• 한국어병학회지
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• 제35권1호
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• pp.129-133
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• 2022

The full genome sequence of a Korean white spot syndrome virus (WSSV, isolate: WSSV-GoC18) is presented here. We obtained a total of 12,320,554 reads with 291,172 bases, 170 gene, and 170 coding DNA sequence, which were assembled in 1 contig. Phylogenetic analysis revealed that the WSSV-GoC18 was closely related to Chinese isolate (WSSV-PC) and distinctly different with previously reported a Korean isolate (WSSV K-LV1). The complete genome sequence of WSSV isolates will be of great help in molecular epidemiological studies, contributing to molecular diagnosis and disease prevention in shrimp aquaculture.

• 생명과학회지
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• 제20권10호
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• pp.1427-1432
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• 2010

본 연구는 WSSV의 재조합단백질 rVP466에 대하여 생산된 항혈청을 사용하여 WSSV에 대한 neutralization (중화) 효과를 확인하고자 수행하였다. 먼저 재조합단백질 rVP466의 생산을 위해 WSSV의 구성단백질 VP466을 암호화하는 유전자인 VP466을 포함하는 재조합 플라스미드 pCold-VP466을 제작한 다음 이것을 발현용 숙주인 E. coli RIPL에서 발현하였다. 발현된 rVP466에 대한 항혈청은 토끼를 사용하여 생산하였으며, 항원 rVP466에 대한 특이면역반응은 Western blot을 통해 확인하였다. WSSV에 대한 항혈청의 중화효과를 확인하기 위해 항혈청과 반응시킨 바이러스액($1{\times}10^4$ 배로 희석된 WSSV)을 이용하여 실험용 새우(Penaeus chinensis)에게 주사 감염을 통해 공격실험(challenge test)을 수행하였다. 실험 결과, WSSV로 공격실험한 감염대조구(positive control)의 새우들은 감염 후 17일째에 100% 누적폐사율을 보였으며, preimmune serum과 WSSV의 혼합액을 challenge한 preimmune control의 새우들은 감염 후 25일째에 83%의 누적폐사율을 보였다. WSSV와 rVP466 항혈청을 1:0.01, 1:0.1, 1:1로 혼합한 액으로 challenge한 새우들은 감염 후 25일째에 각각 73%, 53%, 46%의 누적폐사율을 보였다. 이상의 결과를 통해 WSSV가 rVP466 항혈청에 의해 농도의존적으로 neutralization됨을 확인하였으며, 이는 WSSV 감염과정에 VP466이 관여함을 나타내는 것이다.

• 생명과학회지
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• 제20권8호
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• pp.1181-1185
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• 2010

본 연구는 WSSV의 주요 구조단백질인 VP19와 VP28을 모두 포함하는 VP19+28 fusion protein을 제조하여, Litopenaeus vannamei에서 WSSV에 대한 백신으로서의 효능을 평가하고자 수행하였다. VP19와 VP28 유전자를 fusion하여 제작한 VP19+28 유전자를 pET-28a(+) vector에 삽입하고 단일단백질로서 제작된 VP19+28 유전자를 E. coli BL21 (DE3)에서 발현시켰다. 백신실험을 위해 새우에게 2주 동안 실험용 사료를 급이하였으며, 그 후 바이러스액($1{\times}10^2$배로 희석한 WSSV)을 이용하여 새우에게 주사 감염에 의해 in vivo 공격실험(challenge test)을 수행하였다. 실험결과, vaccination을 하지 않은 새우들은 감염 후 11일째에 100%의 누적폐사율을 보였으며, host control로써 E. coli BL21을 사용하여 vaccination한 새우들은 감염 후 17일째에 100%의 누적폐사율을 보였다. rVP19, rVP28, rVP19+28을 이용하여 vaccination한 새우들의 경우 감염 후 21일째에 각각 66.7%, 41.7%, 41.7%의 누적폐사율을 보였다. 이상의 결과를 통해 rVP28과 rVP19+28이 WSSV에 대해 높은 백신효능을 가짐을 확인하였다. 또한 감염 후 21일째에 fusion protein rVP19+28과 rVP28의 누적폐사율은 동일하였지만 공격실험기간 동안 폐사율이 rVP19+28을 투여 한 실험군이 낮게 나타나는 것을 보아 WSSV에 대한 새우의 방어효능은 rVP19+28이 더 높음을 나타내는 것이다.

