To deduce the antibacterial mechanism of ethyl acetate extract of Amaranthus tricolor L. against Xanthomonas axonopodis pv.citri, the antibacterial activity of ethyl acetate extract of Amaranthus tricolor L. against X. axonopodis pv. citri was determined and the effects of ethyl acetate extract of Amaranthus tricolor L. on OD600nm, conductivity, protein content and respiratory metabolism enzyme activity of X. axonopodis pv. citri strain liquid were investigated. The results show that the inhibition zone diameter on X. axonopodis pv. citri with ethyl acetate extract of Amaranthus tricolor L. was above 15 mm, with MIC 3.68 mg/mL. With ethyl acetate extract of Amaranthus tricolor L., the growth of X. axonopodis pv. citri was delayed, and the conductivity of X. axonopodis pv. citri was increased by 15% and enzyme activities of POD, CAT, SDH and MDH were inhibited by 78.43%, 68.63%, 63.14%, 68%, respectively. Scanning electron microscopy showed that cell membrane of X. axonopodis pv. citri was damaged, the bacteria was lysed, and the cell was cracked with ethyl acetate extract of Amaranthus tricolor L.. Transmission electron microscopy showed that ethyl acetate extract of Amaranthus tricolor L. caused the damage of cell wall and cell membrane structure of X. axonopodis pv. citri, and caused the increase of intracellular blank area, the leak of cytoplasm, and the lysis of bacteria.