First Report of Verticillium Wilt on <i>Siraitia grosvenorii</i> Caused by <i>Verticillium dahliae</i> in China

Siraitia grosvenorii, commonly known as Monk fruit, is the fruit of S. grosvenorii (Swingle) C. Jeffrey ex A. M. Lu & Zhi Y. Zhang (Jeffrey C. 1980). As a traditional medicinal and edible homologous herb, S. grosvenorii is widely distributed in southern China (Li et al. 2025). In July 2024, wilting plants were found in farmed fields in Longsheng Nationalities Autonomous County (25.714443°N, 110.032812°E), Guilin, Guangxi, China, with an estimated incidence rate of 10 to 30%. Initially the leaves exhibited signs of water loss. The stems and leaves gradually turned yellow and withered, ultimately leading to the death of the entire plant. To isolate the causal agent, 12 small pieces of discolored stem tissues were cut, and then they were surface sterilized by using 75% EtOH for 30s and placed in 2.5% NaOCl for 2 min, and then rinsed three times in sterilized distilled water. These samples were then placed on potato dextrose agar (PDA) and they were incubated at 25℃ in the darkness for 7 days. The hyphal tips were transferred to fresh PDA in order to obtain pure cultures. 26 isolates were obtained, 19 of which had similar morphology. These were isolated from all the symptomatic stems and they were considered to be pathogens, and named L1. After 14 days, the colonies were white, fluffy, with white aerial mycelium, while the reverse side turned dark black, due to the formation of microsclerotia. The hyphae were hyaline and formed whorled branches. The conidia were hyaline, aseptate, cylindrical to oval and measured 4.3 to 6.6 × 1.6 to 3.1 μm in size ( n = 60). The microsclerotia were immersed in the agar, and they were composed of rounded cells and were elongated or irregular in shape. These morphological features were consistent with Verticillium dahliae (Isaac 1967). The L1 isolate was subjected to sequencing of the internal transcribed spacer (ITS), actin (ACT) and glyceraldehyde-3-phosphate dehydrogenase (GPD) sections (Inderbitzin et al. 2011) for molecular identification. The ITS, ACT and GPD sequences (Genebank accession nos. PQ836512, PQ847485 and PQ847486) displayed 99 to 100% (529/531, 475/475, 640/640 bp) similarity to V. dahliae (Genebank accession nos. KX058040, OK513223 and OK513236, respectively). Phylogenetic analysis was based on a combination of ITS, ACT and GPD sequence data using MEGA 11 software to construct a phylogenetic tree with neighbor-joining. In the phylogenetic tree, the combined sequences attributed L1 to V. dahliae. A pathogenicity test was performed by soaking 6 1-month-old S. grosvenorii cut seedlings with wounded root tips in a fungal conidial suspension (1 × 106 conidium/mL) for 1 h (Choi et al. 2022). Six additional seedlings were treated with sterile distilled water as controls. All the seedlings were kept in a greenhouse at 25°C with 80% humidity. After 3 weeks, the inoculated plant leaves turned yellow and dried off. After 4 weeks, all the treated plants were dead with vascular discoloration in the roots and stems. V. dahliae was re-isolated from the symptomatic stem tissues and confirmed by morphology and DNA sequencing (ITS, ACT and GPD sequences). The control plants remained healthy and no V. dahliae was isolated from them, thereby fulfilling the Koch postulate. V. dahliae is an economically important pathogen with a wide host range worldwide (Lu et al. 2021). To our knowledge, this is the first report of V. dahliae causing Verticillium wilt of S. grosvenorii in China and elsewhere, thereby providing the possibility for the integrated management of Verticillium wilt on this herb.