• Journal of Microbiology and Biotechnology
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• 제17권3호
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• pp.511-515
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• 2007

White-spot syndrome virus (WSSV) is a devastating, infectious virus affecting shrimp. Although sensitive techniques involving PCR have been developed to assist farmers in screening shrimp (brood stock) for WSSV prior to stocking ponds, such practices have not yet been applied in Korea. Despite the rationality of implementing screening, there has been some doubt as to whether the stocking of WSSV-PCR-negative fly epidemiologically decreases white-spot disease outbreaks. Here, we report a retrospective analysis of data from shrimp farms in the western coast of Korea where WSSV-PCR-negative brood stocks were used to stock rearing ponds. A total of 366 shrimp from Heuksan Island were sampled for WSSV with PCR. Of the tested shrimp, 7.2% (28 brood stocks) were identified as WSSV positive; only WSSV-PCR-negative shrimp were used for brood stocks. Total unit production (final shrimp production/ the area of the ponds) was higher, at 1.96, in ponds where WSSV-PCR-negative shrimp were used, as compared with 1.02 in other ponds in Korea in 2004. This retrospective analysis of WSSV in Korea may be useful to the shrimp aquaculture industry, suggesting a testable hypothesis that may contribute to the eventual control of WSSV outbreaks.

• 한국어병학회지
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• 제32권1호
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• pp.45-48
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• 2019

We developed and subsequently characterized mouse monoclonal antibodies (MAbs) against recombinant VP28 structural protein (rVP28) of white spot syndrome virus (WSSV). We established six hybridoma clones secreting MAbs against rVP28: 15A11, 20G6, 31H2, 34H6, 38D1 and 43A1. All six MAbs recognized the 25 kDa of protein in gill homogenates of WSSV-infected shrimp by western blot analysis, while no reactivity was observed in gill homogenates of normal shrimp. Moreover, high enzyme-linked immunosorbent assay (ELISA) optical density (OD) values (0.8-2.68) were observed in the hemolymphs from WSSV-infected shrimp, while low OD values (less than 0.24) were recorded in the hemolymphs from normal shrimp, by using these six MAbs produced in this study. These results suggest that these six MAbs are useful for the detection of WSSV.

• 한국수의병리학회:학술대회논문집
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• 한국수의병리학회 2003년도 추계학술대회초록집
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• pp.41-41
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• 2003

White spot syndrome virus (WSSV) is the causative agent of a disease that has led to severe mortalities of cultured shrimps in Korea and many other countries. Since 1993, massive mortalities due to the viral infection have also occurred in the penaeid shrimps cultured in Korea. WSSV is a large, circular, double stranded (ds) DNA virus and an enveloped, ellipsoid virus with a rod-shaped nucleocapsid with flat ends. In order to identify the characteristics of this Korean isolate of WSSV, the genes for four virion proteins, VP15, VP19, VP28 and VP35 were cloned and their sequences were compared with the available pool of WSSV gene sequences in the GenBank/EMBL databases. From these comparisons, we confirm the occurrence of WSSV in Korea and deduce that, VP15, VP28 and VP35 genes are identically conserved among the Korean isolate and geographically different foreign isolates, but VP19 amino acid sequences of the Korean WSSV isolates changed valine of the foreign isolates into aspartate. (omitted)

• Journal of Microbiology and Biotechnology
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• 제21권2호
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• pp.170-175
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• 2011

