MaAsLin2 User Manual

MaAsLin2 is the next generation of MaAsLin (Microbiome Multivariable Association with Linear Models).

MaAsLin2 is comprehensive R package for efficiently determining multivariable association between clinical metadata and microbial meta-omics features. MaAsLin2 relies on general linear models to accommodate most modern epidemiological study designs, including cross-sectional and longitudinal, along with a variety of filtering, normalization, and transform methods.

If you use the MaAsLin2 software, please cite our manuscript:

Mallick H, Rahnavard A, McIver LJ, Ma S, Zhang Y, Nguyen LH, Tickle TL, Weingart G, Ren B, Schwager EH, Chatterjee S, Thompson KN, Wilkinson JE, Subramanian A, Lu Y, Waldron L, Paulson JN, Franzosa EA, Bravo HC, Huttenhower C (2021). Multivariable Association Discovery in Population-scale Meta-omics Studies. PLoS Computational Biology, 17(11):e1009442.

Check out the MaAsLin 2 tutorial for an overview of analysis options.

If you have questions, please direct it to :
MaAsLin2 Forum
Google Groups (Read only)

Description
Requirements
Installation
How to Run
Visualization
Troubleshooting

Description

MaAsLin2 finds associations between microbiome multi-omics features and complex metadata in population-scale epidemiological studies. The software includes multiple analysis methods (with support for multiple covariates and repeated measures), filtering, normalization, and transform options to customize analysis for your specific study.

Requirements

MaAsLin2 is an R package that can be run on the command line or as an R function.

Installation

MaAsLin2 can be run from the command line or as an R function. If only running from the command line, you do not need to install the MaAsLin2 package but you will need to install the MaAsLin2 dependencies.

From command line

Download the source: MaAsLin2.tar.gz
Decompress the download:
- $ tar xzvf maaslin2.tar.gz
Install the Bioconductor dependencies edgeR and metagenomeSeq.
Install the CRAN dependencies:
- $ R -q -e "install.packages(c('lmerTest','pbapply','car','dplyr','vegan','chemometrics','ggplot2','pheatmap','hash','logging','data.table','glmmTMB','MASS','cplm','pscl'), repos='http://cran.r-project.org')"
Install the MaAsLin2 package (only r,equired if running as an R function):
- $ R CMD INSTALL maaslin2

From R

Install Bioconductor and then install Maaslin2

if(!requireNamespace("BiocManager", quietly = TRUE))
    install.packages("BiocManager")
BiocManager::install("Maaslin2")

How to Run

MaAsLin2 can be run from the command line or as an R function. Both methods require the same arguments, have the same options, and use the same default settings.

Input Files

MaAsLin2 requires two input files.

Data (or features) file
- This file is tab-delimited.
- Formatted with features as columns and samples as rows.
- The transpose of this format is also okay.
- Possible features in this file include taxonomy or genes.
Metadata file
- This file is tab-delimited.
- Formatted with features as columns and samples as rows.
- The transpose of this format is also okay.
- Possible metadata in this file include gender or age.

The data file can contain samples not included in the metadata file (along with the reverse case). For both cases, those samples not included in both files will be removed from the analysis. Also the samples do not need to be in the same order in the two files.

NOTE: If running MaAsLin2 as a function, the data and metadata inputs can be of type data.frame instead of a path to a file.

Output Files

MaAsLin2 generates two types of output files: data and visualization.

Data output files
- all_results.tsv
  - This includes the same data as the data.frame returned.
  - This file contains all results ordered by increasing q-value.
  - The first columns are the metadata and feature names.
  - The next two columns are the value and coefficient from the model.
  - The next column is the standard deviation from the model.
  - The N column is the total number of data points.
  - The N.not.zero column is the total of non-zero data points.
  - The pvalue from the calculation is the second to last column.
  - The qvalue is computed with p.adjust with the correction method.
- significant_results.tsv
  - This file is a subset of the results in the first file.
  - It only includes associations with q-values <= to the threshold.
- ``features```
  - This folder includes the filtered, normalized, and transformed versions of the input feature table.
  - These steps are performed sequentially in the above order.
  - If an option is set such that a step does not change the data, the resulting table will still be output.
- models.rds
  - This file contains a list with every model fit object.
  - It will only be generated if save_models is set to TRUE.
- residuals.rds
  - This file contains a data frame with residuals for each feature.
- fitted.rds
  - This file contains a data frame with fitted values for each feature.
- ranef.rds
  - This file contains a data frame with extracted random effects for each feature (when random effects are specified).
- maaslin2.log
  - This file contains all log information for the run.
  - It includes all settings, warnings, errors, and steps run.
Visualization output files
- heatmap.pdf
  - This file contains a heatmap of the significant associations.
- [a-z/0-9]+.pdf
  - A plot is generated for each significant association.
  - Scatter plots are used for continuous metadata.
  - Box plots are for categorical data.
  - Data points plotted are after filtering but prior to normalization and transform.

