| Currently,the tomato powdery mildew is becoming more and more severe.In this case,planting the disease-resistant varieties is the most economical and effective solution.The identification of the pathogenic bacteria and the observation of the infection process are the basis for the innovation of the disease-resistant germplasm resources,the breeding of new varieties,and the prevention and control of diseases.In this study,the pathogen isolate of the tomato powdery mildew in Yinchuan area is applied as the test material to conduct morphological identification from the aspects of conidia,germ tube,conidiophores,and appressorium,and the sequence of ITS region is utilized for molecular biological identification so as to the species of pathogen causing tomato powdery mildew in Yinchuan area;the staining effects of the I2-KI staining method,the TTC(2,3,5-triphenyltetrazolium chloride)staining method,the methylene blue staining method,and the fast green staining method are compared to screen out the optimum method to detect the spore vigor of fungus from the tomato powdery mildew;the spore germination rate and disease index of 9 treatments are compared under the three time periods(30 days,60 days and 90 days)through different methods(the cryopreservation method,the glycerol preservation method,and dimethylsulfoxide preservation method)in combination with different preservation conditions,to screen out the optimum preservation method for the bacteria of the tomato powdery mildew;the Coomassie brilliant blue staining is performed on Moneymaker leaves inoculated with powdery mildew at 0 hour,4 hours,8 hours,12 hours,24 hours,48 hours,96 hours,120 hours,and 7 days,and the infection process of the tomato powdery mildew is observed under the light microscope;L9(34)Orthogonal Test is applied to screen out the optimum method for identifying the resistance of the tomato powdery mildew at the indoor seedling stage;on this basis,the disease-resistant germplasm resource evaluation is conducted on 30 tomato germplasm resources.The research results reveal as follows:1.The conidia from the pathogenic bacteria of the tomato powdery mildew in Yinchuan are solitary,oval or drum-shaped;the apical position germinates the bud tube,and the papillary appressorium grows at the end of the germ tube;the appressorium is also living on the hyphae;the conidiophore is upright and unbranched,and no occluded shell is found.According to the morphological characteristics of the above pathogens,it is preliminarily judged to be Oidium neolycopersici;the phylogenetic tree is established through the multiple sequence alignments in the ITS region,whose consistency with the new tomato Onosporium fungus is more than 99%.After the morphological and molecular biological identification,the results consistently show that the pathogen causing tomato powdery mildew in Yinchuan City is the fungus of the new tomato powdery mildew.2.The staining effect of the spore vigor measured through the methylene blue staining method is stable,and the conidial vigor reaches 53.08%,which is not significantly different from the I2-KI staining method and CK,and is the most suitable rapid detection method for the conidia vigor of the tomato powdery mildew;the conidia vigor determined by I2-KI staining method is the highest,which is 54.49%,while the coloring of the I2-KI staining method is uneven,and unfavorable to conduct observation and statistics;the vigor measured by the fast green staining method is the lowest;the TTC staining method has no effect on the staining of the powdery mildew conidia,and the spores are not stained.3.Spore germination rate at 30 d and 60 d was 4 4.2%,22.5 and 39.6%,respectively,significantly higher than the other 8 treatments.At 90 d,the spore germination rate of 4℃ and disease index was 27.5%and 16.3,with no significant difference from-20℃(24 h)to-80 ℃dimethylsulfoxide,which were significantly higher than the other 7 treatments.Of the 8 treatments,the best preservation effect of tomato powdery mildew bacteria was the 4℃ cryogenic preservation method.4.The tomato powdery mildew begins to germinate 4 hours after the inoculation on the leaves,and the top of the conidia germinates to produce germ tubes;at 8 hours after the inoculation,the appressorium grows on the germ tubes;at 12 hours after the inoculation,the haustorium forms under the appressorium;at 24 hours after the inoculation,the primary hyphae are produced;at 48 hours after the inoculation,the secondary hyphae are produced,and the appressorium is also present on the hyphae;at 96 hour after the inoculation,abundant hyphae are formed on the surface of tomato leaves,and conidiophores appear;At 120 hours after the inoculation,the conidia form at the apex of the conidial stalk on hyphae.At this time,the entire growth cycle of the tomato powdery mildew pathogen has completed;at 7 days after the inoculation,a large number of hyphae are formed,and each hypha is continuously branching and spreading radially on the tomato leaves.5.The optimum method for inoculating tomato powdery mildew pathogens in indoor tomato seedling stage is as follows:the inoculation method is the brush leaf method;the seedling age is at the 4-leaf stage;the inoculation concentration is 106 spores/mL,and the moisturizing time is 24 hours.Through the evaluation on the disease-resistant germplasm resources of 30 tomato germplasm resources,the results are as follows:there are 0 immune varietie,1 disease-resistant varieties(62850-1),12 medium-resistant varieties,18 copies of susceptible materials,and 1 copies of highly susceptible materials(62754-1).The above research provides theoretical basis and reference for the control and the disease-resistant breeding for tomato powdery mildew. |
