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"
## 2025-04-15 17:59:44.532149 INFO::Writing function arguments to log file
## 2025-04-15 17:59:44.541123 INFO::Verifying options selected are valid
## 2025-04-15 17:59:44.554925 INFO::Determining format of input files
## 2025-04-15 17:59:44.559353 INFO::Input format is data samples as rows and metadata samples as rows
## 2025-04-15 17:59:44.561484 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2025-04-15 17:59:44.561907 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2025-04-15 17:59:44.562706 INFO::Filter data based on min abundance and min prevalence
## 2025-04-15 17:59:44.56296 INFO::Total samples in data: 1595
## 2025-04-15 17:59:44.563197 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2025-04-15 17:59:44.564972 INFO::Total filtered features: 0
## 2025-04-15 17:59:44.565287 INFO::Filtered feature names from abundance and prevalence filtering:
## 2025-04-15 17:59:44.568416 INFO::Total filtered features with variance filtering: 0
## 2025-04-15 17:59:44.568711 INFO::Filtered feature names from variance filtering:
## 2025-04-15 17:59:44.568962 INFO::Running selected normalization method: TSS
## 2025-04-15 17:59:45.060624 INFO::Bypass z-score application to metadata
## 2025-04-15 17:59:45.061482 INFO::Running selected transform method: LOG
## 2025-04-15 17:59:45.070287 INFO::Running selected analysis method: LM
## 2025-04-15 17:59:45.198865 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:45.339203 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2025-04-15 17:59:45.399147 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2025-04-15 17:59:45.503163 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:45.698755 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2025-04-15 17:59:45.758897 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:45.817795 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2025-04-15 17:59:45.878705 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2025-04-15 17:59:45.936355 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2025-04-15 17:59:45.992701 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2025-04-15 17:59:46.051044 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: -4.2e+01
## 2025-04-15 17:59:46.102766 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: -4.2e+01
## 2025-04-15 17:59:46.158484 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:46.213103 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:46.282941 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2025-04-15 17:59:46.338955 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:46.395316 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2025-04-15 17:59:46.452151 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2025-04-15 17:59:46.510972 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:46.563315 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:46.616361 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:46.674961 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:46.730075 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2025-04-15 17:59:46.783763 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2025-04-15 17:59:46.841128 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2025-04-15 17:59:46.895868 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2025-04-15 17:59:46.94804 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.005552 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2025-04-15 17:59:47.060212 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2025-04-15 17:59:47.115508 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.17594 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2025-04-15 17:59:47.233302 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2025-04-15 17:59:47.288001 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.350866 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.406936 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## Feature Alistipes.onderdonkii : simpleWarning: Model failed to converge with 1 negative eigenvalue: -2.3e+00
## 2025-04-15 17:59:47.46069 WARNING::Fitting problem for feature 34 a warning was issued
## boundary (singular) fit: see help('isSingular')
## Warning: Model failed to converge with 1 negative eigenvalue: -2.3e+00
## 2025-04-15 17:59:47.517542 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.573493 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2025-04-15 17:59:47.634423 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2025-04-15 17:59:47.687254 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2025-04-15 17:59:47.74811 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.804615 INFO::Fitting model to feature number 40, Clostridium.citroniae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.860701 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.919173 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:47.974636 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2025-04-15 17:59:48.15269 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2025-04-15 17:59:48.207844 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.264195 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.324528 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2025-04-15 17:59:48.377742 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2025-04-15 17:59:48.432552 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.489883 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.546815 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.600545 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.652602 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.710996 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2025-04-15 17:59:48.762567 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2025-04-15 17:59:48.814855 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2025-04-15 17:59:48.875535 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.929968 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:48.98419 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.041917 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.097355 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2025-04-15 17:59:49.147687 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.205932 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.257033 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2025-04-15 17:59:49.311904 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2025-04-15 17:59:49.369454 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2025-04-15 17:59:49.423773 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.479875 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2025-04-15 