Effect size of delayed freezing, diurnal variation, and hindgut location on the mouse fecal microbiome relative to a standardized biological variable

While murine fecal collection is central to microbiome research, there are a number of practical considerations that may vary during fecal sample collection, including time to sample storage, time of day the sample is collected, and position within the colon during terminal collections. While the need to control these factors is recognized, the relative effect on microbial community of duration at room temperature, time of day, and hindgut position, in the context of a known biological variable, is unclear. To answer these questions, and assess reproducibility of results across different microbiome compositions, parallel experiments were performed to investigate the effect of those factors on the microbiome of age- and sex-matched isogenic mice colonized with two different vendor-origin microbiomes.