신생자견에 있어서 Canine parvovirus에 대한 혈청학적 연구

박경옥 ( Kyung Ok Park ),김상윤 ( Sang Yun Kim ),조옥숙 ( Ok Sook Cho ),김정화 ( Jeong Hwa Kim ),김대원 ( Dae Won Kim ) 한국가축위생학회 1999 韓國家畜衛生學會誌 Vol.21 No.1

The present study was conducted to characterize maternal antibody status which haemagglutination inhibition(HI) titers against canine parvovirus(CPV) in the 15 puppies delivered from 3 dams. The range of HI titers of 5 puppies delivered from a mother dog(A) with HI titer of 1:1,024 were 1:16~1:64 at 1 day old before suckling, 1:512~1:1,024 at 2 days old after suckling, 1:512~1:2,048 at 1 week old, 1:256~1:1,024 at 2 weeks old, 1:128~1:512 at 3 weeks old, 1:128~1:256 at 4 weeks old, 1:32~1:128 at 5 weeks old, 1:16~1:64 at 6 weeks old, 1:16~1:64 at 7 weeks old, and 1:16~1:32 at 8 weeks old. After vaccination w1th DHPPL to canine parvovirus in 60 days and 80 days old puppies, 1:8~1:32 at 9 weeks old, 1:16~1:128 at 10 weeks old, 1:32~1:256 at 11 weeks old, 1:16~1:256 at 12 weeks old, 1:128~1:256 at 13 weeks old, 1:64~1:512 at 14 weeks old, and 1:128~1:512 at 15 weeks old. The HI titers of 3 puppies delivered from a mother dog(B) with HI titer of 1:512 were 1:16 at 1 day old before suckling, 1:256~1:512 at 2 days old after suckling, 1:512 at 1 week old, 1:128~1:256 at 2 weeks old, 1:64~1:128 at 3 weeks old, 1:64~1:128 at 4 weeks old, 1:128 at 5 weeks old, 1:64~1:128 at 6 weeks old, 1:16 at 7 weeks old, and 1:8 at 8 weeks old. After vaccination with DHPPL to canine parvovirus in 60 day and 80 days old puppies, <:8~1:8 at 9 weeks old, <:8~1:16 at 10 weeks old, 1:64~1:128 at 11 weeks old, and 1:256~1:512 at 12 weeks old. The HI titers of 7 puppies delivered from mother dog(C) with HI titer 1:1,024 were 1:512~1:1,024 at 2 days old after suckling, 1:256~1:1,024 at 1 week old, 1:256~1:1,024 at 2 weeks old, 1:64~1:512 at 3 weeks old, 1:64~1:512 at 4weeks old, 1:8~1:64 at 5 weeks old, 1:8~1:64 at 6 weeks old, 1:8~1:32 at 7 weeks old, and <1:8~1:8 at 8 weeks old. Antibody to CPV was transferred mainly from mother to progeny through the colostrums and the transferred maternal antibody was in proportion to the HI titer of the mother. As the HI titer of maternal antibody in puppies was low, puppies have a rapid immune response and a massive rise in HI titer to vaccination against CPV compared with puppies having high level of maternal antibody.

Production of recombinant new canine parvovirus 2a viral protein 2 in SF9 cells using a baculovirus expression system

Hyeonhae Choi,Seyeon Park,Yoon-Hee Lee,Ju-Yeon Lee,Jae Young Song,Seo Young Moon,Suyeong Yun,Soo-dong Cho,Jienny Lee,Bang-hun Hyun,In-Ohk Ouh 한국예방수의학회 2020 예방수의학회지 Vol.44 No.3

Canine parvovirus (CPV) remains a leading infectious cause of death in canines, especially in young puppies. Though vaccination is being carried out regularly, immunization failures occur, and puppies may be exposed to infection. Virus-like particles (VLPs) act like a subunit vaccine, mimicking the structure of authentic viruses. Therefore, VLPs have the potential to be used as vaccine candidates. Since Viral Protein 2 (VP2), a major structural protein of CPV, is the crucial antigen for CPV, the purpose of this study was to produce a recombinant VP2 of new canine parvovirus-2a using the baculovirus expression system in SF9 insect cells. The results revealed that recombinant VP2 assembles to form VLPs with antigenic properties similar to those of natural CPV, the recombinant VLP can produce a hemagglutination assay (HA) titer (1:2¹⁰) in SF9 cells. Expression of the recombinant 6-His-tagged VP2 in SF9 cells was confirmed by western blotting. These findings suggest that the recombinant VP2 expressed in this study could be used as an efficient subunit vaccine against CPV infection.

