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92 changes: 46 additions & 46 deletions README.md
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Expand Up @@ -8,13 +8,13 @@ pending. Please refer to the PATENTS file for additional inforamation
regarding licencing and use.


Reference
Reference
*http://jordan.biology.gatech.edu/page/software/stringmlst/*

Abstract
Abstract
*http://bioinformatics.oxfordjournals.org/content/early/2016/09/06/bioinformatics.btw586.short?rss=1*

Application Note
Application Note
*http://bioinformatics.oxfordjournals.org/content/early/2016/09/06/bioinformatics.btw586.full.pdf+html*

[![install with bioconda](https://img.shields.io/badge/install%20with-bioconda-brightgreen.svg?style=flat-square)](http://bioconda.github.io/recipes/stringmlst/README.html) [![PyPI version](https://badge.fury.io/py/stringMLST.svg)](https://badge.fury.io/py/stringMLST) ![downloads](https://img.shields.io/conda/dn/bioconda/stringmlst.svg?style=flat) [![container ready](https://quay.io/repository/biocontainers/stringmlst/status)](https://quay.io/repository/biocontainers/stringmlst)
Expand All @@ -26,8 +26,8 @@ keeping your databases updated, stringMLST can download and build databases from
and profile definitions. Please see the "Included databases and automated retrieval of databases from pubMLST" section
below for instructions. *The databases bundled here are for convenience only, do not rely on them being up-to-date*.

stringMLST is licensed and distributed under [CC Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)](https://creativecommons.org/licenses/by-nc-sa/4.0)
and is free for academic users and requires permission before any commercial use for any version of this code/algorithm.
stringMLST is licensed and distributed under [CC Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0)](https://creativecommons.org/licenses/by-nc-sa/4.0)
and is free for academic users and requires permission before any commercial use for any version of this code/algorithm.
If you are a commercial user, please contact [email protected] for permissions

## Recommended installation method
Expand All @@ -41,7 +41,7 @@ pip install stringMLST

```
git clone https://github.com/jordanlab/stringMLST
# Optional, download prebuilt databases
# Optional, download prebuilt databases
# We don't recommend this method, instead build the databases locally
cd stringMLST
git submodule init
Expand All @@ -50,13 +50,13 @@ git submodule update

## Quickstart guide

