Abstract: The plane-parasitic nematode Bursaphelenchus xylophilus is the causal agent of pine wilt disease, which is the most serious conifer disease in the world. However, it is difficult to differentiate isolates of B. xylophilus from the closely related species B. mucronatus, which is not pathogenic to pine trees. So it is very important to distinguish B. mucronatus from B. xylophilus rapidly. We applied the two-dimensional electrophoresis and mass spectrometry to study the proteomics of the two nematodes. And we have obtained 45 differentiate proteins, 22 of which were from B. xylophilus and others were from B. mucronatus. The results supply new insights into the rapid identification of B. xylophilus the virulence of B. xylophilus.