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"
## 2024-11-08 06:00:30.358611 INFO::Writing function arguments to log file
## 2024-11-08 06:00:30.397221 INFO::Verifying options selected are valid
## 2024-11-08 06:00:30.469095 INFO::Determining format of input files
## 2024-11-08 06:00:30.469863 INFO::Input format is data samples as rows and metadata samples as rows
## 2024-11-08 06:00:30.47247 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2024-11-08 06:00:30.473109 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2024-11-08 06:00:30.474449 INFO::Filter data based on min abundance and min prevalence
## 2024-11-08 06:00:30.474919 INFO::Total samples in data: 1595
## 2024-11-08 06:00:30.475339 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2024-11-08 06:00:30.478361 INFO::Total filtered features: 0
## 2024-11-08 06:00:30.478935 INFO::Filtered feature names from abundance and prevalence filtering:
## 2024-11-08 06:00:30.485263 INFO::Total filtered features with variance filtering: 0
## 2024-11-08 06:00:30.485763 INFO::Filtered feature names from variance filtering:
## 2024-11-08 06:00:30.48619 INFO::Running selected normalization method: TSS
## 2024-11-08 06:00:31.149731 INFO::Bypass z-score application to metadata
## 2024-11-08 06:00:31.150419 INFO::Running selected transform method: LOG
## 2024-11-08 06:00:31.164381 INFO::Running selected analysis method: LM
## 2024-11-08 06:00:31.358921 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis

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

## 2024-11-08 06:00:31.582515 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2024-11-08 06:00:31.685592 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2024-11-08 06:00:31.768435 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum

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

## 2024-11-08 06:00:31.855014 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2024-11-08 06:00:31.946596 INFO::Fitting model to feature number 6, Bacteroides.caccae

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

## 2024-11-08 06:00:32.03479 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2024-11-08 06:00:32.116909 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2024-11-08 06:00:32.210327 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2024-11-08 06:00:32.294494 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2024-11-08 06:00:32.385756 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

## 2024-11-08 06:00:32.455371 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

## 2024-11-08 06:00:32.544501 INFO::Fitting model to feature number 12, Bacteroides.fragilis

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

## 2024-11-08 06:00:32.627723 INFO::Fitting model to feature number 13, Bacteroides.intestinalis

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

## 2024-11-08 06:00:32.718012 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2024-11-08 06:00:32.804837 INFO::Fitting model to feature number 15, Bacteroides.ovatus

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

## 2024-11-08 06:00:32.885222 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2024-11-08 06:00:32.975735 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2024-11-08 06:00:33.063834 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron

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

## 2024-11-08 06:00:33.149776 INFO::Fitting model to feature number 19, Bacteroides.uniformis

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

## 2024-11-08 06:00:33.255617 INFO::Fitting model to feature number 20, Bacteroides.vulgatus

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

## 2024-11-08 06:00:33.340102 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens

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

## 2024-11-08 06:00:33.424023 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2024-11-08 06:00:33.511795 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2024-11-08 06:00:33.597392 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2024-11-08 06:00:33.689704 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2024-11-08 06:00:33.774322 INFO::Fitting model to feature number 26, Parabacteroides.distasonis

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

## 2024-11-08 06:00:33.862954 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2024-11-08 06:00:33.949053 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2024-11-08 06:00:34.039393 INFO::Fitting model to feature number 29, Parabacteroides.unclassified

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

## 2024-11-08 06:00:34.122456 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2024-11-08 06:00:34.213498 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2024-11-08 06:00:34.296728 INFO::Fitting model to feature number 32, Prevotella.copri

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

## 2024-11-08 06:00:34.385001 INFO::Fitting model to feature number 33, Alistipes.finegoldii

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

## 2024-11-08 06:00:34.47207 INFO::Fitting model to feature number 34, Alistipes.onderdonkii

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

## 2024-11-08 06:00:34.556988 INFO::Fitting model to feature number 35, Alistipes.putredinis

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

## 2024-11-08 06:00:34.646598 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2024-11-08 06:00:34.73761 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2024-11-08 06:00:34.817838 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2024-11-08 06:00:35.053585 INFO::Fitting model to feature number 39, Clostridium.bolteae

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

## 2024-11-08 06:00:35.1402 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

## 2024-11-08 06:00:35.210407 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

## 2024-11-08 06:00:35.29952 INFO::Fitting model to feature number 41, Clostridium.clostridioforme

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

## 2024-11-08 06:00:35.379349 INFO::Fitting model to feature number 42, Clostridium.hathewayi

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

## 2024-11-08 06:00:35.463474 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2024-11-08 06:00:35.556722 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2024-11-08 06:00:35.644729 INFO::Fitting model to feature number 45, Clostridium.symbiosum

