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"
## 2026-04-28 19:29:47.330024 INFO::Writing function arguments to log file
## 2026-04-28 19:29:47.36075 INFO::Verifying options selected are valid
## 2026-04-28 19:29:47.392001 INFO::Determining format of input files
## 2026-04-28 19:29:47.393202 INFO::Input format is data samples as rows and metadata samples as rows
## 2026-04-28 19:29:47.397567 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2026-04-28 19:29:47.398628 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2026-04-28 19:29:47.400766 INFO::Filter data based on min abundance and min prevalence
## 2026-04-28 19:29:47.401493 INFO::Total samples in data: 1595
## 2026-04-28 19:29:47.402182 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2026-04-28 19:29:47.406983 INFO::Total filtered features: 0
## 2026-04-28 19:29:47.407868 INFO::Filtered feature names from abundance and prevalence filtering:
## 2026-04-28 19:29:47.416032 INFO::Total filtered features with variance filtering: 0
## 2026-04-28 19:29:47.416878 INFO::Filtered feature names from variance filtering:
## 2026-04-28 19:29:47.41757 INFO::Running selected normalization method: TSS
## 2026-04-28 19:29:48.455164 INFO::Bypass z-score application to metadata
## 2026-04-28 19:29:48.456205 INFO::Running selected transform method: LOG
## 2026-04-28 19:29:48.469737 INFO::Running selected analysis method: LM
## 2026-04-28 19:29:48.794626 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:49.668941 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2026-04-28 19:29:49.825326 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2026-04-28 19:29:49.980524 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:50.122346 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2026-04-28 19:29:50.272139 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:50.405885 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2026-04-28 19:29:50.548736 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2026-04-28 19:29:50.694936 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2026-04-28 19:29:50.831911 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2026-04-28 19:29:50.974255 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
## 2026-04-28 19:29:51.090184 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
## 2026-04-28 19:29:51.226056 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:51.357405 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:51.499542 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2026-04-28 19:29:51.633568 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:51.760178 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2026-04-28 19:29:51.901733 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2026-04-28 19:29:52.038009 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:52.168746 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:52.307794 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:52.458668 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:52.608238 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2026-04-28 19:29:52.753126 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2026-04-28 19:29:52.892101 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2026-04-28 19:29:53.036978 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2026-04-28 19:29:53.174055 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:53.308899 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2026-04-28 19:29:53.451688 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2026-04-28 19:29:53.59032 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:53.962062 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2026-04-28 19:29:54.099298 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2026-04-28 19:29:54.23482 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:54.366798 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:54.513211 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:54.68102 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:54.820995 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2026-04-28 19:29:54.965351 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2026-04-28 19:29:55.097042 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2026-04-28 19:29:55.244082 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:55.389143 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
## 2026-04-28 19:29:55.500635 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
## 2026-04-28 19:29:55.631811 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:55.759888 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:55.898281 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2026-04-28 19:29:56.043581 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2026-04-28 19:29:56.175401 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:56.305198 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:56.437959 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2026-04-28 19:29:56.57763 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2026-04-28 19:29:56.717025 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:56.855004 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:56.984318 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:57.120445 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:57.243458 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:57.394429 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2026-04-28 19:29:57.544957 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2026-04-28 19:29:57.694381 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2026-04-28 19:29:57.830421 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:57.964264 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:58.104585 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:58.243936 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:58.378403 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2026-04-28 19:29:58.506886 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:58.636858 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:58.77466 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2026-04-28 19:29:58.917326 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2026-04-28 19:29:59.049955 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2026-04-28 19:29:59.186821 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:59.318202 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2026-04-28 19:29:59.449376 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2026-04-28 19:29:59.587633 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:29:59.720818 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2026-04-28 19:29:59.856171 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2026-04-28 19:29:59.99506 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:30:00.12378 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2026-04-28 19:30:00.286614 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:30:00.427557 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:30:00.561219 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2026-04-28 19:30:00.693835 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2026-04-28 19:30:00.85785 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2026-04-28 19:30:00.990036 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2026-04-28 19:30:01.131837 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2026-04-28 19:30:01.281758 INFO::Fitting model to feature number 82, Escherichia.coli
## 2026-04-28 19:30:01.432189 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2026-04-28 19:30:01.561588 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:30:01.692605 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2026-04-28 