Studies On Risk Of Resistance To Oxadixyl, Dimethomorph And Cymoxanil In Air-bone Pathogens Causing Downy Mildew And Phytophthora Blight

To evaluate risk of resistance to fungicide is of great importance in developing, introducing, applying novel fungicides, and managing resistance, however, systemic evaluation to resistance risk has not been made yet in China, examples of risk evaluation are rare abroad and prediction is often not in accordance with actual situation.In the thesis, a method of systematicly assessing risk of resistance to oxadixyl, dimethomorph and cymoxanil in air-borne pathogens causing downy mildew and Phytophthora blight was established, it was showed that high risk of resistance to oxadixyl, moderate risk to dimethomorph and low risk to cymoxanil, which could lay a foundation for starting evaluation of resistance risk in China; Based on the fact that no cross resistance between low-risk cymoxanil and high-risk metalaxyl or oxadixyl, cymoxanil mixtures with mancozeb was synergistic and able to delay development of resistance to oxadixyl in PI, and excellent control efficacy against oxadixyl-resistant (OR) downy mildew on cucumber, a strategy was established that cymoxanil mixture with mancozeb (1:8), should be used as substitution for managing resistance and decreasing risk of resistance to phenylamides in target fungi.With sporangia irradiated by Ultraviolet (UV), frequency of oxadixyl-resistant (OR) and dimethomorph-resistant (DR) mutants of PI was 4.2× 10~-7 and 1.8 × 10~-7, respectively. Frequency of OR and DR mutants of PV was 1.4× 10~-7 and 1.8 × 10~-7, no cymoxanil-resistant (CR) mutants was found. With mycelia of PI irradiated by UV, frequency of CR, DR and OR mutants was 1.25%, 8.75% and 18.1%, respectively. With sporangia treated by MNNG, frequency of CR, DR and OR mutants of PI was 3.3 × 10~-7, 5.0 × 10~-7 and 5.0 × 10~-7, frequency of OR and DR mutants of PV was 5.0 × 10~-7 and 1.7 × 10~-7 no CR mutant was found.Resistance to oxadixyl developed after PI was subcultured for 3 asexual cycles on oat agar medium (OA) containing oxadixyl, lower resistance developed after PI was transferred for 5 generations on OA containing cymoxanil and concentration of cymoxanil increased gradually. No resistance to dimethomorph was trained out. Resistance to oxadixyl and dimethomorph developedafter PV was tansferred for 3 and 5 generations on grape leaves sprayed with oxadixyl and dimethomorph, respectively, no resistance to cymoxanil were trained out.Fitness test showed that by induction PI and PV easily developed OR mutants as fit as their original strains, most DR mutants less fit than their parent strains, CR mutants lost or decreased pathogenicity. Of 35 OR mutants of PI, 5.7% had compound fitness index (CFI) significantly higher than their parent strains, 28.6% with CFI as high as parent, 65.6% with obviously lower CFI. Of 7 OR mutants of PV, one's pathogenic fitness nearly stable, but 6 mutants less fit. 14 DR strains of PI and 6 DR strains of PV much less fit than their parent, 2 DR mutants of PI nearly as fit as parent. Of 9 CR strains of PI, CFI of 2 mutants was significantly lower, 7 mutants lost pathogenicity. After subculturing free of fungicide, one OR mutant of PI was still more fit than its parent strain , 3 OR mutants of P V fit as before, 2 DR mutants of PI much less fit.Stability result indicated that induced OR mutants stable in resistance and pathogenicity, but DR and CR mutants unstable, their subcultures in the absence of fungicide or single-zoospore clones became less pathogenic or lost pathogenicity. On potato discs, of 10 OR mutants of PI with resistance level (RL) of 28-3226 fold, one mutant became 10 times less resistant, 9 mutants nearly as resistant as before; after 3 subcultures free of fungicide, 2 mutants of PI with RL of 4 fold to cymoxanil lost pathogenicity. 