Archivio Journal of Plant Pathology

Abstract:

Leaf spot on buckwheat seriously occurred in Baoji, Shaanxi, China, in August 2015. The disease incidence ranged from 60 to 80%. Fungal isolates were isolated on potato-dextrose-agar media at 25±1°C (Sinclair and Dhingra, 1995). After 6 days of incubation, colonies developed dark pycnidia. The pycnidia were ostiolate and 100-125 µm in diameter. Conidia were elliptic, colorless, single cell, containing one or two lipid droplets, 4.2-5.8 ×2.7-3.7 µm. DNA was extracted from a representative monosporic culture and the internal transcribed spacer (ITS) and beta-tublin region of the nuclear ribosomal DNA (rDNA) was amplified and sequenced using primers ITS-1F/ITS4 and betatub3/betatub4r, respectively. The obtained sequences of 485 bp and 213 bp were deposited in the GenBank (accession No. LC198572.1 and LC198184.1). Phylogenetic tree based on ITS and beta-tublin showed 98% similarity to Didymella rhei. Pathogenicity tests were performed on four-leaf stage buckwheat seedlings ‘Xinong 9920’. Leaves of 20 plants were inoculated by either wounding or non-wounding. For the wounding test, 5 µl of 105 -106 spores/ml conidial suspension was dropped on the leaf sites with needle-prick wound. For the non-wounding test, the conidial suspension was sprayed on the leaf surface to runoff. Control plants were sprayed or dropped with sterilized water. Leaf lesions were observed after 2-3 weeks post-inoculation. The symptom development on wound-inoculated plants was faster than the non-wounding inoculation. The fungus was reisolated only from diseased leaves, fulfilling Koch’s postulates. Though many species of Didymella cause spot diseases in the world (Rothwell, 1982; Jellis and Punithalingam, 2007; Pearce et al., 2016), this is the first report of D. rhei infecting buckwheat.