PUMA (Pipeline for Unifying Microbiome Analysis) (Version 1.2)

Pre-print: https://doi.org/10.1101/482380

Frontiers in Microbiology Paper: https://www.frontiersin.org/articles/10.3389/fmicb.2020.584699/full

• The Manual provided below will provide setup instructions as well as important tutorials for students/researchers for jumpstarting the analysis process with the various tools shown in the schema above.
  • Manual: https://ucla.app.box.com/s/ybehndj8phvj8eooqpcwxbqaxc3iaif2

How to install: (MacOSx and Linux supported)

• If you are on Windows you can run the program by installing the Linux subsystem as shown here: https://www.onmsft.com/how-to/how-to-install-windows-10s-linux-subsystem-on-your-pc or you can use a VM of your choice.

1. Install Conda for your operating system (https://docs.conda.io/projects/conda/en/latest/user-guide/install/)
   • for mac download the .pkg file
2. Run git clone https://github.com/keithgmitchell/PUMA.git by opening terminal in the folder of your choice.
3. conda config --append channels conda-forge; conda config --append channels bioconda
4. cd PUMA
5. conda create -n puma_env --file requirements.txt
   • Be sure to select "Yes" by clicking "y" when prompted.
   • This may take 5-10 mins.
6. conda activate puma_env
   • You should now see (puma_env) at the start of your terminal instead of (base)
7. pip install -r pip_requirements.txt; mkdir temp
8. For the CLI and the GUI run python PumaCLI.py --help and python RunPuma.py respectively.
9. A window will pop in your default browser, if it says "This site can not be reached" click refresh.

How to update your PUMA version:

1. Navigate to the directory where you ran part 2 of the installation process.
   • For example:cd /Users/keithmitchell/Desktop/Repositories/PUMA
   • You should see files such as pip_requirements.txt, RunPuma.py, etc.
2. Run git pull
3. Run steps 1, 3-8 from "How to Install"

How to run:

Results from running the PUMA scripts will produce output in the directory ‘output/’, or a specified output directory, with a prefix for the time the job was completed YEAR_MONTH_DAY_HOUR_MINUTE as well as a descriptive suffix such as ‘functional_profile’, ‘anacapa’, ‘MrDNA’, or ‘QIIME 2’ such as that shown in the Protocol sections. In addition; the CLI version provides the option for a user to specify an additional unique string to enable automation and parallel processing.

Be sure all sample names in the metadata and in the ASV/OTU table are matching an are only alphanumeric titles.

• Running MrDNA example data: (using files from examples/mrdna_F15) (rarefaction depth and iterations randomly chosen)

  OR

  python PumaCLI.py -type MrDNA -metadata examples/mrdna_F15/mrdna_F15_mock_metadata.tsv \
  -otutable examples/mrdna_F15/112415KR515F-pr.fasta.otus.fa.OTU.txt \
  -seqs examples/mrdna_F15/sequences.fasta -rarefactioniter 4 -rarefactiondepth 2000 -msa_phylo False
  

  • The output will look something like this where the top .zip file is the most recent job ran and check the log to make sure the job finished ok:

• Running Anacapa Data: (using files in examples/anacapa_skirball_S18) (rarefaction depth and iterations randomly chosen)

  

  OR

  python PumaCLI.py -otutable examples/anacapa_skirball_S18/16S_ASV_raw_taxonomy_70.tsv \
  -fwdseq examples/anacapa_skirball_S18/nochim_forward16S.fasta \
  -mergeseq examples/anacapa_skirball_S18/nochim_merged16S.fasta \
  -reverseseq examples/anacapa_skirball_S18/nochim_reverse16S.fasta \
  -rarefactioniter 0 -rarefactiondepth 0 -metadata examples/anacapa_skirball_S18/anacapa_skirball_metadata_3_11_18.tsv \
  -type anacapa -unique_id demo -msa_phylo False
  

  • The output will look something like this where the top .zip file is the most recent job ran and check the log to make sure the job finished ok:

• Running QIIME2 data: (rarefaction depth and iterations randomly chosen)

  python PumaCLI.py -type QIIME2 -metadata examples/qiime2_demo/qiime2_moving_pictures_metadata.tsv \
  -otutable examples/qiime2_demo/table_moving_pictures.qza \
  -taxonomy examples/qiime2_demo/taxonomy_moving_pictures.qza \
  -seqs examples/qiime2_demo/rep-seqs.qza \ 
  -rarefactioniter 4 -rarefactiondepth 2000 -msa_phylo False
  

  • QIIME2 Output view for example files:

• Running the Functional Profile following input of file sets to piphillin (https://piphillin.secondgenome.com/):

    python functional_profile.py \
    -i examples/piphillin/Keith_20180723210057.tar,examples/piphillin/Keith_20180723214258.tar \
    -o UCLA -metadata examples/anacapa_skirball_metadata_S18/anacapa_skirball_metadata_3_11_18.tsv      
  

Version 1.3 (In progress)

• Automated Rarefaction Depth Option
• Capability to select output files (rather then produce all mentioned)
• Include more tools such as the iTOL tool and any other requested (please create Issue if you have one in mind)
• Include more file input types from other sequencing services (please create Issue if you have one in mind)
• Fix the anacapa input functions.
• Add picrust 2.0
• Add merge files option that includes the metadata.
• upload standard otu and seqs
• add rarefaction_depth/iter_ to the file names
• double check that no rarefaction option still works
• fix background process
• finish piphillin view and logging

NEAR FUTURE: