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"
## 2024-12-18 17:10:48.547998 INFO::Writing function arguments to log file
## 2024-12-18 17:10:48.578944 INFO::Verifying options selected are valid
## 2024-12-18 17:10:48.615451 INFO::Determining format of input files
## 2024-12-18 17:10:48.617505 INFO::Input format is data samples as rows and metadata samples as rows
## 2024-12-18 17:10:48.623167 INFO::Formula for random effects: expr ~ (1 | site) + (1 | subject)
## 2024-12-18 17:10:48.62515 INFO::Formula for fixed effects: expr ~  diagnosis + dysbiosisnonIBD + dysbiosisUC + dysbiosisCD + antibiotics + age
## 2024-12-18 17:10:48.628296 INFO::Filter data based on min abundance and min prevalence
## 2024-12-18 17:10:48.629249 INFO::Total samples in data: 1595
## 2024-12-18 17:10:48.630241 INFO::Min samples required with min abundance for a feature not to be filtered: 159.500000
## 2024-12-18 17:10:48.636386 INFO::Total filtered features: 0
## 2024-12-18 17:10:48.63808 INFO::Filtered feature names from abundance and prevalence filtering:
## 2024-12-18 17:10:48.648063 INFO::Total filtered features with variance filtering: 0
## 2024-12-18 17:10:48.649154 INFO::Filtered feature names from variance filtering:
## 2024-12-18 17:10:48.649983 INFO::Running selected normalization method: TSS
## 2024-12-18 17:10:49.713125 INFO::Bypass z-score application to metadata
## 2024-12-18 17:10:49.714673 INFO::Running selected transform method: LOG
## 2024-12-18 17:10:49.753699 INFO::Running selected analysis method: LM
## 2024-12-18 17:10:50.061108 INFO::Fitting model to feature number 1, Bifidobacterium.adolescentis
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:50.399985 INFO::Fitting model to feature number 2, Bifidobacterium.bifidum
## 2024-12-18 17:10:50.569875 INFO::Fitting model to feature number 3, Bifidobacterium.longum
## 2024-12-18 17:10:50.778864 INFO::Fitting model to feature number 4, Bifidobacterium.pseudocatenulatum
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:51.089146 INFO::Fitting model to feature number 5, Collinsella.aerofaciens
## 2024-12-18 17:10:51.238611 INFO::Fitting model to feature number 6, Bacteroides.caccae
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:51.398466 INFO::Fitting model to feature number 7, Bacteroides.cellulosilyticus
## 2024-12-18 17:10:51.548779 INFO::Fitting model to feature number 8, Bacteroides.dorei
## 2024-12-18 17:10:51.70692 INFO::Fitting model to feature number 9, Bacteroides.eggerthii
## 2024-12-18 17:10:51.844745 INFO::Fitting model to feature number 10, Bacteroides.faecis
## 2024-12-18 17:10:51.983151 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
## 2024-12-18 17:10:52.086711 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
## 2024-12-18 17:10:52.22306 INFO::Fitting model to feature number 12, Bacteroides.fragilis
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:52.368975 INFO::Fitting model to feature number 13, Bacteroides.intestinalis
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:52.51186 INFO::Fitting model to feature number 14, Bacteroides.massiliensis
## 2024-12-18 17:10:52.667442 INFO::Fitting model to feature number 15, Bacteroides.ovatus
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:52.803428 INFO::Fitting model to feature number 16, Bacteroides.salyersiae
## 2024-12-18 17:10:52.93477 INFO::Fitting model to feature number 17, Bacteroides.stercoris
## 2024-12-18 17:10:53.071875 INFO::Fitting model to feature number 18, Bacteroides.thetaiotaomicron
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:53.208297 INFO::Fitting model to feature number 19, Bacteroides.uniformis
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:53.339088 INFO::Fitting model to feature number 20, Bacteroides.vulgatus
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:53.470555 INFO::Fitting model to feature number 21, Bacteroides.xylanisolvens
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:53.611747 INFO::Fitting model to feature number 22, Bacteroidales.bacterium.ph8
## 2024-12-18 17:10:53.764924 INFO::Fitting model to feature number 23, Barnesiella.intestinihominis
## 2024-12-18 17:10:53.924487 INFO::Fitting model to feature number 24, Coprobacter.fastidiosus
## 2024-12-18 17:10:54.108999 INFO::Fitting model to feature number 25, Odoribacter.splanchnicus
## 2024-12-18 17:10:54.256965 INFO::Fitting model to feature number 26, Parabacteroides.distasonis
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:54.397634 INFO::Fitting model to feature number 27, Parabacteroides.goldsteinii
## 2024-12-18 17:10:54.557982 INFO::Fitting model to feature number 28, Parabacteroides.merdae
## 2024-12-18 17:10:54.709769 INFO::Fitting model to feature number 29, Parabacteroides.unclassified
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:54.868513 INFO::Fitting model to feature number 30, Paraprevotella.clara
