MaAsLin2 User Manual

MaAsLin2 is the next generation of MaAsLin (Microbiome Multivariable Association with Linear Models).

MaAsLin2 is comprehensive R package for efficiently determining multivariable association between clinical metadata and microbial meta-omics features. MaAsLin2 relies on general linear models to accommodate most modern epidemiological study designs, including cross-sectional and longitudinal, along with a variety of filtering, normalization, and transform methods.

If you use the MaAsLin2 software, please cite our manuscript:

Mallick H, Rahnavard A, McIver LJ, Ma S, Zhang Y, Nguyen LH, Tickle TL, Weingart G, Ren B, Schwager EH, Chatterjee S, Thompson KN, Wilkinson JE, Subramanian A, Lu Y, Waldron L, Paulson JN, Franzosa EA, Bravo HC, Huttenhower C (2021). Multivariable Association Discovery in Population-scale Meta-omics Studies. PLoS Computational Biology, 17(11):e1009442.

Check out the MaAsLin 2 tutorial for an overview of analysis options.

If you have questions, please direct it to :
MaAsLin2 Forum
Google Groups (Read only)


Description

MaAsLin2 finds associations between microbiome multi-omics features and complex metadata in population-scale epidemiological studies. The software includes multiple analysis methods (with support for multiple covariates and repeated measures), filtering, normalization, and transform options to customize analysis for your specific study.

Requirements

MaAsLin2 is an R package that can be run on the command line or as an R function.

Installation

MaAsLin2 can be run from the command line or as an R function. If only running from the command line, you do not need to install the MaAsLin2 package but you will need to install the MaAsLin2 dependencies.

From command line

  1. Download the source: MaAsLin2.tar.gz
  2. Decompress the download:
    • $ tar xzvf maaslin2.tar.gz
  3. Install the Bioconductor dependencies edgeR and metagenomeSeq.
  4. Install the CRAN dependencies:
    • $ R -q -e "install.packages(c('lmerTest','pbapply','car','dplyr','vegan','chemometrics','ggplot2','pheatmap','hash','logging','data.table','glmmTMB','MASS','cplm','pscl'), repos='http://cran.r-project.org')"
  5. Install the MaAsLin2 package (only r,equired if running as an R function):
    • $ R CMD INSTALL maaslin2

From R

Install Bioconductor and then install Maaslin2

if(!requireNamespace("BiocManager", quietly = TRUE))
    install.packages("BiocManager")
BiocManager::install("Maaslin2")

How to Run

MaAsLin2 can be run from the command line or as an R function. Both methods require the same arguments, have the same options, and use the same default settings.

Input Files

MaAsLin2 requires two input files.

  1. Data (or features) file
    • This file is tab-delimited.
    • Formatted with features as columns and samples as rows.
    • The transpose of this format is also okay.
    • Possible features in this file include taxonomy or genes.
  2. Metadata file
    • This file is tab-delimited.
    • Formatted with features as columns and samples as rows.
    • The transpose of this format is also okay.
    • Possible metadata in this file include gender or age.

The data file can contain samples not included in the metadata file (along with the reverse case). For both cases, those samples not included in both files will be removed from the analysis. Also the samples do not need to be in the same order in the two files.

NOTE: If running MaAsLin2 as a function, the data and metadata inputs can be of type data.frame instead of a path to a file.

Output Files

MaAsLin2 generates two types of output files: data and visualization.

