Introduction and initialization

This is an R Markdown file containing code to parse the results of a dada2 analysis into phyloseq for further analysis. It is separated into chunks that may be run independently by pressing the play button. You will need 3 files in the same location in order to run this pipeline successfully:

Recommended use: Set the individual chunks until you are content with the ouput, then knit the whole document into a PDF/html, so you have a full record of a successful run.

Optional custom Taxonomy file

A custom taxonomy file may be provided instead of using the taxonomy output from dada2. This may be used to supply taxonomy derived e.g. from BLAST searches of the ASVs. Custom taxonomy files must be tab-delimited text with as many rows as the original, colum headers (for all columns except for the first column). For example:

Kingdom Phylum Class Order Family Genus Species
ESV1 Kingdomx Phylumx Classx Orderx Familyx Genusx Speciesx
ESV2 Kingdomy Phylumy Classy Ordery Familyy Genusy Speciesy
ESV3 Kingdomz Phylumz Classz Orderz Familyz Genusz Speciesz
… ESVn Kingdomy Phylumy Classy Ordery Familyy Genusy Speciesy

Friendly warning: Parsing the results of a BLAST search into this format may require some effort.

Descriptor table

‘descriptors.txt’ should be a tab-delimited .txt table describing the samples. It must have the same length and order as the samples in seqtab_nochim.rds. To check the order and length of samples in seqtab_nochim.rds and generate a template to fill out, you may run the chunk below with “optional_sample_check” set to “TRUE”.

Any number of descriptors is possible. The sample names may be retained as one descriptor, but this is not necessary, as they will be added during parsing. For example, if there are 4 samples (order: s1, s2, s3, s4), the txt file could look as follows:

Subject Species Time
Kar1 A.thaliana 24hpi
Kar1 A.thaliana 72hpi
Mec2 S.tuberosum 24hpi
Mec3 S.tuberosum 24hpi

Finally, the file should end with an empty line, since it may throw an error otherwise. However, this is usually not a serious problem.

If you choose to use the blank file, you MUST retain the original order of the samples!

Setup

This chunk also loads required packages and defines the location of the input files. It requires the correct path as input, and allows setting the pruning of control samples and choosing generation of a phylogenetic tree. Beware: The generation of a phylogenetic tree may take several days for >1000 sequences, it is therefore recommended to only use this feature for the final analysis or small sample sets. This scricpt assumes the packages Biostrings, dada2, DECIPHER, ggplot2, ggsci, phangorn, phyloseq and stringr to be installed.

# CHANGE ME to the directory that contains 'seqtab_nochim.rds'
path = "16S_DADA2_results_260821/"

# CHANGE ME to TRUE to list all samples and generate an empty metadata file 
optional_sample_check = TRUE

# CHANGE ME to TRUE to update cuphyr
update_cuphyr = TRUE

# Initiate by loading packages and setting knit options
################# NO CHANGES NECESSARY BELOW #################
knitr::opts_chunk$set(echo = TRUE)
knitr::opts_chunk$set(root.dir = paste0(path))
knitr::opts_chunk$set(message = FALSE)
knitr::opts_chunk$set(warning = FALSE)

if (update_cuphyr) {
  devtools::install_github("simeross/cuphyr")
}

# Sequence and microbiome specific libraries
library(dada2)
library(Biostrings)
library(DECIPHER)
library(cuphyr)
# The export of phyloseq objects to a BIOM format and the generation of fancier 
# ordination plots require the phyloseq-extended package. The first command 
# installs the package that is currently on the dev brach of the author's 
# repository, the second command sources some extra functions, including the 
# better ordination plot implementation.
remotes::install_github("mahendra-mariadassou/phyloseq-extended", ref = "dev")
source("https://raw.githubusercontent.com/mahendra-mariadassou/phyloseq-extended/master/load-extra-functions.R" )

library(phyloseq)
library(SIAMCAT)

# Phylogeny libraries
library(phangorn)
library(ape)

# Plotting and figure export
library(gridExtra)
library(viridis)
library(ggpubr)

# Tidyverse
library(tidyverse)
library(stringr)

# Various packages for specific analysis
library(readxl)
library(openxlsx)
library(ggpmisc)
library(betareg)
library(BBmisc)
library(aod)
library(betareg)
#install.packages('MicrobiomeStat')
library(MicrobiomeStat)

# Checks whether output path exists and creates it if not. Throws warning if 
# directory exists.
outp <- paste0(path,"/analysis_output")
dir.create(file.path(outp))

if (optional_sample_check) {
  seqtabcheck <- readRDS(paste0(path,"/seqtab_nochim.rds")) 
  samps <- rownames(seqtabcheck)
  lensamps <- length(samps)
  blankcol <- vector(mode = "character", length = lensamps)
  blanktable <- data.frame(SampleIDs = samps, ExampleProperty1 = blankcol, 
                           ExampleProperty2 = blankcol, 
                           ExampleProperty3 = blankcol)
  write.table(blanktable, file = paste0(path, "/descriptors_blank.txt"), 
              sep = "\t", row.names = F)
  cat("'seqtab_nochim.rds' contains samples in the following order:\n", 
      samps, "\nThe number of samples in the file is:", lensamps, sep = "\n")
  rm(optional_sample_check, seqtabcheck, samps, 
     lensamps, blankcol, blanktable, update_cuphyr)
  }else{rm(optional_sample_check, update_cuphyr)}
## 'seqtab_nochim.rds' contains samples in the following order:
## 
## 1
## 10
## 11
## 12
## 13
## 14
## 15
## 16
## 17
## 18
## 2
## 3
## 4
## 5
## 6
## 7
## 8
## 9
## NegativK-2
## NegativK-4
## PositivK-1
## PositivK-2
## PositivK-3
## PositivK-4
## Vann1
## Vann2
## 
## The number of samples in the file is:
## 26

Parameters

This chunk allows the adjustment of several parameters, such as setting the pruning of control samples based on keywords, requiring that a phylogenetic tree be provided or generated, defining a minimum ASV count and providing an alternative taxonomy.

# Dedicated environment containing all global analysis settings for better
# overview and collected export of settings
parameters <- new.env()

# CHANGE ME to 'TRUE' to remove control samples from the analysis or 'FALSE' to
# analyse all samples.
parameters$prune_controls = "TRUE"
# CHANGE ME to a list of unique identifiers that only occur in the names of
# samples you do NOT want to analyse. Common examples are provided.
parameters$controls = c("Pos", "NegativK", "PositivK", "Vann", "H2O", "Neg", "Kontr",
    "Contr", "POSK")

# CHANGE ME to 'TRUE' to remove certain taxonomic groups from the analysis by
# name. This is useful to exclude non-target organisms or noise from organelles
# such as Chloroplasts and Mitochondria. It is recommended to first look at all
# data before using this setting.
parameters$prune_noise_taxgroups = "FALSE"
# CHANGE ME to define the taxonomic groups to be removed as noise.
parameters$noise_taxgroups = c("Chloroplast", "Mitochondria")

# CHANGE ME to a number of ASV counts [~reads] that analyzed samples should
# minimally have. Samples with lower ASV counts than 'minread' will be pruned.
# Set to 0 to not prune any samples.
parameters$minASVcount = 3000

# CHANGE ME to 'TRUE', if you want to provide a custom taxonomy table instead
# of using the default dada2 output ('taxa.rds').
parameters$customTax = "TRUE"
# CHANGE ME to the location of the custom taxonomy file. This only matters if
# parameters$customTax='TRUE', otherwise it will be ignored.
parameters$taxfile = "16S_DADA2_results_260821/custom_BLAST_taxonomy_nt.txt"

# CHANGE ME to 'TRUE' to generate a phylogenetic tree. This process takes a
# long time depending on the number of sequences (up to days for thousands).
# If a tree is provided as 'phylotree.rds' in 'path', then it will be used
# regardless of the value of 'parameters$maketree'
parameters$maketree = "FALSE"

# CHANGE ME to 'TRUE' to root the used phylogenetic tree (if one exists) on the
# leaf with the longest branch (outgroup). This makes analyses that rely on the
# phylogenetic tree reproducible instead of picking a random leaf as root when
# calculating UNIFRAC distances. Implementation based on
# http://john-quensen.com/r/unifrac-and-tree-roots/ and answers in
# https://github.com/joey711/phyloseq/issues/597
parameters$roottree = "TRUE"

## CHANGE ME to 'TRUE' to export all generated phyloseq objects as .biom
## objects
parameters$biom_export = "FALSE"

Parsing input data

This chunk loads the input data into a usable format.This chunk does not require any user inputs. If no phylogenetic tree with the name ‘phylotree.rds’ was provided and ‘parameters$maketree=“TRUE”’, it will be calculated here. The phylogenetic tree is necessary for certain plots that incorporate ‘true’ taxonomic relationships beyond the annotations, such as PCoA.

############### NO NEED FOR CHANGES BELOW ############### Make dedicated
############### environments to contain temporary values and manage other
############### objects
tmp <- new.env()
plots <- new.env()
set <- new.env()

# Read in variables
tmp$seqtabp <- readRDS(paste0(path, "/seqtab_nochim.rds"))
if (parameters$customTax == "TRUE") {
    tmp$taxap <- read.delim(parameters$taxfile, header = TRUE, sep = "\t")
    rownames(tmp$taxap) <- colnames(tmp$seqtabp)
    tmp$taxap <- as.matrix(tmp$taxap)
} else {
    tmp$taxap <- readRDS(paste0(path, "/taxa.rds"))
}
tmp$samp_table <- read.delim(paste0(path, "/descriptors.txt"), header = TRUE, sep = "\t")
tmp$samp_list <- rownames(tmp$seqtabp)

# Check if descriptors has the same samples as seqtabp
if (length(tmp$samp_table[, 1]) != length(tmp$samp_list)) {
    stop("There are ", length(tmp$samp_table[, 1]), " samples in 'descriptors.txt', but ",
        length(tmp$samp_list), " samples in 'seqtab_nochim.rds'. Please make sure that the correct samples 
    are contained in descriptors.txt.
       
    You may use 'optional_sample_check <- TRUE' in the first chunk to generate an 
    empty template for 'descriptors.txt'")
} else if (!identical(tmp$samp_table[, 1], tmp$samp_list)) {
    warning("Warning: The samples in 'descriptors.txt' do not have the same names 
          or order as the samples in 'seqtab_nochim.rds'. This may be fine if 
          abbreviated names were used or the sample names are not contained in 
          the first column of 'descriptors.txt'. Double-checking never hurts!")
}


# generate phylogenetic tree of ASVs only if there is no file called
# 'phylotree.rds' in the working directory and 'parameters$maketree' is 'TRUE'
if (!file.exists(paste0(path, "/phylotree.rds"))) {
    if (parameters$maketree == "TRUE") {
        tmp$ASVs <- getSequences(tmp$seqtabp)
        names(tmp$ASVs) <- tmp$ASVs
        tmp$ASV_align <- AlignSeqs(DNAStringSet(tmp$ASVs), anchor = NA)
        tmp$ASV_phang <- phyDat(as(tmp$ASV_align, "matrix"), type = "DNA")
        tmp$dm <- dist.ml(tmp$ASV_phang)
        tmp$treeNJ <- NJ(tmp$dm)
        tmp$fit <- pml(tmp$treeNJ, data = tmp$ASV_phang)
        tmp$fitGTR <- update(tmp$fit, k = 4, inv = 0.2)
        tmp$fitGTR <- optim.pml(tmp$fitGTR, model = "GTR", optInv = TRUE, optGamma = TRUE,
            rearrangement = "stochastic", control = pml.control(trace = 0))
        saveRDS(tmp$fitGTR, file = paste0(path, "/phylotree.rds"))
    }
}

## parse into phyloseq object
row.names(tmp$samp_table) <- tmp$samp_list
if (file.exists(paste0(path, "/phylotree.rds"))) {
    tmp$treep <- readRDS(paste0(path, "/phylotree.rds"))
    p <- phyloseq(otu_table(tmp$seqtabp, taxa_are_rows = FALSE), sample_data(tmp$samp_table),
        tax_table(tmp$taxap), phy_tree(tmp$treep$tree))
} else {
    p <- phyloseq(otu_table(tmp$seqtabp, taxa_are_rows = FALSE), sample_data(tmp$samp_table),
        tax_table(tmp$taxap))
}

## Adding nucleotide info and giving sequences ASV## identifiers
tmp$ASV_sequences <- Biostrings::DNAStringSet(taxa_names(p))
taxa_names(p) <- paste0("ASV", seq(ntaxa(p)))
names(tmp$ASV_sequences) <- taxa_names(p)
p <- merge_phyloseq(p, tmp$ASV_sequences)

## optional pruning
if (parameters$prune_controls == "TRUE") {
    if (!is.null(parameters$controls)) {
        tmp$samp_clean <- tmp$samp_list[!tmp$samp_list %in% grep(paste0(parameters$controls,
            collapse = "|"), tmp$samp_list, value = T)]
        tmp$contr_pruned <- setdiff(tmp$samp_list, tmp$samp_clean)
        ps <- prune_samples(tmp$samp_clean, p)
        # Physeq object for Just controls
        ps.contr <- prune_samples(tmp$contr_pruned, p)
        ps.contr <- prune_taxa(taxa_sums(ps.contr) > 0, ps.contr)
        ps.transcontr <- transform_sample_counts(ps.contr, function(ASV) ASV/sum(ASV))

        message(cat("\n", "Number of control samples that were pruned and will not be analysed:\n",
            length(tmp$samp_list) - length(tmp$samp_clean), "\n", "The following controls were pruned:\n",
            tmp$contr_pruned, "The controls are contained in a separate phyloseq object: ps.contr",
            "\n", sep = "\n"))
    } else {
        warning(cat("\n\nparameters$prune_controls is TRUE but 'parameters$controls' is empty. 
    No samples were pruned.\n\n"))
    }
} else {
    ps <- p
}
## 
## 
## Number of control samples that were pruned and will not be analysed:
## 
## 8
## 
## 
## The following controls were pruned:
## 
## NegativK-2
## NegativK-4
## PositivK-1
## PositivK-2
## PositivK-3
## PositivK-4
## Vann1
## Vann2
## The controls are contained in a separate phyloseq object: ps.contr
# Prune ASVs defined as noise
if (parameters$prune_noise_taxgroups == "TRUE") {
    tmp$ps_taxlvls <- colnames(tax_table(ps))
    tmp$noise_ASVs <- character(0)
    for (lvl in tmp$ps_taxlvls) {
        tmp$noise_ASVs <- c(tmp$noise_ASVs, cuphyr::list_subset_ASVs(physeq = ps,
            subv = parameters$noise_taxgroups, taxlvlsub = lvl))
    }
    tmp$noise_ASVs <- unique(tmp$noise_ASVs)
    tmp$no_noise_ASVs <- colnames(otu_table(ps))
    tmp$no_noise_ASVs <- setdiff(tmp$no_noise_ASVs, tmp$noise_ASVs)
    if (length(tmp$noise_ASVs) > 0) {
        ps <- prune_taxa(tmp$no_noise_ASVs, ps)
        tmp$no_noise_ps <- ps
        cat(length(tmp$noise_ASVs), "ASVs were pruned because they belonged to the following 
        taxonomic groups:\n")
        cat(parameters$noise_taxgroups, "\n", sep = "\n")
    } else {
        cat("No ASVs were recognized as belonging to the following taxonomic groups 
        defined as noise:\n")
        cat(parameters$noise_taxgroups, "\n", sep = "\n")
    }
}

# Prune samples with fewer than reads than minASVcount
if (parameters$minASVcount > 0) {
    tmp$samp_pruned <- names(which(sample_sums(ps) < parameters$minASVcount))
    ps <- prune_samples(sample_sums(ps) >= parameters$minASVcount, ps)
    if (length(tmp$samp_pruned) > 0) {
        cat("The following samples were pruned because ASV counts were lower than",
            parameters$minASVcount, ":\n")
        cat(tmp$samp_pruned, "\n", sep = "\n")
    }
}

# Remove 0 count ASVs (e.g. control ASVs that remain) from the base object
ps <- prune_taxa(taxa_sums(ps) > 0, ps)

# Get a tbl of the base object for easier access in some phyloseq-independent
# analyses. Takes some seconds, potentially up to minutes.
ps_tbl <- as_tibble(psmelt(ps))

# Transformed per sample (per-sample relative abundance)
ps.trans <- transform_sample_counts(ps, function(ASV) ASV/sum(ASV))

# Read NDVI values as numeric
sample_data(ps)[["ndvi"]] <- as.numeric(sample_data(ps)[["ndvi"]])
sample_data(ps.trans)[["ndvi"]] <- as.numeric(sample_data(ps.trans)[["ndvi"]])


# Get a tbl of the base object for easier access in some phyloseq-independent
# analyses. Takes some seconds, potentially up to minutes.
ps_tbl <- as_tibble(psmelt(ps))
ps_trans_tbl <- as_tibble(psmelt(ps.trans))

# Condensing to Abundance per Genus and Sample
genus_abundance_tbl_per_sample <- ps_trans_tbl %>%
    group_by(Genus, Sample) %>%
    mutate(Genus_Sample_Abundance = sum(Abundance)) %>%
    select(Genus, Sample, ndvi, Genus_Sample_Abundance, Alias) %>%
    unique()

if (parameters$roottree == "TRUE" && parameters$maketree == "TRUE") {
    phyloseq::phy_tree(ps) <- cuphyr::root_tree_in_outgroup(physeq = ps)
}

if (parameters$biom_export == "TRUE") {
    suppressWarnings(phyloseq.extended::write_phyloseq(p, biom_file = paste0(path,
        "all_samples.biom"), biom_format = "standard"))
    suppressWarnings(phyloseq.extended::write_phyloseq(ps, biom_file = file.path(path,
        "samples_without_controls.biom"), biom_format = "standard"))
    suppressWarnings(phyloseq.extended::write_phyloseq(ps.trans, biom_file = file.path(path,
        "samples_without_controls_rel_abundance.biom"), biom_format = "standard"))
    suppressWarnings(phyloseq.extended::write_phyloseq(ps.contr, biom_file = file.path(path,
        "just_controls.biom"), biom_format = "standard"))
}

ps
## phyloseq-class experiment-level object
## otu_table()   OTU Table:         [ 1821 taxa and 18 samples ]
## sample_data() Sample Data:       [ 18 samples by 15 sample variables ]
## tax_table()   Taxonomy Table:    [ 1821 taxa by 8 taxonomic ranks ]
## refseq()      DNAStringSet:      [ 1821 reference sequences ]

Output

The chunks below will produce various plots and other output. Each chunk is headed by a description of the output and may contain some parameters to adjust the output.