Run a Demo

Example input files can be found in the inst/extdata folder of the MaAsLin2 source. The files provided were generated from the HMP2 data which can be downloaded from https://ibdmdb.org/ .

HMP2_taxonomy.tsv: is a tab-demilited file with species as columns and samples as rows. It is a subset of the taxonomy file so it just includes the species abundances for all samples.

HMP2_metadata.tsv: is a tab-delimited file with samples as rows and metadata as columns. It is a subset of the metadata file so that it just includes some of the fields.

Command line

$ Maaslin2.R --fixed_effects="diagnosis,dysbiosisnonIBD,dysbiosisUC,dysbiosisCD,antibiotics,age" --random_effects="site,subject" --standardize=FALSE inst/extdata/HMP2_taxonomy.tsv inst/extdata/HMP2_metadata.tsv demo_output

Make sure to provide the full path to the MaAsLin2 executable (ie ./R/Maaslin2.R).
In the demo command:
- HMP2_taxonomy.tsv is the path to your data (or features) file
- HMP2_metadata.tsv is the path to your metadata file
- demo_output is the path to the folder to write the output

In R

library(Maaslin2)
input_data <- system.file(
    'extdata','HMP2_taxonomy.tsv', package="Maaslin2")
input_metadata <-system.file(
    'extdata','HMP2_metadata.tsv', package="Maaslin2")
fit_data <- Maaslin2(
    input_data, input_metadata, 'demo_output',
    fixed_effects = c('diagnosis', 'dysbiosisnonIBD','dysbiosisUC','dysbiosisCD', 'antibiotics', 'age'),
    random_effects = c('site', 'subject'),
    reference = "diagnosis,nonIBD",
    standardize = FALSE)

## [1] "Creating output folder"
## [1] "Creating output feature tables folder"
## [1] "Creating output fits folder"
## [1] "Creating output figures folder"
## 2024-12-08 06:04:04.080822 INFO::Writing function arguments to log file
## 2024-12-08 06:04:04.152485 INFO::Verifying options selected are valid
## 2024-12-08 06:04:04.171702 INFO::Determining format of input files
## 2024-12-08 06:04:04.17236 INFO::Input format is data samples as rows and metadata samples as rows
## 2024-12-08 06:04:04.175415 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2024-12-08 06:04:04.176117 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2024-12-08 06:04:04.177676 INFO::Filter data based on min abundance and min prevalence
## 2024-12-08 06:04:04.17816 INFO::Total samples in data: 1595
## 2024-12-08 06:04:04.178634 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2024-12-08 06:04:04.182158 INFO::Total filtered features: 0
## 2024-12-08 06:04:04.182755 INFO::Filtered feature names from abundance and prevalence filtering:
## 2024-12-08 06:04:04.189105 INFO::Total filtered features with variance filtering: 0
## 2024-12-08 06:04:04.189621 INFO::Filtered feature names from variance filtering:
## 2024-12-08 06:04:04.190057 INFO::Running selected normalization method: TSS
## 2024-12-08 06:04:04.866048 INFO::Bypass z-score application to metadata
## 2024-12-08 06:04:04.866816 INFO::Running selected transform method: LOG
## 2024-12-08 06:04:04.888018 INFO::Running selected analysis method: LM
## 2024-12-08 06:04:05.105823 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:05.293129 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2024-12-08 06:04:05.398059 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2024-12-08 06:04:05.483388 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:05.5731 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2024-12-08 06:04:05.668071 INFO::Fitting model to feature number 6, Bacteroides.caccae

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:05.758931 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2024-12-08 06:04:05.844251 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2024-12-08 06:04:05.940553 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2024-12-08 06:04:06.02947 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2024-12-08 06:04:06.123834 INFO::Fitting model to feature number 11, Bacteroides.finegoldii

## boundary (singular) fit: see help('isSingular')

## Feature Bacteroides.finegoldii : simpleWarning: Model failed to converge with 1 negative eigenvalue: -7.9e+01

## 2024-12-08 06:04:06.195387 WARNING::Fitting problem for feature 11 a warning was issued

## boundary (singular) fit: see help('isSingular')

## Warning: Model failed to converge with 1 negative eigenvalue: -7.9e+01

## 2024-12-08 06:04:06.287549 INFO::Fitting model to feature number 12, Bacteroides.fragilis

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:06.37321 INFO::Fitting model to feature number 13, Bacteroides.intestinalis

