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Hölzer, M.

Publications and source records attributed to Hölzer, M..

At least 19 recordsLinked to original sources
bioRxiv2026

Leveraging a hybrid cross-disciplinary training model to accelerate global bioinformatics capacity

Disparities in formal bioinformatics training exacerbate the global skills gap, impeding the democratized application of advanced genomic technologies. To bridge this divide, we introduce a scalable, hybrid training framework designed to rapidly accelerate regional bioinformatics capacity. We exemplify this approach through the Eastern European Bioinformatics and Genomics (EEBG) workshop series -- a cross-disciplinary initiative that pairs international faculty with local institutions to deliver modular, hands-on curricula. Functioning as a structured knowledge-transfer pipeline, the series has catalyzed a sustainable educational ecosystem, evidenced by the establishment of multiple independent summer schools across the region. The assessment of the 2025 EEBG workshop in Krakow, Poland, validates the models viability; participant metrics confirm high efficacy in skill acquisition (mean satisfaction: 4.4/5.0) and community building. Crucially, the hybrid delivery mode dismantled geographic barriers, serving as a vital mechanism for maintaining scientific continuity for researchers facing displacement and crisis. Synthesizing these outcomes, we define the core features of a replicable blueprint for scientific readiness in resource-constrained environments. We conclude by presenting a strategic roadmap -- organized around infrastructure standardization, governance sustainability, and geographical expansion -- for adapting this regional proof-of-concept into a global export-ready model, offering a critical path toward ensuring universal access to genomic innovation.

scientific communication and education↗
bioRxiv2025

Decoding bacterial methylomes in four public health-relevant microbial species: Nanopore sequencing enables reproducible analysis of DNA modifications

Investigating bacterial methylation profiles provides essential complementary information to the native DNA sequence, significantly extending our understanding of how DNA modifications influence virulence, antibiotic resistance, and the ability of bacteria to evade the immune system. Recent advancements in real-time Nanopore sequencing and basecalling algorithms have enabled the direct detection of modified bases from raw signal data, eliminating the need for bisulfite treatment of DNA. However, decoding methylation signals remains challenging due to rapid technological and methodological progress. In this study, we focus on public health-relevant bacterial strains to analyze their methylation profiles and identify methylation motifs. Our dataset includes samples from Staphylococcus aureus, Listeria monocytogenes, Enterococcus faecium, and Klebsiella pneumoniae, sequenced on the Nanopore GridION platform using the latest flow cell chemistry (R10.4.1) and modification basecalling models (Dorado basecalling SUP model v5). We investigated distinct methylation patterns within and between species, focusing on heavily modified genes or genomic regions. Our results reveal distinct species-specific methylation profiles, with each strain exhibiting unique modification patterns. We developed a modular pipeline using Nextflow and the Nanopore Modkit tool to streamline the detection of methylated motifs. We compared the results with outputs from MicrobeMod, a recent toolkit for exploring prokaryotic methylation and base modifications in nanopore sequencing. Our pipeline is publicly available for further use (github.com/rki-mf1/ont-methylation). We identified known methylation motifs already described in the literature and novel de novo motifs, providing deeper insights into the diversity of bacterial DNA modifications. Furthermore, we identified genomic regions that are extensively methylated, which could have implications for bacterial behavior and pathogenicity. We also assess improvements in basecalling accuracy, specifically how methylated bases can influence neighboring basecalls. Recent advances in basecalling models, particularly v5 models as part of Dorado, have reduced these issues, improving the reliability of methylation detection in bacterial genomes. In conclusion, our study highlights the potential of current nanopore sequencing tools for detecting DNA modifications in prokaryotes. By making our pipeline and results publicly available, we facilitate further research into bacterial DNA modifications and their role in microbial pathogenesis.

