Virus-like particles (VLPs) are similar to pathogenic viruses, but because they have no nucleic acid, they have excellent safety and immunogenicity and are used as a good vaccine material. However, in the selection of various structural proteins of pathogenic viruses to form VLPs, all expression systems consume a lot of time in common. Among them, the baculovirus expression system causes additional time consumption to construct the recombinant baculovirus. Therefore, there is a need for a system that can rapidly determine the structural proteins required for effective VLP production. This study aims at solving this problem by constructing a BmNPV inducible expression platform through the construction of vectors induced by BmNPV. The platform was evaluated for overexpression using EGFP. We also confirmed the formation of virus-like particles through overexpression of canine parvovirus structural proteins.

Thrips from the genus Frankliniella (Thysanoptera, Thripidae) are a serious insect pest of various crops, including vegetables, fruits and ornamental plants. Thrips cause significant economic damage plants directly by feeding, and indirectly by acting as vectors for the tospoviruses such as Tomato spotted wilt virus (TSWV) and Chrysanthemum stem necrosis virus (CSNV) in chrysanthemum. To investigate the associations of thrips with tospoviruses and their ability to transmit, we have developed a protocol for identifying tospovirus and thrips species simultaneously in an individual thrips sample was successfully conducted. Total RNA was extracted from thrips according to manufacturer’s insturctions of RNeasy mini kit (Quiagen co.), and then TSWV was identified by reverse transcription-polymerase chain reaction (RT-PCR) using TSWV specific primers for the N genes. The residual genomic DNA in thrips RNA extract was used as a template to identify thrips species by PCR using universal primers for ITS2 region and subsequently digesting the PCR product by an restriction enzyme (RsaI) In addition, a classification into the species of thrips was confirmed using the nucleotide sequence of PCR products. The developed protocol was applied to investigate the occurrence of viruliferous thrips species in thrips populations collected from chrysanthemum fields. In this study, most of thrips were identified to Frankliniella spp. and thirps that acquire TSWV was 14 % of 65 thrips.

Heamaphysalis longicornis is a major vector for Severe Fever with Thrombocytopenia Syndrome (SFTS) virus and the density of the vector has been increasing because of the climate change. The incidence of fatalities due to SFTS is increasing every year. In this study, to evaluate the SFTS transmission by ticks, the density of ticks mediating SFTS was monitored. Tick was collected every month from four different sites (Grass land, Mountain path, Grave, Copse) in Andong with the traps containing dry ice as CO2 attractants. Among 2,572 ticks of 3 species; H. longicornis, H. flava, and Ixodes nipponensis were most abundantly collected from April to August. H. longicornis is the richest species (92.8%), whereas Ixodes nipponensis was the least species (0.8%). The 54.5% of the sample were nymph stage and female/male ratio was 64.3%. According to the pathogene analysis, SFTS virus was detected from H. longicornis adult and larvae stages collected in July and August. In July samples, SFTS virus was detected only from grassland site, but the virus was observed in the sample from all four sites in August. For effective prevention of SFTS, the tick density should be continuously monitored based on the onset time of SFTS with the consideration of habitate, habit & life history of ticks.

South Korea has over 0.38 million of managed honey bee (Apis cerana) colonies before 2009 years ago, which produce the highest quantity of honey in the Korea; however, almost colony (90%) were collapsed by Korean Sacbrood Virus (KSBV) in South Korea. Korean Sacbrood Virus (KSBV) is the pathogen of A. cerana Sacbrood disease, which poses a serious threat to honeybee A. cerana, and tends to cause bee colony and even the whole apiary collapse. Colony collapse of A. cerana was first reported on the Pyeong-Chang of the South Korea in 2009. Several scientists and governments has been tried research for cure the sacbrood disease in A. cerana colony by medicines and management techniques. Unfortunately, The sacbrood disease dosen`t improve. So, we were developed a better breed of A. cerana for resistance of sacbrood virus by selection and then artificial insemination. A. cerana breeding technique was first successful applied with A. cerana in Korean. Queens was grafted from sacbrood resistance line and then it was growing in sacbrood disease colony that was survived 100%. Altogether selected 18 queens were artificially inseminated and 2,000 drones of A. cerana in Korea was used to evaluate amount of semen collection. We are select two scabrood resistance A. cerana line (R and H). R line be used for rearing the Queen. Drone was reared in H line colony. The RH hybrid were not infected sacbrood virus even spread sacbrood virus (2×106). RH colonies have very excellent hygienic behavior, brood, and sacbrood disease resistance activity.