국내에서 분리된 canine parvovirus의 구조유전자 cloning과 염기서열 분석

박종현,송재영,이중복,현방훈,안수환,전무형 충남대학교 생물공학연구소 1993 생물공학연구지 Vol.3 No.-

In this study gene encoding structural proteins of a CPV isolate was cloned and saquenced to elucidate the molecular genetical properties of the canine parvoviruses isolated from the field. Six recombinant plasmids of pEP3, p1471, p2070, pEP069, pEP338 and p14711p were constructed from the map positions 22 to 98 of RF DNA to clone the VP1 and VP2 genes of CPV-V20. Sequentialy the gene comprising 3780 nucleotides were sequenced by dideoxy chain termination method. When nucleotide sequence of gene encoding the structural proteins of CPV-V20 was compared with those of other strains, CPV-N, CPV-d and CPV-780929 published previously. DNA homologies to CPV-V20 were 99.87% with CPV-NM, 99.73% with CPV-d, 96.85% with CPV-780929 AND 98.4% with FPLV-Carl, respectively. The DNA sequence data of CPV-V20 showed seven point mutations and also deletion of 135 nucleotides from the nucleotide position 4745 to 4879 located in the 3-noncoding region of CPV-N.

Baculovirus를 利用한 Canine Parvovirus VP2蛋自質의 發現

朴鍾賢,宋載永,玄芳動,安動濬,姜永源,全茂炯,安壽煥 충남대학교 생물공학연구소 1998 생물공학연구지 Vol.6 No.-

국내에서 分離된 개파보바이러스주(V20주)의 VP2 遺傳子를 baculovirus system을 이용하여 발현시켜 다음의 結果를 얻었다. 1. 개파보바이러스의 VP2 유전자를 PCR에 의해 增幅하여 1755bp의 VP2遺傳子를 pUC19에 클로닝하여, 클로닝된 遺傳子를 polyhedrin promoter를 가지는 baculovirus expression vector인 pVL1393에 옮겨 VP2 발현벡터인 pVL1393-VP2를 얻을 수 있었다. 2. pVL1393-VP2 plasmid와 baculovirus DNA와의 homologous recombination에 의해 재조합바이러스인 VP2-BV를 얻을 수 있었으며, 그 발현효율은 2.000-5.000 HAU/0.05 ml이었다. 3. 免疫沈澱法에 의해 발현된 단백질은 개파보바이러스의 VP2단백질과 유사한 64 kb에 달하는 것이었으며, 血球凝集能을 지니고 있었다 4. 血球凝集能을 지닌 蛋白質이 여러 陽性血淸에 있어서 抗體수준을 測定할 수 있는지 개파보바이러스항원과의 相關性을 比較한 바 0.94 (n=125. p<0.01)의 相關係數를 보였다 5. 발현 VP2白은 virus-like particles를 形成하였으며, 그 크기로는 개파보바이러스와 비슷한 25 ㎚의 크기를 갖았다. Canine parvovirus(CPV) is a member of autonomous replicating parvoviruses and is aetiologically associated with enteritis and mycoarditis in puppies. The capsids of CPV are composed of three structural proteins: VP1, VP2 and VP3. The VP2 protein is the major component of capsid. The VP2 gene of a canine parvovirus. V20 strain isolated in Korea was cloned into baculovirus expression vector, and subsequently the VP2 protein was expressed by a recombinant baculovirus under the control of polyhedrin promoter. The recombinant VP2 protein expressed in Spodoptera frugiperda 9(Sf9) cells was detected by haemagglutination(HA) test and immunofluorescent antibody assay. Molecular weight of the recombinant VP2 protein expressed was estimated as 64Kd when tested by immunoprecipitation test using anti-CPV monoclonal antibody. In haemagglutination inhibition(Ⅲ) test. 8 HA units of the recombinant VP2 protein antigen was successfully utilized to determine a level of antibody against CPV in various positive sera. The recombinant VP2 protein showed also the capability to form virus like particles similar in size and appearance to the CPV virions.

Production of VP2 protein of canine parvovirus-2c expressed in baculovirus expression system

Seyeon Park,Ju-Yeon Lee,Jae Young Song,Yoon-Hee Lee,Hyeonhae Choi,Seo Young Moon,Min Ji Kim,Suyeong Yun,Soo-dong Cho,Jienny Lee,Bang-hun Hyun,In-ohk Ouh 한국예방수의학회 2020 예방수의학회지 Vol.44 No.2

Viral protein 2 (VP2), which is the structural protein of parvovirus, can produce virus-like particles (VLPs) by a self-assembly process in vitro, making VLPs attractive vaccine candidates. VP2 of canine parvovirus (CPV) is responsible for neutralizing antibodies in immunized animals. In this study, VP2 protein of canine parvovirus-2c was expressed using a baculovirus expression system and assembled into parvovirus-like particles in insect cells. The results show that VP2 proteins assembled into virus-like particles (VLPs) with antigenic properties similar to those of natural CPV and a high hemagglutination (HA) titer (1:2⁷). The recombinant 6-His-tagged VP2 protein with a molecular mass of about 65 kDa was detected by anti-His antibody and anti-PPV serum. This study provides a foundation for the application of VP2 protein in the clinical diagnosis of CPV and in the vaccination against CPV.

