국내 분리주를 이용한 오리 바이러스성 간염 생백신주의 개발

성환우,김재홍,송창선,한명국,이윤정,모인필,김기석,Sung, Haan-woo,Kim, Jae-hong,Song, Chang-seon,Han, Myung-guk,Lee, Youn-jeong,Mo, In-pil,Kim, Ki-seuk 대한수의학회 2000 大韓獸醫學會誌 Vol.40 No.1

Duck viral hepatitis is an acutic, highly infectious viral disease of young ducklings. The most practical means for controlling duck viral hepatitis is the vaccination of ducklings or of a breeding stock. We attempted to develop a vaccine strain of duck hepatitis virus (DHV) using a Korean isolate by serial chicken embryo passages. The propagation of DHV in chicken embryos was carried 140 passages. After the $50^{th}$ passage, of which the virus was non-pathogenic for ducklings, approximately every $20^{th}$ passage of the virus was tested for vaccinal efficacy. Both the $70^{th}$ and $90^{th}$ passage of the virus gave good protection against challenge infection to a DHV-DRL reference strain(type 1) and a virulent Korean isolate. The $110^{th}$, $125^{th}$ and $140^{th}$ passage of the virus were less protective than the $70^{th}$ and $90^{th}$ passage, which means that more than $110^{th}$ passage may lead to over-attenuation of the virus. Ducklings vaccinated with the chicken-embryo-adapted virus by oral, intramuscular or eye drop administration showed earlier resistance to challenge infection from 3 to 7 days postvaccination. Of the above methods, ducklings vaccinated intramuscularly presented the most rapid resistance against challenge. The minimum immune dose of the chicken-embryo-adapted virus in ducklings was also studied. Ducklings inoculated with a dose of $10^{2.0}\;ELD_{50}$ and below were not fully protected against challenge with a virulent DHV, showing a protection rate of 67% to 73%, but ducklings inoculated with a dose of $10^{3.0}\;ELD_{50}$ and over were completely protected. The virus yield of the chicken-embryo-adapted DHV was examined at 24hrs and 48hrs of the incubation time in the allantoic fluid, embryo head and embryo minus head of the embryonating egg. In all three components, the titer of the virus was higher at 48 hours than that at 24 hours after incubation. And the titer of the virus was higher in the embryo minus head, embryo head and the allantoic fluid, in order. Field trials for evaluating the efficacy of the attenuated DHV as a live vaccine were done in duck farms with about 25% mortality of flocks resulting from duck viral hepatitis. After the use of the experimental vaccine, the mortality due to duck viral hepatitis was dramatically reduced in the farms. These results indicated that the attenuated DHV using a Korean isolate could be a good candidate as a live vaccine strain of DHV in Korea.

산란계에서의 칸디다증 국내 발생 사례

성환우,권혁무,한정희,모인필,Sung, Haan-Woo,Kwon, Hyuk-Moo,Han, Jeong-Hee,Mo, In-Pil 대한수의학회 2010 大韓獸醫學會誌 Vol.50 No.4

Candidiasis is a mycosis caused by the mycelial yeast of the Candida genus which is opportunistic pathogen of humans, animals, and birds. Under some conditions such as prolonged antibiotic therapy, overcrowding, and immunosuppression, the opportunistic Candida can cause disease. Chicken candidiasis is sporadically occurred and characterized by unsatisfactory growth, listlessness, roughness of feathers, and death. A case of 23 weeks old layer with history of increased mortality and anemia was submitted to our Lab. At necropsy, the characteristic lesions were observed in the crop and proventriculus. The whitish pseudomembrane, that are peeled easily, was found in the crop. Proventriculus was swollen and the mucosa was covered with hemorrhagic exudate. The histological changes of the affected crop are epithelial hyperplasia, hydropic degeneration, and mycelia formation. Smears made from the necrotic mucosal surfaces of the crop revealed the presence of large number of yeast cells and mycelia. Pure cultures of yeast colonies were obtained from the potato dextrose agar. The yeast cells were identified as Candida albicans by gene sequencing. To our knowledge, this is the first report of candidiasis in chickens with anemia in Korea.

