• Journal of Microbiology and Biotechnology
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• 제11권4호
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• pp.619-627
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• 2001

n order to increase the virus safety of a human intravenous immunoglobulin (IVIg) that was manufactured by a successive process of cold ethanol fractionation, polyethylene glycol precipitation, and pasteurization ($60^{\circ}C$ heat treatment for 10h), a low pH incubation process (pH 3.9 at $25{\circ}C$ for 14 days) was employed as the final step. The efficacy and mechanism of the fraction III cold ethanol fractionation, pasteurization, and low pH treatment steps in the removal and/or inactivation of blood-borne viruses were closely examined. A variety of experimental model viruses for human pathogenic viruses, including the Bovine herpes virus (BHV), Bovine viral diarrhoea virus (BVDV), Murine encephalomyocarditis virus (EMCV), and Porcine parvovirus (PPV), were selected for this study. The mechanism of reduction for the enveloped viruses (BHV and BVDV) during fraction III fractionation was both inactivation and partitioning, however, it was partitioning in the case of the nonenveloped viruses (EMCV and PPV). The log reduction factors achieved during fraction III fractionation were ${\geqq}$6.7 for BHV, ${\geqq}4.7$ for BVDV, 4.5 for EMCV, and 4.4 for PPV. Pasteurization was found to be a robust and effective step in inactivating all the viruses tested. The log reduction factors achieved during the pasteurization process were ${\geqq}7.5$ for BHV, ${\geqq}4.8$ for BVDV, 3.0 for EMCV, and 3.3 for PPV. A low pH incubation was very effective in inactivating the enveloped viruses as well as EMCV. The log reduction factors achieved during low pH incubation were ${\geqq}7.4$ for BHV, ${\geqq}3.9$ for BVDV, 5.2 for EMCV, and 2.0 for PPV. These results indicate that the low pH treatment successfully improved the viral safety of the final products.

• 한국식품위생안전성학회지
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• 제18권4호
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• pp.183-188
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• 2003

식중독 원인균인 S. aureus, E. coli O157:H7, S. typhimurium, S. enteritidis, L. monocytogenes, 장티프스 원인균 S. typhi, 패혈증 원인균 V. parahaemolyticus, 세균성 이질 원인균 S. sonnei를 $20^{\circ}C$에서 30분간 $그리존^{TM}$과 접촉시킨 결과 최소 3배 희석액부터 최대 24배 희석액에서 최대 24배 희석액에서 균에 대한 살멸 효과를 나타내었다. $그리존^{TM}$ 3배 희석액을 이용하여 30초, 1분, 5분간 균과 접촉시킨 결과 S. aureus만 제외하고 모두 30초에 100% 살멸하는 효과를 나타내었다. $그리존^{TM}$을 이용하여 사스의 원인체와 동일한 바이러스인 코로나 바이러스에 대한 살바이러스 효과를 본 결과 제품의ㅣ 5배 희석액까지 유효간 살바이러스 효과를 보였다. 사람과 접촉이 많은 애완견 바이러스인 파보바이러스 (CPV), 디스템퍼바이러스 (CDV)에 대하여 시험한 결과 유기물과 경수 등의 악조건에서도 바이러스를 살멸하는 효과를 나타내었다. $그리존^{TM}$을 이용한 회와 냉장육 등에서의 적용 시험결과 미생물 생육이 현저히 저해됨을 알 수 있었다. 체소내의 미생물에 대한 살멸효과 시험에서도 미생물의 수가 현저하게 감소하였고, 특히 대장균의 수가 현저하게 감소하였다. $그리존^{TM}$은 세균, 바이러스의 살멸에 탁원한 효과를 보이며, 음식물에 직접 처리할 시에도 그 안전성과 효능이 입증되었다.