17:59:49.531161 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2025-04-15 17:59:49.585784 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.642921 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2025-04-15 17:59:49.69218 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2025-04-15 17:59:49.750102 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.796883 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2025-04-15 17:59:49.859902 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.914567 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:49.973456 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2025-04-15 17:59:50.026773 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2025-04-15 17:59:50.082344 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2025-04-15 17:59:50.253728 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2025-04-15 17:59:50.307293 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2025-04-15 17:59:50.368059 INFO::Fitting model to feature number 82, Escherichia.coli
## 2025-04-15 17:59:50.542693 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2025-04-15 17:59:50.592464 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:50.64386 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 17:59:50.699188 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2025-04-15 17:59:50.760497 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2025-04-15 17:59:50.837463 INFO::Counting total values for each feature
## 2025-04-15 17:59:50.850347 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2025-04-15 17:59:50.899521 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2025-04-15 17:59:50.956937 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2025-04-15 17:59:51.035809 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2025-04-15 17:59:51.065919 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2025-04-15 17:59:51.082804 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2025-04-15 17:59:51.08584 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2025-04-15 17:59:51.088841 INFO::Writing the significant results (those which are less than or equal to the threshold of 0.250000 ) to file (ordered by increasing q-values): demo_output/significant_results.tsv
## 2025-04-15 17:59:51.090155 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2025-04-15 17:59:51.149666 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2025-04-15 17:59:51.152406 INFO::Plotting associations from most to least significant, grouped by metadata
## 2025-04-15 17:59:51.152851 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2025-04-15 17:59:51.153526 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2025-04-15 17:59:51.273593 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2025-04-15 17:59:51.376862 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2025-04-15 17:59:51.466338 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2025-04-15 17:59:51.559185 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2025-04-15 17:59:51.646142 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2025-04-15 17:59:51.739687 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2025-04-15 17:59:51.827705 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2025-04-15 17:59:51.923777 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2025-04-15 17:59:52.016835 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2025-04-15 17:59:52.111552 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2025-04-15 17:59:52.200908 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2025-04-15 17:59:52.294045 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2025-04-15 17:59:52.385097 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2025-04-15 17:59:52.476709 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2025-04-15 17:59:52.567625 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2025-04-15 17:59:52.659872 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2025-04-15 17:59:52.75058 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2025-04-15 17:59:52.844747 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2025-04-15 17:59:52.934058 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-04-15 17:59:53.028414 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2025-04-15 17:59:53.123865 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2025-04-15 17:59:53.211079 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2025-04-15 17:59:53.300427 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2025-04-15 17:59:53.392496 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2025-04-15 17:59:53.484882 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2025-04-15 17:59:53.570685 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2025-04-15 17:59:53.661535 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2025-04-15 17:59:53.748844 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2025-04-15 17:59:53.839046 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2025-04-15 17:59:53.922359 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2025-04-15 17:59:54.012556 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2025-04-15 17:59:54.101521 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2025-04-15 17:59:54.182418 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2025-04-15 17:59:54.311266 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2025-04-15 17:59:54.401404 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2025-04-15 17:59:54.49256 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2025-04-15 17:59:54.578899 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2025-04-15 17:59:54.672872 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2025-04-15 17:59:54.762336 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2025-04-15 17:59:54.854429 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2025-04-15 17:59:54.94608 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2025-04-15 17:59:55.046171 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2025-04-15 17:59:55.137897 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2025-04-15 17:59:55.234911 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2025-04-15 17:59:55.327303 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2025-04-15 17:59:55.428703 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2025-04-15 17:59:55.521097 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2025-04-15 17:59:55.612422 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2025-04-15 17:59:55.706472 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2025-04-15 17:59:55.789879 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2025-04-15 17:59:55.88374 