  1. Data output files
    • all_results.tsv
      • This includes the same data as the data.frame returned.
      • This file contains all results ordered by increasing q-value.
      • The first columns are the metadata and feature names.
      • The next two columns are the value and coefficient from the model.
      • The next column is the standard deviation from the model.
      • The N column is the total number of data points.
      • The N.not.zero column is the total of non-zero data points.
      • The pvalue from the calculation is the second to last column.
      • The qvalue is computed with p.adjust with the correction method.
    • significant_results.tsv
      • This file is a subset of the results in the first file.
      • It only includes associations with q-values <= to the threshold.
    • ``features```
      • This folder includes the filtered, normalized, and transformed versions of the input feature table.
      • These steps are performed sequentially in the above order.
      • If an option is set such that a step does not change the data, the resulting table will still be output.
    • models.rds
      • This file contains a list with every model fit object.
      • It will only be generated if save_models is set to TRUE.
    • residuals.rds
      • This file contains a data frame with residuals for each feature.
    • fitted.rds
      • This file contains a data frame with fitted values for each feature.
    • ranef.rds
      • This file contains a data frame with extracted random effects for each feature (when random effects are specified).
    • maaslin2.log
      • This file contains all log information for the run.
      • It includes all settings, warnings, errors, and steps run.
  2. Visualization output files
    • heatmap.pdf
      • This file contains a heatmap of the significant associations.
    • [a-z/0-9]+.pdf
      • A plot is generated for each significant association.
      • Scatter plots are used for continuous metadata.
      • Box plots are for categorical data.
      • Data points plotted are after filtering but prior to normalization and transform.

Run a Demo

Example input files can be found in the inst/extdata folder of the MaAsLin2 source. The files provided were generated from the HMP2 data which can be downloaded from https://ibdmdb.org/ .

HMP2_taxonomy.tsv: is a tab-demilited file with species as columns and samples as rows. It is a subset of the taxonomy file so it just includes the species abundances for all samples.

HMP2_metadata.tsv: is a tab-delimited file with samples as rows and metadata as columns. It is a subset of the metadata file so that it just includes some of the fields.

Command line

$ Maaslin2.R --fixed_effects="diagnosis,dysbiosisnonIBD,dysbiosisUC,dysbiosisCD,antibiotics,age" --random_effects="site,subject" --standardize=FALSE inst/extdata/HMP2_taxonomy.tsv inst/extdata/HMP2_metadata.tsv demo_output

  • Make sure to provide the full path to the MaAsLin2 executable (ie ./R/Maaslin2.R).
  • In the demo command:
    • HMP2_taxonomy.tsv is the path to your data (or features) file
    • HMP2_metadata.tsv is the path to your metadata file
    • demo_output is the path to the folder to write the output

