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"
## 2024-05-01 00:25:05.352238 INFO::Writing function arguments to log file
## 2024-05-01 00:25:05.384316 INFO::Verifying options selected are valid
## 2024-05-01 00:25:05.418949 INFO::Determining format of input files
## 2024-05-01 00:25:05.420068 INFO::Input format is data samples as rows and metadata samples as rows
## 2024-05-01 00:25:05.424108 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2024-05-01 00:25:05.42516 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2024-05-01 00:25:05.427305 INFO::Filter data based on min abundance and min prevalence
## 2024-05-01 00:25:05.427938 INFO::Total samples in data: 1595
## 2024-05-01 00:25:05.428543 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2024-05-01 00:25:05.433249 INFO::Total filtered features: 0
## 2024-05-01 00:25:05.434022 INFO::Filtered feature names from abundance and prevalence filtering:
## 2024-05-01 00:25:05.442754 INFO::Total filtered features with variance filtering: 0
## 2024-05-01 00:25:05.44347 INFO::Filtered feature names from variance filtering:
## 2024-05-01 00:25:05.444083 INFO::Running selected normalization method: TSS
## 2024-05-01 00:25:06.521398 INFO::Bypass z-score application to metadata
## 2024-05-01 00:25:06.522326 INFO::Running selected transform method: LOG
## 2024-05-01 00:25:06.54107 INFO::Running selected analysis method: LM
## 2024-05-01 00:25:06.827647 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:07.14232 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2024-05-01 00:25:07.295986 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2024-05-01 00:25:07.494962 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:07.626441 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2024-05-01 00:25:07.78192 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:07.925205 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2024-05-01 00:25:08.059849 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2024-05-01 00:25:08.210983 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2024-05-01 00:25:08.355565 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2024-05-01 00:25:08.507679 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
## 2024-05-01 00:25:08.619657 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
## 2024-05-01 00:25:08.765019 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:08.898777 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:09.042726 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2024-05-01 00:25:09.184019 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:09.321088 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2024-05-01 00:25:09.460268 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2024-05-01 00:25:09.606002 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:09.743998 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:09.886996 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:10.021407 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:10.173694 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2024-05-01 00:25:10.343547 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2024-05-01 00:25:10.492695 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2024-05-01 00:25:10.636582 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2024-05-01 00:25:10.777559 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:10.917724 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2024-05-01 00:25:11.056262 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2024-05-01 00:25:11.201123 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:11.343596 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2024-05-01 00:25:11.488629 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2024-05-01 00:25:11.624792 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:11.767161 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:11.908717 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:12.045906 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:12.187304 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2024-05-01 00:25:12.332986 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2024-05-01 00:25:12.46331 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2024-05-01 00:25:12.608966 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:12.756423 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
## 2024-05-01 00:25:12.868642 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
## 2024-05-01 00:25:13.011914 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:13.151757 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:13.296615 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2024-05-01 00:25:13.463285 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2024-05-01 00:25:13.610116 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:13.756318 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:13.903338 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2024-05-01 00:25:14.054923 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2024-05-01 00:25:14.21 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:14.359445 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:14.703281 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:14.855913 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:14.995363 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:15.14557 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2024-05-01 00:25:15.298081 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2024-05-01 00:25:15.43158 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2024-05-01 00:25:15.569798 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:15.711088 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:15.847078 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:15.982542 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:16.126977 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2024-05-01 00:25:16.252654 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:16.384761 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:16.523431 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2024-05-01 00:25:16.665776 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2024-05-01 00:25:16.8011 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2024-05-01 00:25:16.941711 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:17.066797 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2024-05-01 00:25:17.19678 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2024-05-01 00:25:17.339645 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:17.476674 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2024-05-01 00:25:17.615598 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2024-05-01 00:25:17.763471 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:17.895376 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2024-05-01 00:25:18.042967 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:18.177854 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:18.312386 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2024-05-01 00:25:18.45024 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2024-05-01 00:25:18.58527 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2024-05-01 00:25:18.717866 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2024-05-01 00:25:18.86382 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2024-05-01 00:25:19.00584 INFO::Fitting model to feature number 82, Escherichia.coli
## 2024-05-01 00:25:19.144632 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2024-05-01 00:25:19.283092 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:19.412446 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2024-05-01 00:25:19.546967 