• Molecules and Cells
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• v.26 no.1
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• pp.81-86
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• 2008

RNA interference (RNAi) was performed on several essential genes in the pinewood nematode Bursaphelenchus xylophilus, which causes pine wilt disease. Double-stranded RNA (dsRNA) was delivered to larvae or adult worms by soaking, electroporation, or microinjection. Soaking and electroporation of L2-L3 stage worms in solutions containing dsRNA for essential genes induced over 25% lethality after 5 days, and gene-specific phenotypes were observed. This lethality agreed with significant reductions of the targeted transcripts, as assayed by reverse-transcription coupled with real time PCR. Microinjection was the most efficient route as measured by the hatching rate of F1 embryos, which was reduced by 46%. When adult worms were soaked in dsRNA, lethality was induced in the F1 larvae, revealing the persistence of knockdown phenotypes. The penetrance of the RNAi phenotypes for essential genes was relatively low but consistent, indicating that RNAi should be useful for studying the in vivo functions of B. xylophilus gene products.

• Korean journal of applied entomology
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• v.31 no.4
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• pp.427-451
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• 1992

The nematodes associated with forest trees were surveyed in the National Parks and Mountains in Korea. A total of 73 species on 28 genera were identified from rhizosphere of 154 tree s species. Platycarya strobilacea, Ulmus davidiana and Acer ginnala were known to new host p plants of Meloinema kerongens. Bursaphelenchus xylophilus was detected only in Pusan on Pi$\pi$us t thunbergii, B. mucronatus was detected in Cheju, Masan, and Chinhae. Insect parasitic nemat todes, Prothallonema intermedium, on P. rigida in Daegu, P. thunbergii in Cheju city, Namwonu up, Gujoa-up and Aewol-up. The number of nematode species according to forest trees was t the highest on P. thunbergii to 32 species, followed by 24 species on Abies holophylla, 20 species on Zelkova serrata, 19 species on Diospyros kaki, 18 species on Quercus acutissima, respectively. L Localities of distribution according to nematode species were as follows; Crico$\pi$emella i$\pi$formis i in 81, X. americanum in 74, Tylenchorhynchus claytoni in 51, C. macrodora in 35, H Hemicriconemoides varionodus and Ogma serratum in 31, Pararotylenchus pini in 25, H Hemicycliophora koreana in 24, Aphelenchus avenae and C. pseudohercyniensis in 24, respectively.

• Journal of Korean Society of Forest Science
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• v.102 no.2
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• pp.300-304
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• 2013

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 1988. Pine wood nematode is transferred to healthy trees by Monochamus alternatus (Coleoptera: Cerambycidae) during its maturation feeding and female oviposition. A typical control method against insect vectors in Korea is fumigation of the dead trees by using metam-sodium SL (25%). However, this method is not environment friendly because of the forest contamination by chemical application and destroying landscape by plastic cover. Portable Bark Remover (PBR) was invented to reduce these environmental problems. The vectors oviposit under the bark of the newly dead trees only. Debarking infested trees prevents the vectors from laying eggs and eventually, they can not complete their life cycle. The PBR is a modified debarking device that is attached on the top of the electrical chain saw, which allows ease and rapid debarking of the infested trees. The new method by PBR is expected to be more economic and effective than other conventional methods such as "crushing", "burning" and "fumigation".

• Korean Journal of Breeding Science
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• v.41 no.4
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• pp.377-384
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• 2009

Three-year-old seedlings of susceptible Pinus densiflora and resistant Pinus x rigitaeda were each inoculated with the pinewood nematode, Bursaphelenchus xylophilus, to compare disease development. Needle dehydration was evident on seedlings of P. densiflora by 20 days after inoculation, 10 days earlier than this symptom was observed on P. ${\times}$ rigitaeda. Xylem drying was more frequent in seedlings of P. densiflora than in that of P. ${\times}$ rigitaeda between 20 and 60 days after inoculation. No significant differences were found between P. densiflora and P. ${\times}$ rigitaeda for stem water content or for stem and leaf relative water content in current-year branches after nematode inoculation, but the average number of B. xylophilus recovered from stems differed significantly between the two groups. The number of B. xylophilus recovered from stems was negatively correlated with the stem water content and with stem and leaf relative water content. By the time the experiment was terminated at 60 days after inoculation, all 3 of the last group of P. densiflora seedlings had died, but 2 of the 3 remaining P. ${\times}$ rigitaeda hybrid seedlings were still alive. Additional studies are needed to further explore the specific mechanisms preventing nematode multiplication in the seedlings of resistant P. ${\times}$ rigitaeda.

