Often the new/latest species/data/results (AMR/Virulence/MLST…etc,.) not included in the databases (DBs) which are used by tools. But it is possible to use the latest data with some of the tools. This documentation will walk you through "How to create your own database" or "add the new data to existing database" for three of the most commonly used tools:
- ARIBA: ReadMapping Approach
- ResFinder and PointFinder: Local alignment Approach
- MLST: Local alignment
Tool Input: Assembly (.fasta) or reads (.fastq). For creating DB: fasta sequence with gene name, which wi
Login to SAGA
ssh username@saga.sigma2.no
Activate ARIBA conda environment
conda activate ARIBA
Go into test directory.
cd /cluster/projects/nn9305k/tutorial/300522_User_Defined_DBs/
ls -lh
total 1.0K
drwxrwsr-x 2 jeevka nn9305k 1 May 27 11:12 Additional_Virulence_Genes
drwxrwsr-x 3 jeevka nn9305k 28 May 27 13:04 db_resfinder
drwxrwsr-x 3 jeevka nn9305k 1 May 27 13:49 MLST
drwxrwsr-x 2 jeevka nn9305k 2 May 27 11:11 Test_Fastq_Files
Additional_Virulence_Genes: contains the fasta sequences of additional virulence genes.
Test_Fastq_Files: contains a test fastq paired end sequences
Creating data base from given fasta file
ariba prepareref --all_coding yes -f Add_Virulence_Genes/Additional_Vir_genes.fasta ariba_virulence_genes_DB
Add_Virulence_Genes/Additional_Vir_genes.fasta : input files. ariba_virulence_genes_DB : output folder which contains the db
Mapping the reads to newly created db for new virulence genes
ariba run ariba_virulence_genes_DB/ Test_Fastq_Files/Vibrio_R1.fastq Test_Fastq_Files/Vibrio_R2.fastq Virulence_Output
Virulence_Output/ contains the output files.
"report.tsv" - file contains the results
Deactivate ARIBA conda environment
conda deactivate
Input: Assembly (.fasta) or reads (.fastq)
To ADD new genes to DB:
- <resistance_phenotye_class>.fsa file: multifasta file of AMR genes. f. ex. aminoglycoside.fsa
head aminoglycoside.fsa
>aac(6')-Ib_2_M23634
ATGAGTATTCAACATTTCCAAAGAAAGTTAGGCATCACAAAGTACAGCATCGTGACCAAC
AGCAACGATTCCGTCACACTGCGCCTCATGACTGAGCATGACCTTGCGATGCTCTATGAG
TGGCTAAATCGATCTCATATCGTCGAGTGGTGGGGCGGAGAAGAAGCACGCCCGACACTT
GCTGACGTACAGGAACAGTACTTGCCAAGCGTTTTAGCGCAAGAGTCCGTCACTCCATAC
ATTGCAATGCTGAATGGAGAGCCGATTGGGTATGCCCAGTCGTACGTTGCTCTTGGAAGC
GGGGACGGATGGTGGGAAGAAGAAACCGATCCAGGAGTACGCGGAATAGACCAGTTACTG
GCGAATGCATCACAACTGGGCAAAGGCTTGGGAACCAAGCTGGTTCGAGCTCTGGTTGAG
TTGCTGTTCAATGATCCCGAGGTCACCAAGATCCAAACGGACCCGTCGCCGAGCAACTTG
CGAGCGATCCGATGCTACGAGAAAGCGGGGTTTGAGAGGCAAGGTACCGTAACCACCCCA