• 대한수의학회지
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• 제47권2호
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• pp.175-185
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• 2007

This study was carried out to investigate the pathogenesis and pathogenicity of the porcine circovirus type 2 (PCV2) Korean isolate from weaned pigs. Twenty four weaned pigs, PCV2, porcine reproductive and respiratory syndrome virus (PRRSV) and porcine parvovirus (PPV) antibodies free, were allocated to 4 groups (n = 6). Six pigs were inoculated intranasally with PCV2 alone, 6 with PCV2 and PRRSV, 6 with the combined PCV2/PRRSV/PPV inoculum, and 6 were remained as a uninoculated negative control. Pigs were killed 3 and 6 weeks after inoculation and tissue samples examined for gross and microscopic lesions and for the presence of PCV2 antigens and nucleic acids. Experimentally inoculated pigs were evaluated for 3 considerations: 1. development of postweaning multisystemic wasting syndrome (PMWS), 2. distribution of viral antigens by immunohistochemistry and polymerase chain reaction (PCR), and 3. cytokine mRNA levels in lymph nodes. Pigs inoculated with PCV2/PRRSV/PPV showed typical clinical signs, gross findings, and histopathologic characteristics of PMWS. In the PCV2/PRRSV/PPV inoculated group, the PCV2 antigen was widely distributed in various parenchymal organs such as brain, spinal cord, tonsil, lymph nodes, lung, heart, liver, kidney, spleen, and peyer's patch. Lymph node mRNA expression of IL-$1{\alpha}$, IL-2R and IL-8 was determined by real-time PCR. The pigs of PCV2/PRRSV and PCV2/PRRSV/PPV inoculation group, the mRNA expression was characterized by a decrease of IL-$1{\alpha}$, IL-2R and IL-8. The decrease of cytokine mRNA represent the state of T cell immuno-suppression in pig, and nicely support the evidence for the impairment of immune system in pigs with PMWS. In conclusion, PCV2 infection and some additional infectious causes such as PRRSV and/or PPV are warranted for the presence of PMWS in weaned pigs 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.

• Journal of Microbiology and Biotechnology
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• 제11권3호
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• pp.497-503
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• 2001

The purpose of this study was to examine the efficacy and mechanism of the cryo-precipitation, solvent/detergent (S/D) treatment, monoclonal anti-FVIIIc antibody (mAb) column chromatography, Q-Sepharose column chromatography, and lyophilization involved in the manufacture of antithemophilic factor VII(GreenMono) from human plasma, in the removal and/or inactivation of blood-borne viruses. A variety of experimental model viruses for human pathogenic viruses, including the bovine viral diarrhoea virus (BVDV), bovine herpes virus (BHV), murine encephalomyocarditis virus (EMCV), and porcine parvovirus (PPV), were all selected for this study. BHV and EMCV were effectively partitioned from a factor VII during the cryo-precipitation with a log reduction factor of 2.83 and 3.24, respectively. S/D treatment using the organic solvent, tri(n-butyl) phosphate (TNBP), and the detergent, Triton X-100, was a robust and effective step in inactivating enveloped viruses. The titers of BHV and BVDV were reduced from the initial titer of 8.85 and $7.89{log_10} {TCID_50}$, respectively, reaching undetectable levels within 1 min of the S/D treatment. The mAb chromatography was the most effective step for removing nonenveloped viruses, EMCV and PPV, with the log reduction factors of 4.86 and 3.72, respectively. Q-Sepharose chromatography showed a significant efficacy for partitioning BHV, BVDV, EMCV, and PPV with the log reduction the log reduction factors of 2.32, 2.49, 2.60, and 1.33 respectively. Lyophilization was an effective step in inactivating g nonenveloped viruses rather than enveloped viruses, where the log reduction factors of BHV, BVDV, DMCV, and PPV were 1.41, 1.79, 4.76, and 2.05, respectively. The cumulative log reduction factors of BHV, BVDV, EMCV, and PPV were ${\geqq}$11.12, ${\geqq}$7.88, 15.46, and 7.10, respectively. These results indicate that the production process for GreenMono has a sufficient virus-reducing capacity to achieve a high margin of the virus safety.

