Abstract: In order to screen highly effective antagonistic strains against tobacco Fusarium spp., the antagonistic bacterium were isolated and screened from the rhizosphere soil of tobacco plants in Pingdingshan tobacco-growing area by plate confrontation, pot culture and field screening tests. The antagonistic bacterium were identified by morphological observation, physiological and biochemical tests and 16S rDNA sequence analysis. The mechanism of antagonistic bacteria against Fusarium spp. root rot was also investigated. The results showed that a total of 512 soil microbial strains were isolated from 10 soil samples, of which strain L210 had the best inhibitory effect on Fusarium spp. root rot with a bacteriostatic diameter in plate confrontation test of 71.4 mm and an inhibitory rate of 68.2%. The disease incidence rate in pot and field tests were 0 and 6%, the disease index was 0 and 1.5, and the control effect was 100% and 77.9%, respectively. L210 was identified as Serratia liquefaciens, and the antibacterial mechanism was on the growth inhibition of pathogenic bacteria, dissolution of Fusarium oxysporum mycelia, which caused the pathogen to lose its infectivity. The fermentation broth contained chitinase and cellulase, which could dissolve the cell wall of pathogenic bacteria; the fermentation broth of this strain could also significantly induce and increase the activities of defense enzymes such as polyphenol oxidase (PPO), peroxidase (POD), and phenylalanine ammonia-lyase (PAL). Strain L210 thus has great potential for biological control of Fusarium root rot in tobacco.