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2025-04-15 17:59:55.97737 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2025-04-15 17:59:56.07771 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2025-04-15 17:59:56.163845 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2025-04-15 17:59:56.260305 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2025-04-15 17:59:56.348539 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2025-04-15 17:59:56.441955 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2025-04-15 17:59:56.525981 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2025-04-15 17:59:56.619658 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2025-04-15 17:59:56.712976 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2025-04-15 17:59:56.806622 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2025-04-15 17:59:56.901085 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2025-04-15 17:59:57.860943 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2025-04-15 17:59:57.861721 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2025-04-15 17:59:57.970735 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2025-04-15 17:59:58.067626 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2025-04-15 17:59:58.169818 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2025-04-15 17:59:58.264586 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2025-04-15 17:59:58.372942 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2025-04-15 17:59:58.465476 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2025-04-15 17:59:58.565398 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2025-04-15 17:59:58.655032 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2025-04-15 17:59:58.760472 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2025-04-15 17:59:58.848253 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2025-04-15 17:59:58.951631 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2025-04-15 17:59:59.063314 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-04-15 17:59:59.189943 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2025-04-15 17:59:59.301902 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2025-04-15 17:59:59.392153 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2025-04-15 17:59:59.493484 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2025-04-15 17:59:59.580655 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2025-04-15 17:59:59.676133 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2025-04-15 17:59:59.760753 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2025-04-15 17:59:59.858127 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2025-04-15 17:59:59.945631 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2025-04-15 18:00:00.043116 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2025-04-15 18:00:00.137813 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2025-04-15 18:00:00.227939 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2025-04-15 18:00:00.329311 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2025-04-15 18:00:00.421602 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2025-04-15 18:00:00.512046 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2025-04-15 18:00:00.595314 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2025-04-15 18:00:00.689874 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2025-04-15 18:00:00.774971 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2025-04-15 18:00:00.865628 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2025-04-15 18:00:00.966753 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2025-04-15 18:00:01.054551 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2025-04-15 18:00:01.16158 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2025-04-15 18:00:01.249737 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2025-04-15 18:00:01.352385 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2025-04-15 18:00:01.442113 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2025-04-15 18:00:01.544049 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2025-04-15 18:00:01.682055 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2025-04-15 18:00:01.778973 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2025-04-15 18:00:01.891169 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2025-04-15 18:00:01.98583 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2025-04-15 18:00:02.090559 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2025-04-15 18:00:03.072683 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2025-04-15 18:00:03.073541 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2025-04-15 18:00:03.155689 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2025-04-15 18:00:03.254143 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2025-04-15 18:00:03.359136 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2025-04-15 18:00:03.44791 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2025-04-15 18:00:03.555501 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-04-15 18:00:03.656821 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2025-04-15 18:00:03.760076 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2025-04-15 18:00:03.856055 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2025-04-15 18:00:03.96091 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2025-04-15 18:00:04.050296 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2025-04-15 18:00:04.151735 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2025-04-15 18:00:04.252158 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2025-04-15 18:00:04.342313 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2025-04-15 18:00:04.444599 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2025-04-15 18:00:04.536677 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2025-04-15 18:00:04.628144 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2025-04-15 18:00:04.71377 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2025-04-15 18:00:04.814845 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2025-04-15 18:00:04.909898 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2025-04-15 18:00:05.001861 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2025-04-15 18:00:05.110832 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2025-04-15 18:00:05.20892 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2025-04-15 18:00:05.30818 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2025-04-15 18:00:05.434008 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2025-04-15 18:00:05.534996 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2025-04-15 18:00:05.636555 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2025-04-15 