Fatty acids of Ralstonia solanacearum cultured under different temperatures, times, pH values and cultural media were detected by using gas chromatography (GC) method. Rs-J.1.4-010704-01v, a virulent strain of R. solanacearum isolated from ginger was chosen for the experiment. The results showed that the kind of fatty acid of Rs-J.1.4-010704-01v fluctuated from 14 to 33. The contents of its three plentiful fatty acids, C16:1ω7c/C15:0 ISO 2OH, C16:0 and C18:1ω7c (with retain times of 10.644, 10.950 and 14.177 min, respectively), also varied in a range of 55.66% to 75.69%. The diversification of the bacterium’s fatty acids at various cultural conditions was clustered into four groups by cluster analysis, according to the kinds and percentage contents of the fatty acids detected. The pathogenicities of Rs-J.1.4-010704-01v under 20°C and 25°C were deduced to be mid-virulent, with C16:0 less than C16:1ω7c/C15:0 ISO 2OH. The bacterium showed as a virulent strain under the other cultural conditions including 30°C~40°C, 24 h~96 h, pH 5~9 and four cultural media (LB、NA、TTC and TSB), with C16:0 more than C16:1ω7c/C15:0 ISO 2OH. However, the difference between C16:0 and C16:1ω7c/C15:0 ISO 2OH raised significantly from 2.35 to 13.23 under 40°C and 48 h~96 h. Meanwhile, the kind of fatty acid increased more than 30 as the cultural time increased. It was concluded that temperature and cultural time had more significant effects on the fatty acid composition of R. solanacearum than pH value and cultural medium.