Pine wilt disease (PWD) is one of the most important forest tree diseases, especially in the East Asian countries of Japan, China, and Korea. The Japanese pine sawyer, Monochamus alternatus Hope, is the PWN vector for Japanese red pine and Japanese black pine while Monochamus saltuarius Gebler is the vector for Korean white pine. Various control methods, such as aerial pesticide applications, biological control using parasitoids and fumigant such as methyl bromide are used. But the PWD still has spread. Therefore, we were selected effective aerial insecticides. Susceptibility of M. saltuarius and M. alternatus adults were investigated using 9 insecticides which are available in the market in Korea. And then, we tested them to the Apis mellifera to vertify the environmental impact.

The nematode Bursaphelenchus xylophilus is the causal pathogen of pine wilt disease in Korea. Currently, it is reported that B. mucronatus also has a low pathogenicity. Despite this difference in pathogenicity, it is very difficult to differentiate these two species due to similarity in morphological characters. The sequence variation of internal transcribed spacer (ITS) and intergenic spacer (IGS) regions of ribosomal DNA has been used for species identification and phylogeny of B. xylophilus and B. mucronatus. But, the IGS region sequence data of B. mucronatus has been only reported in Portugal. In this study, We analyzed genetic variation on ITS and IGS regions of B. xylophilus and B. mucronatus (Asian genotype, European genotype) based on rDNA gene sequences, and conducted phylogeography using TCS network. When the each isolates was determined the phylogenetic analysis using the neighbor-joining method, Bursaphelenchus species were divided into each groups, and showed low variation within each species. In the TCS analysis, The isolate of B. xylophilus and B. mucronatus (Asian genotype and European genotype) were divided into each groups and confirmed slightly genetic distance within species. B. mucronatus European genotype has possibility of new species different with Asian genotype.

Metam-sodium has been generally applied to control Pinus koraiensis, P. densiflora and P. thunbergii infected by pine wilt disease in the forest field of Korea. We determined the atmospheric concentration of nematocidal and insecticidal gaseous methyl isothiocyanate (MITC) and more volatile and toxic gaseous methyl isocyanate (MIC) in the metam-sodium fumigated fields depending on the distance from metam-sodium treated site, time and season by OSHA No. 52 method. Determined atmospheric MIC was below acute exposure limit (50 ㎍/㎥) and chronic exposure limit (1.0 ㎍/㎥) of EPA regulation in all detection point. In summer, the maximum of MITC and MIC was observed to 105.2 ㎍/㎥ in the 1st day and 0.41 ㎍/㎥ in the 4th day after metam-sodium treatment, respectively. On the other hand, in winter, the maximum of MITC and MIC was observed to 4.6 ㎍/㎥ in the 4th day and 0.52 ㎍/㎥ in the 5th day after metam-sodium treatment, respectively. Atmospheric concentrations of MITC and MIC were higher the closer to the metam-sodium treated site, but decreased below LOQ with distance (~ 50 m). This result showed that the reaction rate of metam-sodium to MITC is more affected by temperature and the translation rate of MITC to MIC is generally very low, regardless of season.

This study was conducted to examine the occurrence characteristics of pine trees infected by pine wood nematode in Gyeongsangbuk-do, Korea. Based on the GPS coordinate data of the infected trees at Yeongdeok-gun from 2013 to 2014 and at Andong-si in 2014, nearest neighbor analysis was conducted to analyze nearest distance between the infected trees. And the influence of geographical factors such as altitude, slope, and the distance from the road on the position of infected trees were identified. Our results showed that over 60% of the newly infected trees were found within 50m of the trees infected in previous year. Over 70% of infected trees occurred below 100m altitude in Yoeongdeok-gun whereas less 32% of total pine trees distributed below the altitude. 98% of the infected trees and 74% of total pine trees in Andong-si were found between 150 and 300m altitude, and altitudinal distribution of the infected trees was similar to that of pine forests. Only less 5% of infected trees found in the forests located below 5 degree of slope in Yeongdeok-gun whereas 18% of total pine trees existed in the areas. Over 50% of infected trees in the forests located from 10 to 20 degree of slope, and 47% of total pine trees were found in the forests located from 5 to 15 degree of slope. Most infected trees and total pine trees were found in the forests located below 15 degree of slope in Andong-si. Over 80% of the infected trees occurred within 1km of road in both study areas.

