Frequently Asked Questions

How to report an issue?

If you encounter a problem while running MacSyFinder, please submit an issue on the dedicated page of the GitHub project

To ensure we have all elements to help, please provide:

  • a concise description of the issue

  • the expected behavior VS observed one

  • the exact command-line used

  • the version of MacSyFinder used

  • the exact error message, and if applicable, the macsyfinder.log and macsyfinder.conf files

  • if applicable, an archive (or link to it) with the output files obtained

  • if possible, the smallest dataset there is to reproduce the issue

  • if applicable, this would also include the macsy-models (XML models plus HMM profiles) used (or precise version of the models if there are publicly available). Same as above, if possible, please provide the smallest set possible of models and HMM profiles.

All these will definitely help us to help you! ;-)

How to cite MacSyFinder and published macy-models?

  • Abby et al. 2014, PLoS ONE for the general principles of MacSyFinder (version 1), and the corresponding set of Cas systems (CasFinder, 1st version).

  • Abby and Rocha 2012, PLoS Genetics, for the study of the evolutionary relationship between the T3SS and the bacterial flagellum, and how were designed the corresponding HMM protein profiles.

  • Abby et al. 2016, Scientific Reports, for the description of bacterial protein secretion systems’ models (TXSScan: T1SS, T2SS, T5SS, T6SS, T9SS, Tad, T4P).

  • Denise et al. 2019, PLoS Biology, for the description of type IV-filament super-family models (TFF-SF: T2SS, T4aP, T4bP, Com, Tad, archaeal T4P).

  • Rendueles et al. 2017, PLoS Pathogens, for the CapsuleFinder set of models.

  • Couvin, Bernheim et al. 2018, Nucleic Acids Research, for the updated version of the set of Cas systems’ models, CasFinder.

What do MacSyFinder command lines look like?

Here are a few examples of command line formation:

To browse interactive help:

macsyfinder -h

The minimal command line, to search all systems with models from the “TFF-SF” set of models (installed with macsydata):

macsyfinder --db-type ordered_replicon --sequence-db genome.fasta --models TFF-SF all

To search for several systems (ModelA and ModelB) from the “model_family” set of models that can be found in the “./my-models” folder:

macsyfinder --db-type ordered_replicon --sequence-db genome.fasta --models model_family ModelA ModelB --models-dir ./my-models

To alter the search parameters and allow a maximal distance between components of 20 for the T2SS and 15 for the Tad pilus:

macsyfinder --db-type ordered_replicon --sequence-db genome.fasta --models TFF-SF all --inter-gene-max-space T2SS 20 --inter-gene-max-space Tad 15

To alter the search parameters and allow the Tad pilus to be made of multiple loci:

macsyfinder --db-type ordered_replicon --sequence-db genome.fasta --models TFF-SF all --multi-loci Tad

In gembase or ordered_replicon mode macsyfinder need to index the sequence-db. By default, this index is write beside the sequence-db file. But sometimes the directory where the sequence-db is located is not writable, in centralized shared data in multi user environement for instance. To avoid to copy sequences in other location, you could specify an alternate directory for the index with --index-dir (This directory must exists):

macsyfinder --db-type ordered_replicon --sequence-db genome.fasta --index-dir my-indexes --models TFF-SF all

See also the MacSyFinder Quick Start section for more examples.

What search mode to be used?

Depending on the type of dataset you have, you will have to adapt MacSyFinder’s search mode.

  • If you have a fasta file from a complete genome where proteins are ordered according to the corresponding genes’ order along the replicon, your dataset is entitled to the most powerful search mode (see below): ordered_replicon and use the following option –db-type ordered_replicon.

  • If you have a fasta file of proteins with no sense of the order of the corresponding genes along the chromosome(s) or replicon(s), you will have to use the unordered search mode with the following option: –db-type unordered

  • If you have multiple ordered replicons to analyse at once, you can follow the Gembase convention to name the proteins in the fasta file, so that the original replicons can be assessed from their name: see here for a description.

Note

  • When the gene order is known (ordered_replicon search mode) the power of the analysis is maximal, since both the genomic content and context are taken into account for the search.

  • When the gene order is unknown (unordered search mode) the power of the analysis is more limited since the presence of systems can only be suggested on the basis of the quorum of components - and not based on genomic context information.

More on command-line options here and on MacSyFinder’s functioning here.

How to deal with fragmented genomes (MAGs, SAGs, draft genomes)?

There are more and more genomes available which are not completely assembled, or are fragmented and incomplete. In this case, several options can be considered.

