Effects Of Boscalid On The Growth And Virulence Of Sclerotinia Sclerotiorum

Sclerotinia sclerotiorum?Lib.?de Bary is an ascomycetous plant pathogenic fungus with a broad host range.The primary method to manage S.sclerotiorum is chemical control.In this thesis,we investigated the inhibitory activities and hormetic effects of boscalid on mycelial growth and virulence of S.sclerotiorum.These studies have important implications for judicious application of boscalid to control S.sclerotiorum.To establish the baseline sensitivity of S.sclerotiorum to boscalid,the effective concentrations causing mycelial growth inhibition by 50%(EC₅₀ values)of isolates collected in Anhui Province were determined.The mean EC₅₀0 values of 76 isolates collected in 2008 and 77 isolates collected in 2014 were 0.0383±0.0196?standard deviation,SD??g/mL and 0.0395±0.0180?SD??g/mL,respectively.There was no significant difference?P=0.695?in boscalid sensitivity between 2008 and 2014,and the frequency distribution of EC₅₀ values was unimodal.Correlation analysis showed that there was a statistically significant positive correlation?R²=0.856,P<0.001?of sensitivity between boscalid and thifluzamide,but no significant correlation?R²=0.010,P=0.650?between boscalid and prochloraz was obtained.Both preventive and curative efficacies of boscalid were significantly higher than those of the carbendazim at the same concentrations.Boscalid at0.04 to 0.16?g/mL in PDA significantly reduced the number of sclerotia of S.sclerotiorum?P<0.05?,but the weight of sclerotia increased slightly?P=0.084?.The activities of succinate dehydrogenase?SDH?and superoxide dismutase?SOD?were decreased for mycelia grown on PDA amended with boscalid at 0.01 and 0.05?g/mL,and the hyphae were contorted and branched more.Furthermore,mitochondria of hyphal cell were deformed.However,boscalid at 0.01 and 0.05?g/mL had no significant effect on mycelial cell membrane permeability of S.sclerotiorum.Sublethal doses of boscalid had stimulatory effects on mycelial growth and virulence of S.sclerotiorum.Boscalid at 0.0005 to 0.002?g/mL in PDA had stimulatory effects on mycelial growth of isolate HLJ3H and the percent stimulation ranged from 2.3%to 5.1%.Boscalid sprayed at 0.0001 to 0.1?g/mL increased the virulence of isolate HLJ3H by 0.6%to 18.7%.The maximum stimulation amplitudes of boscalid on mycelial growth and virulence for dimethachlone-resistant isolates were lower than those of dimethachlone,whereas the maximum stimulation amplitudes of boscalid for sensitive isolates were similar to those of dimethachlone.The maximum percent stimulations of boscalid on mycelial growth of dimethachlone-resistant isolates HLJ3H and HLJ4H were 5.1%and 5.9%,respectively,while the maximum percent stimulations of dimethachlone were 48.7%and13.9%,respectively.The maximum percent stimulations of boscalid on mycelial growth for sensitive isolates HLJMG1 and JD391 were 6.4%and 4.7%,respectively,and the maximum percent stimulations of dimethachlone were 5.3%and 5.2%,respectively.Boscalid sprayed alone at 0.1?g/mL and dimethachlone sprayed alone at 40?g/mL increased the virulence of isolate HLJ3H by 2.9%and 43.3%respectively,however,the mixture of boscalid at 0.1?g/mL and dimethachlone at 40?g/mL reduced the virulence by2.9%.Compared with either boscalid or dimethachlone alone,the virulence stimulation curve of the mixture of the two fungicides for dimethachlone-resistant isolates shifted to the left,and there were dose-additive interactions between the two fungicides.After the mycelia grown on PDA amended with boscalid,dimethachlone and the mixture of two fungicides for 48 h were inoculated on the fresh PDA or rapeseed leaves,both the mycelial growth and virulence were increased.Studies with 12 isolates of S.sclerotiorum showed that there was a positive correlation between the maximum virulence stimulation amplitudes and EC₅₀ values?R²=0.473,P=0.013?for dimethachlone,whereas there were negative correlations between the maximum virulence stimulation amplitudes and the mycelial growth rates?R²=0.631,P=0.002?,and between the maximum stimulation amplitudes and virulence at the absence of fungicide?R²=0.760,P<0.001?.Moreover,there was a negative correlation between the maximum virulence stimulation amplitudes of boscalid with the virulence of isolates?R²=0.439,P=0.019?,but there was no significant correlation of the maximum virulence stimulation amplitudes with the mycelial growth?R²=0.139,P=0.232?or with boscalid EC₅₀ values?R²=0.035,P=0.560?.Studies on hormesis mechanisms showed that mycelia grown on PDA amended with boscalid,dimethachlone and the mixture of the two fungicides had no significant effect on the tolerance of S.sclerotiorum to H₂O₂.Molecular studies showed that sublethal doses of boscalid and dimethachlone had no significant effect on the expression of cutinase gene SsCut,polygalacturonases gene SsPG1 or oxaloacetate acetylhydrolase gene SsOah1 of S.sclerotiorum.