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

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

  1. Download the source: MaAsLin2.tar.gz
  2. Decompress the download:
    • $ tar xzvf maaslin2.tar.gz
  3. Install the Bioconductor dependencies edgeR and metagenomeSeq.
  4. 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')"
  5. 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.

  1. 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.
  2. 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.

  1. 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.
  2. 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"
## 2026-04-08 18:31:05.34584 INFO::Writing function arguments to log file
## 2026-04-08 18:31:05.377945 INFO::Verifying options selected are valid
## 2026-04-08 18:31:05.41017 INFO::Determining format of input files
## 2026-04-08 18:31:05.411464 INFO::Input format is data samples as rows and metadata samples as rows
## 2026-04-08 18:31:05.416136 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2026-04-08 18:31:05.417214 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2026-04-08 18:31:05.419412 INFO::Filter data based on min abundance and min prevalence
## 2026-04-08 18:31:05.420103 INFO::Total samples in data: 1595
## 2026-04-08 18:31:05.42077 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2026-04-08 18:31:05.425366 INFO::Total filtered features: 0
## 2026-04-08 18:31:05.426209 INFO::Filtered feature names from abundance and prevalence filtering:
## 2026-04-08 18:31:05.434402 INFO::Total filtered features with variance filtering: 0
## 2026-04-08 18:31:05.435225 INFO::Filtered feature names from variance filtering:
## 2026-04-08 18:31:05.43591 INFO::Running selected normalization method: TSS
## 2026-04-08 18:31:06.498181 INFO::Bypass z-score application to metadata
## 2026-04-08 18:31:06.499273 INFO::Running selected transform method: LOG
## 2026-04-08 18:31:06.514769 INFO::Running selected analysis method: LM
## 2026-04-08 18:31:06.85343 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:07.75968 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2026-04-08 18:31:07.925239 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2026-04-08 18:31:08.08544 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:08.215978 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2026-04-08 18:31:08.368284 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:08.509864 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2026-04-08 18:31:08.658165 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2026-04-08 18:31:08.807302 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2026-04-08 18:31:08.949129 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2026-04-08 18:31:09.098185 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
## 2026-04-08 18:31:09.212141 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
## 2026-04-08 18:31:09.353493 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:09.497648 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:09.638456 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2026-04-08 18:31:09.776279 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:09.911151 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2026-04-08 18:31:10.051611 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2026-04-08 18:31:10.190864 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:10.3435 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:10.492596 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:10.628043 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:10.780492 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2026-04-08 18:31:10.917309 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2026-04-08 18:31:11.054094 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2026-04-08 18:31:11.203284 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2026-04-08 18:31:11.341146 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:11.477937 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2026-04-08 18:31:11.619167 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2026-04-08 18:31:11.759681 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:11.896589 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2026-04-08 18:31:12.039679 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2026-04-08 18:31:12.186965 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:12.320768 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:12.456543 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:12.60124 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:12.752185 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2026-04-08 18:31:12.893743 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2026-04-08 18:31:13.024729 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2026-04-08 18:31:13.168248 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:13.306047 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
## 2026-04-08 18:31:13.421308 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
## 2026-04-08 18:31:13.554107 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:13.686655 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:13.822589 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2026-04-08 18:31:13.968863 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2026-04-08 18:31:14.107449 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:14.242495 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:14.376999 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2026-04-08 18:31:14.544108 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2026-04-08 18:31:14.685539 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:14.828612 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:14.96788 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:15.101336 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:15.238883 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:15.399238 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2026-04-08 18:31:15.538806 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2026-04-08 18:31:15.674388 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2026-04-08 18:31:16.066914 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:16.20066 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:16.338976 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:16.478198 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:16.633706 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2026-04-08 18:31:16.773516 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:16.904991 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:17.038786 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2026-04-08 18:31:17.186183 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2026-04-08 18:31:17.330472 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2026-04-08 18:31:17.467692 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:17.594038 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2026-04-08 18:31:17.728411 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2026-04-08 18:31:17.877193 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:18.034182 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2026-04-08 18:31:18.17371 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2026-04-08 18:31:18.315212 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:18.452862 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2026-04-08 18:31:18.605944 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:18.739146 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:18.867855 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2026-04-08 18:31:19.00078 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2026-04-08 18:31:19.147506 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2026-04-08 18:31:19.279601 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2026-04-08 18:31:19.418004 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2026-04-08 18:31:19.563582 INFO::Fitting model to feature number 82, Escherichia.coli
## 2026-04-08 18:31:19.931473 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2026-04-08 