## 2024-12-18 17:10:55.032784 INFO::Fitting model to feature number 31, Paraprevotella.unclassified
## 2024-12-18 17:10:55.184031 INFO::Fitting model to feature number 32, Prevotella.copri
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:55.332004 INFO::Fitting model to feature number 33, Alistipes.finegoldii
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:55.49016 INFO::Fitting model to feature number 34, Alistipes.onderdonkii
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:55.664896 INFO::Fitting model to feature number 35, Alistipes.putredinis
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:55.814683 INFO::Fitting model to feature number 36, Alistipes.shahii
## 2024-12-18 17:10:56.005842 INFO::Fitting model to feature number 37, Alistipes.unclassified
## 2024-12-18 17:10:56.152945 INFO::Fitting model to feature number 38, Streptococcus.salivarius
## 2024-12-18 17:10:56.296787 INFO::Fitting model to feature number 39, Clostridium.bolteae
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:56.454589 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
## 2024-12-18 17:10:56.574198 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
## 2024-12-18 17:10:56.719377 INFO::Fitting model to feature number 41, Clostridium.clostridioforme
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:56.868888 INFO::Fitting model to feature number 42, Clostridium.hathewayi
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:57.028755 INFO::Fitting model to feature number 43, Clostridium.leptum
## 2024-12-18 17:10:57.183156 INFO::Fitting model to feature number 44, Clostridium.nexile
## 2024-12-18 17:10:57.32576 INFO::Fitting model to feature number 45, Clostridium.symbiosum
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:57.477313 INFO::Fitting model to feature number 46, Flavonifractor.plautii
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:57.641175 INFO::Fitting model to feature number 47, Eubacterium.eligens
## 2024-12-18 17:10:57.783327 INFO::Fitting model to feature number 48, Eubacterium.hallii
## 2024-12-18 17:10:57.944458 INFO::Fitting model to feature number 49, Eubacterium.rectale
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:58.093858 INFO::Fitting model to feature number 50, Eubacterium.siraeum
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:58.231977 INFO::Fitting model to feature number 51, Eubacterium.sp.3.1.31
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:58.390167 INFO::Fitting model to feature number 52, Eubacterium.ventriosum
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:58.516071 INFO::Fitting model to feature number 53, Ruminococcus.gnavus
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:58.663068 INFO::Fitting model to feature number 54, Ruminococcus.obeum
## 2024-12-18 17:10:58.811342 INFO::Fitting model to feature number 55, Ruminococcus.torques
## 2024-12-18 17:10:58.957173 INFO::Fitting model to feature number 56, Coprococcus.comes
## 2024-12-18 17:10:59.091936 INFO::Fitting model to feature number 57, Dorea.longicatena
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:59.451887 INFO::Fitting model to feature number 58, Lachnospiraceae.bacterium.1.1.57FAA
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:59.616677 INFO::Fitting model to feature number 59, Lachnospiraceae.bacterium.3.1.46FAA
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:59.758077 INFO::Fitting model to feature number 60, Roseburia.hominis
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:10:59.910224 INFO::Fitting model to feature number 61, Roseburia.intestinalis
## 2024-12-18 17:11:00.041835 INFO::Fitting model to feature number 62, Roseburia.inulinivorans
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:00.188373 INFO::Fitting model to feature number 63, Roseburia.unclassified
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:00.336921 INFO::Fitting model to feature number 64, Oscillibacter.unclassified
## 2024-12-18 17:11:00.481317 INFO::Fitting model to feature number 65, Peptostreptococcaceae.noname.unclassified
## 2024-12-18 17:11:00.613929 INFO::Fitting model to feature number 66, Faecalibacterium.prausnitzii
## 2024-12-18 17:11:00.756732 INFO::Fitting model to feature number 67, Ruminococcus.bromii
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:00.911641 INFO::Fitting model to feature number 68, Ruminococcus.callidus
## 2024-12-18 17:11:01.094618 INFO::Fitting model to feature number 69, Ruminococcus.lactaris
## 2024-12-18 17:11:01.251934 INFO::Fitting model to feature number 70, Subdoligranulum.unclassified
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:01.398907 INFO::Fitting model to feature number 71, Coprobacillus.unclassified
## 2024-12-18 17:11:01.53589 INFO::Fitting model to feature number 72, Acidaminococcus.unclassified
## 2024-12-18 17:11:01.683092 INFO::Fitting model to feature number 73, Dialister.invisus
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:01.840774 INFO::Fitting model to feature number 74, Veillonella.atypica
