Optimizing 16S rRNA gene profile analysis from low biomass nasopharyngeal and induced sputum specimens

High throughput sequencing of the 16S rRNA gene poses a risk for biased or inacurate bacterial profiling. This study demonstrates the value of the inclusion, and detailed exploratory analyses, of sequencing controls to reduce the experimental error associated with 16S rRNA gene high throughput sequencing profiles from respiratory specimens. Our study aimed to 1) compare two DNA extraction methods for 16S rRNA gene sequencing using high and low biomass bacterial mock communities, each generated using two storage buffers; and to 2) evaluate the quality of 16S rRNA sequencing data from low-biomass nasopharyngeal and induced sputum specimens. We also aimed to 3) address the use of the decontam package, implemented through R software, to correct for potential contamination inherent to 16S rRNA sequencing.