세망내피증 바이러스 감염 닭에서의 혈액화학치 변화

성환우,권혁무,김선중,Sung, Haan-Woo,Kwon, Hyuk-Moo,Kim, Sun-Jung 대한수의학회 2008 大韓獸醫學會誌 Vol.48 No.4

Body weights and blood biochemical values in chickens infected with reticuloendotheliosis virus (REV)-HI, a Korean isolate, were studied. REV-HI causes severe body weight depression in chickens inoculated but not in chicken contact-infected. Body weights of infected chickens in 3, 4, and 5 weeks after infection were 78%, 76% and 65% of those of control respectively. Blood glucose levels in REVinfected chickens were extremely high compared with those in control (226 $\geq$ 21 vs. 814 $\geq$91.3 mg/dl in week 2) during the experiment period. Triglyceride levels in REV-infected chickens were significantly higher in week 2 and 3, whereas in week 4, REV-infected chickens showed significantly lower levels than the control. Blood lipase, amylase and alkaline phosphatase levels of REV-infected chickens in week 2 were significantly higher, whereas cholesterol, magnesium and calcium values in week 4 were significantly lower than the control. Other blood biochemical values such as alkaline aminotransferase, aspartate aminotransferase, and $\gamma$-glutamyltransferase were nonsignificantly different from the control. These above results suggest that weight depression by REV may be related with increase of blood glucose, which indicated that REV-infected chickens could not use blood glucose as energy source.

세망내피증 바이러스 항체검출을 위한 ELISA 표준화

성환우,이수정,Sung, Haan Woo,Lee, Su Jeong 대한수의학회 2005 大韓獸醫學會誌 Vol.45 No.4

Enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to reticuloendotheliosis virus (REV) at single serum dilution was standardized. REV HI, one of the Korean field isolates, was inoculated into chicken embryo fibroblast (CEF) cells and was harvested from the culture fluids and cells after 10 to 12 days. Viruses were purified by centrifugation at the $107,000{\times}g$ for 12 hours on 20, 30, 45% (W/V) sucrose gradient. Virus specific fraction was collected and used as ELISA antigen. To standardize ELISA, the optimal concentration of coating antigen ($1{\mu}g/well$) and conjugate (1/1000) was determined by corrected OD (OD value of positive serum-OD value of negative serum) and P/N ratio (OD value of positive serum/OD value of negative serum). To calculate ELISA titer by measuring absorbance at 1/400 single serum dilution, serum titrations were carried out for various sample sera together with standard positive and negative sera. The observed titers of serum samples were plotted against sample/positive (s/p) ratios at 1/400 serum dilution. From the above data, the ELISA titers could be calculated by the equation of $log_{10}$ ELISA titer = 2.2763 ($log_{10}$ s/p) + 3.482 (r = 0.93). For evaluating the sensitivity, the standardized method were compared with conventional agar gel immunodiffusion (AGID) test method using serum samples collected from REV infected field chicken flocks. Fifty seven of 60 samples (95%) were positive for REV by ELISA, whereas only 11 (18.3%) samples were positive by AGID test. This results suggested that the ELISA tests developed in this study could be used for detection of antibodies to REV with high sensitivity.

오리 간염 바이러스의 분리와 국내 분리주의 약독화

성환우,김재홍,Sung, Haan-woo,Kim, Jae-hong 대한수의학회 2000 大韓獸醫學會誌 Vol.40 No.1

Duck viral hepatitis is an acute, highly infectious viral disease of young dacklings aged from two days to three weeks. The significant lesion associated with the disease was enlarged liver including necrotic foci and numerous hemorrhagic spots. We have isolated five strains of duck hepatitis virus (DHV) from field cases showing about 20% mortality with a sign of opisthotonos. When a-day-old ducklings were intramuscularly inoculated with one of the isolates, 92% of the birds were died within 5 days. We attempted to develop an attenuated strain of duck hepatitis virus (DHV) using one of the isolates by serial chicken embryo passages. The propagation of DHV in chicken embryos was carried 140 passages. The virus titer increased gradually from the $21^{st}$ through the $50^{th}$ passage, but there was no significant increase of virus titer in subsequent passages after then. Through the serial passages, the virulence of the virus for chicken embryos was gradually increased but decreased for ducklings. The pathogenicity of the virus for ducklings was preserved up to the $21^{st}$ passage but disappeared at the $50^{th}$passage. An attenuated Korean isolate which was passaged 140 times in chicken embryos gave good protection in ducklings against both challenge infection to a Korean virulent strain and to a DHV-DRL strain, a type 1 reference strain of DHV, which indicated that the Korean isolates could be classified as DHV type 1. And the above results suggest that an attenuated Korean isolate can be used for developing a live DHV vaccine.