```bash
pip install stringMLST
mkdir -p stringMLST_analysis; cd stringMLST_analysis
stringMLST.py --getMLST -P neisseria/nmb --species neisseria
# Download all available databases with:
# stringMLST.py --getMLST -P mlst_dbs --species all
wget ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR026/ERR026529/ERR026529_1.fastq.gz ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR026/ERR026529/ERR026529_2.fastq.gz
```bash
pip install stringMLST
mkdir -p stringMLST_analysis; cd stringMLST_analysis
stringMLST.py --getMLST -P neisseria/nmb --species neisseria
# Download all available databases with:
# stringMLST.py --getMLST -P mlst_dbs --species all
wget ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR026/ERR026529/ERR026529_1.fastq.gz ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR026/ERR026529/ERR026529_2.fastq.gz
stringMLST.py --predict -P neisseria/nmb -1 ERR026529_1.fastq.gz -2 ERR026529_2.fastq.gz
Sample abcZ adk aroE fumC gdh pdhC pgm ST
ERR026529 231 180 306 612 269 277 260 10174
Expand All @@ -65,9 +65,9 @@ ERR026529 231 180 306 612 269 277 260 10174

## Python dependencies and external programs

stringMLST does not require any python dependencies for basic usage (Building databases and predicting STs).
stringMLST does not require any python dependencies for basic usage (Building databases and predicting STs).

For advanced used (genome coverage), stringMLST depends on the `pyfaidx` python module and `bamtools`, `bwa`, and `samtools`.
For advanced used (genome coverage), stringMLST depends on the `pyfaidx` python module and `bamtools`, `bwa`, and `samtools`.
See the coverage section for more information

stringMLST has been tested with:
Expand Down Expand Up @@ -139,18 +139,18 @@ profile datasets/Neisseria_spp/neisseria.txt
* Run stringMLST.py --buildDB to create DB. Choose a k value and prefix (optional).
```
stringMLST.py --buildDB -c databases/Neisseria_spp/config.txt -k 35 -P NM
stringMLST.py --buildDB -c databases/Neisseria_spp/config.txt -k 35 -P NM
```
### Predict:
#### Single sample :
```
stringMLST.py --predict -1 tests/fastqs/ERR026529_1.fastq -2 tests/fastqs/ERR026529_2.fastq -k 35 -P NM
stringMLST.py --predict -1 tests/fastqs/ERR026529_1.fastq -2 tests/fastqs/ERR026529_2.fastq -k 35 -P NM
```
#### Batch mode (all the samples together):
```
stringMLST.py --predict -d ./tests/fastqs/ -k 35 -P NM
stringMLST.py --predict -d ./tests/fastqs/ -k 35 -P NM
```
#### List mode:
Create a list file (list_paired.txt) as :
Expand Down Expand Up @@ -185,7 +185,7 @@ stringMLST's workflow is divided into two routines:
Readme for stringMLST
=============================================================================================
Usage
./stringMLST.py
./stringMLST.py
[--buildDB]
[--predict]
[-1 filename_fastq1][--fastq1 filename_fastq1]
Expand Down Expand Up @@ -217,7 +217,7 @@ There are two steps to predicting ST using stringMLST.
Synopsis:
stringMLST.py --buildDB -c <config file> -k <kmer length(optional)> -P <DB prefix(optional)>
config file : is a tab delimited file which has the information for typing scheme ie loci, its multifasta file and profile definition file.
Format :
Format :
[loci]
locus1 locusFile1
locus2 locusFile2
Expand All @@ -232,13 +232,13 @@ Required arguments
--buildDB
Identifier for build db module
-c,--config = <configuration file>
Config file in the format described above.
All the files follow the structure followed by pubmlst. Refer extended document for details.
Config file in the format described above.