소나무재선충병에 대한 다양한 방제방법에도 고사목은 해마다 급격히 증가하고 있다. 그러나 현 방제방법 문제에 대한 연구는 매우 미흡한 실정이다. 재선충 확산 방지를 위해 가장 널리 쓰이는 방법은 병의 매개충인 솔수염하늘소 유충 폐사를 위한 감염목 훈증처리방법이다. 본 연구는 산림 내 소나무재선충 감염목 훈증처리지역에서 훼손 정도를 확인하고자 경상남도 밀양시 일대에 100㎡의 조사구 32개소를 설치하여 근원단면적과 수관단면적을 분석하였다. 훼손되지 않은 수목에서 도출한 근원단면적과 수관단면적의 회귀식(침엽수: R2=57.4, 낙엽활엽수: R2=63.8)을 바탕으로 훼손된 수목의 수관단면적을 유추하였다. 분석결과 수관단면적을 기준으로 재선충감염목은 18.7%이었으며 재선충과 무관하게 베어진 수목은 35.5%에 달했다. 소나무류의 경우 8.4%가 재선충과 관계없이 훼손되었으나 참나무류를 포함한 낙엽활엽수류는 훼손비율이 62%에 달했다. 소나무재선충 방재를 위한 훈증처리작업에 따른 아교목층 낙엽활엽수의 무분별한 제거는 산림의 온도저감, 습도조절 등 공익적 기능을 저하할 우려가 높았다.

The pine wilt disease that blocks the path for water and nutrition in pine trees is caused by the nematode, Bursaphelenchus xylophilus (Nematoda: Aphelenchoididae). The nematode relies on the longhorn pine sawyer beetle Monochamus alternatus and Monochamus saltuaris (Coleoptera: Cerambycidae) as vectors. Recently, 2-(undecyloxy)ethanol was identified as a male-produced aggregation pheromone of Monochamus species. In this study, we investigated the effect of 2-(undecyloxy) ethanol along with host plant volatiles -pinene and ethanol on attracting M. alternatus at a pine forest in Pohang, Korea from May, 2014 to July, 2014. To sustain the volatility of 2-(undecyloxy)ethanol and host plant volatiles, a superabsorbent polymer based on polyacrylic acids and water were added to the pheromone mixture. A total of 46 M. alternatus were collected from two field bioassays. Our results indicate that 2-(undecyloxy)ethanol is effective in attracting M. alternatus in Korea. Our study suggests that the aggregation pheromone could be used for detection and population monitoring of the beetles as well as for the effective mass trapping in outbreak situations.

After pine wilt disease (PWD) at Korean pine, Pinus koraiensis has been first reported in the middle area of South Korea in 2006, it spread rapidly to the nearest area. However, it is difficult to predict the spread of damage and set up the proper control strategy specific to P. koreaiensis because the pathology and the biology of PWD in P. koraiensis are not the same with PWD in P. densiflora and P. thunbergii distributed in middle southern area. So, we selected 25 of dead P. koraiensis in PWD damaged area and analyzed the inner-tree distribution of Bursaphelenchus xylophilus and the entrance holes of Monochamus saltuarius. There was no statistical significance between height or direction of wood extracted and the distribution of B. xylophilus among 25 dead pines. The entrance holes of M. saltuarius were concentrated on 7 dead pines without leaves and the maximum number of entrance holes in individual tree was 543. It indicates that the event of spawning of M. saltuarius adult and entrance of hatched M. saltuarius larvae to wood may be dependent on the time of death of host. The entrance holes were frequently found in a southward direction. These result shows that M. salutuarius larvae prefer relatively weak woody parts to enter easily. However, the relation between the thickness of bark of P. koraiensis and the frequencies of entrance holes of M. saltuarius was not observed.

