Pipolins constitute a new group of mobile genetic elements (MGEs) that encode a primer-independent DNA polymerase (piPolB). They are widespread among the major bacterial phyla and mitochondria.
ExplorePipolin is a search tool that identifies and analyses pipolins within bacterial genomes.
Citing
- Install the requirements (see above).
wget https://github.com/pipolinlab/ExplorePipolin/archive/0.0.1.zipunzip 0.0.1.zip && cd ExplorePipolin-0.0.1pip install .(install in user site-packages) orsudo pip install .(requires superuser privileges)
NOTE: before installing, it is possible to run unit tests:
pytest or python setup.py test (from the source root directory).
How to uninstall:
(sudo) pip uninstall ExplorePipolin
- Before installing ExplorePipolin, make sure you're running the latest version of Conda:
conda update conda
- Install into a new environment in one-step:
conda create -n <new_env_name> -c pipolinlab -c conda-forge -c bioconda -c defaults explore-pipolin
NOTE: solving the environment takes time. Be patient.
As input, ExplorePipolin takes FASTA file(s) with genome sequence(s). Due to restrictions of some of the libraries used, filename must not exceed 16 characters long. A genome sequence can be either a single complete chromosome (preferred) or contigs (in a single multiFASTA file).
Usage: explore_pipolin [OPTIONS] GENOMES...
ExplorePipolin is a search tool for prediction and analysis of pipolins,
bacterial mobile genetic elements.
Options:
--out-dir-prefix TEXT Use this prefix for the output directory,
instead of the default "results" prefix.
--out-dir PATH Use this output directory instead.
--pipolb-hmm-profile PATH The HMM profile for the piPolB sequence to use as
1st priority. If not provided, the default
profile will be used instead.
--only-find-pipolbs Only find piPolB genes.
--ref-att PATH ATT sequence in FASTA file to use as 1st
priority. If not provided, the default ATT will
be used instead.
--percent-identity INTEGER Minimum percent identity for direct repeats
search [default: 85]
--max-inflate INTEGER If no borders of pipolin are found (no ATTs),
inflate the analysed region from both sides of
piPolB by this length. [default: 30000]
--skip-annotation Do not run the annotation step (i.e. Prokka).
--proteins PATH Prokka param: FASTA or GBK file to use as 1st
priority. If not provided, the default file will
be used instead.
--colours PATH A TSV file describing features to colour. Please,
refer to
https://github.com/pipolinlab/ExplorePipolin for
more information about the file structure.
--skip-colours Do not add an Easyfig-compatible colouring
scheme to the final Genbank files.
--cpus INTEGER Prokka param: Number of CPUs to use [0=all]
[default: 8]
--keep-tmp Preserve intermediate files produced during the
run (it might be useful for debugging).
--keep-going Do not stop analysis if an error is raised for
one of the genomes.
-h, --help Show this message and exit.
The output directory will contain a separate folder for each analysed genome. In the genome-specific folder, the following files can be found:
| Folder/File | Content description |
|---|---|
<genome>.log |
log file |
pipolins |
extracted pipolin sequences in FASTA format, GenBank and GFF annotations of pipolin genes with atts included |
Files in pipolins folder have the following scheme: <genome>_NvN.<type>.<ext>,
where:
<genome>— genome name, retrieved from a provided FASTA file without extension. Should not exceed 16 characters, due to Biopython restrictions!N— index of a found pipolinvN— index of a reconstructed variant (each pipolin can be reconstructed in different ways)
NOTE: When alternative orientations of the piPolB-containing fragment are possible, both reconstructions are generated. The structure denoted as v0 will include piPolB gene pointing toward the attR (usually overlaps with tRNA). The alternative pipolin with the piPolB gene leftwards referred as v1.
<type>— can becomplete(att---pol---att),truncated(att---pol---) orminimal(---pol---)<ext>— file extension (.fa,.gff,.gbk)
NOTE: A file with single_record.gbk ending is produced when separate contigs are
concatenated into a single record (after the reconstruction step). Contigs are joined
with an assembly_gap feature that can be distinguished by the presence of
/inference="ExplorePipolin" field. Also, by default, these assembly gaps have
black /colour, while all other assembly gaps are pink
(see Easyfig-compatible colouring).
Additional files, when --keep-tmp option is present:
| Folder/File | Content description |
|---|---|
<genome>.fa |
genome file copied |
pipolbs |
HMMER search results for piPolB genes |
atts |
BLAST search results for the known att sites |
atts_denovo |
Results of de novo search for att sites |
trnas |
ARAGORN search results for tRNAs/tmRNAs |
prokka |
Prokka annotation results (check files description here) |
By default, a /colour field with an RGB colour code will be added to each feature
in the output Genbank files. The colours will be assigned according to
this scheme. To modify the colouring, download
and change this TSV file and provide it using --colours option.
An example command to run Easyfig:
# IMPORTANT: only run with python2.x!
python <path>/Easyfig.py -o my_figure.svg -svg -f1 T \
-f CDS arrow -f repeat_region rect -f tRNA rect -f assembly_gap pointer \
-tblastx -min_length 500 -ann_height 100 \
single_record1.gbk single_record2.gbk ...All options can be checked by running python <path>/Easyfig.py --help.
NOTE: Use --skip-colours when running ExplorePipolin if you do not need the colours.
Genetic structure of diverse pipolins from diverse genomes from a wide range of bacteria reconstructed by ExplorePipolin.
Predicted protein-coding genes are represented by arrows, indicating the direction of transcription and more common genes are colored following Prokka annotation (piPolB in red, tyrosine recombinase in brown, UDG in cyan, excisionase in purple and metallohydrolase in magenta). When detected, other features are indicated as colored arrowheads: sequence gaps (black), E. coli related-atts (navy blue), de novo detected direct repeats (steel) and tRNAs (green). The greyscale on the right reflects the percent of amino acid identity between pairs of sequences. The image was generated by EasyFig software using tBlastX for elements comparison (options -tblastx -i 20). Names of the analized genomes are indicated on the left and colored by taxonomy: magenta, Alphaproteobacteria; red, Gammaproteobacteria; purple, fungi, forest green, Actinobacteria and green, Firmicutes. Genome assemblies and the pipeline output can be found in the example_genomes folder.
Plotting pipolins in R is possible using the GGGenomes R package. However, plotting requires a previous processing of the GBK files with a custom script located in the "gggenomes_lrec_plotting" folder, along with example files.
More details and instructions for plotting pipolins with gggenomes can be found in the README.txt file from that folder.
Early work on ExplorePipolins was supported by Grant PGC2018-093723-A-I00 from MCIN/AEI/10.13039/501100011033/ and FEDER ‘A way to make Europe’.
During 2022-2023 this project is supported by the Madrid Government (Comunidad de Madrid-Spain) under the Multiannual Agreement with Universidad Autónoma de Madrid in the line of stimulation or Research by young Researchers, in the context of the V PRICIT (Regional Programme of Research and Technological Innovation). (SI3/PJI/2021-00271).”