zAMPExplorer

zAMPExplorer is an interactive Shiny application designed to facilitate the downstream analysis of outputs generated by the zAMP pipeline . zAMP itself is a comprehensive DADA2-based bioinformatics pipeline for processing 16S rRNA gene amplicon metagenomic sequences, offering convenient, reproducible, and scalable analysis from raw sequencing reads to taxonomic profiles. The output of zAMP is a phyloseq object, which serves as the input for zAMPExplorer.

A typical phyloseq object contains the following components:

## phyloseq-class experiment-level object
## otu_table()   OTU Table:         [ 5 taxa and 6 samples ]
## sample_data() Sample Data:       [ 6 samples by 4 sample variables ]
## tax_table()   Taxonomy Table:    [ 5 taxa by 7 taxonomic ranks ]
## phy_tree()    Phylogenetic Tree: [ 5 tips and 4 internal nodes ]
## refseq()      DNAStringSet:      [ 5 reference sequences ]

For more information on the phyloseq package, please visit the official phyloseq documentation.

zAMPExplorer enables users to perform a wide range of microbiota and statistical analyses, including compositional barplot, relative abundance heatmap, community diversity (alpha diversity), community similarity through unsupervised (NMDS/PCoA) and supervised (RDA) ordinations, and community typing (or clustering) of microbial profiles using Dirichlet Multinomial Mixtures (DMM). All of these analyses are made accessible through an intuitive graphical interface, bridging the gap between complex command-line bioinformatics processing and user-friendly data exploration.

Prerequisites and installation

Prerequisites

Operating system: Windows, macOS, or Linux
R: Version 4.0 or later
RStudio: Recommended for running the Shiny app
zAMP: zAMPExplorer is designed to work with output generated from the zAMP pipeline.

Installation

Clone the repository:

git clone git@github.com:metagenlab/zAMPExplorer.git

Navigate to the project directory:
```
cd zAMPExplorer
```

Install conda environment: Create a conda environment with the necessary r-packages:

conda env create --name zAMPexplorer --file zAMPexplorer.yaml
conda activate zAMPexplorer
./install_additional_rPackages.sh

Launch the app: After installation, launch zAMPExplorer with:
```
shiny::runApp("path_to_your_app")
```

Overview of the interface

zAMPExplorer is divided into several tabs, each dedicated to a specific type of analysis:

Upload Data: Upload your phyloseq object here. This object is the output of the zAMP pipeline and serves as the input for all downstream analyses in the app.
Reads and Taxa Overview: Visualize the total number of reads per sample and the estimated number of organisms in each sample at different taxonomic levels.
Compositional Barplot: Create interactive barplots to visualize the relative abundance of taxa within your samples.
Heatmap: Generate heatmaps to visualize the relative abundance of taxa across samples and sample groups.
Alpha Diversity: Analyze and visualize alpha diversity metrics, comparing diversity within groups.
Beta Diversity: Explore beta diversity using different distance matrices to assess similarities/differences in microbial communities between samples.
Shared Core Taxa (Venn Diagram): Identify and visualize core taxa shared across different groups.
Community Typing (DMM): Perform community typing using Dirichlet Multinomial Mixture models to infer the optimal number of community types inside the dataset.
RDA Plot: Perform redundancy analysis (RDA) to explore the association between your samples and explanatory variables.

Troubleshooting and FAQ

Common issues

Issue: App fails to launch.
- Solution: Ensure all dependencies are installed by running source("Dependencies.R").
Issue: Plots are not displaying correctly.
- Solution: Verify your R version is 4.0 or later and that all necessary libraries are installed.

FAQ

Q: What file types can I upload?
- A: zAMPExplorer supports .rds files containing phyloseq objects.
Q: How do I update the app with new features?
- A: Pull the latest updates from the GitHub repository, or fork the project and make changes to your fork.

References

The following R packages are integral to the functionality of zAMP Explorer. We highly recommend consulting their respective documentation:

shiny: Shiny Web Application Framework for R
shinydashboard: Create Dashboards with 'Shiny'
hinyWidgets: Custom Inputs Widgets for Shiny
webshot2: Taking Screenshots of Web Pages
htmlwidgets: HTML Widgets for R
dplyr: A grammar of data manipulation
phyloseq: Handling and Analysis of High-Throughput Microbial Community Data
DT: An R interface to the JavaScript library DataTables
ggplot2: Create Elegant Data Visualisations Using the Grammar of Graphics
ggpubr: ggpubr: ‘ggplot2’ Based Publication Ready Plots
plotly: Create Interactive Web Graphics via 'plotly.js'
vegan: vegan: an R package for community ecologists
RColorBrewer: ColorBrewer Palettes
ComplexHeatmap: Make Complex Heatmaps
InteractiveComplexHeatmap: Make Interactive Complex Heatmaps in R
vegan: Community Ecology Package
microViz: Tools to make microbial community data more accessible
microbiome: Microbiome Analytics
MicEco: Various functions for analysis for microbial community data
metagMisc: Miscellaneous functions for metabarcoding and metagenomic analysis
mia: mia - Microbiome analysis
DirichletMultinomial: Dirichlet-Multinomial Mixture Model Machine Learning for Microbiome Data

For detailed information about each package, visit the provided links.

Contributing

Thank you for using zAMPExplorer. We hope it helps you gain deeper insights into your microbiome data (16S amplicons). Please feel free to contribute, suggest features, or report any issues you encounter.

Acknowledgments

We would like to thank the developers of the R packages and tools integrated into zAMPExplorer. Please make sure to acknowledge their contributions in any publications or projects using this tool.