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"
## 2026-03-16 18:19:59.999236 INFO::Writing function arguments to log file
## 2026-03-16 18:20:00.052148 INFO::Verifying options selected are valid
## 2026-03-16 18:20:00.108674 INFO::Determining format of input files
## 2026-03-16 18:20:00.110704 INFO::Input format is data samples as rows and metadata samples as rows
## 2026-03-16 18:20:00.118195 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2026-03-16 18:20:00.119387 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2026-03-16 18:20:00.121614 INFO::Filter data based on min abundance and min prevalence
## 2026-03-16 18:20:00.122332 INFO::Total samples in data: 1595
## 2026-03-16 18:20:00.123007 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2026-03-16 18:20:00.127675 INFO::Total filtered features: 0
## 2026-03-16 18:20:00.128594 INFO::Filtered feature names from abundance and prevalence filtering:
## 2026-03-16 18:20:00.137261 INFO::Total filtered features with variance filtering: 0
## 2026-03-16 18:20:00.138246 INFO::Filtered feature names from variance filtering:
## 2026-03-16 18:20:00.138959 INFO::Running selected normalization method: TSS
## 2026-03-16 18:20:01.197703 INFO::Bypass z-score application to metadata
## 2026-03-16 18:20:01.198604 INFO::Running selected transform method: LOG
## 2026-03-16 18:20:01.215108 INFO::Running selected analysis method: LM
## 2026-03-16 18:20:01.546784 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:02.56499 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2026-03-16 18:20:02.738828 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2026-03-16 18:20:02.909764 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:03.052404 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2026-03-16 18:20:03.209244 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:03.364101 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2026-03-16 18:20:03.512259 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2026-03-16 18:20:03.665748 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2026-03-16 18:20:03.824085 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2026-03-16 18:20:03.98218 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
## 2026-03-16 18:20:04.102195 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
## 2026-03-16 18:20:04.244047 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:04.397343 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:04.590486 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2026-03-16 18:20:04.739288 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:04.974549 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2026-03-16 18:20:05.122056 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2026-03-16 18:20:05.267181 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:05.407553 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:05.549411 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:05.693934 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:05.827586 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2026-03-16 18:20:05.961872 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2026-03-16 18:20:06.130839 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2026-03-16 18:20:06.279161 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2026-03-16 18:20:06.431847 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:06.564682 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2026-03-16 18:20:06.704487 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2026-03-16 18:20:06.853496 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:06.992379 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2026-03-16 18:20:07.143775 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2026-03-16 18:20:07.280402 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:07.418334 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:07.54885 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:07.690064 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:07.823303 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2026-03-16 18:20:07.969005 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2026-03-16 18:20:08.100803 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2026-03-16 18:20:08.28011 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:08.427725 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
## 2026-03-16 18:20:08.545696 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
## 2026-03-16 18:20:08.687858 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:08.817993 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:08.957899 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2026-03-16 18:20:09.140284 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2026-03-16 18:20:09.283751 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:09.427949 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:09.581526 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2026-03-16 18:20:09.72015 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2026-03-16 18:20:09.867658 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:10.02948 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:10.1726 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:10.316662 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:10.468976 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:10.606631 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2026-03-16 18:20:10.749174 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2026-03-16 18:20:10.91062 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2026-03-16 18:20:11.064035 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:11.204714 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:11.647011 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:11.784633 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:11.926085 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2026-03-16 18:20:12.068605 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:12.236411 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:12.376222 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2026-03-16 18:20:12.52764 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2026-03-16 18:20:12.671866 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2026-03-16 18:20:12.818686 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:12.956398 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2026-03-16 18:20:13.089902 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2026-03-16 18:20:13.231246 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:13.367621 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2026-03-16 18:20:13.508579 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2026-03-16 18:20:13.648767 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:13.780486 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2026-03-16 18:20:13.92081 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:14.061795 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:14.190637 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2026-03-16 18:20:14.324328 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2026-03-16 18:20:14.488251 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2026-03-16 18:20:14.644815 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2026-03-16 18:20:14.787095 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2026-03-16 18:20:14.939797 INFO::Fitting model to feature number 82, Escherichia.coli
## 2026-03-16 18:20:15.085458 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2026-03-16 18:20:15.224802 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:15.355884 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2026-03-16 