ABSTRACT: Probiotics are consumed in fermented dairy products or as capsules for their putative health benefits. However, little research has been done to evaluate the effects of the delivery matrix on the health benefits of probiotics in humans. To examine the effects of delivering Bifidobacterium animalis subsp. lactis BB-12 (BB-12) (log10 10 ± 0.5 CFU/day) via a yogurt smoothie versus a capsule, we monitored the fecal microbiota, gut transit times (GTTs), and fecal excretion of short-chain fatty acids (SCFAs) in healthy adults. In a randomized, four-period, crossover study performed in a partially blind manner, 36 adults were recruited and randomly assigned to four treatments: control yogurt smoothie (YS), yogurt smoothie with BB-12 added prefermentation (PRE), yogurt smoothie with BB-12 added postfermentation (POST), and capsule containing BB-12 (CAP). Participants' fecal microbiota was assessed using 16S rRNA sequencing, GTTs via SmartPill, and fecal SCFAs by gas chromatography (GC) before (baseline) and after each intervention. Participants had significantly higher percentage of Streptococcus after consuming YS versus CAP (P = 0.01). Bifidobacterium-specific terminal restriction fragment length polymorphism analysis revealed a significantly higher percentage of B. animalis after consuming PRE and POST compared to baseline, YS, CAP, and final washout (P < 0.0001). The predominant SCFAs were negatively correlated with GTTs. Consumption of BB-12 delivered in a yogurt smoothie or capsule did not significantly alter the composition of the gut microbiota, GTTs, or fecal SCFA concentration of the study cohort. However, daily consumption of BB-12 in yogurt smoothie may result in higher relative abundance of B. animalis in healthy adults. (This trial has been registered at ClinicalTrials.gov under identifier NCT01399996.) IMPORTANCE Bifidobacterium animalis subsp. lactis BB-12 is a probiotic strain that has been used worldwide since 1985. It has commonly been delivered in fermented dairy products for perceived benefits associated with gut health and enhanced immune function. In addition to fermented dairy products, many new probiotic-containing alternatives such as probiotic-containing juice, probiotic-containing chocolate, and capsules have been developed. While these products provide more options for people to access probiotics, little research has been done on the effect of delivery matrix (dairy versus nondairy) on their efficacy in humans. In addition, it was unclear how yogurt fermentation may influence the survival of BB-12 in the product or on its performance in vivo. The significance of our study is in simultaneously assessing the effect of BB-12, alone and in different delivery vehicles, on the gut transit time, fecal short-chain fatty acids, and the composition of the gut microbiota of the study cohort.