Although the fertilization effect on soil microbial communities have been studied extensively, its influence combined with plastic-film mulching on seasonal variations of bacterial communities remains unknown. High throughput sequencing was used to explore seasonal bacterial communities in a longterm fertilization experiment that commenced in 1987. Non-fertilization and the application of manure combined with nitrogen fertilization significantly (P<0.05) increased bacterial abundance at phylum level without film mulching in July and with film mulching in October. Also, the relative abundance of dominant bacterial groups in October exceeded those in July. Nitrogen fertilization (N) significantly (P<0.05) decreased soil bacterial diversity and richness with or without film mulching compared to nonfertilization controls during July and October. Plastic film mulching and long-term manure fertilization significantly (P<0.05) increased bacterial diversity and richness as well as the relative abundance of dominant bacterial groups. Statistical analysis revealed that soil pH was the main driving force influencing bacterial groups. In conclusion, our results highlight the relative importance of long-term application of manure fertilization and plastic film mulching on seasonal variations of bacterial diversity and structure. This finding also highlights the relevance of soil pH as the main deterministic abiotic factor for soil bacterial community structure regardless of film mulching.