Pathogenic Differentiation And Genetic Diversity Of Fusarium Oxysporum F. Sp. Vasinfectum

Fusarium wilt of cotton, caused by Fusarium oxysporum f. sp. vasinfectum, is widespread in the most of the major cotton-producing areas of the world and causing heavy cotton production losses. To date, eight races of the pathogen have been identified based on a set of differential cotton cultivars in the world, and only races 3, 7 and 8 were reported from China. Race 7 was the most predominant race and distributed throughout cotton growing areas in the china, but this race has a obvious difference in the pathogenicity. Therefore, clarification the pathogenic differentiation and genetic diversity of F. oxysporum f. sp. vasinfectum is valuable for cotton breeding, the distribution of different cotton cultivars, and the integrated control of cotton fusarium wilt.The amplified fragment length polymorphism (AFLP) analysis system was optimized for the DNA digestion with the restriction endonuclease HaeIII/PstI, and the concentration of templates with pre-amplification and selective amplification. Six F. oxysporum f. sp. vasinfectum isolates, including race 3, 7 and 8 and three tested isolates, could be separated clearly by using the AFLP analysis system. The optimized AFLP reaction system for F. oxysporum f. sp. vasinfectum were as followed: 250 ng template DNA was digested by HaeIII/PstI at 37℃for 2 h, add 1μL PstⅠand digested for 2 h further. The digestion was ligated with adaptor at 25℃for 16 h, and the ligation was diluted to 10 times for the pre-amplification, and the pre-amplification production was diluted to 50 times for the selective amplification. And cluster analysis using data generated by AFLP makers showed the system could clearly separate all three F. oxysporum f. sp. vasinfectum races.Genetic diversity of eighty five isolates of F. oxyporum f. sp. vasinfectum was evaluated by AFLP techonology. Great genetic diversity was discovered based on the 0.51 to 1.00 Genetic distance of tested isolates. Phylogenetic tree for the 85 isolates was constructed and four genetic groups (AGsⅠto AGsⅣ) at 0.51 level of Dice similarity coefficients were divided. AGsⅠincluded one isolate of representing physiological race 3, AGsⅡincluded one isolate of representing physiological race 8; AGsⅢincluded one isolate of representing physiological race 7 and seventy four tested isolates, but the great genetic diversities were also observed among different isolates in the AGsⅢphysiological group at the same time. AGsⅣincluded eight isolates, which different from representing physiological races 3, 7 and 8 of F. oxysporum f. sp. vasinfectum from China. AFLP groups were generally related with the different races. However, there was no relationship between the AFLP groups and geographic regions and pathogenicity of F. oxysporum f. sp. vasinfectum isolates.The eight isolates belonging to AGsⅣwere analysised further by the sequences of EF-1α, PHO, and BT genes. All isolates were described for F. oxysporum f. sp. vasinfectum and five isolates had all most consistent sequences homology with the Australian origined isolates, therefore, the five isolates maybe a new genotype of Australian origined race. The remaining three isolates had all most consistent sequences homology with the the race 7, and should be a new genotype of race 7.All the 85 tested F. oxysporum f. sp. vasinfectum isolates were evaluated for the pathogenicity in greenhouse. Results indicated that the pathogenicity were different among isolates when tested against Jimian 11 cotton cultivar and three different pathogenicity groups (strong, middle, weak) were observed. Fifty six isolates belong to the strong pathogenicity group and covered 65.88% to the tested isolates. Twenty isolates belong to the moderate pathogenicity group and covered 23.53% to the tested isolates. Nine isolates belong to the weak pathogenicity group and covered 10.59% to the tested isolates. The five presumable new genotype isolates of Australian origined race were weakly virulent on Jimian 11 cotton cultivar. The remaining three isolates performed diverse virulents.