• IMMUNE NETWORK
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• 제7권2호
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• pp.66-74
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• 2007

Background: The genus Flavivirus consists of many emerging arboviruses, including Dengue virus (DV), Japanese encephalitis virus (JEV) and West Nile virus (WNV). Effective preventive vaccines remain elusive for these diseases. Mice are being increasingly used as the animal model for vaccine studies. However, the pathogenic mechanisms of these viruses are not clearly understood. Here, we investigated the interaction of DV and JEV with murine bone marrow-derived dendritic cells (bmDC). Methods: ELISA and FACS analysis were employed to investigate cytokine production and phenotypic changes of DCs obtained from bone marrow following flavivirus infection. Results: We observed that these viruses altered the cytokine profile and phenotypic markers. Although both viruses belong to the same family, JEV-infected bmDC produced anti-inflammatory cytokine (IL-10) along with pro-inflammatory cytokines, whereas DV infection induced production of large amounts of pro-inflammatory cytokines (IL-6 and TNF-${\alpha}$) and no IL-10 from murine bmDCs. Both flaviviruses also up-regulated the expression of co-stimulatory molecules such as CD40, CD80 and CD86. JEV infection led to down-regulation of MHC II expression on infected bmDCs. We also found that cytokine production induced by JEV and DV is MyD88-dependent. This dependence was complete for DV, as cytokine production was completely abolished in the absence of MyD88. With regard to JEV, the absence of MyD88 led to a partial reduction in cytokine levels. Conclusion: Here, we demonstrate that MyD88 plays an important role in the pathogenesis of flaviviruses. Our study provides insight into the pathogenesis of JEV and DV in the murine model.

• 미생물학회지
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• 제46권2호
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• pp.140-147
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• 2010

일본뇌염바이러스(Japanese encephalitis virus)는 모기 매개성 플라비바이러스에 속하며, 주로 동남아시아 지역에서 유행성 바이러스성 뇌염을 일으킨다. 일본뇌염바이러스는 외피를 가진 작은 바이러스로서, 양성가닥 RNA 게놈을 가지고 있다. 감염성을 띤 바이러스 입자는 capsid (C), membrane (M; prM 전구체로부터 생성), 및 envelope (E)과 같은 3개의 구조단백질로 이루어져 있다. 본 연구에서는 일본뇌염바이러스 생산과 정에 M 단백질의 N-말단부위에 위치한 극성 아미노산 잔기의 역할을 분석하였다. 일본뇌염바이러스의 infectious cDNA를 활용하여, M 단백질의 $E^9$와 $K^{15}K^{16}E^{17}$ 잔기를 알라닌으로 치환시킨 2개의 mutant cDNA (Mm1과 Mm2)를 각각 제작하였다. 각각의 cDNA로부터 합성된 mutant RNA를 세포에 트랜스펙션시킴으로써, 비록 세포 내에 축적된 3개의 구조단백질양은 변화가 없으나, 이들 세포로부터 생산된 바이러스의 양은 Mm2 RNA의 경우 ~1,000배 감소됨을 관찰하였다. 흥미롭게도, Mm2 RNA로부터 발현된 mutant M 단백질은 wild-type M 단백질을 인지하는 항혈청에는 반응하지 않았으나, mutant M 단백질을 항원으로 제작된 항혈청에는 반응하는 것을 알 수 있었다. 본 실험결과는 일본뇌염바이러스 M 단백질을 구성하는 3개의 극성 아미노산 잔기($K^{15}K^{16}E^{17}$)가 바이러스의 생산과정에 관여한다는 것을 암시한다. 앞으로, wild-type 또는 mutant M 단백질(Mm2)을 인식하는 2개의 항혈청은 이 단백질의 기능연구에 유용한 재료로 사용될 것으로 기대된다.