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

## 2024-11-08 06:00:35.726263 INFO::Fitting model to feature number 46, Flavonifractor.plautii

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

## 2024-11-08 06:00:35.811677 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2024-11-08 06:00:35.898531 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2024-11-08 06:00:35.983887 INFO::Fitting model to feature number 49, Eubacterium.rectale

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

## 2024-11-08 06:00:36.072179 INFO::Fitting model to feature number 50, Eubacterium.siraeum

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

## 2024-11-08 06:00:36.157531 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31

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

## 2024-11-08 06:00:36.240295 INFO::Fitting model to feature number 52, Eubacterium.ventriosum

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

## 2024-11-08 06:00:36.31969 INFO::Fitting model to feature number 53, Ruminococcus.gnavus

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

## 2024-11-08 06:00:36.411916 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2024-11-08 06:00:36.492647 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2024-11-08 06:00:36.575622 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2024-11-08 06:00:36.66144 INFO::Fitting model to feature number 57, Dorea.longicatena

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

## 2024-11-08 06:00:36.763113 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA

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

## 2024-11-08 06:00:36.849693 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA

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

## 2024-11-08 06:00:36.934603 INFO::Fitting model to feature number 60, Roseburia.hominis

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

## 2024-11-08 06:00:37.025189 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2024-11-08 06:00:37.103591 INFO::Fitting model to feature number 62, Roseburia.inulinivorans

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

## 2024-11-08 06:00:37.186247 INFO::Fitting model to feature number 63, Roseburia.unclassified

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

## 2024-11-08 06:00:37.273784 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2024-11-08 06:00:37.361967 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2024-11-08 06:00:37.44701 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2024-11-08 06:00:37.536635 INFO::Fitting model to feature number 67, Ruminococcus.bromii

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

## 2024-11-08 06:00:37.614917 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2024-11-08 06:00:37.696218 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2024-11-08 06:00:37.787227 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified

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

## 2024-11-08 06:00:37.869738 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2024-11-08 06:00:37.948955 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2024-11-08 06:00:38.037501 INFO::Fitting model to feature number 73, Dialister.invisus

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

## 2024-11-08 06:00:38.117103 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2024-11-08 06:00:38.201451 INFO::Fitting model to feature number 75, Veillonella.dispar

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

## 2024-11-08 06:00:38.289039 INFO::Fitting model to feature number 76, Veillonella.parvula

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

## 2024-11-08 06:00:38.368574 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2024-11-08 06:00:38.453497 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2024-11-08 06:00:38.538562 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2024-11-08 06:00:38.62048 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2024-11-08 06:00:38.709872 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2024-11-08 06:00:38.798594 INFO::Fitting model to feature number 82, Escherichia.coli
## 2024-11-08 06:00:38.888788 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2024-11-08 06:00:38.968973 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae

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

## 2024-11-08 06:00:39.049646 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae

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

## 2024-11-08 06:00:39.137442 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2024-11-08 06:00:39.222064 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2024-11-08 06:00:39.335786 INFO::Counting total values for each feature
## 2024-11-08 06:00:39.355814 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2024-11-08 06:00:39.41809 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2024-11-08 06:00:39.480731 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2024-11-08 06:00:39.575962 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2024-11-08 06:00:39.613607 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2024-11-08 06:00:39.635225 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2024-11-08 06:00:39.638536 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2024-11-08 06:00:39.641677 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
## 2024-11-08 06:00:39.6432 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2024-11-08 06:00:39.770238 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2024-11-08 06:00:39.77366 INFO::Plotting associations from most to least significant, grouped by metadata
## 2024-11-08 06:00:39.774213 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2024-11-08 06:00:39.775175 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii

## 2024-11-08 06:00:39.997503 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified

## 2024-11-08 06:00:40.166072 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis

## 2024-11-08 06:00:40.316122 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale

## 2024-11-08 06:00:40.470412 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus

## 2024-11-08 06:00:40.633215 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus

## 2024-11-08 06:00:40.790182 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum

## 2024-11-08 06:00:40.946848 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified

## 2024-11-08 06:00:41.096586 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans

## 2024-11-08 06:00:41.251235 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis

## 2024-11-08 06:00:41.405428 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron

## 2024-11-08 06:00:41.560139 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis

## 2024-11-08 06:00:41.713982 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis

## 2024-11-08 06:00:41.872461 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei

## 2024-11-08 06:00:42.02007 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii

## 2024-11-08 06:00:42.171789 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens

## 2024-11-08 06:00:42.322726 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum

## 2024-11-08 06:00:42.476212 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena

## 2024-11-08 06:00:42.630806 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae

## 2024-11-08 06:00:42.786862 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-11-08 06:00:42.946005 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli

## 2024-11-08 06:00:43.101164 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae

## 2024-11-08 06:00:43.249184 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified

## 2024-11-08 06:00:43.395314 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii

## 2024-11-08 06:00:43.549943 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens

## 2024-11-08 06:00:43.702026 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris

## 2024-11-08 06:00:43.856259 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes

## 2024-11-08 06:00:44.008733 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis

## 2024-11-08 06:00:44.161002 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii

## 2024-11-08 06:00:44.332407 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii

## 2024-11-08 06:00:44.486517 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis

## 2024-11-08 06:00:44.644679 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara

## 2024-11-08 06:00:44.794082 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum

## 2024-11-08 06:00:44.948591 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae

## 2024-11-08 06:00:45.106263 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified

## 2024-11-08 06:00:45.264373 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens

## 2024-11-08 06:00:45.416959 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus

## 2024-11-08 06:00:45.576584 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme

## 2024-11-08 06:00:45.724903 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii

## 2024-11-08 06:00:45.880953 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii

## 2024-11-08 06:00:46.035916 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum

## 2024-11-08 06:00:46.195629 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris

## 2024-11-08 06:00:46.354308 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47

## 2024-11-08 06:00:46.508896 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus

## 2024-11-08 06:00:46.668698 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii

## 2024-11-08 06:00:46.827938 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis

## 2024-11-08 06:00:46.991598 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified

## 2024-11-08 06:00:47.142073 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques

## 2024-11-08 06:00:47.295883 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified

## 2024-11-08 06:00:47.459622 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus

## 2024-11-08 06:00:47.615689 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis

## 2024-11-08 06:00:47.771823 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified

## 2024-11-08 06:00:47.931589 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified

## 2024-11-08 06:00:48.110167 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis

## 2024-11-08 06:00:48.267442 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis

## 2024-11-08 06:00:48.426535 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31

## 2024-11-08 06:00:48.58129 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae

## 2024-11-08 06:00:48.733793 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8

## 2024-11-08 06:00:48.890013 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae

## 2024-11-08 06:00:49.048675 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii

## 2024-11-08 06:00:49.207216 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii

## 2024-11-08 06:00:49.372015 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum

## 2024-11-08 06:00:49.530647 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA

## 2024-11-08 06:00:51.539313 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2024-11-08 06:00:51.540835 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified

## 2024-11-08 06:00:51.676493 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii

## 2024-11-08 06:00:51.843755 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae

## 2024-11-08 06:00:52.010626 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified

## 2024-11-08 06:00:52.180731 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis

## 2024-11-08 06:00:52.523787 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum

## 2024-11-08 06:00:52.677037 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus

## 2024-11-08 06:00:52.823351 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis

## 2024-11-08 06:00:52.977944 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii

## 2024-11-08 06:00:53.123071 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale

## 2024-11-08 06:00:53.271474 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis

## 2024-11-08 06:00:53.425568 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis

## 2024-11-08 06:00:53.57717 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-11-08 06:00:53.726789 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii

## 2024-11-08 06:00:53.877483 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens

## 2024-11-08 06:00:54.022049 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris

## 2024-11-08 06:00:54.172379 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii

## 2024-11-08 06:00:54.321407 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis

## 2024-11-08 06:00:54.471777 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum

## 2024-11-08 06:00:54.617622 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus

## 2024-11-08 06:00:54.766958 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii

## 2024-11-08 06:00:54.917756 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii

## 2024-11-08 06:00:55.065524 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae

## 2024-11-08 06:00:55.217137 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus

## 2024-11-08 06:00:55.367508 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum

## 2024-11-08 06:00:55.515015 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae

## 2024-11-08 06:00:55.667582 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae

## 2024-11-08 06:00:55.811437 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum

## 2024-11-08 06:00:55.958054 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis

## 2024-11-08 06:00:56.126296 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii

## 2024-11-08 06:00:56.281812 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques

## 2024-11-08 06:00:56.425125 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8

## 2024-11-08 06:00:56.585976 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum

## 2024-11-08 06:00:56.73768 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum

## 2024-11-08 06:00:56.899359 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis

## 2024-11-08 06:00:57.054762 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens

## 2024-11-08 06:00:57.209502 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi

## 2024-11-08 06:00:57.357818 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified

## 2024-11-08 06:00:57.512628 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens

## 2024-11-08 06:00:57.669344 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus

## 2024-11-08 06:00:57.827806 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae

## 2024-11-08 06:00:57.987868 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus

## 2024-11-08 06:00:58.145046 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris

## 2024-11-08 06:00:58.299345 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47

## 2024-11-08 06:01:00.241632 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2024-11-08 06:01:00.242872 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii

## 2024-11-08 06:01:00.378322 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale

## 2024-11-08 06:01:00.538422 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31

## 2024-11-08 06:01:00.705467 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis

## 2024-11-08 06:01:00.862475 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified

## 2024-11-08 06:01:01.026776 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-11-08 06:01:01.194406 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques

## 2024-11-08 06:01:01.352979 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum

## 2024-11-08 06:01:01.51346 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus

## 2024-11-08 06:01:01.676402 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli

## 2024-11-08 06:01:01.837151 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum

## 2024-11-08 06:01:01.998401 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified

## 2024-11-08 06:01:02.163767 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis

## 2024-11-08 06:01:02.325897 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens

## 2024-11-08 06:01:02.492738 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae

## 2024-11-08 06:01:02.65735 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii

## 2024-11-08 06:01:02.808698 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus

## 2024-11-08 06:01:02.963805 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified

## 2024-11-08 06:01:03.131889 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis

## 2024-11-08 06:01:03.287634 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii

## 2024-11-08 06:01:03.449313 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans

## 2024-11-08 06:01:03.610145 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii

## 2024-11-08 06:01:03.775698 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified

## 2024-11-08 06:01:03.939383 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii

## 2024-11-08 06:01:04.122829 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis

## 2024-11-08 06:01:04.290633 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri

## 2024-11-08 06:01:04.457096 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei

## 2024-11-08 06:01:04.605961 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis

## 2024-11-08 06:01:04.775878 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii

## 2024-11-08 06:01:06.787184 INFO::Plotting data for metadata number 4, antibiotics
## 2024-11-08 06:01:06.788364 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans

## 2024-11-08 06:01:06.924079 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis

## 2024-11-08 06:01:07.080803 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale

## 2024-11-08 06:01:07.234801 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus

## 2024-11-08 06:01:07.393063 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis

## 2024-11-08 06:01:07.550478 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena

## 2024-11-08 06:01:07.708802 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus

## 2024-11-08 06:01:07.865342 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii

## 2024-11-08 06:01:08.015477 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum

## 2024-11-08 06:01:08.167445 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae

## 2024-11-08 06:01:08.343639 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis

## 2024-11-08 06:01:08.484318 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii

## 2024-11-08 06:01:08.645074 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii

## 2024-11-08 06:01:08.811577 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified

## 2024-11-08 06:01:08.972347 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum

## 2024-11-08 06:01:09.133492 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-11-08 06:01:09.284443 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii

## 2024-11-08 06:01:09.441756 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31

## 2024-11-08 06:01:09.598711 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis

## 2024-11-08 06:01:09.754786 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii

## 2024-11-08 06:01:09.917406 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis

## 2024-11-08 06:01:10.082158 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens

## 2024-11-08 06:01:10.232231 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens

## 2024-11-08 06:01:10.387155 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron

## 2024-11-08 06:01:10.547793 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii

## 2024-11-08 06:01:10.714036 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae

## 2024-11-08 06:01:10.871024 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum

## 2024-11-08 06:01:11.032156 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques

## 2024-11-08 06:01:11.18732 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum

## 2024-11-08 06:01:11.349688 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis

## 2024-11-08 06:01:11.503886 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified

## 2024-11-08 06:01:11.657665 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar

## 2024-11-08 06:01:11.80856 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica

## 2024-11-08 06:01:11.966419 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8

## 2024-11-08 06:01:12.122839 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile

## 2024-11-08 06:01:12.309184 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47

## 2024-11-08 06:01:12.471364 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA

## 2024-11-08 06:01:12.636018 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila

## 2024-11-08 06:01:12.790342 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae

## 2024-11-08 06:01:12.943953 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus

## 2024-11-08 06:01:14.925157 INFO::Plotting data for metadata number 5, age
## 2024-11-08 06:01:14.926592 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()`).