• 한국임상수의학회지
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• 제31권5호
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• pp.361-366
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• 2014

본 연구는 상용화되어 판매되고 있는 CPV-2 백신과 CPV-2b 백신이 국내에서 주로 유행하고 있는 CPV-2a 분리주에 대해 방어능력을 가지는 지를 평가하기 위하여, CPV에 대한 예방접종을 실시하지 않은 생후 9주령의 잡종 강아지 20두를 예방접종 미실시군(대조군) 4두와 예방접종군 16두로 편성하였다. 예방접종 군의 강아지 8두씩은 3주 간격으로 3회에 걸쳐 CPV-2 또는 CPV-2b 백신을 각각 예방접종하였다. 3차 예방접종 2주 후에 모든 강아지에 $1{\times}10^6\;TCID_{50}$의 CPV-2a (VR00174 strain) 바이러스를 경구접종하였으며 강아지들의 임상증상, CPV의 변 내 배출, CPV에 대한 혈청학적 반응에 대하여 2주 동안 관찰하였다. 국내 분리 CPV-2a를 공격접종한 후에, 예방접종을 실시한 모든 강아지에서는 어떠한 임상증상도 나타나지 않았지만 예방접종을 실시하지 않은 강아지들은 식욕부진, 우둔, 구토, 점액 또는 혈액성 설사를 나타내었으며 2두는 공격접종 6일째에 폐사하였다. 예방접종을 실시하지 않은 강아지들에서는 공격접종 4일 후부터 변에서 CPV가 검출되었지만 예방접종을 실시한 강아지에서는 변에서 CPV가 검출되지 않았다. 그리고 예방접종 실시한 강아지들은 1차 예방접종에 의하여 방어수준 이상으로 CPV에 대한 항체를 형성하였다. 본 연구는 현재 국내에서 시판 중인 CPV-2와 CPV-2b 백신이 최근 국내에서 주로 분리되는 CPV-2a에 대하여 교차 방어력을 나타내는 것으로 확인되었다.

• 미생물학회지
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• 제41권4호
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• pp.269-274
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• 2005

동물용 생 바이러스 백신 내에 mycoplasma를 검출하기 위해 polymerase chain reaction (PCR)기법과 2가지의 상품화된 PCR 검출킷트를 평가하였다. PCR기법은 시험에 사용된 모든 mycoplasma를 특이적으로 검출할 수 있었으나, 2가지의 상품화된 PCR 검출킷트는 일부의 mycoplasma를 검출하지 못하였다. 또한, PCR기법의 검출 특이도는 조류 유래 mycoplasma에 속한 4주의 표준주 및 7주의 야외분리주를 모두 검출할 수 있었다. PCR기법의 민감도는 9 CFR Mycoplasma액체배지에서 배양한 Mycoplasma 속균 및 Acholeplasma속균에 대해 $1\~100$ colony forming units/ml까지 검출할 수 있었다. 동물용생 바이러스 백신에 대해 PCR기법의 적용가능성을 평가하기 위해, 돼지 전염성위장염 및 로타바이러스 흔합백신과 개 파보바이러스 백신내에 A. laidlawii를 인공적으로 접종한 후, PCR기법의 민감도를 조사하였을 때 배양액을 이용한 검출한계와 유사하였다. 본 연구에서 사용된 PCR 기법은 동물용 생 바이러스 백신내의 mycoplasma를 신속하고 민감하게 검출할 수 있을 것으로 판단되었다.

• 방사선산업학회지
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• 제5권3호
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• pp.279-283
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• 2011

This study was compared microbiological safety with gamma-irradiated porcine tendon and skin, as materials for the development of xenografts to regenerate damaged tissues and protect secondary contamination. The porcine tendon and skin were gamma-irradiated after inoculation of bacteria and virus to evaluate irradiation sensitivity of microorganisms. The result showed that the porcine tendon and skin were not different on the sensitivity of microorganisms by gamma irradiation. Bacteria inoculated in the porcine tendon and skin were confirmed that E. coli was the $D_{10}$ values of $0.32{\pm}0.082$ and $0.25{\pm}0.1kGy$ on tendon and skin, and B. subtilis was $4.00{\pm}0.312$ and $3.88{\pm}0.3kGy$ on gamma irradiation, respectively. Moreover, Virus inoculated in the porcine tendon and skin was observed that poliovirus (PV) was $6.26{\pm}0.332$ and $6.88{\pm}0.3kGy$, and porcine parvovirus (PPV) was $1.75{\pm}0.131$ and $1.73{\pm}0.2kGy$ and bovine viral diarrhoea virus (BVDV) was $3.70{\pm}0.212$ and $3.81{\pm}0.2kGy$ on gamma irradiation, respectively. Virus showed higher resistance compared to bacteria on gamma irradiation, but was not detected CPE (cytopathic effect) by virus both tendon and skin at 25 kGy, a standard dose recommended from IAEA for sterilization of medical products. Therefore, These results were considered that gamma irradiation could control effectively bacteria and virus to develop safe porcine xenograft, and apply same irradiation doses to all tissues including tendon and skin of porcine.