• 대한바이러스학회지
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• 제27권2호
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• pp.197-207
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• 1997

To investigate the NS4 region of JEV, NS4 cDNA of K94P05 (JEV strain isolated from Korea in 1994) was amplified by RT-PCR and analyzed by sequencing PCR product. Genomic size of NS4 was 1212bp and nucleotide sequence was compared with that of other JEV strains. Nucleotide homology between JaOAr582 and K94P05 was 91.1% and that between Beijing and K94P05 was 89.8%, respectively. But the nucleotide sequence of E region of JaOAr582 and K94P05 showed 97.0% homology and that of Beijing and K94P05 did 95.8% homology. NS4 protein was expressed as a form of fusion protein by a prokaryotic expression system. The induced fusion product showed a lower molecular weight than predicted size and remained insoluble. The NS4 protein might be cleavaged by E. coli protease. Concluding above results, high hydrophobicity of the NS4 protein supported the fact that this protein played a role as a membrane component and the poor nucleotide sequence conservativity among JEV strains suggested that this region might be important to adapt each viral growth environment.

• KSBB Journal
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• 제13권3호
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• pp.231-237
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• 1998

세계보건기구 (WHO)가 백신 생산에 권장하고 있는 표준세포 주인 Vero 세포에 약독화 일본뇌염바이러스인 SA14-14-2 ( (PDK)를 연속 계대배양을 통해 적응(adaptation)시켜, tIter가 $10^7$pfu/mL을 넘는 SA-14-14-2(Vero)을 분리하였다 바이러스 배양 최적온도는 $35^{\circ}C$이며, T -flask에서 배양된 바이러스의 최고 tIter는 감염 후 4일째에 $4\times10^7$ pfu/mL로 관찰되었다. 또한 무혈청배지에서도 바이러스 증식이 활발하여 2% 혈청이 보충된 정우와 거의 비슷한 바이라스 tIter를 보였다. 바이러스 대량 배양을 위해 roller bottle culture와 미 립 담체 플 이용한 spinner flask culture 가능성에 대하여 고찰하였다 바이러스 감염을 위한 미립담체에서의 Vera cell monolayer는 초기 세포 농도 $4\times10$ cells/mL로 접종하여 50 rpm에서 7일간 배양하여 얻을 수 있었다. 바이러스의 roller bottle 배양이 spinner flask 배양보다 바이러스 tIter변에서 2배 내지 3배 높 았고, $10^7$pfu/mL을 넘는 배양 기간도 하루 죄었다. 하지만 두 배양 방법 모두 T -flask 배양에서와 같이 무혈청 배지를 사용 하여도 바이라스 증식이 활발했고, 최고조의 tIter를 보이는 배 양기간은 감염 후 2일째로써 T -flask 배양에서 보다 2일 빨랐다. Roller bottle culture의 경우, 감염 후 3일부터 17일까지 2 일 간격으로 배양액을 무혈청 EMEM으로 100% 교체하면서 매 양을 지속한 결과 3일부터 9일까지 $10^7$pfu/mL을 념는 tIter가 유지되는 것이 확인되어 바이러스의 multi-harvest가 가능한 것 로 고찰되었다. 상기의 결과는 생산성 면에서 매우 유리한 결 과로 제품의 생산 단가플 낮추고 작업 노력을 절감하는 기대 효 과가 클 것으로 예측된다.

• 한국임상수의학회지
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• 제30권4호
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• pp.236-240
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• 2013

A total of 170 litters (575 samples) of aborted and stillbirth fetuses submitted to the Gyeongsangbuk-Do Veterinary Service Laboratory (GVSL) between January 2006 and December 2010 from pig farms in Gyeongbuk province were studied to identify porcine abortion- and stillbirth-associated viruses such as Porcine parvovirus (PPV), Encephalomyocarditis Virus (EMCV), Japanese Encephalitis Virus (JEV), Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), and Aujeszky's Disease Virus (ADV). Virus was not detected by PCR in 36 litters, but viral antibody was detected by HI and ELISA in 93 litters. The majority of etiological viruses were PPV (67 litters, 39.4%), EMCV (50 litters, 29.4%), PRRSV (15 litters, 8.8%), and JEV (11 litters, 6.5%); ADV was not detected by either PCR or ELISA. Single infection occurred in 52 litters (30.6%), co-infection occurred in 41 litters (24.1%), and unknown cases with no detection of any of the five viruses occurred in 77 litters (45.3%).