In R

library(Maaslin2)
input_data <- system.file(
    'extdata','HMP2_taxonomy.tsv', package="Maaslin2")
input_metadata <-system.file(
    'extdata','HMP2_metadata.tsv', package="Maaslin2")
fit_data <- Maaslin2(
    input_data, input_metadata, 'demo_output',
    fixed_effects = c('diagnosis', 'dysbiosisnonIBD','dysbiosisUC','dysbiosisCD', 'antibiotics', 'age'),
    random_effects = c('site', 'subject'),
    reference = "diagnosis,nonIBD",
    standardize = FALSE)
## [1] "Creating output folder"
## [1] "Creating output feature tables folder"
## [1] "Creating output fits folder"
## [1] "Creating output figures folder"
## 2023-06-08 20:02:18.775656 INFO::Writing function arguments to log file
## 2023-06-08 20:02:18.81247 INFO::Verifying options selected are valid
## 2023-06-08 20:02:18.849331 INFO::Determining format of input files
## 2023-06-08 20:02:18.854855 INFO::Input format is data samples as rows and metadata samples as rows
## 2023-06-08 20:02:18.867647 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2023-06-08 20:02:18.869836 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2023-06-08 20:02:18.872988 INFO::Filter data based on min abundance and min prevalence
## 2023-06-08 20:02:18.884164 INFO::Total samples in data: 1595
## 2023-06-08 20:02:18.886056 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2023-06-08 20:02:18.895844 INFO::Total filtered features: 0
## 2023-06-08 20:02:18.897775 INFO::Filtered feature names from abundance and prevalence filtering:
## 2023-06-08 20:02:18.907021 INFO::Total filtered features with variance filtering: 0
## 2023-06-08 20:02:18.909321 INFO::Filtered feature names from variance filtering:
## 2023-06-08 20:02:18.910922 INFO::Running selected normalization method: TSS
## 2023-06-08 20:02:20.01818 INFO::Bypass z-score application to metadata
## 2023-06-08 20:02:20.019996 INFO::Running selected transform method: LOG
## 2023-06-08 20:02:20.041863 INFO::Running selected analysis method: LM
## 2023-06-08 20:02:20.593023 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:21.00235 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2023-06-08 20:02:21.190793 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2023-06-08 20:02:21.355085 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:21.494578 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2023-06-08 20:02:21.643409 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:21.794925 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2023-06-08 20:02:21.926452 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2023-06-08 20:02:22.068573 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2023-06-08 20:02:22.206333 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2023-06-08 20:02:22.375904 INFO::Fitting model to feature number 11, Bacteroides.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:22.512592 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:22.653572 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:22.862046 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2023-06-08 20:02:23.001334 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:23.131772 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2023-06-08 20:02:23.317513 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2023-06-08 20:02:23.471605 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:23.617609 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:23.791829 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:23.956968 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:24.09694 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2023-06-08 20:02:24.232887 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2023-06-08 20:02:24.377211 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2023-06-08 20:02:24.519177 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2023-06-08 20:02:24.667173 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:24.833155 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2023-06-08 20:02:24.970524 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2023-06-08 20:02:25.131248 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:25.293915 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2023-06-08 20:02:25.460962 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2023-06-08 20:02:25.631477 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:25.826144 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:25.979455 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:26.146321 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:26.29385 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2023-06-08 20:02:26.457639 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2023-06-08 20:02:26.602979 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2023-06-08 20:02:26.746454 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:26.892952 INFO::Fitting model to feature number 40, Clostridium.citroniae
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:27.034862 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:27.167213 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:27.313397 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2023-06-08 20:02:27.464923 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2023-06-08 20:02:27.604948 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:27.75522 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:27.894334 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2023-06-08 20:02:28.046459 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2023-06-08 20:02:28.184939 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:28.331607 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:28.464445 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:28.604591 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:28.744508 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:28.892703 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2023-06-08 20:02:29.032969 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2023-06-08 20:02:29.176837 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2023-06-08 20:02:29.343981 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:29.489114 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:29.635331 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:29.785213 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:29.937468 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2023-06-08 20:02:30.094714 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:30.228397 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:30.362045 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2023-06-08 20:02:30.50365 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2023-06-08 