Hsp Genes Expression Level Of Blumeria Graminis F. Sp. Tritici Under Heat Stress And Host-induced Gene Silencing (HIGS) System Establishment

Temperature is an important environmental factor influencing epidemic of diseases,that not only influences the occurrence of diseases,but also the evolution of the pathogen population.Wheat powdery mildew caused by Blumeria graminis f.sp.tritici(Bgt),an obligate parasitic fungus,is one of the important wheat diseases.The previous research showed that high temperature tolerant isolates of Bgt had been produced in the natural population in china.In this study,based on determining temperature sensitivity of 55 Bgt isolates collected from Beijing,Henan,Shaanxi and Yunnan provinces from 2012 to 2013,five temperature sensitive isolates and four temperature tolerant isolates were screened,and the infection process of Bgt isolates with different temperature sensitivity under optimum temperature(18℃)and high temperature(25℃)was observed by histology and staining technique;The expression of heat shock protein genes Hsp40,Hsp70,Hsp90 and Hsp104(BgtHsp40,BgtHsp70,BgtHsp90 and BgtHsp104)of Bgt isolates with different temperature sensitivity was analyzed by using Real-time RTPCR,and the correlation between BgtHsp40,BgtHsp70,BgtHsp90 and BgtHsp104 and high temperature tolerance of Bgt isolates was explored.At the same time,the BSMV-HIGS system for gene function of Bgt in response to high temperature was established.These studies laid a foundation for a further study on genes of wheat powdery mildew pathogen relating to heat resistance and their functions.The main research results are as follows:1.Based on determining the temperature sensitivity of 55 Bgt isolates collected from Beijing,Henan,Shaanxi and Yunnan provinces from 2012 to 2013 by setting 5 different temperature gradients,five temperature sensitive isolates including 13-14-7-2-(2),13-10-11-1-(1),13-11-4-2-2-(1),13-1-1-(1),13-1-5-5-1-(2)and four temperature tolerant isolates including 13-1-4-1-1-(1),13-14-8-2-(2),13-10-3-2-(2),13-11-4-2-1-(1)were screened.2.The results of observing the infection process of the sensitive temperature isolate 13-14-7-2-(2),13-11-4-2-2-(1)and the temperature tolerant isolate 13-14-8-2-(2),13-11-4-2-1-(1)under high temperature stress,showed that high temperature had an inhibitory effect on spore germination of Bgt isolates,and with the extension of high temperature stress time,the inhibition was increasingly obvious,but there was no significant difference between spore germination rate of the temperature sensitive isolates and that of temperature tolerant isolates under high temperature stress;High temperature also affected the formation of normal appressorium of Bgt isolates.The formation rate of normal appressorium of Bgt isolates under high temperature stress was lower than that under normal temperature.However,the normal appressorium formation rate of the temperature tolerant isolates was no significantly different from that of the temperature sensitive isolates under high temperature stress.That high temperature also significantly inhibited the formation of haustorium of Bgt isolates,and the haustorium’s formation rate of sensitive isolates under high temperature stress were significantly lower than those of temperature tolerant isolates.3.The results of analysing the expression of BgtHsp40,BgtHsp70,BgtHsp90 and BgtHsp104 of Bgt isolates with different temperature sensitivity showed that under high temperature stress at 6 hour post inoculation(hpi)(Spore germination stage),24 hpi(Appressorium formation stage)and 48 hpi(Haustorial formation stage)the relative expressions of BgtHsp40,and BgtHsp70 of temperature tolerant isolates were higher than those of temperature sensitive isolates,but the relative expression of BgtHsp40 and BgtHsp70 of temperature tolerant isolates were not significantly different from those of temperature sensitive isolates.Therefore,BgtHsp40 and BgtHsp70 could play an important role in the high temperature resistance of Bgt under high temperature stress during and before the haustorium formation stage,while BgtHsp90 and BgtHsp104 could play a minor role.4.The BSMV-HIGS system for the genes function study of wheat powdery mildew was preliminarily established by analyzing the affinity of “Xue Zao” to BMSV virus.The relative expressions of BgtTUB and the severity of wheat leaves infected by Bgt isolates were significantly lower than those of the control group,the efficiency of silence is up to 77%.