18:31:20.061267 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:20.188665 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2026-04-08 18:31:20.32489 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2026-04-08 18:31:20.467722 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2026-04-08 18:31:20.869992 INFO::Counting total values for each feature
## 2026-04-08 18:31:20.900105 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2026-04-08 18:31:21.008458 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2026-04-08 18:31:21.120983 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2026-04-08 18:31:21.315343 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2026-04-08 18:31:21.371797 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2026-04-08 18:31:21.404043 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2026-04-08 18:31:21.40927 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2026-04-08 18:31:21.414513 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
## 2026-04-08 18:31:21.417025 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2026-04-08 18:31:21.624286 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2026-04-08 18:31:21.630249 INFO::Plotting associations from most to least significant, grouped by metadata
## 2026-04-08 18:31:21.631304 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2026-04-08 18:31:21.632872 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2026-04-08 18:31:22.105868 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2026-04-08 18:31:22.493434 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2026-04-08 18:31:22.865895 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2026-04-08 18:31:23.249971 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2026-04-08 18:31:23.654916 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2026-04-08 18:31:24.023622 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2026-04-08 18:31:24.389251 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2026-04-08 18:31:24.753505 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2026-04-08 18:31:25.119326 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2026-04-08 18:31:25.494869 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2026-04-08 18:31:25.860465 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2026-04-08 18:31:26.220196 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2026-04-08 18:31:26.583742 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2026-04-08 18:31:26.9838 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2026-04-08 18:31:27.361982 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2026-04-08 18:31:27.717333 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2026-04-08 18:31:28.148239 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2026-04-08 18:31:28.518056 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2026-04-08 18:31:28.870656 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-04-08 18:31:29.234129 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2026-04-08 18:31:29.598467 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2026-04-08 18:31:29.963206 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2026-04-08 18:31:30.321335 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2026-04-08 18:31:30.679919 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2026-04-08 18:31:31.036346 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2026-04-08 18:31:31.397492 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2026-04-08 18:31:35.403295 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2026-04-08 18:31:35.769681 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2026-04-08 18:31:36.141131 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2026-04-08 18:31:36.50519 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2026-04-08 18:31:36.876056 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2026-04-08 18:31:37.23986 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2026-04-08 18:31:37.599171 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2026-04-08 18:31:37.964926 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2026-04-08 18:31:38.340625 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2026-04-08 18:31:38.723995 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2026-04-08 18:31:39.102458 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2026-04-08 18:31:39.504261 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2026-04-08 18:31:39.884484 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2026-04-08 18:31:40.253091 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2026-04-08 18:31:40.627961 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2026-04-08 18:31:41.000083 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2026-04-08 18:31:41.370456 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2026-04-08 18:31:41.746046 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2026-04-08 18:31:42.123433 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2026-04-08 18:31:42.526807 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2026-04-08 18:31:42.903393 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2026-04-08 18:31:43.273412 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2026-04-08 18:31:43.66188 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2026-04-08 18:31:44.029833 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2026-04-08 18:31:44.400748 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2026-04-08 18:31:44.777397 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2026-04-08 18:31:45.16132 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2026-04-08 18:31:45.537781 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2026-04-08 18:31:45.920469 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2026-04-08 18:31:46.2907 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2026-04-08 18:31:46.65588 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2026-04-08 18:31:47.064381 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2026-04-08 18:31:47.487993 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2026-04-08 18:31:47.866245 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2026-04-08 18:31:48.257582 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2026-04-08 18:31:48.638472 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2026-04-08 18:31:52.518922 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2026-04-08 18:31:52.520794 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2026-04-08 18:31:52.865132 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2026-04-08 18:31:53.246576 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2026-04-08 18:31:53.627158 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2026-04-08 18:31:54.007011 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2026-04-08 18:31:54.400524 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2026-04-08 18:31:54.790248 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2026-04-08 18:31:55.168133 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2026-04-08 18:31:55.569428 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2026-04-08 18:31:55.978158 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2026-04-08 18:31:56.398341 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2026-04-08 18:31:56.792395 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2026-04-08 18:31:57.18421 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-04-08 18:31:57.572092 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2026-04-08 18:31:57.969858 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2026-04-08 18:31:58.361574 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2026-04-08 18:31:58.755772 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2026-04-08 18:31:59.144897 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2026-04-08 18:31:59.530476 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2026-04-08 18:31:59.907485 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2026-04-08 18:32:00.29184 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2026-04-08 18:32:00.693241 