## 2024-12-18 17:11:01.98392 INFO::Fitting model to feature number 75, Veillonella.dispar
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:02.12203 INFO::Fitting model to feature number 76, Veillonella.parvula
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:02.482066 INFO::Fitting model to feature number 77, Veillonella.unclassified
## 2024-12-18 17:11:02.62775 INFO::Fitting model to feature number 78, Burkholderiales.bacterium.1.1.47
## 2024-12-18 17:11:02.799614 INFO::Fitting model to feature number 79, Parasutterella.excrementihominis
## 2024-12-18 17:11:02.963399 INFO::Fitting model to feature number 80, Sutterella.wadsworthensis
## 2024-12-18 17:11:03.125406 INFO::Fitting model to feature number 81, Bilophila.unclassified
## 2024-12-18 17:11:03.294763 INFO::Fitting model to feature number 82, Escherichia.coli
## 2024-12-18 17:11:03.455267 INFO::Fitting model to feature number 83, Escherichia.unclassified
## 2024-12-18 17:11:03.599485 INFO::Fitting model to feature number 84, Klebsiella.pneumoniae
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:03.753383 INFO::Fitting model to feature number 85, Haemophilus.parainfluenzae
## boundary (singular) fit: see help('isSingular')
## 2024-12-18 17:11:03.906401 INFO::Fitting model to feature number 86, Akkermansia.muciniphila
## 2024-12-18 17:11:04.05336 INFO::Fitting model to feature number 87, C2likevirus.unclassified
## 2024-12-18 17:11:04.251829 INFO::Counting total values for each feature
## 2024-12-18 17:11:04.29984 INFO::Writing filtered data to file demo_output/features/filtered_data.tsv
## 2024-12-18 17:11:04.419947 INFO::Writing filtered, normalized data to file demo_output/features/filtered_data_norm.tsv
## 2024-12-18 17:11:04.566024 INFO::Writing filtered, normalized, transformed data to file demo_output/features/filtered_data_norm_transformed.tsv
## 2024-12-18 17:11:04.786937 INFO::Writing residuals to file demo_output/fits/residuals.rds
## 2024-12-18 17:11:04.844512 INFO::Writing fitted values to file demo_output/fits/fitted.rds
## 2024-12-18 17:11:04.87253 INFO::Writing extracted random effects to file demo_output/fits/ranef.rds
## 2024-12-18 17:11:04.879303 INFO::Writing all results to file (ordered by increasing q-values): demo_output/all_results.tsv
## 2024-12-18 17:11:04.886345 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
## 2024-12-18 17:11:04.890078 INFO::Writing heatmap of significant results to file: demo_output/heatmap.pdf
## 2024-12-18 17:11:05.045582 INFO::Writing association plots (one for each significant association) to output folder: demo_output
## 2024-12-18 17:11:05.052522 INFO::Plotting associations from most to least significant, grouped by metadata
## 2024-12-18 17:11:05.053628 INFO::Plotting data for metadata number 1, dysbiosisCD
## 2024-12-18 17:11:05.055349 INFO::Creating boxplot for categorical data, dysbiosisCD vs Faecalibacterium.prausnitzii
## 2024-12-18 17:11:05.426898 INFO::Creating boxplot for categorical data, dysbiosisCD vs Subdoligranulum.unclassified
## 2024-12-18 17:11:05.739354 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.uniformis
## 2024-12-18 17:11:06.002931 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.rectale
## 2024-12-18 17:11:06.259006 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.vulgatus
## 2024-12-18 17:11:06.497736 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.ovatus
## 2024-12-18 17:11:06.745681 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.obeum
## 2024-12-18 17:11:06.971387 INFO::Creating boxplot for categorical data, dysbiosisCD vs Oscillibacter.unclassified
## 2024-12-18 17:11:07.22276 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.inulinivorans
## 2024-12-18 17:11:07.484376 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.hominis
## 2024-12-18 17:11:07.744914 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.thetaiotaomicron
## 2024-12-18 17:11:07.997113 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.putredinis
## 2024-12-18 17:11:08.285498 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.distasonis
## 2024-12-18 17:11:08.582428 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.dorei
## 2024-12-18 17:11:08.858955 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.shahii
## 2024-12-18 17:11:09.102362 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.xylanisolvens
## 2024-12-18 17:11:09.368406 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.leptum
## 2024-12-18 17:11:09.630003 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dorea.longicatena
## 2024-12-18 17:11:09.892915 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.caccae
## 2024-12-18 17:11:10.153249 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2024-12-18 17:11:10.413821 INFO::Creating boxplot for categorical data, dysbiosisCD vs Escherichia.coli
## 2024-12-18 17:11:10.667414 INFO::Creating boxplot for categorical data, dysbiosisCD vs