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"
## 2025-03-08 05:18:20.844386 INFO::Writing function arguments to log file
## 2025-03-08 05:18:20.859645 INFO::Verifying options selected are valid
## 2025-03-08 05:18:20.90172 INFO::Determining format of input files
## 2025-03-08 05:18:20.902437 INFO::Input format is data samples as rows and metadata samples as rows
## 2025-03-08 05:18:20.905406 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2025-03-08 05:18:20.906077 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2025-03-08 05:18:20.907587 INFO::Filter data based on min abundance and min prevalence
## 2025-03-08 05:18:20.964493 INFO::Total samples in data: 1595
## 2025-03-08 05:18:20.965084 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2025-03-08 05:18:20.968209 INFO::Total filtered features: 0
## 2025-03-08 05:18:20.968735 INFO::Filtered feature names from abundance and prevalence filtering:
## 2025-03-08 05:18:20.973983 INFO::Total filtered features with variance filtering: 0
## 2025-03-08 05:18:20.974464 INFO::Filtered feature names from variance filtering:
## 2025-03-08 05:18:20.974898 INFO::Running selected normalization method: TSS
## 2025-03-08 05:18:21.655656 INFO::Bypass z-score application to metadata
## 2025-03-08 05:18:21.656367 INFO::Running selected transform method: LOG
## 2025-03-08 05:18:21.67248 INFO::Running selected analysis method: LM
## 2025-03-08 05:18:21.890126 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis

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

## 2025-03-08 05:18:22.109897 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2025-03-08 05:18:22.214876 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2025-03-08 05:18:22.300389 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum

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

## 2025-03-08 05:18:22.389928 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2025-03-08 05:18:22.484187 INFO::Fitting model to feature number 6, Bacteroides.caccae

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

## 2025-03-08 05:18:22.574991 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2025-03-08 05:18:22.659882 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2025-03-08 05:18:22.75697 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2025-03-08 05:18:22.843483 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2025-03-08 05:18:22.936657 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

## 2025-03-08 05:18:23.008766 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

## 2025-03-08 05:18:23.100328 INFO::Fitting model to feature number 12, Bacteroides.fragilis

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

## 2025-03-08 05:18:23.186316 INFO::Fitting model to feature number 13, Bacteroides.intestinalis

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

## 2025-03-08 05:18:23.28456 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2025-03-08 05:18:23.373291 INFO::Fitting model to feature number 15, Bacteroides.ovatus

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

## 2025-03-08 05:18:23.455318 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2025-03-08 05:18:23.548951 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2025-03-08 05:18:23.64521 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron

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

## 2025-03-08 05:18:23.73405 INFO::Fitting model to feature number 19, Bacteroides.uniformis

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

## 2025-03-08 05:18:23.819728 INFO::Fitting model to feature number 20, Bacteroides.vulgatus

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

## 2025-03-08 05:18:23.927986 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens

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

## 2025-03-08 05:18:24.01522 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2025-03-08 05:18:24.105479 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2025-03-08 05:18:24.194538 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2025-03-08 05:18:24.289768 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2025-03-08 05:18:24.377114 INFO::Fitting model to feature number 26, Parabacteroides.distasonis

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

## 2025-03-08 05:18:24.468484 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2025-03-08 05:18:24.55774 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2025-03-08 05:18:24.651053 INFO::Fitting model to feature number 29, Parabacteroides.unclassified

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

## 2025-03-08 05:18:24.736488 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2025-03-08 05:18:24.830133 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2025-03-08 05:18:24.916807 INFO::Fitting model to feature number 32, Prevotella.copri

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

## 2025-03-08 05:18:25.008344 INFO::Fitting model to feature number 33, Alistipes.finegoldii

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

## 2025-03-08 05:18:25.248744 INFO::Fitting model to feature number 34, Alistipes.onderdonkii

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

## 2025-03-08 05:18:25.335989 INFO::Fitting model to feature number 35, Alistipes.putredinis

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

## 2025-03-08 05:18:25.422991 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2025-03-08 05:18:25.512245 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2025-03-08 05:18:25.603266 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2025-03-08 05:18:25.71148 INFO::Fitting model to feature number 39, Clostridium.bolteae

