MOB-suite: Software tools for clustering, reconstruction and typing of plasmids from draft assemblies
Plasmids are mobile genetic elements (MGEs), which allow for rapid evolution and adaption of bacteria to new niches through horizontal transmission of novel traits to different genetic backgrounds. The MOB-suite is designed to be a modular set of tools for the typing and reconstruction of plasmid sequences from WGS assemblies.
The MOB-suite depends on a series of databases which are too large to be hosted in git-hub. They can be downloaded or updated by running mob_init or if running any of the tools for the first time, the databases will download and initialize automatically. However, they are quite large so the first run will take a long time depending on your connection and speed of your computer. The databases can be downloaded from figshare here: https://ndownloader.figshare.com/articles/5841882/versions/1
On first run of MOB-typer or MOB-recon, MOB-init should run to download the databases from figshare, sketch the databases and setup the blast databases. However, it can be run manually if the databases need to be re-initialized.
% mob_init
This tool creates plasmid similarity groups using fast genomic distance estimation using MASH. Plasmids are grouped into clusters using single-linkage clustering and the cluster codes provided by the tool provide an approximation of operational taxonomic units OTU’s
This tool reconstructs individual plasmid sequences from draft genome assemblies using the clustered plasmid reference databases provided by MOB-cluster.
Provides in silico predictions of the replicon family, relaxase type, mate-pair formation type and predicted transferability of the plasmid
Python v. 3.6 +
blast+ v. 2.3.0 + circlator mash
We recommend MOB-Suite installation as a conda package due to large number of dependencies. The package is available through bioconda channel.
% conda config --add channels defaults
% conda config --add channels conda-forge
% conda config --add channels bioconda
% conda install blast amos mash circlator mob_suite
The MOB-suite uses the minimus2 pipeline from Circlator but there are some hardcoded links which need to be created in order for the tool to work correctly. After installing circlator and amos run the following as root.
option 1: from https://github.com/sanger-pathogens/circlator/issues/65
You will need to change ${CONDA_PREFIX}/bin/minimus2 for both SHOWCOORDS and DELTAFILTER
Interestingly the two variables have also been hard coded, but fortunately using the ${CONDA_PREFIX} variable.
Run the following line fixed the code whilst inside the environment.
sed -i 's%/usr/local%${CONDA_PREFIX}%g' ${CONDA_PREFIX}/bin/minimus2
option 2:
% which show-coords
using the path above as "conda-show-coords-path"
% ln -s conda-show-coords-path /usr/local/bin/show-coords
We recommend installing via bioconda but you can install it via pip using the command below
% pip3 install mob_suite
Clone this repository and install via setuptools.
% git clone https://github.com/phac-nml/mob-suite.git
% cd mob-suite
% python setup.py install
You can perform plasmid typing using a fasta formated file containing a single plasmid represented by one or more contigs. Do not include multiple unrelated plasmids in the file as they will be treated as a single plasmid.
# Basic Mode
% mob_typer --infile assembly.fasta --outdir my_out_dir
# Look for a file called mobtyper_(input_file)_report.txt
% cat my_out_dir/mobtyper_(input_file)_report.txt
This procedure works with draft or complete genomes and is agnostic of assembler choice but if unicycler is used, then the circularity information can be parsed directly from the header of the unmodified assembly.