This study was carried out for the molecular level identification of recombinant protein vaccine efficacy, by oral feeding against white spot syndrome virus infection, with the comparison of viral mRNA transcriptional levels in shrimp cells. For the determination of WSSV dilution ratio for the vaccination experiment by oral feeding, in vivo virus titration was carried out using different virus dilutions of virus stock ($1{\times}10^2$, $2{\times}10^2$, and $1{\times}10^3$). Among the dilution ratios, $2{\times}10^2$ diluted WSSV stock was chosen as the optimal condition because this dilution showed 90% mortality at 10 days after virus injection. Recombinant viral proteins, rVP19 and rVP28, produced as protein vaccines were delivered in shrimps by oral feeding. The cumulative mortalities of the shrimps vaccinated with rVP19 and rVP28 at 21 days after the challenge with WSSV were 66.7% and 41.7%, respectively. This indicates that rVP28 showed a better protective effect against WSSV in shrimp than rVP19. Through the comparison of mRNA transcriptional levels of viral genes from collected shrimp organ samples, it was confirmed that viral gene transcriptions of vaccinated shrimps were delayed for 4~10 days compared with those of unvaccinated shrimps. Protection from WSSV infection in shrimp by the vaccination with recombinant viral proteins could be accomplished by the prevention of entry of WSSV due to the shrimp immune system activated by recombinant protein vaccines.

• Bulletin of the Korean Chemical Society
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• 제33권1호
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• pp.69-75
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• 2012

Five-channel electrochemical chips were fabricated based on the Micro-electromechanical System (MEMS) technology and were used as platforms to develop DNA arrays. Different kinds of thiolated DNA strands, whose sequences were related to white spot syndrome virus (WSSV) gene, were separately immobilized onto different working electrodes to fabricate a combinatorial biosensor system. As a result, different kinds of target DNA could be analyzed on one chip via a simultaneous recognition process using potassium ferricyanide as an indicator. To perform quantitative target DNA detection, a limit of 70 nM (S/N=3) was found in the presence of 600 nM coexisting noncomplementary ssDNA. The real samples of loop-mediated isothermal amplification (LAMP) products were detected by the proposed method with satisfactory result, suggesting that the multichannel chips had the potential for a high effective microdevice to recognize specific gene sequence for pointof-care applications.

• 한국양식학회지
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• 제15권1호
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• pp.7-13
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• 2002

Since 1993, the White Spot Syndrome Virus (WSSV) disease occurred in China among cultured shrimps resulting in mass mortality. Epizootiological surveys undertaken during the outbreak period of 1993-1994 indicated that all stages of Penaeus chinensis, P. japonicus and P. monodon were infected. Consequent to the transport of contaminated shrimp seedlings and seawater, the disease spread all over the farms of China. The disease was more rapidly transmitted at temperatures above $25^{\circ}C$. Challenge experiments showed the causative agent was highly virulent. White spots appeared on the carapace of both span-taneous and experimentally infected shrimps. Moribund shrimps contained turbid hemolymph, hypertrophied Iymphoid organ and a necrotic mid-gut gland. Electron microscopy showed the presence of viral particles in the gills, stomach, lymphoid organ, and epidermal tissue of the infected shrimp. The visions were slightly ovoid with an envelope and averaged 350 $\times$ 150 nm; nucleocapsids measured 375 $\times$ 157 nm. With discontinuous sucrose gradient of 35, 50 and 60% (w/v), the virus was separated from hemolymph of the infected shrimp. The estimated molecular weight of genomic DNA was 237 Kb with EcoR I, 247 Kb with Hind III and 241kb with Pst I. A total of 9 hybridoma colones secreting monoclonal antibodies (MAbs) were produced from mouse myeloma and spleen cells immunized with WSSV. The immunofluorescence assay of gill tissue showed that the MAbs reacted with diseased but not with healthy shrimp. The MAbs belonged to IgGl, IgG2b subclass and IgM class, all with kappa light Immune-electron-microscopy with colloidal gold marker showed the presence of 5 MAbs epitopes on the envelope and one on the capsid of the virus. Baculoviral mid-gut gland necrosis showed the specificity of the MAbs produced. For diagnosis 5 different methods were selected. Using Kimura primers for PCR, or MAbs for immunoblot, ELISA or FAT method, in situ hybridization was carried out to show the gene. All these methods detected WSSV in the organ samples of the diseased shrimp but not in healthy one.