19:30:01.826822 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2026-04-28 19:30:01.966563 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2026-04-28 19:30:02.15318 INFO::Counting total values for each feature
## 2026-04-28 19:30:02.182193 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2026-04-28 19:30:02.278404 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2026-04-28 19:30:02.376287 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2026-04-28 19:30:02.553384 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2026-04-28 19:30:02.609139 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2026-04-28 19:30:02.641248 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2026-04-28 19:30:02.646355 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2026-04-28 19:30:02.651181 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
## 2026-04-28 19:30:02.653493 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2026-04-28 19:30:02.853535 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2026-04-28 19:30:02.859111 INFO::Plotting associations from most to least significant, grouped by metadata
## 2026-04-28 19:30:02.860062 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2026-04-28 19:30:02.86156 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2026-04-28 19:30:03.328954 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2026-04-28 19:30:03.71996 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2026-04-28 19:30:04.083658 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2026-04-28 19:30:04.447459 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2026-04-28 19:30:04.803158 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2026-04-28 19:30:05.166303 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2026-04-28 19:30:05.539766 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2026-04-28 19:30:05.943458 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2026-04-28 19:30:06.31145 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2026-04-28 19:30:06.681333 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2026-04-28 19:30:07.038922 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2026-04-28 19:30:07.400564 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2026-04-28 19:30:07.766993 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2026-04-28 19:30:08.178301 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2026-04-28 19:30:09.785198 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2026-04-28 19:30:11.120627 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2026-04-28 19:30:11.776716 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2026-04-28 19:30:12.13545 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2026-04-28 19:30:12.501318 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-04-28 19:30:12.866182 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2026-04-28 19:30:13.293003 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2026-04-28 19:30:13.657092 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2026-04-28 19:30:14.017617 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2026-04-28 19:30:14.383597 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2026-04-28 19:30:14.749127 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2026-04-28 19:30:15.117642 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2026-04-28 19:30:15.487799 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2026-04-28 19:30:15.85758 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2026-04-28 19:30:16.214834 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2026-04-28 19:30:16.57534 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2026-04-28 19:30:16.945009 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2026-04-28 19:30:17.310386 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2026-04-28 19:30:17.66924 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2026-04-28 19:30:18.036687 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2026-04-28 19:30:18.402374 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2026-04-28 19:30:18.760847 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2026-04-28 19:30:19.12958 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2026-04-28 19:30:19.495716 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2026-04-28 19:30:19.859554 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2026-04-28 19:30:20.222075 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2026-04-28 19:30:20.59323 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2026-04-28 19:30:20.962128 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2026-04-28 19:30:21.327468 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2026-04-28 19:30:22.019464 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2026-04-28 19:30:22.378061 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2026-04-28 19:30:22.734214 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2026-04-28 19:30:23.091167 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2026-04-28 19:30:23.482308 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2026-04-28 19:30:23.841637 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2026-04-28 19:30:24.191537 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2026-04-28 19:30:24.545501 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2026-04-28 19:30:24.908371 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2026-04-28 19:30:25.26291 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2026-04-28 19:30:25.609789 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2026-04-28 19:30:25.963004 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2026-04-28 19:30:26.318627 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2026-04-28 19:30:26.66888 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2026-04-28 19:30:27.013655 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2026-04-28 19:30:27.367434 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2026-04-28 19:30:27.718225 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2026-04-28 19:30:28.116944 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2026-04-28 19:30:28.469357 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2026-04-28 19:30:32.213163 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2026-04-28 19:30:32.214974 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2026-04-28 19:30:32.552137 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2026-04-28 19:30:32.926019 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2026-04-28 19:30:33.298667 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2026-04-28 19:30:33.668039 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2026-04-28 19:30:34.045217 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2026-04-28 19:30:34.420142 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2026-04-28 19:30:34.777133 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2026-04-28 19:30:35.15316 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2026-04-28 19:30:35.523988 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2026-04-28 19:30:35.891762 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2026-04-28 19:30:36.270085 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2026-04-28 19:30:36.641079 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-04-28 19:30:37.043034 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2026-04-28 19:30:37.431075 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2026-04-28 19:30:37.805752 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2026-04-28 19:30:38.176208 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2026-04-28 19:30:38.544682 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2026-04-28 19:30:38.913139 