First Report of Stemphylium lycopersici Causing Leaf Spot on Polygonatum kingianum in China.

Abstract

Polygonatum kingianum, a member of the Liliaceae, is valued in traditional medicine and as a vegetable food crop. In July 2019, more than 50% of P. kingianum growth was suppressed in several field nurseries in Simao, Mojiang, Jingdong and Lancang County, Puer City, China. At the early stage of infection, symptoms manifested as a small circular brown spot. As the lesion matured, the spot gradually enlarged, forming an oval to irregular lesion with reddish-brown and dark green borders. In serious cases, the leaves were withered, and became brittle with cracks. The infected plants were collected from six major fields. The tissues of diseased leaves were soaked in 75% ethanol for 10 s, 0.1% mercuric chloride for 2 min, rinsed with sterilized water, and placed on potato dextrose agar (PDA) at 25℃ for 7 days. On PDA, four strains were isolated, and the colony was gray to dark yellowish-brown, flocculent, regular with concentric growth rings. Strain PKLS06 produced a dark red to brown pigment in the agar medium. On lesions, the conidiophores were solitary or in fascicles, straight or slightly curved, brown, with a conical apex, with three to five septa. The conidiogenous cells were pale-brown and swollen at the apex. On PDA, spores were solitary, oblong, bluntly rounded or sometimes with a point at the apex, with two to five transverse septa and one to two longitudinal septa with contractions at the main transverse septum. Morphological characteristics were consistent with published descriptions of Stemphylium lycopersici (Kee et al. 2017; Xie et al. 2018). For molecular identification, rDNA internal transcribed spacer (ITS) and the glyceraldehyde-3-phosphate dehydrogenase (gpd) gene were amplified and sequenced (ITS accessions: MW243098, MW243099, MW243100, MW243101; gpd accessions: MW246803, MW246804, MW246805, MW246806) using published primers (White et al. 1990; Câmara et al. 2002). A phylogenetic tree was developed by Maximum Parsimony (MP), Maximum Likelihood (ML) and Bayesian inference (BI). These four isolates fall into the S. lycopersici clade with strong support and all isolates were distinguished clearly from other species. Pathogenicity tests were performed using these four isolates. Each isolate was cultured on PDA and shake-cultured in V-8 juice broth (Nasehi et al. 2014). Conidia were resuspended in sterilized water (1×106 conidia/mL) and inoculated on intact leaves with injury of 1-year-old P. kingianum. The plants were incubated at 25℃ with a 12 h photoperiod and 90% humidity. A small spot began to appear after 3 days, and symptoms were similar to the those observed in the nursery after 10 days. Interestingly, the pathogenicity of strain PKLS06 was relatively weaker. Control plants treated with sterile water showed no disease symptoms. Re-isolated strains had the same morphological characteristics and the same ITS and gpd sequences as the original isolates, thus fulfilling Koch's postulates. S. lycopersici, an important pathogen, is widely distributed, and can cause a variety of plant diseases. It is noteworthy that the disease was observed on a plant in the Liliaceae, expanding the host range of S. lycopersici, which previously was reported to primarily infect plants in the Solanaceae. Based on the results presented above, P. kingianum is a new host plant of S. lycopersici in China. This disease is a threat for nursery production of P. kingianum, leading to a reduction in yields and economic losses. References Kee, Y. J., et al. 2017. Plant Disease 102 (2): 445-446 Xie, X. W., et al. 2019. Canadian Journal of Plant Pathology-Revue Canadienne De Phytopathologie 41 (1): 124-128 White, T. J., et al. 1990. PCR Protocols: A Guide to Methods and Applications PCR Protocols: A Guide to Methods and Applications 18: 315-322 Câmara M. P. S., et al. 2002. Mycologia 94 (4): 660-672 Nasehi A., et al. 2014. Archives of Phytopathology & Plant Protection, 47 (14): 1658-1665.