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"
## 2025-01-02 19:57:04.275564 INFO::Writing function arguments to log file
## 2025-01-02 19:57:04.308988 INFO::Verifying options selected are valid
## 2025-01-02 19:57:04.34102 INFO::Determining format of input files
## 2025-01-02 19:57:04.342999 INFO::Input format is data samples as rows and metadata samples as rows
## 2025-01-02 19:57:04.349837 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2025-01-02 19:57:04.351584 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2025-01-02 19:57:04.354514 INFO::Filter data based on min abundance and min prevalence
## 2025-01-02 19:57:04.358703 INFO::Total samples in data: 1595
## 2025-01-02 19:57:04.360044 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2025-01-02 19:57:04.365487 INFO::Total filtered features: 0
## 2025-01-02 19:57:04.367019 INFO::Filtered feature names from abundance and prevalence filtering:
## 2025-01-02 19:57:04.38379 INFO::Total filtered features with variance filtering: 0
## 2025-01-02 19:57:04.385451 INFO::Filtered feature names from variance filtering:
## 2025-01-02 19:57:04.386843 INFO::Running selected normalization method: TSS
## 2025-01-02 19:57:05.342125 INFO::Bypass z-score application to metadata
## 2025-01-02 19:57:05.343782 INFO::Running selected transform method: LOG
## 2025-01-02 19:57:05.364784 INFO::Running selected analysis method: LM
## 2025-01-02 19:57:05.649747 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:06.077209 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2025-01-02 19:57:06.444417 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2025-01-02 19:57:06.596275 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:06.753116 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2025-01-02 19:57:06.983287 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:07.200309 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2025-01-02 19:57:07.413997 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2025-01-02 19:57:07.595321 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2025-01-02 19:57:07.746971 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2025-01-02 19:57:07.892851 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
## 2025-01-02 19:57:08.013074 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
## 2025-01-02 19:57:08.178525 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:08.326733 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:08.529472 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2025-01-02 19:57:08.726664 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:08.942885 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2025-01-02 19:57:09.138283 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2025-01-02 19:57:09.330701 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:09.464603 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:09.591851 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:09.739718 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:10.153778 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2025-01-02 19:57:10.2923 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2025-01-02 19:57:10.423081 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2025-01-02 19:57:10.5612 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2025-01-02 19:57:10.71108 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:10.869081 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2025-01-02 19:57:10.996341 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2025-01-02 19:57:11.155511 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:11.280064 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2025-01-02 19:57:11.410379 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2025-01-02 19:57:11.534838 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:11.669972 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:11.792288 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:11.91937 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:12.055178 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2025-01-02 19:57:12.182113 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2025-01-02 19:57:12.304331 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2025-01-02 19:57:12.446603 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:12.576045 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
## 2025-01-02 19:57:12.680439 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
## 2025-01-02 19:57:12.807368 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:12.939619 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:13.06074 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2025-01-02 19:57:13.197384 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2025-01-02 19:57:13.330728 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:13.454089 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:13.585016 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2025-01-02 19:57:13.723002 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2025-01-02 19:57:13.86516 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:14.003045 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:14.139782 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:14.265907 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:14.397427 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:14.538545 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2025-01-02 19:57:14.660081 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2025-01-02 19:57:14.803173 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2025-01-02 19:57:14.946465 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:15.083578 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:15.221747 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:15.375586 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:15.50404 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2025-01-02 19:57:15.627557 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:15.763321 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:15.89475 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2025-01-02 19:57:16.100451 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2025-01-02 19:57:16.308505 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2025-01-02 19:57:16.440871 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:16.572285 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2025-01-02 19:57:16.702667 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2025-01-02 19:57:16.878356 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:17.013593 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2025-01-02 19:57:17.225942 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2025-01-02 19:57:17.393948 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:17.806068 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2025-01-02 19:57:18.02529 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:18.173974 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:18.309223 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2025-01-02 19:57:18.441661 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2025-01-02 19:57:18.567768 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2025-01-02 19:57:18.691375 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2025-01-02 19:57:18.837346 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2025-01-02 19:57:18.967714 INFO::Fitting model to feature number 82, Escherichia.coli
## 2025-01-02 19:57:19.102107 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2025-01-02 19:57:19.245906 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:19.408767 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2025-01-02 