Figure 1.
Frequency distributions of EC50 values (the effective concentration causing 50% inhibition of mycelial growth of Phytophthora melonis) for flumorph, dimethomorph and iprovalicarb.
In total, 80 isolates of P. melonis were collected from areas never exposed to carboxylic acid amide fungicides.
Figure 2.
Genetic relationships among 15 isolates of Phytophthora melonis.
The denrogram (UPGMA) shows the relationships among the isolates of P. melonis based on randomly amplified polymorphic DNA (RAPD) analysis with 16 decamer primers. Scale at the bottom depicts the genetic distance.
Table 1.
Nucleotide sequences and characteristics of primers used in this study.
Table 2.
Isolates of Phytophthora melonis used for RAPD analysis and their sensitivities to flumorph, dimethomorph and iprovalicarb.
Table 3.
Results of the experiments conducted to induce resistance against flumorph, dimethomorph, and iprovalicarb in Phytophthora melonis.
Table 4.
Fitness of CAA-resistant and -sensitive isolates of Phytophthora melonis in vitro.
Figure 3.
Cross-resistance among flumorph, dimethomorph and iprovalicarb.
Log-transformed EC50 values (the effective concentration for causing 50% inhibition of mycelial growth inhibition of Phytophthora melonis) for isolates of P. melonis were compared among the three carboxylic acid amide fungicides using Spearman’s rank correlation coefficients. (A), (B), and (C) indicate positive cross-resistance among flumorph, dimethomorph, and iprovalicarb; (D-F) include only the higher EC50 values from (A-C), i.e., EC50 values from CAA-resistant isolates. (D) reveals a positive correlation between the EC50 values for dimethomorph and flumorph among CAA-resistant isolates, while (E) and (F) reveals a negative correlation between iprovalicarb and flumorph and between iprovalicarb and dimethomorph among CAA-resistant isolates.
Figure 4.
Structure and site of mutation in the PmCesA3 gene associated with carboxylic acid amide (CAA) fungicide resistance.
(A) Intron/exon structure of the PmCesA3 gene. Numbers represent the size in base pairs. Point mutations in CAA-resistant mutants and the predicted amino acid substitution in the mutant gene products are indicated. (B) Alignment of partial amino acid sequences of CesA3 in P. melonis (PmCesA3), P. infestans (PiCesA3), and P. viticola (PvCesA3). TJ-90, TX-21, and TX-33 were wild-type isolates. D63-1 and D70-3 were dimethomorph-resistant mutants. F58-4 and F63-11 were flumorph-resistant mutants. I63-2 and I70-5 were iprovalicarb-resistant mutants. Mutations in CAA-resistant mutants of P. infestans, P. viticola and P. melonis are indicated by asterisks.
Table 5.
Predicted amino acid sequence identities (%) among known CesA3s from four Phytophthora species, Plasmopara viticola, and Arabidopsis thaliana.
Figure 5.
Specificity of four allele-specific PCR primer pairs for the detection of carboxylic acid amide (CAA)-resistant isolates of Phytophthora melonis.
(A) Specificity of the four primer pairs for the CAA-sensitive isolate TJ-58 (S) and the CAA-resistant isolate F58-4 (R) at gradient annealing temperatures. (B) Specificity of primer pair (PMF + PMR1109B) for four CAA-sensitive and five CAA-resistant isolates at 68.5°C.