Field Investingation Of Wheat Wellow Mosaic Disease During 2015-2016 And Prelinginary Establishment Of Sand Culture System Of Polymyxa Graminis

Polymyxa graminis is a soil-borne,obligate intracellular parasite of plant roots which can transmits more than 10 agronomically important viruses to cereal crops.Wheat yellow mosaic virus(WYMV)and Chinese wheat mosaic virus(CWMV),both transmitted by P.graminis,have been identified as the disease agents in winter wheat regions of China,and outbreak of wheat yellow mosaic disease has a serious impact on the safety of wheat production.Confirmation the distribution of wheat yellow mosaic disease in China is important for the control of the disease.During March 2015 and 2016,more than 200 suspected wheat disease samples from the fields were collected or posted from the regions including 7 provinces in China and were detected for the presence WYMV and/or CWMV with RT-PCR in the lab.Our results indicated that the distribution of WYMV is similar compare to the previous years,however,more serious symptoms were observed in Zhumadian and Linyi(Henan province)during the investigation.Beside Rongcheng and Yantai(Shandong province)which is known to be infected with CWMV,wheat samples from Linyi and Tai’an(Shandong province)were also identified to be mixed infected with CWMV and WYMV.It is the first report of CWMV detected in Linyi and Tai’an(Shandong province),indicating a trend of proliferation for CWMV in Shandong province.Due to the limited number of wheat cultivars that shows resistant to CWMV,screening of wheat germplasm resources resistant to CWMV needs to be accelerated.Screening and planting resistant wheat varieties is now proved to be the only ecological friendly and effective method for the control of wheat yellow mosaic disease in field.At present,the mechanism for resistant varieties to virus is mainly focused between plant and virus,whereas little attention has been paid to the transmission vector P.graminis.In this study,a real-time fluorescence quantitative PCR method(absolute quantification)was established and applied for the quantification analysis of P.graminis and WYMV for five mediums resistant(soil and roots)and three high susceptible(leaves and roots)wheat cultivars.Our results showed that there was no significant difference for the copy number of P,graminis in the rhizosphere soil of medium resistant and high susceptible wheat cultivars.The content of P.graminis in the roots of four medium resistant wheat cultivars,Yannong 999,Dongpin 10,Linxuan 1309 and Yannong 24,was significantly lower than that of the other two high susceptible cultivars,Linmai 4 and Qian 0513.However,there was no significant difference in the copy number of P.graminis between the medium resistant cultivar Tainong 18 and these two high susceptible cultivars.Furthermore,the content of WYMV in leaves and roots of wheat showed clear correlation between leaves viral titer and wheat cultivar resistance,but no obvious linear relationship was observed between root content of P.graminis and viral titer of wheat plant.Our results preliminarily elucidated that wheat cultivars exhibit different resistance to P.graminis,and the resistance showed no clear correlations with disease symptom and viral titer of the wheat plants.Due to obligate nature of P.graminis,it’s important to establish stable sand culture system for the maintenance of P.graminis and transmission of the virus under lab condition to a better understanding the relationship within the vector,plant host and viruses.In this study,a preliminary sand culture system for P.graminis infection is established using wheat roots infected with P.graminis and viruses collected during 2015-2016 as inoculum.Though we failed to observe resting spores of P.graminis in wheat roots in the late stage of infection and no clear disease symptoms was observed in wheat leaves during the infection,our molecular detection clearly indicated the successful infection of P.graminis to the root of healthy wheat and transmission of viruses(WYMV and/or CWMV)from vector to wheat plant the newly established sand culture system.Further experiments showed that CWMV might be more infective to susceptible wheat plant than WYMV with increased proportion of infected plants under same condition.Temperature gradient experiments(9℃,15℃and 22 ℃)revealed that low temperature conditions(9℃)contributed to the infection for both P.graminis and viruses(abbreviated infection times).