• Journal of Microbiology and Biotechnology
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• v.29 no.7
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• pp.1117-1123
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• 2019

Control of pine wilt disease, which is caused by pine wilt nematode Bursaphelenchus xylophilus, is heavily dependent on the use of chemicals such as abamectin. Although such chemicals are highly effective, demands for alternatives that are derived preferentially from natural sources, are increasing out of environmental concerns. One of the challenges to discovery of alternative control agents is lack of fast and efficient screening method that can be used in a high-throughput manner. Here we described the development of colorimetric assay for the rapid and accurate screening of candidate nematicidal compounds/biologics targeting B. xylophilus. Contrary to the conventional method, which relies on laborious visual inspection and counting of nematode population under microscope, our method utilizes a redox dye that changes its color in response to metabolic activity of nematode population in a given sample. In this work, we optimized parameters of our colorimetric assay including number of nematodes and amount of redox dye, and tested applicability of our assay for screening of chemicals and biologics. We demonstrated that our colorimetric assay can be applied to rapid and accurate quantification of nematode viability/mortality in a nematode population treated with candidate chemicals/biologics. Application of our method would facilitate high-throughput endeavors aiming at finding environment-friendly control agents for deadly disease of pine trees.

• The Plant Pathology Journal
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• v.39 no.3
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• pp.290-302
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• 2023

Abamectin offers great protection against Bursaphelenchus xylophilus, a well-known devastating pathogen of pine tree stands. Trunk injection of nematicides is currently the most preferred method of control. This study aimed to evaluate the potency of the commonly used formulations of abamectin against B. xylophilus. Twenty-one formulations of abamectin were evaluated by comparing their sublethal toxicities and reproduction inhibition potentials against B. xylophilus. Nematodes were treated with diluted formulation concentrations in multi-well culture plates. And, populations preexposed to pre-determined concentrations of the formulations were inoculated onto Botrytis cinerea culture, and in pine twig cuttings. Potency was contrastingly different among formulations, with LC₉₅ of 0.00285 and 0.39462 mg/ml for the most, and the least potent formulation, respectively. Paralysis generally occurred at an application dose of 0.06 ㎍/ml or higher, and formulations with high sublethal toxicities caused significant paralysis levels at the tested doses, albeit the variations. Nematode reproduction was evident at lower doses of 0.00053-0.0006 ㎍/ml both on Botrytis cinerea and pine twigs, with significant variations among formulations. Thus, the study highlighted the inconsistencies in the potency of similar product formulations with the same active ingredient concentration against the target organism, and the need to analyze the potential antagonistic effects of the additives used in formulations.

• The Plant Pathology Journal
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• v.39 no.1
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• pp.75-87
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• 2023

The pine wood nematode (PWN), Bursaphelenchus xylophilus is a well-known devastating pathogen of economic importance in the Republic of Korea and other countries. In the Republic of Korea, trunk injection of nematicides is the preferred method of control. In this study, the efficacy of 16 locally produced formulations of emamectin benzoate against the PWN are compared through determining their sublethal toxicities and reproduction inhibition potentials. Nematodes were treated with varying concentrations of the tested chemicals in multi-well culture plates, and rates of paralysis and mortality were determined after 24 h. Reproduction inhibition potential was tested by inoculating pre-treated nematodes onto Botrytis cinerea, and in pine twig cuttings. Despite the uniformity in the concentration of the active ingredient, efficacy was contrastingly different among formulations. The formulations evidently conformed to three distinct groups based on similarities in sublethal activity (group 1: LC₉₅ of 0.00768-0.01443 mg/ ml; group 2: LC₉₅ of 0.03202-0.07236 mg/ml, and group 3: LC₉₅ of as high as 0.30643-0.40811 mg/ml). Nematode paralysis generally occurred at the application dose of 0.0134-0.1075 ㎍/ml, and there were significant differences in nematode paralysis rates among the products. Nematode reproduction was only evident at lower doses both on B. cinerea and pine twigs, albeit the variations among formulations. Group 1 formulations significantly reduced nematode reproduction even at a lower dose of 0.001075 ㎍/ml. The variations in efficacy might be attributed to differences in inert ingredients. Therefore, there is need to analyze the potential antagonistic effects of the large number of additives used in formulations.

