A Quantitative Sequencing Framework for Absolute Abundance Measurements of Mucosal and Lumenal Microbial Communities

Abstract

A fundamental goal in microbiome studies is to determine which microbes affect host physiology. Standard methods for determining changes in microbial taxa measure relative microbial abundances, which cannot capture absolute changes. Moreover, studies often focus on a single site (usually stool), although microbial demographics differ substantially among gastrointestinal (GI) locations. Here, we developed a quantitative framework to accurately measure absolute abundances of individual bacterial taxa by combining the precision of digital PCR with the high-throughput nature of 16S rRNA gene amplicon sequencing. In a murine ketogenic-diet study, we compared microbial loads in lumenal and mucosal samples at several sites along the GI tract. Measurements of absolute (but not relative) abundances revealed decreases in total microbial loads on the ketogenic diet and enabled us to accurately determine the effect of the diet on each taxon at each GI location. Quantitative measurements also revealed different patterns in how the ketogenic diet affected each taxon's abundance in stool and small-intestine mucosa samples. This rigorous quantitative microbial analysis framework applied to samples from relevant GI locations will enable mapping microbial biogeography of the mammalian GI tract and more accurately capture the changes of microbial taxa in experimental microbiome studies.

Bacteriophages Could Be a Potential Game Changer in the Trajectory of Coronavirus Disease (COVID-19)

“…42 If needed, quantitative microbiome sequencing could potentially be used. 43 There are assumptions that need to be met during the clinical trials for the approach to work. (1) The cohort has to be chosen to have a high probability of developing bacterial infections.…”