Plot looks

This chunk sets the background structure and color palette. Viridis was chosen because it is optimized for grey-scale printing and various types of color blindness and More info on the Viridis palette can be found on the Viridis info page. It also establishes save_plot as a shorter variant of ggsave with customized date-time structure to save plots with the same name mulitple times instead of overwriting them (overwriting can be triggered with overwrite=TRUE).

##### Optional settings (sensible defaults) #####
# Can be changed to adjust the output format for all plots. Default "pdf", 
# possible "eps"/"ps", "tex" (pictex), "jpeg", "tiff", "png", "bmp" and "svg"
parameters$output_format = "pdf"

# Can be changed to preferred ggplot2 theme. Recommended: "theme_bw()".
theme_set(theme_bw())

############### NO NEED FOR CHANGES BELOW ###############

my_scale_col <- scale_color_viridis(discrete = TRUE)
my_scale_fill <- scale_fill_viridis(discrete = TRUE)

# Custom, more narrow color ranges based on viridis
# Base order to have adjacent colors be distinct from each other
tmp$sort_colors <- c(rbind(c(1:5), c(6:10), c(11:15), c(16:20)))

# Customized vectors
tmp$n_col <- 20
tmp$viridis_greens <- viridis(tmp$n_col,  option = "D", begin = 0.85, 
                              end = 0.7)[tmp$sort_colors]
tmp$viridis_reds <- viridis(tmp$n_col,  option = "B", begin = 0.7, 
                            end = 0.5)[tmp$sort_colors]
tmp$viridis_blues <- viridis(tmp$n_col,  option = "D", begin = 0.2, 
                             end = 0.4)[tmp$sort_colors]
tmp$viridis_yellows <- viridis(tmp$n_col,  option = "D", begin = 1, 
                               end = 0.9)[tmp$sort_colors]
tmp$viridis_dark <- viridis(tmp$n_col,  option = "A", begin = 0, 
                            end = 0.1)[tmp$sort_colors]
tmp$viridis_light <- viridis(tmp$n_col,  option = "A", begin = 1, 
                             end = 0.9)[tmp$sort_colors]
# Collected list that is available in the global environment
sub_viridis <- list(tmp$viridis_greens, tmp$viridis_blues, tmp$viridis_yellows, 
                    tmp$viridis_light, tmp$viridis_reds, tmp$viridis_dark)
names(sub_viridis) <- c("greens", "blues", "yellows", "lights", "reds", "darks")

tmp$out <- paste0(".", parameters$output_format)

#################### Function ############################

# Generic save function for plots that checks whether file exists and if so, 
# creates a new one with d/m/y+time info to avoid overwriting. Overwriting can 
# be triggered with overwrite = TRUE. Width, height and resolution are taken 
# from parameters in the 'set' environment or set to 20x20 cm with 300dpi.
save_plot <- function(
  pl, filetype = ".pdf", plot_name = "my_plot", overwrite=FALSE){
  wp <- if (!is.null(set$wp)) set$wp else 20
  hp <- if (!is.null(set$hp)) set$hp else 20
  res <- if (!is.null(set$res)) set$res else 300
  name <- paste0("/", plot_name,filetype)
  if (file.exists(paste0(outp, name)) & !overwrite) {
  name <- paste0("/", plot_name, "_", 
                 format(Sys.time(), "%d-%m-%y_%H%M%S"),filetype)}
  ggsave(file.path(outp, name), pl, 
         width = wp, height = hp, unit = "cm", dpi = res)
}

################################################

Total ASV counts ranked

This chunk plots the absolute abundance of all samples (including controls) and all samples without controls and other trimmed samples.

# CHANGE ME to the sample group for color coding. Accepted values are the column 
# headers in the descriptor file.
set$color_by = "mel_mol_asv"

##### Optional settings (sensible defaults) #####

# Can be changed to change the width (in cm) of the saved plot.
set$wp = 17
# Can be changed to change the height (in cm) of the saved plot.
set$hp = 20  
# Can be changed to change the resolution (in dpi) of the saved plot.
set$res = 300

############### NO NEED FOR CHANGES BELOW ###############
# Rank samples
set$ranked <- cuphyr::make_ranked_sums(p, myset = tmp$subset_id)
set$ranked_ps <- cuphyr::make_ranked_sums(ps, myset = tmp$subset_id)
set$ymax <- max(set$ranked$Abundance)
set$ymax <- set$ymax + round(set$ymax/10)
set$xmax <- nrow(set$ranked) + 1
set$title2 <- "Samples (without controls)"

# Stabilize colors
set$color_vars <- set$ranked[,set$color_by]  %>% 
  unlist() %>% as.character() %>% unique()
set$color_vars <- sort(set$color_vars)
set$color_varsPalette <- viridis(length(set$color_vars))
names(set$color_varsPalette) <- set$color_vars
set$my_scale_fill <- scale_fill_manual(values = set$color_varsPalette)

# plot
plots$overview_all <- ggplot(data = set$ranked, aes(x = Rank, y = Abundance)) + 
  aes_string(fill = set$color_by) + 
  geom_col() + set$my_scale_fill + ggtitle("All samples") + ylim(0, set$ymax) + 
  xlim(0,set$xmax) + ylab("ASV counts ('reads')")

if (length(tmp$noise_ASVs) > 0) {
  set$ranked_nonoise <- cuphyr::make_ranked_sums(
    tmp$no_noise_ps, myset = tmp$subset_id)
  plots$overview_noise <- ggplot(
    data = set$ranked_nonoise, aes(x = Rank, y = Abundance)) + 
  aes_string(fill = set$color_by) + 
  geom_col() + set$my_scale_fill + 
    ggtitle("Samples (without controls), noise ASVs removed") + 
    ylim(0, set$ymax) + 
    xlim(0,set$xmax) + ylab("ASV counts ('reads')")
}

if (parameters$minASVcount > 0) {
plots$overview_all <- plots$overview_all + 
  geom_hline(yintercept = parameters$minASVcount, linetype = "dashed") + 
    ggtitle("All samples (ASV count cutoff indicated)")
set$title2 <- "Samples (without controls and low count samps)"
}

plots$overview_ps <- ggplot(data = set$ranked_ps, aes(x = Rank, y = Abundance)) + 
  aes_string(fill = set$color_by) + 
  geom_col() + set$my_scale_fill + ggtitle(set$title2) + ylim(0, set$ymax) + 
  xlim(0,set$xmax) + ylab("ASV counts ('reads')")
plots$combo_overview <- ggarrange(
  plots$overview_all, plots$overview_ps, nrow = 2, align = "v", 
  common.legend = TRUE, legend = "right")

if (parameters$minASVcount > 0) {
plots$combo_overview <- ggarrange(
  plots$overview_all, plots$overview_noise, plots$overview_ps,
  nrow = 3, align = "v", 
  common.legend = TRUE, legend = "right")
}

#Save plots
save_plot(plots$combo_overview, plot_name = "Overview_all_and_pruned", 
          filetype = tmp$out)

#Clean up plot parameters
rm(list = ls(set), envir = set)

#Print plots
plots$combo_overview

Controls

This chunk generates an overview over the controls (positive AND negative)

# CHANGE ME to the desired sample categories on the x-axis. In this case it 
# should be the Sample names.
set$x_axis_value = "SampleIDs"

# CHANGE ME to the taxonomic level for color coding. Use "OTU" for ASVs, 
# "Genus", "Species" or "OTU" recommended to compare pos. controls.
set$color_by_taxlvl = "Species"

# CHANGE ME to the taxonomic level for labeling the tree tips (if phylogenetic 
# tree is available). Use "OTU" for ASVs.
set$label_by_taxlvl = "OTU"

# CHANGE ME to a sample category to shape the tree tip labels by (if 
# phylogenetic tree is available).
set$label_shape_by = "Symptoms"

##### Optional settings (sensible defaults) #####

# Can be changed to generate a tree for just the control sequences IF no 
# phylogenetic tree for all seuquences is provided. This may slow down this 
# chunk when running it for the first time
set$control_tree = TRUE

# Can be changed to change the width (in cm) of the saved plot.
set$wp = 17
# Can be changed to change the height (in cm) of the saved plot.
set$hp = 20  
# Can be changed to change the resolution (in dpi) of the saved plot.
set$res = 300

############### NO NEED FOR CHANGES BELOW ###############
if (set$control_tree & class(try(phy_tree(ps.transcontr), 
                                 silent = TRUE)) == "try-error") {
  # generate phylogenetic tree of ASVs only if there is no file called 
  # 'phylotree.rds' in the working directory and 'parameters$maketree' is "TRUE"
  if (!file.exists(paste0(path, "/controls_phylotree.rds"))) {
    set$ASVs <- phyloseq::refseq(ps.transcontr)
    set$ASV_align <- AlignSeqs(set$ASVs, anchor = NA)
    set$ASV_phang <- phyDat(as(set$ASV_align, "matrix"), type = "DNA")
    set$dm <- dist.ml(set$ASV_phang)
    set$treeNJ <- NJ(set$dm)
    set$fit <- pml(set$treeNJ, data = set$ASV_phang)
    set$fitGTR <- update(set$fit, k = 4, inv = 0.2)
    set$fitGTR <- optim.pml(set$fitGTR, model = "GTR", 
                            optInv = TRUE, optGamma = TRUE,
                            rearrangement = "stochastic", 
                            control = pml.control(trace = 0))
    saveRDS(set$fitGTR, file = paste0(path, "/controls_phylotree.rds"))}
  set$fitGTR <- readRDS(paste0(path, "/controls_phylotree.rds"))
  phyloseq::phy_tree(ps.transcontr) <- set$fitGTR$tree
}

plots$topnpplot <- plot_bar(ps.contr, x = set$x_axis_value, 
                            fill = set$color_by_taxlvl) + my_scale_fill + 
  theme(axis.title.x = element_blank(), legend.position = "none", 
        legend.key.size = unit(3, "mm")) + 
  ylab("ASV counts") + guides(col = guide_legend(ncol = 3))

plots$topntplot <- plot_bar(ps.transcontr, x = set$x_axis_value, 
                            fill = set$color_by_taxlvl) + my_scale_fill + 
  theme(axis.title.x = element_blank(), legend.position = "none", 
        legend.key.size = unit(3, "mm")) + 
  ylab("Relative abundance") + guides(col = guide_legend(ncol = 3))

plots$combo_contr <- ggarrange(plots$topnpplot, plots$topntplot, ncol = 2, 
                               labels = c("A", "B"), align = "hv", 
                               common.legend = TRUE, legend = "right")

if (class(try(phy_tree(ps.transcontr), silent = TRUE)) != "try-error") {
plots$tre <- plot_tree(
          ps.transcontr, ladderize = "left", label.tips = set$label_by_taxlvl, 
          color = "abundance", text.size = 2.5, shape = set$label_shape_by) + 
          scale_color_viridis_c(aesthetics = c("color","fill")) + 
          theme(legend.position = "left", panel.border = element_blank())
plots$combo_contr <- ggarrange(plots$tre, ggarrange(plots$topnpplot, 
                                                    plots$topntplot, ncol = 2, 
                               labels = c("B", "C"), align = "hv", 
                               common.legend = TRUE, legend = "right"), 
                               nrow = 2, legend = "right", labels = c("A")) 
}

# save
save_plot(plots$combo_contr, plot_name = "Controls", filetype = tmp$out)

plots$combo_contr

Get a list of Top N taxa at a given level

This chunk lists the top n most abundant taxonomic terms at a given level. Change the function parameters to the desired values. For more info, check help page of the function with ?cuphyr::abundant_tax_physeq(). Change ‘ignore_na’ to include/exclude NA values at the given level.

#The character vector can later be accessed by calling 'tmp$tops'
tmp$tops <- cuphyr::abundant_tax_physeq(physeq = ps, 
                            lvl = "Superkingdom",
                            top = 10,
                            output_format = "tops",
                            ignore_na = TRUE,
                            silent = FALSE)
## 
## The top 10 most abundant annotated groups at the taxonomic level 'Superkingdom' are:
## Bacteria
## Archaea
## Eukaryota

ASVs belonging to phylum

This chunk gives an overview of the number of ASVs belonging to which Phylum

ps_trans_tbl %>% select(OTU, Superkingdom) %>% unique() %>% group_by(Superkingdom) %>% summarise(length(OTU))
## # A tibble: 4 × 2
##   Superkingdom `length(OTU)`
##   <chr>                <int>
## 1 Archaea                  6
## 2 Bacteria              1700
## 3 Eukaryota               54
## 4 <NA>                    61
# Total ASV counts

ps_tbl %>% select(OTU, Abundance) %>% summarise(total_sum = sum(Abundance))
## # A tibble: 1 × 1
##   total_sum
##       <int>
## 1    209313
# Total ASV counts per class 

ps_tbl %>% select(OTU, Superkingdom, Abundance) %>% group_by(Superkingdom) %>% summarise(Abundance_sum = sum(Abundance))
## # A tibble: 4 × 2
##   Superkingdom Abundance_sum
##   <chr>                <int>
## 1 Archaea               4091
## 2 Bacteria            201304
## 3 Eukaryota             1822
## 4 <NA>                  2096
# Percentage of ASV counts belonging to Bacteria

bacteria_percentage = (201304 /209313)*100 # ASV counts Bacteria / Total ASV counts
bacteria_percentage
## [1] 96.17367

Top N ASVs/taxa Bar plot

This chunk plots abundance of the Top n ASVs or taxa at a given level as a bar plot, giving an insight into the presence of the n ASV and most common taxa for the primary and secondary parameters. The default for n is set at 20, a larger n may lead to delay/skipping of the plot in standard out, but it should be saved as a PDF regardless for ASVs. For taxa, a large n may lead to unreadable plots. The chunk does not require any input, but it is possible to adjust the default ‘n’, and to change width, height and resolution of the PDF-output if necessary.

# CHANGE ME to the sample category that will be shown in separate panels.
# Accepted values are the column headers in your descriptor file.
set$panel_by = "Symptoms"

# CHANGE ME to the desired sample categories on the x-axis.  Accepted values
# are the column headers in the descriptor file.
set$x_axis_value = "Alias"

# CHANGE ME to the count of top ASVs you want to plot (e.g. 'set$topASVs = 20'
# plots the 20 most abundant ASVs)
set$topASVs = 100

# CHANGE ME to the taxonomic level of interest (color coding). Accepted values
# are the column headers in your descriptor file.
set$taxlvl = "Genus"

# CHANGE ME to change the number of Top n taxa to be plotted at taxlvl.
set$top_n = 100

# CHANGE ME to an entry at the chosen taxonomic level you want to highlight.
# Comment out to not highlight anything.
set$highlight = "Arthrobotryss"

##### Optional settings (sensible defaults) #####

# Can be changed to turn unified coloring on or off (same taxonomy term = same
# color in both plots). Highlighting will unify colors even if unify_colors is
# FALSE.
set$unify_colors = TRUE

# Can be changed to include (FALSE) or exclude (TRUE) NA values in the barplot
set$ignore_na = TRUE

# Can be changed to remove ASV segmentation in the top n taxlvl plot. This
# improves visual clarity when a bar segment appears black due to the border of
# many small ASVs overlapping.
set$fuse_ASVs = TRUE

# Can be changed to change the width (in cm) of the saved plot.
set$wp = 40
# Can be changed to change the height (in cm) of the saved plot.
set$hp = 20
# Can be changed to change the resolution (in dpi) of the saved plot.
set$res = 300

# Can be changed to change the y-axis label
set$y_axis_label = "Relative abundance"

############### NO NEED FOR CHANGES BELOW ###############

# Make physeq objects of top n taxa and top n ASVs
set$ps.topnTax <- cuphyr::abundant_tax_physeq(ps.trans, lvl = set$taxlvl, top = set$top_n,
    ignore_na = set$ignore_na)
set$topnASVs <- names(sort(taxa_sums(ps), decreasing = TRUE))[1:set$topASVs]
set$ps.topnASVs <- prune_taxa(set$topnASVs, ps.trans)

if (set$unify_colors | exists("highlight", envir = set) | set$fuse_ASVs) {
    set$toptax <- union(phyloseq::tax_table(set$ps.topnTax)[, set$taxlvl], phyloseq::tax_table(set$ps.topnASVs)[,
        set$taxlvl])
    set$toptax <- sort(set$toptax)
    set$taxlvlPalette <- viridis(length(set$toptax))
    names(set$taxlvlPalette) <- set$toptax
    if (exists("highlight", envir = set)) {
        # It is possible to change the highlight color here by substituting
        # 'sub_viridis$reds[4]' with a hexcode-string, e.g. '#ff7f7f''
        set$taxlvlPalette[set$highlight] <- sub_viridis$reds[4]
    }
    set$taxlvlPalette <- set$taxlvlPalette[sort(names(set$taxlvlPalette))]
    set$my_scale_fill <- scale_fill_manual(values = set$taxlvlPalette, na.value = "grey")
} else {
    set$my_scale_fill <- my_scale_fill
}

# Plot
if (set$unify_colors | exists("highlight", envir = set) | set$fuse_ASVs) {
    set$my_scale_fill <- scale_fill_manual(values = set$taxlvlPalette[sort(unique(phyloseq::tax_table(set$ps.topnTax)[,
        set$taxlvl]))], na.value = "grey")
}
plots$topn_tax <- plot_bar(set$ps.topnTax, x = set$x_axis_value, fill = set$taxlvl,
    title = paste0("Top", set$top_n, "_", set$taxlvl)) + facet_grid(paste0("~", set$panel_by),
    scales = "free", space = "free") + set$my_scale_fill + ylab(set$y_axis_label)

if (set$fuse_ASVs) {
    plots$topn_tax <- plots$topn_tax + geom_bar(aes_string(color = set$taxlvl, fill = set$taxlvl),
        stat = "identity", position = "stack") + scale_color_manual(values = set$taxlvlPalette,
        na.value = NA)
}

if (set$unify_colors | exists("highlight", envir = set) | set$fuse_ASVs) {
    set$my_scale_fill <- scale_fill_manual(values = set$taxlvlPalette[sort(unique(phyloseq::tax_table(set$ps.topnASVs)[,
        set$taxlvl]))], na.value = "grey")
}
plots$topn_ASVs <- plot_bar(set$ps.topnASVs, x = set$x_axis_value, fill = set$taxlvl,
    title = paste0("Top", set$topASVs, "_ASVs")) + facet_wrap(paste0("~", set$panel_by),
    scales = "free_x") + set$my_scale_fill + ylab(set$y_axis_label)

# save
save_plot(plots$topn_tax, plot_name = paste0("Top", set$top_n, "_", set$taxlvl),
    filetype = tmp$out)
save_plot(plots$topn_ASVs, plot_name = paste0("Top", set$topASVs, "_ASVs"), filetype = tmp$out)

# Clean up plot parameters
rm(list = ls(set), envir = set)

# Print to standard out
plots$topn_tax

plots$topn_ASVs

#### Customized Top N Genus Bar plot

This chunk plots abundance of the Top n ASVs or taxa at a given level as a bar plot, giving an insight into the presence of the n ASV and most common taxa for the primary and secondary parameters. The default for n is set at 20, a larger n may lead to delay/skipping of the plot in standard out, but it should be saved as a PDF regardless for ASVs. For taxa, a large n may lead to unreadable plots. The chunk does not require any input, but it is possible to adjust the default ‘n’, and to change width, height and resolution of the PDF-output if necessary.

