• Journal of Microbiology
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• 제43권6호
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• pp.537-545
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• 2005

Even though neutralizing antibodies against the Hantaan virus (HTNV) has been proven to be critical against viral infections, the cellular immune responses to HTNV are also assumed to be important for viral clearance. In this report, we have examined the cellular and humoral immune responses against the HTNV nucleocapsid protein (NP) elicited by virus infection or DNA vaccination. To examine the cellular immune response against HTNV NP, we used $H-2K^b$ restricted T-cell epitopes of NP. The NP-specific $CD8^+$ T cell response was analyzed using a $^{51}Cr-release$ assay, intracellular cytokine staining assay, enzyme-linked immunospot assay and tetramer binding assay in C57BL/6 mice infected with HTNV. Using these methods, we found that HTNV infection elicited a strong NP-specific $CD8^+$ T cell response at eight days after infection. We also found that several different methods to check the NP-specific $CD8^+$ T cell response showed a very high correlation among analysis. In the case of DNA vaccination by plasmid encoding nucleocapsid gene, the NP-specific antibody response was elicited $2\~4$ weeks after immunization and maximized at $6\~8$ weeks. NP-specific $CD8^+$ T cell response reached its peak 3 weeks after immunization. In a challenge test with the recombinant vaccinia virus expressing NP (rVV-HTNV-N), the rVV-HTNV-N titers in DNA vaccinated mice were decreased about 100-fold compared to the negative control mice.

• Journal of Microbiology
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• 제45권1호
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• pp.41-47
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• 2007

The Hantaan virus (HTNV) is an enveloped virus that is capable of inducing low pH-dependent cell fusion. We molecularly cloned the viral glycoprotein (GP) and nucleocapsid (NP) cDNA of HTNV and expressed them in Vero E6 cells under the control of a CMV promoter. The viral gene expression was assessed using an indirect immunofluorescence assay and immunoprecipitation. The transfected Vero E6 cells expressing GPs, but not those expressing NP, fused and formed a syncytium following exposure to a low pH. Monoclonal antibodies (MAbs) against envelope GPs inhibited cell fusion, whereas MAbs against NP did not. We also investigated the N-linked glycosylation of HTNV GPs and its role in cell fusion. The envelope GPs of HTNV are modified by N-linked glycosylation at five sites: four sites on G1 (N134, N235, N347, and N399) and one site on G2 (N928). Site-directed mutagenesis was used to construct eight GP gene mutants, including five single N-glycosylation site mutants and three double-site mutants, which were then expressed in Vero E6 cells. The oligosaccharide chain on residue N928 of G2 was found to be crucial for cell fusion after exposure to a low pH. These results suggest that G2 is likely to be the fusion protein of HTNV.

• 대한바이러스학회지
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• 제28권4호
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• pp.327-335
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• 1998

Hantaan (HTN) and Seoul (SEO) viruses, murid rodent-borne hantaviruses, are known to causes hemorrhagic fever with renal syndrome (HFRS) in Korea. To determine the genomic diversity and molecular phylogeny of HTN and SEO viruses found in Korea, we amplified for part of M and S genomic segments of hantaviruses from sera of HFRS patients and lung tissues of hantavirus seropositive striped-field mice. Both M and S segment of 16 HTN and 2 SEO viruses were amplified by nested reverse transcription-polymerase chain reaction. Based on 324 nucleotides in the M genomic segment, the HTN and SEO strains showed $93.8{\sim}100%$ and $99.1{\sim}99.4%$ homologies, respectively. Similarly, based on 230 nucleotides in the S genomic segment, HTN and SEO strains showed $90.9{\sim}100%$ and 100% homologies, respectively. Phylogenetic analysis of M and S segments indicated that HTN strains could be divided into at least two main groups in M and S trees and the sequence differences detected among the Sand M genomic segments of HTN viruses are consistent with reassortment having taken place between HTN virus strains.

• 대한미생물학회지
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• 제22권1호
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• pp.1-8
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• 1987

Hantaan virus(HV) 76-118 strain was inoculated into suckling ICR mice by intra-nasal route with an inoculum of $10LD_{50}$. Mortality was 65% at the 3rd week after inoculation, but declined to 35% at the 4th week. Infectivity was determined by the measuring immuno-fluorescent antibody in sera. The peak of infectivity was 80% at the 4'th week after inoculation. Viremia was reached peak level of $1.7{\times}10^4\;PFU/ml$ by day 10. Immunofluorescent antibody and neutralizing antibody appeared by 2 weeks and 15-17 days respectively, but achieved similar titer by 35 days. By using a monoclonal antibody to HV 76-118, viral antigens were initially detected in inguinal and axillary lymph node by 2 days. Viral antigens in bone marrow and lung were delayed much more than in those of lymph node. These were similar with those of intra-peritoneal and intra-muscular route. Immune complex against IgG, IgM and C3 appeared by 16 days, 14 days, and 18 days respectively. The pattern of immunofluorescence in the basement membrane of glomeruli was diffuse membranous. Spotted pattern was also observed in the tissue stained with anti-mouse C3 antibody. By 20 days, control tissue was also shown immune complex in the glomeruli.

