kluyveri as a critical barrier to DNA transfer and overcame it by identifying and characterizing the crucial methyltransferase. To mimic the native DNA methylation pattern of C. kluyveri, we performed in-vivo methylation of the shuttle vector plasmid by expressing the methyltransferase in Escherichia coli, followed by DNA transfer via conjugation. After validating the genetic system, we demonstrated heterologous expression of different combinations of both NADH and NADPH-dependent alcohol dehydrogenases from Clostridium acetobutylicum. The expression of these genes was controlled by the Pthl promoter, which is commonly used in Clostridia, and the PadhE2 promoter, leading to n-butanol and n-hexanol production of the mutant strains. This genetic system for C. kluyveri will not only enable further research on the metabolism of this microbe but also enable more profound insights into ethanol-based chain elongation in general. IMPORTANCEMedium-chain carboxylates are required in various everyday products, including cosmetics, pharmaceuticals, and fragrances, and show a natural antimicrobial property. Further, they represent food additives and serve as chemical building blocks for several other compounds. Traditionally, these carboxylates are produced from fossil resources, contributing to increased greenhouse gas emissions. Alternatively, they are derived from animal- or plant-based fat (e.g., coconut oil), which competes with agricultural land that is needed for food production. However, microbial chain elongation, which is a biotechnological approach relying on microbes, such as Clostridium kluyveri, is sustainable and a promising alternative to the conventional production of medium-chain carboxylates. Notably, it enables the use of industrial waste streams (e.g., off-gases, carbohydrate-rich industrial waste) as substrates, making the process more environmentally friendly. By applying our genetic system for C. kluyveri, a better understanding of microbial chain elongation can be achieved, and potentially even an extension of its product portfolio.

microbiology↗

bioRxiv2024

Toward industrial C8 production: Oxygen intrusion drives renewable n caprylate production from ethanol and acetate via intermediate metabolite production

Previous bioreactor studies achieved high volumetric n-caprylate (i.e., n-octanoate) production rates and selectivities from ethanol and acetate with chain-elongating microbiomes. However, the metabolic pathways from the substrates to n-caprylate synthesis were unclear. We operated two n-caprylate-producing upflow bioreactors with a synthetic medium to study the underlying metabolic pathways. The operating period exceeded 2.5 years, with a peak volumetric n-caprylate production rate of 190 {+/-} 8.4 mmol C L-1 d-1 (0.14 g L-1 h-1). We identified oxygen availability as a critical performance parameter, facilitating intermediate metabolite production from ethanol. Bottle experiments in the presence and absence of oxygen with 13C-labeled ethanol suggest acetyl-coenzyme A-based derived production of n-butyrate (i.e., n-butanoate), n-caproate (i.e., n-hexanoate), and n-caprylate. Here, we postulate a trophic hierarchy within the bioreactor microbiomes based on metagenomics, metaproteomics, and metabolomics data, as well as experiments with a Clostridium kluyveri isolate. First, the aerobic bacterium Pseudoclavibacter caeni and the facultative anaerobic fungus Cyberlindnera jadinii converted part of the ethanol pool into the intermediate metabolites succinate, lactate, and pyroglutamate. Second, the strict anaerobic C. kluyveri elongated acetate with the residual ethanol to n-butyrate. Third, Caproicibacter fermentans and Oscillibacter valericigenes elongated n-butyrate with the intermediate metabolites to n-caproate and then to n-caprylate. Among the carbon chain-elongating pathways of carboxylates, the tricarboxylic acid cycle and the reverse {beta}-oxidation pathways showed a positive correlation with n-caprylate production. The results of this study inspire the realization of a chain-elongating production platform with separately controlled aerobic and anaerobic stages to produce n-caprylate renewably as an attractive chemical from ethanol and acetate as substrates. Broader contextNext to renewable electric energy, carbon-based chemicals have to be produced sustainably and independently from fossil sources. To meet this goal, we must expand the portfolio of bio-based conversion technologies on an industrial scale to cover as many target chemicals as possible. We explore the bioprocess of chain elongation to provide medium-chain carboxylates that can function as future platform chemicals in the circular economy. The most valuable medium-chain carboxylate produced with chain elongation is n-caprylate (i.e., n-octanoate). This molecule with eight carbon atoms in a row (C8) is challenging to produce renewably for the chemical industry. Previous reports elucidated that elevated ethanol-to-acetate ratios, which are found in syngas-fermentation effluent, stimulated n-caprylate production. Until now, studies have suggested that chain elongation from high concentrations of ethanol and acetate is a fully anaerobic process. We refine this view by showing a trophic hierarchy of aerobic and anaerobic microbes capable of facilitating this process. Appropriate oxygen supplementation enables the synthesis of succinate, lactate, and pyroglutamate that permit high-rate chain elongation to n-caprylate under anaerobic conditions. Given these results, future research should focus on the segregated study of aerobic and anaerobic microbes to further enhance the process performance to produce n-caprylate renewably at an industrial scale.

bioengineering↗

bioRxiv2024

Performance effects from different shutdown methods of three electrode materials for the power-to-gas application with electromethanogenesis

Industrial applications of microbial electrochemical systems will require regular maintenance shutdowns, involving inspections and component replacements to extend the lifespan of the system. Here, we examined the impact of such shutdowns on the performance of three electrode materials (i.e., platinized titanium, graphite, and nickel) as cathodes in a microbial electrochemical system that would be used for electromethanogenesis in power-to-gas applications. We focused on methane (CH4) production from hydrogen (H2) and carbon dioxide (CO2) using Methanothermobacter thermautotrophicus. We showed that the platinized titanium cathode resulted in high volumetric CH4 production rates and Coulombic efficiencies. Using a graphite cathode would be more cost-effective than using the platinized titanium cathode in microbial electrochemical systems but showed an inferior performance. The microbial electrochemical system with the nickel cathode showed improvements compared to the graphite cathode. Additionally, this system with a nickel cathode demonstrated the fastest recovery during a shutdown experiment compared to the other two cathodes. Fluctuations in pH and nickel concentrations in the catholyte during power interruptions affected CH4 production recovery in the system with the nickel cathode. This research enhances understanding of the integration of biological and electrochemical processes in microbial electrochemical systems, providing insights into electrode selection and operating strategies for effective and sustainable CH4 production.

bioengineering↗