Abstract: This study was conducted to analyze the risk of resistance of Alternaria alternata to azoxystrobin. Resistant mutants were induced by incubating mycelial plugs and mutating conidia on poisoned agar plates. Mutant characterization and mechanism of resistance were also studied and the target gene cyt b was analyzed using partial sequence analysis. Four mutants of A. alternata with resistance to azoxystrobin were isolated. No mutant was obtained through mycelial plug incubation while four mutants were isolated through conidia mutation, with a frequency of around 0.004%. The resistance factors for the four mutants were 42.50, 282.50, 515 and 107.50, respectively. All resistant strains displayed a highly reduced sensitivity to azoxystrobin. Studies of fitness parameters in the wild-type and mutant strains of A. alternata showed that all resistant isolates had equal abilities on conidia germination and virulence, and less ability in mycelial growth. Three mutants showed enhanced ability of conidia production, while the other one decreased its' conidia production. Sequence analysis of the target gene cyt b showed that there was no point mutation at position 129 or 143 on neither the four mutants or the sensitive isolate of A. alternata. This is believed to be the first report of different levels of azoxystrobin resistance in A. alternata. These studies suggested that the risk of resistance development in A. alternata was low for azoxystrobin.