Klebsiella.pneumoniae
## 2024-12-18 17:11:10.932676 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bilophila.unclassified
## 2024-12-18 17:11:11.168676 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.finegoldii
## 2024-12-18 17:11:11.421662 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.eligens
## 2024-12-18 17:11:11.672721 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.stercoris
## 2024-12-18 17:11:11.923109 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprococcus.comes
## 2024-12-18 17:11:12.189478 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.fragilis
## 2024-12-18 17:11:12.429835 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.finegoldii
## 2024-12-18 17:11:12.695255 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.hallii
## 2024-12-18 17:11:12.935034 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.faecis
## 2024-12-18 17:11:13.179522 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.clara
## 2024-12-18 17:11:13.423992 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.siraeum
## 2024-12-18 17:11:13.751161 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.merdae
## 2024-12-18 17:11:13.99504 INFO::Creating boxplot for categorical data, dysbiosisCD vs Paraprevotella.unclassified
## 2024-12-18 17:11:14.245377 INFO::Creating boxplot for categorical data, dysbiosisCD vs Collinsella.aerofaciens
## 2024-12-18 17:11:14.489063 INFO::Creating boxplot for categorical data, dysbiosisCD vs Odoribacter.splanchnicus
## 2024-12-18 17:11:14.762398 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.clostridioforme
## 2024-12-18 17:11:15.020384 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.eggerthii
## 2024-12-18 17:11:15.278915 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.onderdonkii
## 2024-12-18 17:11:15.53125 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.ventriosum
## 2024-12-18 17:11:15.798694 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.lactaris
## 2024-12-18 17:11:16.055361 INFO::Creating boxplot for categorical data, dysbiosisCD vs Burkholderiales.bacterium.1.1.47
## 2024-12-18 17:11:16.304286 INFO::Creating boxplot for categorical data, dysbiosisCD vs Dialister.invisus
## 2024-12-18 17:11:16.559692 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.bromii
## 2024-12-18 17:11:16.814291 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parasutterella.excrementihominis
## 2024-12-18 17:11:17.08467 INFO::Creating boxplot for categorical data, dysbiosisCD vs Alistipes.unclassified
## 2024-12-18 17:11:17.340935 INFO::Creating boxplot for categorical data, dysbiosisCD vs Ruminococcus.torques
## 2024-12-18 17:11:17.599512 INFO::Creating boxplot for categorical data, dysbiosisCD vs Coprobacillus.unclassified
## 2024-12-18 17:11:17.886254 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.cellulosilyticus
## 2024-12-18 17:11:18.156511 INFO::Creating boxplot for categorical data, dysbiosisCD vs Roseburia.intestinalis
## 2024-12-18 17:11:18.431299 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.unclassified
## 2024-12-18 17:11:18.698874 INFO::Creating boxplot for categorical data, dysbiosisCD vs Acidaminococcus.unclassified
## 2024-12-18 17:11:18.946364 INFO::Creating boxplot for categorical data, dysbiosisCD vs Barnesiella.intestinihominis
## 2024-12-18 17:11:19.245148 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.massiliensis
## 2024-12-18 17:11:19.499179 INFO::Creating boxplot for categorical data, dysbiosisCD vs Eubacterium.sp.3.1.31
## 2024-12-18 17:11:19.762372 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroides.salyersiae
## 2024-12-18 17:11:20.01357 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bacteroidales.bacterium.ph8
## 2024-12-18 17:11:20.292608 INFO::Creating boxplot for categorical data, dysbiosisCD vs Clostridium.citroniae
## 2024-12-18 17:11:20.553502 INFO::Creating boxplot for categorical data, dysbiosisCD vs Flavonifractor.plautii
## 2024-12-18 17:11:20.824752 INFO::Creating boxplot for categorical data, dysbiosisCD vs Parabacteroides.goldsteinii
## 2024-12-18 17:11:21.080948 INFO::Creating boxplot for categorical data, dysbiosisCD vs Bifidobacterium.longum
## 2024-12-18 17:11:21.349528 INFO::Creating boxplot for categorical data, dysbiosisCD vs Lachnospiraceae.bacterium.1.1.57FAA
## 2024-12-18 17:11:24.298165 INFO::Plotting data for metadata number 2, dysbiosisUC
## 2024-12-18 17:11:24.301076 INFO::Creating boxplot for categorical data, dysbiosisUC vs Subdoligranulum.unclassified
## 2024-12-18 17:11:24.565168 INFO::Creating boxplot for categorical data, dysbiosisUC vs Faecalibacterium.prausnitzii
## 2024-12-18 17:11:24.841681 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.caccae
## 2024-12-18 17:11:25.111882 INFO::Creating boxplot for categorical data, dysbiosisUC vs Oscillibacter.unclassified
## 2024-12-18 17:11:25.386841 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.uniformis
## 2024-12-18 17:11:25.676183 