## 2024-11-08 06:01:15.111422 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()`).

## 2024-11-08 06:01:15.335039 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()`).

## 2024-11-08 06:01:15.504084 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()`).

## 2024-11-08 06:01:15.683198 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()`).

## 2024-11-08 06:01:15.845911 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()`).

## 2024-11-08 06:01:16.010012 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()`).

## 2024-11-08 06:01:16.183215 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()`).

## 2024-11-08 06:01:16.527977 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()`).

## 2024-11-08 06:01:16.669382 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()`).

## 2024-11-08 06:01:16.810821 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()`).

## 2024-11-08 06:01:16.949914 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()`).

## 2024-11-08 06:01:17.089714 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()`).

## 2024-11-08 06:01:17.232366 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()`).

## 2024-11-08 06:01:17.36915 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()`).

## 2024-11-08 06:01:17.503074 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()`).

## 2024-11-08 06:01:17.641255 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()`).

## 2024-11-08 06:01:17.776908 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()`).

## 2024-11-08 06:01:17.911145 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()`).

## 2024-11-08 06:01:18.049156 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()`).

## 2024-11-08 06:01:18.190456 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()`).

## 2024-11-08 06:01:18.327814 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()`).

## 2024-11-08 06:01:20.207949 INFO::Plotting data for metadata number 6, diagnosis
## 2024-11-08 06:01:20.209145 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis

## 2024-11-08 06:01:20.346953 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2024-11-08 06:01:20.501996 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2024-11-08 06:01:20.657906 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2024-11-08 06:01:20.81057 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2024-11-08 06:01:20.967849 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme

## 2024-11-08 06:01:21.123032 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2024-11-08 06:01:21.277842 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2024-11-08 06:01:21.434693 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2024-11-08 06:01:21.589543 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2024-11-08 06:01:21.750264 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans

## 2024-11-08 06:01:21.907985 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2024-11-08 06:01:22.066389 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2024-11-08 06:01:22.220299 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2024-11-08 06:01:22.377027 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii

## 2024-11-08 06:01:22.532819 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified

## 2024-11-08 06:01:22.6841 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2024-11-08 06:01:22.843357 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2024-11-08 06:01:23.006654 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2024-11-08 06:01:23.166609 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2024-11-08 06:01:23.33738 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum

## 2024-11-08 06:01:23.495874 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2024-11-08 06:01:23.652701 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2024-11-08 06:01:23.818125 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2024-11-08 06:01:23.979469 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2024-11-08 06:01:24.14009 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified

## 2024-11-08 06:01:24.299506 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2024-11-08 06:01:24.467767 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2024-11-08 06:01:24.630987 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified

## 2024-11-08 06:01:24.796782 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris

## 2024-11-08 06:01:24.959453 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2024-11-08 06:01:25.120717 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus

## 2024-11-08 06:01:25.280683 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis

## 2024-11-08 06:01:25.44314 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2024-11-08 06:01:25.606545 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus

## 2024-11-08 06:01:25.775757 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis

## 2024-11-08 06:01:25.936562 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2024-11-08 06:01:26.100204 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii

## 2024-11-08 06:01:26.265681 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2024-11-08 06:01:26.424001 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2024-11-08 06:01:26.583806 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2024-11-08 06:01:26.746044 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2024-11-08 06:01:26.931111 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_0.49          
##  [4] bslib_0.8.0         ggplot2_3.5.1       lattice_0.22-6     
##  [7] numDeriv_2016.8-1.1 vctrs_0.6.5         tools_4.4.2        
## [10] generics_0.1.3      parallel_4.4.2      getopt_1.20.4      
## [13] tibble_3.2.1        fansi_1.0.6         DEoptimR_1.1-3     
## [16] cluster_2.1.6       pkgconfig_2.0.3     logging_0.10-108   
## [19] pheatmap_1.0.12     Matrix_1.7-1        data.table_1.16.2  
## [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_1.9.0        nloptr_2.1.1       
## [37] crayon_1.5.3        jquerylib_0.1.4     MASS_7.3-61        
## [40] cachem_1.1.0        vegan_2.6-8         boot_1.3-31        
## [43] nlme_3.1-166        robustbase_0.99-4-1 tidyselect_1.2.1   
## [46] digest_0.6.37       mvtnorm_1.3-2       dplyr_1.1.4        
## [49] labeling_0.4.3      maketools_1.3.1     splines_4.4.2      
## [52] pcaPP_2.0-5         fastmap_1.2.0       grid_4.4.2         
## [55] colorspace_2.1-1    cli_3.6.3           magrittr_2.0.3     
## [58] utf8_1.2.4          withr_3.0.2         scales_1.3.0       
## [61] rmarkdown_2.29      lme4_1.1-35.5       pbapply_1.7-2      
## [64] evaluate_1.0.1      knitr_1.48          mgcv_1.9-1         
## [67] rlang_1.1.4         Rcpp_1.0.13-1       glue_1.8.0         
## [70] optparse_1.7.5      DBI_1.2.3           minqa_1.2.8        
## [73] 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