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"
## 2024-08-29 03:37:04.496736 INFO::Writing function arguments to log file
## 2024-08-29 03:37:04.571378 INFO::Verifying options selected are valid
## 2024-08-29 03:37:04.59052 INFO::Determining format of input files
## 2024-08-29 03:37:04.59117 INFO::Input format is data samples as rows and metadata samples as rows
## 2024-08-29 03:37:04.59437 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2024-08-29 03:37:04.595067 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2024-08-29 03:37:04.596599 INFO::Filter data based on min abundance and min prevalence
## 2024-08-29 03:37:04.597064 INFO::Total samples in data: 1595
## 2024-08-29 03:37:04.597532 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2024-08-29 03:37:04.6011 INFO::Total filtered features: 0
## 2024-08-29 03:37:04.601682 INFO::Filtered feature names from abundance and prevalence filtering:
## 2024-08-29 03:37:04.60804 INFO::Total filtered features with variance filtering: 0
## 2024-08-29 03:37:04.608541 INFO::Filtered feature names from variance filtering:
## 2024-08-29 03:37:04.60899 INFO::Running selected normalization method: TSS
## 2024-08-29 03:37:05.302446 INFO::Bypass z-score application to metadata
## 2024-08-29 03:37:05.303205 INFO::Running selected transform method: LOG
## 2024-08-29 03:37:05.325177 INFO::Running selected analysis method: LM
## 2024-08-29 03:37:05.546495 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis

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

## 2024-08-29 03:37:05.734553 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2024-08-29 03:37:05.849116 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2024-08-29 03:37:05.934094 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum

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

## 2024-08-29 03:37:06.022445 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2024-08-29 03:37:06.117738 INFO::Fitting model to feature number 6, Bacteroides.caccae

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

## 2024-08-29 03:37:06.20888 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2024-08-29 03:37:06.294837 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2024-08-29 03:37:06.391542 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2024-08-29 03:37:06.479718 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2024-08-29 03:37:06.573673 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

## 2024-08-29 03:37:06.645238 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

## 2024-08-29 03:37:06.736937 INFO::Fitting model to feature number 12, Bacteroides.fragilis

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

## 2024-08-29 03:37:06.822421 INFO::Fitting model to feature number 13, Bacteroides.intestinalis

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

## 2024-08-29 03:37:06.915785 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2024-08-29 03:37:07.000916 INFO::Fitting model to feature number 15, Bacteroides.ovatus

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

## 2024-08-29 03:37:07.088043 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2024-08-29 03:37:07.180511 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2024-08-29 03:37:07.267455 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron

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

## 2024-08-29 03:37:07.356163 INFO::Fitting model to feature number 19, Bacteroides.uniformis

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

## 2024-08-29 03:37:07.604378 INFO::Fitting model to feature number 20, Bacteroides.vulgatus

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

## 2024-08-29 03:37:07.690175 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens

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

## 2024-08-29 03:37:07.776395 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2024-08-29 03:37:07.868542 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2024-08-29 03:37:07.95521 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2024-08-29 03:37:08.044522 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2024-08-29 03:37:08.136411 INFO::Fitting model to feature number 26, Parabacteroides.distasonis

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

## 2024-08-29 03:37:08.220257 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2024-08-29 03:37:08.308166 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2024-08-29 03:37:08.399577 INFO::Fitting model to feature number 29, Parabacteroides.unclassified

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

## 2024-08-29 03:37:08.482556 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2024-08-29 03:37:08.572176 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2024-08-29 03:37:08.66182 INFO::Fitting model to feature number 32, Prevotella.copri

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

## 2024-08-29 03:37:08.748862 INFO::Fitting model to feature number 33, Alistipes.finegoldii

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

## 2024-08-29 03:37:08.831993 INFO::Fitting model to feature number 34, Alistipes.onderdonkii

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

## 2024-08-29 03:37:08.919976 INFO::Fitting model to feature number 35, Alistipes.putredinis

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

## 2024-08-29 03:37:09.008708 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2024-08-29 03:37:09.096114 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2024-08-29 03:37:09.178699 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2024-08-29 03:37:09.270502 INFO::Fitting model to feature number 39, Clostridium.bolteae

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

## 2024-08-29 03:37:09.359434 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

## 2024-08-29 03:37:09.430158 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

## 2024-08-29 03:37:09.520452 INFO::Fitting model to feature number 41, Clostridium.clostridioforme

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

## 2024-08-29 03:37:09.603437 INFO::Fitting model to feature number 42, Clostridium.hathewayi

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

## 2024-08-29 03:37:09.688816 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2024-08-29 03:37:09.789043 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2024-08-29 03:37:09.874216 INFO::Fitting model to feature number 45, Clostridium.symbiosum