• 대한바이러스학회지
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• 제27권2호
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• pp.209-216
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• 1997

Three dimensional structures of envelope protein from Korean isolates and Nakayama-NIH strain of Japanese encephalitis virus (JEV) were deduced by a computer program (HyperChem 4.0 Chemplus 1.0) based on the data of the three dimentional structure of Tick-borne encephalitis virus. In the three dimensional structure of envelope protein, neutralizing epitope and T-helper cell recognition site of C-terminal region of Korean isolates were structually similar to those of Nakayama-NIH but the N-terminal region was not. Korean JE isolates were compared with Nakayama-NIH strain by using cross-neutralization antibody test. Neutralizing activities of Korean isolates derived from guinea pigs were higher than those of Nakayama-NIH strain against Korean isolates, although the polyclonal antibody titers of Nakayama-NlH showed 1:160 to 1:640 against Korean isolates. According to the results from three dimentional structures and cross-neutralization analyses, the antigenic difference between Korean JE isolates and Nakayama-NIH strain may be dependent on structural difference of envelope protein.

• 한국동물위생학회지
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• 제24권4호
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• pp.359-367
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• 2001

A serological survey was performed to establish basic data for the prevalence of antibodies to some major diseases of domesticated boar serum samples from January to December 2000. Sera collected in breeding farms in Gyeongbuk province were tested for Aujeszky's disease virus(ADV), Porcine reproductive and respiratory syndrome virus(PRRSV), Porcine parvovirus(PPV), Japanese encephalitis virus (JEV), Bordetella bronchiseptica(B bronchiseptica), Mycoplasma ; APP), Toxoplasma, and Brucella. There was no antibody to ADV in domesticated boars serum samples detected by Anti-ADV-gpI assay kit. Sero-positive samples to PRRS by IFA were 0.9%(3/330) The HI titers to PPV ranged variously from less than 10 to over 1,280. Two hundred ninety-four out of 330 tested sera showed HI titer of less than 10. In HI test to JEV, 90.3% of the sera (298/330) were below 10. The majority of the serum samples had low prevalence of the antibody B bronchiseptica. ELISA titers to M hyopneumoniae ranged variously from $\leq$ 10 to $\geq$ 1,280. Antibody titers to A pleuropneumoniae type 2(APP2) and type 5(APP5) were 58.2% and 52.7%, respectively, and the tested samples showing ELISA antibody titers of less than 20. There was no significant geographical difference between APP2 and APP5 in this study. In the antibody test of Toxoplasma, 11.5%(38/330) were positive and samples were all negative in sera test of Brucella.

• 대한수의학회지
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• 제29권2호
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• pp.109-114
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• 1989

