| Summary: | <i>Bursaphelenchus xylophilus</i> (Steiner & Buhrer) Nickle, is a serious forest pest, causing enormous economic losses in pine trees in Korea, China, Japan, and countries in Western Europe. To prevent pine wilt disease (PWD), trunk injection with nematicide is performed in Korea. Although these nematicidal agents are quite efficient, the development of new nematicidal agents is needed to prevent pesticide resistance and reduce pest management costs. The aim of this study was to investigate nematicidal activities of pure naphthoquinones (NTQs)−1,4-NTQ, juglone, and plumbagin—against <i>B. xylophilus</i> via in vitro and semi-in vivo assays to identify new candidate agents for trunk injection. Estimated LC<sub>50</sub> values (48 h exposure) were 100.0 ppm, 57.0 ppm, and 104.0 ppm for 1,4-NTQ, juglone, and plumbagin, respectively. In the semi-in vivo assay on pine bolt of the Japanese black pine, <i>Pinus thunbergii</i>, the population of inoculated <i>B. xylophilus</i> was significantly decreased at two weeks after treatment with juglone when compared with the effects of treatment with 1,4-NTQ and plumbagin. We also observed that naphthoquinones could generate reactive oxygen species, which presumably indicated that naphthoquinones caused significant oxidative stress in <i>B. xylophilus</i>. The findings of this study suggest the nematicidal potential of naphthoquinones and their possible use in further in vivo assays to test their nematicidal efficacy against <i>B. xylophilus</i> when injected through trunk injection.
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