Detection of Bursaphelenchus xylophilus in Mid- to High-Elevation Coniferous Forests of Yunnan, China, and Symptom Development in Inoculated Seedlings

Mid- to high-elevation coniferous forests of Yunnan are characterized by complex topography, but direct evidence remains limited regarding the occurrence of the pine wood nematode, Bursaphelenchus xylophilus, under montane forest conditions. In this study, 45 wood segment samples were collected from trees in three mid- to high-elevation coniferous forest sites in northwestern Yunnan whose crowns showed approximately one-third needle discoloration. Nematodes were extracted using the Baermann funnel method and identified based on the 28S rDNA D2–D3 region, phylogenetic analysis, and auxiliary morphological observations. A total of 10 molecular nematode taxa were detected. Bursaphelenchus mucronatus-like was the most frequent taxon, whereas B. xylophilus was detected as a single individual from one Pinus densata sample. In inoculation experiments with the pine wood nematode strain PNFT01, Pinus yunnanensis seedlings developed progressive needle chlorosis and wilting, P. densata showed only mild symptoms, and Picea likiangensis remained symptomless during the 45-day observation period. Sterile-water controls remained symptomless throughout the experiment. Symptom scores differed significantly among host species at 30 and 45 d after inoculation (Kruskal–Wallis test, p = 0.001), and pine wood nematodes were re-isolated from all three host species. These results provide molecular evidence for rare B. xylophilus occurrence in wood from weakened trees in mid- to high-elevation coniferous forests of Yunnan and indicate contrasting symptom development among inoculated seedlings. The findings do not demonstrate established pine wilt disease outbreaks in the region, but they support continued monitoring of the nematode, potential vector insects, and susceptible hosts under montane forest conditions.