20:02:30.652267 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2023-06-08 20:02:30.822411 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:30.979255 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2023-06-08 20:02:31.121116 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2023-06-08 20:02:31.262986 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:31.39731 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2023-06-08 20:02:31.535517 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2023-06-08 20:02:31.675582 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:31.836381 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2023-06-08 20:02:31.990005 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:32.130415 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:32.264969 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2023-06-08 20:02:32.407373 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2023-06-08 20:02:32.559126 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2023-06-08 20:02:32.716458 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2023-06-08 20:02:32.866084 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2023-06-08 20:02:33.013741 INFO::Fitting model to feature number 82, Escherichia.coli
## 2023-06-08 20:02:33.166239 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2023-06-08 20:02:33.299194 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:33.434883 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2023-06-08 20:02:33.582244 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2023-06-08 20:02:33.736784 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2023-06-08 20:02:33.998264 INFO::Counting total values for each feature
## 2023-06-08 20:02:34.050483 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2023-06-08 20:02:34.351686 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2023-06-08 20:02:34.64636 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2023-06-08 20:02:35.043025 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2023-06-08 20:02:35.092121 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2023-06-08 20:02:35.123683 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2023-06-08 20:02:35.131346 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2023-06-08 20:02:35.145947 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
## 2023-06-08 20:02:35.152731 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2023-06-08 20:02:35.371872 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2023-06-08 20:02:35.383158 INFO::Plotting associations from most to least significant, grouped by metadata
## 2023-06-08 20:02:35.385034 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2023-06-08 20:02:35.387681 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2023-06-08 20:02:35.766434 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2023-06-08 20:02:36.029319 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2023-06-08 20:02:36.331109 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2023-06-08 20:02:36.595516 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2023-06-08 20:02:36.874726 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2023-06-08 20:02:37.12284 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2023-06-08 20:02:37.379161 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2023-06-08 20:02:37.626391 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2023-06-08 20:02:37.902607 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2023-06-08 20:02:38.159427 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2023-06-08 20:02:38.426008 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2023-06-08 20:02:38.949213 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2023-06-08 20:02:39.187625 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2023-06-08 20:02:39.434182 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2023-06-08 20:02:39.671427 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2023-06-08 20:02:39.924788 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2023-06-08 20:02:40.181437 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2023-06-08 20:02:40.432269 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2023-06-08 20:02:40.695772 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2023-06-08 20:02:40.996092 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2023-06-08 20:02:41.265332 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2023-06-08 20:02:41.510338 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2023-06-08 20:02:41.769224 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2023-06-08 20:02:42.029657 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2023-06-08 20:02:42.274468 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2023-06-08 20:02:42.532638 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2023-06-08 20:02:42.78179 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2023-06-08 20:02:43.039745 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2023-06-08 20:02:43.285559 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2023-06-08 20:02:43.549432 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2023-06-08 20:02:43.830785 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2023-06-08 20:02:44.094952 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2023-06-08 20:02:44.376314 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2023-06-08 20:02:44.627348 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2023-06-08 20:02:44.892168 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2023-06-08 20:02:45.138083 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2023-06-08 20:02:45.396889 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2023-06-08 20:02:45.661983 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2023-06-08 20:02:46.102511 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2023-06-08 20:02:46.372574 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2023-06-08 20:02:46.63139 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2023-06-08 20:02:46.89239 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2023-06-08 20:02:47.161146 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2023-06-08 20:02:47.424993 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2023-06-08 20:02:47.706512 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2023-06-08 20:02:48.010685 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2023-06-08 20:02:48.26217 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2023-06-08 20:02:48.52429 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2023-06-08 20:02:48.868574 