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2026-04-28 19:30:39.336898 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2026-04-28 19:30:39.710417 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2026-04-28 19:30:40.08262 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2026-04-28 19:30:40.452137 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2026-04-28 19:30:40.822923 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2026-04-28 19:30:41.195266 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2026-04-28 19:30:41.565134 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2026-04-28 19:30:41.93742 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2026-04-28 19:30:42.301311 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2026-04-28 19:30:42.685326 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2026-04-28 19:30:43.080793 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2026-04-28 19:30:43.4489 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2026-04-28 19:30:43.811032 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2026-04-28 19:30:44.190025 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2026-04-28 19:30:44.556525 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2026-04-28 19:30:44.949856 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2026-04-28 19:30:45.322988 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2026-04-28 19:30:45.696748 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2026-04-28 19:30:46.068562 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2026-04-28 19:30:46.443836 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2026-04-28 19:30:46.853723 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2026-04-28 19:30:47.235273 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2026-04-28 19:30:47.621524 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2026-04-28 19:30:48.005017 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2026-04-28 19:30:48.392961 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2026-04-28 19:30:52.253981 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2026-04-28 19:30:52.255887 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2026-04-28 19:30:52.608263 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2026-04-28 19:30:52.995113 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2026-04-28 19:30:53.377787 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2026-04-28 19:30:53.750152 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2026-04-28 19:30:54.129832 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-04-28 19:30:54.515305 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2026-04-28 19:30:54.911597 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2026-04-28 19:30:55.347257 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2026-04-28 19:30:55.728344 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2026-04-28 19:30:56.107889 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2026-04-28 19:30:56.489862 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2026-04-28 19:30:56.881048 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2026-04-28 19:30:57.264035 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2026-04-28 19:30:57.648678 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2026-04-28 19:30:58.031496 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2026-04-28 19:30:58.401816 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2026-04-28 19:30:58.775335 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2026-04-28 19:30:59.156283 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2026-04-28 19:30:59.53107 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2026-04-28 19:30:59.922459 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2026-04-28 19:31:00.308362 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2026-04-28 19:31:00.691325 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2026-04-28 19:31:01.070425 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2026-04-28 19:31:01.446536 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2026-04-28 19:31:01.846341 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2026-04-28 19:31:02.229214 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2026-04-28 19:31:02.643767 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2026-04-28 19:31:03.040774 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2026-04-28 19:31:06.97967 INFO::Plotting data for metadata number 4, antibiotics
## 2026-04-28 19:31:06.981595 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2026-04-28 19:31:07.330527 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2026-04-28 19:31:07.709718 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2026-04-28 19:31:08.087502 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2026-04-28 19:31:08.470011 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2026-04-28 19:31:08.847837 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2026-04-28 19:31:09.230093 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2026-04-28 19:31:09.605307 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2026-04-28 19:31:09.988166 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2026-04-28 19:31:10.369567 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2026-04-28 19:31:11.073941 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2026-04-28 19:31:11.411169 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2026-04-28 19:31:11.742452 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2026-04-28 19:31:12.087655 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2026-04-28 19:31:12.429061 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2026-04-28 19:31:12.769018 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2026-04-28 19:31:13.109242 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2026-04-28 19:31:13.455933 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2026-04-28 19:31:13.784888 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2026-04-28 19:31:14.117184 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2026-04-28 19:31:14.458603 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2026-04-28 19:31:14.799786 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2026-04-28 19:31:15.138982 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2026-04-28 19:31:15.48269 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2026-04-28 19:31:15.823713 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2026-04-28 19:31:16.167413 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2026-04-28 19:31:16.507733 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2026-04-28 19:31:16.844237 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2026-04-28 19:31:17.199946 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2026-04-28 19:31:17.56843 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2026-04-28 19:31:17.908477 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2026-04-28 19:31:18.249537 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2026-04-28 19:31:18.590963 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2026-04-28 19:31:18.934458 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2026-04-28 19:31:19.308486 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2026-04-28 19:31:19.661046 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2026-04-28 19:31:20.009374 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2026-04-28 19:31:20.366665 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2026-04-28 19:31:20.704546 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2026-04-28 19:31:21.054632 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2026-04-28 19:31:24.61267 INFO::Plotting data for metadata number 5, age