1. If your genome is at least partially assembled and contigs are not too short, you might “feel lucky” and first consider to run MacSyFinder with the ordered_replicon mode. It could be particularly efficient if you are investigating systems encoded by compact loci (Cas systems, some secretion systems…), as they might be encoded by a single contig.

2. On top of the ordered_replicon mode, you might add the option “multi-loci” to the systems to annotate (if not already the case), in order to maximize the chance to annotate an entire system, even if encoded across several contigs.

3. The unordered mode can be used in complement of the two above options, e.g. to retrieve some of the missing components. It will enable to assess the genetic potential and possible presence of a system, independently of the quality of assembly of the genome. It might also be the only reasonable option if the genome is too fragmented and/or too incomplete.

Note

  • The results obtained with the ordered_replicon mode on a fragmented genome have to be considered carefully, especially with respect to the contigs’ borders, as some proteins from different contigs might be artificially considered as closely encoded.

  • To retrieve “fragments” of a system not found to reach the quorum in the ordered_replicon mode, it is possible to retrieve clusters of genes from the rejected_candidates.tsv file.

How to search for multiple systems at once?

  • It is possible to search for only some systems from a macsy-model package. In this case, the command-line should be formed as follows:

macsyfinder --models TXSS Flagellum T2SS --sequence-db mygenomes.fasta --db-type gembase

This would run the search of the systems “Flagellum” and “T2SS” in the dataset “mygenomes.fasta”.

  • To run the search of all the models contained in a macsy-model package, use the following:

macsyfinder --models TXSS all --sequence-db mygenomes.fasta --db-type gembase
macsyfinder --models CRISPRCas all --sequence-db mygenomes.fasta --db-type gembase
macsyfinder --models CRISPRCas/typing all --sequence-db mygenomes.fasta --db-type gembase

You can see that the all keyword can not only be applied to an entire macsy-model package and its entire hierarchy, but can also be ran on all the systems from a macsy-model sub-directory.

When can the option –previous-run be used?

The option –previous-run enables to avoid running the HMM profile search and the hits extraction when the set of systems to search and the replicons to analyse are exactly the same between runs. This enables to alter the features of the systems to be searched for, i.e. basically any feature found in the XML file of the corresponding models:

  • the maximal distance allowed between components to be considered as part of a same locus –inter-gene-max-space

  • the minimal number of components to be found to infer a full system –min-mandatory-genes-required and –min-genes-required

  • the general genomic architecture of the system –multi-loci

This also means that there are a number of options that are incompatible with –previous-run, including:

--config, --sequence-db, --profile-suffix, --res-extract-suffix, --e-value-res, --db-type, --hmmer

Which output file to be used to get ONE solution?

Since version 2 of MacSyFinder, a combinatorial exploration of the possible sets of systems is performed. A scoring scheme has been set up to differentiate between solutions, in order to provide the user with the most complete set of systems as possible given the searched models. This score is maximal for the “best solution”. This also means that some solutions might get the same maximal score. In this case, one can wonder how to find all the equivalent solutions, and an other, how to simply pick one solution among the best, whichever it is. We thus propose several kind of output files.

  • All equivalent best solutions are found in the all_best_solutions.tsv file.

  • One best solution is given in the best_solution.tsv file.

Note

For those more familiar with the output files from MacSyFinder v1, the file best_solution.tsv is the closest from the previous output file macsyfinder.report.

Where to find MacSyFinder models?

Since version 2, there is a tool to enable the download and installation of published models from a repository: the macsydata tool.

See here for details on how to use it.

What are the rules for options precedence?

MacSyFinder offers many ways to parametrize the systems’ search: through the command-line, through various configuration files (for the models, for the run, etc…). It offers a large control over the search engine. But it also means you can get lost in configuration. ;-)

Here is a recap of the rules for options precedence. In a general manner, the command line always wins.

The precedence rules between the different levels of configuration are:

system < home < model < project < --cfg-file | --previous-run < command line options

  • system: the macsyfinder.conf file either in /etc/macsyfinder/ or in ${VIRTUAL_ENV}/etc/macsyfinder/ in case of a virtualenv this configuration affects only the MacSyFinder version installed in this virtualenv

  • home: the ~/.macsyfinder/macsyfinder.conf file

  • model: the model_conf.xml file at the root of the model package

  • project: the macsyfinder.conf file found in the directory where the macsyfinder command was run

  • cfgfile: any configuration file specified by the user on the command line (conflicts with the –previous-run option)

  • previous-run: the macsyfinder.conf file found in the results directory of the previous run (conflicts with the –cfg-file option)

  • command line: any option specified directly in the command line