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2026-04-08 18:32:01.088662 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2026-04-08 18:32:01.499579 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2026-04-08 18:32:01.908011 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2026-04-08 18:32:02.292556 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2026-04-08 18:32:02.686831 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2026-04-08 18:32:03.080067 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2026-04-08 18:32:03.459614 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2026-04-08 18:32:03.897348 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2026-04-08 18:32:04.298953 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2026-04-08 18:32:04.671412 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2026-04-08 18:32:05.060688 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2026-04-08 18:32:05.44007 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2026-04-08 18:32:05.83213 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2026-04-08 18:32:06.211158 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2026-04-08 18:32:06.600034 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2026-04-08 18:32:06.999101 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2026-04-08 18:32:07.396755 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2026-04-08 18:32:07.788312 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2026-04-08 18:32:08.179456 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2026-04-08 18:32:08.570175 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2026-04-08 18:32:08.955646 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2026-04-08 18:32:09.336424 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2026-04-08 18:32:13.393201 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2026-04-08 18:32:13.394972 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2026-04-08 18:32:13.714764 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2026-04-08 18:32:14.068327 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2026-04-08 18:32:14.422714 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2026-04-08 18:32:14.768885 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2026-04-08 18:32:15.129529 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-04-08 18:32:15.4928 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2026-04-08 18:32:15.851297 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2026-04-08 18:32:16.215614 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2026-04-08 18:32:16.56655 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2026-04-08 18:32:16.914696 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2026-04-08 18:32:17.267729 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2026-04-08 18:32:17.626386 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2026-04-08 18:32:17.979789 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2026-04-08 18:32:18.338811 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2026-04-08 18:32:18.690578 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2026-04-08 18:32:19.032356 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2026-04-08 18:32:19.3779 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2026-04-08 18:32:19.780529 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2026-04-08 18:32:20.135277 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2026-04-08 18:32:20.493805 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2026-04-08 18:32:20.853971 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2026-04-08 18:32:21.219843 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2026-04-08 18:32:21.585832 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2026-04-08 18:32:21.945318 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2026-04-08 18:32:22.313682 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2026-04-08 18:32:22.677824 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2026-04-08 18:32:23.032864 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2026-04-08 18:32:23.399584 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2026-04-08 18:32:27.124653 INFO::Plotting data for metadata number 4, antibiotics
## 2026-04-08 18:32:27.12647 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2026-04-08 18:32:27.494558 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2026-04-08 18:32:27.865425 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2026-04-08 18:32:28.233158 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2026-04-08 18:32:28.599498 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2026-04-08 18:32:28.959278 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2026-04-08 18:32:29.32379 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2026-04-08 18:32:29.69211 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2026-04-08 18:32:30.050113 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2026-04-08 18:32:30.4109 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2026-04-08 18:32:30.779812 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2026-04-08 18:32:31.127404 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2026-04-08 18:32:31.481491 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2026-04-08 18:32:31.850717 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2026-04-08 18:32:32.207638 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2026-04-08 18:32:32.564439 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-04-08 18:32:32.926642 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2026-04-08 18:32:33.288959 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2026-04-08 18:32:33.651195 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2026-04-08 18:32:34.001618 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2026-04-08 18:32:34.357207 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2026-04-08 18:32:34.763136 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2026-04-08 18:32:35.124024 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2026-04-08 18:32:35.4996 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2026-04-08 18:32:35.863215 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2026-04-08 18:32:36.231465 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2026-04-08 18:32:36.586715 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2026-04-08 18:32:36.960474 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2026-04-08 18:32:37.330999 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2026-04-08 18:32:37.703736 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2026-04-08 18:32:38.067101 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2026-04-08 18:32:38.428693 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2026-04-08 18:32:38.782687 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2026-04-08 18:32:39.154454 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2026-04-08 18:32:39.526504 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2026-04-08 18:32:39.887837 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2026-04-08 18:32:40.245262 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2026-04-08 18:32:40.627512 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2026-04-08 18:32:40.996698 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2026-04-08 18:32:41.394001 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2026-04-08 18:32:45.159584 INFO::Plotting data for metadata number 5, age
## 2026-04-08 18:32:45.161639 INFO::Creating scatter plot for continuous data, age vs Haemophilus.parainfluenzae
## Warning: Using `size` aesthetic for lines was deprecated in ggplot2 3.4.0.
## ℹ Please use `linewidth` instead.
## ℹ The deprecated feature was likely used in the Maaslin2 package.
##   Please report the issue at <https://github.com/biobakery/maaslin2/issues>.
## This warning is displayed once per session.
## Call `lifecycle::last_lifecycle_warnings()` to see where this warning was
## generated.
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2026-04-08 18:32:45.616791 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()`).
