Artiifcial Inoculation Techniques Of Rice False Smut And Analysis Of Genetic Diversity Of Ustilaginoidea Virens

The Rice False Smut caused by Ustilaginoidea virens (Cooke) Tak was a spikedisease in rice, which not only lead to a great loss in rice yields and qualities, but alsomade people or animals disabled or dead by the fungal toxin. In recent10years, RiceFalse Smut has been a dominant disease in different rice producing regions, andgreatly influenced the rice production.In order to understand the biological characteristics of Ustilaginoidea virens,the mycelial growth and conidia germination were tested in various culture conditions,including different media, temperature and pH value. The results showed that PSA(potato sucrose agar) was the optimal medium for pathogen growth. The optimaltemperature and pH value for mycelial growth, chlamydospore germination andconidia germination were28℃and6, respectively. When exposed to temperature of15℃to33.5℃and those of13℃-23℃for48h, the conidia germination rates were81.79%and44.20%respectively. The conidia germination rate was reduced to43.38%after treated by low temperature (4℃) for168h. When exposed to hightemperature of36℃-42℃for1-6h, the pathogen grew worsely and the colony inplates became smaller. The germinated conidia could not been found after treated byhigh temperatures over40℃, The germinated chlamydospore could not been detectedafter they exposed to41℃for4h or more.The dry weight of pathogen mycelial reached its peak (16.48mg/ml) whencultured in PDB for13days and the conidia number was most abundent whenpathogen cultured for9days. However, the dry weight of mycelia, conidia number,and pH value obviously decreased with the prolong of culture period, and the culturesexhibited inhibition effects on the germination of rice seeds. The extracts from paniclein booting stage and preliminary grout stage of rice varieties 'Liangyoupeijiu','Shanyou63' and 'Yongyou6' significantly promoted conidia germination, butinhibited chlamydospore germination.The hyphal growth rates, sporulating abilities and conidia germination abilitiesof102isolates from rices in different growing regions were examined. The results showed that the hyphal growth rates of all isolates ranged from28.7mm/15d to45mm/15d, the sporulating number varied from2.5×10₄/mL to2.4×10₇/mL, andconidia germination rates ranged from9%to97.5%.To understand the factors influencing inoculation effects, the inoculation effectsof different inoculums including those cultured in different media, those cultured fordifferent time course and those inoculated at different concentrations were tested,meanwhile the inoculation effects of pathogens inoculated into rice at different lifestages were examined as well. It was found that the pathogen cultured in PSB mediumcaused100%panicles infected. Inoculation effect of the pathogens cultured in PSBmedium for5-7d was better than other treatments. The lower conidia concentration ofinoculums resulted in lower percentages of diseased panicles and diseased grains.When rice variety "Liangyoupeijiu" inoculated with pathogen (at a concentration of4×10₆conidia/mL) at the middle and later stages of booting, the percentage ofdiseased panicles was100%. In booting stage, the inoculation effect was the bestwhen the ligule distance reached8cm-9cm. The spraying method can improve theinoculation effect. In the six rice varieties, Liangyoupeijiu was infected most seriouslyand Ⅱyouming86was attacked with milder symptoms. The resistance of15rice newvarieties from Fujian province was tested against rice false smut by using the methodsof artificial injected inoculation in greenhouse and natural inducing in field,respectively. The results showed that the resistance to rice false smut and rice neckblast disease of artificial injected in greenhouse was better than those of naturalinducing in field, and the average percentage of diseased panicles and the number ofdiseased grains were36.56%and4.2, respectively. The disease effect was more stablewith the techniques of artificial inoculation, which could be used to distinguishdifferences of resistance to rice false smut among rice varieties in greenhouse.In order to know the relationship among genetic diversity of Ustilaginoidea virens,rice varieties, rice growing season and geographic origin,102isolates of the pathogencollected from19cities or counties in Fujian province were assessed withRAPD(random amplified polymorphic DNA) and cluster analysis. A total of157bands were amplified by RAPD primers from different isolates, with the percentage of polymorphic bands of82.17%and genetic distance between0.02⁰.66. The102isolates could be classified into7genetic groups at0.349genetic distance level, andR1was the predominant group containing80isolates, in which some subgroupsexisted.10isolates from Ninghua could be classified into2genetic groups, earlyseason and late season, at0.330genetic distance level. The preliminary analysisshowed that geographic origin of isolates, rice varieties, their growing season andgrowing years might be the main factors influencing the genetic diversity of U. virensin Fujian, and play an important role in genetic variation of the rice false smut.