An RNA virus strain-level identification tool for short reads.
E-mail: heruiliao2-c@my.cityu.edu.hk
- Old Version - V1.14: Fix some bugs but lack virstrain_contig and virstrain_merge.
Click here to check the log of all updates
- V1.12: VirStrain is able to take gzipped FASTQs as input now!
- Fix one bug of the perl script about head name problem.
- Update a new parameter '-s' that allows sorting the most possible strain by matches to the sites.
- The web server extension of VirStrain - StrainDetect (https://strain.ee.cityu.edu.hk) is online now!
- V1.13: Fix a database generation bug in V1.12 of bioconda version!
- A new function that allows comprehensive (including 45619 strains of 28 viral species) viral strain identification for assembled contigs is available!
- V1.14: Fix a bug (about handling gzipped FASTQs) in V1.13!
- Tem_Vs files are named randomly (only GitHub version) and links for pre-built databases are provided.
- V1.17: All the changes made so far have been updated in both GitHub and Conda.
- Python >=3.6 (3.7.3 is recommanded and 3.9 is not supprted now!)
- Perl
- Required python package: networkx==2.4, numpy==1.17.3, pandas==1.0.1, biopython==1.74, Plotly==3.10.0
- Bowtie2 (for virstrain version >= V1.17)
(If you have installed conda, then you can run sh install_package.sh to install all required packages automatically.)
Make sure these programs have been installed before using VirStrain. (However, if you use bioconda/pip to install VirStrain, ignore this.)
The first way to install VirStrain, is to use bioconda. Once you have bioconda environment installed, install package virstrain:
conda install -c bioconda virstrain
The second way to install VirStrain, is to use pip:
pip install virstrain==1.17
It should be noted that some commands have been replaced if you install VirStrain using bioconda/pip. (See below)
| Command (Not bioconda/pip) | Command (bioconda/pip) |
|---|---|
| python VirStrain.py -h | virstrain -h |
| python VirStrain_build.py -h | virstrain_build -h |
| python VirStrain_contig.py -h | virstrain_contig -h |
| python VirStrain_contigDB_merge.py -h | virstrain_merge -h |
Or you can install VirStrain mannually (Make sure all dependencies have been installed before this step).
git clone https://github.com/liaoherui/VirStrain.git
cd VirStrain
chmod 755 bin/jellyfish-linux
rm VirStrain_DB.tar.gz
Then, you can download the reference database of 3 RNA viruses used in the paper.
There are three ways to download the reference database.
-> Method-1:
Run:
cd VirStrain
sh download.sh
- -> Method-2:
Run:
cd VirStrain
wget -qO- "https://figshare.com/ndownloader/files/34002479" | tar -zx
Or, download the database from figshare mannually, and then extract it using the commandtar -zxvf.
If all failed, please email to the author to get the database.
- The databases of two DNA viruses (HBV and HCMV) used in the paper can be downloaded now!
sh download_dna.sh - Besides, a larger database with more SARS-CoV-2 strains (see Supplementary Section 1.1 in the paper) can also be downloaded now.
sh download_scov2_big.sh
You can also build the VirStrain database with your own genomes, the mannual is written in Usage section.
In the event that the download scripts fail to retrieve the pre-built database, we also provide Google drive inks to access all pre-built databases. The table below offers information about the public pre-built databases. Users can download these databases and use them to identify viral strains directly.
| Name | Description | Download link |
|---|---|---|
| VirStrain_DB.tar.gz | Databases containing SCOV2, H1N1, and HIV viral strains used in the paper | Google drive |
| SCOV2_newBig.tar.gz | Databases containing more SCOV2 viral strains used in the paper | Google drive |
| VirStrain_DNA_DB.tar.gz | Databases containing two DNA viral (HBV and HCMV) strains used in the paper | Google drive |
| VirStrain_contig_DB.tar.gz | Contig-level database | Google drive |
It should be noted if you install VirStrain using bioconda/pip, you should replace the commands. (see below)
| Command (Not bioconda/pip) | Command (bioconda/pip) |
|---|---|
| python VirStrain.py -h | virstrain -h |
| python VirStrain_build.py -h | virstrain_build -h |
| python VirStrain_contig.py -h | virstrain_contig -h |
| python VirStrain_contigDB_merge.py -h | virstrain_merge -h |
For SE reads:
python VirStrain.py -i Test_Data/MT451123_1.fq -d VirStrain_DB/SCOV2 -o MT451123_SE_Test
For PE reads:
python VirStrain.py -i Test_Data/MT451123_1.fq -p Test_Data/MT451123_2.fq -d VirStrain_DB/SCOV2 -o MT451123_PE_Test
When the virus has high mutation rate, like HIV, you may need to add -m parameter.
For HIV:
SE reads: python VirStrain.py -i <Read1> -d VirStrain_DB/HIV -o <Output_dir> -m
PE reads: python VirStrain.py -i <Read1> -p <Read2> -d VirStrain_DB/HIV -o <Output_dir> -m
python VirStrain_contig.py -i <Input_Contig_fasta> -d VirStrain_contig_DB -o VirStrain_Contig_Res
You can use the command below to download the pre-built comprehensive viral strain database for contig identification:
sh download_contig_db.sh
If you want to convert pre-built VirStrain databases for reads (e.g. VirStrain_DB/SCOV2 and VirStrain_DB/H1N1) to database for contigs. Then you can try the command below:
python VirStrain_contigDB_merge.py -i VirStrain_DB/SCOV2,VirStrain_DB/H1N1 -o VirStrain_contig_DB_merge
python VirStrain_build.py -i <Input_MSA> -d <Database_Dir>
Important note: "," and "|" are not allowed in your <Input_MSA>. For example, ">Strain_A, 2022" or ">Strain_A|2022" is not allowed but ">Strain_A_2022" is allowed.
