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1.
5种脑炎人兽共患病病毒多重RT-PCR检测方法的建立 总被引：3，自引：0，他引：3
为建立同时检测流行性乙型脑炎病毒(JEV)、森林脑炎病毒(TBEV)、东方马脑炎病毒(EEEV)、西方马脑炎病毒(WEEV)和基孔肯雅病毒(CHIKV)5种人兽共患脑炎病病毒的多重RT-PCR方法,本研究根据GenBank登录的相关病毒基因序列设计特异引物,通过优化引物组合及PCR反应条件,建立可同时检测5种病毒的方法,扩增片段长度分别为411 bp(JEV)、945 bp(TBEV)、193 bp(EEEV)、545 bp(WEEV)和769 bp(CHIKV);该方法具有良好的特异性,对病毒核酸最低检测拷贝数分别为7.1×103、3.6×103、2.2×103、5.6×103和5.1×103.该方法具有特异性强、灵敏度高、操作简便等优点,为以上5种人兽共患脑炎病病毒提供快速检测手段. 相似文献
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秦红刚肖敏李伟谢红玲漆世华温文生吴玉石 《中国兽药杂志》2012,46(7):58-61
猪乙型脑炎是一种嗜神经性虫媒病毒引起的人兽共患传染病。疫苗的免疫接种是控制猪乙型脑炎发生的主要手段。阐述了猪乙型脑炎常规疫苗和新型疫苗的研究进展,为进一步研究猪乙型脑炎疫苗提供参考。 相似文献
3.
云南省乙型脑炎病毒宿主和媒介研究 总被引：18，自引：0，他引：18
于１９７８年至１９９７年,从云南猪血液和乳猪脑组织中分离到乙型脑炎（ＪＥ）病毒５株,从蝙蝠脑组织中分得ＪＥ病毒１０株,从鸟类脑组织中分得ＪＥ病毒５株,从１５种蚊虫体内分离到ＪＥ病毒６３株,从２种蠓类中分得ＪＥ病毒２株。分析认为,猪是ＪＥ病毒的主要扩散宿主,蝙蝠和鸟类在该病毒保存和传播中起重要作用;三带喙库蚊是该病毒的主要传播媒介,伪杂鳞库蚊、霜背库蚊、蚊腿库蚊、白蚊伊和刺扰伊蚊为重要媒介,蠓类亦可 相似文献
4.
狐狸脑炎病毒细胞培养灭活疫苗的研究 总被引：1，自引：0，他引：1
采用分离到的狐狸脑炎病毒ＦＥＶ－Ｈ作种毒，经ＭＤＣＫ细胞培养，经最终含量为０．２％的甲醛灭活，制备狐狸脑炎病毒细胞培养灭活疫苗。皮下接种疫苗１０ｍｌ，试验狐，貉，犬均无临床特异反应，局部吸收良好。最小免疫剂量为１６倍稀释的疫苗原液１ｍｌ，使用剂量定为４倍稀释的疫苗原液１ｍｌ，试验狐狸免疫后３０天，血清中和抗体效价达到高峰值１：１４５，能耐受张毒攻击。免疫后６个月，抗体水平降至１：３２，但此时仍能耐 相似文献
5.
对伪狂犬病病毒（PRV）、猪细小病毒（PPV）和流行性乙型脑炎病毒（JEV）检测基因芯片的制备及该芯片的检测技术进行了研究。选定靶基因最佳点样质量浓度为200 mg/L,用基因芯片点样仪将其点制在氨基化基片上,经干燥、水合、紫外线交联和洗涤后,成功制备了PRV-PPV-JEV检测基因芯片。以CY3荧光素标记的dCTP经PCR扩增制备探针,对芯片的质量进行了评价。结果表明,制备的芯片质量好,探针最佳使用质量浓度为3 000μg/L,芯片系统检测灵敏度可达3μg/L。该芯片可同时检测PRV、PPV和JEV,其灵敏度高、特异性强,芯片可重复使用,室温下至少可保存4个月。 相似文献
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检测CSFV、JEV、PRRSV三种RNA病毒多重RT-PCR方法的建立 总被引：1，自引：0，他引：1
猪瘟病毒(CSFV)、流行性乙型脑炎病毒(JEV)和猪繁殖与呼吸综合征病毒(PRRSV)是引起严重的种猪繁殖障碍的病原,而且经常混合感染,及时准确诊断是防治的前提。根据GenBank发表序列选取3对引物建立检测CSFV、JEV和PRRSV病毒的多重RT-PCR方法,扩增产物分别为508 bp、380 bp、263 bp。经与IDEXX商品化的检测CSW抗原试剂盒比较,二者的符合率为96.7%;扩增JEV和PRRSV PCR产物分别经EcoR V和Sau3A I酶切得到预期的片段。建立的多重RT-PCR检测JEV、PRRSV和CSFV敏感度分别为12.5个TCID_(50)、10个TCID_(50)和10~(-3)ng总RNA。结果表明该多重RT-PCR方法具有很好的特异性和敏感性,可用于临床三种病毒核酸的检测。 相似文献
8.
