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IMAP: Integrated Microbiome Analysis Pipelines: End-to-End Practical User Guides Using Integrated Approaches

6 Reviewing Phyloseq Objects

In this section, we demonstrate basic functions for reviewing and exploring the contents of phyloseq objects. By familiarizing ourselves with these functions, we gain insights into the structure, composition, and metadata associated with phyloseq objects, facilitating effective analysis and interpretation of microbiome data.

library(phyloseq)
library(ape)

ps <- ps_dietswap
ps
phyloseq-class experiment-level object
otu_table()   OTU Table:         [ 130 taxa and 222 samples ]
sample_data() Sample Data:       [ 222 samples by 8 sample variables ]
tax_table()   Taxonomy Table:    [ 130 taxa by 3 taxonomic ranks ]
phy_tree()    Phylogenetic Tree: [ 130 tips and 129 internal nodes ]

6.1 Taxonomy rank names 

rank_names(ps)
[1] "Phylum" "Family" "Genus"

6.2 Number of taxa 

ntaxa(ps)
[1] 130

6.3 Split by taxon name, e.g. Firmicutes 

library(metagMisc) # foe phyloseq_sep_tax()

taxa_stats <- phyloseq_sep_tax(ps, TaxRank = "Phylum", drop_NA = FALSE)
cat("\n")
taxa_stats$Firmicutes
phyloseq-class experiment-level object
otu_table()   OTU Table:         [ 76 taxa and 222 samples ]
sample_data() Sample Data:       [ 222 samples by 8 sample variables ]
tax_table()   Taxonomy Table:    [ 76 taxa by 3 taxonomic ranks ]
phy_tree()    Phylogenetic Tree: [ 76 tips and 75 internal nodes ]
cat("\n")
taxa_stats$Bacteroidetes
phyloseq-class experiment-level object
otu_table()   OTU Table:         [ 16 taxa and 222 samples ]
sample_data() Sample Data:       [ 222 samples by 8 sample variables ]
tax_table()   Taxonomy Table:    [ 16 taxa by 3 taxonomic ranks ]
phy_tree()    Phylogenetic Tree: [ 16 tips and 15 internal nodes ]
cat("\n")
taxa_stats$Actinobacteria
phyloseq-class experiment-level object
otu_table()   OTU Table:         [ 8 taxa and 222 samples ]
sample_data() Sample Data:       [ 222 samples by 8 sample variables ]
tax_table()   Taxonomy Table:    [ 8 taxa by 3 taxonomic ranks ]
phy_tree()    Phylogenetic Tree: [ 8 tips and 7 internal nodes ]

6.4 Number of samples 

nsamples(ps)
[1] 222

6.5 Sample names 

head(sample_names(ps), 20)
 [1] "Sample-1"  "Sample-2"  "Sample-3"  "Sample-4"  "Sample-5"  "Sample-6" 
 [7] "Sample-7"  "Sample-8"  "Sample-9"  "Sample-10" "Sample-11" "Sample-12"
[13] "Sample-13" "Sample-14" "Sample-15" "Sample-16" "Sample-17" "Sample-18"
[19] "Sample-19" "Sample-20"

6.6 Sample variables 

sample_variables(ps)
[1] "subject"                "sex"                    "nationality"           
[4] "group"                  "sample"                 "timepoint"             
[7] "timepoint.within.group" "bmi_group"

6.7 Abundance table 

otu_table(ps)[1:5, 1:5]
OTU Table:          [5 taxa and 5 samples]
                     taxa are rows
                                      Sample-1 Sample-2 Sample-3 Sample-4
Phascolarctobacterium faecium et rel.        4        5       13        4
Actinomycetaceae                             0        1        0        1
Eubacterium hallii et rel.                  10      102       39       31
Clostridium stercorarium et rel.             6        3        9        8
Clostridium symbiosum et rel.              220      247      214      111
                                      Sample-5
Phascolarctobacterium faecium et rel.       33
Actinomycetaceae                             0
Eubacterium hallii et rel.                  28
Clostridium stercorarium et rel.             3
Clostridium symbiosum et rel.              155

