How to analyze microbiomes in QIIME?

Install QIIME 2

• Begin by installing QIIME 2, the latest version is recommended for up-to-date features. Follow the installation guide from the official QIIME 2 website, ensuring you meet all system requirements.

 

• QIIME 2 is primarily designed for Unix-based systems, so ensure you have access to such an environment. If using Windows, consider using a virtual machine or Docker installation.

Set Up Your Environment

• Activate your QIIME 2 conda environment with the command: conda activate qiime2-.

 

• Confirm that QIIME 2 is working by running: qiime --help. This should display the list of available commands, indicating a successful setup.

Import Your Data

• Ensure your sequencing data is in the appropriate format. QIIME 2 supports various formats, including FASTQ for demographic data.

 

• Use the QIIME 2 import command to convert your data into a .qza artifact: qiime tools import --type 'SampleData[SequencesWithQuality]' --input-path --output-path demux.qza.

Visualize Raw Data

• Use qiime demux summarize to generate a visualization of your demultiplexed sequences: qiime demux summarize --i-data demux.qza --o-visualization demux.qzv.

 

• View the results using QIIME’s visualization tools or the QIIME 2 View platform online.

Quality Control and Feature Table Construction

• Use DADA2 or Deblur to perform quality filtering and denoising to create a feature table and representative sequences file. For DADA2: qiime dada2 denoise-single --i-demultiplexed-seqs demux.qza --p-trim-left 0 --p-trunc-len 150 --o-representative-sequences rep-seqs.qza --o-table table.qza.

 

• Explore your feature table, use: qiime feature-table summarize --i-table table.qza --o-visualization table.qzv and for representative sequences: qiime feature-table tabulate-seqs --i-data rep-seqs.qza --o-visualization rep-seqs.qzv.

Taxonomic Analysis

• Classify the sequences taxonomically, generally using a pretrained classifier: qiime feature-classifier classify-sklearn --i-classifier --i-reads rep-seqs.qza --o-classification taxonomy.qza.

 

• Generate a visualization of the taxonomy table with: qiime metadata tabulate --m-input-file taxonomy.qza --o-visualization taxonomy.qzv.

Diversity Analysis

• Initiate core-metrics phylogenetic analysis to assess alpha and beta diversity: qiime diversity core-metrics-phylogenetic --i-phylogeny --i-table table.qza --p-sampling-depth --m-metadata-file --output-dir core-metrics-results.

 

• Explore results, such as the PCoA plots and alpha diversity, through the visualizations available within your core-metrics results.

Export Data

• Export processed data (tables, sequences, etc.) for external analysis. For instance, export a feature table by: qiime tools export --input-path table.qza --output-path exported-feature-table.

 

• Convert QIIME-produced files into formats compatible with other bioinformatics tools if needed, using the appropriate conversion commands.