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
Requirements
Installation
How to Run
Visualization
Troubleshooting

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

Download the source: MaAsLin2.tar.gz
Decompress the download:
- $ tar xzvf maaslin2.tar.gz
Install the Bioconductor dependencies edgeR and metagenomeSeq.
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')"
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.

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.
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.

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.
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"
## 2025-02-06 05:44:01.530006 INFO::Writing function arguments to log file
## 2025-02-06 05:44:01.546155 INFO::Verifying options selected are valid
## 2025-02-06 05:44:01.595336 INFO::Determining format of input files
## 2025-02-06 05:44:01.596126 INFO::Input format is data samples as rows and metadata samples as rows
## 2025-02-06 05:44:01.599197 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2025-02-06 05:44:01.599893 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2025-02-06 05:44:01.669605 INFO::Filter data based on min abundance and min prevalence
## 2025-02-06 05:44:01.670289 INFO::Total samples in data: 1595
## 2025-02-06 05:44:01.670761 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2025-02-06 05:44:01.673693 INFO::Total filtered features: 0
## 2025-02-06 05:44:01.674245 INFO::Filtered feature names from abundance and prevalence filtering:
## 2025-02-06 05:44:01.679586 INFO::Total filtered features with variance filtering: 0
## 2025-02-06 05:44:01.680107 INFO::Filtered feature names from variance filtering:
## 2025-02-06 05:44:01.680552 INFO::Running selected normalization method: TSS
## 2025-02-06 05:44:02.390628 INFO::Bypass z-score application to metadata
## 2025-02-06 05:44:02.391406 INFO::Running selected transform method: LOG
## 2025-02-06 05:44:02.405503 INFO::Running selected analysis method: LM
## 2025-02-06 05:44:02.625137 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:02.848742 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2025-02-06 05:44:02.95791 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2025-02-06 05:44:03.047102 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:03.13988 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2025-02-06 05:44:03.238215 INFO::Fitting model to feature number 6, Bacteroides.caccae

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:03.33223 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2025-02-06 05:44:03.421463 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2025-02-06 05:44:03.519993 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2025-02-06 05:44:03.610044 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2025-02-06 05:44:03.707858 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

## 2025-02-06 05:44:03.782023 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

## 2025-02-06 05:44:03.878778 INFO::Fitting model to feature number 12, Bacteroides.fragilis

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:03.96805 INFO::Fitting model to feature number 13, Bacteroides.intestinalis

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:04.068639 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2025-02-06 05:44:04.165587 INFO::Fitting model to feature number 15, Bacteroides.ovatus

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:04.25574 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2025-02-06 05:44:04.353774 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2025-02-06 05:44:04.452812 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:04.54772 INFO::Fitting model to feature number 19, Bacteroides.uniformis

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:04.638818 INFO::Fitting model to feature number 20, Bacteroides.vulgatus

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:04.753834 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:04.843652 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2025-02-06 05:44:04.936561 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2025-02-06 05:44:05.028901 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2025-02-06 05:44:05.127256 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2025-02-06 05:44:05.218254 INFO::Fitting model to feature number 26, Parabacteroides.distasonis

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:05.313544 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2025-02-06 05:44:05.406439 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2025-02-06 05:44:05.502583 INFO::Fitting model to feature number 29, Parabacteroides.unclassified

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:05.591702 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2025-02-06 05:44:05.688939 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2025-02-06 05:44:05.778175 INFO::Fitting model to feature number 32, Prevotella.copri

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:06.025381 INFO::Fitting model to feature number 33, Alistipes.finegoldii

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:06.112494 INFO::Fitting model to feature number 34, Alistipes.onderdonkii

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:06.204102 INFO::Fitting model to feature number 35, Alistipes.putredinis

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:06.299675 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2025-02-06 05:44:06.392814 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2025-02-06 05:44:06.487067 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2025-02-06 05:44:06.578935 INFO::Fitting model to feature number 39, Clostridium.bolteae