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:06.466493 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2024-12-08 06:04:06.55157 INFO::Fitting model to feature number 15, Bacteroides.ovatus

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:06.639743 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2024-12-08 06:04:06.731694 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2024-12-08 06:04:06.819537 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:06.911068 INFO::Fitting model to feature number 19, Bacteroides.uniformis

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:07.002567 INFO::Fitting model to feature number 20, Bacteroides.vulgatus

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:07.093303 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:07.200639 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2024-12-08 06:04:07.286243 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2024-12-08 06:04:07.379616 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2024-12-08 06:04:07.472187 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2024-12-08 06:04:07.558687 INFO::Fitting model to feature number 26, Parabacteroides.distasonis

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:07.650259 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2024-12-08 06:04:07.740467 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2024-12-08 06:04:07.834924 INFO::Fitting model to feature number 29, Parabacteroides.unclassified

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:07.921202 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2024-12-08 06:04:08.161872 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2024-12-08 06:04:08.247298 INFO::Fitting model to feature number 32, Prevotella.copri

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:08.333148 INFO::Fitting model to feature number 33, Alistipes.finegoldii

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:08.42439 INFO::Fitting model to feature number 34, Alistipes.onderdonkii

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:08.520751 INFO::Fitting model to feature number 35, Alistipes.putredinis

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:08.607279 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2024-12-08 06:04:08.700096 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2024-12-08 06:04:08.781926 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2024-12-08 06:04:08.870313 INFO::Fitting model to feature number 39, Clostridium.bolteae

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:08.964871 INFO::Fitting model to feature number 40, Clostridium.citroniae

## boundary (singular) fit: see help('isSingular')

## Feature Clostridium.citroniae : simpleWarning: Model failed to converge with 1 negative eigenvalue: -3.4e+01

## 2024-12-08 06:04:09.036912 WARNING::Fitting problem for feature 40 a warning was issued

## boundary (singular) fit: see help('isSingular')

## Warning: Model failed to converge with 1 negative eigenvalue: -3.4e+01

## 2024-12-08 06:04:09.123967 INFO::Fitting model to feature number 41, Clostridium.clostridioforme

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:09.211601 INFO::Fitting model to feature number 42, Clostridium.hathewayi

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:09.295343 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2024-12-08 06:04:09.388608 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2024-12-08 06:04:09.472803 INFO::Fitting model to feature number 45, Clostridium.symbiosum

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:09.561099 INFO::Fitting model to feature number 46, Flavonifractor.plautii

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:09.648723 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2024-12-08 06:04:09.733659 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2024-12-08 06:04:09.830934 INFO::Fitting model to feature number 49, Eubacterium.rectale

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:09.923065 INFO::Fitting model to feature number 50, Eubacterium.siraeum

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:10.008614 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:10.099245 INFO::Fitting model to feature number 52, Eubacterium.ventriosum

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:10.180737 INFO::Fitting model to feature number 53, Ruminococcus.gnavus

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:10.269124 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2024-12-08 06:04:10.357439 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2024-12-08 06:04:10.441222 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2024-12-08 06:04:10.52782 INFO::Fitting model to feature number 57, Dorea.longicatena

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:10.619113 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:10.707329 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:10.794102 INFO::Fitting model to feature number 60, Roseburia.hominis

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:10.88793 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2024-12-08 06:04:10.968578 INFO::Fitting model to feature number 62, Roseburia.inulinivorans

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:11.058704 INFO::Fitting model to feature number 63, Roseburia.unclassified

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:11.302229 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2024-12-08 06:04:11.394147 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2024-12-08 06:04:11.478266 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2024-12-08 06:04:11.56966 INFO::Fitting model to feature number 67, Ruminococcus.bromii

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:11.650274 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2024-12-08 06:04:11.732665 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2024-12-08 06:04:11.824384 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:11.90955 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2024-12-08 06:04:11.992009 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2024-12-08 06:04:12.081607 INFO::Fitting model to feature number 73, Dialister.invisus

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:12.16855 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2024-12-08 06:04:12.254385 INFO::Fitting model to feature number 75, Veillonella.dispar

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:12.339756 INFO::Fitting model to feature number 76, Veillonella.parvula

