Crocus sativus L. (Iridaceae) is a perennial, bulbous plant whose dried stigma is one of the most valuable spices and herbal medicines. In China, it is cultivated with a “two-segment” method, a field stage for corm propagation and an indoor stage for flowering. During October and November of 2023, symptoms such as browning, necrosis and rot were observed on C. sativus corms in Nantong (31°51′36″N, 121°28′48″E), Jiangsu Province, China, with an estimated disease incidence of 12% (mean of three replicates, 100 corms examined per replicate). Symptomatic corms were surface-sterilized with 75% ethanol for 60 s, followed by 4% NaClO for 20 min (minimize the contamination without explant damage), and rinsed with sterile water five times. The junction of symptomatic and healthy areas of corms was cut into 5 × 5 mm pieces, placed on potato dextrose agar (PDA), and incubated at 28°C for 4 days in the dark. Thirty-two fungal strains were obtained from 4 symptomatic corms, including 7 Clonostachys-like monospore isolates. The representative isolate (LZ-1) formed colonies ≈40 mm in diameter on PDA after 7 days at 28°C (dark), and displayed felty to tomentose to granulose aerial mycelia with a white or light-yellow appearance (reverse light brown). The conidiophores were Verticillium-like, with 1 to 3 levels, and stipe lengths ranged from 30 to 71.9 µm. Secondary conidiophores exhibited 2 or 3 whorls of phialides. The penicillate-like conidiophores were absent. Conidia were colorless, round or oval, and aseptate, with an average size of 3.4×5.7 µm. Molecular identification of LZ-1 was performed by amplifying and sequencing the nuclear ribosomal internal transcribed spacer (ITS), β-tubulin (TUB2), and the translation elongation factor 1-alpha (TEF1-a) fragments (Broberg et al. 2020; Moreira et al. 2016), respectively. BLASTn against the GenBank showed 100% (569/569 bp) ITS, 99.43% (524/527 bp) TUB2, and 99.14% (689/695 bp) TEF1-a identity with Clonostachys rosea (HQ731632.1ITS, PQ462664.1TUB2, OR593726.1TEF1-α). Phylogenetic analysis of combined ITS, TUB2, and TEF1-α sequences confirmed LZ-1 as C. rosea. For pathogenicity tests, surface-sterilized healthy corms were wounded and inoculated with 7 mm in diameter LZ-1 mycelial plugs. The control corms were treated with PDA. Inoculated corms were placed in a chamber at 28°C (dark) and 70% humidity. The pathogenicity test was performed three times. After 7 days, all inoculated corms developed brown necrosis symptoms, while control corms remained asymptomatic. Re-isolated pathogens from the symptomatic corms matched the morphological and molecular characteristics of LZ-1. Next, potted healthy corms were inoculated by applying 1 mL of 1×108 conidia/mL conidial suspension (Zheng et al., 2021) in soil; control corms received sterile water. After 24 h, 50 mL water was added into each pot for regular irrigating. The test was replicated three times under 28℃, 14 h light/10 h dark. After 25 days, scattered brown necrosis spots appeared on the inoculated corms but not on the control corms, and LZ-1 was re-isolated and confirmed. Clonostachys rosea was previously reported to cause bulb rot in garlic (Rocío Díaz et al., 2022) and Fritillaria taipaiensis (Wang et al., 2024). To our knowledge, this is the first report of C. rosea causing corm rot in C. sativus in China, providing a basis for further studies on the prevention and control of this disease.
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