• 방사선산업학회지
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• 제5권1호
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• pp.75-80
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• 2011

This study was to determine the microbiological safety and tensile strength of gammairradiated porcine tendon for the development of safe xenografts. Escherichia coli and Bacillus subtilis were used as model pathogens and inoculated as $10^6{\sim}10^7log$ colonies forming unit $(CFU)g^{-1}$. As model virus from porcine, porcine parvovirus (PPV), bovine viral diarrhoea virus (BVDV) and poliovirus were inoculated as $10^5{\sim}10^6$ tissue culture infectious dose $(TCID)_{50}g^{-1}$ into porcine skin. The $D_{10}$ value of E. coli and B. subtilis was measured as $0.32{\pm}0.082kGy$ and $4.0{\pm}0.312kGy$, respectively. Additionally, the $D_{10}$ values of PPV, BVDV and poliovirus were also shown as $1.75{\pm}0.131kGy$, $3.70{\pm}0.212kGy$ and $6.26{\pm}0.332kGy$, respectively. Gamma irradiation decreased the tensile strength of porcine tendon. Results indicate that microbiological safety of porcine tendon can be improved significantly by gamma irradiation. However, further studies are needed to improve the tensile strength of gamma-irradiated porcine tendon.

• Journal of Veterinary Science
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• 제22권6호
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• pp.87.1-87.12
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• 2021

Background: African swine fever virus (ASFV), classical swine fever virus (CSFV), and porcine reproductive and respiratory syndrome virus (PRRSV) are still prevalent in many regions of China. Co-infections make it difficult to distinguish their clinical symptoms and pathological changes. Therefore, a rapid and specific method is needed for the differential detection of these pathogens. Objectives: The aim of this study was to develop a multiplex real-time quantitative reverse transcription polymerase chain reaction (multiplex qRT-PCR) for the simultaneous differential detection of ASFV, CSFV, and PRRSV. Methods: Three pairs of primers and TaqMan probes targeting the ASFV p72 gene, CSFV 5' untranslated region, and PRRSV ORF7 gene were designed. After optimizing the reaction conditions, including the annealing temperature, primer concentration, and probe concentration, multiplex qRT-PCR for simultaneous and differential detection of ASFV, CSFV, and PRRSV was developed. Subsequently, 1,143 clinical samples were detected to verify the practicality of the assay. Results: The multiplex qRT-PCR assay could specifically and simultaneously detect the ASFV, CSFV, and PRRSV with a detection limit of 1.78 × 10⁰ copies for the ASFV, CSFV, and PRRSV, but could not amplify the other major porcine viruses, such as pseudorabies virus, porcine circovirus type 1 (PCV1), PCV2, PCV3, foot-and-mouth disease virus, porcine parvovirus, atypical porcine pestivirus, and Senecavirus A. The assay had good repeatability with coefficients of variation of intra- and inter-assay of less than 1.2%. Finally, the assay was used to detect 1,143 clinical samples to evaluate its practicality in the field. The positive rates of ASFV, CSFV, and PRRSV were 25.63%, 9.36%, and 17.50%, respectively. The co-infection rates of ASFV+CSFV, ASFV+PRRSV, CSFV+PRRSV, and ASFV+CSFV+PRRSV were 2.45%, 2.36%, 1.57%, and 0.17%, respectively. Conclusions: The multiplex qRT-PCR developed in this study could provide a rapid, sensitive, specific diagnostic tool for the simultaneous and differential detection of ASFV, CSFV, and PRRSV.