All the files follow the structure followed by pubmlst. Refer extended document for details.

Optional arguments
Optional arguments
-k = <kmer length>
Kmer size for which the db has to be formed(Default k = 35). Note the tool works best with kmer length in between 35 and 66
for read lengths of 55 to 150 bp. Kmer size can be increased accordingly. It is advised to keep lower kmer sizes
for read lengths of 55 to 150 bp. Kmer size can be increased accordingly. It is advised to keep lower kmer sizes
if the quality of reads is not very good.
-P,--prefix = <prefix>
Prefix for db and log files to be created(Default = kmer). Also you can specify folder where you want the dbb to be created.
Expand All @@ -248,9 +248,9 @@ Optional arguments
Prints the help manual for this application

--------------------------------------------------------------------------------------------

2. stringMLST.py --predict

stringMLST --predict : can run in three modes
1) single sample (default mode)
2) batch mode : run stringMLST for all the samples in a folder (for a particular specie)
Expand All @@ -263,7 +263,7 @@ stringMLST.py --predict -1 <fastq file> -2 <fastq file> -d <directory location>
Required arguments
--predict
Identifier for predict miodule

Optional arguments
-1,--fastq1 = <fastq1_filename>
Path to first fastq file for paired end sample and path to the fastq file for single end file.
Expand All @@ -277,7 +277,7 @@ Optional arguments
Calculate seqence coverage for each allele. Turns on read generation (-r) and turns off fuzzy (-z 1)
Requires bwa, bamtools and samtools be in your path
-k = <kmer_length>
Kmer length for which the db was created(Default k = 35). Could be verified by looking at the name of the db file.
Kmer length for which the db was created(Default k = 35). Could be verified by looking at the name of the db file.
Could be used if the reads are of very bad quality or have a lot of N's.
-l,--list = <list_file>
LIST MODE : Location of list file and flag for list mode.
Expand Down Expand Up @@ -311,15 +311,15 @@ Optional arguments

3. stringMLST.py --getMLST

Synopsis:
Synopsis:
stringMLST.py --getMLST --species= <species> [-k kmer length] [-P DB prefix]

Required arguments
--getMLST
Identifier for getMLST module
--species= <species name>
Species name from the pubMLST schemes (use --schemes to get list of available schemes)
"all" will download and build all
Species name from the pubMLST schemes (use "--species show" to get list of available schemes)
"all" will download and build all

Optional arguments
-k = <kmer length>
Expand Down Expand Up @@ -355,27 +355,27 @@ NA10831_ATCACG_L002_R1_001.fastq.gz
#### Included databases and automated retrieval of databases from pubMLST
stringMLST includes all the pubMLST databases as of **February 15, 2017**, built with the default kmer (*35*). They can be found in the `datasets/` folder.
stringMLST includes all the pubMLST databases as of **February 15, 2017**, built with the default kmer (*35*). They can be found in the `datasets/` folder.
Simply unzip the databases you need and begin using stringMSLT as described below.
All the databases from pubMLST can be downloaded and prepared with your kmer choice
*Getting all pubMLST schemes*
*Getting all pubMLST schemes*
```
stringMLST.py --getMLST -P datasets/ --species all
```
Individual databases from pubMLST can also be downloaded as needed, using the scheme identifiers
*Downloading a scheme*
*Downloading a scheme*
```
# List available schemes
stringMLST.py --getMLST --schemes

