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First Report of Fusarium proliferatum Causing Trunk Canker on Ilex cornuta in China
- Feng, F. S., Li, H.
- Plant disease 2019 v.103 no.1 pp. 155
- DNA primers, Fusarium proliferatum, Ilex cornuta, branches, color, conidia, culture media, defoliation, dieback, fungal diseases of plants, genes, greenhouse production, hyphae, internal transcribed spacers, leaves, mycelium, pathogenicity, plant pathogenic fungi, polymerase chain reaction, relative humidity, ribosomal RNA, sequence analysis, small fruits, sodium hypochlorite, spraying, tissues, woodlands, China
- Ilex cornuta, commonly known as Chinese holly, is widely distributed in China, where it is valued horticulturally for its attractive and distinctive rectangular foliage and large red berries. In March 2018, I. cornuta showing symptoms of an unknown disease was obtained from native woodlands in Tiao-ma prefecture of Changsha City, China (28°8′N, 112°59′E). Initial symptoms included trunk cankers with shallow black-colored rot as well as early defoliation. Over time, the cankers extended to lateral branches. Necrotic lesions from trunks with canker of I. cornuta were surface disinfested with 0.4% sodium hypochlorite solution for 1 min, and diseased tissue pieces were plated on potato dextrose agar. A Fusarium sp. (code HNCSTM4) was consistently isolated from diseased tissues, and a monosporal culture was obtained by single-spore isolation. The fungal colonies appeared fluffy; hyphae were densely branched and developed white to flavescent. Aerial mycelium was dense with a purple color on the lower surface of the plates when kept for 7 days at 25 ± 2°C. The average growth of colonies was 4.6 mm per day (n = 7). Macroconidia were 28.2 to 39.8 × 2.4 to 3.5 μm (length × width), straight to slightly curved, sickle shaped, with three to five septa. These morphological characteristics were consistent with previous descriptions of Fusarium (Mun et al. 2012; Summerell et al. 2003) and correspond to the species F. proliferatum (Wang et al. 2015). To identify the isolate of the pathogen molecularly, a polymerase chain reaction (PCR) was conducted using primers targeted to the internal transcribed spacer (ITS) regions of the rRNA gene (ITS4, 5′-TCCTCCGCTTATTGATATGC-3′; ITS5, 5′-GGAAGTAAAAGTCGTAACAAGG-3′) (Rampersad et al. 2011). The PCR product was sequenced and aligned with GenBank sequences. BLAST analysis of ITS sequences showed that the sequences were 100% identical to F. proliferatum, and the ITS region obtained in this study was deposited under the GenBank accession number MH332769. In addition, an F. proliferatum-specific amplicon was obtained and sequenced (GenBank accession no. MH700253) using the primer pairs PRO1 (5′-CTTTCCGCCAAGTTTCTTC-3′) and PRO2 (5′-TGTCAGTAACTCGACGTTGTTG-3′) for F. proliferatum (Mulè et al. 2004). Thus, both morphological and molecular characteristics identified the pathogen as F. proliferatum. Pathogenicity was confirmed by spraying a conidial suspension (1 × 10⁶ conidia/ml) onto nine branches of 1-year-old potted I. cornuta plants, which were wounded with a sterilized needle. Control branches were sprayed with sterile distilled water. Inoculated plants and the controls were kept under greenhouse conditions at 25°C and 90% relative humidity (day/night). The experiment was repeated three times on different days. Twenty days following inoculation, F. proliferatum-infected plants exhibited canker development and branch dieback, whereas control plants remained healthy. The pathogen was repeatedly isolated from inoculated I. cornuta plants, thus satisfying Koch’s postulates. To our knowledge, this is the first report of F. proliferatum causing trunk canker on I. cornuta in China. These findings will help in developing better preventive measures in accordance with the emergence of the new pathogen.