Ironing out the wrinkles in the rare biosphere through improved OTU clustering

Deep sequencing of PCR amplicon libraries facilitates the detection of low-abundance populations in environmental DNA surveys of complex microbial communities. At the same time, deep sequencing can lead to overestimates of microbial diversity through the generation of low-frequency, error-prone reads. Even with sequencing error rates below 0.005 per nucleotide position, the common method of generating operational taxonomic units (OTUs) by multiple sequence alignment and complete-linkage clustering significantly increases the number of predicted OTUs and inflates richness estimates. We show that a 2% single-linkage preclustering methodology followed by an average-linkage clustering based on pairwise alignments…

• NS
  National Science Foundation

  Awards: NSF/OCE 0430724, 0430724
• AP
  Alfred P. Sloan Foundation
• UO
  University of Michigan
• NI
  National Institutes of Health