깔짚 교체 및 재사용 육계농장 분리 대장균의 항생제 내성 양상

성환우,최강석,권혁무,이영주,Sung, Haan-Woo,Choi, Kang-Seuk,Kwon, Hyuk-Moo,Lee, Young-Ju 대한수의학회 2017 大韓獸醫學會誌 Vol.57 No.3

The isolation rate of Escherichia (E.) coli in poultry litter was investigated at 44 broiler farms, 20 that used fresh litter and 24 that used recycled litter. The patterns of resistance to antibiotics of the E. coli isolates were compared. In litter sampled before the rearing period, the isolation rate of E. coli was higher at farms that used fresh litter; E. coli was present in the litter in 94.5% (35 out of 37 flocks tested) of the farms that used fresh litter vs. 51.2% (21 out of 41 flocks) of the farms that used recycled litter. The susceptibility of the 93 isolates of E. coli to 13 antibiotics was studied. Before the rearing period, E. coli isolates from the farms that recycled litter showed higher resistance rates than isolates from farms that replaced litter with fresh litter. Comparing the antibiotic resistance patterns of isolates from litter sampled before and at the end of the rearing period, the antibiotic resistance rates at the end of the rearing period increased dramatically compared with rates before the rearing period.

Vaccines for the Control of Avian influenza

성환우(Haan Woo Sung) 한국실험동물학회 2006 한국실험동물학회 학술발표대회 논문집 Vol.2006 No.-

닭의 혈액내 단핵세포 표면항원 특이 단클론성 항체 생산

최준구,성환우,김선중,Choi, Jun-Gu,Sung, Haan-Woo,Kim, Sun-Joong 대한수의학회 2002 大韓獸醫學會誌 Vol.42 No.2

This study was performed to produce monoclonal antibodies (mAb) specifically reacting with chicken leukocyte surface antigens. Popliteal lymph node cells of BALB/c mice previously immunized through foot-pad with peripheral blood mononuclear cells (PBMC) of chickens separated by Ficoll-Histopaque method. They were fused with P3X63Ag14 mouse myeloma cells. A total of 34 hybridomas secreted antibodies specifically binding to the PBMC. According to the reactivity patterns with PBMC, the mAbs were divided into 4 groups. Group 1 mAbs (IIB3, IIB10, IIE10) specifically reacted with non-adherent lymphocytes but not with adherent cells which were mainly composed of thrombocytes and monocytes in PBMC culture. These mAbs were reactive with 25-59% of thymus cells and 42-64% of spleen cells of chickens. They did not show any significant reactivity with cells in the bursa of Fabricius, T-cell (MDCC-MSB1) and B-cell (LSCC-1104B1) lines. These results indicate that Group I mAbs specifically reacted with T-lymphocyte subpopulation. Monoclonal antibodies in Group II (IC6, IG2-2 and IID9) showed specific reactivity with monocytes but not with thrombocytes or non-adherent cells in PBMC culture. These mAbs, though not reacted with the chicken macrophage cell line, HD11, also bound to macrophages of the spleen and lung in immunohistochemical staining. Five mAbs in Group III showed characteristics of binding to lymphocytes and monocytes, but not to thrombocytes. Twenty-three mAbs in Group IV showed specific reactivity to lymphocytes, monocytes, and thrombocytes. Two mAbs (IC3 and IE9) in Group IV reacted with most of PBMC.

닭의 전염성 F낭병 바이러스 유전자백신에 의한 방어 면역에 Genetic Adjuvant (Chicken Interleukin-6)와 Chemical Adjuvant (Levamisole)의 효과

박정호,성환우,윤병일,박선일,권혁무,Park, Jeong-Ho,Sung, Haan-Woo,Yoon, Byung-Il,Pak, Son-Il,Kwon, Hyuk-Moo 한국미생물학회 2009 미생물학회지 Vol.45 No.2

