CopyRighter: a rapid tool for improving the accuracy of microbial community profiles through lineage-specific gene copy number correction

Abstract

BackgroundCulture-independent molecular surveys targeting conserved marker genes, most notably 16S rRNA, to assess microbial diversity remain semi-quantitative due to variations in the number of gene copies between species.ResultsBased on 2,900 sequenced reference genomes, we show that 16S rRNA gene copy number (GCN) is strongly linked to microbial phylogenetic taxonomy, potentially under-representing Archaea in amplicon microbial profiles. Using this relationship, we inferred the GCN of all bacterial and archaeal lineages in the Greengenes database within a phylogenetic framework. We created CopyRighter, new software which uses these estimates to correct 16S rRNA amplicon microbial profiles and associated quantitative (q)PCR total abundance. CopyRighter parses microbial profiles and, because GCN estimates are pre-computed for all taxa in the reference taxonomy, rapidly corrects GCN bias. Software validation with in silico and in vitro mock communities indicated that GCN correction results in more accurate estimates of microbial relative abundance and improves the agreement between metagenomic and amplicon profiles. Analyses of human-associated and anaerobic digester microbiomes illustrate that correction makes tangible changes to estimates of qPCR total abundance, α and β diversity, and can significantly change biological interpretation. For example, human gut microbiomes from twins were reclassified into three rather than two enterotypes after GCN correction.ConclusionsThe CopyRighter bioinformatic tools permits rapid correction of GCN in microbial surveys, resulting in improved estimates of microbial abundance, α and β diversity.

Allelochemicals Produced by Brown Macroalgae of the Lobophora Genus Are Active against Coral Larvae and Associated Bacteria, Supporting Pathogenic Shifts to Vibrio Dominance

“…A taxon summary was created using QIIME's summarize_taxa_th-rough_plots.py (93). A second OTU table was generated using the CopyRighter tool to improve the accuracy of microbial community profiles (94).…”

Evaluating community–environment relationships along fine to broad taxonomic resolutions reveals evolutionary forces underlying community assembly

“…To avoid inaccurate taxonomic assignments because of short reads, we only used OTUs with significant hits (similarity 40.97 and coverage 40.97) to the GreenGenes otu97 core set for community profiles. Then, we used CopyRighter (Angly et al, 2014) to adjust relative abundance profiles, which correct the bias of the 16S rRNA gene copies among different prokaryotic genomes (Acinas et al, 2004;Vetrovsky and Baldrian, 2013). This initial site-byotu97 matrix was then aggregated into a series of community matrices with fine to broad taxonomic resolutions, according to GreenGenes OTU linkage maps (DeSantis et al, 2006b) and GreenGenes explicit taxonomic ranks (McDonald et al, 2012).…”

Microbial Community Analysis with Ribosomal Gene Fragments from Shotgun Metagenomes

“…There are up to 15 SSU rRNA gene copies in some bacteria and up to 5 in archaea (59). This pipeline utilizes the SSU rRNA copy database in CopyRighter (36). As expected, Firmicutes in soil samples have the highest fold change (see Fig.…”

“…We used the SSU rRNA gene copy number database in CopyRighter (36), which provides the copy number for each taxon in the Greengenes database (37). In the taxonomic summary, the abundance of each taxon is weighted by the inverse of its SSU rRNA gene copy number.…”