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

## 2024-08-29 03:37:09.958747 INFO::Fitting model to feature number 46, Flavonifractor.plautii

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

## 2024-08-29 03:37:10.050023 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2024-08-29 03:37:10.135741 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2024-08-29 03:37:10.231058 INFO::Fitting model to feature number 49, Eubacterium.rectale

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

## 2024-08-29 03:37:10.320088 INFO::Fitting model to feature number 50, Eubacterium.siraeum

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

## 2024-08-29 03:37:10.403896 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31

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

## 2024-08-29 03:37:10.493602 INFO::Fitting model to feature number 52, Eubacterium.ventriosum

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

## 2024-08-29 03:37:10.57405 INFO::Fitting model to feature number 53, Ruminococcus.gnavus

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

## 2024-08-29 03:37:10.820177 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2024-08-29 03:37:10.903088 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2024-08-29 03:37:10.987352 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2024-08-29 03:37:11.076017 INFO::Fitting model to feature number 57, Dorea.longicatena

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

## 2024-08-29 03:37:11.166947 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA

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

## 2024-08-29 03:37:11.254344 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA

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

## 2024-08-29 03:37:11.341791 INFO::Fitting model to feature number 60, Roseburia.hominis

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

## 2024-08-29 03:37:11.434304 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2024-08-29 03:37:11.51466 INFO::Fitting model to feature number 62, Roseburia.inulinivorans

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

## 2024-08-29 03:37:11.600054 INFO::Fitting model to feature number 63, Roseburia.unclassified

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

## 2024-08-29 03:37:11.689234 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2024-08-29 03:37:11.783604 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2024-08-29 03:37:11.86939 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2024-08-29 03:37:11.960539 INFO::Fitting model to feature number 67, Ruminococcus.bromii

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

## 2024-08-29 03:37:12.040677 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2024-08-29 03:37:12.124301 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2024-08-29 03:37:12.216552 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified

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

## 2024-08-29 03:37:12.301995 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2024-08-29 03:37:12.387836 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2024-08-29 03:37:12.479695 INFO::Fitting model to feature number 73, Dialister.invisus

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

## 2024-08-29 03:37:12.561357 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2024-08-29 03:37:12.647697 INFO::Fitting model to feature number 75, Veillonella.dispar

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

## 2024-08-29 03:37:12.737814 INFO::Fitting model to feature number 76, Veillonella.parvula

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

## 2024-08-29 03:37:12.819902 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2024-08-29 03:37:12.902308 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2024-08-29 03:37:12.992379 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2024-08-29 03:37:13.076128 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2024-08-29 03:37:13.168062 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2024-08-29 03:37:13.258564 INFO::Fitting model to feature number 82, Escherichia.coli
## 2024-08-29 03:37:13.345926 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2024-08-29 03:37:13.429842 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae

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

## 2024-08-29 03:37:13.511731 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae

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

## 2024-08-29 03:37:13.601829 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2024-08-29 03:37:13.689312 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2024-08-29 03:37:13.808479 INFO::Counting total values for each feature
## 2024-08-29 03:37:13.828302 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2024-08-29 03:37:13.891 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2024-08-29 03:37:13.955106 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2024-08-29 03:37:14.045531 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2024-08-29 03:37:14.245234 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2024-08-29 03:37:14.267519 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2024-08-29 03:37:14.270981 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2024-08-29 03:37:14.274218 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
## 2024-08-29 03:37:14.275759 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2024-08-29 03:37:14.404575 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2024-08-29 03:37:14.408311 INFO::Plotting associations from most to least significant, grouped by metadata
## 2024-08-29 03:37:14.408863 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2024-08-29 03:37:14.409889 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii

## 2024-08-29 03:37:14.618581 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified

## 2024-08-29 03:37:14.789578 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis

## 2024-08-29 03:37:14.946855 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale

## 2024-08-29 03:37:15.103658 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus

## 2024-08-29 03:37:15.260555 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus

## 2024-08-29 03:37:15.416359 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum

## 2024-08-29 03:37:15.571906 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified

## 2024-08-29 03:37:15.728663 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans

## 2024-08-29 03:37:15.90846 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis

## 2024-08-29 03:37:16.066623 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron

## 2024-08-29 03:37:16.225583 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis

## 2024-08-29 03:37:16.382862 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis

## 2024-08-29 03:37:16.538564 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei

## 2024-08-29 03:37:16.695231 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii

## 2024-08-29 03:37:16.85232 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens

## 2024-08-29 03:37:17.007874 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum

## 2024-08-29 03:37:17.165645 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena

## 2024-08-29 03:37:17.321342 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae

## 2024-08-29 03:37:17.478834 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-08-29 03:37:17.636821 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli

## 2024-08-29 03:37:17.797517 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae

## 2024-08-29 03:37:17.953401 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified

## 2024-08-29 03:37:18.102547 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii

## 2024-08-29 03:37:18.258923 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens

## 2024-08-29 03:37:18.414025 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris

## 2024-08-29 03:37:18.572772 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes

## 2024-08-29 03:37:18.729955 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis

## 2024-08-29 03:37:18.889884 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii

## 2024-08-29 03:37:19.067221 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii

## 2024-08-29 03:37:19.224586 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis

## 2024-08-29 03:37:19.387588 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara

## 2024-08-29 03:37:19.542243 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum

## 2024-08-29 03:37:19.705208 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae

## 2024-08-29 03:37:19.866225 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified

## 2024-08-29 03:37:20.028581 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens

## 2024-08-29 03:37:20.187421 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus

## 2024-08-29 03:37:20.35027 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme

## 2024-08-29 03:37:20.511772 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii

## 2024-08-29 03:37:20.67754 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii

## 2024-08-29 03:37:20.837594 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum

## 2024-08-29 03:37:21.003351 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris

## 2024-08-29 03:37:21.166254 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47

## 2024-08-29 03:37:21.324969 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus

## 2024-08-29 03:37:21.489521 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii

## 2024-08-29 03:37:21.655042 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis

## 2024-08-29 03:37:21.817541 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified

## 2024-08-29 03:37:21.980017 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques

## 2024-08-29 03:37:22.136611 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified

## 2024-08-29 03:37:22.305067 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus

## 2024-08-29 03:37:22.485411 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis

## 2024-08-29 03:37:22.653754 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified

## 2024-08-29 03:37:22.825634 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified

## 2024-08-29 03:37:22.993759 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis

## 2024-08-29 03:37:23.156745 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis

## 2024-08-29 03:37:23.323038 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31

## 2024-08-29 03:37:23.48769 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae

## 2024-08-29 03:37:23.65068 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8

## 2024-08-29 03:37:23.815315 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae

## 2024-08-29 03:37:23.981167 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii

## 2024-08-29 03:37:24.146326 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii

## 2024-08-29 03:37:24.315589 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum

## 2024-08-29 03:37:24.485449 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA

## 2024-08-29 03:37:26.59298 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2024-08-29 03:37:26.594359 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified

## 2024-08-29 03:37:26.74559 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii

## 2024-08-29 03:37:26.92443 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae

## 2024-08-29 03:37:27.100762 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified

## 2024-08-29 03:37:27.273946 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis

## 2024-08-29 03:37:27.450044 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum

## 2024-08-29 03:37:27.626894 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus

## 2024-08-29 03:37:27.797133 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis

## 2024-08-29 03:37:27.974021 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii

## 2024-08-29 03:37:28.14656 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale

## 2024-08-29 03:37:28.324722 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis

## 2024-08-29 03:37:28.505987 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis

## 2024-08-29 03:37:28.684007 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-08-29 03:37:28.857318 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii

## 2024-08-29 03:37:29.034639 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens

## 2024-08-29 03:37:29.209798 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris

## 2024-08-29 03:37:29.384137 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii

## 2024-08-29 03:37:29.556577 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis

## 2024-08-29 03:37:29.749012 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum

## 2024-08-29 03:37:29.928831 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus

## 2024-08-29 03:37:30.106302 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii

## 2024-08-29 03:37:30.284923 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii

## 2024-08-29 03:37:30.463721 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae

## 2024-08-29 03:37:30.649356 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus

## 2024-08-29 03:37:30.828713 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum

## 2024-08-29 03:37:31.006899 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae

## 2024-08-29 03:37:31.189991 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae

## 2024-08-29 03:37:31.364548 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum

## 2024-08-29 03:37:31.540942 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis

## 2024-08-29 03:37:31.715226 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii

## 2024-08-29 03:37:31.8912 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques

## 2024-08-29 03:37:32.065088 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8

## 2024-08-29 03:37:32.246937 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum

## 2024-08-29 03:37:32.417207 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum

## 2024-08-29 03:37:32.604071 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis

## 2024-08-29 03:37:32.78196 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens

## 2024-08-29 03:37:32.960524 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi

## 2024-08-29 03:37:33.3027 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified

## 2024-08-29 03:37:33.44978 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens

## 2024-08-29 03:37:33.626963 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus

## 2024-08-29 03:37:33.77566 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae

## 2024-08-29 03:37:33.932997 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus

## 2024-08-29 03:37:34.081631 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris

## 2024-08-29 03:37:34.229485 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47

## 2024-08-29 03:37:36.102864 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2024-08-29 03:37:36.10409 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii

## 2024-08-29 03:37:36.236025 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale

## 2024-08-29 03:37:36.386882 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31

## 2024-08-29 03:37:36.543669 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis

## 2024-08-29 03:37:36.685745 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified

## 2024-08-29 03:37:36.841829 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-08-29 03:37:36.994587 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques

## 2024-08-29 03:37:37.145055 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum

## 2024-08-29 03:37:37.301661 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus

## 2024-08-29 03:37:37.45131 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli

## 2024-08-29 03:37:37.601454 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum

## 2024-08-29 03:37:37.75059 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified

## 2024-08-29 03:37:37.906991 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis

## 2024-08-29 03:37:38.057445 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens

## 2024-08-29 03:37:38.213784 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae

## 2024-08-29 03:37:38.365039 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii

## 2024-08-29 03:37:38.512173 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus

## 2024-08-29 03:37:38.65884 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified

## 2024-08-29 03:37:38.81605 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis

## 2024-08-29 03:37:38.962289 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii

## 2024-08-29 03:37:39.114143 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans

## 2024-08-29 03:37:39.264953 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii

## 2024-08-29 03:37:39.420793 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified

## 2024-08-29 03:37:39.571661 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii

## 2024-08-29 03:37:39.722034 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis

## 2024-08-29 03:37:39.873039 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri

## 2024-08-29 03:37:40.031966 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei

## 2024-08-29 03:37:40.178075 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis

## 2024-08-29 03:37:40.366906 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii

## 2024-08-29 03:37:42.270169 INFO::Plotting data for metadata number 4, antibiotics
## 2024-08-29 03:37:42.271425 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans

## 2024-08-29 03:37:42.391841 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis

## 2024-08-29 03:37:42.534354 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale

## 2024-08-29 03:37:42.6853 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus

## 2024-08-29 03:37:42.82914 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis

## 2024-08-29 03:37:42.976992 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena

## 2024-08-29 03:37:43.118944 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus

## 2024-08-29 03:37:43.268943 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii

## 2024-08-29 03:37:43.405044 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum

## 2024-08-29 03:37:43.550493 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae

## 2024-08-29 03:37:43.694993 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis

## 2024-08-29 03:37:43.835858 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii

## 2024-08-29 03:37:43.975914 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii

## 2024-08-29 03:37:44.12117 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified

## 2024-08-29 03:37:44.269717 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum

## 2024-08-29 03:37:44.410264 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA

## 2024-08-29 03:37:44.557639 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii

## 2024-08-29 03:37:44.699997 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31

## 2024-08-29 03:37:44.844866 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis

## 2024-08-29 03:37:44.980498 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii

## 2024-08-29 03:37:45.12977 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis

## 2024-08-29 03:37:45.271807 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens

## 2024-08-29 03:37:45.416786 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens

## 2024-08-29 03:37:45.559454 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron

## 2024-08-29 03:37:45.706655 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii

## 2024-08-29 03:37:45.850771 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae

## 2024-08-29 03:37:45.987377 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum

## 2024-08-29 03:37:46.139277 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques

## 2024-08-29 03:37:46.276017 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum

## 2024-08-29 03:37:46.429484 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis

## 2024-08-29 03:37:46.570498 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified

## 2024-08-29 03:37:46.718808 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar

## 2024-08-29 03:37:46.857126 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica

## 2024-08-29 03:37:47.004852 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8

## 2024-08-29 03:37:47.145226 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile

## 2024-08-29 03:37:47.315988 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47

## 2024-08-29 03:37:47.456063 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA

## 2024-08-29 03:37:47.610019 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila

## 2024-08-29 03:37:47.751622 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae

## 2024-08-29 03:37:47.900503 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus

## 2024-08-29 03:37:49.745631 INFO::Plotting data for metadata number 5, age
## 2024-08-29 03:37:49.746875 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()`).

