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

Abstract

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 for quantification and annotation by providing RNA-Seq data (optionally). 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 metatranscript annotations across the metagenomic sample and for each bin. MUFFIN is available on github under GNUv3 licence: https://github.com/RVanDamme/MUFFIN.

Introduction to the principles and methods underlying the recovery of metagenome‐assembled genomes from metagenomic data

“…In our experience, results are mixed, as current software has trouble incorporating all the input data into a cohesive whole, but as highlighted by Van Damme et al ( 2021 ) when proposing their metagenomics pipeline called MUFFIN, a thorough review of hybrid assemblers is still pending. Moreover, publications presenting new assemblers typically show them performing very well in a limited context and with a selected number of reference genomes.…”

nf-core/mag: a best-practice pipeline for metagenome hybrid assembly and binning

“…Several pipelines have been developed for the assembly and binning of metagenomes ( 4–6 ). However, only a few pipelines such as Muffin ( 7 ) and ATLAS ( 8 ) make use of workflow management systems, such as Snakemake ( 9 ) or Nextflow ( 10 ), which facilitate scalability, portability, reproducibility and ease of application. These pipelines have different strengths and weaknesses but only Muffin supports hybrid assembly and none of them supports co-assembly.…”

MIntO: A Modular and Scalable Pipeline For Microbiome Metagenomic and Metatranscriptomic Data Integration

“…Standard metagenomic and metatranscriptomic approaches entail 1) read curation, 2) de novo assembly and/or co-assembly, 3) binning, 4) gene prediction, 5) annotation of predicted genes at taxonomic and functional level and 6) quantification of gene abundances and transcripts. However, most of the computational pipelines developed so far can only analyze metagenomic or metatranscriptomic data individually and only few, reported in Table 1 , can handle both meta-omics data ( Kim et al, 2016 ; Narayanasamy et al, 2016 ; Tamames and Puente-Sánchez, 2018 ; Salazar et al, 2019 ; Sequeira et al, 2019 ; Van Damme et al, 2021 ). Furthermore, only one of them ( Van Damme et al, 2021 ) can combine two sequencing technologies (Nanopore or long-sequences and Illumina or short-sequences) to recover MAGs.…”