Pine wood nematodes (Bursaphelenchus xylophilus, PWNs) are a major threat to Pinus koraiensis in northeast China, and emamectin benzoate (EB) is commonly used for their control. Although high doses of EB can alleviate symptoms of pine wilt disease (PWD), they do not fully eradicate PWNs due to their detoxification mechanisms. This study investigates the content of EB in P. koraiensis and its efficacy in controlling PWNs after exogenous application of EB. We found that while EB significantly reduced PWN populations, it did not eliminate them. Transcriptomic analysis of PWNs treated with concentration at 20% (LC₂₀) revealed that PWNs exhibit detoxification responses to low EB concentrations (LC₂₀), driven by the Bx-SDR3 gene. RNA interference (RNAi)-mediated silencing of this gene decreased the detoxification ability of PWNs and enhanced the toxic effects of LC₂₀ EB by 20.9%. These results highlight the key role of Bx-SDR3 in PWN detoxification and suggest that targeting this gene could improve the effectiveness of EB, offering a promising strategy for more efficient and eco-friendly pest management.