Using FLORAL for Microbiome Analysis

library(FLORAL)
library(dplyr)
library(patchwork)

Data

We will be using data from the curatedMetagenomicData package. For easier installation, we saved a flat copy of the data, but the steps below show how that was created.

if (! "BiocManager" %in% installed.packages()) install.packages("BiocManager")
if (! "curatedMetagenomicData" %in% installed.packages()) BiocManager::install("curatedMetagenomicData")
if (! "patchwork" %in% installed.packages()) install.packages("patchwork")
library(curatedMetagenomicData)

# Take a look at the summary of the studies available:
curatedMetagenomicData::sampleMetadata  |> group_by(.data$study_name, .data$study_condition) |> count() |> arrange(.data$study_name) 
#As an example, let us look at the `YachidaS_2019` study between healthy controls and colorectal cancer (CRC) patients.
curatedMetagenomicData::curatedMetagenomicData("YachidaS_2019") 
rawdata <- curatedMetagenomicData::curatedMetagenomicData("2021-03-31.YachidaS_2019.relative_abundance", dryrun = FALSE, counts = TRUE) |>  mergeData()
x <- SummarizedExperiment::assays(rawdata)$relative_abundance %>% t()
y <- rawdata@colData$disease

save(list = c("x", "y"),  file = file.path("inst", "extdata", "YachidaS_2019.Rdata"))
load(system.file("extdata", "YachidaS_2019.Rdata", package="FLORAL"))

Running FLORAL

We extracted the data from the TreeSummarizedExperiment object to two objects: the taxa matrix x and the “outcomes” vector y of whether a patient is healthy or has colorectal cancer (CRC). Note that for binary outcomes, the input vector y needs to be formatted with entries equal to either 0 or 1. In addition, we need to specify family = "binomial" in FLORAL to fit the logistic regression model. To print the progress bar as the algorithm runs, please use progress = TRUE.

  
x <- x[y %in% c("CRC","healthy"),]
x <- x[,colSums(x >= 100) >= nrow(x)*0.2] # filter low abundance taxa

y <- as.numeric(as.factor(y[y %in% c("CRC","healthy")]))-1
fit <- FLORAL(x = x, y = y, family="binomial", ncv=10, progress=TRUE)
#> Algorithm running for full dataset: 
#> Algorithm running for cv dataset 1 out of 10: 
#> Algorithm running for cv dataset 2 out of 10: 
#> Algorithm running for cv dataset 3 out of 10: 
#> Algorithm running for cv dataset 4 out of 10: 
#> Algorithm running for cv dataset 5 out of 10: 
#> Algorithm running for cv dataset 6 out of 10: 
#> Algorithm running for cv dataset 7 out of 10: 
#> Algorithm running for cv dataset 8 out of 10: 
#> Algorithm running for cv dataset 9 out of 10: 
#> Algorithm running for cv dataset 10 out of 10:

Interpreting the Model

FLORAL, like other methods that have an optimization step, has two “best” solutions for \(\lambda\) available: one minimizing the mean squared error (\(\lambda_\min\)), and one maximizing the value of \(\lambda\) withing 1 standard error of the minimum mean squared error (\(\lambda_{\text{1se}}\)). These are referred to as the min and 1se solutions, respectively.

We can see the mean squared error (MSE) and the coefficients vs log(\(\lambda\)) as follows:

fit$pmse + fit$pcoef

In both plots, the vertical dashed line and dotted line represent \(\lambda_\min\) and \(\lambda_{\text{1se}}\), respectively. In the MSE plot, the bands represent plus minus one standard error of the MSE. In the coefficient plot, the colored lines represent individual taxa, where taxa with non-zero values at \(\lambda_\min\) and \(\lambda_{\text{1se}}\) are selected as predictive of the outcome.

To view specific names of the selected taxa, please see fit$selected.feature$min or fit$selected.feature$1se vectors. To view all coefficient estimates, please see fit$best.beta$min or fit$best.beta$1se. Without looking into ratios, one can crudely interpret positive or negative association between a taxon and the outcome by the positive or negative sign of the coefficient estimates. However, we recommend referring to the two-step procedure discussed below for a more rigorous interpretation based on ratios, which is derived from the log-ratio model assumption.


head(fit$selected.feature$min)
#> [1] "k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02"                
#> [2] "k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_ICM47"                     
#> [3] "k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium|s__Bifidobacterium_pseudocatenulatum"
#> [4] "k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Micrococcales|f__Micrococcaceae|g__Rothia|s__Rothia_mucilaginosa"                               
#> [5] "k__Bacteria|p__Actinobacteria|c__Coriobacteriia|o__Coriobacteriales|f__Coriobacteriaceae|g__Collinsella|s__Collinsella_aerofaciens"                
#> [6] "k__Bacteria|p__Actinobacteria|c__Coriobacteriia|o__Eggerthellales|f__Eggerthellaceae|g__Asaccharobacter|s__Asaccharobacter_celatus"

head(sort(fit$best.beta$min))
#>                  k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra 
#>                                                                                                                        -0.08130690 
#> k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02 
#>                                                                                                                        -0.04913093 
#> k__Bacteria|p__Actinobacteria|c__Coriobacteriia|o__Coriobacteriales|f__Coriobacteriaceae|g__Collinsella|s__Collinsella_aerofaciens 
#>                                                                                                                        -0.04660536 
#>  k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis 
#>                                                                                                                        -0.03803803 
#>          k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Ruminococcaceae|g__Anaerotruncus|s__Anaerotruncus_colihominis 
#>                                                                                                                        -0.03261738 
#>      k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_cellulosilyticus 
#>                                                                                                                        -0.02280316

