The scripts 'attC-screening.sh' and 'IntI-screening.sh' can be used to filter DNA sequences to ensure that they represent amplicons from genuine integrons.
The 'attC-screening.sh' script filters input sequences for gene cassette recombination sites (attCs) and should be used for attC-attC amplicon data. The 'IntI-screening.sh' script filters input sequences for intergron integrase (IntI) protein sequences and should be used for intI-attC amplicon data.
This script uses the HattCI + Infernal pipeline that has been previously described here and here. In short, 'attC-screening.sh' searches for the sequence and secondary structures conserved among attCs and retains any input sequence that has at least one attC site. The script can be used on data generated from any sequencing technology.
usage: ./attC-screening.sh
Mandatory arguments:
-m : universal attC CM ('attCs.cm')
-i : input fasta of DNA sequences
Optional arguments:
-b : bit score cut-off for cmsearch | default: 20
-l : minimum length of output sequences | default: 500 bp
-t : number of CPUs | default: 1
-o : output directory | default: current directory
Dependencies (Script may work with older or newer versions):
INFERNAL version 1
SAMtools v1.12
SeqKit v0.7.2
The script uses a profile HMM used by the Integron Finder program to detect the additional domain that is unique to integron integrases. The pipeline first uses Prodigal to predict all encoded protein sequences, and then screens them for the IntI-specific domain using hmmsearch from the HMMER v3 software package. Any sequences that do not contain a recognisable integron integrase are discarded. Similarly, intI_screening.sh can be used on data generated from any sequencing technology.
usage: ./IntI-screening.sh
Mandatory arguments:
-m : IntI HMM ('IntI-Cterm.hmm')
-i : input fasta of DNA sequences
Optional arguments:
-l : minimum length of output sequences | default: 800 bp
-t : number of CPUs | default: 1
-o : output directory | default: current directory
Dependencies (Script may work with older or newer versions):
Prodigal v2.6.3
HMMER version 3
SAMtools v1.12
SeqKit v0.7.2
If you use either script (attC-screening.sh or IntI-screening.sh), please cite:
Ghaly, T.M., Penesyan, A., Pritchard, A., Qi, Q., Rajabal, V., Tetu, S.G. and Gillings, M.R. 2021. Methods for the targeted sequencing and analysis of integrons and their gene cassettes from complex microbial communities. Microbial Genomics, 8(3): 000788. doi: 10.1099/mgen.0.000788.
Please also cite all of their dependencies listed above.
IntI-screening.sh makes use of the IntI-Cterm.hmm profile used by Integron Finder. If you use this script, please also cite their paper:
Cury, J., Jové, T., Touchon, M., Néron, B. and Rocha, E.P. 2016. Identification and analysis of integrons and cassette arrays in bacterial genomes. Nucleic Acids Research, 44(10):4539-4550.