19:57:19.599682 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2025-01-02 19:57:20.097415 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2025-01-02 19:57:20.345201 INFO::Counting total values for each feature
## 2025-01-02 19:57:20.381053 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2025-01-02 19:57:20.702231 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2025-01-02 19:57:21.159616 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2025-01-02 19:57:21.603283 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2025-01-02 19:57:21.65776 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2025-01-02 19:57:21.696051 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2025-01-02 19:57:21.705461 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2025-01-02 19:57:21.728623 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
## 2025-01-02 19:57:21.74006 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2025-01-02 19:57:22.337386 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2025-01-02 19:57:22.346505 INFO::Plotting associations from most to least significant, grouped by metadata
## 2025-01-02 19:57:22.349279 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2025-01-02 19:57:22.35288 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2025-01-02 19:57:22.88823 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2025-01-02 19:57:23.320522 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2025-01-02 19:57:23.629349 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2025-01-02 19:57:24.009311 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2025-01-02 19:57:24.27192 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2025-01-02 19:57:24.573317 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2025-01-02 19:57:24.831125 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2025-01-02 19:57:25.107312 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2025-01-02 19:57:25.369408 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2025-01-02 19:57:25.78735 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2025-01-02 19:57:26.131144 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2025-01-02 19:57:26.533495 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2025-01-02 19:57:26.785925 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2025-01-02 19:57:27.079522 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2025-01-02 19:57:27.336188 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2025-01-02 19:57:27.667698 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2025-01-02 19:57:27.947145 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2025-01-02 19:57:28.224621 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2025-01-02 19:57:28.507724 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-01-02 19:57:28.803803 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2025-01-02 19:57:29.082681 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2025-01-02 19:57:29.510897 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2025-01-02 19:57:29.83351 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2025-01-02 19:57:30.10576 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2025-01-02 19:57:30.343601 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2025-01-02 19:57:30.626887 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2025-01-02 19:57:30.909398 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2025-01-02 19:57:31.180728 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2025-01-02 19:57:31.463477 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2025-01-02 19:57:31.769784 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2025-01-02 19:57:32.148227 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2025-01-02 19:57:32.547188 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2025-01-02 19:57:32.893784 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2025-01-02 19:57:33.346078 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2025-01-02 19:57:33.673106 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2025-01-02 19:57:34.052217 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2025-01-02 19:57:34.439872 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2025-01-02 19:57:34.722224 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2025-01-02 19:57:35.002674 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2025-01-02 19:57:35.278243 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2025-01-02 19:57:35.632081 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2025-01-02 19:57:35.919366 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2025-01-02 19:57:36.227391 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2025-01-02 19:57:36.501743 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2025-01-02 19:57:36.75645 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2025-01-02 19:57:37.318575 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2025-01-02 19:57:37.709185 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2025-01-02 19:57:38.058058 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2025-01-02 19:57:38.381135 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2025-01-02 19:57:38.733162 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2025-01-02 19:57:39.200048 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2025-01-02 19:57:39.454733 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2025-01-02 19:57:39.788402 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2025-01-02 19:57:40.067363 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2025-01-02 19:57:40.382487 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2025-01-02 19:57:40.698938 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2025-01-02 19:57:41.04379 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2025-01-02 19:57:41.351441 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2025-01-02 19:57:41.637957 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2025-01-02 19:57:41.945067 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2025-01-02 19:57:42.319871 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2025-01-02 19:57:42.936614 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2025-01-02 19:57:46.663262 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2025-01-02 19:57:46.667203 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2025-01-02 19:57:46.959881 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2025-01-02 19:57:47.241782 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2025-01-02 19:57:47.500842 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2025-01-02 19:57:47.800515 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2025-01-02 19:57:48.064082 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2025-01-02 19:57:48.414376 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2025-01-02 19:57:48.708568 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2025-01-02 19:57:49.088232 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2025-01-02 19:57:49.345769 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2025-01-02 19:57:49.656338 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2025-01-02 19:57:49.969249 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2025-01-02 19:57:50.400603 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-01-02 19:57:50.801591 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2025-01-02 19:57:51.087162 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2025-01-02 19:57:51.465692 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2025-01-02 19:57:51.883026 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2025-01-02 19:57:52.17437 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2025-01-02 19:57:52.481338 