bioinformatics↗
bioRxiv2023

Targeted decontamination of sequencing data with CLEAN

Many biological and medical questions are answered based on the analysis of sequence data. However, we can find contamination, artificial spike-ins, and overrepresented rRNA sequences in various read collections and assemblies. In particular, spike-ins used as controls, as those known from Illumina or Nanopore data, are often not considered as contaminants and also not appropriately removed during analyses. Additionally, removing human host DNA may be necessary for data protection and ethical considerations to ensure that individuals cannot be identified. We developed CLEAN, a pipeline to remove unwanted sequences from both long- and short-read sequencing techniques. While focusing on Illumina and Nanopore data with their technology-specific control sequences, the pipeline can also be used for host decontamination of metagenomic reads and assemblies, or the removal of rRNA from RNA-Seq data. The results are the purified sequences and sequences identified as contaminated with statistics summarized in a report. The output can be used directly in subsequent analyses, resulting in faster computations and improved results. Although decontamination seems mundane, many contaminants are routinely overlooked, cleaned by steps that are not fully reproducible or difficult to trace. CLEAN facilitates reproducible, platform-independent data analysis in genomics and transcriptomics and is freely available at https://github.com/rki-mf1/clean under a BSD3 license. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=93 SRC="FIGDIR/small/552089v3_ufig1.gif" ALT="Figure 1"> View larger version (16K): org.highwire.dtl.DTLVardef@133c638org.highwire.dtl.DTLVardef@1ab448org.highwire.dtl.DTLVardef@1320f21org.highwire.dtl.DTLVardef@17f7a9c_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗
bioRxiv2023

Impact of reference design on estimating SARS-CoV-2 lineage abundances from wastewater sequencing data

BackgroundSequencing of SARS-CoV-2 RNA from wastewater samples has emerged as a valuable tool for detecting the presence and relative abundances of SARS-CoV-2 variants in a community. By analyzing the viral genetic material present in wastewater, public health officials can gain early insights into the spread of the virus and inform timely intervention measures. The construction of reference datasets from known SARS-CoV-2 lineages and their mutation profies has become state-of-the-art for assigning viral lineages and their relative abundances from wastewater sequencing data. However, the selection of reference sequences or mutations directly affects the predictive power. ResultsHere, we show the impact of a mutation- and sequence-based reference reconstruction for SARS-CoV-2 abundance estimation. We benchmark three data sets: 1) synthetic "spike-in" mixtures, 2) German samples from early 2021, mainly comprising Alpha, and 3) samples obtained from wastewater at an international airport in Germany from the end of 2021, including 1rst signals of Omicron. The two approaches differ in sub-lineage detection, with the marker-mutation-based method, in particular, being challenged by the increasing number of mutations and lineages. However, the estimations of both approaches depend on selecting representative references and optimized parameter settings. By performing parameter escalation experiments, we demonstrate the effects of reference size and alternative allele frequency cutoffs for abundance estimation. We show how different parameter settings can lead to different results for our test data sets, and illustrate the effects of virus lineage composition of wastewater samples and references. ConclusionsHere, we compare a mutation- and sequence-based reference construction and assignment for SARS-CoV-2 abundance estimation from wastewater samples. Our study highlights current computational challenges, focusing on the general reference design, which significantly and directly impacts abundance allocations. We illustrate advantages and disadvantages that may be relevant for further developments in the wastewater community and in the context of higher standardization.

bioinformatics↗
bioRxiv2023

Sequencing accuracy and systematic errors in nanopore direct RNA sequencing

Direct RNA sequencing (dRNA-seq) on the Oxford Nanopore Technologies (ONT) platforms can produce reads covering up to full-length gene transcripts while containing decipherable information about RNA base modifications and poly-A tail lengths. Although many published studies have been exploring and expanding the potential of dRNA-seq, the sequencing accuracy and error patterns remain understudied. We present the first comprehensive evaluation of accuracy and systematic errors in dRNA-seq data from diverse species, as well as synthetic RNA. Deletions significantly outnumbered mismatches/insertions, while the median read accuracy exhibited species-level variation. In addition to homopolymer errors, we observed systematic biases across nucleotides and heteropolymeric motifs in all species. In general, cytosine/uracil-rich regions were more likely to be erroneous than guanines/adenines. Moreover, the systematic errors were strongly dependent on local sequence contexts. By examining raw signal data, we identified underlying signal-level features potentially associated with the error patterns. While read quality scores approximated error rates at base and read levels, failure to detect DNA adapters may lead to data loss. By comparing distinct basecallers, we reason that some sequencing errors are attributable to signal insufficiency rather than algorithmic (base-calling) artefacts. Lastly, we discuss the implications of such error patterns for downstream applications of dRNA-seq data.

bioinformatics↗
bioRxiv2023

Functional comparisons of the virus sensor RIG-I from humans, the microbat Myotis daubentonii, and the megabat Rousettus aegyptiacus, and their response to SARS-CoV-2 infection