설사증 라환견(羅患犬)으로 부터 분리한 Canine Parvovirus의 성상에 관한 연구

최해연 ( Hae Yeon Choi ),정운선 ( Un Sun Chung ),전무형 ( Moo Hyung Jun ),박성국 ( Seong Kuk Park ),민원기 ( Won Gi Min ) 한국가축위생학회 1990 韓國家畜衛生學會誌 Vol.13 No.2

From 1988 to 1989, 8 strains of canine parvovirus-2 (CPV-2) were isolated from the fecal specimens from the dogs that were clinically diagnosed as canine parvoviral enteritis in the veterinary hospitals located in the regions of Taejon and Chungbuk province. Studies on biological and physicochemical properties for the isolates were carried out. The results obtained by experiments are summarized as follows. 1. Among 62 fecal samples collected from the dogs with enteric diseases, 24 (38.7%) showed the haemagglutinating activity against porcine erythrocyte, ranging from 16 to 16,384 of HA titers. 2. When 8 fecal specimens with high HA titer over 1,000 were inocultated into CRFX cells, intranuclear inclusion bodies were obseverd in all of eight specimens, of which three specimens showed cytoplasmic inolusions concurrently with the intranuclear inclusion bodies. 3. In study on species-specificity of haemagglutinating activity of the isolates, TJ-89-1 and TJ-89-2, it was found that the isolates revealed the highest haemagglutinating activity with porcine erythrocytes, showing the relatively lower haemagglutination titers with the erythrocytes from cat and rabbit. None of erythrocytes from other animals reacted with the isolates. 4. By the cross-haemagglutination inhibition test of the isolates with reference viruses and sera, the isolates were evidently identified as the strains of canine parvovirus-2. 5. In physicochemical property test, it was evident that the isolates were stable in lipid solvent, pH and heat treatment at 56℃ for 30 mm. and contain DNA genome. 6. When seven puppies were inoculated intraorally with the isolate at HA titer of 8, 192, all of the puppies showed the symptoms of anorexia, vomiting, diarrhea and died at the 5th to 10th days post inoculation(pi). The fecal samples from all of the puppies revealed significantly high HA titers afterward the 5th days pi. Body temperature and the number of total leucocytes were slightly increased at the early stage of infection, but extremely decreased at the stage of collapse. HI titers of the sera started to increase at the 2nd to 3rd days pi reaching 512 to 1,024 at the 4th to 5th day pi.

설사증 나환견으로 부터 분리한 Canine Parvovirus의 성상에 관한 연구

최해연,정운선,전무형,박성국,민원기 한국동물위생학회 1990 韓國家畜衛生學會誌 Vol.13 No.2

From 1988 to 1989, 8 strains of canine parvovirus-2 (CPV-2) were isolated from the fecal specimens from the dogs that were clinically diagnosed as canine parvoviral enteritis in the veterinary hospitals located in the regions of Taejon and Chungbuk province. Studios on biological and physicochemical properties for the isolates were carried out. The results obtained by experiments are summarized as follows. 1. Among 62 fecal samples collected from the dogs with enteric diseases, 24 (38.7%) showed the haemagglutinating activity against porcine erythrocyte, ranging from 16 to 16, 384 of HA titers. 2. When 8 fecal specimens with high HA titer over 1, 000 were inocultated into CRFX cells, intranuclear inclusion bodies were obseverd in all of eight specimens, of w)lick three specimens showed cytoplasmic inclusions concurrently with the intranuclear inclusion bodies. 3. In study on species-specificity of haemagglutinating activity of the isolates, TJ-89-1 and TJ-89-2, it was found that the isolates revealed the highest haemagglutinating activity with porcine erythrocytes, showing the relatively lower haemagglutination titers with the erythrocytes from cat and rabbit. None of erythrocytes from other animals reacted with the isolates. 4. By the cross-haemagglutination inhibition test of the Isolates with reference viruses and sera, the Isolates were evidently identified as the strains of canine parvovirus-2. 5. In Physicochemical property test, it was evident that the isolates were stable in, lipid solvent, pH and heat treatment at $56^{\circ}C$ for 30 min. and contain DNA genome. 6. When seven puppies were inoculated intraorally with the isolate at HA titer of 8, 192, all of the puppies showed the symptoms of anorexia, vomiting, diarrhea and died at the 5th to 10th days post inoculation(pi). The fecal samples from all of the puppies revealed significantly high HA titers afterward the 5th days pi. Body temperature and the number of total leucocytes were slightly increased at the early stage of infection. but extremely decreased at the stage of collapse. HI titers of the sera started to increase at the 2nd to 3rd days pi reaching 512 to 1, 024 at the 4th to 5th day pi.