• Journal of Microbiology
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• v.43 no.1
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• pp.17-20
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• 2005

Poria cocos, a famous traditional Chinese medicine, was found to have nematicidal activity in experiments searching for nematicidal fungi. The experiment showed it could kill 94.9% of the saprophytic nematode, Panagrellus redivivue, 92.6% of the root-knot nematode, Meloidogyne arenaria, and 93.5% of the pine nematode, Bursaphelenchus xylophilus, on PDA plate within 12 hours. According to the nematicidal activity, three new compounds, 2, 4, 6-triacetylenic octane diacid, 2, 4, 5, 6-tetrahydroxyhexanoic acid and 3, 4-dihydroxy-2-keto-n-butyl 2,4,5,6-tetrahydroxyhexanate, were isolated from submerged cultures of Poria cocos. Of these, 2, 4, 6-triacetylenic octane diacid could kill 83.9% Meloidogyne arenaria and 73.4% Panagrellus redivivus at 500 ppm within 12 hours. Here, it is reported for the first time that Poria cocos has nematicidal activity.

• The Korean Journal of Pesticide Science
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• v.13 no.4
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• pp.267-274
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• 2009

This study was carried out to select effective preventive pesticides against pine wilt disease caused by pinewood nematode (PWN), Bursaphelenchus xylophilus on trunk injection. 1,000 fold aquatic solution of abamectin 1.8% EC and emamectin benzoate 2.15% EC were lower mortality (7.3% and 8.3% respectively) against PWN on 1 day after treatment. However effects of abamectin 1.8% EC, emamectin benzoate 2.15% EC, fosthiazate 30% SL and fenitrothion 30% SL were inhibited the reproduction of PWN over 99.6% in Botrytis cineria media. Effect of trunk injection of abamectin 1.8% EC and emamectin benzoate 2.15% EC at the rate of $10\;m{\ell}$ per 10 cm in diameter of breast height (DBH) on mortality of Japanese black pine, Pinus thungergii by inoculated PWN was 0% and 3.3%, respectively at the applied year however when injection of fosthiazate 30% SL were treated with the rate of $5\;m{\ell}$ per 10 cm tree DBH, mortality of tree was 63.3%. Abamectin 1.8% EC and emamectin benzoate 2.15% EC was showed high preventive efficacy representing >90% against PWN at the following year. PWN preventing efficacy of trunk injection was lower in naturally occurred area (mortality of pine tree in control was 11.7% at the first year) of PWN than artificially infected site (mortality of pine tree in control was >76.7% at the first year), PWN preventing efficacy of trunk injection of abamectin 1.8% EC and emamectin benzoate 2.15% EC at the rate of $10\;m{\ell}$ per 10 cm in DBH was 91.5% and 82.9%, respectively, at the applied year and 89.5% and 82.6% respectively at the following year in PWN naturally occurred site. Control efficacy by trunk injection of abamectin 1.8% EC and emamectin benzoate 2.15% EC was more higher in 10 fold dilution with 10 fold high amount of aquatic solution than no dilution with 10 fold less amount of aquatic solution. The preventive effect of trunk injection of abamectin 1.8% EC and emamectin benzoate 2.15% EC at the rate of $5\;m{\ell}$ per 10 cm in DBH was showed 100% at the applied year in PWN inoculated tree.

• Journal of Korean Society of Forest Science
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• v.112 no.2
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• pp.258-266
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• 2023

Pine wilt disease is caused by the pine wood nematode (Bursaphelenchus xylophilus) and has killed many pine trees in Europe and Northeast Asia, including South Korea. Resistance to pine wilt disease varies among species. Previous studies were mostly conducted in nature or greenhouses and only a few in test fields. In this study, seedlings of six pine species (Pinus thunbergii, P. koraiensis, P. densiflora, P. parviflora, P. rigida × P. taeda, and P. strobus) were artificially inoculated by pine wood nematodes in the test field. The Wilt Index was measured every 2 weeks after inoculation in addition to the mortality rate, detection rate, and pine wood nematode concentration measurement after 24 weeks. The pine wilt disease mortality rates were P. thunbergii (80%), P. koraiensis (77.8%), P. densiflora (62.5%), and P. parviflora (22.0%), and both P. rigida × P. taeda and P. strobus survived. The pine nematode detection rates were the same among the species except for P. rigida × P. taeda pine (22.2%). High Wilt-Index values were obtained for P. thunbergii, P. koraiensis, and P. densiflora, which had mortality rates higher than the other species. Furthermore, there were no significant differences in the Wilt Indexes between P. parviflora, P. rigida × P. taeda, P. strobus, and the control group. Statistically, P. thunbergii and P. koraiensis showed high susceptibility to pine wilt disease, P. densiflora and P. parviflora showed moderate susceptibility, and P. rigida × P. taeda and P. strobus showed apparent resistance. These results provide basic data for pine wood nematode resistance breeding or as evidence of the need for afforestation of P. rigida × P. taeda and P. strobus.