Abstract: Anaerobic fermented organic materials fluid (AFOF) could alleviate apple replant diseases. In this study, bacteria from AFOF was screened which could antagonize the main pathogenic fungi (Fusarium solani, F. proliferatum, F. oxysporum, F. moniliforme). The effects of the antagonistic bacteria were verified by pot experiment. The results showed that AFOF could significantly inhibit the pathogenic fungi growth and reproduction. Four strains (L11, L12, L13, L14) with strong inhibition activity were obtained from AFOF by the confrontation test, with the highest inhibitory rate being up to 57.3%. All the four strains were Bacillus, with no obvious antagonism each other. Both antagonistic bacteria and methyl bromide fumigation could promote the biomass of Malus hupenhensis seedling compared with continuous cropping soil. Methyl bromide fumigation led to the best growth of M. hupenhensis seedling. The antagonistic bacteria cloud improve root activity, with root length and root tip number increased by 25.1% and 70.9%, respectively. Compared with the continuous cropping soil, antagonistic bacteria and methyl bromide fumigation effectively decreased soil fungi by 71.2% and 64.2%, respectively. Treatment with antagonistic bacteria increased soil bacteria and soil actinomycetes by 48.0% and 140.2%, respectively, indicating that soil microbial community was transformed to “bacteriatype”. Treatment with methyl bromide fumigation could significantly reduce soil bacteria and soil actinomycetes, suggesting the antagonistic bacteria could inhibit the growth of pathogenic fungi in soil.