Pine-wilt disease (PWD) is one of the most devastating forest diseases in Eurasia. PWD is known to be caused by pine wood nematode, Bursaphelenchus xylophilus which is indigenous in North America. Ever since PWD was found in Japan, PWD has been continuously spreading to Korea, Taiwan, China, Portugal and Spain. Because B. xylophilus infected trees died within 2 ~ 3 months, the most effective way to control PWD is preventing the translocation of infected trees to other regions. Thus, developing on-site diagnostic methods for identifying B. xylophilus infected tree is utmost important. Even though various cellular and molecular biological techniques were developed to identify B. xylophilus at the laboratory, they had certain limitations to be applied for on-site diagnostic methods. In this presentation, I will overview the recent advances in B. xylophilus detection methods. And then, I will present recent progression for developing on-site diagnostic methods for B. xylophilus made from my laboratory collaboration with Korea Forest Research Institute. The development of on-site diagnosis tool for PWD is one of the most arduous mission to accomplish. Thus, the accomplishment of our mission requires continued interest and support to PWD researches.

Pine wilt disease, caused by pine wood nematode, Bursaphelenchus xylophilus (Steiner et Buhrer) Nickle, has become the most serious threat to pine trees in Korea since first reported in 1988. Pine wood nematode is transferred to uninfested trees by Monochamus alternatus (Coleoptera: Cerambycidae) (vector). A typical controlling method against vectors in Korea is fumigation of the dead trees infested by vector M.alternatus and B.xylophilus using metam-sodium SL(25%). However, this method is harmful to the environment because of the chemical contamination and vinyl waste in the mountain afterward. Portable Bark Remover (PBR) was contrived to reduce these environmental problems. The vectors oviposite under the bark of the freshly dead trees only. Debark infested trees prohibited the vectors from oviposite and finally, vectors can not complete their life cycle. The PBR is a newly designed as a debarking device that is equipped on top of the chain saw engine allowing ease and rapid debarking of the infested trees at the controlling field. Debarking of freshly-dead trees can eliminate egg-laying and hatch places where between the wood and bark from vectors. The new method for vector controlling using the debarking device is expected to lower-price and high-efficiency comparing with other conventional methods such as “crushing”, “burning” and “fumigation”.

Pine wilt disease (PWD) represents a major threat to forest ecosystems worldwide. Although PWD is now better understood, effective control measures for this disease have still not been devised. Here, we report several years of field studies on preventative silvicultural control of PWD. Silvicultural control through preventative clear-cutting and the manual removal of logs was implemented between 2005 and 2009 in 16 Korean districts that had newly PWD-infected stands. Preventative clear-cutting of neighbouring asymptomatic pine trees (within a 10 to 50-m radius ofwilt trees) and the removal of felled logs or branches suppressed spread of PWD. Occurrences of PWD wilt pines in districts (city or county) subjected to this silvicultural control method were significantly reduced compared with those in districts using conventional controls (physical or chemical treatment of wilt pine trees). Through silvicultural control, PWD was successfully suppressed in 11 of 16 districts investigated. In contrast, successful control was achieved in only 1 among 18 districts subjected to conventional control. Our results will be of considerable interest to those engaged in the very difficult battle against the global spread of PWD.

The most effective methods against pine wilt disease (Bursaphelenchus xylophilus, PWD), black pine bast scale (Matsucoccus thunbergianae), pine needle gall midge (Thecodiplosis japonensis) is the trunk injection of insecticides. The period of trunk injection that was considered with ecology and physiology of pests such as pine wilt disease, black pine bast scale and pine needle gall midge, was applied from Dec. to next Feb. and June, Dec. respectively. And there are differences in quantities of being injected chemicals to pine trees by the period of trunk injection. Thus, we investigated to prevent the effect of insecticides, to estimate the effect under the trunk injection of insecticides for pine trees against pests by the injecting period and time, boring site, opening and shutting of injection site. On Pinuns thunbergii, it was examined to figure out the effect of insecticides by the injecting period, twice a month on a sunny day around 10 AM from January to September. Injecting of insecticides was tested at a dose of 5ml per cm dbh of a pine tree after boring with hand drill at 50 cm high from the ground. As a result, abmectin 1.8% EC and phospamidon 50% SL were injected over 90% of mortalities from the end of January to the middle of March but as time goes by, injecting insecticides tended to decrease. To compare the volume of injecting insecticides in a day, between 10 AM and 2 PM on February 23rd, abamectin 1.8% EC was injected at a dose of 5ml per pine trees with micro pipette after boring with a drill machine at 50 cm high from the ground. As a result, injecting insecticide of a pine which was injected around 10 AM was completely injected and about 90% of the insecticide was injected in case of a pine that was injected around 2 PM. Trunk injection of insecticides was examined to figure out differences of the volume of injected insecticides before and after rain. There was no difference not only the rainfall but also the bored direction.