18:20:15.491616 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2026-03-16 18:20:15.633974 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2026-03-16 18:20:15.817838 INFO::Counting total values for each feature
## 2026-03-16 18:20:15.847657 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2026-03-16 18:20:15.939672 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2026-03-16 18:20:16.038065 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2026-03-16 18:20:16.212975 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2026-03-16 18:20:16.268901 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2026-03-16 18:20:16.300892 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2026-03-16 18:20:16.306005 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2026-03-16 18:20:16.310896 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
## 2026-03-16 18:20:16.313185 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2026-03-16 18:20:16.518098 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2026-03-16 18:20:16.52427 INFO::Plotting associations from most to least significant, grouped by metadata
## 2026-03-16 18:20:16.525192 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2026-03-16 18:20:16.526733 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2026-03-16 18:20:17.020225 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2026-03-16 18:20:17.415479 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2026-03-16 18:20:17.783443 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2026-03-16 18:20:18.15128 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2026-03-16 18:20:18.511272 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2026-03-16 18:20:18.878867 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2026-03-16 18:20:19.273973 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2026-03-16 18:20:19.647424 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2026-03-16 18:20:20.012648 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2026-03-16 18:20:20.376592 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2026-03-16 18:20:20.728014 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2026-03-16 18:20:21.10109 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2026-03-16 18:20:21.481054 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2026-03-16 18:20:22.145122 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2026-03-16 18:20:22.502928 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2026-03-16 18:20:22.935826 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2026-03-16 18:20:24.232936 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2026-03-16 18:20:25.417822 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2026-03-16 18:20:26.335672 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-03-16 18:20:26.87912 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2026-03-16 18:20:27.248598 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2026-03-16 18:20:27.62559 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2026-03-16 18:20:28.025298 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2026-03-16 18:20:28.387371 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2026-03-16 18:20:28.750751 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2026-03-16 18:20:29.115258 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2026-03-16 18:20:29.478442 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2026-03-16 18:20:29.853211 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2026-03-16 18:20:30.213278 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2026-03-16 18:20:30.565269 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2026-03-16 18:20:30.929972 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2026-03-16 18:20:31.286782 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2026-03-16 18:20:31.631068 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2026-03-16 18:20:32.021708 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2026-03-16 18:20:32.393389 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2026-03-16 18:20:32.755056 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2026-03-16 18:20:33.12122 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2026-03-16 18:20:33.480654 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2026-03-16 18:20:33.848051 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2026-03-16 18:20:34.218255 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2026-03-16 18:20:34.593044 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2026-03-16 18:20:34.962116 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2026-03-16 18:20:35.332756 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2026-03-16 18:20:35.704985 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2026-03-16 18:20:36.075247 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2026-03-16 18:20:36.448731 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2026-03-16 18:20:36.815948 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2026-03-16 18:20:37.174413 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2026-03-16 18:20:37.560834 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2026-03-16 18:20:37.932881 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2026-03-16 18:20:38.305333 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2026-03-16 18:20:38.672338 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2026-03-16 18:20:39.033332 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2026-03-16 18:20:39.388882 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2026-03-16 18:20:39.794369 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2026-03-16 18:20:40.167035 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2026-03-16 18:20:40.535049 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2026-03-16 18:20:40.907659 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2026-03-16 18:20:41.276942 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2026-03-16 18:20:41.643044 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2026-03-16 18:20:42.018162 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2026-03-16 18:20:42.382054 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2026-03-16 18:20:46.274755 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2026-03-16 18:20:46.276654 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2026-03-16 18:20:46.625289 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2026-03-16 18:20:47.004213 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2026-03-16 18:20:47.379771 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2026-03-16 18:20:47.75024 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2026-03-16 18:20:48.125076 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2026-03-16 18:20:48.540564 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2026-03-16 18:20:48.915441 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2026-03-16 18:20:49.308109 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2026-03-16 18:20:49.703242 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2026-03-16 18:20:50.090332 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2026-03-16 18:20:50.482193 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2026-03-16 18:20:50.86902 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-03-16 18:20:51.250673 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2026-03-16 18:20:51.636065 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2026-03-16 18:20:52.027646 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2026-03-16 18:20:52.412501 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2026-03-16 18:20:52.792063 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2026-03-16 18:20:53.165884 