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.siraeum
## 2024-12-18 17:11:25.978297 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.ovatus
## 2024-12-18 17:11:26.279317 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.fragilis
## 2024-12-18 17:11:26.587129 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.shahii
## 2024-12-18 17:11:26.923678 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.rectale
## 2024-12-18 17:11:27.181815 INFO::Creating boxplot for categorical data, dysbiosisUC vs Roseburia.hominis
## 2024-12-18 17:11:27.454534 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.putredinis
## 2024-12-18 17:11:27.707543 INFO::Creating boxplot for categorical data, dysbiosisUC vs Lachnospiraceae.bacterium.3.1.46FAA
## 2024-12-18 17:11:27.981822 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.hallii
## 2024-12-18 17:11:28.259051 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.xylanisolvens
## 2024-12-18 17:11:28.565361 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.stercoris
## 2024-12-18 17:11:28.832767 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.finegoldii
## 2024-12-18 17:11:29.114031 INFO::Creating boxplot for categorical data, dysbiosisUC vs Barnesiella.intestinihominis
## 2024-12-18 17:11:29.374067 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.leptum
## 2024-12-18 17:11:29.625365 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.gnavus
## 2024-12-18 17:11:29.896734 INFO::Creating boxplot for categorical data, dysbiosisUC vs Alistipes.onderdonkii
## 2024-12-18 17:11:30.151391 INFO::Creating boxplot for categorical data, dysbiosisUC vs Flavonifractor.plautii
## 2024-12-18 17:11:30.405828 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.merdae
## 2024-12-18 17:11:30.665473 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.cellulosilyticus
## 2024-12-18 17:11:30.931275 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.longum
## 2024-12-18 17:11:31.178705 INFO::Creating boxplot for categorical data, dysbiosisUC vs Klebsiella.pneumoniae
## 2024-12-18 17:11:31.437481 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.citroniae
## 2024-12-18 17:11:31.688723 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.ventriosum
## 2024-12-18 17:11:31.942348 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.distasonis
## 2024-12-18 17:11:32.191711 INFO::Creating boxplot for categorical data, dysbiosisUC vs Parabacteroides.goldsteinii
## 2024-12-18 17:11:32.480394 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.torques
## 2024-12-18 17:11:32.751215 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroidales.bacterium.ph8
## 2024-12-18 17:11:33.005532 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.obeum
## 2024-12-18 17:11:33.27544 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.bifidum
## 2024-12-18 17:11:33.567205 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bifidobacterium.adolescentis
## 2024-12-18 17:11:33.845724 INFO::Creating boxplot for categorical data, dysbiosisUC vs Collinsella.aerofaciens
## 2024-12-18 17:11:34.088058 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.hathewayi
## 2024-12-18 17:11:34.369667 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bilophila.unclassified
## 2024-12-18 17:11:34.63383 INFO::Creating boxplot for categorical data, dysbiosisUC vs Eubacterium.eligens
## 2024-12-18 17:11:34.907247 INFO::Creating boxplot for categorical data, dysbiosisUC vs Bacteroides.vulgatus
## 2024-12-18 17:11:35.179901 INFO::Creating boxplot for categorical data, dysbiosisUC vs Clostridium.bolteae
## 2024-12-18 17:11:35.431446 INFO::Creating boxplot for categorical data, dysbiosisUC vs Dialister.invisus
## 2024-12-18 17:11:35.748143 INFO::Creating boxplot for categorical data, dysbiosisUC vs Ruminococcus.lactaris
## 2024-12-18 17:11:36.051345 INFO::Creating boxplot for categorical data, dysbiosisUC vs Burkholderiales.bacterium.1.1.47
## 2024-12-18 17:11:39.307832 INFO::Plotting data for metadata number 3, dysbiosisnonIBD
## 2024-12-18 17:11:39.310566 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Faecalibacterium.prausnitzii
## 2024-12-18 17:11:39.528962 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.rectale
## 2024-12-18 17:11:39.796958 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.sp.3.1.31
## 2024-12-18 17:11:40.062391 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.hominis
## 2024-12-18 17:11:40.300698 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Subdoligranulum.unclassified
## 2024-12-18 17:11:40.566928 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Lachnospiraceae.bacterium.3.1.46FAA
## 2024-12-18 17:11:40.857371 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.torques
## 2024-12-18 17:11:41.132155 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Clostridium.leptum
## 2024-12-18 17:11:41.411439 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Odoribacter.splanchnicus