# CHANGE ME to the desired sample categories on the x-axis. 
# Accepted values are the column headers in the descriptor file.
set$x_axis_value = "ndvi"

# CHANGE ME to the taxonomic level of interest (color coding). Accepted values 
# are the column headers in your descriptor file.
set$taxlvl = "Genus"

# CHANGE ME to change the number of Top n taxa to be plotted at 
# taxlvl.
set$top_n = 10

# Can be changed to include (FALSE) or exclude (TRUE) NA values in the barplot
set$ignore_na = TRUE

# CHANGE ME to an entry at the chosen taxonomic level you want to highlight. 
# Comment out to not highlight anything.
#set$highlight = "Meloidogyne"

##### Optional settings (sensible defaults) #####

# Can be changed to change the width (in cm) of the saved plot.
set$wp = 20
# Can be changed to change the height (in cm) of the saved plot.
set$hp = 13
# Can be changed to change the resolution (in dpi) of the saved plot.
set$res = 300

# Can be changed to change the y-axis label
set$y_axis_label = "Relative abundance"

# Can be changed to change the x-axis label
set$x_axis_label = "Sample"

############### NO NEED FOR CHANGES BELOW ###############
# Estimate Alpha-diversity (Shannon)
set$alpha_div_ps_trans <- estimate_richness(ps.trans, measures = "Shannon") %>%
  as_tibble(rownames = "Sample")

# Make physeq objects of top n taxa and top n ASVs
set$ps.topnTax <- cuphyr::abundant_tax_physeq(ps.trans, lvl = set$taxlvl, 
                                              top = set$top_n, 
                                              ignore_na = set$ignore_na)

# Plot
set$my_scale_fill <- my_scale_fill


set$topntax_tbl <- psmelt(set$ps.topnTax) %>% 
                    as_tibble() %>%
                    left_join(set$alpha_div_ps_trans, by = "Sample") %>% 
                    select(Genus, Alias, ndvi, Abundance, Shannon) %>%
                    filter(Abundance > 0) %>%
                    group_by(Genus, Alias, ndvi, Shannon) %>%
                    summarise(Abundance = sum(Abundance)) %>%
                    arrange(ndvi) %>%
                    mutate(ndvi_rank = c(1:length(ndvi))) 
                    

plots$topn_tax_custom <- ggplot(set$topntax_tbl, aes(x = fct_reorder(Alias, ndvi),
                           y = Abundance,
                           fill = Genus)) + 
                           #title = paste0("Top ", set$top_n, " ", set$taxlvl))) +
  geom_col(color = "black") +
  scale_fill_viridis(discrete = TRUE, na.value = "grey")  +
  ylab(set$y_axis_label) +
  xlab(set$x_axis_label) +
  theme(strip.background = element_blank(),
        # strip.text = element_text(size = 16),
       # axis.title=element_text(size=16),
       # legend.text = element_text(size=14), 
        legend.position = "bottom")

plots$ndvi_dot_plot <- ggplot(set$topntax_tbl, aes(fct_reorder(Alias, ndvi),
                           y = ndvi)) +
                             geom_point() +
    theme(strip.background = element_blank(),
          #strip.text = element_text(size = 16),
        # axis.title=element_text(size=16),
       # legend.text = element_text(size=14)
        axis.title.x=element_blank()) +
  ylab("NDVI")

plots$shannon_dot_plot <- ggplot(set$topntax_tbl, 
                                 aes(fct_reorder(Alias, ndvi),
                           y = Shannon)) +
                             geom_point() +
    theme(strip.background = element_blank(),
          #strip.text = element_text(size = 16),
        #axis.title=element_text(size=16),
        #legend.text = element_text(size=14),
        axis.title.x=element_blank()) +
  ylab("Shannon")


plots$combo_topn_custom <- ggarrange(plots$ndvi_dot_plot,
                                     plots$shannon_dot_plot,
                                     plots$topn_tax_custom,
                                     nrow = 3,
                                     heights = c(1, 1, 3),
                                     align = "v") 

save_plot(plots$combo_topn_custom, plot_name = paste0("Customized_NDVI_Shannon_plot"), 
          filetype = tmp$out)
plots$combo_topn_custom

NDVI+Shannon regression

topntax_data <- set$topntax_tbl %>%
  mutate(Taxa = 'bacteria') %>%
  ungroup() %>%
  select(Alias, ndvi, Shannon, Taxa) %>%
  distinct()

write.csv(topntax_data, file = "../topntax_all_taxa/topntax_data_bacteria.csv")

Abuandance tables per genus

ps_trans_tbl <- as_tibble(psmelt(ps.trans))

# Genera of interest

genera_of_interest <- c("Bacillus", "Paenibacillus", "Pseudarthrobacter", "Bradyrhizobium", "Sporosarcina", "Clostridium", "Candidatus Nitrosocosmicus", "Pseudomonas", "Rhizobium")
#genera_of_interest <- c("Globodera, Meloidogyne, Pristionchus, Rhabditis, Pellioditis, Cephaloboides, Aporcelaimellus, Acrobeloides, Nygolaimus, Cruznemas")

# Taking ps_tbl, grouping by Genus and summarizing (open heart surgery)
# Will give a table with two columns: "Genus" "Genus_total_Abundance"

genus_abundance_tbl <- ps_trans_tbl %>%
  group_by(Genus) %>%
  summarise(Genus_total_Abundance = sum(Abundance))


# If you want per genus and sample
#genus_abundance_tbl_per_sample <- ps_trans_tbl %>%
#  group_by(Genus, Sample) %>%
#  summarise(Genus_Sample_Abundance = sum(Abundance)) %>%
#  filter(Genus %in% genera_of_interest) %>%
#  group_by(Sample)
  

# To get back all the other info, one way is to fuse back to original table
ps_tbl_with_genus_abundance <- left_join(ps_trans_tbl, genus_abundance_tbl_per_sample,
                                         by = c("Genus", "Sample"))

library(readxl)

# Can be made more compact by removing ASV+Species info and original abundance 
# values and then reducing to unique rows
genus_abundance_tbl_full_info <- ps_tbl_with_genus_abundance %>%
  select(-OTU, -Species) %>%
  unique()



write.xlsx(genus_abundance_tbl_per_sample, "genus_abundance_tbl_per_sample.xlsx")

# ndvi <- read_excel("~/Documents/GitHub/metamarte/Data_analysis/2021-patch-metabarcoding/16S/genus_abundance_tbl_per_sample_copy.xlsx")
# 
# 

NDVI regression

# Create properly formatted tibble with columns Sample, ndvi_01 (ndvi translated to (0, 1) interval) and one column for each genus
# containing the sample abundances for that genus.
ldf <- data.frame(genus_abundance_tbl_per_sample %>% pivot_wider(id_cols = c('Sample', 'ndvi'), names_from = 'Genus', values_from = 'Genus_Sample_Abundance'))
ldf_genus_data <- data.frame(ldf) %>% select(!c('Sample', 'ndvi'))
colnames(ldf_genus_data) <- gsub(' ', '.', colnames(ldf_genus_data))
ldf <- cbind.data.frame(
Sample = ldf$Sample,
ndvi_01 = (ldf$ndvi + 1) / 2.0
)
ldf <- tibble(cbind(ldf, ldf_genus_data))

n_samples_by_genus <- data.frame(ldf_genus_data > 0) %>% mutate_if(is.logical, as.numeric) %>% colSums() %>% sort(decreasing = TRUE)
keep_n <- 100 # Maximum number of genuses to include in the analysis
top_n_occurence_genuses <- names(n_samples_by_genus[1:keep_n])
top_n_occurence_genuses <- top_n_occurence_genuses[!is.na(top_n_occurence_genuses)]

l_genus_ldf <- ldf %>% select(all_of(top_n_occurence_genuses))
l_genus_ldf_transposed <- data.frame(t(l_genus_ldf))
l_meta_ldf <- ldf %>% select('ndvi_01')