# Download the Neisseria spp. scheme

stringMLST.py --getMLST -P datasets/nmb --species neisseria
stringMLST.py --getMLST -P datasets/nmb --species Neisseria

```
Expand All @@ -393,7 +393,7 @@ E.g.:
Species of interest: Neisseria spp.
Corresponding definition file: http://pubmlst.org/data/profiles/neisseria.txt
Corresponding multi fasta locus files:
Corresponding multi fasta locus files:
http://pubmlst.org/data/alleles/neisseria/abcZ.tfa
http://pubmlst.org/data/alleles/neisseria/adk.tfa
http://pubmlst.org/data/alleles/neisseria/aroE.tfa
Expand Down Expand Up @@ -425,7 +425,7 @@ TTTGATACCGTTGCGAA...
TTTGATACCGTTGCCAA...
```
These files can be obtained from PubMLST/BIGSdb or can be create by the user themselves.
These files can be obtained from PubMLST/BIGSdb or can be create by the user themselves.
In either case, an accompanying configuration file is also required to describe the profile definition and allele sequence files. An example configuration file is shown below:
```
Expand All @@ -444,7 +444,7 @@ profile /data/home/stringMLST/pubmlst/Neisseria_sp/neisseria.txt
This file is pre-packed on stringMLSTs website and can easily be created by the user for custom database.
#### Database Building
#### Database Building
The next step is for database building is running the buildDB module to create the database files. buildDB module requires the user to specify the config file. The default k-mer size is 35 but can be changed using the -k option. Specifying the prefix for the created database files is optional but is recommended.
The choice of k-mer depends on the size of the sequencing read. In general, the value of k can never be greater than the read length. The application has been tested on a number of read lengths ranging from 55 to 150 bps using k-mer sizes of 21 to 66. In our testing, the k-mer size does not affect the accuracy of the read length. A smaller k-mer size will increase the runtime and a larger k-mer size will increase the file size. The user should ideally pick a k-mer with a length around half of the average read length. For lower quality data, it also advised to choose smaller k-mer values to reduce false hits.
Expand All @@ -463,16 +463,16 @@ This command will produce 3 database files and a log file. The log file is used
For the example above, the following files will be created:
NM_35.txt, NM_weight.txt and NM_profile.txt
Please note that in the prediction routine the database is identified with the prefix.
Please note that in the prediction routine the database is identified with the prefix.
ST discovery routine
As discussed earlier, StringMLST has 3 running modes
As discussed earlier, StringMLST has 3 running modes
* Single sample mode - for running stringMLST on a single sample
* Batch mode - for running stringMLST on all the FASTQ files present in a directory
* List mode - for running stringMLST on all the FASTQ files provided in a list file
#### Single sample mode:
This is the default mode for stringMLST and takes in one sample at a time. The sample can be single-end or paired-end. The sample has to be in FASTQ format. In order to run, the user should know the prefix of the database created and the k-mer size.
#### Single sample mode:
This is the default mode for stringMLST and takes in one sample at a time. The sample can be single-end or paired-end. The sample has to be in FASTQ format. In order to run, the user should know the prefix of the database created and the k-mer size.
By default, the tool expects paired-end samples.
```
Expand All @@ -482,7 +482,7 @@ stringMLST.py --predict -1 <paired-end file 1> -2 <paired-end file 2> -p --prefi
```
stringMLST.py --predict -1 <single-end file> -s --prefix <prefix for the database> -k <k-mer size> -o <output file name>
```
#### Batch Mode:
#### Batch Mode:
This mode can be used for processing multiple files with one command. All the samples will be queried against the same database. Also all samples should be in the same directory. All the samples will be treated either as single-end or paired-end. The paired-end samples should be differentiated with the character _1 and _2 at the end (E.g.: sampleX_1.fastq and sampleX_2.fastq).
*Paired-end samples:*
Expand All @@ -494,7 +494,7 @@ stringMLST.py --predict -d <directory for samples> -p --prefix <prefix for the d
```
stringMLST.py --predict -d <directory for samples> -s --prefix <prefix for the database> -k <k-mer size> -o <output file name>
```
#### List Mode:
#### List Mode:
This mode could be used if user has samples at different locations or if the paired-end samples are not stored in traditional way. All the samples will be queried against the same database. All the samples will be treated either as single-end or paired-end. This mode requires the user to provide a list file which has the list of all samples along with the location. Each line in the list file represents a new sample.
A sample list file for single-end sample looks like the following.
```
Expand Down Expand Up @@ -533,7 +533,7 @@ stringMLST provides two, complimentary methods for determining confidence in an
stringMLST determines an allele based on its kmer support; the more kmers seen for allele 1, the more likely that allele 1 is the allele present in the genome. Unlike SRST2 and other mapping/BLAST based tools, stringMLST always infers an ST, using the maximimally supported allele (allele with most kmer hits). The difference between the maximum support (the reported allele) and the second support (next closest allele) can be informative for low coverage reads. The `-z|--fuzzy` threshold (Default = 300), assigns significance to the difference between supports. Much like SRST2 and Torsten Seemann's popular [pubMLST script](https://github.com/tseemann/mlst), stringMLST reports potentially new or closely supported alleles in allele* syntax. For high coverage reads, we suggest a fuzzy threshold >500. For low coverage reads, a fuzzy threshold of <50.
Coverage mode requires `bedtools`, `bwa`, and `samtools` in your PATH and an additional python module, `pyfaidx` (See the dependencies section for installion information). Coverage mode by default disables display of fuzzy alleles in favor of sequence coverage information made by mapping potential reads to the putative allele sequence. In our testing, coverage mode slightly increases prediction time (<1 sec increase per sample).
Coverage mode requires `bedtools`, `bwa`, and `samtools` in your PATH and an additional python module, `pyfaidx` (See the dependencies section for installion information). Coverage mode by default disables display of fuzzy alleles in favor of sequence coverage information made by mapping potential reads to the putative allele sequence. In our testing, coverage mode slightly increases prediction time (<1 sec increase per sample).
**Please note:** stringMLST *always* infers the ST from the reads, fuzzy matches and/or <100% coverage do not necessarily mean a new allele has been found.
Expand All @@ -546,7 +546,7 @@ stringMLST.py --predict -1 <paired-end file 1> -2 <paired-end file 2> -p --prefi
stringMLST.py --predict -1 <paired-end file 1> -2 <paired-end file 2> -p --prefix <prefix for the database> -k <k-mer size> -r -o <output file name>- -f 50
```
#### Other Examples :
#### Other Examples :
*Reporting time along with the output.*
```
Expand Down
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Expand Up @@ -1186,7 +1186,7 @@ def checkParams(buildDB, predict, config, k, listMode, list, batch, dir, fastq1,
--getMLST
Identifier for getMLST module
--species= <species name>
Species name from the pubMLST schemes (use --schemes to get list of available schemes)
Species name from the pubMLST schemes (use "--species show" to get list of available schemes)
"all" will download and build all
Optional arguments
-k = <kmer length>
Expand Down

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