### Basic Mode
% mob_recon --infile assembly.fasta --outdir my_out_dir
### Full Mode
# In this mode, MOB-typer will be run on each identified plasmid grouping and will produce a summary report
% mob_recon --infile assembly.fasta --outdir my_out_dir --run_typer
Use this tool only to update the plasmid databases or build a new one and should only be completed with closed high quality plasmids. If you add in poor quality data it will severely impact MOB-recon
### Build a new database
% mob_cluster --mode build --infile plasmid.fasta --outdir output_directory
### Add a sequence to an existing database
% mob_cluster --infile update_sequences.fasta --ref_fasta_file original.fasta --ref_mash_db original.msh --ref_cluster_file original_clusters.txt
### Test new plasmid database with MOB-recon
% makeblastdb -in path_to_plasmid_testing_db -dbtype nucl
% mash sketch -i path_to_plasmid_testing_db <---- produces mash sketch file with format "path_to_plasmid_testing_db.msh"
% mob_recon --infile assembly.fasta --outdir my_out_dir --run_typer --plasmid_mash_db path_to_mash_testing_db --plasmid_db path_to_plasmid_testing_db
### Update MOB-suite plasmid databases
% mv new_mob_formated_db.fasta mob_db_path/ncbi_plasmid_full_seqs.fas
% makeblastdb -in mob_db_path/ncbi_plasmid_full_seqs.fas -dbtype nucl
% mash sketch -i mob_db_path/ncbi_plasmid_full_seqs.fas
| file | Description |
|---|---|
| contig_report.txt | This file describes the assignment of the contig to chromosome or a particular plasmid grouping |
| repetitive_blast_report | Summary information of contigs found to consist of nothing but a repetitive element |
| chromosome.fasta | Fasta file of all contigs found to belong to the chromosome |
| plasmid_(X).fasta | Each plasmid group is written to an individual fasta file which contains the assigned contigs |
| mobtyper_(input_file)_report.txt | Individual MOB-typer report files for each identified plasmid |
| mobtyper_aggregate_report.txt | Aggregate MOB-typer report files for all identified plasmid |
| field id | description |
|---|---|
| file_id | Name of the input file |
| cluster_id | MOB-cluster type of reference match |
| contig_id | Unique identifier of the contig |
| contig_length | Length of the contig |
| circularity_status | Circular if Circlator or Unicycler find it to be circular, and incomplete if not |
| rep_type | Replicon types idenfied |
| rep_type_accession | Accessions of replicons identified |
| relaxase_type | Relaxase types identified |
| relaxase_type_accession | Accessions of relaxases identified |
| mash_nearest_neighbor | Mate-pair formation types identified |
| mash_neighbor_distance | Mate-pair formation type accessioons |
| repetitive_dna_id | Repetitive DNA match id |
| match_type | Repetitive element class |
| score | Blast bitscore of match |
| contig_match_start | Start of match on contig |
| contig_match_end | End of match on contig |
| field name | description |
|---|---|
| file_id | Name of the input file |
| num_contigs | Number of sequences identified in the file |
| total_length | Total number of bases in all sequences |
| gc | GC% of all sequences |
| rep_type(s) | Replicon types idenfied |
| rep_type_accession(s) | Accessions of replicons identified |
| relaxase_type(s) | Relaxase types identified |
| relaxase_type_accession(s) | Accessions of relaxases identified |
| mpf_type | Mate-pair formation types identified |
| mpf_type_accession(s) | Mate-pair formation type accessioons |
| orit_type(s) | Relaxase type of oriT sequence |
| orit_accession(s) | Accession for oriT |
| PredictedMobility | Mobility prediction for the plasmid (Conjugative, Mobilizable, Non-mobilizable) |
| mash_nearest_neighbor | Accession of closest database match |
| mash_neighbor_distance | Mash distance from query to match |
| mash_neighbor_cluster | MOB-cluster type of reference match |
| field name | description |
|---|---|
| qseqid | query sequence id |
| sseqid | subject sequence id |
| qlen | query length |
| slen | subject length |
| qstart | match start query |
| qend | match end query |
| sstart | match subject start |
| send | match subject end |
| length | length of alignment |
| mismatch | number of mismatches |
| pident | identity |
| qcovhsp | query coverage by hsp |
| qcovs | query coverage by subject |
| sstrand | strad of hit in subject |
| evalue | evalue of match |
| bitscore | bitscore of match |
James Robertson - [email protected]
Licensed under the Apache License, Version 2.0 (the "License"); you may not use this work except in compliance with the License. You may obtain a copy of the License at:
http://www.apache.org/licenses/LICENSE-2.0
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