Bats (order Chiroptera) are a major reservoir for emerging and re-emerging zoonotic viruses. Their tolerance towards highly pathogenic human viruses led to the hypothesis that bats may possess an especially active antiviral interferon (IFN) system. Here, we cloned and functionally characterized the virus RNA sensor, Retinoic Acid-Inducible Gene-I (RIG-I), from the "microbat" Myotis daubentonii (suborder Yangochiroptera) and the "megabat" Rousettus aegyptiacus (suborder Yinpterochiroptera), and compared them to the human ortholog. Our data show that the overall sequence and domain organization is highly conserved and that all three RIG-I orthologs can mediate a similar IFN induction in response to viral RNA at 37{degrees} and 39{degrees}C, but not at 30{degrees}C. Like human RIG-I, bat RIG-Is were optimally activated by double stranded RNA containing a 5-triphosphate end and required Mitochondrial Antiviral-Signalling Protein (MAVS) for antiviral signalling. Moreover, the RIG-I orthologs of humans and of R. aegyptiacus, but not of M. daubentonii, enable innate immune sensing of SARS-CoV-2 infection. Our results thus show that microbats and megabats express a RIG-I that is not substantially different from the human counterpart with respect to function, temperature dependency, antiviral signaling, and RNA ligand properties, and that human and megabat RIG-I are able to sense SARS-CoV-2 infection. IMPORTANCEA common hypothesis holds that bats (order Chiroptera) are outstanding reservoirs for zoonotic viruses because of a special antiviral interferon (IFN) system. However, functional studies about key components of the bat IFN system are rare. RIG-I is a cellular sensor for viral RNA signatures that activates the antiviral signalling chain to induce IFN. We cloned and functionally characterized RIG-I genes from representatives of the suborders Yangochiroptera and Yinpterochiroptera. The bat RIG-Is were conserved in their sequence and domain organization, and similar to human RIG-I in (i) mediating virus- and IFN-activated gene expression, (ii) antiviral signalling, (iii) temperature dependence, and (iv) recognition of RNA ligands. Moreover, RIG-I of Rousettus aegyptiacus (suborder Yinpterochiroptera) and of humans were found to recognize SARS-CoV-2 infection. Thus, members of both bat suborders encode RIG-Is that are comparable to their human counterpart. The ability of bats to harbour zoonotic viruses therefore seems due to other features.

immunology↗
bioRxiv2022

VIRify: an integrated detection, annotation and taxonomic classification pipeline using virus-specific protein profile hidden Markov models

The study of viral communities has revealed the enormous diversity and impact these biological entities have on a range of different ecosystems. These observations have sparked widespread interest in developing computational strategies that support the comprehensive characterization of viral communities based on sequencing data. Here we introduce VIRify, a new computational pipeline designed to provide a user-friendly and accurate functional and taxonomic characterization of viral communities. VIRify identifies viral contigs and prophages from metagenomic assemblies and annotates them using a collection of viral profile hidden Markov models (HMMs). These include our manually-curated profile HMMs, which serve as specific taxonomic markers for a wide range of prokaryotic and eukaryotic viral taxa and are thus used to reliably classify viral contigs. We tested VIRify on assemblies from two microbial mock communities and a large metagenomics study. The results showed that VIRify was able to identify sequences from both prokaryotic and eukaryotic viruses, and provided taxonomic classifications from the genus to the family rank with an accuracy of at least 95.5%. In addition, VIRify allowed the detection and taxonomic classification of a range of prokaryotic and eukaryotic viruses present in 243 marine metagenomic assemblies. Overall, we demonstrate that VIRify is a novel and powerful resource that offers an enhanced capability to detect a broad range of viral contigs and taxonomically classify them.

bioinformatics↗
bioRxiv2022

Assembling highly repetitive Xanthomonas TALomes using Oxford Nanopore sequencing