3 mutants of PI with RL of 7-14 fold to dimethomorph became as sensitive as parent.3 DR mutants of PV decreased in RL from 30-150 fold to 12-38 fold, 3 OR mutants of PV kept nearly stable in RL. For CR2, one CR mutant of PI, of 20 single-zoospore clones of the 1st generation with RL of 1-7 fold, sensitive clones were 10%; of 20 single-zoospore clones of the 2nd generation isolated from CR2.1 with RL of 2-5 fold, sensitive clones were 25%; of 20 single-zoospore clones of the 3rd generation isolated from CR2.1.1 with RL of 1-2 fold, sensitive clones were 90%. For PHDR1, one DR mutant of PI, 20 single-zoospore clones of the 1st generation had RL of 14-26 fold, 20 single-zoospore clones of the 2nd generation isolated from one single-zoospore clone as resistant as PHDR1 had RL of 11-14 fold, 20 single-zoospore clones of the 3rd generation isolated from one resistant single-zoospore clone had RL of 7-21 fold; For OR1, with RL of 1504 fold, 20 single-zoospore clones of the 1st generation had RL of 938-1812 fold, 20 single-zoospore clones of the 2nd generation isolated from one resistant single-zoospore clone had RL of 1028-1742 fold, 20 single-zoospore clones of the 3rd generation isolated from one resistant single-zoospore clone had RL of 1139-1539 fold.Competition test showed that DR mutants or CR mutants exhibited a very weak competitive ability when mixture of DR or CR strains with their original strains were inoculated on untreated detached leaves, but OR mutants were very competitive as mixture of OR strains with their original strains were subcultured on untreated potato detached leaves; The treatment of mixture of cymoxanil and mancozeb (1:8) delayed development of resistance to oxadixyl in PI. Frequency ofresistance to dimethomorph or cymoxanil significantly decreased when mixture of PHDR1, a DR strain of PI, or CR2, a CR strain of PI, or DR5, a DR strain of PV with their original strains were subcultured on untreated detached leaves. Frequency of resistance to oxadixyl rose obviously as mixture of ORl, an OR strain with NS, its original strain, were transferred on untreated leaves, frequency of resistance to oxadixyl rose from 20% to 100% or from 50% to 100% as mixture of ORl with NS were transferred for 4 or 3 sporulation cycles on detached potato leaves treated with oxadixyl, frequency of resistance to oxadixyl rose from 20% or 50% to 100% as mixture of ORl with NS were subcultured for 5 asexual cycles on the detached potato leaves treated with the mixture of oxadixyl with mancozeb (1:8), frequency kept stable when treated with Curzate-M8. Frequency of resistance to dimethomorph rose from 20% or 50% to 68% or 100%, and frequency of resistance to cymoxanil rose from 20% or 50% to 30% or 81% when mixture of DR or CR strains from fields with their sensitive strains were subcultured for 8 sporulation cycles on intact potato plants treated with dimethomorph or cymoxanil.It was confirmed that cross resistance existed between metalaxyl and oxadixyl, no cross resistance between metalaxyl or oxadixyl and dimethomorph or cymoxanil. Field strains of PI sensitive to metalaxyl were also sensitive to oxadixyl, some strains resistant, intermediate or sensitive to oxadixyl were sensitive to dimethomorph or cymoxanil, others insensitive; 12 field strains of PC were highly resistant to metalaxyl and oxadixyl, but sensitive to dimethomorph, 13 OR strains and 2 sensitive strains of PV were sensitive to dimethomorph and cymoxanil.A leaf disc quantitative spraying method suitable for testing action modes and synergistic action of fungicide mixtures was established. It was found first that cymoxanil mixture with mancozeb (1:7) was synergistic against OR strains of PC, when cucumber leaf discs were treated with curative or eradicative spray. Cymoxanil mixture with mancozeb (1:8) displayed a persistent period of 8 days, but 4 days for cymoxanil. Cymoxanil mixtures with mancozeb (1:8) showed excellent curative and eradicative effects against PV, on an OR strain of PC synergism ratio between cymoxanil with mancozeb (1:8) was 5.0.Resistance detection and control efficacy showed that OR strains of PI, collected from potato plants of Belgium, potato plants in Chongli and Weichang, and tomato plants in Xushui, were 27.3%, 32.7% and 1.1% of the sampled; but OR strains of PC and PV in Suburban of Beijing, Baoding and Xinji were 75% and 85%, with RL of 1474 and 1748 fold, respectively, control efficacy of 72% Curzate-M8 WP on PC was 88.3-94.1%, obviously higher than that of 25% Ridomil WP and 64% Sandofan-M8 WP. No CR or DR strain of three fungi was detected in the areas above. Resistance detection and efficacy performance confirmed resistance risk assessment.