## 2024-12-18 17:11:41.694443 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Escherichia.coli
## 2024-12-18 17:11:41.951395 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.obeum
## 2024-12-18 17:11:42.225196 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bilophila.unclassified
## 2024-12-18 17:11:42.486815 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.uniformis
## 2024-12-18 17:11:42.766182 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.eligens
## 2024-12-18 17:11:43.060395 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Klebsiella.pneumoniae
## 2024-12-18 17:11:43.337486 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Ruminococcus.bromii
## 2024-12-18 17:11:43.611466 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.vulgatus
## 2024-12-18 17:11:43.876467 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs C2likevirus.unclassified
## 2024-12-18 17:11:44.14301 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bifidobacterium.adolescentis
## 2024-12-18 17:11:44.425315 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.finegoldii
## 2024-12-18 17:11:44.707562 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.inulinivorans
## 2024-12-18 17:11:44.992806 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.onderdonkii
## 2024-12-18 17:11:45.282062 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.unclassified
## 2024-12-18 17:11:45.587921 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Eubacterium.hallii
## 2024-12-18 17:11:45.879821 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Roseburia.intestinalis
## 2024-12-18 17:11:46.177251 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Prevotella.copri
## 2024-12-18 17:11:46.468046 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.dorei
## 2024-12-18 17:11:46.761661 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Bacteroides.fragilis
## 2024-12-18 17:11:47.040551 INFO::Creating boxplot for categorical data, dysbiosisnonIBD vs Alistipes.shahii
## 2024-12-18 17:11:49.916007 INFO::Plotting data for metadata number 4, antibiotics
## 2024-12-18 17:11:49.9183 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.inulinivorans
## 2024-12-18 17:11:50.145795 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.hominis
## 2024-12-18 17:11:50.421509 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.rectale
## 2024-12-18 17:11:50.669275 INFO::Creating boxplot for categorical data, antibiotics vs Dialister.invisus
## 2024-12-18 17:11:50.9455 INFO::Creating boxplot for categorical data, antibiotics vs Roseburia.intestinalis
## 2024-12-18 17:11:51.194442 INFO::Creating boxplot for categorical data, antibiotics vs Dorea.longicatena
## 2024-12-18 17:11:51.431086 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.callidus
## 2024-12-18 17:11:51.686928 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.bromii
## 2024-12-18 17:11:51.910664 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.obeum
## 2024-12-18 17:11:52.15464 INFO::Creating boxplot for categorical data, antibiotics vs Klebsiella.pneumoniae
## 2024-12-18 17:11:52.408515 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.adolescentis
## 2024-12-18 17:11:52.632973 INFO::Creating boxplot for categorical data, antibiotics vs Faecalibacterium.prausnitzii
## 2024-12-18 17:11:52.851906 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.hallii
## 2024-12-18 17:11:53.114119 INFO::Creating boxplot for categorical data, antibiotics vs Bilophila.unclassified
## 2024-12-18 17:11:53.358657 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.leptum
## 2024-12-18 17:11:53.599403 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.3.1.46FAA
## 2024-12-18 17:11:53.826491 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.finegoldii
## 2024-12-18 17:11:54.063216 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.sp.3.1.31
## 2024-12-18 17:11:54.285319 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.fragilis
## 2024-12-18 17:11:54.537775 INFO::Creating boxplot for categorical data, antibiotics vs Alistipes.onderdonkii
## 2024-12-18 17:11:54.759702 INFO::Creating boxplot for categorical data, antibiotics vs Sutterella.wadsworthensis
## 2024-12-18 17:11:55.005929 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.eligens
## 2024-12-18 17:11:55.227783 INFO::Creating boxplot for categorical data, antibiotics vs Collinsella.aerofaciens
## 2024-12-18 17:11:55.470024 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.thetaiotaomicron
## 2024-12-18 17:11:55.71655 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroides.eggerthii
## 2024-12-18 17:11:55.97209 INFO::Creating boxplot for categorical data, antibiotics vs Haemophilus.parainfluenzae
## 2024-12-18 17:11:56.196015 INFO::Creating boxplot for categorical data, antibiotics vs Bifidobacterium.pseudocatenulatum
## 2024-12-18 17:11:56.440876 INFO::Creating boxplot for categorical data, antibiotics vs Ruminococcus.torques