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2023-06-08 20:02:49.137687 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2023-06-08 20:02:49.403916 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2023-06-08 20:02:49.709401 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2023-06-08 20:02:50.012471 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2023-06-08 20:02:50.288995 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2023-06-08 20:02:50.591408 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2023-06-08 20:02:50.842529 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2023-06-08 20:02:51.12245 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2023-06-08 20:02:51.366567 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2023-06-08 20:02:51.633634 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2023-06-08 20:02:51.912305 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2023-06-08 20:02:52.185199 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2023-06-08 20:02:52.441195 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2023-06-08 20:02:55.995603 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2023-06-08 20:02:55.998703 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2023-06-08 20:02:56.257392 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2023-06-08 20:02:56.510585 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2023-06-08 20:02:56.797894 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2023-06-08 20:02:57.076053 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2023-06-08 20:02:57.347663 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2023-06-08 20:02:57.629098 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2023-06-08 20:02:57.900216 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2023-06-08 20:02:58.192059 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2023-06-08 20:02:58.486795 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2023-06-08 20:02:58.757082 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2023-06-08 20:02:59.058482 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2023-06-08 20:02:59.362486 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2023-06-08 20:02:59.632568 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2023-06-08 20:02:59.91179 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2023-06-08 20:03:00.19591 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2023-06-08 20:03:00.474718 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2023-06-08 20:03:00.814158 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2023-06-08 20:03:01.193738 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2023-06-08 20:03:01.455728 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2023-06-08 20:03:01.748121 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2023-06-08 20:03:02.012524 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2023-06-08 20:03:02.294363 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2023-06-08 20:03:02.556231 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2023-06-08 20:03:02.860147 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2023-06-08 20:03:03.17561 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2023-06-08 20:03:03.438156 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2023-06-08 20:03:03.712378 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2023-06-08 20:03:03.969078 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2023-06-08 20:03:04.252635 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2023-06-08 20:03:04.525072 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2023-06-08 20:03:04.791095 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2023-06-08 20:03:05.094625 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2023-06-08 20:03:05.358043 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2023-06-08 20:03:05.644603 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2023-06-08 20:03:05.975599 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2023-06-08 20:03:06.24947 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2023-06-08 20:03:06.537739 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2023-06-08 20:03:06.845885 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2023-06-08 20:03:07.133772 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2023-06-08 20:03:07.411818 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2023-06-08 20:03:07.672462 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2023-06-08 20:03:08.022256 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2023-06-08 20:03:08.344139 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2023-06-08 20:03:12.091736 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2023-06-08 20:03:12.094944 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2023-06-08 20:03:12.363182 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2023-06-08 20:03:12.634745 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2023-06-08 20:03:12.960662 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2023-06-08 20:03:13.244336 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2023-06-08 20:03:13.543292 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2023-06-08 20:03:13.952705 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2023-06-08 20:03:14.225453 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2023-06-08 20:03:14.542089 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2023-06-08 20:03:14.837342 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2023-06-08 20:03:15.129064 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2023-06-08 20:03:15.434814 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2023-06-08 20:03:15.735217 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2023-06-08 20:03:16.031398 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2023-06-08 20:03:16.330554 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2023-06-08 20:03:16.609433 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2023-06-08 20:03:16.973187 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2023-06-08 20:03:17.259038 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2023-06-08 20:03:17.54577 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2023-06-08 20:03:17.862737 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2023-06-08 20:03:18.12464 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2023-06-08 20:03:18.425173 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2023-06-08 20:03:18.730978 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2023-06-08 20:03:19.005022 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2023-06-08 20:03:19.311082 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2023-06-08 