Identification and Biological Control of Pestalotiopsis microspora Causing Leaf Spot Disease on Amomum villosum

Abstract

Amomum villosum is an evergreen perennial herb widely distributed in tropical and subtropical regions, with significant economic and medicinal importance. Throughout the cultivation process, it faces severe fungal disease problems that significantly impact its yield and quality. In July 2025, leaf spot disease with a 56% incidence was observed on A. villosum in Wenshan, Yunnan, China. Affected leaves initially developed irregular grayish-white lesions surrounded by brown margins. As symptom development progressed, semi-submerged black spots formed on the lesion surfaces. Severe infections resulted in premature leaf abscission and, ultimately, death of the entire plant. To identify the pathogen responsible, we conducted isolation and pathogenicity studies. Through morphological characterization, phylogenetic analysis (ITS, LSU, and TUB), and pathogenicity tests, Pestalotiopsis microspora was determined as a pathogen. Koch’s postulates were fulfilled on attached leaves. After 15 days, typical necrotic lesions appeared on inoculated leaves, while controls remained symptom-free. This is the first report of A. villosum leaf spot caused by P. microspora in China. Biocontrol assays revealed that Trichoderma harzianum T15 and T. asperellum T16 exhibited significant antagonistic activity against P. microspora, with inhibition rates of 44.77% and 50.70%, respectively. In addition, Bacillus velezensis SWFU41, isolated from healthy A. villosum leaves, showed superior disease suppression, achieving a control efficacy of 60.52%. Compared with the fungal biocontrol agents, B. velezensis demonstrated greater inhibitory activity against the pathogen, suggesting that bacterial antagonists may provide a more effective biological control strategy for managing A. villosum leaf spot disease. This is the first report of P. microspora-caused leaf spot disease on A. villosum. Furthermore, the comparative evaluation of fungal and bacterial antagonists demonstrated that B. velezensis SWFU41 outperformed T. harzianum and T. asperellum in disease suppression, highlighting its potential as a promising biocontrol agent for sustainable disease management. These findings provide a scientific basis for pathogen monitoring, epidemiological studies, and the development of integrated biological control strategies for A. villosum cultivation.