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:06.675442 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

## 2025-02-06 05:44:06.751268 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

## 2025-02-06 05:44:06.844137 INFO::Fitting model to feature number 41, Clostridium.clostridioforme

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:06.938 INFO::Fitting model to feature number 42, Clostridium.hathewayi

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:07.028673 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2025-02-06 05:44:07.128645 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2025-02-06 05:44:07.222334 INFO::Fitting model to feature number 45, Clostridium.symbiosum

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:07.30989 INFO::Fitting model to feature number 46, Flavonifractor.plautii

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:07.400807 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2025-02-06 05:44:07.657188 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2025-02-06 05:44:07.749333 INFO::Fitting model to feature number 49, Eubacterium.rectale

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:07.843356 INFO::Fitting model to feature number 50, Eubacterium.siraeum

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:07.931774 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:08.025923 INFO::Fitting model to feature number 52, Eubacterium.ventriosum

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:08.108609 INFO::Fitting model to feature number 53, Ruminococcus.gnavus

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:08.197384 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2025-02-06 05:44:08.288751 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2025-02-06 05:44:08.374563 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2025-02-06 05:44:08.467088 INFO::Fitting model to feature number 57, Dorea.longicatena

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:08.561491 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:08.652554 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:08.741159 INFO::Fitting model to feature number 60, Roseburia.hominis

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:08.836523 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2025-02-06 05:44:08.919816 INFO::Fitting model to feature number 62, Roseburia.inulinivorans

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:09.009788 INFO::Fitting model to feature number 63, Roseburia.unclassified

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:09.105308 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2025-02-06 05:44:09.201931 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2025-02-06 05:44:09.292521 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2025-02-06 05:44:09.38687 INFO::Fitting model to feature number 67, Ruminococcus.bromii

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:09.471396 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2025-02-06 05:44:09.556699 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2025-02-06 05:44:09.652673 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:09.740501 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2025-02-06 05:44:09.828324 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2025-02-06 05:44:09.91708 INFO::Fitting model to feature number 73, Dialister.invisus

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:10.003518 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2025-02-06 05:44:10.096401 INFO::Fitting model to feature number 75, Veillonella.dispar

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:10.18383 INFO::Fitting model to feature number 76, Veillonella.parvula

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:10.276516 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2025-02-06 05:44:10.362262 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2025-02-06 05:44:10.451868 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2025-02-06 05:44:10.543225 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2025-02-06 05:44:10.635314 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2025-02-06 05:44:10.738412 INFO::Fitting model to feature number 82, Escherichia.coli
## 2025-02-06 05:44:10.830334 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2025-02-06 05:44:10.918692 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:11.00421 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae

## boundary (singular) fit: see help('isSingular')

## 2025-02-06 05:44:11.0988 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2025-02-06 05:44:11.189188 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2025-02-06 05:44:11.312835 INFO::Counting total values for each feature
## 2025-02-06 05:44:11.332873 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2025-02-06 05:44:11.400155 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2025-02-06 05:44:11.464104 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2025-02-06 05:44:11.560905 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2025-02-06 05:44:11.598683 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2025-02-06 05:44:11.620648 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2025-02-06 05:44:11.624202 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2025-02-06 05:44:11.627529 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
## 2025-02-06 05:44:11.629129 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2025-02-06 05:44:11.759434 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2025-02-06 05:44:11.763386 INFO::Plotting associations from most to least significant, grouped by metadata
## 2025-02-06 05:44:11.764008 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2025-02-06 05:44:11.765045 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii

## 2025-02-06 05:44:11.997014 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified

## 2025-02-06 05:44:12.171455 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis

## 2025-02-06 05:44:12.335324 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale

## 2025-02-06 05:44:12.497743 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus

## 2025-02-06 05:44:12.654065 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus

## 2025-02-06 05:44:12.829382 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum

## 2025-02-06 05:44:12.993765 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified

## 2025-02-06 05:44:13.156703 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans

## 2025-02-06 05:44:13.323454 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis

## 2025-02-06 05:44:13.483079 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron

## 2025-02-06 05:44:13.654028 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis

## 2025-02-06 05:44:13.823557 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis

## 2025-02-06 05:44:13.991705 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei

## 2025-02-06 05:44:14.156192 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii

## 2025-02-06 05:44:14.318869 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens

## 2025-02-06 05:44:14.473693 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum

## 2025-02-06 05:44:14.633382 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena

## 2025-02-06 05:44:14.793594 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae

## 2025-02-06 05:44:14.95897 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2025-02-06 05:44:15.122246 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli

## 2025-02-06 05:44:15.285224 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae

## 2025-02-06 05:44:15.448935 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified

## 2025-02-06 05:44:15.603579 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii

## 2025-02-06 05:44:15.764143 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens

## 2025-02-06 05:44:15.916336 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris

## 2025-02-06 05:44:16.080213 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes

## 2025-02-06 05:44:16.239938 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis

## 2025-02-06 05:44:16.40138 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii

## 2025-02-06 05:44:16.564613 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii

## 2025-02-06 05:44:16.754224 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis

## 2025-02-06 05:44:16.927804 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara

## 2025-02-06 05:44:17.095984 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum

## 2025-02-06 05:44:17.246866 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae

## 2025-02-06 05:44:17.422185 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified

## 2025-02-06 05:44:17.598163 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens

## 2025-02-06 05:44:17.761986 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus

## 2025-02-06 05:44:17.931372 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme

## 2025-02-06 05:44:18.10239 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii

## 2025-02-06 05:44:18.262785 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii

## 2025-02-06 05:44:18.423303 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum

## 2025-02-06 05:44:18.594874 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris

## 2025-02-06 05:44:18.762382 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47

## 2025-02-06 05:44:18.925246 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus

## 2025-02-06 05:44:19.093724 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii

## 2025-02-06 05:44:19.260546 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis

## 2025-02-06 05:44:19.431994 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified

## 2025-02-06 05:44:19.60048 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques

## 2025-02-06 05:44:19.755639 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified

## 2025-02-06 05:44:19.933715 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus

## 2025-02-06 05:44:20.104286 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis

## 2025-02-06 05:44:20.274528 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified

## 2025-02-06 05:44:20.445919 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified

## 2025-02-06 05:44:20.614232 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis

## 2025-02-06 05:44:20.816966 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis

## 2025-02-06 05:44:20.99042 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31

## 2025-02-06 05:44:21.160436 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae

## 2025-02-06 05:44:21.316063 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8

## 2025-02-06 05:44:21.485192 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae

## 2025-02-06 05:44:21.654729 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii

## 2025-02-06 05:44:21.830166 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii

## 2025-02-06 05:44:22.006096 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum

## 2025-02-06 05:44:22.172732 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA

## 2025-02-06 05:44:24.271755 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2025-02-06 05:44:24.273162 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified

## 2025-02-06 05:44:24.421824 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii

## 2025-02-06 05:44:24.602481 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae

## 2025-02-06 05:44:24.77709 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified

## 2025-02-06 05:44:24.951271 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis

## 2025-02-06 05:44:25.127076 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum

## 2025-02-06 05:44:25.320256 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus

## 2025-02-06 05:44:25.498576 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis

## 2025-02-06 05:44:25.673876 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii

## 2025-02-06 05:44:25.84136 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale

## 2025-02-06 05:44:26.020753 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis

## 2025-02-06 05:44:26.202212 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis

## 2025-02-06 05:44:26.38226 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA

## 2025-02-06 05:44:26.566526 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii

## 2025-02-06 05:44:26.745323 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens

## 2025-02-06 05:44:26.91865 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris

## 2025-02-06 05:44:27.088414 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii

## 2025-02-06 05:44:27.266799 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis

## 2025-02-06 05:44:27.449718 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum

## 2025-02-06 05:44:27.626572 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus

## 2025-02-06 05:44:27.80336 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii

## 2025-02-06 05:44:27.983177 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii

## 2025-02-06 05:44:28.162537 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae

## 2025-02-06 05:44:28.342185 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus

## 2025-02-06 05:44:28.518658 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum

## 2025-02-06 05:44:28.694541 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae

## 2025-02-06 05:44:28.874629 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae

## 2025-02-06 05:44:29.047939 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum

## 2025-02-06 05:44:29.226563 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis

## 2025-02-06 05:44:29.605994 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii

## 2025-02-06 05:44:29.763193 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques

## 2025-02-06 05:44:29.917006 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8

## 2025-02-06 05:44:30.081432 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum

## 2025-02-06 05:44:30.235385 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum

## 2025-02-06 05:44:30.399333 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis

## 2025-02-06 05:44:30.548219 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens

## 2025-02-06 05:44:30.701788 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi

## 2025-02-06 05:44:30.869842 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified

## 2025-02-06 05:44:31.027091 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens

## 2025-02-06 05:44:31.184114 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus

## 2025-02-06 05:44:31.34514 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae

## 2025-02-06 05:44:31.510926 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus

## 2025-02-06 05:44:31.666851 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris

## 2025-02-06 05:44:31.826351 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47

## 2025-02-06 05:44:33.796144 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2025-02-06 05:44:33.797555 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii

## 2025-02-06 05:44:33.961578 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale

## 2025-02-06 05:44:34.132423 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31

## 2025-02-06 05:44:34.300726 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis

## 2025-02-06 05:44:34.456888 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified

## 2025-02-06 05:44:34.618936 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2025-02-06 05:44:34.789074 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques

## 2025-02-06 05:44:34.95469 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum

## 2025-02-06 05:44:35.124753 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus

## 2025-02-06 05:44:35.295491 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli

## 2025-02-06 05:44:35.459276 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum

## 2025-02-06 05:44:35.616578 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified

## 2025-02-06 05:44:35.787766 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis

## 2025-02-06 05:44:35.959015 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens

## 2025-02-06 05:44:36.129313 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae

## 2025-02-06 05:44:36.299868 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii

## 2025-02-06 05:44:36.457478 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus

## 2025-02-06 05:44:36.621736 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified

## 2025-02-06 05:44:36.791978 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis

## 2025-02-06 05:44:36.952426 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii

## 2025-02-06 05:44:37.115844 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans

## 2025-02-06 05:44:37.278562 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii

## 2025-02-06 05:44:37.452405 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified

## 2025-02-06 05:44:37.63108 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii

## 2025-02-06 05:44:37.797016 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis

## 2025-02-06 05:44:38.012103 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri

## 2025-02-06 05:44:38.18832 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei

## 2025-02-06 05:44:38.358037 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis

## 2025-02-06 05:44:38.53259 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii

## 2025-02-06 05:44:40.607227 INFO::Plotting data for metadata number 4, antibiotics
## 2025-02-06 05:44:40.608806 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans

## 2025-02-06 05:44:40.75558 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis

## 2025-02-06 05:44:40.931072 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale

## 2025-02-06 05:44:41.102618 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus

## 2025-02-06 05:44:41.26927 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis

## 2025-02-06 05:44:41.417361 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena

## 2025-02-06 05:44:41.584715 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus

## 2025-02-06 05:44:41.748587 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii

## 2025-02-06 05:44:41.904361 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum

## 2025-02-06 05:44:42.065216 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae

## 2025-02-06 05:44:42.237094 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis

## 2025-02-06 05:44:42.419333 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii

## 2025-02-06 05:44:42.574222 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii

## 2025-02-06 05:44:42.752214 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified

## 2025-02-06 05:44:42.925056 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum

## 2025-02-06 05:44:43.106499 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA

## 2025-02-06 05:44:43.27981 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii

## 2025-02-06 05:44:43.43549 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31

## 2025-02-06 05:44:43.604414 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis

## 2025-02-06 05:44:43.772579 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii

## 2025-02-06 05:44:43.940987 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis

## 2025-02-06 05:44:44.110586 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens

## 2025-02-06 05:44:44.275673 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens

## 2025-02-06 05:44:44.429777 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron

## 2025-02-06 05:44:44.599803 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii

## 2025-02-06 05:44:44.767792 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae

## 2025-02-06 05:44:44.932301 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum

## 2025-02-06 05:44:45.102566 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques

## 2025-02-06 05:44:45.264014 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum

## 2025-02-06 05:44:45.436249 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis

## 2025-02-06 05:44:45.596721 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified

## 2025-02-06 05:44:45.754469 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar

## 2025-02-06 05:44:45.922125 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica

## 2025-02-06 05:44:46.089917 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8

## 2025-02-06 05:44:46.25863 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile

## 2025-02-06 05:44:46.428279 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47

## 2025-02-06 05:44:46.61535 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA

## 2025-02-06 05:44:46.792965 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila

## 2025-02-06 05:44:46.956502 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae

## 2025-02-06 05:44:47.123656 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus

## 2025-02-06 05:44:49.178529 INFO::Plotting data for metadata number 5, age
## 2025-02-06 05:44:49.18018 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()`).

## 2025-02-06 05:44:49.375771 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()`).

## 2025-02-06 05:44:49.601949 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()`).

## 2025-02-06 05:44:49.766206 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()`).

## 2025-02-06 05:44:49.94979 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()`).

## 2025-02-06 05:44:50.12489 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()`).

## 2025-02-06 05:44:50.299164 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()`).

## 2025-02-06 05:44:50.474641 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()`).

## 2025-02-06 05:44:50.644886 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()`).

## 2025-02-06 05:44:50.842175 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()`).

## 2025-02-06 05:44:51.01687 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()`).

## 2025-02-06 05:44:51.190568 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()`).

## 2025-02-06 05:44:51.369298 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()`).

## 2025-02-06 05:44:51.546693 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()`).

## 2025-02-06 05:44:51.716655 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()`).

## 2025-02-06 05:44:51.894537 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()`).

## 2025-02-06 05:44:52.076825 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()`).

## 2025-02-06 05:44:52.259872 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()`).

## 2025-02-06 05:44:52.43613 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()`).

## 2025-02-06 05:44:52.611528 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()`).

## 2025-02-06 05:44:52.788334 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()`).

## 2025-02-06 05:44:52.965161 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()`).

## 2025-02-06 05:44:55.255824 INFO::Plotting data for metadata number 6, diagnosis
## 2025-02-06 05:44:55.257239 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis

## 2025-02-06 05:44:55.391759 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2025-02-06 05:44:55.554658 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2025-02-06 05:44:55.709399 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2025-02-06 05:44:55.876228 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2025-02-06 05:44:56.040748 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme

## 2025-02-06 05:44:56.20102 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2025-02-06 05:44:56.361161 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2025-02-06 05:44:56.520141 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2025-02-06 05:44:56.676266 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2025-02-06 05:44:56.83454 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans

## 2025-02-06 05:44:56.993993 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2025-02-06 05:44:57.149526 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2025-02-06 05:44:57.306662 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2025-02-06 05:44:57.476193 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii

## 2025-02-06 05:44:57.636602 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified

## 2025-02-06 05:44:57.789481 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2025-02-06 05:44:57.949198 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2025-02-06 05:44:58.109527 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2025-02-06 05:44:58.271147 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2025-02-06 05:44:58.44958 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum

## 2025-02-06 05:44:58.617931 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2025-02-06 05:44:58.781475 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2025-02-06 05:44:58.950699 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2025-02-06 05:44:59.113217 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2025-02-06 05:44:59.265975 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified

## 2025-02-06 05:44:59.430097 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2025-02-06 05:44:59.59947 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2025-02-06 05:44:59.765219 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified

## 2025-02-06 05:44:59.931399 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris

## 2025-02-06 05:45:00.09583 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2025-02-06 05:45:00.257819 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus

## 2025-02-06 05:45:00.418769 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis

## 2025-02-06 05:45:00.588242 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2025-02-06 05:45:00.753424 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus

## 2025-02-06 05:45:00.922109 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis

## 2025-02-06 05:45:01.085772 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2025-02-06 05:45:01.254265 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii

## 2025-02-06 05:45:01.426973 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2025-02-06 05:45:01.590379 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2025-02-06 05:45:01.754525 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2025-02-06 05:45:01.922113 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2025-02-06 05:45:02.112195 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)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04.1 LTS
## 
## Matrix products: default
## BLAS:   /usr/lib/x86_64-linux-gnu/openblas-pthread/libblas.so.3 
## LAPACK: /usr/lib/x86_64-linux-gnu/openblas-pthread/libopenblasp-r0.3.26.so;  LAPACK version 3.12.0
## 
## locale:
##  [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C              
##  [3] LC_TIME=en_US.UTF-8        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: Etc/UTC
## tzcode source: system (glibc)
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
## 
## other attached packages:
## [1] Maaslin2_1.21.0
## 
## loaded via a namespace (and not attached):
##  [1] gtable_0.3.6        biglm_0.9-3         xfun_0.50          
##  [4] bslib_0.9.0         ggplot2_3.5.1       lattice_0.22-6     
##  [7] numDeriv_2016.8-1.1 vctrs_0.6.5         tools_4.4.2        
## [10] Rdpack_2.6.2        generics_0.1.3      parallel_4.4.2     
## [13] getopt_1.20.4       tibble_3.2.1        DEoptimR_1.1-3-1   
## [16] cluster_2.1.8       pkgconfig_2.0.3     logging_0.10-108   
## [19] pheatmap_1.0.12     Matrix_1.7-2        data.table_1.16.4  
## [22] RColorBrewer_1.1-3  lifecycle_1.0.4     farver_2.1.2       
## [25] compiler_4.4.2      munsell_0.5.1       lmerTest_3.1-3     
## [28] permute_0.9-7       htmltools_0.5.8.1   sys_3.4.3          
## [31] buildtools_1.0.0    sass_0.4.9          hash_2.2.6.3       
## [34] yaml_2.3.10         pillar_1.10.1       nloptr_2.1.1       
## [37] crayon_1.5.3        jquerylib_0.1.4     MASS_7.3-64        
## [40] cachem_1.1.0        vegan_2.6-10        reformulas_0.4.0   
## [43] boot_1.3-31         nlme_3.1-167        robustbase_0.99-4-1
## [46] tidyselect_1.2.1    digest_0.6.37       mvtnorm_1.3-3      
## [49] dplyr_1.1.4         labeling_0.4.3      maketools_1.3.1    
## [52] splines_4.4.2       pcaPP_2.0-5         fastmap_1.2.0      
## [55] grid_4.4.2          colorspace_2.1-1    cli_3.6.3          
## [58] magrittr_2.0.3      withr_3.0.2         scales_1.3.0       
## [61] rmarkdown_2.29      lme4_1.1-36         pbapply_1.7-2      
## [64] evaluate_1.0.3      knitr_1.49          rbibutils_2.3      
## [67] mgcv_1.9-1          rlang_1.1.5         Rcpp_1.0.14        
## [70] glue_1.8.0          optparse_1.7.5      DBI_1.2.3          
## [73] minqa_1.2.8         jsonlite_1.8.9      R6_2.5.1

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

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.
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.
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.

- MaAsLin2 User Manual

Maaslin2