Abstract: Five strains pathogenic bacteria were isolated from the diseased plants in major sesame production area of Henan province, these Pathogenic bacteria were identified by Koch's Rule and molecular identification. Disinfection technology was optimized through comparison of disinfectants and the disinfection time, sesame seedlings were cultured with best disinfection technology, and the resistance of 12 sesame germplasm resources to Macrophomina phaseolina was identified by inoculating Macrophomina phaseolina in the disinfection seedling of sesame with fungus cake. The study aimed to screen sesame resistant variety for field application and to excavate resistance resources for sesame resistance breeding. The results showed that five strains were all Macrophomina phaseolina (Tassi) Goid, the seed disinfected with 2% NaClO for 20 min produced the best disinfection seedling. The seedling stage resistance was evaluated with fungus cake method and the result demonstrated that only ZZM0905 and ZZM1205 of 12 resources had stable resistance to five different strains, others had different resistance responses.