l_model <- linda(
l_genus_ldf_transposed,
l_meta_ldf,
formula = '~ ndvi_01',
feature.dat.type = 'proportion',
is.winsor = FALSE,
alpha = 0.05
)
## 0  features are filtered!
## The filtered data has  18  samples and  100  features will be tested!
## Fit linear models ...
## Completed.
# Print model info
l_model
## $variables
## [1] "ndvi_01"
## 
## $bias
## [1] -0.04378204
## 
## $output
## $output$ndvi_01
##                                                baseMean log2FoldChange
## Priestia                                     78049.6994    0.334652673
## Pseudarthrobacter                            31666.2592    0.030823145
## Niallia                                      60101.5018    0.073418096
## Sporosarcina                                 29509.4485    0.266199202
## Bradyrhizobium                               30782.1044   -0.328557694
## X.uncultured.bacterium.                     381081.5444   -0.132049983
## Sphingomonas                                 26450.4787    0.157003615
## Clostridium                                  23641.1064    0.332666861
## Bacillus                                     13441.1227    0.547619226
## Candidatus.Nitrosocosmicus                   13806.7597    0.020842195
## Methylothermalis                             15788.6684   -0.024516629
## Streptomyces                                 13028.4841    0.122576505
## Dokdonella                                    5774.4672   -0.166993631
## Blastococcus                                  4445.1897   -0.127683267
## Hyphomicrobium                                7093.0167   -0.307211440
## Rhizobium                                     7579.1878    0.152358260
## Nocardioides                                  3899.1685    0.263071602
## X.uncultured.prokaryote.                      8478.7331   -0.454380196
## Terrabacter                                   5571.9424    0.028215564
## Rhynchosporium                                2211.6275   -0.652012714
## Massilia                                      6978.8416    0.495467212
## Pseudomonas                                   5840.7432    0.599878311
## Peribacillus                                 11866.6363    0.009862157
## Paenibacillus                                34156.2013    0.434194618
## Oryzihumus                                    4172.8611   -0.185729107
## Rhodanobacter                                 4258.3998    0.100677226
## Rhodococcus                                   2939.9709    0.454353032
## Spongospora                                   2243.7214    0.053709239
## Devosia                                       5701.0382    0.424713742
## NA.                                           9860.2578   -0.466484116
## X.uncultured.archaeon.                        4658.1870    0.062000915
## Pedobacter                                    4329.1638    0.664781461
## Nakamurella                                   4597.4502   -0.166262696
## Chujaibacter                                  3795.0861   -0.445739478
## Variovorax                                    4404.5287   -0.245112454
## Caldibacillus                                 3168.3360   -0.038538194
## Ureibacillus                                  3376.2083    0.067406164
## Terribacillus                                 2094.7482    0.138993392
## Streptosporangium                             2832.0439   -0.073517090
## Pantoea                                       3650.8917    0.726879984
## Flavobacterium                                2387.4575    1.036398785
## Piscinibacter                                 3638.5694   -0.364288408
## X.uncultured.Acidobacteriota.bacterium.       3941.5511   -0.348604273
## Candidatus.Nitrosotalea                       3205.7450   -0.352828729
## Ammoniphilus                                  1683.9409    0.140682569
## Paraburkholderia                              4960.3777   -0.475877870
## Ramlibacter                                   4240.8880    0.530976364
## Mesorhizobium                                 1820.4669   -0.056021933
## Burkholderia                                  3673.0033   -0.529319308
## Caulobacter                                   1734.3867    0.718574929
## X.uncultured.Alphaproteobacteria.bacterium.   2477.5985   -0.421474190
## Nitrospira                                    2145.4208   -0.375483878
## Cohnella                                      2646.8579    0.354509182
## Tumebacillus                                  1580.2807   -0.453930174
## Pedomicrobium                                 2259.9326   -0.089177426
## Niastella                                     1809.1890   -0.351904774
## Shouchella                                    2312.7543    0.298871498
## Stenotrophobacter                             1737.9497    0.276765787
## Geomonas                                       792.4044   -0.262846179
## X.uncultured.Actinomycetes.bacterium.         2042.0359    0.020746893
## Modestobacter                                 2056.1429    0.077847212
## Kaistia                                       1362.4533   -0.372114503
## Arenimonas                                    1103.5320    0.137270130
## Ancylobacter                                  1174.6082    0.059524623
## Chryseobacterium                              1048.8562    1.073586715
## Aetherobacter                                 3673.5038   -0.867262707
## Scopulibacillus                               2037.3717    1.131439742
## Pullulanibacillus                             2622.8216    0.312276978
## Dyella                                        1671.5356   -0.387337917
## Anaerocolumna                                 1289.7446    0.334083197
## Psychrobacillus                               1903.5538    0.091821753
## Phenylobacterium                              1647.1233   -0.201908701
## Reyranella                                    1238.2884   -0.025778960
## Methylobacterium                              1011.0328   -0.318069979
## Asticcacaulis                                  504.8360    0.348204585
## Agrobacterium                                 2761.7554    1.266671269
## Simulacricoccus                               3333.3421   -0.307688817
## Cystobacter                                   1251.6465   -0.421994706
## Candidatus.Solibacter                         1632.1269   -0.394583822
## Achromobacter                                  660.4535    0.879371290
## Stenotrophomonas                              1694.5995    0.892202107
## Mycolicibacterium                             3537.9957    0.167251458
## Noviherbaspirillum                            1820.3628   -0.699193274
## Acidovorax                                    1105.6643    0.041129002
## Paenarthrobacter                               970.8077    0.299985622
## Methylophilus                                 1021.8124    0.730547485
## Exophiala                                      297.8257   -0.145572163
## Knufia                                         215.5495   -0.190402634
## Planococcus                                   4618.8831   -0.043269572
## Chitinophaga                                  1089.1572    0.442934545
## X.uncultured.Acetobacteraceae.bacterium.      1823.2487   -0.317514678
## Geothrix                                      1146.2527   -0.488093558
## Ralstonia                                      679.8978   -1.278911575
## Lysobacter                                     775.2587    0.362996213
## Lacunisphaera                                  785.4520   -0.110530070
## Nonomuraea                                     661.9492   -0.128963435
## Sphingopyxis                                   833.4281    0.788724544
## Nitrosovibrio                                 1787.3750   -0.617468867
## Brevundimonas                                 1942.9170    0.688896801
## Flavitalea                                     790.1982   -0.475246309
##                                                  lfcSE        stat       pvalue
## Priestia                                    0.14437356  2.31796376 0.0340210880
## Pseudarthrobacter                           0.12853104  0.23981091 0.8135220877
## Niallia                                     0.09868748  0.74394542 0.4676951114
## Sporosarcina                                0.09250933  2.87753885 0.0109389297
## Bradyrhizobium                              0.11324072 -2.90140954 0.0104105037
## X.uncultured.bacterium.                     0.07131760 -1.85157648 0.0826320022
## Sphingomonas                                0.08901313  1.76382529 0.0968433349
## Clostridium                                 0.20061272  1.65825407 0.1167386893
## Bacillus                                    0.15402638  3.55536007 0.0026364873
## Candidatus.Nitrosocosmicus                  0.08603885  0.24224166 0.8116705977
## Methylothermalis                            0.11309809 -0.21677316 0.8311249884
## Streptomyces                                0.14842837  0.82582937 0.4210447820
## Dokdonella                                  0.11122317 -1.50142845 0.1527213199
## Blastococcus                                0.15841949 -0.80598204 0.4320697694
## Hyphomicrobium                              0.16644362 -1.84573875 0.0835166575
## Rhizobium                                   0.09392350  1.62215273 0.1243071431
## Nocardioides                                0.14285593  1.84151688 0.0841616672
## X.uncultured.prokaryote.                    0.28679057 -1.58436241 0.1326749189
## Terrabacter                                 0.09136807  0.30881208 0.7614491683
## Rhynchosporium                              0.22740544 -2.86718164 0.0111762363
## Massilia                                    0.17981672  2.75540130 0.0140768619
## Pseudomonas                                 0.33530666  1.78904383 0.0925540682
## Peribacillus                                0.18388736  0.05363151 0.9578926920
## Paenibacillus                               0.14498861  2.99468082 0.0085740993
## Oryzihumus                                  0.24414215 -0.76074166 0.4578768629
## Rhodanobacter                               0.30408682  0.33108053 0.7448801109
## Rhodococcus                                 0.25520695  1.78033176 0.0940166014
## Spongospora                                 0.26241313  0.20467436 0.8404075382
## Devosia                                     0.12167472  3.49056674 0.0030231834
## NA.                                         0.28012377 -1.66527857 0.1153129927
## X.uncultured.archaeon.                      0.09848836  0.62952530 0.5378913629
## Pedobacter                                  0.38565084  1.72379104 0.1040101415
## Nakamurella                                 0.18901575 -0.87962351 0.3920834033
## Chujaibacter                                0.23346580 -1.90922817 0.0743328912
## Variovorax                                  0.15722390 -1.55900250 0.1385543705
## Caldibacillus                               0.13890311 -0.27744659 0.7849900318
## Ureibacillus                                0.15329171  0.43972477 0.6660180724
## Terribacillus                               0.21496809  0.64657684 0.5270728720
## Streptosporangium                           0.19539260 -0.37625320 0.7116711719
## Pantoea                                     0.42814901  1.69772664 0.1089204850
## Flavobacterium                              0.39810833  2.60330847 0.0192112562
## Piscinibacter                               0.16882348 -2.15780647 0.0464879176
## X.uncultured.Acidobacteriota.bacterium.     0.23539464 -1.48093547 0.1580472761
## Candidatus.Nitrosotalea                     0.21062661 -1.67513840 0.1133371488
## Ammoniphilus                                0.24535619  0.57338097 0.5743581975
## Paraburkholderia                            0.19253941 -2.47158683 0.0250648547
## Ramlibacter                                 0.24510494  2.16632261 0.0457306960
## Mesorhizobium                               0.21024349 -0.26646215 0.7932868152
## Burkholderia                                0.18767783 -2.82036143 0.0123128260
## Caulobacter                                 0.28680257  2.50546891 0.0234151016
## X.uncultured.Alphaproteobacteria.bacterium. 0.43243644 -0.97465004 0.3442402056
## Nitrospira                                  0.32513423 -1.15485804 0.2651055880
## Cohnella                                    0.19384387  1.82883874 0.0861252202
## Tumebacillus                                0.30855212 -1.47116206 0.1606410899
## Pedomicrobium                               0.18843022 -0.47326498 0.6424165678
## Niastella                                   0.24721906 -1.42345327 0.1738126288
## Shouchella                                  0.25048798  1.19315707 0.2502010426
## Stenotrophobacter                           0.29004000  0.95423317 0.3541611190
## Geomonas                                    0.45362931 -0.57942944 0.5703685526
## X.uncultured.Actinomycetes.bacterium.       0.28855571  0.07189909 0.9435733043
## Modestobacter                               0.25308444  0.30759382 0.7623591788
## Kaistia                                     0.17421442 -2.13595691 0.0484838342
## Arenimonas                                  0.33544853  0.40921369 0.6878084590
## Ancylobacter                                0.24841535  0.23961733 0.8136695792
## Chryseobacterium                            0.35032824  3.06451662 0.0074103474
## Aetherobacter                               0.26520003 -3.27022097 0.0048117988
## Scopulibacillus                             0.31648129  3.57506046 0.0025290120
## Pullulanibacillus                           0.22638346  1.37941606 0.1867427988
## Dyella                                      0.23740176 -1.63157138 0.1222931573
## Anaerocolumna                               0.32976244  1.01310265 0.3260874110
## Psychrobacillus                             0.22495218  0.40818343 0.6885493762
## Phenylobacterium                            0.19754394 -1.02209513 0.3219422643
## Reyranella                                  0.31362628 -0.08219643 0.9355100155
## Methylobacterium                            0.30684314 -1.03658822 0.3153411539
## Asticcacaulis                               0.28000540  1.24356381 0.2315731884
## Agrobacterium                               0.32193980  3.93449726 0.0011845138
## Simulacricoccus                             0.16737793 -1.83828783 0.0846579716
## Cystobacter                                 0.32609056 -1.29410280 0.2139981460
## Candidatus.Solibacter                       0.28168117 -1.40081717 0.1803646981
## Achromobacter                               0.42288900  2.07943762 0.0540174213
## Stenotrophomonas                            0.47619010  1.87362592 0.0793652244
## Mycolicibacterium                           0.20800855  0.80406052 0.4331468123
## Noviherbaspirillum                          0.23326280 -2.99744866 0.0085247462
## Acidovorax                                  0.34106168  0.12059110 0.9055162938
## Paenarthrobacter                            0.31651749  0.94776950 0.3573428169
## Methylophilus                               0.31114080  2.34796430 0.0320645979
## Exophiala                                   0.30845243 -0.47194364 0.6433391362
## Knufia                                      0.23748917 -0.80173187 0.4344543333
## Planococcus                                 0.20730658 -0.20872262 0.8372988105
## Chitinophaga                                0.35395286  1.25139418 0.2287787062
## X.uncultured.Acetobacteraceae.bacterium.    0.26456586 -1.20013472 0.2475559221
## Geothrix                                    0.22878810 -2.13338699 0.0487237021
## Ralstonia                                   0.30777642 -4.15532673 0.0007449773
## Lysobacter                                  0.33119970  1.09600404 0.2893003704
## Lacunisphaera                               0.22793681 -0.48491539 0.6343084883
## Nonomuraea                                  0.22956989 -0.56176110 0.5820632095
## Sphingopyxis                                0.32196523  2.44971961 0.0261874452
## Nitrosovibrio                               0.21636832 -2.85378591 0.0114905891
## Brevundimonas                               0.21423098  3.21567308 0.0053969468
## Flavitalea                                  0.26118960 -1.81954530 0.0875901983
##                                                   padj reject df
## Priestia                                    0.15464131  FALSE 16
## Pseudarthrobacter                           0.87491353  FALSE 16
## Niallia                                     0.64067823  FALSE 16
## Sporosarcina                                0.08207564  FALSE 16
## Bradyrhizobium                              0.08207564  FALSE 16
## X.uncultured.bacterium.                     0.25025771  FALSE 16
## Sphingomonas                                0.25485088  FALSE 16
## Clostridium                                 0.27148532  FALSE 16
## Bacillus                                    0.06046367  FALSE 16
## Candidatus.Nitrosocosmicus                  0.87491353  FALSE 16
## Methylothermalis                            0.87542452  FALSE 16
## Streptomyces                                0.61190751  FALSE 16
## Dokdonella                                  0.31816942  FALSE 16
## Blastococcus                                0.61190751  FALSE 16
## Hyphomicrobium                              0.25025771  FALSE 16
## Rhizobium                                   0.27623810  FALSE 16
## Nocardioides                                0.25025771  FALSE 16
## X.uncultured.prokaryote.                    0.28842374  FALSE 16
## Terrabacter                                 0.86631725  FALSE 16
## Rhynchosporium                              0.08207564  FALSE 16
## Massilia                                    0.08798039  FALSE 16
## Pseudomonas                                 0.25409892  FALSE 16
## Peribacillus                                0.95789269  FALSE 16
## Paenibacillus                               0.08207564  FALSE 16
## Oryzihumus                                  0.63594009  FALSE 16
## Rhodanobacter                               0.86613966  FALSE 16
## Rhodococcus                                 0.25409892  FALSE 16
## Spongospora                                 0.87542452  FALSE 16
## Devosia                                     0.06046367  FALSE 16
## NA.                                         0.27148532  FALSE 16
## X.uncultured.archaeon.                      0.71718848  FALSE 16
## Pedobacter                                  0.26669267  FALSE 16
## Nakamurella                                 0.58519911  FALSE 16
## Chujaibacter                                0.25025771  FALSE 16
## Variovorax                                  0.29479653  FALSE 16
## Caldibacillus                               0.87491353  FALSE 16
## Ureibacillus                                0.81221716  FALSE 16
## Terribacillus                               0.71226064  FALSE 16
## Streptosporangium                           0.83726020  FALSE 16
## Pantoea                                     0.27148532  FALSE 16
## Flavobacterium                              0.11300739  FALSE 16
## Piscinibacter                               0.18739885  FALSE 16
## X.uncultured.Acidobacteriota.bacterium.     0.32128218  FALSE 16
## Candidatus.Nitrosotalea                     0.27148532  FALSE 16
## Ammoniphilus                                0.74591974  FALSE 16
## Paraburkholderia                            0.13093723  FALSE 16
## Ramlibacter                                 0.18739885  FALSE 16
## Mesorhizobium                               0.87491353  FALSE 16
## Burkholderia                                0.08208551  FALSE 16
## Caulobacter                                 0.13008390  FALSE 16
## X.uncultured.Alphaproteobacteria.bacterium. 0.53787532  FALSE 16
## Nitrospira                                  0.44933151  FALSE 16
## Cohnella                                    0.25025771  FALSE 16
## Tumebacillus                                0.32128218  FALSE 16
## Pedomicrobium                               0.79424585  FALSE 16
## Niastella                                   0.34080908  FALSE 16
## Shouchella                                  0.43138111  FALSE 16
## Stenotrophobacter                           0.54142851  FALSE 16
## Geomonas                                    0.74591974  FALSE 16
## X.uncultured.Actinomycetes.bacterium.       0.95310435  FALSE 16
## Modestobacter                               0.86631725  FALSE 16
## Kaistia                                     0.18739885  FALSE 16
## Arenimonas                                  0.81970164  FALSE 16
## Ancylobacter                                0.87491353  FALSE 16
## Chryseobacterium                            0.08207564  FALSE 16
## Aetherobacter                               0.07709924  FALSE 16
## Scopulibacillus                             0.06046367  FALSE 16
## Pullulanibacillus                           0.35234490  FALSE 16
## Dyella                                      0.27623810  FALSE 16
## Anaerocolumna                               0.51759907  FALSE 16
## Psychrobacillus                             0.81970164  FALSE 16
## Phenylobacterium                            0.51759907  FALSE 16
## Reyranella                                  0.95310435  FALSE 16
## Methylobacterium                            0.51695271  FALSE 16
## Asticcacaulis                               0.41352355  FALSE 16
## Agrobacterium                               0.05922569  FALSE 16
## Simulacricoccus                             0.25025771  FALSE 16
## Cystobacter                                 0.39629286  FALSE 16
## Candidatus.Solibacter                       0.34685519  FALSE 16
## Achromobacter                               0.20006452  FALSE 16
## Stenotrophomonas                            0.25025771  FALSE 16
## Mycolicibacterium                           0.61190751  FALSE 16
## Noviherbaspirillum                          0.08207564  FALSE 16
## Acidovorax                                  0.93352195  FALSE 16
## Paenarthrobacter                            0.54142851  FALSE 16
## Methylophilus                               0.15268856  FALSE 16
## Exophiala                                   0.79424585  FALSE 16
## Knufia                                      0.61190751  FALSE 16
## Planococcus                                 0.87542452  FALSE 16
## Chitinophaga                                0.41352355  FALSE 16
## X.uncultured.Acetobacteraceae.bacterium.    0.43138111  FALSE 16
## Geothrix                                    0.18739885  FALSE 16
## Ralstonia                                   0.05922569  FALSE 16
## Lysobacter                                  0.48216728  FALSE 16
## Lacunisphaera                               0.79424585  FALSE 16
## Nonomuraea                                  0.74623488  FALSE 16
## Sphingopyxis                                0.13093723  FALSE 16
## Nitrosovibrio                               0.08207564  FALSE 16
## Brevundimonas                               0.07709924  FALSE 16
## Flavitalea                                  0.25025771  FALSE 16
## 
## 
## $covariance
## NULL
## 
## $feature.dat.use
##                                                        1            2
## Priestia                                    1.197372e-01 0.1153150647
## Pseudarthrobacter                           2.373290e-02 0.0374182779
## Niallia                                     5.537678e-02 0.0574488802
## Sporosarcina                                2.909627e-02 0.0324106273
## Bradyrhizobium                              1.260392e-02 0.0207261093
## X.uncultured.bacterium.                     3.572003e-01 0.2200584226
## Sphingomonas                                1.930813e-02 0.0344971484
## Clostridium                                 2.963261e-02 0.0303241063
## Bacillus                                    2.319657e-02 0.0165530672
## Candidatus.Nitrosocosmicus                  1.394476e-02 0.0144665461
## Methylothermalis                            1.769912e-02 0.0144665461
## Streptomyces                                1.273800e-02 0.0139101405
## Dokdonella                                  4.827031e-03 0.0050076506
## Blastococcus                                6.033789e-03 0.0054249548
## Hyphomicrobium                              3.620274e-03 0.0083460843
## Rhizobium                                   4.961116e-03 0.0072332731
## Nocardioides                                2.413516e-03 0.0031993323
## X.uncultured.prokaryote.                    8.715473e-03 0.0052858534
## Terrabacter                                 6.167873e-03 0.0062595632
## Rhynchosporium                              6.704210e-04 0.0018083183
## Massilia                                    7.374631e-03 0.0080678815
## Pseudomonas                                 4.022526e-03 0.0141883433
## Peribacillus                                1.568785e-02 0.0196132981
## Paenibacillus                               4.277286e-02 0.0486854917
## Oryzihumus                                  6.170554e-04 0.0033384337
## Rhodanobacter                               4.827031e-03 0.0020865211
## Rhodococcus                                 4.994007e-04 0.0025038253
## Spongospora                                 3.810976e-04 0.0016692169
## Devosia                                     2.949853e-03 0.0080678815
## NA.                                         1.273800e-02 0.0015301155
## X.uncultured.archaeon.                      5.497452e-03 0.0050076506
## Pedobacter                                  3.620274e-03 0.0148838503
## Nakamurella                                 1.198562e-03 0.0050076506
## Chujaibacter                                4.290695e-03 0.0008989213
## Variovorax                                  6.838294e-03 0.0014536127
## Caldibacillus                               3.083937e-03 0.0047294478
## Ureibacillus                                4.692947e-03 0.0044512450
## Terribacillus                               4.912878e-04 0.0044512450
## Streptosporangium                           1.877179e-03 0.0006222979
## Pantoea                                     1.474926e-03 0.0068159688
## Flavobacterium                              2.011263e-03 0.0180831826
## Piscinibacter                               4.692947e-03 0.0012397335
## X.uncultured.Acidobacteriota.bacterium.     