First Report of a New Species, Trichomerium puerense, Causing Sooty Mold on Coffea arabica in China.

Coffea arabica (Rubiaceae) is one of the most popular coffees worldwide (Zhu et al. 2024). In April 2024, symptoms of sooty mold appeared in the largest coffee-growing region in Yunnan Province, China (22°66'N; 100°96'E; 1073 m). The disease initially appears as a black powdery coating on coffee leaves, cherries, and twigs, later forming a thick, adherent layer that cracks or peels off under dry conditions. A field survey was conducted over approximately 20 hectares (300 mu) of coffee plantations, and fresh symptomatic leaves were collected and observed. Micro and macro photographs were prepared for photomicrography. Pure cultures were obtained following the methods in Senanayake et al. (2020). Pure cultures were grown on potato dextrose agar at 28 °C for one month and used for DNA extraction and the pathogenicity test. Morphology and multi-locus phylogeny confirmed that the sooty mold is a distinct new species in Trichomerium: Trichomerium puerense L. Lu, K.D. Hyde & Tibpromma, sp. nov. MycoBank number: MB 857498. Etymology: The epithet refers to the location "Pu'er" from where the holotype was collected. Holotype: MHZU 24-0582. It forms black sheets which are superficial and thin, cover the leaves, and are composed of cylindrical hyphae. Superficial hyphae 4-6 μm wide also form, are septate, constricted at the septum, unbranched, brown, dense, with cylindrical hyphal cells, and produce guttules. Ascostromata are 100-150 × 120-150 μm, superficial, solitary, globose to subglobose, and brown to black. Setae are 40-80 × 4-7 μm, straight or flexuous, brown, and septate. Peridia are 15-20 μm wide, thick-walled, hyaline to brown, and comprised of textura angularis cells. Asci are 45-70 × 20-28 μm (n = 30), 8-spored, bitunicate, ellipsoid to clavate, or cylindrical, with a short pedicle and an ocular chamber. Ascospores are 20-25 × 7-10 μm (n = 10), fusoid to ellipsoid, hyaline, 1-3-septate, constricted at the septum, with narrow ends, somewhat tapering towards the base when mature, smooth-walled, and produce guttules. DNA fragments were amplified and sequenced with ITS, LSU, and SSU (Hyde et al. 2020). The sequences were deposited in GenBank (ITS, PV056513, PV056514; LSU, PV056517, PV056518; SSU, PV056515, PV056516). The morphology of the collection (sm4) was consistent with Trichomerium (Rana et al. 2019; Hyde et al. 2020). Maximum likelihood and Bayesian Inference analyses on CIPRES confirmed that our isolates formed a distinct lineage within Trichomerium with 100% ML/1.00 PP statistical support. Based on morphology and phylogeny, sm4 was identified as a new species, T. puerense. To fulfill Koch's postulates (Xu et al. 2019), ten healthy arabica coffee plants were selected. Five plants were inoculated with two 3-mm fungus plugs per leaf sheath, and the other five plants were inoculated with PDA agar plugs (control group). All plants were sealed in transparent plastic bags and incubated in a greenhouse at 26 ± 2 °C for 28 days, with daily moisturizing using a 5% bee honey solution (with relative humidity maintained at 70%). Four weeks after inoculation, black sooty mold spread on the leaves that were inculated with the fungus plugs, while no symptoms were observed on the control leaves. This study represents the first report of T. puerense associated with coffee sooty mold in China. This finding will facilitate further research on defense strategies against coffee sooty mold and support the development of improved management practices for this disease.