• Toxicological Research
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• 제13권3호
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• pp.281-291
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• 1997

The subacute toxicity of the combined vaccine (KGCC-95VI) for the prophylaxis against Japanese encephalitis and Hantaan virus infection, recently developed by Korea Green Cross Corporation, was investigated. KGCC-95VI was subcutaneously administered into the both sexes of New Zealand White rabbits at the dosage of 0, 10. 50 and 250 ml/kg body weight (20, 100 and 500 times the expected clincal dose) once a day for 30 days. There were no deaths and clinical findings during the experiment period. In both sexes. there were no statistically significant differences between the treated and control groups in urinalysis tests, hematological tests, blood chemistry tests and pathological examinations. The KGCC-95VI is considered not to have the subacute toxicity in the rabbits.

• 대한바이러스학회지
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• 제29권1호
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• pp.1-9
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• 1999

Hantaan virus is widely distributed among rodent populations in Korea. Two antigenically distinct hantaviruses were isolated from Apodemus agrarius in 1976 and Rattus norvegicus in 1980 in Korea. This study was designed to investigate the serological evidence of hantavirus infection among indegenous wild rodents, which were captured in 11 mountains located in Kyunggi, Kangwon, Chungnam, Chunbug and Kyungnam province of South Korea. A total 252 wild rodents of 3 species were trapped from Myungsung Mt., Chumbong Mt., Kali Mt., Hansuk Mt., Chachil peak, Kyebang Mt., Odae Mt., Kyerong Mt., Kaya Mt., Togju Mtand Chiri Mtin 1997. Serologic test for hantavirus infection was performed using hantavirus antigens by indirect immunofluorescent antibody techniqueAmong 122 Apodemus agrarius, 88 Apodemus peninsulae and 42 Eothenomys regulus; 18 A. agrarius (14.8%), 12 A. peninsulae (13.6%) and 4 E. regulus (9.5%) were immunofluorescent antibody (IFA) positive against hantaan virus. IFA titers 3 Eothenomys regulus sera were higher against puumalavirus than hantaan virus. These data imply that above three species of rodent might be natural reservoirs of hantaviruses in Korea.

• 한국군사과학기술학회지
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• 제24권3호
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• pp.348-355
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• 2021

Hantaan virus(HTNV) causes hemorrhagic fever with renal syndrome(HFRS) with a case fatality rate ranging from <1 to 15 % in human. Hantavax^Ⓡ is a vaccine against the Hantavirus, which has been conditionally approved by the Ministry of Food and Drug Safety(MFDS). However, only 50 % of volunteers had neutralizing antibodies 1 year following the boost. Effective antiviral treatments against HTNV infection are limited. Hantaviruses generally cause asymptomatic infection in adult mice. On the other hand, infection of suckling and newborn mice with hantaviruses causes lethal neurological diesease or persistant infection, which is different from the disease in humans. The development of vaccines and antiviral strategies for HTNV has been partly hampered by the lack of an efficient lethal mouse model to evaluate the efficacy of the candidate vaccines or antivirals. In this report, we established a lethal mouse model for HTNV, which may facilitate in vivo studies on the evaluation of candidate drugs against HTNV. The median lethal dose value of HTNV was calculated by probit analysis of deaths occurring within two weeks. Five groups of ten ICR mice were injected intracranially with serial 2-fold dilutions (from 50 to 3.125 PFU/head) of HTNV. Mice injected with HTNV began to die at 8 days post-infection. The lethal dose required to kill 50 % of the mice (LD₅₀) was calculated to be 2.365 PFU/head.

• Journal of Yeungnam Medical Science
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• 제15권1호
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• pp.55-66
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• 1998