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:12.426182 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2024-12-08 06:04:12.507569 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2024-12-08 06:04:12.594775 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2024-12-08 06:04:12.682339 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2024-12-08 06:04:12.768436 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2024-12-08 06:04:12.859633 INFO::Fitting model to feature number 82, Escherichia.coli
## 2024-12-08 06:04:12.950848 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2024-12-08 06:04:13.030759 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:13.1127 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae

## boundary (singular) fit: see help('isSingular')

## 2024-12-08 06:04:13.203073 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2024-12-08 06:04:13.289149 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2024-12-08 06:04:13.404193 INFO::Counting total values for each feature
## 2024-12-08 06:04:13.424069 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2024-12-08 06:04:13.48676 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2024-12-08 06:04:13.551138 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2024-12-08 06:04:13.651009 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2024-12-08 06:04:13.689199 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2024-12-08 06:04:13.711196 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2024-12-08 06:04:13.714557 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2024-12-08 06:04:13.717698 INFO::Writing the significant results (those which are less than or equal to the threshold of 0.250000 ) to file (ordered by increasing q-values): demo_output/significant_results.tsv
## 2024-12-08 06:04:13.719209 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2024-12-08 06:04:13.848234 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2024-12-08 06:04:13.857774 INFO::Plotting associations from most to least significant, grouped by metadata
## 2024-12-08 06:04:13.858423 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2024-12-08 06:04:13.859426 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii

## 2024-12-08 06:04:14.062478 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified

## 2024-12-08 06:04:14.233313 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis

## 2024-12-08 06:04:14.39086 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale

## 2024-12-08 06:04:14.551359 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus

## 2024-12-08 06:04:14.708833 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus

## 2024-12-08 06:04:14.866173 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum

## 2024-12-08 06:04:15.024126 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified

## 2024-12-08 06:04:15.208557 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans

## 2024-12-08 06:04:15.369286 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis

## 2024-12-08 06:04:15.523135 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron

## 2024-12-08 06:04:15.681941 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis

## 2024-12-08 06:04:15.841041 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis

## 2024-12-08 06:04:15.999295 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei

## 2024-12-08 06:04:16.159753 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii

## 2024-12-08 06:04:16.319176 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens

## 2024-12-08 06:04:16.478464 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum

## 2024-12-08 06:04:16.63989 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena

## 2024-12-08 06:04:16.799633 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae

## 2024-12-08 06:04:16.961216 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-12-08 06:04:17.12237 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli

## 2024-12-08 06:04:17.283292 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae

## 2024-12-08 06:04:17.446231 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified

## 2024-12-08 06:04:17.599322 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii

## 2024-12-08 06:04:17.757869 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens

## 2024-12-08 06:04:17.915264 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris

## 2024-12-08 06:04:18.076288 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes

## 2024-12-08 06:04:18.234466 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis

## 2024-12-08 06:04:18.39373 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii

## 2024-12-08 06:04:18.569078 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii

## 2024-12-08 06:04:18.733501 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis

## 2024-12-08 06:04:18.898829 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara

## 2024-12-08 06:04:19.060204 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum

## 2024-12-08 06:04:19.218543 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae

## 2024-12-08 06:04:19.379874 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified

## 2024-12-08 06:04:19.542867 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens

## 2024-12-08 06:04:19.700614 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus

## 2024-12-08 06:04:19.864489 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme

## 2024-12-08 06:04:20.026462 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii

## 2024-12-08 06:04:20.187512 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii

## 2024-12-08 06:04:20.344742 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum

## 2024-12-08 06:04:20.511977 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris

## 2024-12-08 06:04:20.673584 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47

## 2024-12-08 06:04:20.833188 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus

## 2024-12-08 06:04:20.99607 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii

## 2024-12-08 06:04:21.159909 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis

## 2024-12-08 06:04:21.330935 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified

## 2024-12-08 06:04:21.494482 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques

## 2024-12-08 06:04:21.651519 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified

## 2024-12-08 06:04:21.838459 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus

## 2024-12-08 06:04:22.004665 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis

## 2024-12-08 06:04:22.171128 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified

## 2024-12-08 06:04:22.340457 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified

## 2024-12-08 06:04:22.508791 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis

## 2024-12-08 06:04:22.672657 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis

## 2024-12-08 06:04:22.83845 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31

## 2024-12-08 06:04:23.001582 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae

## 2024-12-08 06:04:23.16248 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8

## 2024-12-08 06:04:23.323772 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae

## 2024-12-08 06:04:23.489931 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii

## 2024-12-08 06:04:23.654612 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii

## 2024-12-08 06:04:23.823113 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum

## 2024-12-08 06:04:23.990185 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA

## 2024-12-08 06:04:26.089681 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2024-12-08 06:04:26.091005 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified

## 2024-12-08 06:04:26.243251 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii

## 2024-12-08 06:04:26.423058 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae

## 2024-12-08 06:04:26.603817 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified

## 2024-12-08 06:04:26.780637 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis

## 2024-12-08 06:04:26.959771 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum

## 2024-12-08 06:04:27.139431 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus

## 2024-12-08 06:04:27.310566 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis

## 2024-12-08 06:04:27.486798 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii

## 2024-12-08 06:04:27.659386 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale

## 2024-12-08 06:04:27.831241 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis

## 2024-12-08 06:04:28.008078 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis

## 2024-12-08 06:04:28.182044 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-12-08 06:04:28.36529 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii

## 2024-12-08 06:04:28.54734 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens

## 2024-12-08 06:04:28.72665 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris

## 2024-12-08 06:04:28.904795 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii

## 2024-12-08 06:04:29.245696 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis

## 2024-12-08 06:04:29.396168 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum

## 2024-12-08 06:04:29.56679 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus

## 2024-12-08 06:04:29.714018 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii

## 2024-12-08 06:04:29.868718 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii

## 2024-12-08 06:04:30.017291 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae

## 2024-12-08 06:04:30.175797 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus

## 2024-12-08 06:04:30.328268 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum

## 2024-12-08 06:04:30.4756 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae

## 2024-12-08 06:04:30.63127 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae

## 2024-12-08 06:04:30.775442 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum

## 2024-12-08 06:04:30.926345 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis

## 2024-12-08 06:04:31.075618 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii

## 2024-12-08 06:04:31.229136 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques

## 2024-12-08 06:04:31.372074 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8

## 2024-12-08 06:04:31.532619 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum

## 2024-12-08 06:04:31.678467 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum

## 2024-12-08 06:04:31.837637 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis

## 2024-12-08 06:04:31.983651 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens

## 2024-12-08 06:04:32.138806 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi

## 2024-12-08 06:04:32.289051 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified

## 2024-12-08 06:04:32.443467 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens

## 2024-12-08 06:04:32.593784 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus

## 2024-12-08 06:04:32.75072 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae

## 2024-12-08 06:04:32.904026 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus

## 2024-12-08 06:04:33.06397 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris

## 2024-12-08 06:04:33.214265 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47

## 2024-12-08 06:04:35.114054 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2024-12-08 06:04:35.115358 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii

## 2024-12-08 06:04:35.248615 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale

## 2024-12-08 06:04:35.400082 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31

## 2024-12-08 06:04:35.557539 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis

## 2024-12-08 06:04:35.701167 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified

## 2024-12-08 06:04:35.85785 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-12-08 06:04:36.012219 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques

## 2024-12-08 06:04:36.193055 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum

## 2024-12-08 06:04:36.350654 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus

## 2024-12-08 06:04:36.513284 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli

## 2024-12-08 06:04:36.664548 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum

## 2024-12-08 06:04:36.829287 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified

## 2024-12-08 06:04:36.987818 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis

## 2024-12-08 06:04:37.147347 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens

## 2024-12-08 06:04:37.310285 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae

## 2024-12-08 06:04:37.464411 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii

## 2024-12-08 06:04:37.621057 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus

## 2024-12-08 06:04:37.769621 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified

## 2024-12-08 06:04:37.937604 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis

## 2024-12-08 06:04:38.084822 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii

## 2024-12-08 06:04:38.247301 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans

## 2024-12-08 06:04:38.399263 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii

## 2024-12-08 06:04:38.566092 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified

## 2024-12-08 06:04:38.71907 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii

## 2024-12-08 06:04:38.880055 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis

## 2024-12-08 06:04:39.033219 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri

## 2024-12-08 06:04:39.199463 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei

## 2024-12-08 06:04:39.346815 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis

## 2024-12-08 06:04:39.512787 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii

## 2024-12-08 06:04:41.445546 INFO::Plotting data for metadata number 4, antibiotics
## 2024-12-08 06:04:41.446851 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans

## 2024-12-08 06:04:41.56689 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis

## 2024-12-08 06:04:41.720957 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale

## 2024-12-08 06:04:41.863678 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus

## 2024-12-08 06:04:42.022512 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis

## 2024-12-08 06:04:42.16864 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena

## 2024-12-08 06:04:42.324722 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus

## 2024-12-08 06:04:42.468579 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii

## 2024-12-08 06:04:42.618391 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum

## 2024-12-08 06:04:42.758672 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae

## 2024-12-08 06:04:42.916259 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis

## 2024-12-08 06:04:43.053736 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii

## 2024-12-08 06:04:43.260068 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii

## 2024-12-08 06:04:43.412119 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified

## 2024-12-08 06:04:43.558838 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum

## 2024-12-08 06:04:43.714282 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-12-08 06:04:43.859161 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii

## 2024-12-08 06:04:44.014107 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31

## 2024-12-08 06:04:44.15636 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis

## 2024-12-08 06:04:44.304046 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii

## 2024-12-08 06:04:44.452844 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis

## 2024-12-08 06:04:44.603881 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens

## 2024-12-08 06:04:44.74901 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens

## 2024-12-08 06:04:44.894398 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron

## 2024-12-08 06:04:45.048878 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii

## 2024-12-08 06:04:45.197223 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae

## 2024-12-08 06:04:45.348608 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum

## 2024-12-08 06:04:45.494513 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques

## 2024-12-08 06:04:45.6433 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum

## 2024-12-08 06:04:45.794225 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis

## 2024-12-08 06:04:45.947403 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified

## 2024-12-08 06:04:46.093743 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar

## 2024-12-08 06:04:46.242275 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica

## 2024-12-08 06:04:46.387464 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8

## 2024-12-08 06:04:46.528068 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile

## 2024-12-08 06:04:46.680504 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47

## 2024-12-08 06:04:46.819801 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA

## 2024-12-08 06:04:46.974163 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila

## 2024-12-08 06:04:47.116239 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae

## 2024-12-08 06:04:47.268173 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus

## 2024-12-08 06:04:49.120594 INFO::Plotting data for metadata number 5, age
## 2024-12-08 06:04:49.122205 INFO::Creating scatter plot for continuous data, age vs Haemophilus.parainfluenzae

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:49.295438 INFO::Creating scatter plot for continuous data, age vs Bifidobacterium.pseudocatenulatum

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:49.502774 INFO::Creating scatter plot for continuous data, age vs Faecalibacterium.prausnitzii

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:49.653347 INFO::Creating scatter plot for continuous data, age vs Clostridium.clostridioforme

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:49.80836 INFO::Creating scatter plot for continuous data, age vs Veillonella.parvula

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:49.960806 INFO::Creating scatter plot for continuous data, age vs Subdoligranulum.unclassified

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:50.10635 INFO::Creating scatter plot for continuous data, age vs Clostridium.symbiosum

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:50.302822 INFO::Creating scatter plot for continuous data, age vs Ruminococcus.gnavus

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:50.460963 INFO::Creating scatter plot for continuous data, age vs Dialister.invisus

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:50.617317 INFO::Creating scatter plot for continuous data, age vs Veillonella.dispar

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:50.787606 INFO::Creating scatter plot for continuous data, age vs Veillonella.unclassified

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:50.933211 INFO::Creating scatter plot for continuous data, age vs Bacteroides.thetaiotaomicron

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:51.100749 INFO::Creating scatter plot for continuous data, age vs Ruminococcus.bromii

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:51.259541 INFO::Creating scatter plot for continuous data, age vs Bacteroides.intestinalis

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:51.418955 INFO::Creating scatter plot for continuous data, age vs Eubacterium.siraeum

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:51.561325 INFO::Creating scatter plot for continuous data, age vs Prevotella.copri

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:51.722841 INFO::Creating scatter plot for continuous data, age vs Alistipes.unclassified

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:51.877386 INFO::Creating scatter plot for continuous data, age vs Bacteroidales.bacterium.ph8

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:52.024094 INFO::Creating scatter plot for continuous data, age vs Bifidobacterium.longum

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:52.184479 INFO::Creating scatter plot for continuous data, age vs Akkermansia.muciniphila

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:52.334991 INFO::Creating scatter plot for continuous data, age vs Collinsella.aerofaciens

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:52.496094 INFO::Creating scatter plot for continuous data, age vs Parabacteroides.distasonis

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## `geom_smooth()` using formula = 'y ~ x'

## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).

