Impact of DNA Sequencing and Analysis Methods on 16S rRNA Gene Bacterial Community Analysis of Dairy Products

Abstract

Validated methods are urgently needed to improve DNA sequence-based assessments of complex bacterial communities. In this study, we used 16S rRNA PCR amplicon and gDNA mock community standards, consisting of nine, dairy-associated bacterial species, to evaluate the most commonly applied 16S rRNA marker gene DNA sequencing and analysis platforms used in evaluating dairy and other bacterial habitats. Our results show that bacterial metataxonomic assessments are largely dependent on the DNA sequencing platform and read curation method used. DADA2 improved sequence annotation compared with QIIME 1, and when combined with the Ion Torrent PGM DNA sequencing platform and the Greengenes database for taxonomic assignment, the most accurate representation of the dairy mock community standards was reached. This approach will be useful for validating sample collection and DNA extraction methods and ultimately investigating bacterial population dynamics in milk- and dairy-associated environments.