Pogostemon cablin (Blanco) Benth. is a vital source of patchouli oil, utilized in traditional Chinese medicine, Ayurveda, cosmetics, and hygiene products(Swamy et al. 2016). In 2022, a leaf spot disease outbreak on patchouli plants occurred nearly 10 acres in Yunfu, Guangdong (21°2' N; 110°3' E), with an average incidence rate of 50%. Infected leaves initially showed circular spots with tan centers and yellow halos. Within five to ten days, these spots expanded, crossed leaf veins, and became polygonal with rough surfaces. As the disease progressed, the spots merged, darkening the veins and eventually causing leaf loss. Five symptomatic leaves were soaked in 75% ethanol for 10 s, and 2.5% sodium hypochlorite for 30 s, followed by a final rinse in sterile water. These samples were then placed on potato dextrose agar (PDA) plates. After 3-5 days of incubation at 28 °C, the mycelial growth was transferred to fresh PDA plates and purified by isolating the hyphal tip three times. Four of the five isolates had similar morphology and caused leaf spot symptoms upon inoculation. During culture on PDA, these isolates formed colonies with downy, gray-brown mycelium and darker centers. Microscopic examination revealed branched, septate mycelium; solitary, erect, unbranched grayish-brown conidiophores; and club-shaped conidia. Conidia were faint brown, segmented, contained 2 to 15 septa(predominant number being 3 to 4), ranged from 32 to 220 μm in length and 8.4 to 22.4 μm in width (n = 30). The morphology of these isolates was identical to that of Corynespora cassiicola (Ellis 1971). Pathogenicity tests were conducted on 3-month-old seedlings of P. cablin using the conidium infection method. A suspension of conidia (1 × 106 conidia/ml) was prepared from cultures induced to sporulate by 90-minute near-UV exposure followed by 2-day dark incubation at 28 °C, and 30 ml of this suspension was sprayed onto leaves of each seedling. Inoculated plants were incubated at 28 °C in an incubator. Potted plants treated only with sterile water were used as controls. Each treatment was inoculated into five potted plants. After seven days, all the inoculated leaves displayed symptoms similar to those observed in the fields, whereas the control leaves did not exhibit these symptoms. The pathogenicity test was repeated three times. Following Koch's postulates, the pathogen was re-isolated and identified as C. cassiicola through morphological and ITS sequence each time. To further identify, we selected a representative isolate, LD-TJ, for multi-locus sequence analysis of its ITS, LSU, and TEF1-α genes (GenBank Accession Nos. PQ042036, PQ035023, PQ060235)(Voglmayr et al. 2017). BLASTN analysis of the sequences obtained showed a high similarity of 99 to 100% with the ITS (JAEMHE010000031.1:51214-51777, 481/481 nucleotides), LSU (JAEMHE010000018.1:728464-729373, 910/910 nucleotides), and TEF1-α (JAEMHE010000005.1:656736-657712, 969/970 nucleotides) sequences of C. cassiicola CC01. The phylogenetic tree showed that the LD-TJ strain, C. cassiicola CC01(isolated from rubber trees), and C. cassiicola CC_29(isolated from soybean leaves) clustered into a clade with a 99% bootstrap value. C. cassiicola was identified as the cause of patchouli leaf spot in Hainan(Chen et al. 2010), but it has not yet been reported in Guangdong. Identifying P. cablin leaf spot disease is crucial in Guangdong Province because it is the main growing area for P. cablin in China (Yan et al. 2021).
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