During disease surveys in July 2017 in the Atatürk Arboretum, İstanbul (41°10'30.53"N, 28°59'5.22"E), dieback was observed on branches of a young C. libani. Abundant black pycnidia were observed on dead needles under a binocular microscope. Examination under a compound microscope revealed the presence of non- or one-septate, light to dark brown, oblong-ellipsoid conidiospores, characteristic of the genus Diplodia. Isolations were made by plating spore masses onto 2% malt extract agar in 90 mm diameter Petri dishes under aseptic conditions and incubating at room temperature for ten days in the dark. Blackish-dark grey colonies with grey aerial mycelia were subcultured on 2% malt extract agar covered with cellophane membrane and incubated as described above. DNA was extracted from mycelia using an E.Z.N.A. fungal DNA extraction kit (Omega Bio-tek, USA) and amplified by PCR using species-specific primers for both D. pinea and D. scrobiculata (Smith & Stanosz, 2006). The D. pinea primers amplified a product of 700 bp. The ITS region was also amplified and sequenced (White et al., 1990), and the sequence of a representative isolate was deposited in GenBank (Accession No. MH368280). Comparison of the ITS sequences obtained from C. libani showed 100% nucleotide identity with those of reference strains of D. sapinea (e.g. NR_152452.1).
First report of Diplodia sapinea on Cedrus libani in Turkey
During disease surveys in July 2017 in the Atatürk Arboretum, İstanbul (41°10'30.53"N, 28°59'5.22"E), dieback was observed on branches of a young C. libani. Abundant black pycnidia were observed on dead needles under a binocular microscope. Examination under a compound microscope revealed the presence of non- or one-septate, light to dark brown, oblong-ellipsoid conidiospores, characteristic of the genus Diplodia. Isolations were made by plating spore masses onto 2% malt extract agar in 90 mm diameter Petri dishes under aseptic conditions and incubating at room temperature for ten days in the dark. Blackish-dark grey colonies with grey aerial mycelia were subcultured on 2% malt extract agar covered with cellophane membrane and incubated as described above. DNA was extracted from mycelia using an E.Z.N.A. fungal DNA extraction kit (Omega Bio-tek, USA) and amplified by PCR using species-specific primers for both D. pinea and D. scrobiculata (Smith & Stanosz, 2006). The D. pinea primers amplified a product of 700 bp. The ITS region was also amplified and sequenced (White et al., 1990), and the sequence of a representative isolate was deposited in GenBank (Accession No. MH368280). Comparison of the ITS sequences obtained from C. libani showed 100% nucleotide identity with those of reference strains of D. sapinea (e.g. NR_152452.1).