Version Compatibility

Reference examples tested with: Bracken 2.9+, Kraken2 2.1+, MetaPhlAn 4.1+, pandas 2.2+

Before using code patterns, verify installed versions match. If versions differ:

• Python: pip show <package> then help(module.function) to check signatures
• CLI: <tool> --version then <tool> --help to confirm flags

If code throws ImportError, AttributeError, or TypeError, introspect the installed package and adapt the example to match the actual API rather than retrying.

Abundance Estimation with Bracken

"Get species-level abundances from my Kraken2 results" → Redistribute reads assigned to higher taxonomic levels down to species using Bracken's Bayesian re-estimation for more accurate abundance profiles.

• CLI: bracken -d db -i kraken2.report -o bracken.output -r 150 -l S

Basic Abundance Estimation

bash
# Run Bracken on Kraken2 report
bracken -d /path/to/kraken2_db \
    -i kraken_report.txt \
    -o bracken_output.txt \
    -r 150 \                       # Read length (100, 150, 200, 250, 300)
    -l S                           # Taxonomic level

Full Workflow with Kraken2

bash
# Step 1: Classify with Kraken2
kraken2 --db /path/to/kraken2_db \
    --threads 8 \
    --paired \
    --report sample_kraken_report.txt \
    reads_R1.fastq.gz reads_R2.fastq.gz

# Step 2: Estimate abundances with Bracken
bracken -d /path/to/kraken2_db \
    -i sample_kraken_report.txt \
    -o sample_bracken_species.txt \
    -w sample_bracken_report.txt \
    -r 150 \
    -l S

Different Taxonomic Levels

bash
# Species level (default)
bracken -d db -i report.txt -o species.txt -r 150 -l S

# Genus level
bracken -d db -i report.txt -o genus.txt -r 150 -l G

# Family level
bracken -d db -i report.txt -o family.txt -r 150 -l F

# Phylum level
bracken -d db -i report.txt -o phylum.txt -r 150 -l P

Build Bracken Database

bash
# Build Bracken database for specific read lengths
# Run AFTER building Kraken2 database
bracken-build -d /path/to/kraken2_db -t 8 -l 150

# Build for multiple read lengths
bracken-build -d /path/to/kraken2_db -t 8 -l 100
bracken-build -d /path/to/kraken2_db -t 8 -l 250

Output Format

name                    taxonomy_id    taxonomy_lvl    kraken_assigned_reads    added_reads    new_est_reads    fraction_total_reads
Escherichia coli        562           S               5234                     1245           6479             0.52
Staphylococcus aureus   1280          S               2156                     456            2612             0.21

Filter Low-Abundance Taxa

bash
# Use threshold for minimum reads
bracken -d db \
    -i report.txt \
    -o bracken.txt \
    -r 150 \
    -l S \
    -t 10                          # Minimum reads threshold

Combine Multiple Samples

bash
# Run Bracken on each sample
for report in kraken_reports/*.txt; do
    sample=$(basename $report _kraken_report.txt)
    bracken -d db -i $report -o bracken/${sample}_species.txt -r 150 -l S
done

# Combine into abundance matrix
combine_bracken_outputs.py --files bracken/*_species.txt -o combined_abundance.txt

Parse Bracken Output in Python

python
import pandas as pd

bracken = pd.read_csv('bracken_output.txt', sep='\t')

bracken_sorted = bracken.sort_values('new_est_reads', ascending=False)
bracken_sorted[['name', 'fraction_total_reads']].head(20)

total_reads = bracken['new_est_reads'].sum()
bracken['relative_abundance'] = bracken['new_est_reads'] / total_reads * 100

Convert to Relative Abundance

python
import pandas as pd

df = pd.read_csv('bracken_output.txt', sep='\t')

total = df['new_est_reads'].sum()
df['relative_abundance'] = df['new_est_reads'] / total * 100

df.to_csv('bracken_relative_abundance.txt', sep='\t', index=False)

Create Abundance Matrix

Goal: Merge per-sample Bracken outputs into a single species-by-sample abundance matrix for downstream statistical analysis.

Approach: Load each Bracken output, extract species names and read counts, iteratively outer-merge on species name, and fill missing values with zero.

python
import pandas as pd
import os

files = [f for f in os.listdir('bracken') if f.endswith('_species.txt')]

dfs = []
for f in files:
    sample = f.replace('_species.txt', '')
    df = pd.read_csv(f'bracken/{f}', sep='\t')
    df = df[['name', 'new_est_reads']].rename(columns={'new_est_reads': sample})
    dfs.append(df)

merged = dfs[0]
for df in dfs[1:]:
    merged = merged.merge(df, on='name', how='outer')

merged = merged.fillna(0)
merged.to_csv('abundance_matrix.txt', sep='\t', index=False)

Key Parameters

| Parameter | Description | |-----------|-------------| | -d | Kraken2 database path | | -i | Input Kraken2 report | | -o | Output abundance file | | -w | Output updated report (optional) | | -r | Read length used | | -l | Taxonomic level | | -t | Minimum read threshold |

Taxonomic Levels

| Level | Code | Description | |-------|------|-------------| | Kingdom | K | Bacteria, Archaea | | Phylum | P | Major divisions | | Class | C | Class level | | Order | O | Order level | | Family | F | Family level | | Genus | G | Genus level | | Species | S | Species level |

Read Length Options

Pre-built databases typically include: 50, 75, 100, 150, 200, 250, 300 bp

Choose the length closest to your actual read length.

Related Skills

• kraken-classification - Generate Kraken2 report
• metaphlan-profiling - Alternative profiling method
• metagenome-visualization - Visualize abundances