Pine wood nematode, Bursaphelenchus xylophilus, is a causative organism to induce pine wilt disease in many varieties of pine trees. Until 2006, Monochamus alternatus had been known as the only insect vector of pine wood nematode in Korea which targeted on Pinus densiflora (Japanese red pine) and P. thunbergii (Japanese black pine). However, pine wilt disease was also reported from Korean pine tree (Pinus koraiensis) in 2006 and we found another insect vector, M. saltuarius, was involved to transmit pine wood nematode. Both Monochamus species were confirmed to transfer pine wood nematode to their hosts but, there is no detail information about other transmitted nematode. Especially Bursaphelenchus mucronatus is common species transmitted by Monochamus species which is morphologically closed to B. xylophilus. Moreover B. mucronatus have two genotypes; one is East Asian type and the other is European type. Both genotypes of B. mucronatus were found in Korea but, the host and vector information related to the genotypes of B. mucronatus was not clear. Monochamus saltuarius was collected from three different geographical locations and nematodes were extracted and identified. For the identification of the juveniles, nematode DNA was extracted and ITS-RFLP analysis was done by PCR and gel electrophoresis. The selected enzymes were Hinf I, Alu I, Msp I, Hae III, Rsa I. Most of Bursaphelenchus species carried by M. saltuarius, which collected from pine wilt disease-free area, was determined as European type of B. mucronatus. We will compare the nematode species and genotypes carried by M. alternatus and M. saltuarius. In addition the rate of nematode carrying insect and the average number of nematode per single insect will be counted and compared.

A survey was conducted in the northern conifer forests of Korea from February to May in 2009 for the Monochamus species vectored Bursaphelenchus species. Dead pine trees infested with bark beetle larvaes were collected from Pinus densiflora and P. koraiensis located in the following target areas: Pocheon, Yeongi, Jincheon, Chuncheon, and Bonghwa. A cerambycid beetle, M. saltuarius was only found from sampled log piles in target areas. Bursaphelenchus species carried by M. saltuarius was only isolated in Pocheon, Gyeonggi province. The nematodes recovered from 17 (71%) of 24 beetles from P. koraiensis and 12 (75%) of 16 beetles from P. densiflora. The average number of nematode per adult was 1,974 and 11,823 from P. koraiensis and P. densiflora, respectively. The nematode was also recovered from the inner bark of both Pinus species infested with M. saltuarius larvae. Bursaphelenchus species identification was performed by morphological characters and molecular analysis with ITS-RFLP patterns and sequences of ITS and D2D3 region. Both morphological and molecular characters match well with the original description of the European type of B. mucronatus. This is the first report of M. saltuarius species as a vector of the European type of B. mucronatus on both P. densiflora and P. koraiensis in Korea.

Since pine wilt disease was first reported in Busan in 1988, this disease has become a serious threat to pine forests in Korea. A general survey for Bursaphelenchus xylophilus group (B. xylophilus, B. mucronatus) nematodes were carried out to determine their geographic range, host tree species and insect vectors, Monochamus species. Based on the survey results, B. xylophilus were found over 57 cities or district and a similar nematode, B. mucronatus also has been isolated over 30 cities or district including Jeju and Pocheon. The Asian type of B. mucronatus was founded from Pinus thunbergii and P. densiflora, generally associated with M. alternatus as its insect vector. The European type of B. mucronatus which was limited distribution to the central areas, such as Yeongwol, Bonghwa, Yeongdeok, was also found on P. koraiensis and P. desiflora associated with M. saltuarius. Both Monochamus species transmitted both Bursaphelenchus species and co-existed in Muju and Yeongdeok predicted as their range margin. Their range margins shifted northwards compared to 2006, seemingly as a result of climate change. Recently, the European type of B. mucronatus has been identified in other several areas since it was first reported in Pocheon in 2008. Several reports indicate the potential pathogenicity of European type of B. mucronatus and other species, B. sexdentati and B. lenoni in Europe. Therefore, sustainable survey and the study for the newly invading Bursaphelenchus species as well as the member of B. xylophilus group should be performed to protect Korean forests.

