The gut microbiome plays an important role in healthy infant development, immunity and metabolism. Preterm birth disrupts microbiome development, contributing to acute and chronic disease. To promote microbial and infant development, and to mitigate the risk of disease, premature infants may be treated with probiotics. This study used 16S rRNA high throughout sequencing to characterize the bacterial microbiome of probiotic-supplemented premature infants. The study aimed to identify and understand variation in bacterial gut flora over time, and to explore potential associations with clinical variables.