In vitro toxicity of fungicides with different modes of action to alfalfa anthracnose fungus, Colletotrichum destructivum.

Sensitivity of 24 isolates of Colletotrichum destructivum O'Gara, collected from alfalfa plants in Serbia, to eight selected fungicides, was investigated in this study. Molecular identification and pathogenicity test of isolates tested were also performed. Fungicide sensitivity was evaluated in vitro, using mycelial growth assay method. All isolates exhibited significant pathogenicity, causing necrosis at the alfalfa seedling root tips two days after inoculation. Using the primer pair GSF1-SR1 and by comparing the amplified fragments of the tested isolates with the marker (M), the presence of the amplicon of the expected size of about 900 bp was determined for all isolates. The isolates tested in this study showed different sensitivity towards fungicides in vitro. Mycelial growth was highly inhibited by QoI (quinone outside inhibitors) fungicide pyraclostrobin (mean EC₅₀=0.39 µg mL^-1) and by DMI (demethylation-inhibiting) fungicide tebuconazole (mean EC₅₀=0.61 µg mL^-1), followed by azoxystrobin (mean EC₅₀=2.83 µg mL^-1) and flutriafol (mean EC₅₀=2.11 µg mL^-1). Multi-site fungicide chlorothalonil and MBC (methyl benzimidazole carbamate) fungicide thiophanate-methyl evinced moderate inhibition with mean EC₅₀=35.31 and 62.83 µg mL^-1, respectively. Thirteen isolates were sensitive to SDHI (succinate dehydrogenase inhibitors) fungicide boscalid and fluxapyroxad, (mean EC₅₀=0.49 and 0.19 µg mL^-1, respectively), while the rest of isolates were highly resistant.