The pinewood nematode (PWN, Bursaphelenchus xylophilus) is known as a virulent factor of the pine wilt disease, transmitted to pinewoods by the pine sawyer beetle, Monochamus alternatus. It is very hard to discriminate B. xylophilus from B. mucronatus because these Bursaphelenchus species are genetically and biochemically very close. Therefore, it has been necessary to detect PWN-infected trees for the prevention of pine wilt disease transmission in a short time. We developed polyclonal antibodies against B. xylophilus in BalbC mice and primarily screened with ELISA. Positive clones releasing polyclonal antisera revealed B. xylophilus-specific immuno-reactivity, which were at least two times higher than that of B. mucronatus. Two clones, D9-F10 and 1F3, were finally selected and exhibited specific immuno-reactivity for B. xylophilus, not for B. mucronatus in Western blot analysis. D9-F10 clone was reactive with a 43-kDa whereas 1F3 clone with two proteins, 40- and 45-kDa. Their isotypes against mouse Ig family were identical, kappa-light chain. These results suggest that these monoclonal antibodies can be useful for the development of diagnostic kit for the pine wilt disease.

본 연구는 소나무재선충병의 피해를 받은 토양생태계의 합리적인 관리를 위한 기초정보를 제공할 목적으로 이루어졌다. 토양온도와 수분함량은 완전벌채지 15.3℃, 11.5%, 일부벌채지 14.4℃, 13.5%, 대조구 13.7℃, 14.8%로 나타났다. 토양유기물 함량은 완전벌채지가 2.11~2.64%, 일부벌채지 2.26~3.33%, 대조구 2.27~3.10%의 범위에 있은 것으로 분석되었다. 토양호흡량은 계절적인 차이가 있었는데 여름철에 높았으며, 토양온도와 정의 상관이 있는 것으로 나타났다. 평균 토양호흡량은 완전벌채지 0.24, 일부벌채지 0.36, 대조구 0.32 gCO2/㎡/hr로 나타나 소나무재선충 피해목만 벌채한 일부벌채지의 토양호흡량이 가장 높은 것으로 분석되었다. 소나무재선충병 피해지 산림토양의 Q10값은 2.39에서 2.68 사이에 있는 것으로 나타났다. 조사기간을 통한 년간 토양호흡량은 완전벌채지, 일부벌채지, 대조구에서 각각 8.1, 15.6, 14.6 tCO2/ha/yr으로 분석되었다.

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.

This study is conducted to get current state of data collection and utilization of the trees damaged by pine wilt disease, and to efficiently collect and utilize of the trees damaged by the pine wilt disease and abandoned fumigation-treated trees at forecasts. The method to control pine wilt disease damaged area is mostly fumigation treatment system, and there is no collection and utilization of damaged trees because of absence of efficient collection system, lack of collection cost, and absence of policy, etc. A survey is conducted that asked about the satisfaction degree for the trees damaged by pine wilt disease collection and use divided into 6 topics. It was positively recognized that the need and problem of the trees damaged by pine wilt disease, and results and satisfaction of forestry mechanization while it was mostly negatively recognized that the policy satisfaction for prevention and collection, satisfaction of collection and use, collection and forestry mechanization, and satisfaction of foundation equipment for forestry mechanization, etc. As a result of path analysis, it is necessary to promote a high-tech forestry mechanization to improve satisfaction level of results and satisfaction of forestry mechanization for the trees damaged by pine wilt disease collection and utilization, it is also need to make an effort in various ways for improve satisfaction level of satisfaction of collection and utilization.