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 19:02:01.349291 INFO::Writing function arguments to log file
## 2025-04-15 19:02:01.379579 INFO::Verifying options selected are valid
## 2025-04-15 19:02:01.411057 INFO::Determining format of input files
## 2025-04-15 19:02:01.412244 INFO::Input format is data samples as rows and metadata samples as rows
## 2025-04-15 19:02:01.416535 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2025-04-15 19:02:01.417598 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2025-04-15 19:02:01.419743 INFO::Filter data based on min abundance and min prevalence
## 2025-04-15 19:02:01.420413 INFO::Total samples in data: 1595
## 2025-04-15 19:02:01.421058 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2025-04-15 19:02:01.425734 INFO::Total filtered features: 0
## 2025-04-15 19:02:01.426578 INFO::Filtered feature names from abundance and prevalence filtering:
## 2025-04-15 19:02:01.435396 INFO::Total filtered features with variance filtering: 0
## 2025-04-15 19:02:01.436258 INFO::Filtered feature names from variance filtering:
## 2025-04-15 19:02:01.436951 INFO::Running selected normalization method: TSS
## 2025-04-15 19:02:02.477456 INFO::Bypass z-score application to metadata
## 2025-04-15 19:02:02.478446 INFO::Running selected transform method: LOG
## 2025-04-15 19:02:02.493966 INFO::Running selected analysis method: LM
## 2025-04-15 19:02:02.807429 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:03.395493 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2025-04-15 19:02:03.540771 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2025-04-15 19:02:03.693344 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:03.846581 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2025-04-15 19:02:04.015507 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:04.156324 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2025-04-15 19:02:04.29556 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2025-04-15 19:02:04.444208 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2025-04-15 19:02:04.595386 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2025-04-15 19:02:04.74355 INFO::Fitting model to feature number 11, Bacteroides.finegoldii
## boundary (singular) fit: see help('isSingular')
## Feature Bacteroides.finegoldii : simpleWarning: Model failed to converge with 1 negative eigenvalue: -7.9e+01
## 2025-04-15 19:02:04.860242 WARNING::Fitting problem for feature 11 a warning was issued
## boundary (singular) fit: see help('isSingular')
## Warning: Model failed to converge with 1 negative eigenvalue: -7.9e+01
## 2025-04-15 19:02:05.016103 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:05.164027 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:05.312892 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2025-04-15 19:02:05.455716 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:05.590065 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2025-04-15 19:02:05.732817 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2025-04-15 19:02:05.872432 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:06.04026 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:06.190887 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:06.350743 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:06.503054 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2025-04-15 19:02:06.638117 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2025-04-15 19:02:06.778997 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2025-04-15 19:02:06.930163 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2025-04-15 19:02:07.069305 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:07.210294 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2025-04-15 19:02:07.356065 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2025-04-15 19:02:07.497906 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:07.644483 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2025-04-15 19:02:07.800124 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2025-04-15 19:02:07.941628 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:08.077845 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:08.217353 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:08.358925 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:08.737394 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2025-04-15 19:02:08.87752 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2025-04-15 19:02:09.011322 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2025-04-15 19:02:09.15676 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:09.326834 INFO::Fitting model to feature number 40, Clostridium.citroniae
## boundary (singular) fit: see help('isSingular')
## Feature Clostridium.citroniae : simpleWarning: Model failed to converge with 1 negative eigenvalue: -3.4e+01
## 2025-04-15 19:02:09.448677 WARNING::Fitting problem for feature 40 a warning was issued
## boundary (singular) fit: see help('isSingular')
## Warning: Model failed to converge with 1 negative eigenvalue: -3.4e+01
## 2025-04-15 19:02:09.593893 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:09.74558 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:09.882374 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2025-04-15 19:02:10.03234 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2025-04-15 19:02:10.172147 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:10.306978 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:10.44754 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2025-04-15 19:02:10.816347 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2025-04-15 19:02:10.957942 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:11.100767 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:11.242354 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:11.401753 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:11.539284 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:11.67613 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2025-04-15 19:02:11.816434 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2025-04-15 19:02:11.952727 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2025-04-15 19:02:12.092085 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:12.237442 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:12.379454 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:12.515624 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:12.858633 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2025-04-15 19:02:12.988487 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:13.126184 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:13.27152 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2025-04-15 19:02:13.419142 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2025-04-15 19:02:13.554384 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2025-04-15 19:02:13.691383 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:13.824667 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2025-04-15 19:02:13.955858 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2025-04-15 19:02:14.097513 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:14.240896 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2025-04-15 