• Research in Plant Disease
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• v.29 no.1
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• pp.11-22
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• 2023

Resazurin-based microtiter assay was used to evaluate the inhibitory effect of fungicides on the respiration of Colletotrichum acutatum s. lat. 20JDS8 sensitive and 20CDJ6 resistant to strobilurin fungicides. The spores of C. acutatum s. lat. 20JDS8 were inoculated into potato dextrose broth (PDB) at densities of 1x10⁴, 1x10⁵ and 1x10⁶ spores/ml, respectively. The relative fluorescence unit (RFU) of all treatments inoculated at each spore density started to rise after 12 hr of incubation, and were 1,965.5, 5,412.5, and 10,061.0, respectively, after 24 hr of incubation. To evaluate the inhibitory effect of fungicide on the respiration of the pathogen, the spores of the pathogen were inoculated into the PDB and treated with the fungicides 0, 6, 12, and 24 hr after incubation, respectively. After keeping the pathogen culturing for another 24 hr, PrestoBlue reagent was treated into the PDB culturing the pathogen. The RFU of each treatment was examined 1 hr after the reagent was treated. When dithianon, isopyrazam, pyraclostrobin, and fluazinam were treated at high concentrations in the stages of spores (immediately after inoculation [0 hr]), spore germination (after incubation for 6 hr), and hyphal growth (after incubation for 12 hr), the respiration of pathogens was inhibited by 90-100%. When the fungicides were treated after culturing the pathogen for 24 hr, the respiratory inhibitory effects were greatly reduced. With pyraclostrobin-resistant C. acutatum s. lat. 20CDJ6, azxoystrobin, trifloxystrobin and kresoxim-methyl, which have the same mode of action, had very little or no respiratory inhibitory effect in all growth stages of pathogens. Based on the above results, it was thought that the resazurin-based microtiter assay could quickly and accurately evaluate the inhibitory efficacy of the fungicides that inhibited respiration.

• Research in Plant Disease
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• v.28 no.3
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• pp.122-131
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• 2022

Colletotrichum acutatum s. lat. 20JDS8 sensitive and 20CDJ6 resistant to pylaclostrobin were used to investigate the cross-resistance with fungicides belonging to the strobilurins and the characteristics of fungicidal controlling activities with different mechanisms against the isolate resistant to the fungicide. The resistant isolate of 20CDJ6 also showed the resistance to azoxystrobin, trifloxystrobin, and kresoxim-methyl, suggesting that there is a cross-resistance relationship. All fungicides with different action mechanisms inhibited mycelial growth of both susceptible and resistant isolates of C. acutatum s. lat., but their disease control effects in fruits were different according to the fungicides. The disease control effect of isopyrazam against 20JDS8 and 20CDJ6 was very low, and fluazinam showed a control effect of 91.9% and 88.1% against 20JDS8 and 20CDJ6 only when it was treated before inoculation by spraying spore suspensions on pepper fruits without wounds. Tebuconazole and prochloraz effectively inhibited not only the mycelial growth of 20JDS8 and 20CDJ6 on potato dextrose agar medium, but also disease incidence in red pepper fruits. As a result of this study, C. acutatum s. lat. 20CDJ6 resistant to pyraclostrobin showed cross-resistance with other strobilurin fungicides. In addition, we think that fluazinam, tebuconazole, and prochloraz can be recommended as alternative fungicides for the control of red-pepper pyranthracnose pathogens resistant pyraclostrobin. However, fluazinam can be effective only if it is treated protectively before the occurrence of the disease.

• The Plant Pathology Journal
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• v.38 no.6
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• pp.629-636
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• 2022

Colletotrichum species is known as the major causal pathogen of red pepper anthracnose in Korea and various groups of fungicides are registered for the management of the disease. However, the consistent use of fungicides has resulted in the development of resistance in many red pepper-growing areas of Korea. Effective management of the occurrence of fungicide resistance depends on constant monitoring and early detection. Thus, in this study, various methods such as agar dilution method (ADM), gene sequencing, allele-specific polymerase chain reaction (PCR), and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) were applied for the detection of benzimidazole resistance among 24 isolates of Colletotrichum acutatum s. lat. and Colletotrichum gloeosporioides s. lat. The result of the ADM showed that C. gloeosporioides s. lat. was classified into sensitive and resistant isolates to benomyl while C. acutatum s. lat. was insensitive at ≥1 ㎍/ml of benomyl. The sequence analysis of the β-tubulin gene showed the presence of a single nucleotide mutation at the 198th amino acid position of five isolates (16CACY14, 16CAYY19, 15HN5, 15KJ1, and 16CAYY7) of C. gloeosporioides s. lat. Allele-specific PCR and PCR-RFLP were used to detect point mutation at 198th amino acid position and this was done within a day unlike ADM which usually takes more than one week and thus saving time and resources that are essential in the fungicide resistance management in the field. Therefore, the molecular techniques established in this study can warrant early detection of benzimidazole fungicide resistance for the adoption of management strategies that can prevent yield losses among farmers.