6.972379e-03 0.0009522298
## Candidatus.Nitrosotalea                     8.055854e-04 0.0040339407
## Ammoniphilus                                3.218021e-03 0.0027820281
## Paraburkholderia                            1.573491e-03 0.0070941717
## Ramlibacter                                 3.083937e-03 0.0086242871
## Mesorhizobium                               5.856818e-04 0.0030602309
## Burkholderia                                1.227981e-03 0.0012279810
## Caulobacter                                 3.620274e-03 0.0031993323
## X.uncultured.Alphaproteobacteria.bacterium. 6.436042e-03 0.0038948393
## Nitrospira                                  4.718762e-04 0.0044512450
## Cohnella                                    3.620274e-03 0.0050076506
## Tumebacillus                                3.598848e-04 0.0003598848
## Pedomicrobium                               3.888442e-03 0.0019474197
## Niastella                                   4.156610e-03 0.0012519126
## Shouchella                                  4.290695e-03 0.0050076506
## Stenotrophobacter                           5.095200e-03 0.0026429267
## Geomonas                                    1.206758e-03 0.0001637626
## X.uncultured.Actinomycetes.bacterium.       6.787610e-04 0.0006787610
## Modestobacter                               7.141724e-04 0.0007141724
## Kaistia                                     5.557931e-04 0.0005557931
## Arenimonas                                  2.413516e-03 0.0002992732
## Ancylobacter                                3.967624e-04 0.0003967624
## Chryseobacterium                            2.663471e-04 0.0041730421
## Aetherobacter                               1.112811e-03 0.0022256225
## Scopulibacillus                             4.558863e-03 0.0083460843
## Pullulanibacillus                           4.424779e-03 0.0010847668
## Dyella                                      6.397953e-04 0.0006397953
## Anaerocolumna                               3.620274e-03 0.0057031576
## Psychrobacillus                             8.131231e-04 0.0036166365
## Phenylobacterium                            7.433669e-04 0.0007433669
## Reyranella                                  3.995206e-04 0.0033384337
## Methylobacterium                            3.445187e-04 0.0016692169
## Asticcacaulis                               8.045052e-04 0.0011128112
## Agrobacterium                               7.533080e-04 0.0097370983
## Simulacricoccus                             1.697962e-03 0.0016979624
## Cystobacter                                 4.319388e-04 0.0004319388
## Candidatus.Solibacter                       6.167873e-03 0.0006325742
## Achromobacter                               1.549667e-04 0.0031993323
## Stenotrophomonas                            3.998720e-04 0.0068159688
## Mycolicibacterium                           5.095200e-03 0.0017645492
## Noviherbaspirillum                          7.621951e-04 0.0007621951
## Acidovorax                                  3.995206e-04 0.0003995206
## Paenarthrobacter                            4.010860e-04 0.0004010860
## Methylophilus                               3.598848e-04 0.0003598848
## Exophiala                                   1.190287e-04 0.0001190287
## Knufia                                      9.988014e-05 0.0004173042
## Planococcus                                 8.179136e-03 0.0024805347
## Chitinophaga                                4.261584e-04 0.0043121436
## X.uncultured.Acetobacteraceae.bacterium.    5.229284e-03 0.0008397313
## Geothrix                                    5.504743e-04 0.0005504743
## Ralstonia                                   2.159694e-04 0.0002159694
## Lysobacter                                  3.099334e-04 0.0003099334
## Lacunisphaera                               1.206758e-03 0.0009737098
## Nonomuraea                                  3.393805e-04 0.0003393805
## Sphingopyxis                                3.486750e-04 0.0020865211
## Nitrosovibrio                               9.191826e-04 0.0009191826
## Brevundimonas                               9.833262e-04 0.0043121436
## Flavitalea                                  3.998720e-04 0.0003998720
##                                                        3            4
## Priestia                                    1.029202e-01 0.1059682267
## Pseudarthrobacter                           3.142358e-02 0.0161442679
## Niallia                                     5.372163e-02 0.0532417347
## Sporosarcina                                2.578956e-02 0.0274796050
## Bradyrhizobium                              2.293287e-02 0.0185487334
## X.uncultured.bacterium.                     3.239962e-01 0.2748819236
## Sphingomonas                                2.610697e-02 0.0200944611
## Clostridium                                 3.055071e-02 0.0287677115
## Bacillus                                    1.833042e-02 0.0199227136
## Candidatus.Nitrosocosmicus                  1.190287e-02 0.0131386861
## Methylothermalis                            1.745755e-02 0.0099613568
## Streptomyces                                1.928265e-02 0.0115070846
## Dokdonella                                  4.046977e-03 0.0042078145
## Blastococcus                                2.856689e-03 0.0029197080
## Hyphomicrobium                              2.221870e-03 0.0061829111
## Rhizobium                                   6.983019e-03 0.0097896093
## Nocardioides                                3.967624e-03 0.0024044654
## X.uncultured.prokaryote.                    4.205682e-03 0.0005152426
## Terrabacter                                 5.157911e-03 0.0031773293
## Rhynchosporium                              1.666402e-03 0.0005152426
## Massilia                                    4.602444e-03 0.0037784457
## Pseudomonas                                 1.904460e-03 0.0073851438
## Peribacillus                                7.617838e-03 0.0127093173
## Paenibacillus                               4.062847e-02 0.0439673680
## Oryzihumus                                  3.015394e-03 0.0022327179
## Rhodanobacter                               6.506904e-03 0.0084156290
## Rhodococcus                                 1.190287e-03 0.0038643195
## Spongospora                                 1.031582e-03 0.0015457278
## Devosia                                     3.729567e-03 0.0059252898
## NA.                                         9.681003e-03 0.0071275225
## X.uncultured.archaeon.                      5.634026e-03 0.0054100472
## Pedobacter                                  2.459927e-03 0.0187204809
## Nakamurella                                 3.888272e-03 0.0036066982
## Chujaibacter                                3.332804e-03 0.0031773293
## Variovorax                                  4.761149e-03 0.0037784457
## Caldibacillus                               2.936042e-03 0.0032632031
## Ureibacillus                                3.491509e-03 0.0040360670
## Terribacillus                               3.094747e-03 0.0032632031
## Streptosporangium                           2.697984e-03 0.0043795620
## Pantoea                                     9.522298e-03 0.0054100472
## Flavobacterium                              3.387534e-04 0.0137398025
## Piscinibacter                               3.253452e-03 0.0024903392
## X.uncultured.Acidobacteriota.bacterium.     1.904460e-03 0.0042078145
## Candidatus.Nitrosotalea                     4.523092e-03 0.0052382997
## Ammoniphilus                                2.618632e-03 0.0015457278
## Paraburkholderia                            3.729567e-03 0.0015734913
## Ramlibacter                                 4.919854e-03 0.0046371833
## Mesorhizobium                               1.507697e-03 0.0012022327
## Burkholderia                                4.523092e-03 0.0012279810
## Caulobacter                                 1.745755e-03 0.0030914556
## X.uncultured.Alphaproteobacteria.bacterium. 6.268846e-03 0.0004134224
## Nitrospira                                  4.602444e-03 0.0036066982
## Cohnella                                    3.015394e-03 0.0049806784
## Tumebacillus                                1.190287e-03 0.0030055818
## Pedomicrobium                               3.412157e-03 0.0024044654
## Niastella                                   1.428345e-03 0.0014598540
## Shouchella                                  8.005489e-04 0.0041219407
## Stenotrophobacter                           1.825107e-03 0.0033490769
## Geomonas                                    3.253452e-03 0.0024903392
## X.uncultured.Actinomycetes.bacterium.       2.063165e-03 0.0015457278
## Modestobacter                               1.428345e-03 0.0034349506
## Kaistia                                     5.557931e-04 0.0018892228
## Arenimonas                                  1.825107e-03 0.0018033491
## Ancylobacter                                7.935248e-04 0.0026620867
## Chryseobacterium                            7.141724e-04 0.0029197080
## Aetherobacter                               1.112811e-03 0.0011128112
## Scopulibacillus                             4.761149e-03 0.0053241735
## Pullulanibacillus                           3.808919e-03 0.0039501932
## Dyella                                      1.587050e-03 0.0006397953
## Anaerocolumna                               3.094747e-03 0.0003995206
## Psychrobacillus                             2.063165e-03 0.0008131231
## Phenylobacterium                            2.221870e-03 0.0021468441
## Reyranella                                  2.459927e-03 0.0026620867
## Methylobacterium                            8.728773e-04 0.0003445187
## Asticcacaulis                               1.348992e-03 0.0001763047
## Agrobacterium                               7.533080e-04 0.0222413053
## Simulacricoccus                             3.412157e-03 0.0016979624
## Cystobacter                                 4.319388e-04 0.0004319388
## Candidatus.Solibacter                       6.325742e-04 0.0006325742
## Achromobacter                               1.549667e-04 0.0038643195
## Stenotrophomonas                            3.998720e-04 0.0123658222
## Mycolicibacterium                           1.764549e-03 0.0052382997
## Noviherbaspirillum                          7.621951e-04 0.0007621951
## Acidovorax                                  1.825107e-03 0.0003995206
## Paenarthrobacter                            2.459927e-03 0.0004010860
## Methylophilus                               3.598848e-04 0.0023185917
## Exophiala                                   2.380575e-04 0.0005152426
## Knufia                                      9.988014e-05 0.0002576213
## Planococcus                                 6.427551e-03 0.0024805347
## Chitinophaga                                4.261584e-04 0.0106483469
## X.uncultured.Acetobacteraceae.bacterium.    2.380575e-03 0.0008397313
## Geothrix                                    5.504743e-04 0.0005504743
## Ralstonia                                   2.159694e-04 0.0002159694
## Lysobacter                                  1.904460e-03 0.0003099334
## Lacunisphaera                               4.234417e-04 0.0011163590
## Nonomuraea                                  1.269640e-03 0.0003393805
## Sphingopyxis                                3.486750e-04 0.0023185917
## Nitrosovibrio                               9.191826e-04 0.0009191826
## Brevundimonas                               9.833262e-04 0.0035208244
## Flavitalea                                  3.998720e-04 0.0003998720
##                                                        5            6
## Priestia                                    0.1017953930 0.0956185910
## Pseudarthrobacter                           0.0292174797 0.0207084832
## Niallia                                     0.0470867209 0.0484751631
## Sporosarcina                                0.0321815718 0.0299640431
## Bradyrhizobium                              0.0233739837 0.0283659608
## X.uncultured.bacterium.                     0.3295223577 0.3575043281
## Sphingomonas                                0.0215955285 0.0261020109
## Clostridium                                 0.0220189702 0.0199094420
## Bacillus                                    0.0141429539 0.0159808230
## Candidatus.Nitrosocosmicus                  0.0116869919 0.0135836996
## Methylothermalis                            0.0147357724 0.0179118391
## Streptomyces                                0.0204098916 0.0144493275
## Dokdonella                                  0.0044884824 0.0041283793
## Blastococcus                                0.0033028455 0.0025968837
## Hyphomicrobium                              0.0055894309 0.0091223865
## Rhizobium                                   0.0057588076 0.0041283793
## Nocardioides                                0.0038956640 0.0060593954
## X.uncultured.prokaryote.                    0.0028794038 0.0086562791
## Terrabacter                                 0.0045731707 0.0038620322
## Rhynchosporium                              0.0007621951 0.0009988014
## Massilia                                    0.0077913279 0.0049274204
## Pseudomonas                                 0.0027100271 0.0014649088
## Peribacillus                                0.0129573171 0.0099214276
## Paenibacillus                               0.0452235772 0.0303635637
## Oryzihumus                                  0.0021172087 0.0038620322
## Rhodanobacter                               0.0019478320 0.0032627514
## Rhodococcus                                 0.0022018970 0.0009988014
## Spongospora                                 0.0007621951 0.0028632308
## Devosia                                     0.0038109756 0.0038620322
## NA.                                         0.0138042005 0.0092555600
## X.uncultured.archaeon.                      0.0039803523 0.0048608337
## Pedobacter                                  0.0005598209 0.0011985617
## Nakamurella                                 0.0045731707 0.0023971235
## Chujaibacter                                0.0024559621 0.0017978426
## Variovorax                                  0.0037262873 0.0049274204
## Caldibacillus                               0.0029640921 0.0025302970
## Ureibacillus                                0.0025406504 0.0037288587
## Terribacillus                               0.0033028455 0.0023971235
## Streptosporangium                           0.0034722222 0.0034625117
## Pantoea                                     0.0004848378 0.0004848378
## Flavobacterium                              0.0006775068 0.0016646691
## Piscinibacter                               0.0036415989 0.0038620322
## X.uncultured.Acidobacteriota.bacterium.     0.0033875339 0.0054601145
## Candidatus.Nitrosotalea                     0.0037262873 0.0029964043
## Ammoniphilus                                0.0022018970 0.0023305367
## Paraburkholderia                            0.0038956640 0.0045278999
## Ramlibacter                                 0.0021172087 0.0051271807
## Mesorhizobium                               0.0020325203 0.0031295778
## Burkholderia                                0.0024559621 0.0039952058
## Caulobacter                                 0.0022865854 0.0016646691
## X.uncultured.Alphaproteobacteria.bacterium. 0.0055894309 0.0059262219
## Nitrospira                                  0.0043191057 0.0035290984
## Cohnella                                    0.0038109756 0.0027966440
## Tumebacillus                                0.0038109756 0.0003598848
## Pedomicrobium                               0.0022865854 0.0019310161
## Niastella                                   0.0015243902 0.0029964043
## Shouchella                                  0.0052506775 0.0026634705
## Stenotrophobacter                           0.0024559621 0.0020641896
## Geomonas                                    0.0047425474 0.0023971235
## X.uncultured.Actinomycetes.bacterium.       0.0028794038 0.0017978426
## Modestobacter                               0.0022865854 0.0033959249
## Kaistia                                     0.0005557931 0.0011319750
## Arenimonas                                  0.0018631436 0.0015980823
## Ancylobacter                                0.0003967624 0.0013983220
## Chryseobacterium                            0.0002663471 0.0005326941
## Aetherobacter                               0.0011128112 0.0033293381
## Scopulibacillus                             0.0055047425 0.0043947263
## Pullulanibacillus                           0.0050813008 0.0036622719
## Dyella                                      0.0006397953 0.0006397953
## Anaerocolumna                               0.0017784553 0.0007990412
## Psychrobacillus                             0.0022865854 0.0008131231
## Phenylobacterium                            0.0026253388 0.0007433669
## Reyranella                                  0.0019478320 0.0007990412
## Methylobacterium                            0.0007621951 0.0030629911
## Asticcacaulis                               0.0001763047 0.0006658676
## Agrobacterium                               0.0026253388 0.0023305367
## Simulacricoccus                             0.0016979624 0.0033959249
## Cystobacter                                 0.0004319388 0.0013317353
## Candidatus.Solibacter                       0.0022018970 0.0012651485
## Achromobacter                               0.0001549667 0.0001549667
## Stenotrophomonas                            0.0003998720 0.0003998720
## Mycolicibacterium                           0.0017645492 0.0035290984
## Noviherbaspirillum                          0.0015243902 0.0033293381
## Acidovorax                                  0.0003995206 0.0007990412
## Paenarthrobacter                            0.0044037940 0.0017312558
## Methylophilus                               0.0018631436 0.0003598848
## Exophiala                                   0.0001190287 0.0002663471
## Knufia                                      0.0005081301 0.0001997603
## Planococcus                                 0.0024805347 0.0024805347
## Chitinophaga                                0.0011856369 0.0004261584
## X.uncultured.Acetobacteraceae.bacterium.    0.0008397313 0.0008397313
## Geothrix                                    0.0011009485 0.0005504743
## Ralstonia                                   0.0014397019 0.0017978426
## Lysobacter                                  0.0037262873 0.0023305367
## Lacunisphaera                               0.0008468835 0.0004234417
## Nonomuraea                                  0.0021172087 0.0009988014
## Sphingopyxis                                0.0003486750 0.0003486750
## Nitrosovibrio                               0.0009191826 0.0021307764
## Brevundimonas                               0.0009833262 0.0009833262
## Flavitalea                                  0.0017784553 0.0011319750
##                                                        7            8
## Priestia                                    9.791222e-02 8.484512e-02
## Pseudarthrobacter                           1.901743e-02 2.529927e-02
## Niallia                                     6.180666e-02 3.443169e-02
## Sporosarcina                                2.673465e-02 2.215229e-02
## Bradyrhizobium                              1.584786e-02 1.789461e-02
## X.uncultured.bacterium.                     3.164749e-01 2.886585e-01
## Sphingomonas                                1.770826e-02 2.233741e-02
## Clostridium                                 2.260043e-02 5.899050e-02
## Bacillus                                    1.887963e-02 2.721214e-02
## Candidatus.Nitrosocosmicus                  9.922139e-03 8.823892e-03
## Methylothermalis                            1.708813e-02 1.277305e-02
## Streptomyces                                9.439813e-03 1.209429e-02
## Dokdonella                                  2.893957e-03 4.195977e-03
## Blastococcus                                4.340936e-03 2.838455e-03
## Hyphomicrobium                              3.307380e-03 1.974577e-03
## Rhizobium                                   4.409839e-03 9.811181e-03
## Nocardioides                                3.789706e-03 6.170554e-03
## X.uncultured.prokaryote.                    1.012885e-02 4.381093e-03
## Terrabacter                                 5.236684e-03 4.010860e-03
## Rhynchosporium                              9.646524e-04 6.787610e-04
## Massilia                                    9.715428e-03 5.553499e-03
## Pseudomonas                                 9.370909e-03 1.240281e-02
## Peribacillus                                1.288500e-02 1.437739e-02
## Paenibacillus                               4.933508e-02 5.312847e-02
## Oryzihumus                                  2.687246e-03 1.234111e-03
## Rhodanobacter                               1.240267e-03 1.419227e-03
## Rhodococcus                                 4.065321e-03 1.104529e-02
## Spongospora                                 2.273823e-03 2.283105e-03
## Devosia                                     7.786123e-03 9.194126e-03
## NA.                                         7.923930e-03 1.024312e-02
## X.uncultured.archaeon.                      3.858610e-03 4.381093e-03
## Pedobacter                                  5.650107e-03 7.219548e-03
## Nakamurella                                 3.307380e-03 1.198562e-03
## Chujaibacter                                2.687246e-03 2.097988e-03
## Variovorax                                  3.238476e-03 3.023572e-03
## Caldibacillus                               2.687246e-03 3.085277e-03
## Ureibacillus                                2.825053e-03 3.208688e-03
## Terribacillus                               2.549438e-03 4.504505e-03
## Streptosporangium                           2.756150e-03 2.715044e-03
## Pantoea                                     1.653690e-03 8.762187e-03
## Flavobacterium                              3.031765e-03 1.011971e-02
## Piscinibacter                               2.962861e-03 1.239733e-03
## X.uncultured.Acidobacteriota.bacterium.     9.522298e-04 9.522298e-04
## Candidatus.Nitrosotalea                     4.203128e-03 8.055854e-04
## Ammoniphilus                                1.309171e-03 1.357522e-03
## Paraburkholderia                            1.573491e-03 3.146983e-03
## Ramlibacter                                 9.095294e-03 9.996298e-03
## Mesorhizobium                               1.171364e-03 5.856818e-04
## Burkholderia                                3.858610e-03 1.227981e-03
## Caulobacter                                 1.860401e-03 2.715044e-03
## X.uncultured.Alphaproteobacteria.bacterium. 8.268449e-04 4.134224e-04
## Nitrospira                                  4.718762e-04 4.718762e-04
## Cohnella                                    2.825053e-03 4.319388e-03
## Tumebacillus                                2.825053e-03 2.159694e-03
## Pedomicrobium                               1.998209e-03 2.344811e-03
## Niastella                                   6.259563e-04 6.259563e-04
## Shouchella                                  2.480535e-03 4.134271e-03
## Stenotrophobacter                           3.789706e-03 3.517216e-03
## Geomonas                                    1.998209e-03 3.702332e-04
## X.uncultured.Actinomycetes.bacterium.       2.618342e-03 1.357522e-03
## Modestobacter                               2.687246e-03 1.727755e-03
## Kaistia                                     2.618342e-03 5.557931e-04
## Arenimonas                                  2.992732e-04 2.776749e-03
## Ancylobacter                                1.860401e-03 1.542639e-03
## Chryseobacterium                            1.653690e-03 5.862026e-03
## Aetherobacter                               1.112811e-03 1.112811e-03
## Scopulibacillus                             3.927513e-03 6.293965e-03
## Pullulanibacillus                           3.445187e-03 4.319388e-03
## Dyella                                      2.067112e-03 2.406516e-03
## Anaerocolumna                               1.309171e-03 3.455510e-03
## Psychrobacillus                             5.029973e-03 4.566210e-03
## Phenylobacterium                            2.480535e-03 7.433669e-04
## Reyranella                                  3.995206e-04 2.715044e-03
## Methylobacterium                            6.890374e-04 3.445187e-04
## Asticcacaulis                               6.890374e-04 9.255831e-04
## Agrobacterium                               4.961069e-03 8.268543e-03
## Simulacricoccus                             4.065321e-03 1.697962e-03
## Cystobacter                                 3.376283e-03 8.638776e-04
## Candidatus.Solibacter                       2.962861e-03 3.702332e-03
## Achromobacter                               2.411631e-03 1.974577e-03
## Stenotrophomonas                            3.998720e-04 9.009009e-03
## Mycolicibacterium                           6.476952e-03 4.010860e-03
## Noviherbaspirillum                          7.621951e-04 7.621951e-04
## Acidovorax                                  3.514091e-03 5.615204e-03
## Paenarthrobacter                            8.957486e-04 8.021720e-04
## Methylophilus                               3.927513e-03 4.134271e-03
## Exophiala                                   1.190287e-04 1.190287e-04
## Knufia                                      9.988014e-05 9.988014e-05
## Planococcus                                 4.961069e-03 2.480535e-03
## Chitinophaga                                4.261584e-04 4.261584e-04
## X.uncultured.Acetobacteraceae.bacterium.    8.397313e-04 8.397313e-04
## Geothrix                                    5.504743e-04 1.357522e-03
## Ralstonia                                   2.159694e-04 4.319388e-04
## Lysobacter                                  3.099334e-04 1.666050e-03
## Lacunisphaera                               8.957486e-04 4.234417e-04
## Nonomuraea                                  3.393805e-04 6.787610e-04
## Sphingopyxis                                1.653690e-03 1.727755e-03
## Nitrosovibrio                               9.191826e-04 9.191826e-04
## Brevundimonas                               4.616551e-03 2.653338e-03
## Flavitalea                                  3.998720e-04 3.998720e-04
##                                                        9           10
## Priestia                                    0.0607005758 6.135402e-02
## Pseudarthrobacter                           0.0740563020 2.671016e-02
## Niallia                                     0.0786148433 6.435120e-02
## Sporosarcina                                0.0247120921 2.512341e-02
## Bradyrhizobium                              0.0298304543 4.531030e-02
## X.uncultured.bacterium.                     