The genus Diadegma is a well-known parasitoid group and some are known to have symbiotic virus so called polydnavirus (PDV, more specifically Ichnovirus, IV). The presence of the IV in the Diadegma species a has already been identified more than a decade ago. Previously we reported a DfIV, 62 genomic segments, 247kb from D. fenestrale which parasitized wide range of lepidopteran species (Generalist). However, DsIV from D. semiclausum which parasitized in the P. xylostella (Specialist) has 48 genomic segments, 208kb. Finally, 123 ORFs were re-annotated (repeat element protein, 41; cysteine motif protein, 11; viral innexin, 6; viral ankyrin, 8; polar residue rich protein, 7; N gene, 3; Neuromodulin protein, 2 and not assigned gene, 45). DsIV also have most of lepidopteran immunosuppression gene families and 103 ORFs annotated (repeat element protein, 36; cysteine motif protein, 8; viral innexin, 7; viral ankyrin, 6; polar residue rich protein, 7; N gene, 3; and not assigned gene, 36). Certainly, although viral species specific segments exists, two IVs showed high similarity in most of segments. However DfIV have some more number of that genes such as cysteine motif protein and viral ankyrin. Two Diadegma species also showed difference in mitochondrial genome structure. Therefore we concluded that two species has their own evolutionary linage depending on the lepidopteran hosts with its own symbiotic virus.

Various symptoms of virus diseases are observed on the cultivated and non-cultivated crops in Myanmar. The virus diseases cause a major limiting factor in the tomato and chili production. During 2018, survey on the virus disease and its vectors in tomato and chili fields were conducted in central regions of Myanmar including Naypyidaw, Mandalay, Magway and Sagaing Divisions. The symptoms observed on tomato and chili were chlorosis, puckering of leaf, upward curling of leaves, reduction in leaf size, vein and stunting. Tomato leaf curl (TLC) disease incidence and severity ranged from 10 to 100% and 37 to 86% respectively. Chili leaf curl (CLC) disease incidence and severity was 30 to 100% and 13 to 100% respectively. The highest incidence and severity of TLC and CLC recorded in Lewe townships, Naypyidaw. Insect vectors observed in those areas were whitefly (Bamisia tabaci), jassid (Emposca sp) and mealy bug, and the white fly was common. Whiteflies are very important vectors for virus in tropical and subtropical regions, but not confirmed to this area. Jassids are also vectors for virus. Both nymphs and adults suck the plant sap and introduce toxin to host plants. Therefore, an infestation may be very damaging and show stunted growth and bushy appearance.

Nongwoo Bio has established a company capable of producing vegetable breeding and production in Indonesia. Since 2000, we have conducted a survey on viruses in vegetable crops such as pepper and tomatoes in Indonesia about 7 times. Since there is a possibility that the virus in Southeast Asia may occur in Korea, TSWV resistant substance is secured in pepper to cultivate resistant varieties and cultivate tomato resistant varieties. As a result, we have introduced TSWV resistant varieties of pepper for the first time in Korea, and TYLCV varieties of tomatoes have also been introduced. Through this project, virus analysis on Southeast Asian countries will be conducted and seed companies will be able to fully utilize basic data for cultivating resistant varieties. We will continue to investigate the viruses of vegetable crops in Indonesia and contribute to cultivating resistant varieties.