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

## 2025-03-08 05:18:25.803879 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

## 2025-03-08 05:18:25.880484 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

## 2025-03-08 05:18:25.966504 INFO::Fitting model to feature number 41, Clostridium.clostridioforme

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

## 2025-03-08 05:18:26.051036 INFO::Fitting model to feature number 42, Clostridium.hathewayi

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

## 2025-03-08 05:18:26.141395 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2025-03-08 05:18:26.234217 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2025-03-08 05:18:26.318242 INFO::Fitting model to feature number 45, Clostridium.symbiosum

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

## 2025-03-08 05:18:26.403648 INFO::Fitting model to feature number 46, Flavonifractor.plautii

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

## 2025-03-08 05:18:26.499086 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2025-03-08 05:18:26.583421 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2025-03-08 05:18:26.675094 INFO::Fitting model to feature number 49, Eubacterium.rectale

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

## 2025-03-08 05:18:26.770971 INFO::Fitting model to feature number 50, Eubacterium.siraeum

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

## 2025-03-08 05:18:26.854041 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31

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

## 2025-03-08 05:18:26.940501 INFO::Fitting model to feature number 52, Eubacterium.ventriosum

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

## 2025-03-08 05:18:27.02691 INFO::Fitting model to feature number 53, Ruminococcus.gnavus

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

## 2025-03-08 05:18:27.112647 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2025-03-08 05:18:27.195928 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2025-03-08 05:18:27.280676 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2025-03-08 05:18:27.371613 INFO::Fitting model to feature number 57, Dorea.longicatena

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

## 2025-03-08 05:18:27.457564 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA

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

## 2025-03-08 05:18:27.547024 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA

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

## 2025-03-08 05:18:27.63817 INFO::Fitting model to feature number 60, Roseburia.hominis

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

## 2025-03-08 05:18:27.725065 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2025-03-08 05:18:27.807129 INFO::Fitting model to feature number 62, Roseburia.inulinivorans

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

## 2025-03-08 05:18:27.897151 INFO::Fitting model to feature number 63, Roseburia.unclassified

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

## 2025-03-08 05:18:27.981788 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2025-03-08 05:18:28.078972 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2025-03-08 05:18:28.168045 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2025-03-08 05:18:28.2533 INFO::Fitting model to feature number 67, Ruminococcus.bromii

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

## 2025-03-08 05:18:28.338807 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2025-03-08 05:18:28.421666 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2025-03-08 05:18:28.509245 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified

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

## 2025-03-08 05:18:28.599282 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2025-03-08 05:18:28.683253 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2025-03-08 05:18:28.773561 INFO::Fitting model to feature number 73, Dialister.invisus

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

## 2025-03-08 05:18:28.86028 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2025-03-08 05:18:28.946296 INFO::Fitting model to feature number 75, Veillonella.dispar

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

## 2025-03-08 05:18:29.036286 INFO::Fitting model to feature number 76, Veillonella.parvula

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

## 2025-03-08 05:18:29.11796 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2025-03-08 05:18:29.200132 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2025-03-08 05:18:29.292611 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2025-03-08 05:18:29.375497 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2025-03-08 05:18:29.466637 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2025-03-08 05:18:29.556756 INFO::Fitting model to feature number 82, Escherichia.coli
## 2025-03-08 05:18:29.648142 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2025-03-08 05:18:29.728471 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae

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

## 2025-03-08 05:18:29.814906 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae

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

## 2025-03-08 05:18:29.900196 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2025-03-08 05:18:29.991391 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2025-03-08 05:18:30.10742 INFO::Counting total values for each feature
## 2025-03-08 05:18:30.126906 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2025-03-08 05:18:30.188663 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2025-03-08 05:18:30.251583 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2025-03-08 05:18:30.341076 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2025-03-08 05:18:30.536904 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2025-03-08 05:18:30.558911 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2025-03-08 05:18:30.562318 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2025-03-08 05:18:30.565517 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
## 2025-03-08 05:18:30.567067 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2025-03-08 05:18:30.693711 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2025-03-08 05:18:30.697172 INFO::Plotting associations from most to least significant, grouped by metadata
## 2025-03-08 05:18:30.697703 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2025-03-08 05:18:30.698652 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii

## 2025-03-08 05:18:30.902968 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified

## 2025-03-08 05:18:31.070518 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis

## 2025-03-08 05:18:31.221055 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale

## 2025-03-08 05:18:31.374734 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus

## 2025-03-08 05:18:31.527352 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus

## 2025-03-08 05:18:31.696765 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum

## 2025-03-08 05:18:31.852055 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified

## 2025-03-08 05:18:32.00116 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans

## 2025-03-08 05:18:32.157114 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis

## 2025-03-08 05:18:32.3108 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron

## 2025-03-08 05:18:32.463239 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis

## 2025-03-08 05:18:32.616057 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis

## 2025-03-08 05:18:32.766622 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei

## 2025-03-08 05:18:32.913626 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii

## 2025-03-08 05:18:33.065877 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens

## 2025-03-08 05:18:33.217082 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum

## 2025-03-08 05:18:33.369943 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena

## 2025-03-08 05:18:33.520733 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae

## 2025-03-08 05:18:33.674593 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2025-03-08 05:18:33.82886 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli

## 2025-03-08 05:18:33.977766 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae

## 2025-03-08 05:18:34.1316 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified

## 2025-03-08 05:18:34.281592 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii

## 2025-03-08 05:18:34.451872 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens

## 2025-03-08 05:18:34.605232 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris

## 2025-03-08 05:18:34.759431 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes

## 2025-03-08 05:18:34.911881 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis

## 2025-03-08 05:18:35.06529 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii

## 2025-03-08 05:18:35.218663 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii

## 2025-03-08 05:18:35.38296 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis

## 2025-03-08 05:18:35.534955 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara

## 2025-03-08 05:18:35.689119 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum

## 2025-03-08 05:18:35.840272 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae

## 2025-03-08 05:18:35.995704 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified

## 2025-03-08 05:18:36.154779 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens

## 2025-03-08 05:18:36.298816 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus

## 2025-03-08 05:18:36.455944 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme

## 2025-03-08 05:18:36.611183 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii

## 2025-03-08 05:18:36.766102 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii

## 2025-03-08 05:18:36.91834 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum

## 2025-03-08 05:18:37.077619 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris

## 2025-03-08 05:18:37.233611 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47

## 2025-03-08 05:18:37.378362 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus

## 2025-03-08 05:18:37.535282 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii

## 2025-03-08 05:18:37.691574 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis

## 2025-03-08 05:18:37.84626 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified

## 2025-03-08 05:18:38.003311 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques

## 2025-03-08 05:18:38.157868 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified

## 2025-03-08 05:18:38.320947 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus

## 2025-03-08 05:18:38.474207 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis

## 2025-03-08 05:18:38.628838 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified

## 2025-03-08 05:18:38.78725 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified

## 2025-03-08 05:18:38.957182 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis

## 2025-03-08 05:18:39.12708 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis

## 2025-03-08 05:18:39.281468 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31

## 2025-03-08 05:18:39.439738 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae

## 2025-03-08 05:18:39.596183 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8

## 2025-03-08 05:18:39.751668 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae

## 2025-03-08 05:18:39.910529 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii

## 2025-03-08 05:18:40.064544 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii

## 2025-03-08 05:18:40.222516 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum

## 2025-03-08 05:18:40.377916 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA

## 2025-03-08 05:18:42.380356 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2025-03-08 05:18:42.381562 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified

## 2025-03-08 05:18:42.521607 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii

## 2025-03-08 05:18:42.686579 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae

## 2025-03-08 05:18:42.851005 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified

## 2025-03-08 05:18:43.012154 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis

## 2025-03-08 05:18:43.177724 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum

## 2025-03-08 05:18:43.355275 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus

## 2025-03-08 05:18:43.51471 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis

## 2025-03-08 05:18:43.686144 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii

## 2025-03-08 05:18:43.853787 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale

## 2025-03-08 05:18:44.019894 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis

## 2025-03-08 05:18:44.195964 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis

## 2025-03-08 05:18:44.361904 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA

## 2025-03-08 05:18:44.520867 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii

## 2025-03-08 05:18:44.690067 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens

## 2025-03-08 05:18:44.857497 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris

## 2025-03-08 05:18:45.026392 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii

## 2025-03-08 05:18:45.196184 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis

## 2025-03-08 05:18:45.362761 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum

## 2025-03-08 05:18:45.525021 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus

## 2025-03-08 05:18:45.691844 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii

## 2025-03-08 05:18:45.854911 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii

## 2025-03-08 05:18:46.0188 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae

## 2025-03-08 05:18:46.184414 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus

## 2025-03-08 05:18:46.351433 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum

## 2025-03-08 05:18:46.514567 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae

## 2025-03-08 05:18:46.682004 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae

## 2025-03-08 05:18:46.84152 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum

## 2025-03-08 05:18:47.0044 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis

## 2025-03-08 05:18:47.169668 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii

## 2025-03-08 05:18:47.513287 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques

## 2025-03-08 05:18:47.658518 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8

## 2025-03-08 05:18:47.812925 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum

## 2025-03-08 05:18:47.953593 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum

## 2025-03-08 05:18:48.107011 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis

## 2025-03-08 05:18:48.253371 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens

## 2025-03-08 05:18:48.400532 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi

## 2025-03-08 05:18:48.550467 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified

## 2025-03-08 05:18:48.698042 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens

## 2025-03-08 05:18:48.843854 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus

## 2025-03-08 05:18:48.994561 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae

## 2025-03-08 05:18:49.148881 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus

## 2025-03-08 05:18:49.302617 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris

## 2025-03-08 05:18:49.453091 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47

## 2025-03-08 05:18:51.326611 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2025-03-08 05:18:51.327741 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii

## 2025-03-08 05:18:51.472295 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale

## 2025-03-08 05:18:51.625008 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31

## 2025-03-08 05:18:51.785042 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis

## 2025-03-08 05:18:51.935896 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified

## 2025-03-08 05:18:52.093615 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2025-03-08 05:18:52.254636 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques

## 2025-03-08 05:18:52.410511 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum

## 2025-03-08 05:18:52.564182 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus

## 2025-03-08 05:18:52.719578 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli

## 2025-03-08 05:18:52.873877 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum

## 2025-03-08 05:18:53.029667 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified

## 2025-03-08 05:18:53.190429 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis

## 2025-03-08 05:18:53.346974 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens

## 2025-03-08 05:18:53.506228 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae

## 2025-03-08 05:18:53.656024 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii

## 2025-03-08 05:18:53.804552 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus

## 2025-03-08 05:18:53.956089 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified

## 2025-03-08 05:18:54.1157 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis

## 2025-03-08 05:18:54.268234 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii

## 2025-03-08 05:18:54.422184 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans

## 2025-03-08 05:18:54.577657 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii

## 2025-03-08 05:18:54.735642 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified

## 2025-03-08 05:18:54.892503 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii

## 2025-03-08 05:18:55.045121 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis

## 2025-03-08 05:18:55.224 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri

## 2025-03-08 05:18:55.383082 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei

## 2025-03-08 05:18:55.533999 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis

## 2025-03-08 05:18:55.695476 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii

## 2025-03-08 05:18:57.663568 INFO::Plotting data for metadata number 4, antibiotics
## 2025-03-08 05:18:57.664745 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans

## 2025-03-08 05:18:57.795329 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis

## 2025-03-08 05:18:57.938926 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale

## 2025-03-08 05:18:58.091234 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus

## 2025-03-08 05:18:58.247269 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis

## 2025-03-08 05:18:58.397797 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena

## 2025-03-08 05:18:58.548918 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus

## 2025-03-08 05:18:58.699946 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii

## 2025-03-08 05:18:58.845716 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum

## 2025-03-08 05:18:58.987468 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae

## 2025-03-08 05:18:59.140856 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis

## 2025-03-08 05:18:59.288816 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii

## 2025-03-08 05:18:59.456788 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii

## 2025-03-08 05:18:59.617639 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified

## 2025-03-08 05:18:59.770985 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum

## 2025-03-08 05:18:59.91722 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA

## 2025-03-08 05:19:00.071648 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii

## 2025-03-08 05:19:00.229131 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31

## 2025-03-08 05:19:00.381783 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis

## 2025-03-08 05:19:00.531092 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii

## 2025-03-08 05:19:00.674811 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis

## 2025-03-08 05:19:00.830475 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens

## 2025-03-08 05:19:00.981684 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens

## 2025-03-08 05:19:01.138563 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron

## 2025-03-08 05:19:01.294959 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii

## 2025-03-08 05:19:01.452232 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae

## 2025-03-08 05:19:01.597119 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum

## 2025-03-08 05:19:01.746294 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques

## 2025-03-08 05:19:01.896128 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum

## 2025-03-08 05:19:02.055844 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis

## 2025-03-08 05:19:02.21155 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified

## 2025-03-08 05:19:02.366131 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar

## 2025-03-08 05:19:02.517863 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica

## 2025-03-08 05:19:02.669556 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8

## 2025-03-08 05:19:02.820551 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile

## 2025-03-08 05:19:02.969153 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47

## 2025-03-08 05:19:03.135049 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA

## 2025-03-08 05:19:03.290013 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila

## 2025-03-08 05:19:03.447213 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae

## 2025-03-08 05:19:03.607062 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus

## 2025-03-08 05:19:05.541432 INFO::Plotting data for metadata number 5, age
## 2025-03-08 05:19:05.542599 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()`).

## 2025-03-08 05:19:05.721181 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()`).

## 2025-03-08 05:19:05.947133 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()`).

## 2025-03-08 05:19:06.101229 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()`).

## 2025-03-08 05:19:06.270728 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()`).

## 2025-03-08 05:19:06.426683 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()`).

## 2025-03-08 05:19:06.58506 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()`).

## 2025-03-08 05:19:06.751999 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()`).

## 2025-03-08 05:19:06.916754 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()`).

## 2025-03-08 05:19:07.102881 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()`).

## 2025-03-08 05:19:07.270299 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()`).

## 2025-03-08 05:19:07.432216 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()`).

## 2025-03-08 05:19:07.595151 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()`).

## 2025-03-08 05:19:07.760015 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()`).

## 2025-03-08 05:19:07.918983 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()`).

## 2025-03-08 05:19:08.075156 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()`).

## 2025-03-08 05:19:08.236251 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()`).

## 2025-03-08 05:19:08.397129 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()`).

## 2025-03-08 05:19:08.555273 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()`).

## 2025-03-08 05:19:08.716367 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()`).

## 2025-03-08 05:19:08.887366 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()`).

## 2025-03-08 05:19:09.053739 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()`).

## 2025-03-08 05:19:11.266512 INFO::Plotting data for metadata number 6, diagnosis
## 2025-03-08 05:19:11.267663 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis

## 2025-03-08 05:19:11.397636 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2025-03-08 05:19:11.545235 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2025-03-08 05:19:11.692897 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2025-03-08 05:19:11.845166 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2025-03-08 05:19:11.994624 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme

## 2025-03-08 05:19:12.141727 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2025-03-08 05:19:12.285305 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2025-03-08 05:19:12.435372 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2025-03-08 05:19:12.58303 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2025-03-08 05:19:12.733193 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans

## 2025-03-08 05:19:12.883803 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2025-03-08 05:19:13.036909 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2025-03-08 05:19:13.185471 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2025-03-08 05:19:13.337059 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii

## 2025-03-08 05:19:13.487214 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified

## 2025-03-08 05:19:13.632482 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2025-03-08 05:19:13.783782 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2025-03-08 05:19:13.934999 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2025-03-08 05:19:14.088176 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2025-03-08 05:19:14.252775 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum

## 2025-03-08 05:19:14.40428 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2025-03-08 05:19:14.560558 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2025-03-08 05:19:14.718738 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2025-03-08 05:19:14.872856 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2025-03-08 05:19:15.02766 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified

## 2025-03-08 05:19:15.181138 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2025-03-08 05:19:15.342764 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2025-03-08 05:19:15.499639 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified

## 2025-03-08 05:19:15.65723 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris

## 2025-03-08 05:19:15.807092 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2025-03-08 05:19:15.959604 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus

## 2025-03-08 05:19:16.112051 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis

## 2025-03-08 05:19:16.270855 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2025-03-08 05:19:16.429334 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus

## 2025-03-08 05:19:16.588721 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis

## 2025-03-08 05:19:16.742886 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2025-03-08 05:19:16.900227 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii

## 2025-03-08 05:19:17.059993 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2025-03-08 05:19:17.212436 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2025-03-08 05:19:17.366003 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2025-03-08 05:19:17.52257 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2025-03-08 05:19:17.678434 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 4.4.3 (2025-02-28)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04.2 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_0.51          
##  [4] bslib_0.9.0         ggplot2_3.5.1       lattice_0.22-6     
##  [7] numDeriv_2016.8-1.1 vctrs_0.6.5         tools_4.4.3        
## [10] Rdpack_2.6.2        generics_0.1.3      parallel_4.4.3     
## [13] getopt_1.20.4       tibble_3.2.1        DEoptimR_1.1-3-1   
## [16] cluster_2.1.8       pkgconfig_2.0.3     logging_0.10-108   
## [19] pheatmap_1.0.12     Matrix_1.7-2        data.table_1.17.0  
## [22] RColorBrewer_1.1-3  lifecycle_1.0.4     farver_2.1.2       
## [25] compiler_4.4.3      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_1.10.1       nloptr_2.1.1       
## [37] crayon_1.5.3        jquerylib_0.1.4     MASS_7.3-65        
## [40] cachem_1.1.0        vegan_2.6-10        reformulas_0.4.0   
## [43] boot_1.3-31         nlme_3.1-167        robustbase_0.99-4-1
## [46] tidyselect_1.2.1    digest_0.6.37       mvtnorm_1.3-3      
## [49] dplyr_1.1.4         labeling_0.4.3      maketools_1.3.2    
## [52] splines_4.4.3       pcaPP_2.0-5         fastmap_1.2.0      
## [55] grid_4.4.3          colorspace_2.1-1    cli_3.6.4          
## [58] magrittr_2.0.3      withr_3.0.2         scales_1.3.0       
## [61] rmarkdown_2.29      lme4_1.1-36         pbapply_1.7-2      
## [64] evaluate_1.0.3      knitr_1.49          rbibutils_2.3      
## [67] mgcv_1.9-1          rlang_1.1.5         Rcpp_1.0.14        
## [70] glue_1.8.0          optparse_1.7.5      DBI_1.2.3          
## [73] minqa_1.2.8         jsonlite_1.9.1      R6_2.6.1

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