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2024-05-01 00:25:19.69355 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2024-05-01 00:25:19.865148 INFO::Counting total values for each feature
## 2024-05-01 00:25:19.896312 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2024-05-01 00:25:20.039528 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2024-05-01 00:25:20.188473 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2024-05-01 00:25:20.427457 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2024-05-01 00:25:20.49047 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2024-05-01 00:25:20.52599 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2024-05-01 00:25:20.531413 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2024-05-01 00:25:20.537644 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
## 2024-05-01 00:25:20.540439 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2024-05-01 00:25:20.754366 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2024-05-01 00:25:20.759865 INFO::Plotting associations from most to least significant, grouped by metadata
## 2024-05-01 00:25:20.760717 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2024-05-01 00:25:20.762104 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2024-05-01 00:25:21.138362 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2024-05-01 00:25:21.408837 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2024-05-01 00:25:21.656533 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2024-05-01 00:25:21.891093 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2024-05-01 00:25:22.132152 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2024-05-01 00:25:22.363375 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2024-05-01 00:25:22.601185 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2024-05-01 00:25:22.835525 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2024-05-01 00:25:23.078203 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2024-05-01 00:25:23.313001 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2024-05-01 00:25:23.552245 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2024-05-01 00:25:23.786015 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2024-05-01 00:25:24.020144 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2024-05-01 00:25:24.251074 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2024-05-01 00:25:24.501633 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2024-05-01 00:25:24.748387 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2024-05-01 00:25:24.983234 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2024-05-01 00:25:25.273116 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2024-05-01 00:25:25.514316 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2024-05-01 00:25:25.757616 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2024-05-01 00:25:25.99117 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2024-05-01 00:25:26.231782 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2024-05-01 00:25:26.456176 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2024-05-01 00:25:26.700094 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2024-05-01 00:25:26.934618 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2024-05-01 00:25:27.183188 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2024-05-01 00:25:27.42156 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2024-05-01 00:25:27.664604 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2024-05-01 00:25:27.897872 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2024-05-01 00:25:28.135529 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2024-05-01 00:25:28.370407 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2024-05-01 00:25:28.60877 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2024-05-01 00:25:28.853868 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2024-05-01 00:25:29.129327 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2024-05-01 00:25:29.367361 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2024-05-01 00:25:29.605123 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2024-05-01 00:25:29.83885 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2024-05-01 00:25:30.076232 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2024-05-01 00:25:30.304188 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2024-05-01 00:25:30.53851 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2024-05-01 00:25:30.772513 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2024-05-01 00:25:31.01081 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2024-05-01 00:25:31.235892 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2024-05-01 00:25:31.479848 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2024-05-01 00:25:31.713352 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2024-05-01 00:25:31.956121 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2024-05-01 00:25:32.186182 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2024-05-01 00:25:32.42101 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2024-05-01 00:25:32.66744 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2024-05-01 00:25:32.898548 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2024-05-01 00:25:33.160118 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2024-05-01 00:25:33.39972 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2024-05-01 00:25:33.652102 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2024-05-01 00:25:33.87808 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2024-05-01 00:25:34.115366 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2024-05-01 00:25:34.342247 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2024-05-01 00:25:34.57827 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2024-05-01 00:25:34.801294 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2024-05-01 00:25:35.041071 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2024-05-01 00:25:35.267442 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2024-05-01 00:25:35.513133 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2024-05-01 00:25:35.741451 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2024-05-01 00:25:38.719675 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2024-05-01 00:25:38.721473 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2024-05-01 00:25:38.934173 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2024-05-01 00:25:39.24726 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2024-05-01 00:25:39.488294 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2024-05-01 00:25:39.738511 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2024-05-01 00:25:39.980426 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2024-05-01 00:25:40.235174 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2024-05-01 00:25:40.467588 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2024-05-01 00:25:40.719015 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2024-05-01 00:25:40.964211 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2024-05-01 00:25:41.202729 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2024-05-01 00:25:41.460577 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2024-05-01 00:25:41.700551 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2024-05-01 00:25:41.945408 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2024-05-01 00:25:42.181279 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2024-05-01 00:25:42.426025 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2024-05-01 00:25:42.660823 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2024-05-01 00:25:42.903629 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2024-05-01 00:25:43.14273 