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2025-01-02 19:57:52.736288 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2025-01-02 19:57:53.03642 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2025-01-02 19:57:53.302532 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2025-01-02 19:57:53.561937 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2025-01-02 19:57:54.22652 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2025-01-02 19:57:54.61092 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2025-01-02 19:57:54.937479 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2025-01-02 19:57:55.3337 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2025-01-02 19:57:55.70864 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2025-01-02 19:57:56.006966 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2025-01-02 19:57:56.259919 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2025-01-02 19:57:56.52573 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2025-01-02 19:57:56.782607 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2025-01-02 19:57:57.058428 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2025-01-02 19:57:57.407096 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2025-01-02 19:57:57.815436 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2025-01-02 19:57:58.086991 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2025-01-02 19:57:58.396956 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2025-01-02 19:57:58.659653 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2025-01-02 19:57:58.927367 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2025-01-02 19:57:59.182213 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2025-01-02 19:57:59.540634 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2025-01-02 19:57:59.836735 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2025-01-02 19:58:00.137657 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2025-01-02 19:58:00.503313 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2025-01-02 19:58:04.749606 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2025-01-02 19:58:04.754009 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2025-01-02 19:58:05.09869 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2025-01-02 19:58:05.437411 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2025-01-02 19:58:05.770495 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2025-01-02 19:58:06.180304 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2025-01-02 19:58:06.58242 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-01-02 19:58:06.890155 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2025-01-02 19:58:07.154008 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2025-01-02 19:58:07.483343 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2025-01-02 19:58:07.810627 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2025-01-02 19:58:08.125216 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2025-01-02 19:58:08.430885 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2025-01-02 19:58:08.711966 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2025-01-02 19:58:08.970242 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2025-01-02 19:58:09.260847 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2025-01-02 19:58:09.5115 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2025-01-02 19:58:09.853998 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2025-01-02 19:58:10.168758 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2025-01-02 19:58:10.539474 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2025-01-02 19:58:11.173051 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2025-01-02 19:58:11.462686 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2025-01-02 19:58:11.746299 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2025-01-02 19:58:12.115352 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2025-01-02 19:58:12.480473 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2025-01-02 19:58:12.777355 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2025-01-02 19:58:13.053141 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2025-01-02 19:58:13.337747 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2025-01-02 19:58:13.656111 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2025-01-02 19:58:13.967304 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2025-01-02 19:58:18.044885 INFO::Plotting data for metadata number 4, antibiotics
## 2025-01-02 19:58:18.047757 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2025-01-02 19:58:18.622039 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2025-01-02 19:58:18.943426 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2025-01-02 19:58:19.265789 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2025-01-02 19:58:19.502215 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2025-01-02 19:58:19.788825 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2025-01-02 19:58:20.142735 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2025-01-02 19:58:20.472386 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2025-01-02 19:58:20.745454 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2025-01-02 19:58:20.968347 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2025-01-02 19:58:21.213035 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2025-01-02 19:58:21.435897 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2025-01-02 19:58:21.679042 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2025-01-02 19:58:21.934207 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2025-01-02 19:58:22.243891 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2025-01-02 19:58:22.476769 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2025-01-02 19:58:22.703819 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2025-01-02 19:58:23.004076 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2025-01-02 19:58:23.374354 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2025-01-02 19:58:23.617762 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2025-01-02 19:58:23.863776 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2025-01-02 19:58:24.165415 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2025-01-02 19:58:24.49328 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2025-01-02 19:58:24.786066 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2025-01-02 19:58:25.141162 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2025-01-02 19:58:25.525444 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2025-01-02 19:58:25.787013 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2025-01-02 19:58:26.052456 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2025-01-02 19:58:26.325141 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2025-01-02 19:58:26.662584 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2025-01-02 19:58:26.904502 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2025-01-02 19:58:27.144789 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2025-01-02 19:58:27.399762 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2025-01-02 19:58:27.683691 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2025-01-02 19:58:27.931196 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2025-01-02 19:58:28.181741 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2025-01-02 19:58:28.468944 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2025-01-02 19:58:28.71587 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2025-01-02 19:58:28.960433 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2025-01-02 19:58:29.281713 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2025-01-02 19:58:33.409498 INFO::Plotting data for metadata number 5, age