설사증 이환견(罹患犬)으로 부터 분리한 canine parvovirus의 성상에 관한 연구

최해연,전무형,박성국,Choi, Hae-yeon,Jun, Moo-hyung,Park, Seong-kuk 대한수의학회 1991 大韓獸醫學會誌 Vol.31 No.3

From 1988 to 1989, 8 strains of canine parvovirus-2(CPV-2) were isolated from the fecal specimens from the dogs that were clinically diagnosed as canine parvoviral enteritis in the veterinary hospitals located in the regions of Taejeon and Chungbuk province. The biological and physicochemical properties for the isolates were studied. Among 62 fecal samples collected from the dogs with enteric diseases, 24(38.7%) showed the haemagglutinating activity to porcine erythrocyte ranging from 16 to 16,384 of HA titers. In cytopathological studies with CRFK cells, intranuclear inclusion bodies were observed in all of eight specimens with the high HA titer over 1,000, of which three specimens showed cytoplasmic inclusions concurrently with the intranuclear inclusion bodies. It was found that the isolates revealed the highest haemagglutinating activity with porcine erythrocytes and the relatively lower haemagglutination titers with the erythrocytes from cat and rabbit. None of erythrocytes from the other animals reacted with the isolates. By the cross-haemagglutination inhibition test for the isolates with the reference viruses and sera, the isolates were evidently identified as the strains of CPV-2. In physicochemical property test, the isolates were stable in lipid solvent, pH and heat treatment at $56^{\circ}C$ for 30 min, and showed the virus particle size less than 25 nm, containing a DNA genome.

국내에서 분리된 canine parvovirus의 구조유전자 cloning과 염기서열 분석

박종현,송재영,이중복,현방훈,안수환,전무형,Park, Jong-hyeon,Song, Jae-young,Lee, Jung-bok,Hyun, Bang-hun,An, Soo-hwan,Jun, Moo-hyung 대한수의학회 1992 大韓獸醫學會誌 Vol.32 No.4

In this study gene encoding structural proteins of a CPV isolate was cloned and sequenced to elucidate the molecular genetical properties of the canine parvoviruses isolated from the field. Six recombinant plasmids of pEP3, p1471, p2070, pEP069, pEP338 and p14711p were constructed from the map positions 22 to 98 of RF DNA to clone the VP1 and VP2 genes of CPV-V20. Sequentialy the gene comprising 3780 nucleotides were sequenced by dideoxy chain termination method. When nucleotide sequence of gene encoding the structural proteins of CPV-V20 was compared with those of other strains, CPV-N, CPV-d and CPV-780929 published previously, DNA, homologies to CPV-V20 were 99.87% with CPV-N, 99.73% with CPV-d, 96.85% with CPV-780929 and 98.4% with FPLV-Carl, respectively. The DNA sequence data of CPV-V20 showed seven point mutations and also deletion of 135 nucleotides from the nucleotide position 4745 to 4879 located in the 3'-noncoding region of CPV-N.

울산지역의 개 디스템퍼 및 파보 장염의 항체보유 실태 조사

성기창 ( Ki Chang Sung ),이은우 ( Eun Woo Lee ),박창은 ( Chang Eun Park ) 한국가축위생학회 2010 韓國家畜衛生學會誌 Vol.33 No.3

The results from a total of 412 blood samples consisted of 187 samples from regular visiting group (RV), 94 samples from first visiting group (FV), 52 samples from abandoned group (A), 54 samples from special breeder group (SB), and 25 samples from preliminary breeder group (PB) showed that RV(94.7%) and SB(88.9%) groups had the higher levels of protective antibody, but PB (36.0%) group revealed the lowest level. Among 96 blood samples with lower protective antibody levels, 14 samples (14.6%), 72 samples (75.0%) and 10 samples (10.4%) were below the protective antibody levels to distemper/parvo-virus, distemper only and parvovirus only, respectively. These results implied that antibody to parvovirus was well generated than that to distemper. Eighty six samples (20.9%) showed the protective antibody titer under 1:96 to distemper and 24 samples (5.8%), the protective antibody titer under 1:40 to parvovirus.