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2024-05-01 00:25:43.386242 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2024-05-01 00:25:43.623348 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2024-05-01 00:25:43.867411 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2024-05-01 00:25:44.103654 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2024-05-01 00:25:44.350699 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2024-05-01 00:25:44.595359 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2024-05-01 00:25:44.861709 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2024-05-01 00:25:45.106998 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2024-05-01 00:25:45.35493 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2024-05-01 00:25:45.603586 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2024-05-01 00:25:45.846849 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2024-05-01 00:25:46.093901 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2024-05-01 00:25:46.323261 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2024-05-01 00:25:46.576308 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2024-05-01 00:25:46.809042 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2024-05-01 00:25:47.063075 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2024-05-01 00:25:47.294899 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2024-05-01 00:25:47.545028 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2024-05-01 00:25:47.789356 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2024-05-01 00:25:48.020553 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2024-05-01 00:25:48.270747 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2024-05-01 00:25:48.515696 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2024-05-01 00:25:48.820389 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2024-05-01 00:25:49.07249 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2024-05-01 00:25:49.329254 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2024-05-01 00:25:52.328768 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2024-05-01 00:25:52.330676 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2024-05-01 00:25:52.541044 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2024-05-01 00:25:52.797736 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2024-05-01 00:25:53.041867 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2024-05-01 00:25:53.288084 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2024-05-01 00:25:53.542624 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2024-05-01 00:25:53.801308 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2024-05-01 00:25:54.060926 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2024-05-01 00:25:54.31182 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2024-05-01 00:25:54.577067 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2024-05-01 00:25:54.819132 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2024-05-01 00:25:55.067702 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2024-05-01 00:25:55.310968 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2024-05-01 00:25:55.563835 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2024-05-01 00:25:55.812383 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2024-05-01 00:25:56.068692 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2024-05-01 00:25:56.299537 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2024-05-01 00:25:56.548324 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2024-05-01 00:25:56.806837 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2024-05-01 00:25:57.036824 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2024-05-01 00:25:57.285271 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2024-05-01 00:25:57.523339 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2024-05-01 00:25:57.774351 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2024-05-01 00:25:58.011384 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2024-05-01 00:25:58.257499 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2024-05-01 00:25:58.503347 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2024-05-01 00:25:58.748189 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2024-05-01 00:25:59.2589 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2024-05-01 00:25:59.49877 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2024-05-01 00:26:02.360258 INFO::Plotting data for metadata number 4, antibiotics
## 2024-05-01 00:26:02.361969 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2024-05-01 00:26:02.561249 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2024-05-01 00:26:02.777224 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2024-05-01 00:26:03.003278 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2024-05-01 00:26:03.222649 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2024-05-01 00:26:03.442176 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2024-05-01 00:26:03.670414 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2024-05-01 00:26:03.889657 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2024-05-01 00:26:04.105298 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2024-05-01 00:26:04.319682 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2024-05-01 00:26:04.547962 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2024-05-01 00:26:04.757178 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2024-05-01 00:26:04.976467 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2024-05-01 00:26:05.197926 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2024-05-01 00:26:05.418987 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2024-05-01 00:26:05.632159 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2024-05-01 00:26:05.854146 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2024-05-01 00:26:06.070082 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2024-05-01 00:26:06.289304 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2024-05-01 00:26:06.500367 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2024-05-01 00:26:06.723986 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2024-05-01 00:26:06.939773 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2024-05-01 00:26:07.159906 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2024-05-01 00:26:07.38048 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2024-05-01 00:26:07.601673 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2024-05-01 00:26:07.821324 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2024-05-01 00:26:08.041928 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2024-05-01 00:26:08.256494 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2024-05-01 00:26:08.472041 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2024-05-01 00:26:08.694843 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2024-05-01 00:26:08.919049 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2024-05-01 00:26:09.151138 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2024-05-01 00:26:09.397156 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2024-05-01 00:26:09.62214 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2024-05-01 00:26:09.902412 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2024-05-01 00:26:10.119612 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2024-05-01 00:26:10.343953 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2024-05-01 00:26:10.558439 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2024-05-01 00:26:10.782932 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2024-05-01 00:26:11.000398 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2024-05-01 00:26:13.724614 INFO::Plotting data for metadata number 5, age