## 2024-12-08 06:04:54.472822 INFO::Plotting data for metadata number 6, diagnosis
## 2024-12-08 06:04:54.474393 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis

## 2024-12-08 06:04:54.616877 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2024-12-08 06:04:54.788449 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2024-12-08 06:04:54.949272 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2024-12-08 06:04:55.126145 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2024-12-08 06:04:55.323076 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme

## 2024-12-08 06:04:55.526812 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2024-12-08 06:04:55.682509 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2024-12-08 06:04:55.872288 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2024-12-08 06:04:56.024652 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2024-12-08 06:04:56.194022 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans

## 2024-12-08 06:04:56.347764 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2024-12-08 06:04:56.519715 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2024-12-08 06:04:56.71697 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2024-12-08 06:04:56.873063 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii

## 2024-12-08 06:04:57.043612 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified

## 2024-12-08 06:04:57.197398 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2024-12-08 06:04:57.372436 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2024-12-08 06:04:57.534974 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2024-12-08 06:04:57.707614 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2024-12-08 06:04:57.8616 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum

## 2024-12-08 06:04:58.034666 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2024-12-08 06:04:58.190762 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2024-12-08 06:04:58.362437 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2024-12-08 06:04:58.516602 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2024-12-08 06:04:58.685843 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified

## 2024-12-08 06:04:58.842938 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2024-12-08 06:04:59.021219 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2024-12-08 06:04:59.18048 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified

## 2024-12-08 06:04:59.354477 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris

## 2024-12-08 06:04:59.514036 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2024-12-08 06:04:59.688214 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus

## 2024-12-08 06:04:59.851862 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis

## 2024-12-08 06:05:00.036005 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2024-12-08 06:05:00.211102 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus

## 2024-12-08 06:05:00.389202 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis

## 2024-12-08 06:05:00.567811 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2024-12-08 06:05:00.734269 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii

## 2024-12-08 06:05:00.912526 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2024-12-08 06:05:01.073196 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2024-12-08 06:05:01.247277 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2024-12-08 06:05:01.408667 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2024-12-08 06:05:01.585854 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.siraeum

Session Info

Session info from running the demo in R can be displayed with the following command.

sessionInfo()

## R version 4.4.2 (2024-10-31)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04.1 LTS
## 
## Matrix products: default
## BLAS:   /usr/lib/x86_64-linux-gnu/openblas-pthread/libblas.so.3 
## LAPACK: /usr/lib/x86_64-linux-gnu/openblas-pthread/libopenblasp-r0.3.26.so;  LAPACK version 3.12.0
## 
## locale:
##  [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C              
##  [3] LC_TIME=en_US.UTF-8        LC_COLLATE=C              
##  [5] LC_MONETARY=en_US.UTF-8    LC_MESSAGES=en_US.UTF-8   
##  [7] LC_PAPER=en_US.UTF-8       LC_NAME=C                 
##  [9] LC_ADDRESS=C               LC_TELEPHONE=C            
## [11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C       
## 
## time zone: Etc/UTC
## tzcode source: system (glibc)
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
## 
## other attached packages:
## [1] Maaslin2_1.21.0
## 
## loaded via a namespace (and not attached):
##  [1] gtable_0.3.6        biglm_0.9-3         xfun_0.49          
##  [4] bslib_0.8.0         ggplot2_3.5.1       lattice_0.22-6     
##  [7] numDeriv_2016.8-1.1 vctrs_0.6.5         tools_4.4.2        
## [10] generics_0.1.3      parallel_4.4.2      getopt_1.20.4      
## [13] tibble_3.2.1        fansi_1.0.6         DEoptimR_1.1-3-1   
## [16] cluster_2.1.6       pkgconfig_2.0.3     logging_0.10-108   
## [19] pheatmap_1.0.12     Matrix_1.7-1        data.table_1.16.4  
## [22] RColorBrewer_1.1-3  lifecycle_1.0.4     farver_2.1.2       
## [25] compiler_4.4.2      munsell_0.5.1       lmerTest_3.1-3     
## [28] permute_0.9-7       htmltools_0.5.8.1   sys_3.4.3          
## [31] buildtools_1.0.0    sass_0.4.9          hash_2.2.6.3       
## [34] yaml_2.3.10         pillar_1.9.0        nloptr_2.1.1       
## [37] crayon_1.5.3        jquerylib_0.1.4     MASS_7.3-61        
## [40] cachem_1.1.0        vegan_2.6-8         boot_1.3-31        
## [43] nlme_3.1-166        robustbase_0.99-4-1 tidyselect_1.2.1   
## [46] digest_0.6.37       mvtnorm_1.3-2       dplyr_1.1.4        
## [49] labeling_0.4.3      maketools_1.3.1     splines_4.4.2      
## [52] pcaPP_2.0-5         fastmap_1.2.0       grid_4.4.2         
## [55] colorspace_2.1-1    cli_3.6.3           magrittr_2.0.3     
## [58] utf8_1.2.4          withr_3.0.2         scales_1.3.0       
## [61] rmarkdown_2.29      lme4_1.1-35.5       pbapply_1.7-2      
## [64] evaluate_1.0.1      knitr_1.49          mgcv_1.9-1         
## [67] rlang_1.1.4         Rcpp_1.0.13-1       glue_1.8.0         
## [70] optparse_1.7.5      DBI_1.2.3           minqa_1.2.8        
## [73] jsonlite_1.8.9      R6_2.5.1

Options

Run MaAsLin2 help to print a list of the options and the default settings.

