For Bracken news, updates, and instructions: ccb.jhu.edu/software/bracken/

Bracken's peer-reviewed paper (published Jan 2, 2017): https://peerj.com/articles/cs-104/

Kraken can be downloaded from here: http://ccb.jhu.edu/software/kraken/

Three steps are necessary to set up Kraken abundance estimation. It is assumed that ${KRAKEN_DB} is the path to a built Kraken database, and all sequences are as .fna files in the ${KRAKEN_DB}/library directory.

kraken --db=${KRAKEN_DB} --fasta-input --threads=10 <( find -L library -name "*.fna" -o -name "*.fa" -o -name "*.fasta" -exec cat {} + )  > database.kraken

With ${READ_LENGTH} = 75:

perl count-kmer-abundances.pl --db=${KRAKEN_DB} --read-length=75 database.kraken  > database75mers.kraken_cnts

Set ${READ_LENGTH} to whatever is the most likely perfect read length of your data. E.g., if you are using 100 bp reads, set it to 100.

The kmer distribution file is generated using the following command line:

python generate_kmer_distribution.py -i database75mers.kraken_cnts -o KMER_DISTR.TXT

Given the expected kmer distribution for genomes in a kraken database along with a kraken report file, the number of reads belonging to each species (or genus) is estimated using the estimate_abundance.py file, run with the following command line:

python estimate_abundance.py -i <KRAKEN.REPORT> -k <KMER_DISTR.TXT> -l <CLASSIFICATON_LEVEL> -t <THRESHOLD> -o <OUTPUT_FILE.TXT>

The following required parameters must be specified:

• <KRAKEN.REPORT> - the kraken report generated for a given dataset
• <KMER_DISTR.TXT> - the file generated by generate_kmer_distribution.py
• <OUTPUT_FILE.TXT> - the desired name of the output file to be generated by the code

The following optional parameters may be specified:

• <CLASSIFICATION_LEVEL> - Default = 'S'. This specifies that abundance estimation will calculate estimated reads for each species. Other possible options are K (kingdom level), P (phylum), C (class), O (order), F (family), and G (genus).
• <THRESHOLD> - Default = 10. For species classification, any species with <= 10 (or otherwise specified) reads will not receive any additional reads from higher taxonomy levels when distributing reads for abundance estimation. If another classification level is specified, thresholding will occur at that level.

The following sample input and output files are included in the sample_data/ folder: sample_test.report - Kraken report file generated from the kraken-report command. sample_kmer_distr_75mers.txt - example kmer distribution file generated by generate_kmer_distribution.py sample_output_species_abundance.txt - Bracken species abundance estimation for sample_test.report sample_output_bracken.report - Kraken report style file with all reads redistributed to the species level

Due to size constraints, the following files are not included in the sample_data/ folder: sample_test.kraken - Kraken output file used to generate the Kraken report file database.kraken - Initial Kraken classification of every genome database75mers.kraken_cnts - Counting of kmer abundances

The following commands were used to generate each individual file:

1.  kraken --db${KRAKEN_DB} --threads=10 sample.fa > sample_test.kraken
    kraken-report --db=${KRAKEN_DB} sample_test.kraken > sample_test.report 
   

2.  kraken --db=${KRAKEN_DB} --fasta_input --threads=10 <( find -L library -name "*.fna" -o -name "*.fa" -o -name "*.fasta" -exec cat {} + ) > database.kraken 
    perl count-kmer-abubndances.pl --db=${KRAKEN_DB} --read-length=75 database.kraken > database75mers.kraken_cnts
   

3.  python generate_kmer_distribution.py -i database75mers.kraken_cnts -o sample_kmer_distr_75mers.txt
   

4.  python estimate_abundance.py -i sample_test.report -k sample_kmer_distr_75mers.txt -l S -t 10 -o sample_output_species_abundance.txt 
   

Copyright (C) 2016 Jennifer Lu, [email protected]

Bracken is free software; you can redistribute it and/or modify it under the terms of the GNU General Public License as published by the Free Software Foundation; either version 3 of the license, or (at your option) any later version.

This program is distributed in the hope that it will be useful, but WITHOUT ANY WARRANTY; without even the implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the GNU General Public License for more details.

You should have received a copy of the GNU General Public License along with this program; if not, see http://www.gnu.org/licenses/.

Jennifer Lu ([email protected], ccb.jhu.edu/people/jennifer.lu)

Florian Breitwieser ([email protected], ccb.jhu.edu/people/florian)

Last Updated On: 02/06/2017