本研究以乙型脑炎病毒SA14—14—2疫苗株基因组RNA为模板，采用RT—PCR一步法扩增了PrM基因的全长cDNA（558 bp），用BamHI和EcoRI双酶切PrM基因的扩增产物，回收目的基因后将其克隆至经同样酶切的伪狂犬病病毒通用转移载体pPgG-uni中，获得了转移载体pPgG-PrM，并对其外源片段进行了测序。序列分析结果表明：与已报道的乙型脑炎病毒SA14强毒株和SA14-14-2疫苗株的核苷酸序列比较，PrM基因的同源性为100％。以伪狂犬病病毒Ea株TK^-／gG^-／LacZ^＋突变株为载体构建了一株表达乙型脑炎病毒PrM基因的重组伪狂犬病病毒TK^-／gG^-／PrM^＋，为进一步开展猪乙型脑炎与伪狂犬病二价基因工程疫苗的研究奠定了基础。 相似文献
9.
提取猪日本脑炎病毒(JEV)上海分离株的基因组RNA,反转录合成cDNA,RT-PCR扩增JEV的NS3基因片段,亚克隆到原核表达载体pET-28(a)上,酶切鉴定和PCR鉴定重组原核表达质粒pET-28(a)-JEV-NS3,转化大肠埃希菌BL21(DE3)菌株,IPTG诱导表达重组His-JEV-NS3蛋白,SDS-PAGE分析重组蛋白的表达,his-band试剂盒纯化重组蛋白。获得了1.8kb NS3基因片段,成功构建重组原核表达质粒pET-28(a)-JEV-NS3。IPTG诱导获得分子质量为64ku的His-JEV-NS3蛋白,该蛋白主要以包涵体形式表达,表达量占总菌体蛋白的30%以上,纯化后获得高纯度重组蛋白,其占总蛋白比例达70%以上。 相似文献
10.
日本乙型脑炎病毒的分离与鉴定 总被引：4，自引：1，他引：3
收集一例疑为乙型脑炎病毒感染的种公猪肿大的睾丸病料,并将其处理制成匀浆过滤后,用乳鼠脑内接毒和细胞接毒相结合的方法盲传并分离病毒,然后设计一对PrM/E基因的特异性引物,采用RT-PCR方法对所分离的病毒进行鉴定,并将其PrM/E基因扩增产物测序,测序结果与乙型脑炎GenBank登陆的SA14-14-2株(AF315119)和SA14株(U14163)进行比较。结果表明,所分离病毒的PrM/E基因序列与JEV强毒株SA14和弱毒株SA14-14-2相应序列同源性分别为98.1%和97.1%,证实分离毒株为乙型脑炎病毒。 相似文献
11.