19:02:14.372422 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2025-04-15 19:02:14.513332 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:14.671082 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2025-04-15 19:02:14.821596 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:14.969933 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:15.121405 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2025-04-15 19:02:15.252588 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2025-04-15 19:02:15.395958 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2025-04-15 19:02:15.530008 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2025-04-15 19:02:15.670955 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2025-04-15 19:02:15.823519 INFO::Fitting model to feature number 82, Escherichia.coli
## 2025-04-15 19:02:15.962477 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2025-04-15 19:02:16.092848 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:16.232727 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2025-04-15 19:02:16.370874 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2025-04-15 19:02:16.511984 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2025-04-15 19:02:16.693447 INFO::Counting total values for each feature
## 2025-04-15 19:02:16.722481 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2025-04-15 19:02:16.814937 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2025-04-15 19:02:16.908592 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2025-04-15 19:02:17.083513 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2025-04-15 19:02:17.13962 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2025-04-15 19:02:17.171583 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2025-04-15 19:02:17.176687 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2025-04-15 19:02:17.181447 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 19:02:17.183619 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2025-04-15 19:02:17.389402 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2025-04-15 19:02:17.395544 INFO::Plotting associations from most to least significant, grouped by metadata
## 2025-04-15 19:02:17.396476 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2025-04-15 19:02:17.398051 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2025-04-15 19:02:17.741431 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2025-04-15 19:02:18.035908 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2025-04-15 19:02:18.275631 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2025-04-15 19:02:18.5222 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2025-04-15 19:02:18.762313 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2025-04-15 19:02:19.009204 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2025-04-15 19:02:19.245278 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2025-04-15 19:02:19.489804 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2025-04-15 19:02:19.72899 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2025-04-15 19:02:19.980457 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2025-04-15 19:02:20.218705 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2025-04-15 19:02:20.460904 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2025-04-15 19:02:20.695175 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2025-04-15 19:02:20.942012 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2025-04-15 19:02:21.178135 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2025-04-15 19:02:21.452542 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2025-04-15 19:02:21.689813 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2025-04-15 19:02:21.92994 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2025-04-15 19:02:22.165919 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-04-15 19:02:22.409267 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2025-04-15 19:02:22.64708 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2025-04-15 19:02:22.88838 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2025-04-15 19:02:23.117829 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2025-04-15 19:02:23.362005 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2025-04-15 19:02:23.597432 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2025-04-15 19:02:23.839019 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2025-04-15 19:02:24.076108 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2025-04-15 19:02:24.316823 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2025-04-15 19:02:24.554297 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2025-04-15 19:02:24.788791 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2025-04-15 19:02:25.031666 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2025-04-15 19:02:25.264079 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2025-04-15 19:02:25.767639 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2025-04-15 19:02:26.020166 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2025-04-15 19:02:26.28202 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2025-04-15 19:02:26.521436 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2025-04-15 19:02:26.765549 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2025-04-15 19:02:27.0018 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2025-04-15 19:02:27.244244 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2025-04-15 19:02:27.470106 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2025-04-15 19:02:27.71422 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2025-04-15 19:02:27.946691 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2025-04-15 19:02:28.184147 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2025-04-15 19:02:28.418654 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2025-04-15 19:02:28.658681 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2025-04-15 19:02:28.887855 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2025-04-15 19:02:29.154732 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2025-04-15 19:02:29.393057 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2025-04-15 19:02:29.646677 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2025-04-15 19:02:29.880009 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2025-04-15 19:02:30.120521 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2025-04-15 19:02:30.357825 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2025-04-15 19:02:30.595254 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2025-04-15 19:02:30.823078 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2025-04-15 19:02:31.061432 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2025-04-15 19:02:31.289373 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2025-04-15 19:02:31.529577 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2025-04-15 19:02:31.766539 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2025-04-15 19:02:32.011404 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2025-04-15 19:02:32.248137 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2025-04-15 19:02:32.487466 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2025-04-15 19:02:32.719147 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2025-04-15 