First Report of Cladosporium tenuissimum Causing Blossom Blight on Strawberry in the Brazil

Abstract

Blossom blight in strawberry (Fragaria × ananassa Duch.) was first detected in high tunnel covered field plantings of cultivar Monterey imported from Chile. This detection occurred in the municipality of Barra da Estiva, Bahia, Brazil, during the months of January and February 2018. The disease occurred during all seasons of the year, with an average incidence of ∼20%. Disease symptoms began with gray sporulation of the fungus on the stigmas and anthers of strawberry flowers, followed by necrosis and full flower abortion, similar to that described by Nam et al. (2015) in South Korea. The fungus was isolated directly from affected pistils, which were transferred to potato dextrose agar (PDA) plates at 25°C with a 12-h dark/light cycle. Five isolates on PDA culture presented both immersed and superficial mycelium composed of branched hyphae, 1 to 5 μm wide, septate, subhyaline to pale brown, smooth, with thin walls. The conidiophores are macronematous, solitary, sometimes in groups of two to three, erect, straight or slightly flexuous, cylindrical, 5 to 7 μm wide, having intercalary swellings, sometimes slightly geniculate toward the apex, unbranched or branched, reaching 500 × 3 to 4 μm, septate, medium brown, smooth, possessing walls somewhat thickened, and presenting a swollen base. The conidiogenous cells are integrated, terminal and intercalary, cylindrical-oblong, 10 to 42 μm long, sub-denticulate to denticulate. The ramoconidia are rare, subcylindrical or cylindrical, 17 to 27 × 3 to 4 μm, continuous, sometimes one-septate, with a broadly truncate base, 2 to 3.5 μm wide. The conidia form branched chains with small terminal conidia, subglobose, obovoid, limoniform, sometimes globose, 4 to 5 × 2 to 3 μm, aseptate, apically rounded; the secondary ramoconidia are ellipsoid, fusiform to subcylindrical or cylindrical, 8 to 10 × 4 to 7 μm, with distal hila, zero- to one-septate, pale brown or pale olivaceous-brown, smooth, walls unthickened, attenuated toward apex and showing a basal hilum conspicuous, sub-denticulate to denticulate. Culture characteristics: colonies on PDA attained up to 84 mm in diameter after 14 days, smoke-gray, reverse gray to olivaceous-black, woolly to fluffy, margin glabrous to feathery, gray-olivaceous to white, aerial mycelium abundant, high, fluffy, smoke-gray, dense, without exudates, sporulating. These morphological characteristics fit the description of Cladosporium tenuissimum Cook (Bensch et al. 2012). Morphological identification was confirmed through phylogenetic analyses using the combined internal transcribed spacer (ITS) and partial translation elongation factor 1-alpha gene (TEF) sequences of related species. To amplify the ITS region and the TEF, the respective primer pairs ITS1/ITS4 (White et al. 1990) and EF1-728F/EF1-986R (Carbone and Kohn 1999) were used. The resulting sequences from one isolate have been deposited in GenBank under the accession numbers MK775118 (TEF) and MT241370 (ITS). BLASTn analysis revealed that the sequences were 100% identical to the respective ITS (MH864840) and TEF (HM148442) of the neotype strain of C. tenuissimum (CBS 125995). Phylogenetic analysis further confirmed the results. The pathogenicity test was performed with conidia of a 14-day-old culture, sprayed at a final concentration of 16.5 × 10⁶ conidia/ml on flower clusters of Monterey variety strawberry plants. Nine plants and three flower clusters per plant were used. The inoculated plants were incubated in a dew plastic box at 25°C and 100% relative humidity. The same procedure was done with sterilized water sprayed on two control plants. All inoculated strawberry flowers reproduced symptoms of blossom blight within 2 days. C. tenuissimum was reisolated from the inoculated flowers. This is the first report of the occurrence of this fungus on strawberry blossom in Brazil, and the second report worldwide.