사람과 동일한 증상을 나타내는 동물 모델이 없는 신증후출혈열의 병리 기전을 이해하기 위하여 in vitro culture system을 이용하였다. 신증후출혈열의 주된 병변이 혈관내피세포외에 섬유아세포가 많이 존재하는 신세뇨관 간질 및 폐간질임을 기초로 하여 계태아와 Mongolian gerbil의 섬유아세포에서 fibronectin에 대한 수용체인 ${\alpha}_5{\beta}_1$, integrin에 대한 항체를 처리하여 한탄바이러스 감염에 대한 영향을 관찰하였다. 초대 배양하여 사용한 계태아 섬유아세포 및 MGF 모두 한탄바이러스가 잘 감염되었으나 계태아 섬유아세포에 비하여 MGF에서 바이러스의 증식이 더 우수하였다. 계태아 섬유아세포와 MGF에서 ${\alpha}_5{\beta}_1$, integrin에 대한 항체 처리 5일 후 바이러스의 N 단백의 역가는 항체를 처리하지 않은 대조군에 비하여 각각 32.6%, 72.6%로 virion의 역가는 26.8%, 28.7%로 감소하여 두 세포에서 모두 감염력을 가진 바이러스의 양은 거의 동일하게 감소 되었다. 또한 ${\alpha}_5$ subunit와 ${\beta}_1$, subunit에 대한 항체를 MGF 세포에 처리한 후 바이러스의 N단백의 역가는 65.2%, 59.7%로 ${\alpha}_5{\beta}_1$ integrin에 대한 항체 처리하였을 때(72.6%)와 거의 유사하게 감소하였고 감염력을 가진 바이러스의 양도 26.5%, 29.4%로 감소하여 ${\alpha}_5{\beta}_1$ integrin에 대한 항체 처리시(28.7%)와 거의 유사하게 감소하였다. 이러한 결과는 ${\alpha}_5{\beta}_1$ integrin이 한탄바이러스에 대한 수용체로써 작용할 가능성을 시사하였으며 또다른 기전으로는 ${\alpha}_5{\beta}_1$ integrin의 차단에 의한 이차적인 영향으로 한탄바이러스의 증식에 커다란 영향을 미치는 것을 생각할 수 있었다.

• 대한의생명과학회지
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• 제6권2호
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• pp.141-149
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• 2000

우리 나라에서 발생하고 있는 급성 출혈성 질환인 신증후출혈열의 원인 바이러스는 Family Bunyaviridae의 Genus Hantavirus에 속하는 한탄과 서울바이러스에 의하여 발생되고 있다. 본 연구에서는 신증후출혈열로 의뢰된 환자에서 한탄바이러스에 대한 항체가를 간접면역형광항체법(indirect immunofluorescent antibody technique, IFAT), 면역효소측정법 (enzyme-linked immunosorbent assay, ELISA) (IgG, IgM), 고비중입자응집반응 (high density composite particle agglutination, HDPA) 및 플라크감소중화시험 (plaque reduction neutralization test, PRNT) 등으로 비교 측정하였고, 신증후출혈열 환자로 확진된 15명의 한탄바이러스 혈청형을 PRNT와 혈청형 특이 역전사 효소 중합효소연쇄반응(nested reverse transcriptase polymerase chain reaction, nested RT-PCR)으로 확인하였다. 신증후출혈열로 의뢰된 환자에서의 한탄바이러스에 대한 IFAT, ELISA (IgG, IgM), HDPA그리고 PRNT비교에서 형광항체, ELISA IgG,응집항체 및 중화항체는 8명 모두 높게 나타났으며, ELISA IgM은 5명에서는 현저히 높은 항체를 보유하고 있었다. 신증후출혈 열 환자 15명에서는 높은 형광항체와 중화항체 역가를 나타내었고, 15명 중 12명은 한탄바이러스, 2명은 서울바이러스에 대한 높은 중화항체를 갖고 있었으며, 1명은 두 바이러스에 대하여 동일한 항체 역가를 나타내었으며, 혈청형 특이 primer를 사용한 nested RT-PCR에서는 15명 중 3명과 1명만이 한탄바이러스와 서울바이러스 primer에 대해 RNA가 검출되었다.

• 대한바이러스학회지
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• 제29권4호
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• pp.201-209
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• 1999

Results of the studies on the morphologic and molecular biologic characteristics of Hantaan virus (HTNV), one of the etiologic agents of Hemorrhagic fever with renal syndrome (HFRS), revealed that HTNV was a member of Family Bunyaviridae and its RNA divided into three segments. And the nucleotide sequences of these segments also were known and the differences in nucleotide sequences of HTNV from other members of genus Hantavirus were clearly evaluated. But the morphorgenesis, pathogenesis of HFRS and the replication time had not been clearly determined. In this study, to estimate the replication time of HTNV in Vero E-6 cells, Vero cells were infected with HTNV 76/118 strain, and cells were harvested from two hours post-infection up to 24 hours at two hours-intervals. Harvested cells were treated with ordinary techniques for electron microscopy and immune-electron microscopy. And then thin sections were observed under transmission electron microscope. HTNV particles were not found in the cytoplasm and in the extracellular space between $2{\sim}8$ hours after inoculation of virus, but virus particles were observed in extracellular space near the cell membrane of Vero-E6 cells 10 hours after infection. In immune electron microscopy, mature HTNV particles in extracellular spaces and immature virus labelled with gold particles in the cytoplasm of Vero E-6 cell 10 hours after infection of HTNV could be seen. This results suggest that the replication time of HTNV might be about 10 hours.