19:02:35.756646 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2025-04-15 19:02:35.758467 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2025-04-15 19:02:35.966889 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2025-04-15 19:02:36.218127 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2025-04-15 19:02:36.457525 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2025-04-15 19:02:36.697462 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2025-04-15 19:02:36.986185 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2025-04-15 19:02:37.230877 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2025-04-15 19:02:37.469367 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2025-04-15 19:02:37.722001 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2025-04-15 19:02:37.95966 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2025-04-15 19:02:38.206778 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2025-04-15 19:02:38.45045 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2025-04-15 19:02:38.716863 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-04-15 19:02:38.984353 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2025-04-15 19:02:39.233198 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2025-04-15 19:02:39.48375 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2025-04-15 19:02:39.72417 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2025-04-15 19:02:39.970375 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2025-04-15 19:02:40.212428 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2025-04-15 19:02:40.457492 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2025-04-15 19:02:40.698123 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2025-04-15 19:02:40.945655 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2025-04-15 19:02:41.184178 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2025-04-15 19:02:41.432899 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2025-04-15 19:02:41.678239 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2025-04-15 19:02:41.932112 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2025-04-15 19:02:42.181887 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2025-04-15 19:02:42.425494 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2025-04-15 19:02:42.659276 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2025-04-15 19:02:42.903823 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2025-04-15 19:02:43.14746 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2025-04-15 19:02:43.389726 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2025-04-15 19:02:43.635963 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2025-04-15 19:02:43.879483 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2025-04-15 19:02:44.13203 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2025-04-15 19:02:44.36392 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2025-04-15 19:02:44.607342 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2025-04-15 19:02:44.844891 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2025-04-15 19:02:45.089774 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2025-04-15 19:02:45.326144 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2025-04-15 19:02:45.574983 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2025-04-15 19:02:45.815289 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2025-04-15 19:02:46.063981 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2025-04-15 19:02:46.313858 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2025-04-15 19:02:49.395406 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2025-04-15 19:02:49.397477 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2025-04-15 19:02:49.616073 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2025-04-15 19:02:49.872033 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2025-04-15 19:02:50.12166 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2025-04-15 19:02:50.367777 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2025-04-15 19:02:50.613246 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-04-15 19:02:50.874886 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2025-04-15 19:02:51.123793 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2025-04-15 19:02:51.392992 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2025-04-15 19:02:51.639624 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2025-04-15 19:02:51.895145 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2025-04-15 19:02:52.140981 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2025-04-15 19:02:52.40659 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2025-04-15 19:02:52.655542 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2025-04-15 19:02:52.926103 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2025-04-15 19:02:53.177873 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2025-04-15 19:02:53.42478 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2025-04-15 19:02:53.665045 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2025-04-15 19:02:53.931859 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2025-04-15 19:02:54.168067 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2025-04-15 19:02:54.421026 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2025-04-15 19:02:54.675837 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2025-04-15 19:02:54.924524 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2025-04-15 19:02:55.183066 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2025-04-15 19:02:55.424865 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2025-04-15 19:02:55.702259 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2025-04-15 19:02:55.94944 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2025-04-15 19:02:56.198908 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2025-04-15 19:02:56.443326 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2025-04-15 19:02:59.622144 INFO::Plotting data for metadata number 4, antibiotics
## 2025-04-15 19:02:59.623958 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2025-04-15 19:02:59.828631 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2025-04-15 19:03:00.07495 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2025-04-15 19:03:00.307547 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2025-04-15 19:03:00.555239 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2025-04-15 19:03:00.788972 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2025-04-15 19:03:01.020486 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2025-04-15 19:03:01.265322 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2025-04-15 19:03:01.489598 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2025-04-15 19:03:01.738655 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2025-04-15 19:03:01.977644 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2025-04-15 19:03:02.210354 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2025-04-15 19:03:02.437407 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2025-04-15 19:03:02.686149 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2025-04-15 19:03:02.918121 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2025-04-15 19:03:03.162911 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-04-15 