For small-scale strains (<1000 input strains) or viruses with large genome sizes (like HCMV), you can use manual-covering function to cover more useful sites. For example, in our experiment, we used "-s 0.4" for 328 HCMV strains. Usually, 0.2~0.6 shoule be a suitable range for the parameter "-s". However, if you only have very few strains, like 3 strains, you can also use a greater value like "-s 0.8".
python VirStrain_build.py -i <Input_MSA> -d <Database_Dir> -s 0.4
Besides, if you only want to use SNV sites from "x" to "y" (eg. x=500 to y=1000), then you can add the parameter -r.
python VirStrain_build.py -i <Input_MSA> -d <Database_Dir> -s 0.4 -r 500-1000
Note: The format of input MSA should be same as the format of MSA generated by Mafft (https://mafft.cbrc.jp/alignment/software/).
Identification - VirStrain.py (Default k-mer size: 25)
VirStrain - An RNA virus strain-level identification tool for short reads.
Example: python VirStrain.py -i Test_Data/MT451123_1.fq -p Test_Data/MT451123_2.fq -d VirStrain_DB/SCOV2 -o MT451123_PE_Test
required arguments:
-i, --input_reads Input fastq data.
-d, --database_dir Path of VirStrain database.
optional arguments:
-h, --help Show help message and exit.
-o, --output_dir The output directory. (Default: ./VirStrain_Out)
-p, --input_reads2 Input fastq data for PE reads
-c, --site_filter_cutoff The cutoff of filtering one site when calculate the Vscore. (Default: 0.05)
-s, --rank_by_sites If set to 1, then VirStrain will sort the most possible strain by matches to the sites. (default: 0)
-f, --turn_off_figures If set to 1, then VirStrain will not generate figures. (default: 0)
-m, --high_mutation_virus If the virus has high mutation rate (like HIV), use this option. (Default: off)
Build database - VirStrain_build.py (Default k-mer size: 25)
VirStrain - An RNA virus strain-level identification tool for short reads.
Example: python VirStrain_build.py -i <Input_MSA> -d <Database_Dir>
required arguments:
-i, --input_msa Input MSA file (Must have same format to msa generated by mafft).
optional arguments:
-d, --database_dir The output directory of constructed database. (Default: ./VirStrain_DB)
-c, --dash_cutoff The cutoff of dash in each column of MSA. (Default: 0)
-s, --sites_cutoff The cutoff of sites number for manual-covering function. (eg. 1 means all useful sites will be use and 0.8 means 80% useful sites will be used)
-r, --sites_rcutoff The cutoff of sites range for covering algorithm (eg. 3-500 means the covering algorithm will only consider the SNV sites from 3-500 of MSA.)
The output of VirStrain contains two files. The first is a report file in text format. This file contains all identified strains and their depth and site coverage, etc. The other file is an interactive HTML page to display the depth and uniqueness of sites.
You can check the output file in the folder "MT451123_Sim_PE" in this repository.
The picture below displays an output example of a simulated data (Truth: MT451123.1).
Explaination about the four headers in the output of VirStrain
| Header | Description |
|---|---|
| Most Possible strain* | The most possible strain in the sequencing data detected by VirStrain. (The strains with highest Vscore in the first iteraition.) |
| Other Possible strains* | The other possible strain in the sequencing data detected by VirStrain. (The strains with highest Vscore in the later iteraition, 10 mutation number can be a strong evidence for other possible strains according to our experiment result.) |
| Highest Map Strains | The strain with maximum "Covered SNV site/Total SNV site" in the first iteration. For user's reference. |
| Top 10 Score Strains | The top10 strain sorted by Vscore in the first iteration. For user's reference, and also could be useful information to detect those low abundance strains which are highly similar to the high abundance strain (Eg, only one mutation number). |
（Note: the header with * means the content following this header includes the main identification result.）
Explaination about the columns in the output of VirStrain:
| Column_name | Description |
|---|---|
| Strain_ID | The NCBI (or other public database) accession number of identified strain. |
| Cls_info | The cluster information of identified strain, eg: Cluster2830_2 -> belong Cluster2830, size=2. |
| SubCls_info | The sub-cluster information of identified strain. |
| Vscore | The Vscore generated by VirStrain algorithm. |
| Total_Map_Rate | The covered sites out of total sites in the first iteration of VirStrain. |
| Valid_Map_Rate | The covered sites out of total sites in the remaining iteration of VirStrain. |
| Strain_depth | The sequencing depth of identified strain predicted by VirStrain. |
| Strain_info | The metadata of identified strains, such as region information and subtype, etc. |
| SNV_freq | The SNV frequency of all sites. |
how to cite this tool:
Liao, H., Cai, D. & Sun, Y. VirStrain: a strain identification tool for RNA viruses. Genome Biol 23, 38 (2022). https://doi.org/10.1186/s13059-022-02609-x