닭의 전염성 F낭병 바이러스(IBDV)가 원인 바이러스인 전염성 F낭병은 전 세계 양계산업에 경제적으로 피해가 큰 중요한 질병이다. 이 연구의 목적은 닭에서 IBDV에 대한 방어면역을 유도하기 위한 in ovo 초회항원자극(priming)과 불활화백신에 의한 보강접종 방법에 항원보강제(adjuvant)로 chicken interleukin 6 (pcDNA-ChIL-6;plasmid encoding chicken interleukin-6)와 levamisole (LMS)의 효과를 조사하는 것이다. IBDV의 VP2, VP, VP3 protein을 암호화하는 유전자백신인 plasmid DNA vaccine (pcDNA-VP243) 단독 또는 pcDNA-ChIL-6 또는 LMS와 함께 18일령 부화란의 양막낭(amniotic sac)에 접종하고 부화한 1주령의 병아리에 불활화 IBD 백신을 근육 접종한 다음 3주령에 고병원성 IBDV인 SH/92 주로 공격 접종하고 10일 동안 관찰하였다. 백신하지 않은 공격접종 대조군이 100%의 폐사율을 보인 반면 pcDNA-VP243 단독 접종군과 pcDNA-VP243에 pcDNA-ChIL-6 또는 LMS를 첨가한 실험군은 모두 100%의 생존율을 나타내었다. 그러나 공격접종 후 F낭의 손상을 평가하기 위한 IBDV RNA의 검출, B/B ratio와 F낭의 병변지수(lesion score) 등을 분석한 결과 pcDNA-VP243에 pcDNA-ChIL-6 또는 LMS를 첨가한 실험군은 pcDNA-VP243 단독 접종군보다 향상된 방어효과를 나타내지 않았다. 이 실험결과는 유전자백신에 의한 in ovo 초회항원자극-불활화백신에 의한 보강접종법이 고병원성 IBDV로부터 닭을 보호하기 위한 효과적인 방법이었으나 pcDNA-ChIL-6 또는 LMS의 첨가로 인한 방어효과의 향상은 나타나지 않았다. Infectious bursal disease (IBD) caused by the infectious bursal disease virus (IBDV) has an important economic impact on the poultry industry worldwide. This study examined the adjuvant effects of a plasmid encoding chicken interleukin-6 (pcDNA-ChIL-6) and levamisole (LMS) on in ovo prime-boost vaccination using a genetic vaccine (pcDNA-VP243) to prime in chicken followed by a killed-vaccine boost. A pcDNA-VP243 was injected into the amniotic sac alone or in combination with a pcDNA-ChIL-6 or LMS at embryonation day 18, followed by an intramuscular injection of killed IBD vaccine at 1 week of age. The chicken were orally challenged with very virulent IBDV (vvIBDV) strain at 3 weeks of age and observed for 10 days. No mortality was observed in the groups that received the pcDNA-VP243 alone and pcDNA-VP243 plus pcDNA-ChIL-6 or LMS compared to 100% mortality in unvaccinated challenge control group. However, as determined by bursal damage (the presence of IBDV RNA, B/B ratio, and lesion score), a pcDNA-VP243 alone group was superior to pcDNA-VP243 plus pcDNA-ChIL-6 or LMS groups in the protection against post-challenge. These findings suggest that in ovo priming with genetic vaccine and boosting with killed vaccine is an effective strategy for protecting chicken against vvIBDV and the addition of pcDNA-ChIL-6 or LMS did not enhance protective immunity.

국내 해산양식어 조피볼락에서 분리된 수생버나바이러스 GC-1의 VP2 발현

조성준,성환우,이윤정,김재홍,강신영,Joh, Seong-joon,Sung, Haan-woo,Lee, Yun-jeong,Kim, Jae-hong,Kang, Shien-young 대한수의학회 2003 大韓獸醫學會誌 Vol.43 No.3

The VP2 gene of aquatic birnavirus, Korean isolate (GC-1) was cloned and expressed using the baculovirus expression system. The VP2 gene and VP2 partial gene, which contained a neutralizing epitope, were constructed for recombinant transfer vectors, for baculovirus expression. The expressed recombinant proteins were confirmed by indirect immuno fluorescence antibody (IFA), SDS-PAGE and Western blot. The level of expression was checked at regular time using IFA and Western blot. To measure the neutralizing activity of recombinant proteins against GC-1 strain, the antisera against recombinant proteins were produced by using guinea pigs. The result showed that the antisera neutralized the GC-1 strain. However, the neutralizing titer was higher in antisera against the VP2 gene expressed recombinant protein than that of VP2 partial gene recombinant protein.