GATGGTCCAGCCGTGTACATGGTTCAAACACGCCAGGCATTCGAGCGAACACGCAGGTTT
GCCTAA
>aac(6')-Ib11_1_AY136758
ATGAAAAACACAATACATATCAACAGCAACGATTCCGTCACACTGCGCCTCATGACTGAG
CATGACCTTGCGATGCTCTATGAGTGGCTAAATCGATCTCATATCGTCGAGTGGTGGGGC
GGAGAAGAAGCACGCCCGACACTTGCTGACGTACAGGAACAGTACTTGCCAAGCGTTTTA
GCGCAAGAGTCCGTCACTCCATACATTGCAATGCTGAATGGAGAGCCGATTGGGTATGCC
CAGTCGTACGTTGCTCTTGGAAGCGGGGACGGATGGTGGGAAGAAGAAACCGATCCAGGA
GTACGCGGAATAGACCTGTCACTGGCGAATGCATCACAACTGGGCAAAGGCTTGGGAACC
AAGCTGGTTCGAGCTCTGGTTGAGTTGCTGTTCAATGATCCCGAGGTCACCAAGATCCAA
ACGGACCCGTCGCCGAGCAACTTGCGAGCGATCCGATGCTACGAGAAAGCGGGGTTTGAG
AGGCAAGGTACCGTAACCACCCCAGATGGTCCAGCCGTGTACATGGTTCAAACACGCCAG
GCATTCGAGCGAACACGCAGTGATGCCTAA
- "phenotypes.txt" file :
head phenotypes.txt
Gene_accession no. Class Phenotype PMID Mechanism of resistance Notes Required_gene
ant(2'')-Ia_1_X04555 Aminoglycoside Gentamicin, Tobramycin 3024112 Enzymatic modification Alternative name aadB
ant(2'')-Ia_10_HM367617 Aminoglycoside Gentamicin, Tobramycin 21873033 Enzymatic modification
ant(2'')-Ia_11_HM367620 Aminoglycoside Gentamicin, Tobramycin 21873033 Enzymatic modification
ant(2'')-Ia_12_HQ880250 Aminoglycoside Gentamicin, Tobramycin unpublished Enzymatic modification
ant(2'')-Ia_13_DQ176450 Aminoglycoside Gentamicin, Tobramycin 16304199 Enzymatic modification
ant(2'')-Ia_14_DQ266447 Aminoglycoside Gentamicin, Tobramycin unpublished Enzymatic modification
ant(2'')-Ia_15_EF205594 Aminoglycoside Gentamicin, Tobramycin unpublished Enzymatic modification
ant(2'')-Ia_16_HQ386848 Aminoglycoside Gentamicin, Tobramycin unpublished Enzymatic modification
ant(2'')-Ia_17_JTTZ01000034 Aminoglycoside Gentamicin, Tobramycin unpublished Enzymatic modification
- "phenotype_panels.txt":
head phenotype_panels.txt
:Panel: Campylobacter
Ampicillin
:Panel: Campylobacter jejuni
:Include: Campylobacter
Ciprofloxacin
Erythromycin
Gentamicin
Nalidixic acid
Streptomycin
Tetracycline
:Panel: Campylobacter coli
:Include: Campylobacter
Ciprofloxacin
Erythromycin
Gentamicin
Streptomycin
Tetracycline
Activate conda environment for ResFinder
conda activate resfinder
Go into the db_folder
cd db_resfinder
ls -lh
total 3.6M
-rw-rw-r-- 1 jeevka nn9305k 197K May 27 13:04 aminoglycoside.fsa
-rw-rw-r-- 1 jeevka nn9305k 2.8K May 27 13:04 antibiotic_classes.txt
-rw-rw-r-- 1 jeevka nn9305k 1.8M May 27 13:04 beta-lactam.fsa
-rwxrwxr-x 1 jeevka nn9305k 2.4K May 27 13:04 CHECK-entries.sh
-rw-rw-r-- 1 jeevka nn9305k 92K May 27 13:04 colistin.fsa