## 2024-12-18 17:11:56.666921 INFO::Creating boxplot for categorical data, antibiotics vs Eubacterium.ventriosum
## 2024-12-18 17:11:56.920723 INFO::Creating boxplot for categorical data, antibiotics vs Parasutterella.excrementihominis
## 2024-12-18 17:11:57.178653 INFO::Creating boxplot for categorical data, antibiotics vs Peptostreptococcaceae.noname.unclassified
## 2024-12-18 17:11:57.417462 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.dispar
## 2024-12-18 17:11:57.670906 INFO::Creating boxplot for categorical data, antibiotics vs Veillonella.atypica
## 2024-12-18 17:11:57.911923 INFO::Creating boxplot for categorical data, antibiotics vs Bacteroidales.bacterium.ph8
## 2024-12-18 17:11:58.157046 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.nexile
## 2024-12-18 17:11:58.409974 INFO::Creating boxplot for categorical data, antibiotics vs Burkholderiales.bacterium.1.1.47
## 2024-12-18 17:11:58.659345 INFO::Creating boxplot for categorical data, antibiotics vs Lachnospiraceae.bacterium.1.1.57FAA
## 2024-12-18 17:11:58.882455 INFO::Creating boxplot for categorical data, antibiotics vs Akkermansia.muciniphila
## 2024-12-18 17:11:59.099701 INFO::Creating boxplot for categorical data, antibiotics vs Clostridium.citroniae
## 2024-12-18 17:11:59.321289 INFO::Creating boxplot for categorical data, antibiotics vs Odoribacter.splanchnicus
## 2024-12-18 17:12:01.936257 INFO::Plotting data for metadata number 5, age
## 2024-12-18 17:12:01.938895 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()`).
## 2024-12-18 17:12:02.245157 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()`).
## 2024-12-18 17:12:02.59355 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()`).
## 2024-12-18 17:12:02.847691 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()`).
## 2024-12-18 17:12:03.149091 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()`).
## 2024-12-18 17:12:03.42414 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()`).
## 2024-12-18 17:12:03.708035 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()`).
## 2024-12-18 17:12:03.980218 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()`).
## 2024-12-18 17:12:04.240148 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()`).
## 2024-12-18 17:12:04.513166 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()`).
## 2024-12-18 17:12:04.761592 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()`).
## 2024-12-18 17:12:05.036648 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()`).
## 2024-12-18 17:12:05.287745 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()`).
## 2024-12-18 17:12:05.540021 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()`).
## 2024-12-18 17:12:05.787775 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()`).
## 2024-12-18 17:12:06.025856 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()`).
## 2024-12-18 17:12:06.299214 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()`).
## 2024-12-18 17:12:06.557762 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()`).
## 2024-12-18 17:12:06.804297 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()`).
## 2024-12-18 17:12:07.056158 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()`).
## 2024-12-18 17:12:07.343401 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()`).
## 2024-12-18 17:12:07.584604 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()`).
## 2024-12-18 17:12:10.505241 INFO::Plotting data for metadata number 6, diagnosis
## 2024-12-18 17:12:10.507368 INFO::Creating boxplot for categorical data, diagnosis vs Bifidobacterium.adolescentis
## 2024-12-18 17:12:10.792643 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2024-12-18 17:12:11.066799 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2024-12-18 17:12:11.362574 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2024-12-18 17:12:11.675984 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2024-12-18 17:12:11.949749 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.clostridioforme
## 2024-12-18 17:12:12.226375 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2024-12-18 17:12:12.493712 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2024-12-18 17:12:12.779139 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2024-12-18 17:12:13.035706 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2024-12-18 17:12:13.29104 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.inulinivorans
## 2024-12-18 17:12:13.55404 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.bromii
## 2024-12-18 17:12:13.845611 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.shahii
## 2024-12-18 17:12:14.15 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2024-12-18 17:12:14.429699 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.goldsteinii
## 2024-12-18 17:12:14.733064 INFO::Creating boxplot for categorical data, diagnosis vs Bilophila.unclassified
## 2024-12-18 17:12:15.00979 