First Report of Leaf Anthracnose Caused by Colletotrichum tropicale on Oregano (Origanum vulgare) in Mexico

Abstract

Oregano (Origanum vulgare L.) is a perennial herb belonging to the Lamiaceae family and is commonly grown for its aromatic and medicinal properties. In January 2019, symptoms of leaf anthracnose were observed on approximately 25% of oregano plants located in a nursery in Huitzuco de los Figueroa, Guerrero, Mexico. Lesions on oregano leaves were irregular, brown, and yellow-haloed. For fungal isolation, small pieces from tissue at the margins of lesions of symptomatic leaves were surface disinfested by immersion in a 1% sodium hypochlorite solution for 2 min, rinsed in sterile distilled water, and placed onto potato dextrose agar (PDA) in Petri plates. The plates were incubated at 25°C for 5 days in darkness. Colletotrichum-like colonies were consistently obtained, and five monoconidial isolates were made. Only a single morphotype was observed, and an isolate was randomly selected for characterization, multilocus phylogenetic analysis, and pathogenicity tests. The isolate was deposited in the Culture Collection of Phytopathogenic Fungi at the Colegio Superior Agropecuario del Estado de Guerrero (accession no. CSAEG-CO19). After 8 days on PDA, the colonies were white to gray on the upper surface and had orange conidial masses. Conidia (n = 100) were cylindrical, hyaline, aseptate, with rounded ends, and measured 13.6 to 17.8 × 3.9 to 5.4 μm. Based on morphological features, the isolate was tentatively identified as a member of the Colletotrichum gloeosporioides species complex (Weir et al. 2012). Total DNA was extracted, and the internal transcribed spacer (ITS) region (White et al. 1990) and partial sequences of actin (ACT) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes were amplified by polymerase chain reaction (PCR) (Weir et al. 2012) for sequencing and molecular identification. A phylogenetic tree based on Bayesian inference and including published ITS, GAPDH, and ACT data for Colletotrichum species was constructed. The multilocus phylogenetic analysis clearly distinguished the isolate CSAEG-CO19 as Colletotrichum tropicale, separating it from all other species within the C. gloeosporioides species complex. DNA sequences were deposited in GenBank (accession nos.: ITS, MN947334; GAPDH, MN942958; and ACT, MN942957). Pathogenicity of the fungus was verified by spraying a conidial suspension (1 × 10⁵ spores/ml) on 10 leaves on each of 10 healthy oregano plants. Ten control plants were sprayed with sterile distilled water. All inoculated plants and controls were covered with a plastic bag for 24 h and then were kept under greenhouse conditions at 25 to 30°C for 18 days. Anthracnose symptoms were observed on all inoculated leaves after 15 days, whereas control leaves remained healthy. The fungus was consistently reisolated from leaf lesions, fulfilling Koch’s postulates. To date, only Colletotrichum fioriniae in the Colletotrichum acutatum species complex has been reported as a leaf pathogen on oregano, in Italy (Guarnaccia et al. 2019). However, to our knowledge, this is the first report of C. tropicale causing leaf anthracnose on oregano worldwide.