Remove chimeric sequences using vsearch and a reference database

Applies UCHIME reference-based chimera detection via vsearch to identify and remove chimeric sequences from a phyloseq object. Unlike de novo detection (MiscMetabar::chimera_removal_vs()), this method compares each query sequence against a trusted reference database rather than inferring chimeras from abundance patterns within the sample.

Usage

chimera_removal_vs_ref(
  physeq,
  database,
  vsearchpath = "vsearch",
  vsearch_args = "--fasta_width 0",
  return_a_list = FALSE,
  keep_temporary_files = FALSE
)

Arguments

physeq

(phyloseq, required) A phyloseq object with a refseq slot containing DNA sequences.

database

(character, required) Path to the reference FASTA database used by vsearch for chimera detection (e.g. a curated ITS or 16S database).

vsearchpath

(character, default: "vsearch") Path to the vsearch executable. Defaults to "vsearch" assuming it is on the system PATH.

vsearch_args

(character, default: "--fasta_width 0") Additional arguments passed verbatim to the vsearch command line.

return_a_list

(logical, default: FALSE) If TRUE, returns a list with the filtered phyloseq, kept taxa names, and chimeric taxa names.

keep_temporary_files

(logical, default: FALSE) If TRUE, temporary FASTA files written to tempdir() are kept after the function returns.

Value

If return_a_list = FALSE (default), returns a phyloseq object with chimeric sequences removed. If return_a_list = TRUE, returns a list with:

physeq

The filtered phyloseq object

kept_taxa

Character vector of retained taxa names

chimeric_taxa

Character vector of removed chimeric taxa names

Details

Sequences are written to a temporary FASTA file and passed to vsearch using the --uchime_ref algorithm. Sequences classified as chimeras are removed; sequences classified as non-chimeric or borderline are retained (conservative default). Temporary files are cleaned up unless keep_temporary_files = TRUE.

See also

MiscMetabar::chimera_removal_vs() for de novo vsearch-based chimera removal, chimera_removal_dada2() for dada2-based chimera removal, create_chimera_pq() for creating test data with synthetic chimeras.

Author

Adrien Taudière

Examples

library(MiscMetabar)
data(data_fungi)

# Use the mini UNITE database bundled with MiscMetabar.
# NOTE: this is a very small subset of the full UNITE database,
# so a low number of chimeras detected here is expected and does not
# reflect what you would observe with the complete database.
mini_db <- system.file(
  "extdata", "mini_UNITE_fungi.fasta.gz",
  package = "MiscMetabar"
)

data_nochim <- chimera_removal_vs_ref(data_fungi, database = mini_db)
#> Discard 92 chimeric ASVs including:
#> ASV7;ASV26;ASV93;ASV94;ASV129;ASV170;ASV178;ASV209;ASV267;ASV328...
#> Try return_a_list=TRUE to see all discarded ASVs in the `chimeric_taxa` element.

# \donttest{
# Return detailed output including chimeric taxa names
result <- chimera_removal_vs_ref(
  data_fungi,
  database = mini_db,
  return_a_list = TRUE
)
#> Discard 92 chimeric ASVs including:
#> ASV7;ASV26;ASV93;ASV94;ASV129;ASV170;ASV178;ASV209;ASV267;ASV328...
#> Try return_a_list=TRUE to see all discarded ASVs in the `chimeric_taxa` element.
cat("Removed", length(result$chimeric_taxa), "chimeric ASVs\n")
#> Removed 92 chimeric ASVs

# Benchmark against synthetic chimeras created with create_chimera_pq()
pq_chim <- create_chimera_pq(data_fungi, n_chimeras = 10)
result_bench <- chimera_removal_vs_ref(
  pq_chim$physeq,
  database = mini_db,
  return_a_list = TRUE
)
#> Discard 100 chimeric ASVs including:
#> ASV7;ASV26;ASV93;ASV94;ASV129;ASV170;ASV178;ASV209;ASV267;ASV328...
#> Try return_a_list=TRUE to see all discarded ASVs in the `chimeric_taxa` element.
detected <- pq_chim$chimera_names[
  pq_chim$chimera_names %in% result_bench$chimeric_taxa
]
cat(
  "Detected", length(detected), "/", length(pq_chim$chimera_names),
  "synthetic chimeras\n"
)
#> Detected 8 / 10 synthetic chimeras
# }