Pizza Seminar

15 June 2021

Inês Mendes

MRamirez Lab - iMM

@ines_cim

cimendes

short-read metagenomic assembly

finding the best fit in a world of options

Bacterial Population Genetics

Pathogenesis and Natural History of Infection

Outbreak Investigation and Control

Surveillance of Infectious Diseases

Microbial Genomics

Clinical Microbiology

| The now

Didelot, X., Bowden, R., Wilson, D. et al. Transforming clinical microbiology with bacterial genome sequencing. Nat Rev Genet 13, 601–612 (2012). https://doi.org/10.1038/nrg3226

Clinical Microbiology

| The now

Joana Silva, MRamirez

Clinical Microbiology

| The now

Didelot, X., Bowden, R., Wilson, D. et al. Transforming clinical microbiology with bacterial genome sequencing. Nat Rev Genet 13, 601–612 (2012). https://doi.org/10.1038/nrg3226

Clinical Microbiology

| The now

Loman, N., Pallen, M. Twenty years of bacterial genome sequencing. Nat Rev Microbiol 13, 787–794 (2015). https://doi.org/10.1038/nrmicro3565

Clinical Microbiology

| The future

Metagenomics

Random "shotgun" sequencing of microbial DNA, without selecting a particular gene or species.

Microbial (meta)genomics

Didelot, X., Bowden, R., Wilson, D. et al. Transforming clinical microbiology with bacterial genome sequencing. Nat Rev Genet 13, 601–612 (2012). https://doi.org/10.1038/nrg3226

Microbial (meta)genomics

| Assembly

The assembly methods provide longer sequences that are more informative than shorter sequencing data and can provide a more complete picture of the microbial community in a given sample.

Assembly

de novo

consensus

OLC

de Bruijn graph

de novo

OLC

de Bruijn graph

Microbial (meta)genomics

| de novo Assembly

Martin Ayling, Matthew D Clark, Richard M Leggett, New approaches for metagenome assembly with short reads, Briefings in Bioinformatics, Volume 21, Issue 2, March 2020, Pages 584–594, https://doi.org/10.1093/bib/bbz020

Microbial (meta)genomics

| de novo Assembly

Results are highly dependent on the tools chosen for the analysis - Lack of standardization and proper benchmark.

Major issues

Reads

Contigs

Genomes

Highlights the potential and the limitations of shotgun metagenomics as a diagnostic tool - Lack of reproducibility

https://github.com/cimendes/LMAS

https://lmas.readthedocs.io/

LMAS

| Last Metagenomic Assembler Standing

Automated workflow enabling the benchmarking of genomic and metagenomic prokaryotic de novo assembly software using defined mock communities.

LMAS

| Last Metagenomic Assembler Standing

Git, Nextflow (java) and a container engine (Docker, singularity, shifter...).

apt-get install git

curl -s https://get.nextflow.io | bash

apt-install docker-ce

Clone

git clone https://github.com/cimendes/LMAS.git

Run LMAS

nextflow run LMAS.nf

LMAS

| Last Metagenomic Assembler Standing

The input data is assembled in parallel by the set of genomic and metagenomic de novo assemblers in LMAS.

The global and per reference metrics are grouped in the interactive LMAS report for exploration.

The resulting assembled sequences are processed and assembly quality metrics are computed.

LMAS

| Assembly Quality Metrics

The tabular presentation allows direct comparison of exact values between assemblies, and the interactive plots allow for an intuitive overview and easy exploration of results.