Most plant-pathogenic Xanthomonas bacteria harbor transcription activator-like effector (TALE) genes, which function as transcriptional activators of host plant genes and support infection. The entire repertoire of up to 29 TALE genes of a Xanthomonas strain is also referred to as TALome. The DNA-binding domain of TALEs is comprised of highly conserved repeats and TALE genes often occur in gene clusters, which precludes the assembly of TALE-carrying Xanthomonas genomes based on standard sequencing approaches. Here, we report the successful assembly of the 5 Mbp genomes of five Xanthomonas strains from Oxford Nanopore Technologies (ONT) sequencing data. For one of these strains, Xanthomonas oryzae pv. oryzae (Xoo) PXO35, we illustrate why Illumina short reads and longer PacBio reads are insufficient to fully resolve the genome. While ONT reads are perfectly suited to yield highly contiguous genomes, they suffer from a specific error profile within homopolymers. To still yield complete and correct TALomes from ONT assemblies, we present a computational correction pipeline specifically tailored to TALE genes, which yields at least comparable accuracy as Illumina-based polishing. We further systematically assess the ONT-based pipeline for its multiplexing capacity and find that, combined with computational correction, the complete TALome of Xoo PXO35 could have been reconstructed from less than 20,000 ONT reads. Our results indicate that multiplexed ONT sequencing combined with a computational correction of TALE genes constitutes a highly capable tool for characterizing the TALomes of huge collections of Xanthomonas strains in the future.

bioinformatics↗
bioRxiv2021

Rates of primary production in groundwater rival those in oligotrophic marine systems

The terrestrial subsurface contains nearly all of Earths freshwater reserves1 and harbors upwards of 60% of our planets total prokaryotic biomass2,3. While genetic surveys suggest these organisms rely on in situ carbon fixation, rather than the translocation of photosynthetically derived organic carbon4-6, corroborating measurements of carbon fixation in the subsurface are absent. Using a novel ultra-low level 14C-labeling technique, we show that in situ carbon fixation rates in a carbonate aquifer reached 10% of the median rates measured in oligotrophic marine surface waters, and were up to six-fold greater than those observed in lower euphotic zone waters where deep chlorophyll levels peak. Empirical carbon fixation rates were substantiated by both nitrification and anammox rate data. Metagenomic analyses revealed a remarkable abundance of putative chemolithoautotrophic members of an uncharacterized order of Nitrospiria - the first representatives of this class expected to fix carbon via the Wood-Ljungdahl pathway. Based on these fixation rates, we extrapolate global primary production in carbonate groundwaters to be 0.11 Pg of carbon per year.

microbiology↗
bioRxiv2021

Adaptive nanopore sequencing on miniature flow cell detects extensive antimicrobial resistence

Rapid screening of hospital admissions to detect asymptomatic carriers of resistant bacteria can prevent pathogen outbreaks. However, the resulting isolates rarely have their genome sequenced due to cost constraints and long turn-around times to get and process the data, limiting their usefulness to the practitioner. Here we use real-time, on-device target enrichment ("adaptive") sequencing as a highly multiplexed assay covering 1,147 antimicrobial resistance genes. We compare its utility against standard and metagenomic sequencing, focusing on an isolate of Raoultella ornithinolytica harbouring three carbapenemases (NDM, KPC, VIM). Based on this experimental data, we then model the influence of several variables on the enrichment results and predict a large effect of nucleotide identity (higher is better) and read length (shorter is better). Lastly, we show how all relevant resistance genes are detected using adaptive sequencing on a miniature ("Flongle") flow cell, motivating its use in a clinical setting to monitor similar cases and their surroundings.

microbiology↗
bioRxiv2021

The economical lifestyle of CPR bacteria in groundwater allows little preference for environmental drivers