猪日本脑炎病毒RT-PCR检测方法的建立 总被引：2，自引：0，他引：2
设计了1对引物,利用RT-PCR技术检测乙型脑炎病毒(JEV)。从GenBank中查出收录的31株猪乙型脑炎病毒E基因的已知序列,用DNA star软件对这31株猪乙型脑炎病毒E基因进行同源性分析,以确定扩增的靶序列。以这段靶区域为模板,利用Primer 5软件设计了1对引物,用减毒株SA14-14-2建立了检测乙脑病毒的RT-PCR方法,经敏感性,特异性试验测定,证明该方法敏感,特异;该法可检出样品稀释至256倍的鼠脑毒,相当于0.06个TCID50,对4株河北地区JEV分离株进行检测,结果所设计引物对4株病毒均能扩增出预期的片段。 相似文献
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K. Ergünay M. B. Saygan S. Aydoğan N. Litzba B. Şener S. Lederer M. Niedrig G. Hasçelik D. Us 《Zoonoses and public health》2011,58(3):220-227
Tick‐borne encephalitis virus (TBEV) is the aetiological agent of tick‐borne encephalitis (TBE), a potentially fatal central nervous system infection of humans. TBE is endemic in many areas of Europe and Asia; however, very scarce data on TBEV activity are available from Turkey. We aimed to identify TBEV exposure in healthy blood donors and the impact of TBEV in central nervous system infections in Central/Northern Anatolia. Two‐thousand four hundred and fifty four sera, collected from blood donors at Ankara, Konya, Eski?ehir and Zonguldak branches of the Turkish Red Crescent Middle Anatolia Regional Blood Center, were analysed for TBEV serosurveillance. Paired serum and cerebrospinal fluid samples from 108 patients with the diagnosis of aseptic meningitis/encephalitis of unknown aetiology were also evaluated to identify TBE and neuroborreliosis cases. Commercial enzyme‐linked immunosorbent assays and indirect immunofluorescence tests were employed for antibody detection. Forty‐seven donor samples (1.9%) were reactive for TBEV IgG. In 25 persons with IgG reactivity (53.1%), risk factors for tick‐borne infections were revealed. One sample from Zonguldak province (1/198; 0.5%) in the Black Sea region of Turkey was confirmed to possess neutralizing antibodies via plaque reduction neutralization test. TBEV IgM was detected in 9.2% (8/108) of the patients. IgM was accompanied by IgG reactivity in two persons where, in one, recent history of a tick bite was also identified. Intrathecal antibody production for TBEV could not be demonstrated. No evidence for Borrelia infections could be found. Confirmed exposure to TBEV and/or an antigenically similar tick‐borne flavivirus is documented for the first time in blood donors in Zonguldak in Northern Anatolia. Probable cases of TBE have also been identified from Central Anatolia. The epidemiology of TBEV activity in Turkey needs to be assessed and benefits of vaccination for general population, risk groups or travellers must be considered. 相似文献
13.
乙型脑炎病毒NS1基因重组伪狂犬病毒的构建 总被引：7，自引：0，他引：7
设计1对引物从含有乙型脑炎病毒NS1基因的质粒pNS1上亚克隆NS1基因,将NS1基因插入到中间转移载体pUSK中，获得重组中间转移质粒pUSK—NS1。将pUSK—NS1与伪狂犬病毒Ea株TK／gG／LacZ^ 突变株基因组共转染真核细胞IBRS-2，通过空斑纯化得到了乙型脑炎病毒NS1基因重组伪狂犬病毒株TK／gG^-／NS^ 1。经检测,重组病毒能表达具有生物活性的NS1蛋白。该重组病毒可作为猪乙型脑炎和伪狂犬病双价基因工程疫苗用毒株。 相似文献
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Dong-Kun Yang Yoon-I Oh Hye-Ryoung Kim Youn-Jeong Lee Oun-Kyong Moon Hachung Yoon Byounghan Kim Kyung-Woo Lee Jae-Young Song 《Journal of veterinary science (Suw?n-si, Korea)》2011,12(4):373-377
Climate change induced by recent global warming may have a significant impact on vector-borne and zoonotic diseases. For example, the distribution of Japanese encephalitis virus (JEV) has expanded into new regions. We surveyed the levels of hemagglutination-inhibition (HI) antibodies against JEV (Family Flaviviridae, genus Flavivirus) in wild birds captured in Korea. Blood samples were collected from 1,316 wild birds including the following migratory birds: Oceanodroma castro (n = 4), Anas formosa (n = 7), Anas penelope (n = 20), Fulica atra (n = 30), Anas acuta (n = 89), Anas crecca (n = 154), Anas platyrhynchos (n = 214), Aix galericulata (n = 310), and Anas poecilorhyncha (n = 488). All were captured in 16 locations in several Korea provinces between April 2007 and December 2009. Out of the 1,316 serum samples tested, 1,141 (86.7%) were positive for JEV. Wild birds captured in 2009 had a higher seroprevalence of ant-JEV antibodies than those captured in 2007. Wild birds with an HI antibody titer of 1 : 1,280 or higher accounted for 21.2% (280/1,316) of the animals tested. These findings indicated that wild birds from the region examined in our study have been exposed to JEV and may pose a high risk for introducing a new JEV genotype into Korea. 相似文献
16.
