Brown leaf spot of Jacaranda mimosifolia Caused by Colletotrichum karstii in Sichuan, China.

Abstract

Jacaranda mimosifolia D. Don is widely cultivated in southwest China (Yunnan, Sichuan, and other regions). It is widely applied in papermaking, medicine, environmental monitoring, timber, urban and rural afforestation, and soil and water conservation. In October 2020, a new brown leaf spot disease of J. mimosifolia was discovered in Xichang City (27°49' to 27°56'N, 102°16' to 102°11'E), with approximately 66.23% disease incidence. Firstly, the typical symptoms showed deep yellow necrotic lesions in the center or on the margin of the leaves. Gradually, the necrotic lesions expanded and developed into brown spots. Under humid conditions, the edges of necrotic lesions turned dark brown progressively. Finally, the leaves withered, died, and fell off. Infected tissues from ten samples were cut into small pieces of 2.5 × 2.5 mm. The surfaces of infected tissues were sterilized for 30 s in 3% sodium hypochlorite, 60 s in 75% ethanol, and rinsed three times in sterile water. They were then blot-dried with autoclaved paper towels and cultured on potato dextrose agar (PDA) at 25℃ for 3 to 8 days. After culturing for 8 days at 25℃ and 12 h/12 h light/dark on PDA, the colony diameter reached 78.2 to 82.7 mm. The colonies were light orange, turned pale pink with light orange beneath. The conidia were single-celled, aseptate, cylindrical, smooth-walled, straight, hyaline with both ends bluntly rounded, measuring 12.3 to 16.8 × 4.3 to 5.6 μm (n = 100; average=14.5 × 5.1μm). These morphological characteristics were consistent with the description of C. karstii (Zhao et al. 2021). For molecular identification, the genomic DNA of the representative isolate JM202010 was extracted using a fungal genomic DNA extraction kit (Solarbio, Beijing). The internal transcribed spacer (ITS) [ITS1/ITS4 (White et al., 1990)], calmodulin (CAL) [CL1C/CL2C (Weir et al., 2012)], actin (ACT) [ACT512F/ACT-783R (Carbone & Kohn, 1999)], chitin synthase (CHS-1) [CHS-79F/CHS-345R (Carbone & Kohn, 1999)], β-tubulin (TUB2) [BT2A/BT2B (O'Donnell et al., 1997)], and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) [GDF/GDR (Templeton et al. 1992)] were amplified. Sequences were deposited in GenBank (ITS: OL454787, CAL: OL518966, ACT: OL518967, CHS-1: OL518968, TUB2: OL518969, and GAPDH: OL518970). BLAST results indicated that the ITS, CAL, ACT, CHS-1, TUB2 and GAPDH sequences showed >99% identity with Colletotrichum karstii sequences at NCBI (GenBank MW494453.1, MW495036.1, MG387951.1, MW495038.1, MW495042.1, and MG602034.1). The conidial suspension (1 × 106 conidia/ml) was sprayed on the leaves of 4-year-old J. mimosifolia plants (10 plants) and inoculated for pathogenicity test. Fifteen leaves of each plant (10 pots in total) were inoculated with spore suspensions on both sides of the leaves. An equal number of control leaves was sprayed with sterilized distilled water as a control. Finally, all pots were kept in a greenhouse at 26°C under a 16 h/8 h photoperiod and 60 to 68% relative humidity. The inoculated plants showed symptoms similar to those of the original diseased plants, but the controls remained asymptomatic. Colletotrichum karstii was re-isolated from the infected leaves and identified by both morphological characteristics and DNA sequence analysis. The pathogenicity test was repeated thrice, which showed similar results, confirming Koch's postulates. To our knowledge, this is the first report of brown leaf spot on J. mimosifolia caused by C. karstii in China. C. karstii was previously reported as the causal agent of anthracnose on Fatsia japonica (Xu et al. 2020) and Nandina domestica (Li et al. 2017) in China. This finding provides an important basis for further research on the control of this disease.