0.3115003199 3.627468e-01
## Sphingomonas                                0.0192738324 2.045134e-02
## Clostridium                                 0.0179142674 2.142102e-02
## Bacillus                                    0.0090371081 7.669252e-03
## Candidatus.Nitrosocosmicus                  0.0095169546 1.419252e-02
## Methylothermalis                            0.0141554702 1.198872e-02
## Streptomyces                                0.0087971849 1.190056e-02
## Dokdonella                                  0.0045585413 6.258815e-03
## Blastococcus                                0.0020793346 4.583921e-03
## Hyphomicrobium                              0.0090371081 1.234133e-02
## Rhizobium                                   0.0075175944 4.583921e-03
## Nocardioides                                0.0015994882 1.851199e-03
## X.uncultured.prokaryote.                    0.0128758797 1.392807e-02
## Terrabacter                                 0.0046385157 5.641749e-03
## Rhynchosporium                              0.0029590531 3.349788e-03
## Massilia                                    0.0061580294 5.377292e-03
## Pseudomonas                                 0.0076775432 2.291961e-03
## Peribacillus                                0.0111964171 1.348731e-02
## Paenibacillus                               0.0304702495 2.433004e-02
## Oryzihumus                                  0.0060780550 5.112835e-03
## Rhodanobacter                               0.0026391555 6.201337e-04
## Rhodococcus                                 0.0057581574 1.057828e-03
## Spongospora                                 0.0049584133 3.437941e-03
## Devosia                                     0.0051983365 3.702398e-03
## NA.                                         0.0177543186 1.207687e-02
## X.uncultured.archaeon.                      0.0039187460 3.614245e-03
## Pedobacter                                  0.0011196417 2.909027e-03
## Nakamurella                                 0.0041586692 4.407616e-03
## Chujaibacter                                0.0059181062 3.085331e-03
## Variovorax                                  0.0048784389 6.258815e-03
## Caldibacillus                               0.0037587972 3.878702e-03
## Ureibacillus                                0.0043186180 4.055007e-03
## Terribacillus                               0.0012795905 2.909027e-03
## Streptosporangium                           0.0034388996 2.909027e-03
## Pantoea                                     0.0051183621 9.696756e-04
## Flavobacterium                              0.0014395393 3.387534e-04
## Piscinibacter                               0.0037587972 5.818054e-03
## X.uncultured.Acidobacteriota.bacterium.     0.0040786948 4.231312e-03
## Candidatus.Nitrosotalea                     0.0008055854 4.583921e-03
## Ammoniphilus                                0.0018394114 2.468265e-03
## Paraburkholderia                            0.0074376200 7.140339e-03
## Ramlibacter                                 0.0038387716 1.058604e-03
## Mesorhizobium                               0.0018394114 5.856818e-04
## Burkholderia                                0.0046385157 6.435120e-03
## Caulobacter                                 0.0004183400 4.183400e-04
## X.uncultured.Alphaproteobacteria.bacterium. 0.0050383877 7.845557e-03
## Nitrospira                                  0.0026391555 5.112835e-03
## Cohnella                                    0.0023992322 9.909400e-04
## Tumebacillus                                0.0007197697 2.380113e-03
## Pedomicrobium                               0.0009655081 9.655081e-04
## Niastella                                   0.0019993602 4.055007e-03
## Shouchella                                  0.0008005489 2.291961e-03
## Stenotrophobacter                           0.0005557931 2.909027e-03
## Geomonas                                    0.0001637626 8.815233e-04
## X.uncultured.Actinomycetes.bacterium.       0.0025591811 3.790550e-03
## Modestobacter                               0.0036788228 7.141724e-04
## Kaistia                                     0.0016794626 1.498590e-03
## Arenimonas                                  0.0002992732 1.057828e-03
## Ancylobacter                                0.0015195138 3.967624e-04
## Chryseobacterium                            0.0002663471 2.663471e-04
## Aetherobacter                               0.0111964171 3.702398e-03
## Scopulibacillus                             0.0036788228 4.649000e-04
## Pullulanibacillus                           0.0010847668 1.084767e-03
## Dyella                                      0.0012795905 5.553597e-03
## Anaerocolumna                               0.0003995206 3.995206e-04
## Psychrobacillus                             0.0019993602 2.203808e-03
## Phenylobacterium                            0.0035988484 2.468265e-03
## Reyranella                                  0.0023992322 2.556417e-03
## Methylobacterium                            0.0033589251 3.173484e-03
## Asticcacaulis                               0.0007997441 3.526093e-04
## Agrobacterium                               0.0007533080 7.533080e-04
## Simulacricoccus                             0.0043186180 3.878702e-03
## Cystobacter                                 0.0004319388 2.203808e-03
## Candidatus.Solibacter                       0.0006325742 2.556417e-03
## Achromobacter                               0.0001549667 1.549667e-04
## Stenotrophomonas                            0.0007997441 3.998720e-04
## Mycolicibacterium                           0.0070377479 4.495769e-03
## Noviherbaspirillum                          0.0052783109 3.526093e-03
## Acidovorax                                  0.0003995206 3.995206e-04
## Paenarthrobacter                            0.0053582853 4.010860e-04
## Methylophilus                               0.0007197697 3.598848e-04
## Exophiala                                   0.0011996161 1.190287e-04
## Knufia                                      0.0003998720 9.988014e-05
## Planococcus                                 0.0083973129 9.432299e-03
## Chitinophaga                                0.0004261584 4.261584e-04
## X.uncultured.Acetobacteraceae.bacterium.    0.0016794626 2.732722e-03
## Geothrix                                    0.0005504743 5.504743e-04
## Ralstonia                                   0.0002159694 3.437941e-03
## Lysobacter                                  0.0022392834 3.099334e-04
## Lacunisphaera                               0.0004234417 4.234417e-04
## Nonomuraea                                  0.0003393805 1.498590e-03
## Sphingopyxis                                0.0003486750 3.486750e-04
## Nitrosovibrio                               0.0009191826 5.818054e-03
## Brevundimonas                               0.0009833262 9.833262e-04
## Flavitalea                                  0.0007997441 3.998720e-04
##                                                       11           12
## Priestia                                    6.109029e-02 0.0608244227
## Pseudarthrobacter                           2.736336e-02 0.0273516194
## Niallia                                     5.861557e-02 0.0503641717
## Sporosarcina                                3.259563e-02 0.0229350690
## Bradyrhizobium                              1.831295e-02 0.0323880366
## X.uncultured.bacterium.                     2.852294e-01 0.3534015187
## Sphingomonas                                2.764619e-02 0.0216178522
## Clostridium                                 2.432299e-02 0.0360297536
## Bacillus                                    1.081807e-02 0.0095304510
## Candidatus.Nitrosocosmicus                  1.774730e-02 0.0150317682
## Methylothermalis                            1.216149e-02 0.0106152177
## Streptomyces                                9.050414e-03 0.0131721680
## Dokdonella                                  3.606024e-03 0.0054238339
## Blastococcus                                2.545429e-03 0.0045715171
## Hyphomicrobium                              5.515096e-03 0.0079807841
## Rhizobium                                   8.626175e-03 0.0047264838
## Nocardioides                                3.959556e-03 0.0041841004
## X.uncultured.prokaryote.                    1.576752e-02 0.0170463350
## Terrabacter                                 5.515096e-03 0.0052688672
## Rhynchosporium                              2.262603e-03 0.0039516504
## Massilia                                    7.494874e-03 0.0016048736
## Pseudomonas                                 1.244432e-02 0.0030218503
## Peribacillus                                9.898890e-03 0.0130946846
## Paenibacillus                               3.309057e-02 0.0275840694
## Oryzihumus                                  5.373683e-03 0.0034867503
## Rhodanobacter                               3.111080e-03 0.0048039671
## Rhodococcus                                 7.353461e-03 0.0019370835
## Spongospora                                 2.333310e-03 0.0011622501
## Devosia                                     8.201937e-03 0.0014749263
## NA.                                         9.050414e-03 0.0167364017
## X.uncultured.archaeon.                      4.100969e-03 0.0035642337
## Pedobacter                                  1.414127e-02 0.0013172168
## Nakamurella                                 5.020151e-03 0.0061211839
## Chujaibacter                                1.909072e-03 0.0048814505
## Variovorax                                  3.535318e-03 0.0033317837
## Caldibacillus                               2.686842e-03 0.0035642337
## Ureibacillus                                2.333310e-03 0.0022470169
## Terribacillus                               1.979778e-03 0.0016271502
## Streptosporangium                           3.888850e-03 0.0039516504
## Pantoea                                     8.555469e-03 0.0004848378
## Flavobacterium                              1.223220e-02 0.0041066171
## Piscinibacter                               3.393905e-03 0.0024794669
## X.uncultured.Acidobacteriota.bacterium.     3.323199e-03 0.0054238339
## Candidatus.Nitrosotalea                     1.767659e-03 0.0051139005
## Ammoniphilus                                1.555540e-03 0.0023245002
## Paraburkholderia                            6.222159e-03 0.0054238339
## Ramlibacter                                 5.373683e-03 0.0044165504
## Mesorhizobium                               2.545429e-03 0.0021695336
## Burkholderia                                4.100969e-03 0.0039516504
## Caulobacter                                 2.404016e-03 0.0004183400
## X.uncultured.Alphaproteobacteria.bacterium. 8.484763e-04 0.0057337672
## Nitrospira                                  2.969667e-03 0.0020920502
## Cohnella                                    9.909400e-04 0.0025569503
## Tumebacillus                                4.100969e-03 0.0024794669
## Pedomicrobium                               9.655081e-04 0.0020145669
## Niastella                                   6.259563e-04 0.0006259563
## Shouchella                                  2.191897e-03 0.0034092670
## Stenotrophobacter                           5.557931e-04 0.0005557931
## Geomonas                                    1.637626e-04 0.0040291337
## X.uncultured.Actinomycetes.bacterium.       2.757548e-03 0.0006787610
## Modestobacter                               1.767659e-03 0.0045715171
## Kaistia                                     2.262603e-03 0.0027894003
## Arenimonas                                  1.767659e-03 0.0002992732
## Ancylobacter                                1.555540e-03 0.0020145669
## Chryseobacterium                            3.111080e-03 0.0002663471
## Aetherobacter                               3.252492e-03 0.0092980009
## Scopulibacillus                             3.111080e-03 0.0009298001
## Pullulanibacillus                           1.084767e-03 0.0021695336
## Dyella                                      2.898961e-03 0.0018596002
## Anaerocolumna                               2.474722e-03 0.0015496668
## Psychrobacillus                             1.626246e-03 0.0035642337
## Phenylobacterium                            7.433669e-04 0.0018596002
## Reyranella                                  3.995206e-04 0.0003995206
## Methylobacterium                            1.696953e-03 0.0012397335
## Asticcacaulis                               1.414127e-03 0.0003874167
## Agrobacterium                               7.565580e-03 0.0007533080
## Simulacricoccus                             1.697962e-03 0.0042615838
## Cystobacter                                 1.414127e-03 0.0020145669
## Candidatus.Solibacter                       6.325742e-04 0.0028668836
## Achromobacter                               2.828254e-03 0.0003099334
## Stenotrophomonas                            9.686771e-03 0.0003998720
## Mycolicibacterium                           6.858517e-03 0.0017645492
## Noviherbaspirillum                          1.626246e-03 0.0046490005
## Acidovorax                                  1.626246e-03 0.0029443670
## Paenarthrobacter                            4.010860e-04 0.0004010860
## Methylophilus                               2.191897e-03 0.0003598848
## Exophiala                                   3.535318e-04 0.0005423834
## Knufia                                      9.988014e-05 0.0002324500
## Planococcus                                 7.212048e-03 0.0063536340
## Chitinophaga                                2.757548e-03 0.0008523168
## X.uncultured.Acetobacteraceae.bacterium.    3.888850e-03 0.0056562839
## Geothrix                                    1.909072e-03 0.0020145669
## Ralstonia                                   2.159694e-04 0.0019370835
## Lysobacter                                  3.099334e-04 0.0006198667
## Lacunisphaera                               4.234417e-04 0.0004234417
## Nonomuraea                                  8.484763e-04 0.0008523168
## Sphingopyxis                                3.747437e-03 0.0006973501
## Nitrosovibrio                               1.838365e-03 0.0043390671
## Brevundimonas                               4.171675e-03 0.0009833262
## Flavitalea                                  3.998720e-04 0.0063536340
##                                                       13           14
## Priestia                                    5.994015e-02 5.388220e-02
## Pseudarthrobacter                           2.385635e-02 2.727577e-02
## Niallia                                     5.788799e-02 5.923695e-02
## Sporosarcina                                2.932877e-02 2.016399e-02
## Bradyrhizobium                              3.522873e-02 4.292169e-02
## X.uncultured.bacterium.                     3.271484e-01 4.088019e-01
## Sphingomonas                                2.684908e-02 1.656627e-02
## Clostridium                                 2.736212e-02 1.171352e-02
## Bacillus                                    1.342454e-02 7.697456e-03
## Candidatus.Nitrosocosmicus                  1.265498e-02 9.036145e-03
## Methylothermalis                            1.162890e-02 1.740295e-02
## Streptomyces                                1.419410e-02 1.447456e-02
## Dokdonella                                  5.643437e-03 5.187416e-03
## Blastococcus                                2.736212e-03 5.940428e-03
## Hyphomicrobium                              7.439077e-03 8.032129e-03
## Rhizobium                                   6.669517e-03 5.187416e-03
## Nocardioides                                2.992732e-03 2.259036e-03
## X.uncultured.prokaryote.                    6.412997e-03 1.280120e-02
## Terrabacter                                 5.386917e-03 4.769076e-03
## Rhynchosporium                              1.966652e-03 2.677376e-03
## Massilia                                    8.978196e-03 4.936412e-03
## Pseudomonas                                 3.420265e-03 2.677376e-03
## Peribacillus                                1.556221e-02 7.530120e-03
## Paenibacillus                               3.796494e-02 2.267403e-02
## Oryzihumus                                  4.788371e-03 8.032129e-03
## Rhodanobacter                               2.052159e-02 2.844712e-03
## Rhodococcus                                 1.881146e-03 1.589692e-03
## Spongospora                                 1.881146e-03 2.426372e-03
## Devosia                                     5.985464e-03 5.271084e-03
## NA.                                         1.000428e-02 1.982932e-02
## X.uncultured.archaeon.                      3.762292e-03 4.518072e-03
## Pedobacter                                  5.386917e-03 1.840696e-03
## Nakamurella                                 4.104318e-03 5.689424e-03
## Chujaibacter                                1.094485e-02 2.593708e-03
## Variovorax                                  2.907225e-03 4.016064e-03
## Caldibacillus                               3.078239e-03 2.593708e-03
## Ureibacillus                                2.821719e-03 4.016064e-03
## Terribacillus                               1.282599e-03 1.506024e-03
## Streptosporangium                           3.676785e-03 2.844712e-03
## Pantoea                                     3.334758e-03 1.422356e-03
## Flavobacterium                              1.624626e-03 1.171352e-03
## Piscinibacter                               2.479692e-03 4.350736e-03
## X.uncultured.Acidobacteriota.bacterium.     4.104318e-03 6.442436e-03
## Candidatus.Nitrosotalea                     3.334758e-03 3.430388e-03
## Ammoniphilus                                1.795639e-03 2.008032e-03
## Paraburkholderia                            4.446345e-03 1.464190e-02
## Ramlibacter                                 1.058604e-03 1.058604e-03
## Mesorhizobium                               2.479692e-03 3.012048e-03
## Burkholderia                                4.018811e-03 4.936412e-03
## Caulobacter                                 4.183400e-04 8.366801e-04
## X.uncultured.Alphaproteobacteria.bacterium. 4.275331e-03 6.107764e-03
## Nitrospira                                  4.718762e-04 2.259036e-03
## Cohnella                                    2.736212e-03 2.844712e-03
## Tumebacillus                                3.598848e-04 3.263052e-03
## Pedomicrobium                               9.655081e-04 3.597724e-03
## Niastella                                   2.137666e-03 1.506024e-03
## Shouchella                                  3.078239e-03 8.005489e-04
## Stenotrophobacter                           1.111586e-03 5.557931e-04
## Geomonas                                    1.637626e-04 1.637626e-04
## X.uncultured.Actinomycetes.bacterium.       6.156477e-03 7.028112e-03
## Modestobacter                               3.762292e-03 7.141724e-04
## Kaistia                                     1.111586e-03 1.840696e-03
## Arenimonas                                  5.985464e-04 1.840696e-03
## Ancylobacter                                1.966652e-03 1.757028e-03
## Chryseobacterium                            1.624626e-03 2.663471e-04
## Aetherobacter                               7.695596e-03 8.450469e-03
## Scopulibacillus                             4.649000e-04 4.649000e-04
## Pullulanibacillus                           4.702864e-03 3.765060e-03
## Dyella                                      5.044891e-03 1.924364e-03
## Anaerocolumna                               3.995206e-04 1.589692e-03
## Psychrobacillus                             2.052159e-03 1.757028e-03
## Phenylobacterium                            1.624626e-03 1.924364e-03
## Reyranella                                  3.995206e-04 3.995206e-04
## Methylobacterium                            3.445187e-04 3.445187e-04
## Asticcacaulis                               1.763047e-04 1.763047e-04
## Agrobacterium                               4.531851e-03 7.533080e-04
## Simulacricoccus                             7.353570e-03 1.697962e-03
## Cystobacter                                 4.319388e-04 4.319388e-04
## Candidatus.Solibacter                       6.325742e-04 1.757028e-03
## Achromobacter                               9.405729e-04 1.549667e-04
## Stenotrophomonas                            4.959384e-03 3.998720e-04
## Mycolicibacterium                           1.764549e-03 1.764549e-03
## Noviherbaspirillum                          7.621951e-04 7.621951e-04
## Acidovorax                                  2.479692e-03 1.924364e-03
## Paenarthrobacter                            1.026080e-03 1.757028e-03
## Methylophilus                               1.111586e-03 3.598848e-04
## Exophiala                                   9.405729e-04 8.366801e-04
## Knufia                                      9.988014e-05 9.988014e-05
## Planococcus                                 8.636169e-03 2.480535e-03
## Chitinophaga                                1.368106e-03 4.261584e-04
## X.uncultured.Acetobacteraceae.bacterium.    3.933305e-03 8.397313e-04
## Geothrix                                    5.504743e-04 3.346720e-03
## Ralstonia                                   2.159694e-04 2.593708e-03
## Lysobacter                                  1.966652e-03 3.099334e-04
## Lacunisphaera                               4.234417e-04 4.234417e-04
## Nonomuraea                                  3.393805e-04 1.171352e-03
## Sphingopyxis                                3.486750e-04 3.486750e-04
## Nitrosovibrio                               2.565199e-03 4.099732e-03
## Brevundimonas                               1.966652e-03 9.833262e-04
## Flavitalea                                  8.550663e-04 3.998720e-04
##                                                       15           16
## Priestia                                    0.0506795017 3.651987e-02
## Pseudarthrobacter                           0.0378444696 3.625134e-02
## Niallia                                     0.0528501321 5.531686e-02
## Sporosarcina                                0.0249150623 4.135338e-02
## Bradyrhizobium                              0.0436957342 3.517723e-02
## X.uncultured.bacterium.                     0.3525858815 2.755102e-01
## Sphingomonas                                0.0184975462 3.490870e-02
## Clostridium                                 0.0164212911 1.342642e-02
## Bacillus                                    0.0083993960 9.667025e-03
## Candidatus.Nitrosocosmicus                  0.0149112873 1.074114e-02
## Methylothermalis                            0.0215175538 5.907626e-03
## Streptomyces                                0.0133069083 7.518797e-03
## Dokdonella                                  0.0051906380 7.518797e-03
## Blastococcus                                0.0064175160 8.861439e-03
## Hyphomicrobium                              0.0075500189 8.861439e-03
## Rhizobium                                   0.0081162703 8.055854e-03
## Nocardioides                                0.0029256323 7.787325e-03
## X.uncultured.prokaryote.                    0.0055681389 1.020408e-02
## Terrabacter                                 0.0057568894 6.176155e-03
## Rhynchosporium                              0.0047187618 4.296455e-03
## Massilia                                    0.0067006418 2.416756e-02
## Pseudomonas                                 0.0037750094 4.054780e-02
## Peribacillus                                0.0125519064 2.030656e-03
## Paenibacillus                               0.0198187995 1.638024e-02
## Oryzihumus                                  0.0050962627 1.262084e-02
## Rhodanobacter                               0.0090600227 4.833512e-03
## Rhodococcus                                 0.0033975085 7.787325e-03
## Spongospora                                 0.0008493771 7.518797e-03
## Devosia                                     0.0050962627 7.250269e-03
## NA.                                         0.0121744054 7.650577e-04
## X.uncultured.archaeon.                      0.0043412609 1.604874e-03
## Pedobacter                                  0.0066062665 2.873255e-02
## Nakamurella                                 0.0051906380 1.181525e-02
## Chujaibacter                                0.0110419026 8.989213e-04
## Variovorax                                  0.0035862590 1.453613e-03
## Caldibacillus                               0.0044356361 9.825757e-04
## Ureibacillus                                0.0032087580 1.123508e-03
## Terribacillus                               0.0013212533 4.912878e-04
## Streptosporangium                           0.0025481314 6.222979e-04
## Pantoea                                     0.0015100038 3.410311e-02
## Flavobacterium                              0.0003387534 8.592911e-03
## Piscinibacter                               0.0045300113 1.239733e-03
## X.uncultured.Acidobacteriota.bacterium.     0.0062287656 5.639098e-03
## Candidatus.Nitrosotalea                     0.0033975085 1.611171e-03
## Ammoniphilus                                0.0003930303 3.930303e-04
## Paraburkholderia                            0.0100981502 1.573491e-03
## Ramlibacter                                 0.0056625142 8.592911e-03
## Mesorhizobium                               0.0033031333 5.856818e-04
## Burkholderia                                0.0051906380 1.227981e-03
## Caulobacter                                 0.0036806342 6.444683e-03
## X.uncultured.Alphaproteobacteria.bacterium. 0.0004134224 4.134224e-04
## Nitrospira                                  0.0009437524 4.718762e-04
## Cohnella                                    0.0019818800 9.909400e-04
## Tumebacillus                                0.0036806342 3.598848e-04
## Pedomicrobium                               0.0030200076 9.655081e-04
## Niastella                                   0.0045300113 6.259563e-04
## Shouchella                                  0.0008005489 8.005489e-04
## Stenotrophobacter                           0.0022650057 5.557931e-04
## Geomonas                                    0.0011325028 1.637626e-04
## X.uncultured.Actinomycetes.bacterium.       