20:03:19.611185 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2023-06-08 20:03:19.901628 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2023-06-08 20:03:20.209393 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2023-06-08 20:03:20.524101 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2023-06-08 20:03:24.636627 INFO::Plotting data for metadata number 4, antibiotics
## 2023-06-08 20:03:24.639764 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2023-06-08 20:03:24.882355 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2023-06-08 20:03:25.159732 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2023-06-08 20:03:25.441393 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2023-06-08 20:03:25.720418 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2023-06-08 20:03:26.081909 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2023-06-08 20:03:26.354854 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2023-06-08 20:03:26.662608 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2023-06-08 20:03:26.920736 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2023-06-08 20:03:27.209974 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2023-06-08 20:03:27.520639 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2023-06-08 20:03:27.855095 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2023-06-08 20:03:28.168619 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2023-06-08 20:03:28.448064 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2023-06-08 20:03:28.737335 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2023-06-08 20:03:29.007281 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2023-06-08 20:03:29.287992 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2023-06-08 20:03:29.694494 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2023-06-08 20:03:29.977583 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2023-06-08 20:03:30.261453 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2023-06-08 20:03:30.54537 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2023-06-08 20:03:30.861465 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2023-06-08 20:03:31.167927 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2023-06-08 20:03:31.439682 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2023-06-08 20:03:31.725163 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2023-06-08 20:03:32.018432 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2023-06-08 20:03:32.277032 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2023-06-08 20:03:32.569587 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2023-06-08 20:03:32.891784 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2023-06-08 20:03:33.155439 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2023-06-08 20:03:33.748989 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2023-06-08 20:03:33.989964 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2023-06-08 20:03:34.23675 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2023-06-08 20:03:34.475801 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2023-06-08 20:03:34.727585 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2023-06-08 20:03:35.018129 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2023-06-08 20:03:35.28763 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2023-06-08 20:03:35.578598 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2023-06-08 20:03:35.830066 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2023-06-08 20:03:36.07373 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2023-06-08 20:03:39.44243 INFO::Plotting data for metadata number 5, age
## 2023-06-08 20:03:39.445628 INFO::Creating scatter plot for continuous data, age vs Haemophilus.parainfluenzae
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:39.759613 INFO::Creating scatter plot for continuous data, age vs Bifidobacterium.pseudocatenulatum
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:40.120613 INFO::Creating scatter plot for continuous data, age vs Faecalibacterium.prausnitzii
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:40.376391 INFO::Creating scatter plot for continuous data, age vs Clostridium.clostridioforme
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:40.655328 INFO::Creating scatter plot for continuous data, age vs Veillonella.parvula
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:40.970415 INFO::Creating scatter plot for continuous data, age vs Subdoligranulum.unclassified
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:41.255648 INFO::Creating scatter plot for continuous data, age vs Clostridium.symbiosum
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:41.537932 INFO::Creating scatter plot for continuous data, age vs Ruminococcus.gnavus
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:41.817091 INFO::Creating scatter plot for continuous data, age vs Dialister.invisus
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:42.117339 INFO::Creating scatter plot for continuous data, age vs Veillonella.dispar
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:42.406799 INFO::Creating scatter plot for continuous data, age vs Veillonella.unclassified
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:42.674571 INFO::Creating scatter plot for continuous data, age vs Bacteroides.thetaiotaomicron
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:42.971463 INFO::Creating scatter plot for continuous data, age vs Ruminococcus.bromii
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:43.257949 INFO::Creating scatter plot for continuous data, age vs Bacteroides.intestinalis
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:43.586877 INFO::Creating scatter plot for continuous data, age vs Eubacterium.siraeum
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:43.931828 INFO::Creating scatter plot for continuous data, age vs Prevotella.copri
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:44.226067 INFO::Creating scatter plot for continuous data, age vs Alistipes.unclassified
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:44.488229 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 (`geom_point()`).
## 2023-06-08 20:03:44.772483 INFO::Creating scatter plot for continuous data, age vs Bifidobacterium.longum
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:45.03241 INFO::Creating scatter plot for continuous data, age vs Akkermansia.muciniphila
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:45.302182 INFO::Creating scatter plot for continuous data, age vs Collinsella.aerofaciens
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:45.590438 INFO::Creating scatter plot for continuous data, age vs Parabacteroides.distasonis
