LAUSR

Justicia (Justicia procumbens) is an herbaceous plant widely distributed in tropical and sub-tropical areas of the world. It has been traditionally used to treat fever, edema, cough and pain (Ibrahim et al., 2023). In September 2024, leaf spot symptoms and defoliation, reaching a severity of 25% to 35%, were observed for the first time on Justicia plants in Nanyang (32°58'35'' N; 112°29'30'' E), China. Symptoms started with light brown necrosis surrounded by dark margins, and green areas eventually turned into dark brown necrosis. Leaf pieces (5 × 5 mm) excised from symptomatic tissue (from 25 leaves) were surface disinfested with 75% ethanol for 30 s, 2% NaClO for 1 min, then rinsed three times with sterile water and dried for 30 min on sterile filter paper. Leaf pieces were placed on potato dextrose agar (PDA) medium and incubated at 25℃. Hyphal tips from each isolate originating from Justicia leaf pieces were transferred to a new PDA plate to obtain a pure culture. A total of 34 isolates were obtained, and three isolates named JC-7, JC-11 and JC-16 were selected for further study. The representative isolates were identical by morphological characteristics, with brown, dark colonies in the culture. The colonies had a dark grayish black with cottony mycelia; mycelia were hyaline, smooth and septate. Conidia (n = 50) measured 23.1 ± 5 µm (28.5 to 18.5 µm) in length and 10.5 ± 2.1 µm (13.2 to 8.9 µm) in width and were fusiform, dark brown, melanized with three transverse septa. Colony and conidia morphology were aligned with those of Curvularia spp. (Priwiratama et al. 2024). The genommic DNA was extracted using Chelex 100 method. PCR reactions targeting internal transcribed spacer region (ITS) and large subunit region (LSU) were carried out using the primers ITS1/ITS4 and LR5/LROR (Manamgoda et al. 2012), respectively. BLAST analysis demonstrated that sequences of ITS (accession nos. PQ725734 to PQ725736) and LSU (accession nos. PQ725767 to PQ725769) were 99.64% to 100% identical to the ex-type isolate of C. americana strain UTHSC 08-3414 (Madrid et al. 2014). A maximum likelihood analysis based on the concatenated ITS and LSU sequences per isolate, placed the isolates JC-7, JC-11, and JC-16 in a high-confidence cluster with C. americana. The pathogenicity tests were done on 24 healthy plants. For each isolate, six plants were wounded and inoculated by dropping 10 μL of conidia suspension (1 × 106 conidia/mL), for a total of 18 plants. Ten μL of sterile water was added to the remaining six plants, serving as a negative control. All plants were incubated in a greenhouse at 25 ℃ and 75% relative humidity. After 10 d, similar symptoms to those observed in the field developed on the 18 plants that were inoculated with conidia suspensions, while no symptoms developed on the plants used as negative controls. Pathogens were successfully reisolated from selected symptomatic plants and confirmed to be C. americana based on morphological characterization and molecular analysis. Inoculations were repeated twice with the same results. C. americana is a significant phytopathogenic fungus that causes leaf spot and stem necrosis that has been reported on Bermuda grass (Gilardi et al., 2022), Oryza sativa (Heidarian et al., 2020) and Vitis vinifera. To our knowledge, this is the first report of C. americana infecting Justicia plants in China. Since it poses a serious threat to other hosts, especially to O. sativa, precautions and management strategies may be necessary to minimize the spread of this fungi.