Pepper(Capsicum annuum L.) is the vegetable with the largest production area China (Zou and Zou 2021). In the summer of 2020 and 2021, disease symptoms were observed in C. annuum L. cv. bola in a 10-ha field in Yiyang(28.35°N, 112.56°E), Hunan province of China. The disease incidence ranged from 10% to 30%. The symptoms initially appeared as tan lesions, which were colonized by fast-growing white mycelia, at the soil line. Affected plants eventually became wilted. Wilting was accompanied by girdling of the stem at the base, and signs of the pathogen, mycelia and golden-brown colored sclerotia. The spatial distribution of the disease was either single plants or small foci of affected plants. Diseased stem sections (1.0~1.5 cm) of 20 plants from the field in 2021 with typical symptoms were surface sterilized with 75% ethanol for 30 s, followed by 60 s in 2.5% NaClO, rinsed thrice with sterile water, air dried and plated on potato dextrose agar (PDA), and incubated at 28℃ in the dark for 5 days to isolate the causative pathogen. Twenty fungal isolates with similar colony morphology were collected and purified. These isolates formed radial colonies, and abundant sclerotia were observed after 5 to 10 days of incubation at 28℃. The color of the sclerotia with a diameter of 1.39 ± 0.15 mm (1.15 to 1.60, n=50) gradually changed from white to light yellow, and finally to dark brown. The representative isolate YYBJ20 was selected for further molecular identification. The internal transcribed spacer region and elongation factor-1alpha gene were amplified using the primers, ITS1/ITS4 (White et al. 1990) and EF1-983F/EF1-2218R (Rehner and Buckley 2005), respectively. The ITS and EF1α amplicons were sequenced and deposited in GenBank with the accession numbers OQ186649 and OQ221158, respectively. Sequence analysis revealed that the ITS and EF1α sequences of the YYBJ20 isolate exhibited ≥99% of identity with the ITS (MH260413 and AB075300) and EF1α (OL416131 and MW322687) sequences of Athelia rolfsii, respectively. Phylogenetic analysis classified YYBJ20 into a common clade with different A. rolfsii strains, but different from other Athelia or Sclerotium species. For pathogenicity tests, PDA plugs (6 mm diam.) colonized by 3-day-old mycelia were inoculated into the stem bases of 30-day-old pepper seedlings (n=10). Another 10 seedlings were inoculated with noncolonized PDA plugs were used as noninoculated controls. The pepper seedlings were incubated at 28 ± 2℃ and 60 to 80 % relative humidity under a 14h-10h of light-dark cycle. After 10 days of incubation, ten YYBJ20-inoculated plants were wilted with similar symptoms to those observed in the field, while control plants remained healthy. The pathogenicity tests were repeated three times. The fungal strain re-isolated from the infected seedlings (100% re-isolation frequency) showed the same morphological and molecular traits as the original isolates from the diseased plants. No fungi were isolated from the control plants, which is consistent with the Koch's postulates. Based on the morphological and sequencing results, the causative fungus was identified as A. rolfsii (anamorph Sclerotium rolfsii). To our knowledge, this is the first report of A. rolfsii causing southern blight on pepper in China. Due to the broad host range of and serious consequences caused by A. rolfsii (Lei et al. 2021; Zhang et al. 2022; Zhu et al. 2022), this research will be beneficial to develop strategies to mitigate future losses of pepper in China.
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Jan 4, 2024
Aisyah Nita Yasmin Lubis + 1
Open Access