In Japan, indigo-dyeing has traditionally been performed by natural fermentation. The indigo remains in a reduced state for 6 months, on average, although the fermentation fluid has a greater chance of being contaminated in an open environment, especially during the dyeing process. This fermentation system has several advantages for investigations on certain aspects of microbial ecology such as the functions of individual microorganisms and interactions between individual microorganisms. To understand the mechanisms underlying the changes in the fermentation phase that are concomitant with the indigo reduction, we made 2 batches of indigo fermentation fluid. The bacterial V4 region of 16S rRNA genes were PCR amplified for next-generation sequencing.