Bioactivity And Resistance Risk Assessment Of Three Fungicides Against Sclerotinia Sclerotiorum

The filamentous ascomycete Sclerotinia sclerotiorum(Lib.)de Bary is a cosmopolitan necrotrophic plant fungal pathogen with a worldwide distribution,which can attracks more than 400 plant species belonging to 75 families.Infection can occur on different crop developmental stages.In Australia,United States,Canada and China,sclerotinia stem rot(SSR)can result in serious losses of yield and quality for a number of important field crops.In China,SSR could causes 5 to 80%yield losses of oilseed rape annually.Currently,the application of various fungicides is the primary tool in most oilseed rape managing SSR.The common fungicides carbendazim(benzimidazole fungicides)and dimetachlone(dicarboximide fungicides)have been used for controlling SSR for many years in China.The carbendazim-resistant and dimetachlone-resistant population in the field has been monitored.We established the baseline sensitivity of S.sclerotiorum to fluazinam,cyprodinil and pyraziflumid.The baseline sensitivity of S.sclerotiorum to fluazinam was determined using 103 strains.The sensitivity of S.sclerotiorum populations to fluazinam was determined by the method of mycelial growth test on PDA medium and the resistance risk of fluazinam was evaluated in vitro.Results indicated that the average EC50(50%effective concentration)values against fluazinam were 0.0073±0.0045 ?g/mL for mycelial growth.Nine mutants with low resistance level were obtained by growing sensitivity strains on potato dextrose agar(PDA)amending with minimum inhibitory concentration(MIC)of fluazinam.The resistance of nine mutants was stable after their ten transfers on PDA without the fungicide.The fitness(mycelial growth,sclerotial production,pathogenicity)is decreased compared with the sensitivity parental strains.In addition,cell membrane permeability of nine fluazinam-resistant mutants was higher than that of their sensitivity parental strains.There was no cross-resistance between fluazinam and fludioxonil,dimetachlone,prochloraz,tebuconazole,azoxystrobin,or procymidone in S.sclerotiorum.Besides,there were no mutations in the two-component histidine kinase gene(Shkl)of the resistant mutants.The baseline sensitivity of S.sclerotiorum to cyprodinil was determined using 100 strains.The EC50(50%effective concentration)values ranged from 0.0636?0.8163 ?g/mL for mycelial growth.Nine cyprodinil-resistant mutants(Range of resistance factor:20.22?271.59)were obtained by growing sensitivity strains on potato dextrose agar(PDA)amending with minimum inhibitory concentration(MIC)of cyprodinil and the resistance was stable after their ten transfers on PDA without the fungicide or stored at 4? for 60 days.There was positive cross-resistance between cyprodinil and pyrimethanil but not to fludioxonil,dimetachlone,procymidone,carbendazim and boscalid in S.sclerotiorum.All of the nine cyprodinil-resistant mutants decreased in sclerotial production compared with their sensitivity parental strains.The dry weight of mycelia,pathogenicity and cell membrane permeability of most cyprodinil-resistant mutants decreased.The mycelial growth,oxalic acid content,and the response to various stress for resistant mutants were almost the same as the sensitive parental strains.There was no alteration of amino acid in cystathionine ?-synthase(MetB)and cystathionine ?-lyase(MetC)on all of the strains.Cyprodinil could not only inhibited methionine biosynthesis but also suppressed cystine and cysteine biosynthesis.The baseline sensitivity of S.sclerotiorum to pyraziflumid was determined using 105 strains,and the average EC50 value was 0.0561±0.0263 ?g/mL for mycelial growth.There was no positive cross-resistance between pyraziflumid and carbendazim,dimethachlon and fludioxonil.Pyraziflumid could increased the offshoot of hyphae top.After pyraziflumid treated,cell membrane permeability significantly increased,oxalic acid content decreased markedly and mycelial respiration was inhibited.The pyraziflumid had great control efficiency against S.sclerotiorum on detached rapeseed leaves,and protective activity was better than curative activity.