Immortalization is an essential process of the transformation of cells to a neoplastic growth. High risk human papillomavirus (hrHPV) infection has been the major cause of head and neck squamous cell carcinoma (HNSCC). The aim of this study was to search for a novel pathway causing immortalization in HPV16 E6/E7 transfected immortalized oral keratinocytes (IHOK). hrHPV integration sites were identified through DNA sequencing. HPV16 E6/E7 genes were integrated into 1q32.2, 12q21.2, 15q15.2, and 19q13.43 in IHOKs. Array-CGH was conducted to examine the deranged sites of the genes of IHOK. Of the 587 amplification genes, 70 genes were resided on chromosome 20. We selected PLAGL2 and MAPRE1 as the most amplified genes. PLAGL2 and MAPRE1 mRNA showed higher expression in IHOK than in normal keratinocytes. Knockdown of MAPRE1 significantly reduced telomerase activity. The analysis using a public database substantiated our data, showing the amplification of chromosome 20 and MAPRE1. In conclusion, our results suggest that MAPRE1 could play a crucial role in activating telomerase activity in hrHPV-infected cells. This finding may provide basic data to develop a novel target therapy for hrHPV-related HNSCC.

RNA interference (RNAi) has been considered as an alternative strategy to control agricultural pest whereby double-stranded RNA triggers a potent and specific inhibition of its homologous mRNA. Since small dsRNAs are required for various RNAi applications, there is a need for cost-effective methods for producing large quantities of high-quality dsRNA. In this study, Bacillus thuringiensis (Bt) based dsRNA production platform was established under control of sporulation-dependent promoter and vp1 gene of Sacbrood virus (SBV) was introduced. The dsRNA against vp1 gene produced from the Bt suppressed the replication of SBV. In addition, the dsRNA was assembled into inulin coated-nanoparticle to increase stability of dsRNA in environment.

Bemisia tabaci is a vector of Tomato yellow leaf curl virus (TYLCV) but Trialeurodes vaporariorum is not. To determine the effect of TYLCV acquisition on vector and non-vector, we compared various physiological characteristics between two species. Our results showed that TYLCV acquisition significantly affected B. tabaci but not T. vaporariorum. The B. tabaci increased susceptibility against thermal stress but weakened chill coma recovery, shortening of longevity, low fecundity and abbreviated developmental time by virus acquisition. At the molecular level, B. tabaci increased hsp70 and hsp90 levels by TYLCV ingestion. However, T. vaporariorum did not shown any changes of those characteristics. Therefore, the physiological manipulation of TYLCV was specific to vector species but not non-vector species.

The virus-like particle (VLP) is similar to a pathogenic virus and has a high immunogenicity. However, in the selection of various structural proteins to form VLP, all expression systems commonly consume most of the time and suffer from various difficulties. Therefore, there is a need for a system that can rapidly determine the structural proteins required for effective VLP production. This study aims to construct a transient expression platform using insect cells to solve this problem. Plasmid-based expression vectors and baculovirus-inducible expression vectors were constructed. The vectors were evaluated for overexpression using EGFP. We also confirmed the formation of virus like particles through overexpression of virus structural proteins.

Highly pathogenic avian influenza virus (HPAIV) damages vital organs and tissues, frequently leading to death in birds, and causes serious economic losses in the poultry industry. In addition, HPAIV can infect humans and other mammals, often with fatal outcomes. In this study, the virucidal efficacy of Clean-Zone®, which contains citric acid, malic acid and phosphoric acid, against avian influenza virus (AIV, H9N2) was investigated. Virucidal efficacy was determined by examining the viability of AIV after contact with the disinfectant in the allantoic membrane of chicken embryos. The disinfectant and AIV were reacted under hard water (HW) and organic matter suspension (OM) condition. AIV was inactivated with 200- and 50-fold dilutions of the disinfectant under HW and OM conditions, respectively. As the disinfectant, Clean-Zone®, has a virucidal efficacy against AIV, it can be used to prevent the spread of animal viral diseases.