The highly diverse Cand. Patescibacteria are predicted to have minimal biosynthetic and metabolic pathways, which hinders understanding of how their populations differentiate to environmental drivers or host organisms. Their metabolic traits to cope with oxidative stress are largely unknown. Here, we utilized genome-resolved metagenomics to investigate the adaptive genome repertoire of Patescibacteria in oxic and anoxic groundwaters, and to infer putative host ranges. Within six groundwater wells, Cand. Patescibacteria was the most dominant (up to 79%) super-phylum across 32 metagenomes obtained from sequential 0.2 and 0.1 {micro}m filtration. Of the reconstructed 1275 metagenome-assembled genomes (MAGs), 291 high-quality MAGs were classified as Cand. Patescibacteria. Cand. Paceibacteria and Cand. Microgenomates were enriched exclusively in the 0.1 {micro}m fractions, whereas candidate division ABY1 and Cand. Gracilibacteria were enriched in the 0.2 {micro}m fractions. Patescibacteria enriched in the smaller 0.1 {micro}m filter fractions had 22% smaller genomes, 13.4% lower replication measures, higher fraction of rod-shape determining proteins, and genomic features suggesting type IV pili mediated cell-cell attachments. Near-surface wells harbored Patescibacteria with higher replication rates than anoxic downstream wells characterized by longer water residence time. Except prevalence of superoxide dismutase genes in Patescibacteria MAGs enriched in oxic groundwaters (83%), no major metabolic or phylogenetic differences were observed based on oxygen concentrations. The most abundant Patescibacteria MAG in oxic groundwater encoded a nitrate transporter, nitrite reductase, and F-type ATPase, suggesting an alternative energy conservation mechanism. Patescibacteria consistently co-occurred with one another or with members of phyla Nanoarchaeota, Bacteroidota, Nitrospirota, and Omnitrophota. However, only 8% of MAGs showed highly significant one-to-one association, mostly with Omnitrophota. Genes coding for motility and transport functions in certain Patescibacteria were highly similar to genes from other phyla (Omnitrophota, Proteobacteria and Nanoarchaeota). Other than genes to cope with oxidative stress, we found little genomic evidence for niche adaptation of Patescibacteria to oxic or anoxic groundwaters. Given that we could detect specific host preference only for a few MAGs, we propose that the majority of Patescibacteria can attach to multiple hosts just long enough to loot or exchange supplies with an economic lifestyle of little preference for geochemical conditions.

microbiology↗
bioRxiv2021

Context-aware genomic surveillance reveals hidden transmission of a carbapenemase-producing Klebsiella pneumoniae

Genomic surveillance can inform effective public health responses to pathogen outbreaks. However, integration of non-local data is rarely done. We investigate two large hospital outbreaks of a carbapenemase-carrying Klebsiella pneumoniae strain in Germany and show the value of contextual data. By screening more than ten thousand genomes, 500 thousand metagenomes, and two culture collections using in silico and in vitro methods, we identify a total of 415 closely related genomes reported in 28 studies. We identify the relationship between the two outbreaks through time-dated phylogeny, including their respective origin. One of the outbreaks presents extensive hidden transmission, with descendant isolates only identified in other studies. We then leverage the genome collection from this meta-analysis to identify genes under positive selection. We thereby identify an inner membrane transporter (ynjC) with a putative role in colistin resistance. Contextual data from other sources can thus enhance local genomic surveillance at multiple levels and should be integrated by default when available.

microbiology↗
bioRxiv2021

poreCov - an easy to use, fast, and robust workflow for SARS-CoV-2 genome reconstruction via nanopore sequencing

In response to the SARS-CoV-2 pandemic, a highly increased sequencing effort has been established worldwide to track and trace ongoing viral evolution. Technologies such as nanopore sequencing via the ARTIC protocol are used to reliably generate genomes from raw sequencing data as a crucial base for molecular surveillance. However, for many labs that perform SARS-CoV-2 sequencing, bioinformatics is still a major bottleneck, especially if hundreds of samples need to be processed in a recurring fashion. Pipelines developed for short-read data cannot be applied to nanopore data. Therefore, specific long-read tools and parameter settings need to be orchestrated to enable accurate genotyping and robust reference-based genome reconstruction of SARS-CoV-2 genomes from nanopore data. Here we present poreCov, a highly parallel workflow written in Nextflow, using containers to wrap all the tools necessary for a routine SARS-CoV-2 sequencing lab into one program. The ease of installation, combined with concise summary reports that clearly highlight all relevant information, enables rapid and reliable analysis of hundreds of SARS-CoV-2 raw sequence data sets or genomes. poreCov is freely available on GitHub under the GNUv3 license: github.com/replikation/poreCov.

bioinformatics↗
bioRxiv2021

CovRadar: Continuously tracking and filtering SARS-CoV-2 mutations for molecular surveillance

