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Abstract: The application method for a novel bioorganic fertilizer (BIO) was developed to improve its biocontrol efficacy of Fusarium wilt (Ling et al. 2010). However, its efficacy on controlling Fusarium wilt and the variations of microbial community after long-term application for watermelon production had not been elucidated. To clarify, a 4-years pot experiment of mono-cropping watermelon was conducted. The results revealed that though the disease incidences were increased in all treatments with the increase of continuous cropping years, the treatment of BIO application both in nursery and pot soil always maintained the lowest disease incidence. The real-time PCR results showed that the population of Paenibacillus polymyxa was decreased with continuous cropping years, but in all seasons, the treatment with BIO application both in nursery and pot soil had a highest population of P. polymyxa than the other treatments. On the other hand, the abundance of the pathogen FON was increased with the increase of continuous cropping years and the lowest rate of increase was found by BIO application in both nursery and pot soil. DGGE patterns showed that the bacterial diversity was weakened after mono-cropping of watermelon for 4 years, but the consecutive applications of BIO at nursery and transplanting stage resulted in the minimal change of bacterial diversity. More detailed differences on bacterial diversity between control and double application of BIO treatment after 4-years monoculture were analyzed by 454 pyrosequencing, which showed the dominant phyla found in both samples were Firmicutes, Proteobacteria and Actinobacteria, and the consecutive applications of BIO recruited more beneficial bacteria than control, such as Bacillus, Paenibacillus, Haliangium, Streptomyces. Overall, these results, to a certain extent, approved that the consecutive applications of BIO at nursery and transplanting stage could effectively suppress watermelon Fusarium wilt by regulating the rhizosphere bacterial diversity. These results could give some clues that how to regulate the soil microbial community to an appropriate level which can keep the plant healthy and thus control the soil-borne diseases. [Copyright &y& Elsevier]