Development of Bursaphelenchus xylophillis-specific antibody for detecting the pine wilt disease

The pinewood nematode (PWN, Bursaphelenchus xylophilus) causes the pine wilt disease, transmitted to pinewoods by the pine sawyer beetle, Monochamus alternatus. It is very difficult to discriminate B. xylophilus from B. mucronatus. Therefore, it has been necessary to detect PWN-infected trees for the prevention of pine wilt disease transmission in a short time. The development of biomarkers such as DNA and protein is important for diagnosis of B. xylophilus. However, there have been no reports regarding biomarker identifications of B. xylophilus. In this study, polyclonal antisera were raised against whole proteins of B. xylophilus in BalbC mice and were primarily screened with ELISA. Twenty five among over 500 cell lines releasing polyclonal antisera revealed B. xylophilus-specific immuno-reactivity, which was at least three times higher than that of B. mucronatus. Three cell lines among them were secreting monoclonal antibody through further screening. These cell lines only detect about a 33-kDa protein in B. xylophilus in the western blot. These results suggest that these monoclonal antibodies will be useful for the development of diagnostic kit for the pine wilt disease.