0.0006787610 6.787610e-04
## Modestobacter                               0.0007141724 7.141724e-04
## Kaistia                                     0.0017931295 5.557931e-04
## Arenimonas                                  0.0002992732 2.992732e-04
## Ancylobacter                                0.0003967624 3.967624e-04
## Chryseobacterium                            0.0011325028 9.667025e-03
## Aetherobacter                               0.0077387693 1.112811e-03
## Scopulibacillus                             0.0004649000 4.649000e-04
## Pullulanibacillus                           0.0021706304 1.084767e-03
## Dyella                                      0.0025481314 6.397953e-04
## Anaerocolumna                               0.0018875047 3.995206e-04
## Psychrobacillus                             0.0008131231 8.131231e-04
## Phenylobacterium                            0.0007433669 7.433669e-04
## Reyranella                                  0.0016987542 3.995206e-04
## Methylobacterium                            0.0003445187 3.445187e-04
## Asticcacaulis                               0.0001763047 1.763047e-04
## Agrobacterium                               0.0007533080 2.363050e-02
## Simulacricoccus                             0.0042468856 1.697962e-03
## Cystobacter                                 0.0035862590 6.713212e-03
## Candidatus.Solibacter                       0.0031143828 6.325742e-04
## Achromobacter                               0.0007550019 3.759398e-03
## Stenotrophomonas                            0.0064175160 1.208378e-02
## Mycolicibacterium                           0.0017645492 1.764549e-03
## Noviherbaspirillum                          0.0031143828 7.621951e-04
## Acidovorax                                  0.0003995206 3.995206e-04
## Paenarthrobacter                            0.0013212533 4.010860e-04
## Methylophilus                               0.0003598848 2.685285e-03
## Exophiala                                   0.0009437524 1.190287e-04
## Knufia                                      0.0003775009 9.988014e-05
## Planococcus                                 0.0024805347 2.480535e-03
## Chitinophaga                                0.0035862590 2.953813e-03
## X.uncultured.Acetobacteraceae.bacterium.    0.0045300113 8.397313e-04
## Geothrix                                    0.0022650057 5.504743e-04
## Ralstonia                                   0.0033975085 2.159694e-04
## Lysobacter                                  0.0003099334 3.099334e-04
## Lacunisphaera                               0.0009437524 4.234417e-04
## Nonomuraea                                  0.0003393805 3.393805e-04
## Sphingopyxis                                0.0003486750 8.861439e-03
## Nitrosovibrio                               0.0009191826 9.191826e-04
## Brevundimonas                               0.0009833262 9.667025e-03
## Flavitalea                                  0.0015100038 3.998720e-04
##                                                       17           18
## Priestia                                    0.0371683440 0.0243023713
## Pseudarthrobacter                           0.0179551693 0.0259399974
## Niallia                                     0.0503202196 0.0247609066
## Sporosarcina                                0.0187557182 0.0112013625
## Bradyrhizobium                              0.0372827081 0.0334075724
## X.uncultured.bacterium.                     0.4324107960 0.4693436395
## Sphingomonas                                0.0323650503 0.0185379274
## Clostridium                                 0.0114364135 0.0033407572
## Bacillus                                    0.0054894785 0.0030132320
## Candidatus.Nitrosocosmicus                  0.0131518756 0.0068780296
## Methylothermalis                            0.0172689844 0.0117254029
## Streptomyces                                0.0081198536 0.0033407572
## Dokdonella                                  0.0096065874 0.0070745447
## Blastococcus                                0.0038883806 0.0031442421
## Hyphomicrobium                              0.0078911253 0.0075985851
## Rhizobium                                   0.0082342177 0.0081226254
## Nocardioides                                0.0037740165 0.0029477270
## X.uncultured.prokaryote.                    0.0139524245 0.0116598978
## Terrabacter                                 0.0027447392 0.0047163632
## Rhynchosporium                              0.0059469350 0.0043233329
## Massilia                                    0.0061756633 0.0032097472
## Pseudomonas                                 0.0007324544 0.0195860081
## Peribacillus                                0.0101784081 0.0040613127
## Paenibacillus                               0.0157822507 0.0078911253
## Oryzihumus                                  0.0058325709 0.0036682825
## Rhodanobacter                               0.0061756633 0.0070090397
## Rhodococcus                                 0.0025160110 0.0021616664
## Spongospora                                 0.0034309241 0.0030787371
## Devosia                                     0.0046889296 0.0045198480
## NA.                                         0.0034309241 0.0212891393
## X.uncultured.archaeon.                      0.0038883806 0.0032097472
## Pedobacter                                  0.0005598209 0.0051748985
## Nakamurella                                 0.0045745654 0.0049128783
## Chujaibacter                                0.0081198536 0.0053059086
## Variovorax                                  0.0058325709 0.0074675750
## Caldibacillus                               0.0009825757 0.0019651513
## Ureibacillus                                0.0024016468 0.0011235084
## Terribacillus                               0.0016010979 0.0009825757
## Streptosporangium                           0.0027447392 0.0012445958
## Pantoea                                     0.0139524245 0.0084501507
## Flavobacterium                              0.0008005489 0.0007860605
## Piscinibacter                               0.0067474840 0.0053714136
## X.uncultured.Acidobacteriota.bacterium.     0.0049176578 0.0032752522
## Candidatus.Nitrosotalea                     0.0053751144 0.0030787371
## Ammoniphilus                                0.0003930303 0.0007860605
## Paraburkholderia                            0.0041171089 0.0059609590
## Ramlibacter                                 0.0010586043 0.0046508581
## Mesorhizobium                               0.0018298262 0.0017031311
## Burkholderia                                0.0075480329 0.0064849993
## Caulobacter                                 0.0004183400 0.0021616664
## X.uncultured.Alphaproteobacteria.bacterium. 0.0004134224 0.0037337875
## Nitrospira                                  0.0051463861 0.0030132320
## Cohnella                                    0.0009909400 0.0009909400
## Tumebacillus                                0.0024016468 0.0011790908
## Pedomicrobium                               0.0030878317 0.0025546967
## Niastella                                   0.0026303751 0.0022926765
## Shouchella                                  0.0016010979 0.0008005489
## Stenotrophobacter                           0.0013723696 0.0005557931
## Geomonas                                    0.0005718207 0.0003275252
## X.uncultured.Actinomycetes.bacterium.       0.0006787610 0.0045198480
## Modestobacter                               0.0036596523 0.0035372724
## Kaistia                                     0.0014867338 0.0014411110
## Arenimonas                                  0.0014867338 0.0032097472
## Ancylobacter                                0.0016010979 0.0016376261
## Chryseobacterium                            0.0002663471 0.0007860605
## Aetherobacter                               0.0096065874 0.0123149483
## Scopulibacillus                             0.0004649000 0.0004649000
## Pullulanibacillus                           0.0010847668 0.0010847668
## Dyella                                      0.0006397953 0.0024891917
## Anaerocolumna                               0.0003995206 0.0003995206
## Psychrobacillus                             0.0008131231 0.0008131231
## Phenylobacterium                            0.0014867338 0.0020961614
## Reyranella                                  0.0024016468 0.0009825757
## Methylobacterium                            0.0010292772 0.0030787371
## Asticcacaulis                               0.0001763047 0.0008515656
## Agrobacterium                               0.0007533080 0.0015066160
## Simulacricoccus                             0.0050320220 0.0055679287
## Cystobacter                                 0.0024016468 0.0026202018
## Candidatus.Solibacter                       0.0027447392 0.0006325742
## Achromobacter                               0.0001549667 0.0009825757
## Stenotrophomonas                            0.0003998720 0.0017686362
## Mycolicibacterium                           0.0017645492 0.0067470195
## Noviherbaspirillum                          0.0069762123 0.0028167169
## Acidovorax                                  0.0003995206 0.0028167169
## Paenarthrobacter                            0.0004010860 0.0004010860
## Methylophilus                               0.0003598848 0.0034717673
## Exophiala                                   0.0001190287 0.0001190287
## Knufia                                      0.0004574565 0.0004585353
## Planococcus                                 0.0024805347 0.0024805347
## Chitinophaga                                0.0004261584 0.0004261584
## X.uncultured.Acetobacteraceae.bacterium.    0.0008397313 0.0008397313
## Geothrix                                    0.0043458371 0.0018996463
## Ralstonia                                   0.0002159694 0.0052404035
## Lysobacter                                  0.0003099334 0.0013101009
## Lacunisphaera                               0.0019441903 0.0030132320
## Nonomuraea                                  0.0003393805 0.0003393805
## Sphingopyxis                                0.0003486750 0.0003486750
## Nitrosovibrio                               0.0066331199 0.0009191826
## Brevundimonas                               0.0009833262 0.0009833262
## Flavitalea                                  0.0020585544 0.0003998720
## 
## $meta.dat.use
##       ndvi_01
## 1   0.3558477
## 2   0.5945529
## 3   0.5095020
## 4   0.6619353
## 5   0.6121273
## 6   0.5747102
## 7   0.6887884
## 8   0.8302155
## 9   0.3130208
## 10 -1.9896865
## 11  0.9980715
## 12 -1.9625407
## 13  0.6228720
## 14 -0.8421145
## 15 -1.1410253
## 16  0.9838132
## 17 -0.7571646
## 18 -1.0529250
## 
## $wald
## $wald$beta
##              Priestia Pseudarthrobacter    Niallia Sporosarcina Bradyrhizobium
## (Intercept) 4.6532698        3.35182313 4.27627772    3.2500536      3.3109685
## ndvi_01     0.2908706       -0.01295889 0.02963606    0.2224172     -0.3723397
##             X.uncultured.bacterium. Sphingomonas Clostridium  Bacillus
## (Intercept)                6.940905    3.0921705   2.9301742 2.1155303
## ndvi_01                   -0.175832    0.1132216   0.2888848 0.5038372
##             Candidatus.Nitrosocosmicus Methylothermalis Streptomyces Dokdonella
## (Intercept)                 2.15425145       2.34776618   2.07054591  0.8966364
## ndvi_01                    -0.02293984      -0.06829866   0.07879447 -0.2107757
##             Blastococcus Hyphomicrobium Rhizobium Nocardioides
## (Intercept)    0.5191936      1.1933479 1.2889918    0.3301151
## ndvi_01       -0.1714653     -0.3509935 0.1085762    0.2192896
##             X.uncultured.prokaryote. Terrabacter Rhynchosporium  Massilia
## (Intercept)                1.4507973  0.84512892     -0.4879430 1.1699362
## ndvi_01                   -0.4981622 -0.01556647     -0.6957948 0.4516852
##             Pseudomonas Peribacillus Paenibacillus Oryzihumus Rhodanobacter
## (Intercept)   0.9131006   1.93578772     3.4610242  0.4279855    0.45726000
## ndvi_01       0.5560963  -0.03391988     0.3904126 -0.2295111    0.05689519
##             Rhodococcus  Spongospora   Devosia        NA.
## (Intercept) -0.07724953 -0.467157862 0.8781733  1.6685740
## ndvi_01      0.41057100  0.009927203 0.3809317 -0.5102662
##             X.uncultured.archaeon. Pedobacter Nakamurella Chujaibacter
## (Intercept)             0.58671714  0.4810370   0.5677825    0.2910812
## ndvi_01                 0.01821888  0.6209994  -0.2100447   -0.4895215
##             Variovorax Caldibacillus Ureibacillus Terribacillus
## (Intercept)  0.5059363    0.03067391   0.12235251   -0.56627457
## ndvi_01     -0.2888945   -0.08232023   0.02362413    0.09521136
##             Streptosporangium   Pantoea Flavobacterium Piscinibacter
## (Intercept)        -0.1312078 0.2351975     -0.3775764     0.2303199
## ndvi_01            -0.1172991 0.6830979      0.9926167    -0.4080704
##             X.uncultured.Acidobacteriota.bacterium. Candidatus.Nitrosotalea
## (Intercept)                               0.3457121              0.04760825
## ndvi_01                                  -0.3923863             -0.39661076
##             Ammoniphilus Paraburkholderia Ramlibacter Mesorhizobium
## (Intercept)  -0.88120989        0.6773986   0.4513150   -0.76874289
## ndvi_01       0.09690053       -0.5196599   0.4871943   -0.09980397
##             Burkholderia Caulobacter
## (Intercept)    0.2439088  -0.8386258
## ndvi_01       -0.5731013   0.6747929
##             X.uncultured.Alphaproteobacteria.bacterium. Nitrospira   Cohnella
## (Intercept)                                  -0.3241090 -0.5317908 -0.2287707
## ndvi_01                                      -0.4652562 -0.4192659  0.3107271
##             Tumebacillus Pedomicrobium  Niastella Shouchella Stenotrophobacter
## (Intercept)   -0.9728706    -0.4567717 -0.7777083 -0.4234394        -0.8356651
## ndvi_01       -0.4977122    -0.1329595 -0.3956868  0.2550895         0.2329838
##               Geomonas X.uncultured.Actinomycetes.bacterium. Modestobacter
## (Intercept) -1.9687427                           -0.60304317   -0.59311085
## ndvi_01     -0.3066282                           -0.02303514    0.03406518
##                Kaistia  Arenimonas Ancylobacter Chryseobacterium Aetherobacter
## (Intercept) -1.1868446 -1.49092291  -1.40087181        -1.564235     0.2441053
## ndvi_01     -0.4158965  0.09348809   0.01574259         1.029805    -0.9110447
##             Scopulibacillus Pullulanibacillus     Dyella Anaerocolumna
## (Intercept)      -0.6063422        -0.2419317 -0.8918774    -1.2659660
## ndvi_01           1.0876577         0.2684949 -0.4311200     0.2903012
##             Psychrobacillus Phenylobacterium Reyranella Methylobacterium
## (Intercept)     -0.70435603       -0.9131029  -1.324704        -1.617222
## ndvi_01          0.04803972       -0.2456907  -0.069561        -0.361852
##             Asticcacaulis Agrobacterium Simulacricoccus Cystobacter
## (Intercept)    -2.6191646    -0.1674659       0.1039180  -1.3092242
## ndvi_01         0.3044225     1.2228892      -0.3514709  -0.4657767
##             Candidatus.Solibacter Achromobacter Stenotrophomonas
## (Intercept)            -0.9262982    -2.2315226       -0.8721071
## ndvi_01                -0.4383659     0.8355893        0.8484201
##             Mycolicibacterium Noviherbaspirillum   Acidovorax Paenarthrobacter
## (Intercept)         0.1898809         -0.7688254 -1.488138017       -1.6757940
## ndvi_01             0.1234694         -0.7429753 -0.002653034        0.2562036
##             Methylophilus  Exophiala     Knufia Planococcus Chitinophaga
## (Intercept)    -1.6019211 -3.3805113 -3.8469603  0.57449260   -1.5098392
## ndvi_01         0.6867654 -0.1893542 -0.2341847 -0.08705161    0.3991525
##             X.uncultured.Acetobacteraceae.bacterium.   Geothrix Ralstonia
## (Intercept)                               -0.7665400 -1.4361262 -2.189662
## ndvi_01                                   -0.3612967 -0.5318756 -1.322694
##             Lysobacter Lacunisphaera Nonomuraea Sphingopyxis Nitrosovibrio
## (Intercept) -2.0003018    -1.9814564 -2.2282590   -1.8959218    -0.7952091
## ndvi_01      0.3192142    -0.1543121 -0.1727455    0.7449425    -0.6612509
##             Brevundimonas Flavitalea
## (Intercept)    -0.6748271 -1.9727649
## ndvi_01         0.6451148 -0.5190283
## 
## $wald$sig
##                                    Priestia 
##                                   0.5952674 
##                           Pseudarthrobacter 
##                                   0.5299471 
##                                     Niallia 
##                                   0.4068989 
##                                Sporosarcina 
##                                   0.3814258 
##                              Bradyrhizobium 
##                                   0.4669034 
##                     X.uncultured.bacterium. 
##                                   0.2940500 
##                                Sphingomonas 
##                                   0.3670105 
##                                 Clostridium 
##                                   0.8271474 
##                                    Bacillus 
##                                   0.6350670 
##                  Candidatus.Nitrosocosmicus 
##                                   0.3547473 
##                            Methylothermalis 
##                                   0.4663154 
##                                Streptomyces 
##                                   0.6119858 
##                                  Dokdonella 
##                                   0.4585849 
##                                Blastococcus 
##                                   0.6531803 
##                              Hyphomicrobium 
##                                   0.6862646 
##                                   Rhizobium 
##                                   0.3872565 
##                                Nocardioides 
##                                   0.5890101 
##                    X.uncultured.prokaryote. 
##                                   1.1824678 
##                                 Terrabacter 
##                                   0.3767202 
##                              Rhynchosporium 
##                                   0.9376167 
##                                    Massilia 
##                                   0.7414033 
##                                 Pseudomonas 
##                                   1.3825048 
##                                Peribacillus 
##                                   0.7581870 
##                               Paenibacillus 
##                                   0.5978034 
##                                  Oryzihumus 
##                                   1.0066239 
##                               Rhodanobacter 
##                                   1.2537821 
##                                 Rhodococcus 
##                                   1.0522452 
##                                 Spongospora 
##                                   1.0819570 
##                                     Devosia 
##                                   0.5016777 
##                                         NA. 
##                                   1.1549799 
##                      X.uncultured.archaeon. 
##                                   0.4060779 
##                                  Pedobacter 
##                                   1.5900791 
##                                 Nakamurella 
##                                   0.7793319 
##                                Chujaibacter 
##                                   0.9626041 
##                                  Variovorax 
##                                   0.6482507 
##                               Caldibacillus 
##                                   0.5727122 
##                                Ureibacillus 
##                                   0.6320379 
##                               Terribacillus 
##                                   0.8863362 
##                           Streptosporangium 
##                                   0.8056243 
##                                     Pantoea 
##                                   1.7653036 
##                              Flavobacterium 
##                                   1.6414427 
##                               Piscinibacter 
##                                   0.6960771 
##     X.uncultured.Acidobacteriota.bacterium. 
##                                   0.9705570 
##                     Candidatus.Nitrosotalea 
##                                   0.8684358 
##                                Ammoniphilus 
##                                   1.0116295 
##                            Paraburkholderia 
##                                   0.7938603 
##                                 Ramlibacter 
##                                   1.0105935 
##                               Mesorhizobium 
##                                   0.8668561 
##                                Burkholderia 
##                                   0.7738155 
##                                 Caulobacter 
##                                   1.1825173 
## X.uncultured.Alphaproteobacteria.bacterium. 
##                                   1.7829811 
##                                  Nitrospira 
##                                   1.3405628 
##                                    Cohnella 
##                                   0.7992388 
##                                Tumebacillus 
##                                   1.2721930 
##                               Pedomicrobium 
##                                   0.7769177 
##                                   Niastella 
##                                   1.0193103 
##                                  Shouchella 
##                                   1.0327884 
##                           Stenotrophobacter 
##                                   1.1958655 
##                                    Geomonas 
##                                   1.8703616 
##       X.uncultured.Actinomycetes.bacterium. 
##                                   1.1897457 
##                               Modestobacter 
##                                   1.0434939 
##                                     Kaistia 
##                                   0.7183045 
##                                  Arenimonas 
##                                   1.3830897 
##                                Ancylobacter 
##                                   1.0242427 
##                            Chryseobacterium 
##                                   1.4444403 
##                               Aetherobacter 
##                                   1.0934477 
##                             Scopulibacillus 
##                                   1.3048858 
##                           Pullulanibacillus 
##                                   0.9334029 
##                                      Dyella 
##                                   0.9788325 
##                               Anaerocolumna 
##                                   1.3596454 
##                             Psychrobacillus 
##                                   0.9275016 
##                            Phenylobacterium 
##                                   0.8144945 
##                                  Reyranella 
##                                   1.2931143 
##                            Methylobacterium 
##                                   1.2651467 
##                               Asticcacaulis 
##                                   1.1544918 
##                               Agrobacterium 
##                                   1.3273918 
##                             Simulacricoccus 
##                                   0.6901169 
##                                 Cystobacter 
##                                   1.3445058 
##                       Candidatus.Solibacter 
##                                   1.1614012 
##                               Achromobacter 
##                                   1.7436160 
##                            Stenotrophomonas 
##                                   1.9633821 
##                           Mycolicibacterium 
##                                   0.8576412 
##                          Noviherbaspirillum 
##                                   0.9617672 
##                                  Acidovorax 
##                                   1.4062333 
##                            Paenarthrobacter 
##                                   1.3050350 
##                               Methylophilus 
##                                   1.2828664 
##                                   Exophiala 
##                                   1.2717820 
##                                      Knufia 
##                                   0.9791929 
##                                 Planococcus 
##                                   0.8547469 
##                                Chitinophaga 
##                                   1.4593850 
##    X.uncultured.Acetobacteraceae.bacterium. 
##                                   1.0908330 
##                                    Geothrix 
##                                   0.9433175 
##                                   Ralstonia 
##                                   1.2689947 
##                                  Lysobacter 
##                                   1.3655714 
##                               Lacunisphaera 
##                                   0.9398076 
##                                  Nonomuraea 
##                                   0.9465409 
##                                Sphingopyxis 
##                                   1.3274967 
##                               Nitrosovibrio 
##                                   0.8921094 
##                               Brevundimonas 
##                                   0.8832970 
##                                  Flavitalea 
##                                   1.0769123 
## 
## $wald$X
##    (Intercept)    ndvi_01
## 1            1  0.3558477
## 2            1  0.5945529
## 3            1  0.5095020
## 4            1  0.6619353
## 5            1  0.6121273
## 6            1  0.5747102
## 7            1  0.6887884
## 8            1  0.8302155
## 9            1  0.3130208
## 10           1 -1.9896865
## 11           1  0.9980715
## 12           1 -1.9625407
## 13           1  0.6228720
## 14           1 -0.8421145
## 15           1 -1.1410253
## 16           1  0.9838132
## 17           1 -0.7571646
## 18           1 -1.0529250
## attr(,"assign")
## [1] 0 1
## 
## $wald$bias
## [1] -0.62689352 -0.04378204
# Show effect size and significance plots
linda.plot(
l_model,
variables.plot = c('ndvi_01'),
alpha = 0.05,
lfc.cut = 1,
legend = TRUE
)
## $plot.lfc
## NULL
## 
## $plot.volcano
## $plot.volcano[[1]]