## 2024-05-01 00:26:13.726423 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()`).
## 2024-05-01 00:26:13.982593 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()`).
## 2024-05-01 00:26:14.274988 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()`).
## 2024-05-01 00:26:14.495768 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()`).
## 2024-05-01 00:26:14.738144 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()`).
## 2024-05-01 00:26:14.953583 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()`).
## 2024-05-01 00:26:15.182104 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()`).
## 2024-05-01 00:26:15.413061 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()`).
## 2024-05-01 00:26:15.642892 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()`).
## 2024-05-01 00:26:15.876105 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()`).
## 2024-05-01 00:26:16.09742 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()`).
## 2024-05-01 00:26:16.322592 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()`).
## 2024-05-01 00:26:16.539893 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()`).
## 2024-05-01 00:26:16.769587 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()`).
## 2024-05-01 00:26:16.995871 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()`).
## 2024-05-01 00:26:17.205683 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()`).
## 2024-05-01 00:26:17.430915 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()`).
## 2024-05-01 00:26:17.654971 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()`).
## 2024-05-01 00:26:17.868184 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()`).
## 2024-05-01 00:26:18.098729 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()`).
## 2024-05-01 00:26:18.330548 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()`).
## 2024-05-01 00:26:18.551458 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()`).
## 2024-05-01 00:26:21.522365 INFO::Plotting data for metadata number 6, diagnosis
## 2024-05-01 00:26:21.524106 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2024-05-01 00:26:21.73062 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2024-05-01 00:26:21.980565 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2024-05-01 00:26:22.22749 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2024-05-01 00:26:22.467743 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2024-05-01 00:26:22.7247 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2024-05-01 00:26:22.981477 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2024-05-01 00:26:23.232677 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2024-05-01 00:26:23.470989 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2024-05-01 00:26:23.723512 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2024-05-01 00:26:23.965479 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2024-05-01 00:26:24.228222 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2024-05-01 00:26:24.492668 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2024-05-01 00:26:24.740498 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2024-05-01 00:26:24.993792 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2024-05-01 00:26:25.232818 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2024-05-01 00:26:25.481932 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2024-05-01 00:26:25.720607 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2024-05-01 00:26:25.976971 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2024-05-01 00:26:26.22084 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2024-05-01 00:26:26.473912 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2024-05-01 00:26:26.729633 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2024-05-01 00:26:26.973338 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2024-05-01 00:26:27.239778 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2024-05-01 00:26:27.475441 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2024-05-01 00:26:27.728581 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2024-05-01 00:26:27.994632 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2024-05-01 00:26:28.245755 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2024-05-01 00:26:28.543788 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2024-05-01 00:26:28.791402 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2024-05-01 00:26:29.045855 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2024-05-01 00:26:29.283963 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2024-05-01 00:26:29.534105 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2024-05-01 00:26:29.782778 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2024-05-01 00:26:30.037336 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2024-05-01 00:26:30.29194 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2024-05-01 00:26:30.533756 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2024-05-01 00:26:30.787819 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2024-05-01 00:26:31.02862 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2024-05-01 00:26:31.277662 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2024-05-01 00:26:31.520266 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2024-05-01 00:26:31.791944 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2024-05-01 00:26:32.0338 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 4.4.0 beta (2024-04-15 r86425)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 22.04.4 LTS
## 
## Matrix products: default
## BLAS:   /home/biocbuild/bbs-3.19-bioc/R/lib/libRblas.so 
## LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.10.0
## 
## locale:
##  [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C              
##  [3] LC_TIME=en_US.UTF-8        LC_COLLATE=en_US.UTF-8    
##  [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: America/New_York
## tzcode source: system (glibc)
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
## 
## other attached packages:
## [1] Maaslin2_1.18.0
## 
## loaded via a namespace (and not attached):
##  [1] gtable_0.3.5        biglm_0.9-2.1       xfun_0.43          
##  [4] bslib_0.7.0         ggplot2_3.5.1       lattice_0.22-6     
##  [7] numDeriv_2016.8-1.1 vctrs_0.6.5         tools_4.4.0        
## [10] generics_0.1.3      parallel_4.4.0      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_1.15.4  
## [22] RColorBrewer_1.1-3  lifecycle_1.0.4     farver_2.1.1       
## [25] compiler_4.4.0      munsell_0.5.1       lmerTest_3.1-3     
## [28] permute_0.9-7       htmltools_0.5.8.1   sass_0.4.9         
## [31] hash_2.2.6.3        yaml_2.3.8          pillar_1.9.0       
## [34] nloptr_2.0.3        crayon_1.5.2        jquerylib_0.1.4    
## [37] MASS_7.3-60.2       cachem_1.0.8        vegan_2.6-4        
## [40] boot_1.3-30         nlme_3.1-164        robustbase_0.99-2  
## [43] tidyselect_1.2.1    digest_0.6.35       mvtnorm_1.2-4      
## [46] dplyr_1.1.4         labeling_0.4.3      splines_4.4.0      
## [49] pcaPP_2.0-4         fastmap_1.1.1       grid_4.4.0         
## [52] colorspace_2.1-0    cli_3.6.2           magrittr_2.0.3     
## [55] utf8_1.2.4          withr_3.0.0         scales_1.3.0       
## [58] rmarkdown_2.26      lme4_1.1-35.3       pbapply_1.7-2      
## [61] evaluate_0.23       knitr_1.46          mgcv_1.9-1         
## [64] rlang_1.1.3         Rcpp_1.0.12         glue_1.7.0         
## [67] optparse_1.7.5      DBI_1.2.2           minqa_1.2.6        
## [70] 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

  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.