Monochamus alternatus which is major vector for Bursaphelenchus xylophilus is main pest for pine trees. Moreover, their larvae mainly feed on the phloem of the host during the growth period, and this process makes host debilitate. The reason why the epidemiological investigation of that damage caused by Monochamus alternatus is difficult is that the adult and the larva remains very rarely in the host. Even if the larvae remain in the host, it is not easy to distinguish them from other cerambycidae coleopterans without expertise. For the above reason, we introduce the method of field epidemiological investigation for Monochamus alternatus using the remaining larva frass in the host with Recombinase Polymerase Amplification (RPA) method.

Bursaphelenchus xylophilus (Bx) is the main plant-parasitic nematode of the Japanese Black Pine (Pinus thunbergii), Red Pine (Pinus densiflora) and Korean Pine (Pinus koraiensis) in the South Korea. Until now, the nematode morphological classification or PCR method using specific marker of Bx were used for the diagnosis of pine wilt disease. However, both methods have a disadvantage that these take a long time to confirm the result. Thus, these methods can not be used quickly at the newly damaged regions. For above the reasons, we had been developed the diagnostic method for Bx combining direct gDNA extraction buffer (DAP) with Recombinase Polymerase Amplification (RPA). This method is able to directly use mixed lysates extracted from Bx-infected pinewood by DAP buffer as gDNA template to RPA without another process for increase gDNA yield. Together, our method is able to detect Bx within 20 mins.