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.leptum
## 2024-12-18 17:12:15.334211 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2024-12-18 17:12:15.629944 INFO::Creating boxplot for categorical data, diagnosis vs Coprobacillus.unclassified
## 2024-12-18 17:12:15.938249 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2024-12-18 17:12:16.244618 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.ventriosum
## 2024-12-18 17:12:16.543785 INFO::Creating boxplot for categorical data, diagnosis vs Roseburia.hominis
## 2024-12-18 17:12:16.850535 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2024-12-18 17:12:17.145345 INFO::Creating boxplot for categorical data, diagnosis vs Akkermansia.muciniphila
## 2024-12-18 17:12:17.443662 INFO::Creating boxplot for categorical data, diagnosis vs Sutterella.wadsworthensis
## 2024-12-18 17:12:17.725198 INFO::Creating boxplot for categorical data, diagnosis vs Subdoligranulum.unclassified
## 2024-12-18 17:12:17.974341 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.symbiosum
## 2024-12-18 17:12:18.246564 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.onderdonkii
## 2024-12-18 17:12:18.50449 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.unclassified
## 2024-12-18 17:12:18.76731 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.lactaris
## 2024-12-18 17:12:19.041833 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2024-12-18 17:12:19.312113 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.gnavus
## 2024-12-18 17:12:19.618438 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.fragilis
## 2024-12-18 17:12:19.878455 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2024-12-18 17:12:20.158427 INFO::Creating boxplot for categorical data, diagnosis vs Odoribacter.splanchnicus
## 2024-12-18 17:12:20.423312 INFO::Creating boxplot for categorical data, diagnosis vs Parabacteroides.distasonis
## 2024-12-18 17:12:20.699002 INFO::Creating boxplot for categorical data, diagnosis vs Eubacterium.rectale
## 2024-12-18 17:12:20.948778 INFO::Creating boxplot for categorical data, diagnosis vs Bacteroides.finegoldii
## 2024-12-18 17:12:21.233403 INFO::Creating boxplot for categorical data, diagnosis vs Ruminococcus.callidus
## 2024-12-18 17:12:21.527105 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.finegoldii
## 2024-12-18 17:12:21.770005 INFO::Creating boxplot for categorical data, diagnosis vs Alistipes.putredinis
## 2024-12-18 17:12:22.02683 INFO::Creating boxplot for categorical data, diagnosis vs Clostridium.bolteae
## 2024-12-18 17:12:22.298798 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 Under development (unstable) (2024-11-20 r87352)
## Platform: x86_64-apple-darwin20
## Running under: macOS Monterey 12.7.6
## 
## Matrix products: default
## BLAS:   /Library/Frameworks/R.framework/Versions/4.5-x86_64/Resources/lib/libRblas.0.dylib 
## LAPACK: /Library/Frameworks/R.framework/Versions/4.5-x86_64/Resources/lib/libRlapack.dylib;  LAPACK version 3.12.0
## 
## locale:
## [1] C/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8
## 
## time zone: America/New_York
## tzcode source: internal
## 
## attached base packages:
## [1] stats     graphics  grDevices utils     datasets  methods   base     
## 
## other attached packages:
## [1] Maaslin2_1.21.0
## 
## loaded via a namespace (and not attached):
##  [1] sass_0.4.9          generics_0.1.3      robustbase_0.99-4-1
##  [4] lattice_0.22-6      lme4_1.1-35.5       digest_0.6.37      
##  [7] magrittr_2.0.3      RColorBrewer_1.1-3  evaluate_1.0.1     
## [10] grid_4.5.0          mvtnorm_1.3-2       fastmap_1.2.0      
## [13] jsonlite_1.8.9      Matrix_1.7-1        DBI_1.2.3          
## [16] optparse_1.7.5      biglm_0.9-3         mgcv_1.9-1         
## [19] scales_1.3.0        pbapply_1.7-2       permute_0.9-7      
## [22] numDeriv_2016.8-1.1 getopt_1.20.4       jquerylib_0.1.4    
## [25] cli_3.6.3           rlang_1.1.4         crayon_1.5.3       
## [28] munsell_0.5.1       splines_4.5.0       withr_3.0.2        
## [31] cachem_1.1.0        yaml_2.3.10         vegan_2.6-8        
## [34] tools_4.5.0         parallel_4.5.0      nloptr_2.1.1       
## [37] minqa_1.2.8         dplyr_1.1.4         colorspace_2.1-1   
## [40] ggplot2_3.5.1       boot_1.3-31         hash_2.2.6.3       
## [43] vctrs_0.6.5         R6_2.5.1            lifecycle_1.0.4    
## [46] MASS_7.3-61         logging_0.10-108    pcaPP_2.0-5        
## [49] cluster_2.1.8       pkgconfig_2.0.3     gtable_0.3.6       
## [52] lmerTest_3.1-3      pillar_1.10.0       bslib_0.8.0        
## [55] Rcpp_1.0.13-1       data.table_1.16.4   glue_1.8.0         
## [58] DEoptimR_1.1-3-1    xfun_0.49           tibble_3.2.1       
## [61] tidyselect_1.2.1    knitr_1.49          farver_2.1.2       
## [64] htmltools_0.5.8.1   nlme_3.1-166        labeling_0.4.3     
## [67] rmarkdown_2.29      pheatmap_1.0.12     compiler_4.5.0

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.