l_model_df <- as.data.frame(l_model$output)

write.xlsx(l_model_df, file = "supplementary_table_ndvi_regression_16s.xlsx", rowNames = TRUE, colnames = TRUE)
Credit

This script is based on ideas and code from the dada2 Tutorial by Benjamin Callahan, the publication “Bioconductor Workflow for Microbiome Data Analysis: from raw reads to community analyses” by Callahan et al. (2016) and various pages of the official phyloseq website by Paul J. McMurdie.

sessionInfo()
## R version 4.4.2 (2024-10-31)
## Platform: x86_64-pc-linux-gnu
## Running under: Ubuntu 22.04.5 LTS
## 
## Matrix products: default
## BLAS:   /usr/lib/x86_64-linux-gnu/blas/libblas.so.3.10.0 
## LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.10.0
## 
## locale:
##  [1] LC_CTYPE=C.UTF-8       LC_NUMERIC=C           LC_TIME=C.UTF-8       
##  [4] LC_COLLATE=C.UTF-8     LC_MONETARY=C.UTF-8    LC_MESSAGES=C.UTF-8   
##  [7] LC_PAPER=C.UTF-8       LC_NAME=C              LC_ADDRESS=C          
## [10] LC_TELEPHONE=C         LC_MEASUREMENT=C.UTF-8 LC_IDENTIFICATION=C   
## 
## time zone: Europe/Oslo
## tzcode source: system (glibc)
## 
## attached base packages:
## [1] parallel  stats4    stats     graphics  grDevices utils     datasets 
## [8] methods   base     
## 
## other attached packages:
##  [1] MicrobiomeStat_1.2  aod_1.3.3           BBmisc_1.13        
##  [4] betareg_3.2-1       ggpmisc_0.6.1       ggpp_0.5.8-1       
##  [7] openxlsx_4.2.7.1    readxl_1.4.3        lubridate_1.9.4    
## [10] forcats_1.0.0       stringr_1.5.1       dplyr_1.1.4        
## [13] purrr_1.0.2         readr_2.1.5         tidyr_1.3.1        
## [16] tibble_3.2.1        tidyverse_2.0.0     ggpubr_0.6.0       
## [19] viridis_0.6.5       viridisLite_0.4.2   phangorn_2.12.1    
## [22] SIAMCAT_2.10.0      mlr3_0.22.1         biomformat_1.34.0  
## [25] vegan_2.6-8         lattice_0.22-6      permute_0.9-7      
## [28] scales_1.3.0        gridExtra_2.3       ape_5.8-1          
## [31] reshape2_1.4.4      phyloseq_1.50.0     ggplot2_3.5.1      
## [34] cuphyr_0.3          DECIPHER_3.2.0      Biostrings_2.74.0  
## [37] GenomeInfoDb_1.42.1 XVector_0.46.0      IRanges_2.40.0     
## [40] S4Vectors_0.44.0    BiocGenerics_0.52.0 dada2_1.34.0       
## [43] Rcpp_1.0.13-1      
## 
## loaded via a namespace (and not attached):
##   [1] fs_1.6.5                    matrixStats_1.4.1          
##   [3] bitops_1.0-9                devtools_2.4.5             
##   [5] httr_1.4.7                  RColorBrewer_1.1-3         
##   [7] numDeriv_2016.8-1.1         profvis_0.4.0              
##   [9] tools_4.4.2                 mlr3learners_0.9.0         
##  [11] backports_1.5.0             utf8_1.2.4                 
##  [13] R6_2.5.1                    mgcv_1.9-1                 
##  [15] rhdf5filters_1.18.0         urlchecker_1.0.1           
##  [17] withr_3.0.2                 prettyunits_1.2.0          
##  [19] textshaping_0.4.1           quantreg_5.99.1            
##  [21] cli_3.6.3                   Biobase_2.66.0             
##  [23] formatR_1.14                sandwich_3.1-1             
##  [25] labeling_0.4.3              sass_0.4.9                 
##  [27] systemfonts_1.1.0           Rsamtools_2.22.0           
##  [29] mlr3tuning_1.3.0            paradox_1.0.1              
##  [31] parallelly_1.41.0           sessioninfo_1.2.2          
##  [33] rstudioapi_0.17.1           generics_0.1.3             
##  [35] shape_1.4.6.1               hwriter_1.3.2.1            
##  [37] car_3.1-3                   zip_2.3.1                  
##  [39] Matrix_1.7-1                interp_1.1-6               
##  [41] abind_1.4-8                 infotheo_1.2.0.1           
##  [43] lifecycle_1.0.4             yaml_2.3.10                
##  [45] carData_3.0-5               SummarizedExperiment_1.36.0
##  [47] rhdf5_2.50.0                SparseArray_1.6.0          
##  [49] grid_4.4.2                  LiblineaR_2.10-24          
##  [51] promises_1.3.2              crayon_1.5.3               
##  [53] pwalign_1.2.0               miniUI_0.1.1.1             
##  [55] cowplot_1.1.3               pillar_1.10.0              
##  [57] knitr_1.49                  beanplot_1.3.1             
##  [59] GenomicRanges_1.58.0        statip_0.2.3               
##  [61] boot_1.3-31                 codetools_0.2-20           
##  [63] fastmatch_1.1-4             glue_1.8.0                 
##  [65] ShortRead_1.64.0            data.table_1.16.4          
##  [67] remotes_2.5.0               vctrs_0.6.5                
##  [69] png_0.1-8                   cellranger_1.1.0           
##  [71] gtable_0.3.6                cachem_1.1.0               
##  [73] xfun_0.49                   S4Arrays_1.6.0             
##  [75] mime_0.12                   modeest_2.4.0              
##  [77] survival_3.8-3              timeDate_4041.110          
##  [79] iterators_1.0.14            statmod_1.5.0              
##  [81] ellipsis_0.3.2              nlme_3.1-166               
##  [83] usethis_3.1.0               bbotk_1.5.0                
##  [85] progress_1.2.3              PRROC_1.3.1                
##  [87] fBasics_4041.97             bslib_0.8.0                
##  [89] rpart_4.1.23                colorspace_2.1-1           
##  [91] DBI_1.2.3                   nnet_7.3-19                
##  [93] ade4_1.7-22                 mlr3misc_0.16.0            
##  [95] tidyselect_1.2.1            timeSeries_4041.111        
##  [97] compiler_4.4.2              curl_6.0.1                 
##  [99] glmnet_4.1-8                lgr_0.4.4                  
## [101] SparseM_1.84-2              DelayedArray_0.32.0        
## [103] checkmate_2.3.2             lmtest_0.9-40              
## [105] quadprog_1.5-8              spatial_7.3-17             
## [107] palmerpenguins_0.1.1        digest_0.6.37              
## [109] minqa_1.2.8                 rmarkdown_2.29             
## [111] htmltools_0.5.8.1           pkgconfig_2.0.3            
## [113] jpeg_0.1-10                 lme4_1.1-35.5              
## [115] MatrixGenerics_1.18.0       stabledist_0.7-2           
## [117] fastmap_1.2.0               rlang_1.1.4                
## [119] htmlwidgets_1.6.4           UCSC.utils_1.2.0           
## [121] shiny_1.10.0                farver_2.1.2               
## [123] jquerylib_0.1.4             zoo_1.8-12                 
## [125] jsonlite_1.8.9              BiocParallel_1.40.0        
## [127] magrittr_2.0.3              polynom_1.4-1              
## [129] modeltools_0.2-23           Formula_1.2-5              
## [131] GenomeInfoDbData_1.2.13     Rhdf5lib_1.28.0            
## [133] munsell_0.5.1               stringi_1.8.4              
## [135] pROC_1.18.5                 stable_1.1.6               
## [137] zlibbioc_1.52.0             MASS_7.3-61                
## [139] plyr_1.8.9                  pkgbuild_1.4.5             
## [141] flexmix_2.3-19              ggrepel_0.9.6              
## [143] listenv_0.9.1               deldir_2.0-4               
## [145] splines_4.4.2               multtest_2.62.0            
## [147] hms_1.1.3                   igraph_2.1.2               
## [149] uuid_1.2-1                  ggsignif_0.6.4             
## [151] rmutil_1.1.10               pkgload_1.4.0              
## [153] evaluate_1.0.1              latticeExtra_0.6-30        
## [155] RcppParallel_5.1.9          nloptr_2.1.1               
## [157] tzdb_0.4.0                  foreach_1.5.2              
## [159] httpuv_1.6.15               MatrixModels_0.5-3         
## [161] clue_0.3-66                 future_1.34.0              
## [163] gridBase_0.4-7              broom_1.0.7                
## [165] xtable_1.8-4                rstatix_0.7.2              
## [167] later_1.4.1                 ragg_1.3.3                 
## [169] lmerTest_3.1-3              memoise_2.0.1              
## [171] GenomicAlignments_1.42.0    cluster_2.1.8              
## [173] corrplot_0.95               timechange_0.3.0           
## [175] globals_0.16.3