Background and aims Resistant cultivars are one of the most effective control measures used against bacterial wilt, caused by the soilborne bacterium Ralstonia solanacearum. We aimed to determine the effect of temperature and resistance of tobacco varieties on bacterial wilt occurrence. Methods Five tobacco cultivars, with varying resistance levels, were inoculated and transferred to growth chambers at 10, 15, 20, 25, 30, and 35°C. The growth rate of R. solanacearum was also studied in culture at 10, 20, and 30°C. The mechanism of resistance was further examined in histological studies conducted at 10, 20, and 30°C with strain AW1-gfp38 that expresses green fluorescent protein.Results The highest disease incidence was observed at 30 and 35°C, while no symptoms were observed at 10 and 15°C. Strains grew in culture at all temperatures. At 30°C the low resistant cultivars had the most stems colonized by the bacterium, while the highly resistant cultivars mainly had localized infections in the roots. Conclusions We suggest that the mechanism of resistance in tobacco is associated with the ability to limit colonization of stem tissues and is temperature dependent. Evaluation of restricted root infections in breeding lines may provide a means for early screening of resistance in breeding programs.