## 2026-04-08 18:32:46.071244 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()`).
## 2026-04-08 18:32:46.437993 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()`).
## 2026-04-08 18:32:46.817379 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()`).
## 2026-04-08 18:32:47.173506 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()`).
## 2026-04-08 18:32:47.530235 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()`).
## 2026-04-08 18:32:47.910975 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()`).
## 2026-04-08 18:32:48.280139 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()`).
## 2026-04-08 18:32:48.6427 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()`).
## 2026-04-08 18:32:49.067018 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()`).
## 2026-04-08 18:32:49.484842 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()`).
## 2026-04-08 18:32:49.851876 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()`).
## 2026-04-08 18:32:50.222994 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()`).
## 2026-04-08 18:32:50.580694 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()`).
## 2026-04-08 18:32:50.958628 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()`).
## 2026-04-08 18:32:51.314867 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()`).
## 2026-04-08 18:32:51.66276 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()`).
## 2026-04-08 18:32:52.034541 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()`).
## 2026-04-08 18:32:52.389264 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()`).
## 2026-04-08 18:32:52.76433 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()`).
## 2026-04-08 18:32:53.125633 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()`).
## 2026-04-08 18:32:57.082463 INFO::Plotting data for metadata number 6, diagnosis
## 2026-04-08 18:32:57.084293 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2026-04-08 18:32:57.409648 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2026-04-08 18:32:57.762312 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2026-04-08 18:32:58.116354 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2026-04-08 18:32:58.4775 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2026-04-08 18:32:58.838984 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2026-04-08 18:32:59.199513 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2026-04-08 18:32:59.558884 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2026-04-08 18:32:59.926309 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2026-04-08 18:33:00.304993 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2026-04-08 18:33:00.688377 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2026-04-08 18:33:01.045802 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2026-04-08 18:33:01.406263 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2026-04-08 18:33:01.762368 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2026-04-08 18:33:02.11958 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2026-04-08 18:33:02.478108 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2026-04-08 18:33:02.83356 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2026-04-08 18:33:03.246334 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2026-04-08 18:33:03.614536 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2026-04-08 18:33:03.985991 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2026-04-08 18:33:04.383115 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2026-04-08 18:33:04.766289 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2026-04-08 18:33:05.147533 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2026-04-08 18:33:05.525468 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2026-04-08 18:33:05.888857 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2026-04-08 18:33:06.258936 