18:00:05.724679 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2025-04-15 18:00:05.833058 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2025-04-15 18:00:06.839732 INFO::Plotting data for metadata number 4, antibiotics
## 2025-04-15 18:00:06.840501 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2025-04-15 18:00:06.905572 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2025-04-15 18:00:07.006992 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2025-04-15 18:00:07.098189 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2025-04-15 18:00:07.196908 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2025-04-15 18:00:07.291462 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2025-04-15 18:00:07.390815 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2025-04-15 18:00:07.495658 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2025-04-15 18:00:07.584915 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2025-04-15 18:00:07.68778 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2025-04-15 18:00:07.777836 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2025-04-15 18:00:07.876557 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2025-04-15 18:00:07.960892 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2025-04-15 18:00:08.069398 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2025-04-15 18:00:08.16362 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2025-04-15 18:00:08.262032 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-04-15 18:00:08.344747 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2025-04-15 18:00:08.438938 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2025-04-15 18:00:08.516843 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2025-04-15 18:00:08.609203 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2025-04-15 18:00:08.704426 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2025-04-15 18:00:08.805049 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2025-04-15 18:00:08.906901 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2025-04-15 18:00:08.998263 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2025-04-15 18:00:09.102611 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2025-04-15 18:00:09.187988 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2025-04-15 18:00:09.326937 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2025-04-15 18:00:09.417932 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2025-04-15 18:00:09.517669 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2025-04-15 18:00:09.60964 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2025-04-15 18:00:09.715025 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2025-04-15 18:00:09.811443 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2025-04-15 18:00:09.914815 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2025-04-15 18:00:10.012935 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2025-04-15 18:00:10.123412 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2025-04-15 18:00:10.211638 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2025-04-15 18:00:10.316878 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2025-04-15 18:00:10.407264 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2025-04-15 18:00:10.518436 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2025-04-15 18:00:10.610486 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2025-04-15 18:00:11.521066 INFO::Plotting data for metadata number 5, age
## 2025-04-15 18:00:11.522014 INFO::Creating scatter plot for continuous data, age vs Haemophilus.parainfluenzae
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:11.617088 INFO::Creating scatter plot for continuous data, age vs Bifidobacterium.pseudocatenulatum
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:11.750531 INFO::Creating scatter plot for continuous data, age vs Faecalibacterium.prausnitzii
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:11.847956 INFO::Creating scatter plot for continuous data, age vs Clostridium.clostridioforme
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:11.946172 INFO::Creating scatter plot for continuous data, age vs Veillonella.parvula
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.037203 INFO::Creating scatter plot for continuous data, age vs Subdoligranulum.unclassified
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.139199 INFO::Creating scatter plot for continuous data, age vs Clostridium.symbiosum
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.235248 INFO::Creating scatter plot for continuous data, age vs Ruminococcus.gnavus
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.329518 INFO::Creating scatter plot for continuous data, age vs Dialister.invisus
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.438025 INFO::Creating scatter plot for continuous data, age vs Veillonella.dispar
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.530271 INFO::Creating scatter plot for continuous data, age vs Veillonella.unclassified
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.623107 INFO::Creating scatter plot for continuous data, age vs Bacteroides.thetaiotaomicron
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.728502 INFO::Creating scatter plot for continuous data, age vs Ruminococcus.bromii
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:12.983308 INFO::Creating scatter plot for continuous data, age vs Bacteroides.intestinalis
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:13.060272 INFO::Creating scatter plot for continuous data, age vs Eubacterium.siraeum
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:13.139033 INFO::Creating scatter plot for continuous data, age vs Prevotella.copri
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:13.215408 INFO::Creating scatter plot for continuous data, age vs Alistipes.unclassified
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:13.292943 INFO::Creating scatter plot for continuous data, age vs Bacteroidales.bacterium.ph8
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:13.367563 INFO::Creating scatter plot for continuous data, age vs Bifidobacterium.longum
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:13.451149 INFO::Creating scatter plot for continuous data, age vs Akkermansia.muciniphila
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:13.534161 INFO::Creating scatter plot for continuous data, age vs Collinsella.aerofaciens
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:13.611429 INFO::Creating scatter plot for continuous data, age vs Parabacteroides.distasonis
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values or values outside the scale range
