# wf-metagenomics usage

## Basic commands cheatsheet

- printing the directory you are currently in: `pwd`
- listing the contents of the directory you're in: `ls` (`ls -a` to also list hidden files)
- entering a directory: `cd <directory name>`
- going back one directory: `cd ..` (you can go back two, three or more directories with `../..`, `../../..`, etc)
- opening a file to browse its contents: `less <file name>`. Press `q` once you're done to exit. You can also browse a compressed .gz file without decompressing it by using `zless <file name>`.
- moving a file or directory: `mv <file name> <destination>`. You can also use this command to rename a file or directory: `mv <current name> <new name>`.
- copying a file: `cp <file name> <destination>`. You can copy a directory and its contents with `cp -r <directory name> <destination>`
- removing a file (/!\ THIS IS PERMANENT /!\ ): `rm <file name>`. To remove a directory with all its content: `rm -r <directory name>`.
- creating a directory: `mkdir <name of directory>`.

## Launching the wf-metagenomics pipeline

To launch a command with kraken2 (classification via kmer assignment), filtering for reads between 1300 and 1800 bp:

nextflow run epi2me-labs/wf-metagenomics --fastq <path to data> \
--kraken2 --out_dir <path to output directory> --min_len 1300 --max_len 1800

To launch a command with minimap2 (classification via alignement) with the same filtering:

nextflow run epi2me-labs/wf-metagenomics --fastq data/Elena_cleaned_data \
--out_dir wf-metagenomics_minimap2_filtered \
--min_len 1300 --max_len 1800 --kraken2 false --minimap2 true

Remarks:
- the data directory is supposed to contain one directory per barcode with the fastq files inside - remove any empty files, as they generate errors.
- the workflow will create two working directories, store_dir and work, in the directory you launch the command from.

## Full options list

Core options
  --fastq            [string]  A fastq file or directory containing fastq input files or directories of input files.
  --max_len          [integer] Specify read length upper limit [default: 2000]
  --min_len          [integer] Specify read length lower limit [default: 200]
  --out_dir          [string]  Directory for output of all user-facing files. [default: output]
  --report_name      [string]  Optional report suffix [default: report]
  --sample           [string]  A sample name for non-multiplexed data. Permissible if passing a file or directory of .fastq(.gz).
  --sample_sheet     [string]  CSV file with columns named `barcode`, `sample_name` and `type`. Permissible if passing a directory containing barcodeXX 
                               sub-directories. 
  --store_dir        [string]  Where to store initial download of taxonomy database.
  --source           [string]  Sets the default reference, databases and taxonomy used. Choices: ['TARGLOCI'] [default: ncbi_16s_18s]
  --taxonomy         [string]  Optionally override the taxonomy used [.tar.gz or Dir]

Minimap2 options
  --minimap2         [boolean] Enables classification via alignment
  --reference        [string]  Specifically override reference [.fna]
  --ref2taxid        [string]  Specifically override ref2taxid mapping. Format is .tsv (refname  taxid), no header row.
  --minimap2filter   [string]  Filter output of minimap2 by taxids inc. child nodes, E.g. "9606,1404"
  --minimap2exclude  [boolean] Invert minimap2filter and exclude the given taxids instead
  --split_prefix     [boolean] Enable if using a very large reference with minimap2

Kraken2 options
  --kraken2          [boolean] Enables classification via kmer-assignment [default: true]
  --database         [string]  Specifically override database [.tar.gz or Dir]
  --kraken2exclude   [boolean] Invert kraken2exclude and exclude the given taxids instead
  --kraken2minimap   [boolean] Run minimap2 only on reads classified by Kraken2 [default: true]
  --bracken_dist     [string]  Specifically override bracken kmer dist file
  --bracken_length   [integer] Set the length value bracken will use [default: 1000]
  --watch_path       [boolean] Specify if you want the workflow to continuously process input files as they become available in the input --fastq 
                               directories. 
  --read_limit       [integer] Use with watch_path if you don't want workflow to run indefinitely, will STOP pipeline when this number of reads have been 
                               processed., 
  --run_indefinitely [boolean] When used with watch_path will watch indefinitely if set to true [default: true]
  --batch_size       [integer] Number of fastq files to process in each batch [default: 1]
  --port             [integer] Port number for the kraken server to use [default: 8080]
  --bracken_level    [string]  Set the level that bracken will output at [default: S]

Generic options
  --threads          [integer] Set max number of threads to use. Note: limited by config executor cpus. [default: 8]
  --help             [boolean] Display help text.