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2026-03-16 18:20:53.538504 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2026-03-16 18:20:53.914767 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2026-03-16 18:20:54.290292 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2026-03-16 18:20:54.694151 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2026-03-16 18:20:55.103364 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2026-03-16 18:20:55.478578 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2026-03-16 18:20:55.909375 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2026-03-16 18:20:56.298649 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2026-03-16 18:20:56.675003 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2026-03-16 18:20:57.053057 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2026-03-16 18:20:57.435095 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2026-03-16 18:20:57.843777 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2026-03-16 18:20:58.251863 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2026-03-16 18:20:58.649011 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2026-03-16 18:20:59.041723 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2026-03-16 18:20:59.441337 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2026-03-16 18:20:59.828367 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2026-03-16 18:21:00.218115 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2026-03-16 18:21:00.606341 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2026-03-16 18:21:00.992203 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2026-03-16 18:21:01.378001 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2026-03-16 18:21:01.76946 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2026-03-16 18:21:02.156073 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2026-03-16 18:21:02.539629 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2026-03-16 18:21:02.963301 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2026-03-16 18:21:06.964631 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2026-03-16 18:21:06.966803 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2026-03-16 18:21:07.342016 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2026-03-16 18:21:07.739865 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2026-03-16 18:21:08.140959 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2026-03-16 18:21:08.566222 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2026-03-16 18:21:08.973916 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-03-16 18:21:09.381896 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2026-03-16 18:21:09.783759 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2026-03-16 18:21:10.188957 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2026-03-16 18:21:10.578314 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2026-03-16 18:21:10.972289 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2026-03-16 18:21:11.713276 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2026-03-16 18:21:12.071141 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2026-03-16 18:21:12.427614 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2026-03-16 18:21:12.790568 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2026-03-16 18:21:13.149492 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2026-03-16 18:21:13.496313 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2026-03-16 18:21:13.849284 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2026-03-16 18:21:14.218242 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2026-03-16 18:21:14.570629 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2026-03-16 18:21:14.927127 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2026-03-16 18:21:15.290684 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2026-03-16 18:21:15.661191 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2026-03-16 18:21:16.023397 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2026-03-16 18:21:16.38101 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2026-03-16 18:21:16.75389 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2026-03-16 18:21:17.139571 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2026-03-16 18:21:17.492247 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2026-03-16 18:21:17.849501 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2026-03-16 18:21:21.606164 INFO::Plotting data for metadata number 4, antibiotics
## 2026-03-16 18:21:21.6081 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2026-03-16 18:21:21.943639 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2026-03-16 18:21:22.312082 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2026-03-16 18:21:22.699376 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2026-03-16 18:21:23.065943 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2026-03-16 18:21:23.419449 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2026-03-16 18:21:23.773469 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2026-03-16 18:21:24.131245 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2026-03-16 18:21:24.478645 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2026-03-16 18:21:24.827723 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2026-03-16 18:21:25.184997 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2026-03-16 18:21:25.527385 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2026-03-16 18:21:25.878712 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2026-03-16 18:21:26.242734 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2026-03-16 18:21:26.598383 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2026-03-16 18:21:26.954957 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-03-16 18:21:27.317338 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2026-03-16 18:21:27.761402 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2026-03-16 18:21:28.123524 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2026-03-16 18:21:28.476623 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2026-03-16 18:21:28.847252 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2026-03-16 18:21:29.213283 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2026-03-16 18:21:29.576708 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2026-03-16 18:21:29.955608 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2026-03-16 18:21:30.318283 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2026-03-16 18:21:30.681716 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2026-03-16 18:21:31.035183 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2026-03-16 18:21:31.397354 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2026-03-16 18:21:31.753396 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2026-03-16 18:21:32.121337 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2026-03-16 18:21:32.478957 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2026-03-16 18:21:32.838832 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2026-03-16 18:21:33.249612 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2026-03-16 18:21:33.612374 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2026-03-16 18:21:33.976843 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2026-03-16 18:21:34.336455 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2026-03-16 18:21:34.744199 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2026-03-16 18:21:35.121096 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2026-03-16 18:21:35.498729 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2026-03-16 18:21:35.875685 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2026-03-16 18:21:39.62172 INFO::Plotting data for metadata number 5, age