## 2025-01-02 19:58:33.412551 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()`).
## 2025-01-02 19:58:33.70498 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()`).
## 2025-01-02 19:58:34.100485 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()`).
## 2025-01-02 19:58:34.358956 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()`).
## 2025-01-02 19:58:34.625532 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()`).
## 2025-01-02 19:58:34.896873 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()`).
## 2025-01-02 19:58:35.297143 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()`).
## 2025-01-02 19:58:35.707542 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()`).
## 2025-01-02 19:58:36.026221 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()`).
## 2025-01-02 19:58:36.36687 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()`).
## 2025-01-02 19:58:36.600255 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()`).
## 2025-01-02 19:58:36.864803 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()`).
## 2025-01-02 19:58:37.17523 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()`).
## 2025-01-02 19:58:37.478107 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()`).
## 2025-01-02 19:58:37.753735 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()`).
## 2025-01-02 19:58:38.039245 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()`).
## 2025-01-02 19:58:38.392617 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()`).
## 2025-01-02 19:58:38.714168 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()`).
## 2025-01-02 19:58:39.01304 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()`).
## 2025-01-02 19:58:39.325678 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()`).
## 2025-01-02 19:58:39.588702 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()`).
## 2025-01-02 19:58:39.886283 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()`).
## 2025-01-02 19:58:43.949464 INFO::Plotting data for metadata number 6, diagnosis
## 2025-01-02 19:58:43.952371 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2025-01-02 19:58:44.188127 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2025-01-02 19:58:44.524948 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2025-01-02 19:58:44.945053 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2025-01-02 19:58:45.210215 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2025-01-02 19:58:45.52823 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2025-01-02 19:58:45.795284 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2025-01-02 19:58:46.079325 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2025-01-02 19:58:46.338598 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2025-01-02 19:58:46.705258 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2025-01-02 19:58:47.011346 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2025-01-02 19:58:47.293901 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2025-01-02 19:58:47.562754 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2025-01-02 19:58:47.832149 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2025-01-02 19:58:48.090718 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2025-01-02 19:58:48.366254 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2025-01-02 19:58:48.624228 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2025-01-02 19:58:48.943335 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2025-01-02 19:58:49.207484 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2025-01-02 19:58:49.479346 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2025-01-02 19:58:49.775838 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2025-01-02 19:58:50.20237 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2025-01-02 19:58:50.463334 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2025-01-02 19:58:50.752557 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2025-01-02 19:58:51.014469 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2025-01-02 19:58:51.296291 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2025-01-02 19:58:51.55121 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2025-01-02 19:58:51.851155 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2025-01-02 19:58:52.106842 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2025-01-02 19:58:52.496827 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2025-01-02 19:58:52.888711 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2025-01-02 19:58:53.233379 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2025-01-02 19:58:53.623802 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2025-01-02 19:58:54.061016 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2025-01-02 19:58:54.445699 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2025-01-02 19:58:54.772827 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2025-01-02 19:58:55.116294 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2025-01-02 19:58:55.497425 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2025-01-02 19:58:55.839618 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2025-01-02 19:58:56.189947 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2025-01-02 19:58:56.563013 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2025-01-02 19:58:56.881948 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2025-01-02 19:58:57.286897 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 4.4.2 (2024-10-31 ucrt)
## Platform: x86_64-w64-mingw32/x64
## Running under: Windows Server 2022 x64 (build 20348)
## 
## Matrix products: default
## 
## 
## locale:
## [1] LC_COLLATE=C                          
## [2] LC_CTYPE=English_United States.utf8   
## [3] LC_MONETARY=English_United States.utf8
## [4] LC_NUMERIC=C                          
## [5] LC_TIME=English_United States.utf8    
## 
## time zone: America/New_York
## tzcode source: internal
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
## 
## other attached packages:
## [1] Maaslin2_1.20.0
## 
## loaded via a namespace (and not attached):
##  [1] sass_0.4.9          generics_0.1.3      robustbase_0.99-4-1
##  [4] lattice_0.22-6      lme4_1.1-35.5       digest_0.6.37      
##  [7] magrittr_2.0.3      RColorBrewer_1.1-3  evaluate_1.0.1     
## [10] grid_4.4.2          mvtnorm_1.3-2       fastmap_1.2.0      
## [13] jsonlite_1.8.9      Matrix_1.7-1        DBI_1.2.3          
## [16] optparse_1.7.5      biglm_0.9-3         mgcv_1.9-1         
## [19] scales_1.3.0        pbapply_1.7-2       permute_0.9-7      
## [22] numDeriv_2016.8-1.1 getopt_1.20.4       jquerylib_0.1.4    
## [25] cli_3.6.3           rlang_1.1.4         crayon_1.5.3       
## [28] munsell_0.5.1       splines_4.4.2       withr_3.0.2        
## [31] cachem_1.1.0        yaml_2.3.10         vegan_2.6-8        
## [34] tools_4.4.2         parallel_4.4.2      nloptr_2.1.1       
## [37] minqa_1.2.8         dplyr_1.1.4         colorspace_2.1-1   
## [40] ggplot2_3.5.1       boot_1.3-31         hash_2.2.6.3       
## [43] vctrs_0.6.5         R6_2.5.1            lifecycle_1.0.4    
## [46] MASS_7.3-63         logging_0.10-108    pcaPP_2.0-5        
## [49] cluster_2.1.8       pkgconfig_2.0.3     gtable_0.3.6       
## [52] lmerTest_3.1-3      pillar_1.10.0       bslib_0.8.0        
## [55] Rcpp_1.0.13-1       data.table_1.16.4   glue_1.8.0         
## [58] DEoptimR_1.1-3-1    xfun_0.49           tibble_3.2.1       
## [61] tidyselect_1.2.1    knitr_1.49          farver_2.1.2       
## [64] htmltools_0.5.8.1   nlme_3.1-166        labeling_0.4.3     
## [67] rmarkdown_2.29      pheatmap_1.0.12     compiler_4.4.2

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.