## 2024-08-29 03:37:49.917353 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()`).

## 2024-08-29 03:37:50.118554 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()`).

## 2024-08-29 03:37:50.269268 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()`).

## 2024-08-29 03:37:50.422525 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()`).

## 2024-08-29 03:37:50.577164 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()`).

## 2024-08-29 03:37:50.732263 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()`).

## 2024-08-29 03:37:50.886286 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()`).

## 2024-08-29 03:37:51.04234 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()`).

## 2024-08-29 03:37:51.189916 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()`).

## 2024-08-29 03:37:51.347083 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()`).

## 2024-08-29 03:37:51.487608 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()`).

## 2024-08-29 03:37:51.646974 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()`).

## 2024-08-29 03:37:51.805289 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()`).

## 2024-08-29 03:37:51.947452 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()`).

## 2024-08-29 03:37:52.099075 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()`).

## 2024-08-29 03:37:52.242261 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()`).

## 2024-08-29 03:37:52.394211 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()`).

## 2024-08-29 03:37:52.543916 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()`).

## 2024-08-29 03:37:52.687733 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()`).

## 2024-08-29 03:37:52.846531 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()`).

## 2024-08-29 03:37:52.989655 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()`).

## 2024-08-29 03:37:55.036061 INFO::Plotting data for metadata number 6, diagnosis
## 2024-08-29 03:37:55.037475 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis

## 2024-08-29 03:37:55.173558 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2024-08-29 03:37:55.340259 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2024-08-29 03:37:55.493648 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2024-08-29 03:37:55.665429 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2024-08-29 03:37:55.823853 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme

## 2024-08-29 03:37:55.990674 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2024-08-29 03:37:56.143419 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2024-08-29 03:37:56.3115 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2024-08-29 03:37:56.463735 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2024-08-29 03:37:56.633535 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans

## 2024-08-29 03:37:56.795636 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii

## 2024-08-29 03:37:56.959381 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii

## 2024-08-29 03:37:57.123001 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2024-08-29 03:37:57.281352 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii

## 2024-08-29 03:37:57.447979 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified

## 2024-08-29 03:37:57.598773 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum

## 2024-08-29 03:37:57.765618 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2024-08-29 03:37:57.923406 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified

## 2024-08-29 03:37:58.092557 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2024-08-29 03:37:58.244867 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum

## 2024-08-29 03:37:58.413771 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis

## 2024-08-29 03:37:58.567691 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2024-08-29 03:37:58.743937 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila

## 2024-08-29 03:37:58.896727 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis

## 2024-08-29 03:37:59.062608 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified

## 2024-08-29 03:37:59.219589 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum

## 2024-08-29 03:37:59.385374 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii

## 2024-08-29 03:37:59.551876 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified

## 2024-08-29 03:37:59.710132 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris

## 2024-08-29 03:37:59.876457 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2024-08-29 03:38:00.029766 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus

## 2024-08-29 03:38:00.193208 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis

## 2024-08-29 03:38:00.349687 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2024-08-29 03:38:00.543936 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus

## 2024-08-29 03:38:00.704505 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis

## 2024-08-29 03:38:00.875517 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale

## 2024-08-29 03:38:01.033298 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii

## 2024-08-29 03:38:01.211692 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus

## 2024-08-29 03:38:01.36605 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii

## 2024-08-29 03:38:01.535543 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis

## 2024-08-29 03:38:01.692466 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae

## 2024-08-29 03:38:01.86565 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 4.4.1 (2024-06-14)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 24.04 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_1.19.0
## 
## loaded via a namespace (and not attached):
##  [1] gtable_0.3.5        biglm_0.9-3         xfun_0.47          
##  [4] bslib_0.8.0         ggplot2_3.5.1       lattice_0.22-6     
##  [7] numDeriv_2016.8-1.1 vctrs_0.6.5         tools_4.4.1        
## [10] generics_0.1.3      parallel_4.4.1      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_1.7-0        data.table_1.16.0  
## [22] RColorBrewer_1.1-3  lifecycle_1.0.4     farver_2.1.2       
## [25] compiler_4.4.1      munsell_0.5.1       lmerTest_3.1-3     
## [28] permute_0.9-7       htmltools_0.5.8.1   sys_3.4.2          
## [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   tidyselect_1.2.1   
## [46] digest_0.6.37       mvtnorm_1.2-6       dplyr_1.1.4        
## [49] labeling_0.4.3      maketools_1.3.0     splines_4.4.1      
## [52] pcaPP_2.0-5         fastmap_1.2.0       grid_4.4.1         
## [55] colorspace_2.1-1    cli_3.6.3           magrittr_2.0.3     
## [58] utf8_1.2.4          withr_3.0.1         scales_1.3.0       
## [61] rmarkdown_2.28      lme4_1.1-35.5       pbapply_1.7-2      
## [64] evaluate_0.24.0     knitr_1.48          mgcv_1.9-1         
## [67] rlang_1.1.4         Rcpp_1.0.13         glue_1.7.0         
## [70] optparse_1.7.5      DBI_1.2.3           minqa_1.2.8        
## [73] jsonlite_1.8.8      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