1982년(年)부터 1984년도(年度)까지 3년간(年間)에 걸처, 일본내(日本內) 중부(中部) Saitama현(縣)의 우(牛) 1,306두(頭)와 남부(南部)의 Kagoshima현(縣)의 우(牛) 536두(頭)를 대상(對象)으로 하여 일본뇌염(日本腦炎)바이라스 (JEV)의 적혈구응집억제(赤血球凝集抑制) (HI) 항체양성율(抗體陽性率)을 검사(檢査)한 바, Kagoshima현(縣)에선 68.8% 그리고 Saitama 현(縣)에서는 65.5%가 양성(陽性)이었다. 계절별(季節別)로는 양지역(兩地域)이 공(供)히 하절(夏節)에 항체양성율(抗體陽性率)이 높았고, 연령별(年齡別) 양성율(陽性率)은 Saitama 현(縣)의 경우 64.0%부터 82.8%까지 분포(分布)하고, Kagoshima현(縣)의 우(牛)는 1세군(歲群)에서 29.4%, 2세군(歲群)에선 50.0%, 3세군(歲群)에선 47.4% 그리고 4세군(歲群)에선 74.5%의 양성율(陽性率)을 나타내었다. 그리고 양지역(兩地域)의 연령(年齡)과 항체력가간(抗體力價間)에는 상관성(相關性)이 없었고, Saitama현(縣) 우(牛)의 역가(力價)는 연령(年齡)에 따라 15.3~22.5, Kagoshima 현우(縣牛)은 20.0~32.3이었다.

• 한국동물위생학회지
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• 제34권2호
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• pp.111-116
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• 2011

Artificial insemination (AI) of swine is a very useful reproductive tool and that offers convenience in the Korean swine industry. Since many viruses have been reported to be excreted through boar semen, we investigated the presence of antibodies and antigens against viruses causing reproductive failure in semen of boar in 349 semen samples collected from six Korean AI centers. Viral antigens were detected by polymerase chain reaction (PCR) or reverse transcription-PCR predominantly. The results was as follows. The major reproductive failure causing factor was porcine circovirus type 2 (PCV2), followed by porcine reproductive and respiratory syndrome virus (PRRSV) ($X^2$=166.64, P<0.001). PCV2 and PRRSV, Japanese encephalitis virus (JEV), encephalomyocarditis virus (EMCV) was detected in 73 samples (20.9%), 44 samples (12.6%), 4 samples (1.1%), 3 samples (0.9%), respectively and porcine parvovirus in one sample (0.3%) Classical swine fever virus (CSFV), bovine viral diarrhea virus and Aujeszky's disease virus (ADV) were not detected. Enzyme-linked immunosorbent assay was carried out in 111 serum samples from three AI centers. In most pigs, antibodies response was showed prominently in CSFV (105 sera, 94.6%) ($X^2$=82.580, P<0.001), followed by, in PRRSV (100 sera, 90.1%), PCV2 (92 sera, 90.1%), and PPV (8 sera, 82.9%). ADV antibody was not detected. Thus, the experimental results will be used for the base data, with respect to the state of viral stillbirth in general pig farms, as well as AI centers and breeding farms in Korea.

• 대한수의학회지
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• 제34권1호
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• pp.77-83
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• 1994

유사산 태아의 폐, 청색증을 나타내는 자돈으로부터 돼지생식기 및 호흡기증후군(PRRS)의 원인체로 추정되는 바이러스주(KPRRSV) 들을 분리하였다. 분리된 바이러스주는 돼지콜레라, 돼지오제스키병, 돼지뇌심근염바이러스에 대한 형광항체반응에서는 음성이었으며 기니픽혈구에 대한 혈구응집 능력을 나타내지 않았다. 그리고 포유 마우스의 뇌내 접종시 이상을 나타내지 않았으나 돼지생식기 및 호흡기증후군에 대한 형광항체검사시 양성반응을 나타내었다. 분리된 바이러스는 돼지폐포탐식세포(porcine alveola macrophages)에서 세포변성효과(cytopathic effect)를 나타내었으며 세포변성효과를 나타내었던 바이러스주중 일주(KPRRSV-1)를 돼지폐포탐식세포에서 7대 연속 계대하여 돼지에 접종한 후 혈청을 분리하여 미국 및 유럽지역에서 분리된 돼지유행성 유사산 및 호흡기증후군의 바이러스를 탐식세포에 감염시켜 효소면역방법 (immunoperoxidase monolayer assay)으로 분석한 결과 분리된 바이러스는 미국형 돼지호흡기 및 유사산증후군에 가까운 항원형으로서 판명되었다.