## 2026-04-28 19:31:24.614438 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()`).
## 2026-04-28 19:31:25.068805 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()`).
## 2026-04-28 19:31:25.503469 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()`).
## 2026-04-28 19:31:25.831311 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()`).
## 2026-04-28 19:31:26.177652 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()`).
## 2026-04-28 19:31:26.508923 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()`).
## 2026-04-28 19:31:26.839864 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()`).
## 2026-04-28 19:31:27.182795 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()`).
## 2026-04-28 19:31:27.517656 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()`).
## 2026-04-28 19:31:27.857269 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()`).
## 2026-04-28 19:31:28.254159 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()`).
## 2026-04-28 19:31:28.586095 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()`).
## 2026-04-28 19:31:28.923114 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()`).
## 2026-04-28 19:31:29.266145 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()`).
## 2026-04-28 19:31:29.601756 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()`).
## 2026-04-28 19:31:29.937541 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()`).
## 2026-04-28 19:31:30.280878 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()`).
## 2026-04-28 19:31:30.612152 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()`).
## 2026-04-28 19:31:30.945227 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()`).
## 2026-04-28 19:31:31.276765 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()`).
## 2026-04-28 19:31:31.617393 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()`).
## 2026-04-28 19:31:31.954328 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()`).
## 2026-04-28 19:31:35.52997 INFO::Plotting data for metadata number 6, diagnosis
## 2026-04-28 19:31:35.531825 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2026-04-28 19:31:35.9117 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2026-04-28 19:31:36.294931 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2026-04-28 19:31:36.680395 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2026-04-28 19:31:37.072263 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2026-04-28 19:31:37.453171 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2026-04-28 19:31:37.831431 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2026-04-28 19:31:38.208512 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2026-04-28 19:31:38.594429 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2026-04-28 19:31:38.972799 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2026-04-28 19:31:39.357587 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2026-04-28 19:31:39.755371 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2026-04-28 19:31:40.17112 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2026-04-28 19:31:40.547912 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2026-04-28 19:31:40.927578 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2026-04-28 19:31:41.308641 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2026-04-28 19:31:41.68011 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2026-04-28 19:31:42.074793 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2026-04-28 19:31:42.458709 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2026-04-28 19:31:42.840306 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2026-04-28 19:31:43.215672 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2026-04-28 19:31:43.665854 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2026-04-28 19:31:44.049121 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2026-04-28 19:31:44.441497 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2026-04-28 19:31:44.835951 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2026-04-28 19:31:45.22851 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2026-04-28 19:31:45.611553 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2026-04-28 19:31:45.999577 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2026-04-28 19:31:46.381925 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2026-04-28 19:31:46.766163 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2026-04-28 19:31:47.152572 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2026-04-28 19:31:47.529658 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2026-04-28 19:31:47.927558 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2026-04-28 19:31:48.31479 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2026-04-28 19:31:48.698679 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2026-04-28 19:31:49.088264 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2026-04-28 19:31:49.485057 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2026-04-28 19:31:49.918951 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2026-04-28 19:31:50.309465 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2026-04-28 19:31:50.74178 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2026-04-28 19:31:51.151949 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2026-04-28 19:31:51.564925 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2026-04-28 19:31:51.981351 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 RC (2026-04-17 r89917)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04.4 LTS
## 
## Matrix products: default
## BLAS:   /home/biocbuild/bbs-3.24-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_1.27.0
## 
## 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_4.0.3       lattice_0.22-9     
##  [7] numDeriv_2016.8-1.1 vctrs_0.7.3         tools_4.6.0        
## [10] Rdpack_2.6.6        generics_0.1.4      parallel_4.6.0     
## [13] tibble_3.3.1        DEoptimR_1.1-4      cluster_2.1.8.2    
## [16] pkgconfig_2.0.3     logging_0.10-108    pheatmap_1.0.13    
## [19] Matrix_1.7-5        data.table_1.18.2.1 RColorBrewer_1.1-3 
## [22] S7_0.2.2            lifecycle_1.0.5     compiler_4.6.0     
## [25] farver_2.1.2        lmerTest_3.2-1      permute_0.9-10     
## [28] htmltools_0.5.9     sass_0.4.10         hash_2.2.6.4       
## [31] yaml_2.3.12         pillar_1.11.1       nloptr_2.2.1       
## [34] crayon_1.5.3        jquerylib_0.1.4     MASS_7.3-65        
## [37] cachem_1.1.0        vegan_2.7-3         reformulas_0.4.4   
## [40] boot_1.3-32         nlme_3.1-169        robustbase_0.99-7  
## [43] tidyselect_1.2.1    digest_0.6.39       mvtnorm_1.3-7      
## [46] dplyr_1.2.1         labeling_0.4.3      splines_4.6.0      
## [49] pcaPP_2.0-5         fastmap_1.2.0       grid_4.6.0         
## [52] cli_3.6.6           magrittr_2.0.5      dichromat_2.0-0.1  
## [55] withr_3.0.2         scales_1.4.0        rmarkdown_2.31     
## [58] otel_0.2.0          lme4_2.0-1          pbapply_1.7-4      
## [61] evaluate_1.0.5      knitr_1.51          rbibutils_2.4.1    
## [64] mgcv_1.9-4          rlang_1.2.0         Rcpp_1.1.1-1.1     
## [67] glue_1.8.1          optparse_1.8.2      DBI_1.3.0          
## [70] minqa_1.2.8         jsonlite_2.0.0      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.