The SARS-CoV-2 pandemic underlined the importance of molecular surveillance to track the evolution of the virus and inform public health interventions. Fast analysis, easy visualization and convenient filtering of the latest virus sequences are essential for this purpose. However, access to computational resources, the lack of bioinformatics expertise, and the sheer volume of sequences in public databases complicate surveillance efforts. CovRadar combines an analytical pipeline and a web application designed for the molecular surveillance of the spike gene of SARS-CoV-2, an important vaccine target. The intuitive web front-end focuses on mutations rather than viral lineages and provides easy access to frequencies and spatio-temporal distributions from global sample collections. The data is regularly updated based on a scalable and reproducible analytical back-end. With this platform, we aim to give users, those with or without bioinformatics skills or sufficient computational resources, the possibility to track and explore mutational changes in the SARS-CoV-2 spike gene and to filter, download, and further analyze data that meet their questions and needs. Advanced computational users have the ability to apply the analytical pipeline and data visualization methods locally on their own data. CovRadar is freely accessible at https://covradar.net, source code is available at https://gitlab.com/dacs-hpi/covradar. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/429146v3_ufig1.gif" ALT="Figure 1"> View larger version (52K): org.highwire.dtl.DTLVardef@194a562org.highwire.dtl.DTLVardef@1f5f0d3org.highwire.dtl.DTLVardef@195b5dborg.highwire.dtl.DTLVardef@1d65231_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗
bioRxiv2020

DarkQ: Continuous genomic monitoring using message queues

MotivationNewly sequenced genomes are often not noticed by potential stakeholders because submission to public databases is delayed, and search options are limited. However, the discovery of genomes can be vital: In pathogen outbreaks, fast updates are essential to coordinate containment efforts and prevent spread. ResultsHere we introduce DarkQ, a message queue that allows for instant sharing and discovery of genomes. AvailabilityDarkQ is released under the BSD-2 license at github.com/phiweger/darkq.

genomics↗
bioRxiv2020

What the Phage: A scalable workflow for the identification and analysis of phage sequences

Phages are among the most abundant and diverse biological entities on earth. Phage prediction from sequence data is a crucial first step to understanding their impact on the environment. A variety of bacteriophage prediction tools have been developed over the years. They differ in algorithmic approach, results, and ease of use. We, therefore, developed "What the Phage" (WtP), an easy-to-use and parallel multitool approach for phage prediction combined with an annotation and classification downstream strategy, thus, supporting the users decision-making process by summarizing the results of the different prediction tools in charts and tables. WtP is reproducible and scales to thousands of datasets through a workflow manager (Nextflow). WtP is freely available under a GPL-3.0 license (https://github.com/replikation/What_the_Phage).

bioinformatics↗
bioRxiv2020

PoSeiDon: a Nextflow pipeline for the detection of evolutionary recombination events and positive selection

SummaryPoSeiDon is an easy-to-use pipeline that helps researchers to find recombination events and sites under positive selection in protein-coding sequences. By entering homologous sequences, PoSeiDon builds an alignment, estimates a best-fitting substitution model, and performs a recombination analysis followed by the construction of all corresponding phylogenies. Finally, significantly positive selected sites are detected according to different models for the full alignment and possible recombination fragments. The results of PoSeiDon are summarized in a user-friendly HTML page providing all intermediate results and the graphical representation of recombination events and positively selected sites. Availability and implementationPoSeiDon is freely available at https://github.com/hoelzer/poseidon. The pipeline is implemented in Nextflow with Docker support and processes the output of various tools. Contacthoelzer.martin@gmail.com

bioinformatics↗
bioRxiv2020

Metagenomics workflow for hybrid assembly, differential coverage binning, transcriptomics and pathway analysis (MUFFIN)

Metagenomics has redefined many areas of microbiology. However, metagenome-assembled genomes (MAGs) are often fragmented, primarily when sequencing was performed with short reads. Recent long-read sequencing technologies promise to improve genome reconstruction. However, the integration of two different sequencing modalities makes downstream analyses complex. We, therefore, developed MUFFIN, a complete metagenomic workflow that uses short and long reads to produce high-quality bins and their annotations. The workflow is written by using Nextflow, a workflow orchestration software, to achieve high reproducibility and fast and straightforward use. This workflow also produces the taxonomic classification and KEGG pathways of the bins and can be further used by providing RNA-Seq data (optionally) for quantification and annotation. We tested the workflow using twenty biogas reactor samples and assessed the capacity of MUFFIN to process and output relevant files needed to analyze the microbial community and their function. MUFFIN produces functional pathway predictions and if provided de novo transcript annotations across the metagenomic sample and for each bin. Author SummaryRVD did the development and design of MUFFIN and wrote the first draft; BM and EBR did the critical reading and correction of the manuscript; MH did the critical reading of the manuscript and the general adjustments for the metagenomic workflow; AV did the critical reading of the manuscript and adjustments for the taxonomic classifications. CB supervised the project, did the workflow design, helped with the implementation, and revised the manuscript.

bioinformatics↗