6.8 Taxonomy table 

tax_table(ps)[1:5, ]
Taxonomy Table:     [5 taxa by 3 taxonomic ranks]:
                                      Phylum          
Phascolarctobacterium faecium et rel. "Firmicutes"    
Actinomycetaceae                      "Actinobacteria"
Eubacterium hallii et rel.            "Firmicutes"    
Clostridium stercorarium et rel.      "Firmicutes"    
Clostridium symbiosum et rel.         "Firmicutes"    
                                      Family                    
Phascolarctobacterium faecium et rel. "Clostridium cluster IX"  
Actinomycetaceae                      "Actinobacteria"          
Eubacterium hallii et rel.            "Clostridium cluster XIVa"
Clostridium stercorarium et rel.      "Clostridium cluster III" 
Clostridium symbiosum et rel.         "Clostridium cluster XIVa"
                                      Genus                                  
Phascolarctobacterium faecium et rel. "Phascolarctobacterium faecium et rel."
Actinomycetaceae                      "Actinomycetaceae"                     
Eubacterium hallii et rel.            "Eubacterium hallii et rel."           
Clostridium stercorarium et rel.      "Clostridium stercorarium et rel."     
Clostridium symbiosum et rel.         "Clostridium symbiosum et rel."

6.9 Phylogenetic tree if available

6.10 Taxa names 

taxa_names(ps)[1:10]
 [1] "Phascolarctobacterium faecium et rel."
 [2] "Actinomycetaceae"                     
 [3] "Eubacterium hallii et rel."           
 [4] "Clostridium stercorarium et rel."     
 [5] "Clostridium symbiosum et rel."        
 [6] "Wissella et rel."                     
 [7] "Peptostreptococcus anaerobius et rel."
 [8] "Ruminococcus bromii et rel."          
 [9] "Prevotella ruminicola et rel."        
[10] "Yersinia et rel."

6.11 Summarize Phyloseq Object 


microbiome::summarize_phyloseq(ps)
[[1]]
[1] "1] Min. number of reads = 1776"

[[2]]
[1] "2] Max. number of reads = 28883"

[[3]]
[1] "3] Total number of reads = 2949085"

[[4]]
[1] "4] Average number of reads = 13284.1666666667"

[[5]]
[1] "5] Median number of reads = 13255"

[[6]]
[1] "7] Sparsity = 0.205647955647956"

[[7]]
[1] "6] Any OTU sum to 1 or less? YES"

[[8]]
[1] "8] Number of singletons = 10"

[[9]]
[1] "9] Percent of OTUs that are singletons \n        (i.e. exactly one read detected across all samples)2.30769230769231"

[[10]]
[1] "10] Number of sample variables are: 8"

[[11]]
[1] "subject"                "sex"                    "nationality"           
[4] "group"                  "sample"                 "timepoint"             
[7] "timepoint.within.group" "bmi_group"

6.12 Sort Samples in ascending and descending 

cat("In ascending order\n")
In ascending order
head(sort(phyloseq::sample_sums(ps), decreasing = F))
 Sample-56 Sample-195 Sample-196  Sample-42 Sample-164 Sample-147 
      1776       4477       4606       5006       5541       5624 
cat("\nIn descending order\n")

In descending order
head(sort(phyloseq::sample_sums(ps), decreasing = T))
  Sample-3  Sample-11  Sample-12  Sample-54 Sample-208  Sample-20 
     28883      26895      25238      23529      23131      22413

6.13 Drop Samples Below a Threshold 

ps1perc0 <- phyloseq::subset_samples(ps, phyloseq::sample_sums(ps) > 1)
ps1perc0
phyloseq-class experiment-level object
otu_table()   OTU Table:         [ 130 taxa and 222 samples ]
sample_data() Sample Data:       [ 222 samples by 8 sample variables ]
tax_table()   Taxonomy Table:    [ 130 taxa by 3 taxonomic ranks ]
phy_tree()    Phylogenetic Tree: [ 130 tips and 129 internal nodes ]

6.14 Drop Taxa Below a Threshold 

pstaxa01 <- phyloseq::prune_taxa(phyloseq::taxa_sums(ps) > 1, ps)
pstaxa01 
phyloseq-class experiment-level object
otu_table()   OTU Table:         [ 120 taxa and 222 samples ]
sample_data() Sample Data:       [ 222 samples by 8 sample variables ]
tax_table()   Taxonomy Table:    [ 120 taxa by 3 taxonomic ranks ]
phy_tree()    Phylogenetic Tree: [ 120 tips and 119 internal nodes ]