EbolaSeq

EbolaSeq is a command-line tool that simplifies the process of analyzing Ebola virus sequences. It automates the complete workflow from downloading sequences to creating phylogenetic trees. The tool retrieves Ebola virus sequences from NCBI GenBank, processes them according to user specifications, performs multiple sequence alignment and generates phylogenetic trees.

Installation

1. First, install conda if you haven't already:

   wget https://repo.anaconda.com/miniconda/Miniconda3-latest-Linux-x86_64.sh
   bash Miniconda3-latest-Linux-x86_64.sh

2. Create and activate a new conda environment:

   conda create -n ebola_env python=3.9
   conda activate ebola_env

3. Install required tools in the environment:

   conda install -c bioconda mafft trimal iqtree

4. Install ebolaseq:

   git clone https://github.com/DaanJansen94/ebolaseq.git
   cd ebolaseq
   pip install .

Re-installation

When updates are pushed to GitHub, or when you want to use your own modifications to the code, you'll need to reinstall the package:

conda activate ebola_env  # Make sure you're in the right environment
cd ebolaseq
git pull  # Get the latest updates from GitHub
pip uninstall ebolaseq
pip install .

Note: Any time you modify the code or pull updates from GitHub, you need to reinstall the package using these commands for the changes to take effect.

Usage

First, make sure your conda environment is activated:

conda activate ebola_env

EbolaSeq can be run in two modes:

1. Interactive Mode (default) - for interactive use
2. Non-Interactive Mode - for HPC submissions or automated runs

Interactive Mode

Basic command structure:

ebolaseq --output-dir my_analysis [optional arguments]

Example commands:

# Basic usage
ebolaseq --output-dir my_analysis

# With phylogenetic analysis
ebolaseq --output-dir my_analysis --phylogeny

# Complete analysis with sequence removal and consensus
ebolaseq --output-dir my_analysis \
         --consensus-file path/to/consensus.fasta \
         --remove remove.txt \
         --phylogeny

The tool will interactively prompt you for choices about:

• Virus species
• Genome completeness
• Host type
• Metadata options

Non-Interactive Mode (HPC)

For HPC submissions or automated runs, specify all parameters via command line:

ebolaseq --output-dir my_analysis \
         --virus 1 \
         --genome 2 \
         --completeness 80 \
         --host 1 \
         --metadata 3 \
         --beast 2 \
         --phylogeny \
         --consensus-file path/to/consensus.fasta \
         --remove remove.txt

Required parameters for non-interactive mode:

• --virus: Virus type

  • 1 = Zaire ebolavirus
  • 2 = Sudan ebolavirus
  • 3 = Bundibugyo ebolavirus
  • 4 = Tai Forest ebolavirus
  • 5 = Reston ebolavirus

• --genome: Genome completeness

  • 1 = Complete genomes only
  • 2 = Partial genomes (requires --completeness)
  • 3 = All genomes

• --completeness: Required when --genome=2

  • Value between 1-100 (percentage)

• --host: Host filter

  • 1 = Human only
  • 2 = Non-human only
  • 3 = All hosts

• --metadata: Metadata filter

  • 1 = Location only
  • 2 = Date only
  • 3 = Both location and date
  • 4 = None

• --beast: Required when --metadata is 2 or 3

  • 1 = No
  • 2 = Yes

Optional arguments (both modes):

• --output-dir: Output directory (required)
• --consensus-file: Path to consensus FASTA file
• --remove: Path to sequence removal list
• --phylogeny: Create phylogenetic tree

Input File Formats

1. Consensus file (optional):

   • FASTA format
   • Example:

   >Consensus_sequence_name
   ATGCATGCATGC...
   

2. Remove file (optional):

   • Text file with one GenBank accession number per line
   • Example:

   KM034562.1
   KM034563.1
   MK114118.1
   

Important Notes

It is strongly recommended to always use a remove.txt file with the --remove option when running EbolaSeq. This file should contain sequence IDs that should be excluded from the analysis, particularly sequences obtained from cell culture passages, laboratory-adapted strains, artificially modified sequences, sequences from experimental infections, and other non-natural viral sequences. These sequences can bias analyses as they may not represent natural viral diversity.

When creating large phylogenetic trees for Zaire ebolavirus, it is recommended to root the tree using sequences from the 1976 Yambuku outbreak, as this represents the first documented outbreak of the virus.

Dependencies

• Python ≥ 3.6
• BioPython ≥ 1.79
• NumPy ≥ 1.21.0
• MAFFT
• TrimAl
• IQTree2

Citation

If you use EbolaSeq in your research, please cite:

Jansen, D., & Vercauteren, K. (2025). EbolaSeq: A Command-Line Tool for Downloading, Processing, and Analyzing Ebola Virus Sequences for Phylogenetic Analysis (Version v0.1.1) [Computer software]. https://doi.org/10.5281/zenodo.14851804

License

This project is licensed under the GNU General Public License v3.0 (GPL-3.0) - see the LICENSE file for details.

Contributing

Contributions are welcome! Please feel free to submit a Pull Request.

Support

If you encounter any problems or have questions, please open an issue on GitHub.