-rw-rw-r-- 1 jeevka nn9305k 912 May 27 13:04 config
-rw-rw-r-- 1 jeevka nn9305k 40K May 27 13:04 history.txt
-rwxrwxr-x 1 jeevka nn9305k 3.8K May 27 13:04 INSTALL.py
-rw-rw-r-- 1 jeevka nn9305k 173K May 27 13:04 macrolide.fsa
-rw-rw-r-- 1 jeevka nn9305k 6.9K May 27 13:04 nitroimidazole.fsa
-rw-rw-r-- 1 jeevka nn9305k 89K May 27 13:04 notes.txt
-rw-rw-r-- 1 jeevka nn9305k 45K May 27 13:04 oxazolidinone.fsa
-rw-rw-r-- 1 jeevka nn9305k 44K May 27 13:04 phenicol.fsa
-rw-rw-r-- 1 jeevka nn9305k 2.6K May 27 13:04 phenotype_panels.txt
-rw-rw-r-- 1 jeevka nn9305k 504K May 27 13:04 phenotypes.txt
-rw-rw-r-- 1 jeevka nn9305k 9.3K May 27 13:04 pseudomonicacid.fsa
-rw-rw-r-- 1 jeevka nn9305k 92K May 27 13:04 quinolone.fsa
-rw-rw-r-- 1 jeevka nn9305k 5.4K May 27 13:04 README.md
Make/Build DB
python3 INSTALL.py </path/to/kma_index> non_interactive
Input File: reads (.fastq) To ADD new schemes: Fasta file with allele sequences
Create a folder for new scheme
mkdir sareus
create/copy the necessary files insie the "sareus" folder
ls
saureus.txt
arcC.tfa
aroE.tfa
glpF.tfa
gmk.tfa
pta.tfa
tpi.tfa
yqiL.tfa
head -n 5 saureus.txt
ST arcC aroE glpF gmk pta tpi yqiL clonal_complex
1 1 1 1 1 1 1 1
2 2 2 2 2 2 2 26
3 1 1 1 9 1 1 12
4 10 10 8 6 10 3 2
Alle sequences
head -n 20 arcC.tfa
>arcC_1
TTATTAATCCAACAAGCTAAATCGAACAGTGACACAACGCCGGCAATGCCATTGGATACT
TGTGGTGCAATGTCACAGGGTATGATAGGCTATTGGTTGGAAACTGAAATCAATCGCATT
TTAACTGAAATGAATAGTGATAGAACTGTAGGCACAATCGTTACACGTGTGGAAGTAGAT
AAAGATGATCCACGATTCAATAACCCAACCAAACCAATTGGTCCTTTTTATACGAAAGAA
GAAGTTGAAGAATTACAAAAAGAACAGCCAGACTCAGTCTTTAAAGAAGATGCAGGACGT
GGTTATAGAAAAGTAGTTGCGTCACCACTACCTCAATCTATACTAGAACACCAGTTAATT
CGAACTTTAGCAGACGGTAAAAATATTGTCATTGCATGCGGTGGTGGCGGTATTCCAGTT
ATAAAAAAAGAAAATACCTATGAAGGTGTTGAAGCG
>arcC_2
TTATTAATCCAACAAGCTAAATCGAACAGTGACACAACGCCGGCAATGCCATTGGATACT
TGTGGTGCAATGTCACAAGGTATGATAGGCTATTGGTTGGAAACTGAAATCAATCGCATT
TTAACTGAAATGAATAGTGATAGAACTGTAGGCACAATCGTAACACGTGTGGAAGTAGAT
AAAGATGATCCACGATTTGATAACCCAACTAAACCAATTGGTCCTTTTTATACGAAAGAA
GAAGTTGAAGAATTACAAAAAGAACAGCCAGGCTCAGTCTTTAAAGAAGATGCAGGACGT
GGTTATAGAAAAGTAGTTGCGTCACCACTACCTCAATCTATACTAGAACACCAGTTAATT
CGAACTTTAGCAGACGGTAAAAATATTGTCATTGCATGCGGTGGTGGCGGTATTCCAGTT
ATAAAAAAAGAAAATACCTATGAAGGTGTTGAAGCG
Activate conda environment for MLST
Activate MLST conda environment
conda activate MLST
Adding the scheme
mlst/scripts/mlst-make_blast_db
Check if the scheme is added
mlst --longlist | grep SCHEME
Deactivate the conda environment
conda deactivate MLST