19:03:03.395571 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2025-04-15 19:03:03.638309 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2025-04-15 19:03:03.863999 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2025-04-15 19:03:04.102422 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2025-04-15 19:03:04.336394 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2025-04-15 19:03:04.578533 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2025-04-15 19:03:04.818949 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2025-04-15 19:03:05.098796 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2025-04-15 19:03:05.338182 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2025-04-15 19:03:05.585728 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2025-04-15 19:03:05.810953 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2025-04-15 19:03:06.058983 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2025-04-15 19:03:06.295368 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2025-04-15 19:03:06.5404 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2025-04-15 19:03:06.800524 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2025-04-15 19:03:07.039243 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2025-04-15 19:03:07.279702 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2025-04-15 19:03:07.511117 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2025-04-15 19:03:07.749066 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2025-04-15 19:03:07.976762 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2025-04-15 19:03:08.245128 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2025-04-15 19:03:08.478647 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2025-04-15 19:03:08.728765 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2025-04-15 19:03:08.956108 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2025-04-15 19:03:11.95682 INFO::Plotting data for metadata number 5, age
## 2025-04-15 19:03:11.958632 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 19:03:12.221165 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 19:03:12.558288 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 19:03:12.801869 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 19:03:13.056434 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 19:03:13.29238 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 19:03:13.541114 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 19:03:13.786599 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 19:03:14.036062 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 19:03:14.284135 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 19:03:14.52153 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 19:03:14.768358 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 19:03:15.007489 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 19:03:15.268025 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 19:03:15.51864 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 19:03:15.747899 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 19:03:16.003132 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 19:03:16.244963 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 19:03:16.478877 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 19:03:16.722802 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 19:03:16.955801 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 19:03:17.204677 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 19:03:20.477536 INFO::Plotting data for metadata number 6, diagnosis
## 2025-04-15 19:03:20.479402 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2025-04-15 19:03:20.686804 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2025-04-15 19:03:20.932408 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2025-04-15 19:03:21.165316 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2025-04-15 19:03:21.410313 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2025-04-15 19:03:21.643655 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2025-04-15 19:03:21.879023 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2025-04-15 19:03:22.1095 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2025-04-15 19:03:22.34962 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2025-04-15 19:03:22.58573 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2025-04-15 19:03:22.82989 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2025-04-15 19:03:23.068032 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2025-04-15 19:03:23.315591 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2025-04-15 19:03:23.568246 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2025-04-15 19:03:23.806426 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2025-04-15 19:03:24.041899 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2025-04-15 19:03:24.277812 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2025-04-15 19:03:24.530668 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2025-04-15 19:03:24.773885 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2025-04-15 19:03:25.029742 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2025-04-15 19:03:25.260828 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2025-04-15 19:03:25.506325 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2025-04-15 19:03:25.740795 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2025-04-15 19:03:25.989397 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2025-04-15 19:03:26.254577 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2025-04-15 19:03:26.513658 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2025-04-15 19:03:26.765149 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2025-04-15 19:03:27.004974 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2025-04-15 19:03:27.256226 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2025-04-15 19:03:27.492691 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2025-04-15 19:03:27.734578 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2025-04-15 19:03:27.9687 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2025-04-15 19:03:28.238032 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2025-04-15 19:03:28.481515 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2025-04-15 19:03:28.734856 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2025-04-15 19:03:28.979049 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2025-04-15 19:03:29.23453 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2025-04-15 19:03:29.471374 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2025-04-15 19:03:29.72138 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2025-04-15 19:03:29.95318 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2025-04-15 19:03:30.194875 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2025-04-15 19:03:30.430806 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2025-04-15 19:03:30.677649 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: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04.2 LTS
## 
## Matrix products: default
## BLAS:   /home/biocbuild/bbs-3.21-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_1.22.0
## 
## 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.