## 2026-03-16 18:21:39.623767 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()`).
## 2026-03-16 18:21:40.075243 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()`).
## 2026-03-16 18:21:40.551785 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()`).
## 2026-03-16 18:21:40.900799 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()`).
## 2026-03-16 18:21:41.277233 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()`).
## 2026-03-16 18:21:41.632432 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()`).
## 2026-03-16 18:21:41.986708 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()`).
## 2026-03-16 18:21:42.346703 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()`).
## 2026-03-16 18:21:42.694836 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()`).
## 2026-03-16 18:21:43.106378 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()`).
## 2026-03-16 18:21:43.47011 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()`).
## 2026-03-16 18:21:43.819329 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()`).
## 2026-03-16 18:21:44.168075 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()`).
## 2026-03-16 18:21:44.519207 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()`).
## 2026-03-16 18:21:44.865189 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()`).
## 2026-03-16 18:21:45.213322 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()`).
## 2026-03-16 18:21:45.593733 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()`).
## 2026-03-16 18:21:45.938257 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()`).
## 2026-03-16 18:21:46.288932 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()`).
## 2026-03-16 18:21:46.655041 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()`).
## 2026-03-16 18:21:47.021736 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()`).
## 2026-03-16 18:21:47.376458 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()`).
## 2026-03-16 18:21:51.199454 INFO::Plotting data for metadata number 6, diagnosis
## 2026-03-16 18:21:51.2016 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2026-03-16 18:21:51.58786 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2026-03-16 18:21:51.983667 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2026-03-16 18:21:52.385895 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2026-03-16 18:21:52.815448 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2026-03-16 18:21:53.225064 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2026-03-16 18:21:53.627622 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2026-03-16 18:21:54.076977 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2026-03-16 18:21:54.488481 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2026-03-16 18:21:54.891238 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2026-03-16 18:21:55.315418 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2026-03-16 18:21:55.704068 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2026-03-16 18:21:56.468944 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2026-03-16 18:21:56.82601 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2026-03-16 18:21:57.184017 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2026-03-16 18:21:57.540288 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2026-03-16 18:21:57.890252 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2026-03-16 18:21:58.250535 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2026-03-16 18:21:58.611055 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2026-03-16 18:21:58.976959 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2026-03-16 18:21:59.333726 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2026-03-16 18:21:59.697365 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2026-03-16 18:22:00.055556 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2026-03-16 18:22:00.416983 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2026-03-16 18:22:00.774602 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2026-03-16 18:22:01.140668 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2026-03-16 18:22:01.491498 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2026-03-16 18:22:01.856256 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2026-03-16 18:22:02.215787 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2026-03-16 18:22:02.575897 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2026-03-16 18:22:02.937832 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2026-03-16 18:22:03.331736 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2026-03-16 18:22:03.730124 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2026-03-16 18:22:04.103925 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2026-03-16 18:22:04.470524 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2026-03-16 18:22:04.84293 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2026-03-16 18:22:05.211253 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2026-03-16 18:22:05.58362 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2026-03-16 18:22:05.960933 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2026-03-16 18:22:06.32636 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2026-03-16 18:22:06.696307 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2026-03-16 18:22:07.066606 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2026-03-16 18:22:07.434406 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 4.5.2 (2025-10-31)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04.3 LTS
## 
## Matrix products: default
## BLAS:   /home/biocbuild/bbs-3.22-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_1.24.1
## 
## loaded via a namespace (and not attached):
##  [1] gtable_0.3.6        biglm_0.9-3         xfun_0.56          
##  [4] bslib_0.10.0        ggplot2_4.0.2       lattice_0.22-9     
##  [7] numDeriv_2016.8-1.1 vctrs_0.7.1         tools_4.5.2        
## [10] Rdpack_2.6.6        generics_0.1.4      parallel_4.5.2     
## [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_1.7-4        data.table_1.18.2.1
## [22] RColorBrewer_1.1-3  S7_0.2.1            lifecycle_1.0.5    
## [25] compiler_4.5.2      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_3.1-168       
## [43] robustbase_0.99-7   tidyselect_1.2.1    digest_0.6.39      
## [46] mvtnorm_1.3-6       dplyr_1.2.0         labeling_0.4.3     
## [49] splines_4.5.2       pcaPP_2.0-5         fastmap_1.2.0      
## [52] grid_4.5.2          cli_3.6.5           magrittr_2.0.4     
## [55] dichromat_2.0-0.1   withr_3.0.2         scales_1.4.0       
## [58] rmarkdown_2.30      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_1.1.7        
## [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.