High-throughput sequencing of the 16S rRNA gene of microbial communities was used to investigate how biofloc technology (BFT) improved water quality and fish yield in freshwater pond aquaculture. Three treatments included conventional treatment (CT), biofloc treatment (BT) and biofloc systems augmented with floc-forming bacterium (Zoogloea resiniphila MMB) treatment (BZT). The feeding trial lasted for 8 weeks with triplicate groups of cyprinid species in nine enclosures, which were placed in an outdoor pond to simulate a real aquaculture environment. Biofloc was generated by adding organic carbon and continuous aeration, which resulted in reducing dissolved nitrogen from 22% to 14% and sedimentary nitrogen from 49% to 5% and increasing nitrogen accumulation in biofloc and fish biomass. Nitrogen content in fish biomass increased from 28% in a CT to 42% when BFT was applied. Changes in the bacterial community and stimulation of biofloc formation played a central part in transferring nitrogen from remaining food, excreta and feces to bacteria and fish biomass. Z. resiniphila MMB did not outcompete indigenous communities and had no impact on floc formation by direct addition to the pond. Filamentous bacteria comprised the major structure of biofloc, while the proportion of zoogloea-forming microorganisms was very low. The proportion of aerobic heterotrophic microorganisms increased in both BFT treatments. Actinobacteria promoted the formation of floc and could have secondary benefits of protecting against fish pathogens, such as Aeromonas sp., though may increase the accumulation of off-flavor in the water and fish tissue.