First Report of Rhizoctonia solani AG-4 HG-I Causing Crown and Stem Rot on Purple Coneflower (Echinacea purpurea) in Italy

In July to August 2018, crown and stem rot symptoms were observed on purple coneflower (Echinacea purpurea L.) grown as an ornamental in a garden located near Biella, 45°36′00″N 8°03′00″E (northern Italy). Water-soaked lesions developed on the crown directly above the soil line in 5 to 10% of 60-day-old plants grown in 25-liter pots. The disease was more common and severe in shaded or very humid areas, leading plants to death. Diseased tissue was surface disinfested for 10 s in 1% NaOCl, rinsed with sterile water, and plated on potato dextrose agar (PDA) amended with 25 mg/liter of streptomycin sulfate. A fungus with the morphological characters of Rhizoctonia solani (Sneh et al. 1991) was recovered from 70% of the samples, transferred, and maintained in pure culture. Mycelium of 10-day-old cultures grown on PDA, at 22 ± 1°C, was light brown, compact, with radial growth. Sclerotia were not present. DNA was extracted from a pure culture of a single isolate coded 43 with the E.Z.N.A. Fungal DNA Mini Kit (Omega Bio-Tek, Darmstadt, Germany). The internal transcribed spacer (ITS) region of rDNA was amplified by using the primers ITS1/ITS4 (White et al. 1990). The purified 668-bp PCR product was sequenced at the BMR Genomics Centre (Padova, Italy) and deposited in GenBank under accession number MK430998. A BLASTn search of this sequence showed 100% identity with the reference sequence KF907733 of R. solani AG-4 HGI isolated from Brassica oleracea (Hua et al. 2014). Anastomosis group was defined by pairing one strain of R. solani from E. purpurea with R. solani isolates belonging to already known anastomosis groups (AG1, AG2, AG4, AG7, and AG11) and observing microscopically the hyphal fusions. Three replicates for each pairing were made. R. solani from E. purpurea anastomosed (fusion frequency < 30%) with R. solani isolate AG4 (Sneh et al. 1991). This characterization was in accordance with the features of the isolates described above and with characteristics reported for AG4 group. Moreover, the ITS analysis permitted identification of the subgroup HGI of R. solani AG4. Five pots of 30-day-old plants of E. purpurea were used to evaluate the pathogenicity of isolate 43 of R. solani. The test was repeated once. Healthy plants were transplanted into steam-disinfested peat substrate (Cyclamen, Turco S.p.a., Italy) artificially infested by mixing 1 g/liter of R. solani grown on autoclaved wheat kernels at 25°C for 10 days. Plants transplanted in noninoculated substrate and maintained in the same greenhouse at 20 to 25°C served as a control. The first symptoms, similar to those observed in the garden, developed 10 to 12 days after inoculation, and 60 to 80% of plants died in both replications. Control plants remained healthy. R. solani was always reisolated from affected crowns and stems. Although R. solani was reported on E. purpurea in Canada (Chang et al. 1997), this is the first report of the disease in Italy and in Europe. The impact of the disease is unknown because it was observed for the first time in a private garden.