Free-living (FL) and cell-associated (CA) microbial communities play important roles in the outburst, maintenance and decline of harmful algal blooms (HABs). However, there is still limited knowledge about how FL and CA bacterial communities differ and change during bloom succession and whether the phycosphere bacterial community of an algal species remains similar in different geographic populations. In this study, 16S rRNA gene amplicon sequencing of size-fractionated samples was employed to investigate the community assembly mechanism, dynamic shifts and co-occurrence patterns of both FL and CA bacterial communities at different stages during two independent Prorocentrum shikokuense (Dinophyceae) bloom events (2014 and 2018) in the East China Sea. The CA and FL communities exhibit distinct structure composition and dynamics changes, influenced by varying degrees of assembly processes and environmental factors. Proteobacteria, Bacteroidota, Actinobacteriota and Planctomycetota represented four core phyla, accounting for 81.8-95.8% relative abundance of the total observed amplicon sequence variants (ASVs) in both events. Among these, Proteobacteria and Bacteroidota have also been reported to be associated with P. shikokuense, suggesting they are “resident” bacteria in this dinoflagellate’s phycosphere. The CA community structure changed more evidently during algal bloom development compared to the FL community, sharing a similar community structure despite spatial and temporal differences. Meanwhile, environmental factors account for a higher proportion of the variance in FL communities. Homogeneous selection had a more significant impact on shaping the composition of the FL communities, whereas the CA communities were more strongly influenced by stochastic processes, mainly dispersal limitation. The common taxonomic characteristics and ecological functions of the CA communities between the two events were further explored. Results showed that the diversity and structure of CA bacterial communities were determined by residence space rather than geographic locations or bloom stages, with simpler co-occurrence networks compared with the FL communities. Moreover, in the CA community, the ecological function was significantly enriched in nutrient cycling and cytolysis. The ecological implications of the elevated abundance of Rhizobales regarding nitrogen-fixation are also discussed. Our work unraveled the different assembly and dynamic composition between the FL and CA communities. The findings derived from two independent bloom events provided more in-depth insight for further studies of algal-bacteria interactions.

Harmful Algal Blooms (HABs),bacterial community diversity,cell-associated bacteria,community assembly mechanism,Phycosphere