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## `geom_smooth()` using formula = 'y ~ x'
## Warning: Removed 19 rows containing missing values (`geom_point()`).
## 2023-06-08 20:03:49.395148 INFO::Plotting data for metadata number 6, diagnosis
## 2023-06-08 20:03:49.398241 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2023-06-08 20:03:49.621751 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2023-06-08 20:03:49.955772 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2023-06-08 20:03:50.232914 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2023-06-08 20:03:50.500704 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2023-06-08 20:03:50.834483 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2023-06-08 20:03:51.131236 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2023-06-08 20:03:51.385636 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2023-06-08 20:03:51.67452 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2023-06-08 20:03:51.946306 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2023-06-08 20:03:52.201483 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2023-06-08 20:03:52.479138 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2023-06-08 20:03:52.775176 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2023-06-08 20:03:53.087389 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2023-06-08 20:03:53.35714 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2023-06-08 20:03:53.650225 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2023-06-08 20:03:53.925886 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2023-06-08 20:03:54.214431 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2023-06-08 20:03:54.503104 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2023-06-08 20:03:54.79411 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2023-06-08 20:03:55.086755 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2023-06-08 20:03:55.36515 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2023-06-08 20:03:55.667373 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2023-06-08 20:03:56.004182 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2023-06-08 20:03:56.280745 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2023-06-08 20:03:56.562397 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2023-06-08 20:03:56.869898 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2023-06-08 20:03:57.132058 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2023-06-08 20:03:57.408104 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2023-06-08 20:03:57.687052 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2023-06-08 20:03:57.955069 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2023-06-08 20:03:58.228023 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2023-06-08 20:03:58.501291 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2023-06-08 20:03:58.779285 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2023-06-08 20:03:59.086714 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2023-06-08 20:03:59.363777 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2023-06-08 20:03:59.616038 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2023-06-08 20:03:59.917082 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2023-06-08 20:04:00.196652 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2023-06-08 20:04:00.462242 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2023-06-08 20:04:00.719293 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2023-06-08 20:04:01.003017 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2023-06-08 20:04:01.3008 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 4.3.0 RC (2023-04-13 r84269 ucrt)
## Platform: x86_64-w64-mingw32/x64 (64-bit)
## Running under: Windows Server 2022 x64 (build 20348)
## 
## Matrix products: default
## 
## 
## locale:
## [1] LC_COLLATE=C                          
## [2] LC_CTYPE=English_United States.utf8   
## [3] LC_MONETARY=English_United States.utf8
## [4] LC_NUMERIC=C                          
## [5] LC_TIME=English_United States.utf8    
## 
## time zone: America/New_York
## tzcode source: internal
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
## 
## other attached packages:
## [1] Maaslin2_1.14.1
## 
## loaded via a namespace (and not attached):
##  [1] gtable_0.3.3        biglm_0.9-2.1       xfun_0.39          
##  [4] bslib_0.4.2         ggplot2_3.4.2       lattice_0.21-8     
##  [7] numDeriv_2016.8-1.1 vctrs_0.6.2         tools_4.3.0        
## [10] generics_0.1.3      parallel_4.3.0      getopt_1.20.3      
## [13] tibble_3.2.1        fansi_1.0.4         DEoptimR_1.0-13    
## [16] cluster_2.1.4       pkgconfig_2.0.3     logging_0.10-108   
## [19] pheatmap_1.0.12     Matrix_1.5-4.1      data.table_1.14.8  
## [22] RColorBrewer_1.1-3  lifecycle_1.0.3     farver_2.1.1       
## [25] compiler_4.3.0      munsell_0.5.0       lmerTest_3.1-3     
## [28] permute_0.9-7       htmltools_0.5.5     sass_0.4.6         
## [31] hash_2.2.6.2        yaml_2.3.7          pillar_1.9.0       
## [34] nloptr_2.0.3        crayon_1.5.2        jquerylib_0.1.4    
## [37] MASS_7.3-60         cachem_1.0.8        vegan_2.6-4        
## [40] boot_1.3-28.1       nlme_3.1-162        robustbase_0.95-1  
## [43] tidyselect_1.2.0    digest_0.6.31       mvtnorm_1.2-2      
## [46] dplyr_1.1.2         labeling_0.4.2      splines_4.3.0      
## [49] pcaPP_2.0-3         fastmap_1.1.1       grid_4.3.0         
## [52] colorspace_2.1-0    cli_3.6.1           magrittr_2.0.3     
## [55] utf8_1.2.3          withr_2.5.0         scales_1.2.1       
## [58] rmarkdown_2.22      lme4_1.1-33         pbapply_1.7-0      
## [61] evaluate_0.21       knitr_1.43          mgcv_1.8-42        
## [64] rlang_1.1.1         Rcpp_1.0.10         glue_1.6.2         
## [67] optparse_1.7.3      DBI_1.1.3           minqa_1.2.5        
## [70] jsonlite_1.8.5      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

  1. Question: When I run from the command line I see the error Maaslin2.R: command not found. How do I fix this?
    • Answer: Provide the full path to the executable when running Maaslin2.R.
  2. Question: When I run as a function I see the error Error in library(Maaslin2): there is no package called 'Maaslin2'. How do I fix this?
    • Answer: Install the R package and then try loading the library again.
  3. Question: When I try to install the R package I see errors about dependencies not being installed. Why is this?
    • Answer: Installing the R package will not automatically install the packages MaAsLin2 requires. Please install the dependencies and then install the MaAsLin2 R package.