Research On Resistance Of Pseudoperonospora Cubensis And Alternaria Solani To Azoxystrobin

Azoxystrobin,developed by Syngenta(Zeneca) company,was the first product of the strobilurin fungicides.It combines high level and broad-spectrum of fungicidal activity with excellent safety in the environment.Strobilurin fungicides inhibit mitochondrial respiration in fungi at a specific site on complexâ…¢and break energy synthesization of fungi.Strobilurins were called QoIs as they combined at Qo site of Cytb.This kind of fungicide was widely used because of their novel action mechanism and non-cross-resistance with any other presented fungicides.However,the frequency of spontaneous resistance mutation was high according to several research reports.It's easy to form resistant-population under high fungicidal selection pressure.Cucumber downy mildew(Pseudoperonospora cubensis) and Tomato early blight (Alternaria solani) are one of the serious diseases on China cucumber and tomato production,respectively.Aimed to control the popularity of Cucumber downy mildew and Tomato early blight,fungicidal workers are searching new fungicides.So,it is essential to embark on bioactivity and toxicology at molecular biochemical level of azoxystrobin to Pseudoperonospora cubensis and Alternaria solani.The bioactivity of azoxystrobin to Alternaria solani which had never been exposured to QoIs or QiIs was determined and also its influence on the fungus's biological properties was determined.We also made clear the synergistic effect of SHAM.The result was that azoxystrobin worked alone expressed less inhibition on mycelial growth and spore germination.Sensitivity of Alternaria solani to azoxystrobin could be decreased by the induction of alternative respiration.In the absence of a host,alternative respiration provided a＞27.03-fold rescue potential from azoxystrobin during the germination of conidia derived from five wild-type isolates ofAlternaria solani.And for mycelial growth, the rescue potential was 4.83～9.95-fold.SHAM had synergistic effect on the bioactivity of Azoxystrobin.But,azoxystrobin alone could strongly inhibit conidia production and delay the pigment formation.Oxygen consumption test of mycelia of Alternaria solani showed that azoxystrobin strongly inhibited mycelial respiration after 1 hour of the treatment at 12.5μg/mL and the effect became stronger when the concentration was increased.As treatment time went on, the respiration inhibition on the mycelia decreased,while the mycelium growth was still inhibited strongly.Results showed that the sensitivity decrease of mycelium respiration to azoxystrobin was not caused by induction of alternative respiration or degrade of the fungicide in medium.Cucumber downy mildew caused by Pseudoperonospora cubensis limits crop production in Shandong province of China.Since management of downy mildew is strongly dependent on fungicides,a rational design of control programs requires a good understanding of the fungicide resistance phenomenon in field populations of the pathogen. A total of 106 and 97 isolates of P.cubensis were obtained in 2006 and 2007,respectively. The EC₅₀ values for the growth of all the 106 isolates collected in 2006 were 0.0063-0.0688μg/mL with an average ECs0 value of 0.0196±0.0048μg/mL azoxystrobin which were considered sensitive isolates.However,57 field isolates of P.cubensis out of 97 collected in 2007 with EC₅₀ that ranged from 0.609 to＞51.2μg/mL were considered resistant to azoxystrobin.A total of 97 isolates of P.cubensis were obtained in 2007。There are 57 resistant isolates.The EC₅₀ values for resistant isolates ranged from 0.609 to＞51.2μg/mL,with the resistance factor 21.15-＞2618.9.Of all the 57 resistant isolates,the EC₅₀ values of 20 isolates,23 isolates and 14 isolates were 0.609-10μg/mL,10-50μg/mL and＞51.2μg/mL,respectively.One isolate was randomly selected from each of the three categories and one sensitive isolate was randomly selected for the molecular testing.A single point mutation(GGT to GCT) in the cytochrome b gene,resulting in substitution of glycine by alanine at position 143,was found in the three selected azoxystrobin-resistant isolates of downy mildew.This substitution in cytochrome b exhibited different resistance levels,with the resistance factor from 21.15 to greater than 2618.9.In addition,the different resistance levels seemed to appear within 1 year(within 2006 to 2007).