## (`geom_point()`).
## 2025-04-15 18:00:14.507856 INFO::Plotting data for metadata number 6, diagnosis
## 2025-04-15 18:00:14.508692 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2025-04-15 18:00:14.581084 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2025-04-15 18:00:14.666346 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2025-04-15 18:00:14.760965 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2025-04-15 18:00:14.852854 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2025-04-15 18:00:14.974191 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2025-04-15 18:00:15.059072 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2025-04-15 18:00:15.162841 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2025-04-15 18:00:15.25193 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2025-04-15 18:00:15.347049 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2025-04-15 18:00:15.448813 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2025-04-15 18:00:15.54396 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2025-04-15 18:00:15.635893 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2025-04-15 18:00:15.71927 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2025-04-15 18:00:15.80921 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2025-04-15 18:00:15.892666 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2025-04-15 18:00:15.980431 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2025-04-15 18:00:16.072142 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2025-04-15 18:00:16.156555 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2025-04-15 18:00:16.282523 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2025-04-15 18:00:16.373146 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2025-04-15 18:00:16.47158 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2025-04-15 18:00:16.562455 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2025-04-15 18:00:16.658887 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2025-04-15 18:00:16.74309 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2025-04-15 18:00:16.832018 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2025-04-15 18:00:16.925192 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2025-04-15 18:00:17.036566 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2025-04-15 18:00:17.143783 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2025-04-15 18:00:17.227919 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2025-04-15 18:00:17.321897 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2025-04-15 18:00:17.406062 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2025-04-15 18:00:17.493369 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2025-04-15 18:00:17.579591 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2025-04-15 18:00:17.669583 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2025-04-15 18:00:17.763577 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2025-04-15 18:00:17.848352 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2025-04-15 18:00:17.94453 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2025-04-15 18:00:18.036815 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2025-04-15 18:00:18.135076 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2025-04-15 18:00:18.218297 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2025-04-15 18:00:18.310188 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2025-04-15 18:00:18.403231 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 4.5.0 RC (2025-04-04 r88126)
## Platform: aarch64-apple-darwin20
## Running under: macOS Ventura 13.7.1
## 
## Matrix products: default
## BLAS:   /Library/Frameworks/R.framework/Versions/4.5-arm64/Resources/lib/libRblas.0.dylib 
## LAPACK: /Library/Frameworks/R.framework/Versions/4.5-arm64/Resources/lib/libRlapack.dylib;  LAPACK version 3.12.1
## 
## locale:
## [1] C/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8
## 
## time zone: America/New_York
## tzcode source: internal
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
## 
## other attached packages:
## [1] Maaslin2_1.22.0
## 
## loaded via a namespace (and not attached):
##  [1] gtable_0.3.6        biglm_0.9-3         xfun_0.52          
##  [4] bslib_0.9.0         ggplot2_3.5.2       lattice_0.22-7     
##  [7] numDeriv_2016.8-1.1 vctrs_0.6.5         tools_4.5.0        
## [10] Rdpack_2.6.4        generics_0.1.3      parallel_4.5.0     
## [13] getopt_1.20.4       tibble_3.2.1        DEoptimR_1.1-3-1   
## [16] cluster_2.1.8.1     pkgconfig_2.0.3     logging_0.10-108   
## [19] pheatmap_1.0.12     Matrix_1.7-3        data.table_1.17.0  
## [22] RColorBrewer_1.1-3  lifecycle_1.0.4     farver_2.1.2       
## [25] compiler_4.5.0      munsell_0.5.1       lmerTest_3.1-3     
## [28] permute_0.9-7       htmltools_0.5.8.1   sass_0.4.10        
## [31] hash_2.2.6.3        yaml_2.3.10         pillar_1.10.2      
## [34] nloptr_2.2.1        crayon_1.5.3        jquerylib_0.1.4    
## [37] MASS_7.3-65         cachem_1.1.0        vegan_2.6-10       
## [40] reformulas_0.4.0    boot_1.3-31         nlme_3.1-168       
## [43] robustbase_0.99-4-1 tidyselect_1.2.1    digest_0.6.37      
## [46] mvtnorm_1.3-3       dplyr_1.1.4         labeling_0.4.3     
## [49] splines_4.5.0       pcaPP_2.0-5         fastmap_1.2.0      
## [52] grid_4.5.0          colorspace_2.1-1    cli_3.6.4          
## [55] magrittr_2.0.3      withr_3.0.2         scales_1.3.0       
## [58] rmarkdown_2.29      lme4_1.1-37         pbapply_1.7-2      
## [61] evaluate_1.0.3      knitr_1.50          rbibutils_2.3      
## [64] mgcv_1.9-3          rlang_1.1.6         Rcpp_1.0.14        
## [67] glue_1.8.0          optparse_1.7.5      DBI_1.2.3          
## [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.