The Two-step Procedure

In the previous section, we checked the lasso estimates without identifying specific ratios that are predictive of the outcome (CRC in this case). By default, FLORAL performs a two-step selection procedure to use glmnet and step regression to further identify taxa pairs which form predictive log-ratios. To view those pairs, use fit$step2.ratios$min or fit$step2.ratios$1se for names of ratios and fit$step2.ratios$min.idx or fit$step2.ratios$1se.idx for the pairs of indices in the original input count matrix x. Note that one taxon can occur in multiple ratios.


head(fit$step2.ratios$`1se`)
#> [1] "k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_plebeius/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_dorei"                                             
#> [2] "k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_eligens/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_cellulosilyticus"                                       
#> [3] "k__Bacteria|p__Actinobacteria|c__Coriobacteriia|o__Coriobacteriales|f__Coriobacteriaceae|g__Collinsella|s__Collinsella_aerofaciens/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium|s__Bifidobacterium_pseudocatenulatum"
#> [4] "k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Streptococcaceae|g__Streptococcus|s__Streptococcus_salivarius/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02"                          
#> [5] "k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Erysipelatoclostridium|s__Erysipelatoclostridium_ramosum/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra"             
#> [6] "k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_sp_CAG_38"

fit$step2.ratios$`1se.idx`
#>      [,1] [,2] [,3] [,4] [,5] [,6] [,7] [,8] [,9] [,10] [,11]
#> [1,]    1    4   12   22   26   27   27   44   73    81   114
#> [2,]    2  113  102  128  131   90  151   64   80   128   131

To further interpret the positive or negative associations between the outcome, please refer to the output step regression tables, where the effect sizes of the ratios can be found.

While the corresponding p-values are also available, we recommend only using the p-values as a criterion to rank the strength of the association. We do not recommend directly reporting the p-values for inference, because these p-values were obtained after running the first step lasso model without rigorous post-selective inference. However, it is still valid to claim these selected log-ratios are predictive of the outcome, as demonstrated by the improved 10-fold cross-validated prediction errors.