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2026-04-08 18:33:06.622679 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2026-04-08 18:33:06.997758 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2026-04-08 18:33:07.375009 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2026-04-08 18:33:07.755769 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2026-04-08 18:33:08.126745 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2026-04-08 18:33:08.490975 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2026-04-08 18:33:08.854342 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2026-04-08 18:33:09.227298 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2026-04-08 18:33:09.610311 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2026-04-08 18:33:10.019348 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2026-04-08 18:33:10.403441 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2026-04-08 18:33:10.789231 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2026-04-08 18:33:11.178708 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2026-04-08 18:33:11.560855 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2026-04-08 18:33:11.937542 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2026-04-08 18:33:12.319924 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2026-04-08 18:33:12.703732 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 4.6.0 alpha (2026-04-05 r89794)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04.4 LTS
## 
## Matrix products: default
## BLAS:   /home/biocbuild/bbs-3.23-bioc/R/lib/libRblas.so 
## LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.12.0  LAPACK version 3.12.0
## 
## locale:
##  [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C              
##  [3] LC_TIME=en_GB              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: America/New_York
## tzcode source: system (glibc)
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
## 
## other attached packages:
## [1] Maaslin2_1.25.1
## 
## loaded via a namespace (and not attached):
##  [1] gtable_0.3.6        biglm_0.9-3         xfun_0.57          
##  [4] bslib_0.10.0        ggplot2_4.0.2       lattice_0.22-9     
##  [7] numDeriv_2016.8-1.1 vctrs_0.7.2         tools_4.6.0        
## [10] Rdpack_2.6.6        generics_0.1.4      parallel_4.6.0     
## [13] getopt_1.20.4       tibble_3.3.1        DEoptimR_1.1-4     
## [16] cluster_2.1.8.2     pkgconfig_2.0.3     logging_0.10-108   
## [19] pheatmap_1.0.13     Matrix_1.7-5        data.table_1.18.2.1
## [22] RColorBrewer_1.1-3  S7_0.2.1            lifecycle_1.0.5    
## [25] compiler_4.6.0      farver_2.1.2        lmerTest_3.2-1     
## [28] permute_0.9-10      htmltools_0.5.9     sass_0.4.10        
## [31] hash_2.2.6.4        yaml_2.3.12         pillar_1.11.1      
## [34] nloptr_2.2.1        crayon_1.5.3        jquerylib_0.1.4    
## [37] MASS_7.3-65         cachem_1.1.0        vegan_2.7-3        
## [40] reformulas_0.4.4    boot_1.3-32         nlme_3.1-169       
## [43] robustbase_0.99-7   tidyselect_1.2.1    digest_0.6.39      
## [46] mvtnorm_1.3-6       dplyr_1.2.1         labeling_0.4.3     
## [49] splines_4.6.0       pcaPP_2.0-5         fastmap_1.2.0      
## [52] grid_4.6.0          cli_3.6.5           magrittr_2.0.5     
## [55] dichromat_2.0-0.1   withr_3.0.2         scales_1.4.0       
## [58] rmarkdown_2.31      otel_0.2.0          lme4_2.0-1         
## [61] pbapply_1.7-4       evaluate_1.0.5      knitr_1.51         
## [64] rbibutils_2.4.1     mgcv_1.9-4          rlang_1.2.0        
## [67] Rcpp_1.1.1          glue_1.8.0          optparse_1.7.5     
## [70] DBI_1.3.0           minqa_1.2.8         jsonlite_2.0.0     
## [73] 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

  1. 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.
  2. 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.
  3. 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.