$ Maaslin2.R –help Usage: ./R/Maaslin2.R options <data.tsv> <metadata.tsv>

Options: -h, –help Show this help message and exit

-a MIN_ABUNDANCE, --min_abundance=MIN_ABUNDANCE
    The minimum abundance for each feature [ Default: 0 ]

-p MIN_PREVALENCE, --min_prevalence=MIN_PREVALENCE
    The minimum percent of samples for which a feature 
    is detected at minimum abundance [ Default: 0.1 ]

-b MIN_VARIANCE, --min_variance=MIN_VARIANCE
    Keep features with variance greater than [ Default: 0.0 ]

-s MAX_SIGNIFICANCE, --max_significance=MAX_SIGNIFICANCE
    The q-value threshold for significance [ Default: 0.25 ]

-n NORMALIZATION, --normalization=NORMALIZATION
    The normalization method to apply [ Default: TSS ]
    [ Choices: TSS, CLR, CSS, NONE, TMM ]

-t TRANSFORM, --transform=TRANSFORM
    The transform to apply [ Default: LOG ]
    [ Choices: LOG, LOGIT, AST, NONE ]

-m ANALYSIS_METHOD, --analysis_method=ANALYSIS_METHOD
    The analysis method to apply [ Default: LM ]
    [ Choices: LM, CPLM, NEGBIN, ZINB ]

-r RANDOM_EFFECTS, --random_effects=RANDOM_EFFECTS
    The random effects for the model, comma-delimited
    for multiple effects [ Default: none ]

-f FIXED_EFFECTS, --fixed_effects=FIXED_EFFECTS
    The fixed effects for the model, comma-delimited
    for multiple effects [ Default: all ]

-c CORRECTION, --correction=CORRECTION
    The correction method for computing the 
    q-value [ Default: BH ]

-z STANDARDIZE, --standardize=STANDARDIZE
    Apply z-score so continuous metadata are 
    on the same scale [ Default: TRUE ]

-l PLOT_HEATMAP, --plot_heatmap=PLOT_HEATMAP
    Generate a heatmap for the significant 
    associations [ Default: TRUE ]

-i HEATMAP_FIRST_N, --heatmap_first_n=HEATMAP_FIRST_N
    In heatmap, plot top N features with significant 
    associations [ Default: TRUE ]

-o PLOT_SCATTER, --plot_scatter=PLOT_SCATTER
    Generate scatter plots for the significant
    associations [ Default: TRUE ]
    
-g MAX_PNGS, --max_pngs=MAX_PNGS
    The maximum number of scatter plots for signficant associations 
    to save as png files [ Default: 10 ]

-O SAVE_SCATTER, --save_scatter=SAVE_SCATTER
    Save all scatter plot ggplot objects
    to an RData file [ Default: FALSE ]

-e CORES, --cores=CORES
    The number of R processes to run in parallel
    [ Default: 1 ]
    
-j SAVE_MODELS --save_models=SAVE_MODELS
    Return the full model outputs and save to an RData file
    [ Default: FALSE ]

-d REFERENCE, --reference=REFERENCE
    The factor to use as a reference level for a categorical variable 
    provided as a string of 'variable,reference', semi-colon delimited for 
    multiple variables. Not required if metadata is passed as a factor or 
    for variables with less than two levels but can be set regardless.
    [ Default: NA ]

Troubleshooting

Question: When I run from the command line I see the error Maaslin2.R: command not found. How do I fix this?
- Answer: Provide the full path to the executable when running Maaslin2.R.
Question: When I run as a function I see the error Error in library(Maaslin2): there is no package called 'Maaslin2'. How do I fix this?
- Answer: Install the R package and then try loading the library again.
Question: When I try to install the R package I see errors about dependencies not being installed. Why is this?
- Answer: Installing the R package will not automatically install the packages MaAsLin2 requires. Please install the dependencies and then install the MaAsLin2 R package.

- MaAsLin2 User Manual

Maaslin2