fit$step2.tables$`1se`
#>                                                                                                                                                                                                                                                                                                                                    Estimate
#> k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_plebeius/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_dorei                                                                                        -0.02659576
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_eligens/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_cellulosilyticus                                                                                   0.04043200
#> k__Bacteria|p__Actinobacteria|c__Coriobacteriia|o__Coriobacteriales|f__Coriobacteriaceae|g__Collinsella|s__Collinsella_aerofaciens/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium|s__Bifidobacterium_pseudocatenulatum                                           -0.04886115
#> k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Streptococcaceae|g__Streptococcus|s__Streptococcus_salivarius/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02                                                                      0.09235497
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Erysipelatoclostridium|s__Erysipelatoclostridium_ramosum/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra                                                         0.04784129
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_sp_CAG_38                                                                            -0.03798158
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Proteobacteria|c__Proteobacteria_unclassified|o__Proteobacteria_unclassified|f__Proteobacteria_unclassified|g__Proteobacteria_unclassified|s__Proteobacteria_bacterium_CAG_139 -0.03610014
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Ruminococcaceae|g__Anaerotruncus|s__Anaerotruncus_colihominis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Lachnoclostridium|s__Clostridium_aldenense                                                                             -0.07966635
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Lachnospira|s__Lachnospira_pectinoschiza/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Blautia|s__Ruminococcus_torques                                                                                           0.03247124
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Roseburia|s__Roseburia_faecis/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02                                                                                  0.05287895
#> k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Barnesiellaceae|g__Coprobacter|s__Coprobacter_fastidiosus/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra                                                                                      0.06757447
#>                                                                                                                                                                                                                                                                                                                                 Std. Error
#> k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_plebeius/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_dorei                                                                                        0.01099163
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_eligens/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_cellulosilyticus                                                                                  0.01385914
#> k__Bacteria|p__Actinobacteria|c__Coriobacteriia|o__Coriobacteriales|f__Coriobacteriaceae|g__Collinsella|s__Collinsella_aerofaciens/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium|s__Bifidobacterium_pseudocatenulatum                                           0.01507927
#> k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Streptococcaceae|g__Streptococcus|s__Streptococcus_salivarius/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02                                                                     0.02625928
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Erysipelatoclostridium|s__Erysipelatoclostridium_ramosum/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra                                                        0.01965003
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_sp_CAG_38                                                                            0.01622550
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Proteobacteria|c__Proteobacteria_unclassified|o__Proteobacteria_unclassified|f__Proteobacteria_unclassified|g__Proteobacteria_unclassified|s__Proteobacteria_bacterium_CAG_139 0.01837978
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Ruminococcaceae|g__Anaerotruncus|s__Anaerotruncus_colihominis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Lachnoclostridium|s__Clostridium_aldenense                                                                             0.02173633
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Lachnospira|s__Lachnospira_pectinoschiza/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Blautia|s__Ruminococcus_torques                                                                                          0.01300914
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Roseburia|s__Roseburia_faecis/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02                                                                                 0.01743096
#> k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Barnesiellaceae|g__Coprobacter|s__Coprobacter_fastidiosus/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra                                                                                     0.01818976
#>                                                                                                                                                                                                                                                                                                                                   z value
#> k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_plebeius/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_dorei                                                                                        -2.419638
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_eligens/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_cellulosilyticus                                                                                   2.917353
#> k__Bacteria|p__Actinobacteria|c__Coriobacteriia|o__Coriobacteriales|f__Coriobacteriaceae|g__Collinsella|s__Collinsella_aerofaciens/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium|s__Bifidobacterium_pseudocatenulatum                                           -3.240286
#> k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Streptococcaceae|g__Streptococcus|s__Streptococcus_salivarius/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02                                                                      3.517042
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Erysipelatoclostridium|s__Erysipelatoclostridium_ramosum/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra                                                         2.434668
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_sp_CAG_38                                                                            -2.340857
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Proteobacteria|c__Proteobacteria_unclassified|o__Proteobacteria_unclassified|f__Proteobacteria_unclassified|g__Proteobacteria_unclassified|s__Proteobacteria_bacterium_CAG_139 -1.964123
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Ruminococcaceae|g__Anaerotruncus|s__Anaerotruncus_colihominis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Lachnoclostridium|s__Clostridium_aldenense                                                                             -3.665124
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Lachnospira|s__Lachnospira_pectinoschiza/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Blautia|s__Ruminococcus_torques                                                                                           2.496033
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Roseburia|s__Roseburia_faecis/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02                                                                                  3.033621
#> k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Barnesiellaceae|g__Coprobacter|s__Coprobacter_fastidiosus/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra                                                                                      3.714973
#>                                                                                                                                                                                                                                                                                                                                     Pr(>|z|)
#> k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_plebeius/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_dorei                                                                                        0.0155359842
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_eligens/k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Bacteroidaceae|g__Bacteroides|s__Bacteroides_cellulosilyticus                                                                                  0.0035301655
#> k__Bacteria|p__Actinobacteria|c__Coriobacteriia|o__Coriobacteriales|f__Coriobacteriaceae|g__Collinsella|s__Collinsella_aerofaciens/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Bifidobacteriales|f__Bifidobacteriaceae|g__Bifidobacterium|s__Bifidobacterium_pseudocatenulatum                                           0.0011940974
#> k__Bacteria|p__Firmicutes|c__Bacilli|o__Lactobacillales|f__Streptococcaceae|g__Streptococcus|s__Streptococcus_salivarius/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02                                                                     0.0004363849
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Erysipelatoclostridium|s__Erysipelatoclostridium_ramosum/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra                                                        0.0149054787
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Eubacteriaceae|g__Eubacterium|s__Eubacterium_sp_CAG_38                                                                            0.0192395438
#> k__Bacteria|p__Firmicutes|c__Erysipelotrichia|o__Erysipelotrichales|f__Erysipelotrichaceae|g__Holdemania|s__Holdemania_filiformis/k__Bacteria|p__Proteobacteria|c__Proteobacteria_unclassified|o__Proteobacteria_unclassified|f__Proteobacteria_unclassified|g__Proteobacteria_unclassified|s__Proteobacteria_bacterium_CAG_139 0.0495158342
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Ruminococcaceae|g__Anaerotruncus|s__Anaerotruncus_colihominis/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Lachnoclostridium|s__Clostridium_aldenense                                                                             0.0002472185
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Lachnospira|s__Lachnospira_pectinoschiza/k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Blautia|s__Ruminococcus_torques                                                                                          0.0125590844
#> k__Bacteria|p__Firmicutes|c__Clostridia|o__Clostridiales|f__Lachnospiraceae|g__Roseburia|s__Roseburia_faecis/k__Bacteria|p__Actinobacteria|c__Actinobacteria|o__Actinomycetales|f__Actinomycetaceae|g__Actinomyces|s__Actinomyces_sp_HMSC035G02                                                                                 0.0024163753
#> k__Bacteria|p__Bacteroidetes|c__Bacteroidia|o__Bacteroidales|f__Barnesiellaceae|g__Coprobacter|s__Coprobacter_fastidiosus/k__Bacteria|p__Firmicutes|c__Tissierellia|o__Tissierellales|f__Peptoniphilaceae|g__Parvimonas|s__Parvimonas_micra                                                                                     0.0002032250