Hao Zheng Tongling Shan Yu Deng Chunqing Sun Shishan Yuan Yang Yin Guangzhi Tong 《Journal of veterinary science (Suw?n-si, Korea)》2013,14(1):27-36
Japanese encephalitis virus (JEV) is the leading cause of viral encephalitis in Asia and domestic pigs serve as the amplifying hosts. In the present study, the full genomic sequences of two JEV strains (HEN0701 and SH0601) isolated from pigs in China were determined and compared with other 12 JEV strains deposited in GenBank. These two strains had an 88.8% nucleotide sequence similarity and 97.9% deduced amino acid sequence homology. HEN0701 had high nucleotide sequence and high amino acid sequence identity with genotype I (GI) strains, while SH0601 had high nucleotide sequence and high amino acid sequence identity with GIII strains at both the gene and full genome levels. Further phylogenetic analysis showed that HEN0701 belonged to the JEV GI group and SH0601 was classified as a GIII strain. Analysis of codon usage showed there were a few differences between the GI and GIII strains in nucleotide composition and codon usage for the open reading frames. 相似文献
17.
JEV分子生物学与新型疫苗研究进展 总被引：1，自引：0，他引：1
流行性乙型脑炎病毒（Japanese encephalitis virus,JEV)是一种严惩危害人畜健康的虫媒病毒。本文对JEV的基因组结构、结构蛋白、非结构蛋白及其功能、重组活疫苗和核酸疫苗的研究现状以及与常规疫苗相比所具有的优缺点等方面进行了较为详尽的综述，并且提出JEV复制的确切机制及其蛋白尤其是非结构蛋白的结构与功能尚需深入研究，以期为坦步改良JE新型疫苗打下基础。 相似文献
18.
为鉴定乙型脑炎病毒(JEV)E蛋白I,II结构域抗原表位,本研究设计了一系列针对E蛋白的I、II结构域(1aa～296aa)部分重叠短肽,分别进行GST融合表达,用JEV阳性血清进行western blot和ELISA反应性筛选,获得了5个线性抗原表位分别为:E1(1FNCLGMGNRDFIEGAS16)、E10-4(77TGEAHNEK84)、E11-2(83EKRADSS YVCKQ94)、E19(145GTTTSENHGNYSAQVG160)和E33(257SQEGGLHHALAGAIVV272)。除E10和E19外,其它表位均为第一次通过实验方法确定的。将这5个融合蛋白对小鼠进行免疫,获得了高滴度的针对5个融合短肽的抗血清,研究证明了这些表位具有良好的免疫原性。本研究为乙型脑炎特异性诊断试剂及表位疫苗的开发奠定了基础。 相似文献
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Chutchai Piewbang Sabrina Wahyu Wardhani Surangkanang Chaiyasak Jakarwan Yostawonkul Tanit Kasantikul Somporn Techangamsuwan 《Zoonoses and public health》2022,69(1):55-60
Japanese encephalitis virus (JEV) infection has been recognized as a serious disease in humans. Wildlife animal infections due to JEV have not been well described. This study identified JEV infection in two deceased meerkats in Thailand, with clinical signs of neurological disease. Histopathology of brains revealed severe lymphoplasmacytic necrotizing meningoencephalitis, while similar inflammation was observed in the lung and liver. Partial JEV sequences were identified from the formalin-fixed paraffin-embedded-derived brain sections of two meerkats and were found to be genetically similar to a JEV strain detected in China but not from a local strain. Using immunohistochemistry, the virus was identified in neurons and glial cells, and also found in bronchial glands, Kupffer's cells in liver, lymphocytes in the spleen and pancreatic acini, which suggests extraneural infection. Transmission electron microscopy confirmed the presence of spheroid viral particles in the lungs. These findings may suggest that infection of extraneural organs in meerkats is similar to that described in JEV-infected humans. In conclusion, this study identified the first JEV infection in meerkats as an interesting case study. The JEV should be considered as an important differential diagnosis in meerkats with encephalitis. Further surveillance on JEV infection in meerkats and other wildlife species in a large cohort is needed in the future study. 相似文献
20.
Baldev R. Gulati Harisankar Singha Birendra K. Singh Nitin Virmani Sandip K. Khurana Raj K. Singh 《Journal of veterinary science (Suw?n-si, Korea)》2011,12(4):341-345
The seroprevalence of Japanese encephalitis virus (JEV) among equines was evaluated from January 2006 to December 2009 in 13 different states of India by hemagglutination inhibition (HI) test and virus neutralization test (VNT). Antibodies against JEV were detected in 327 out of 3,286 (10%) equines with a maximum prevalence reported in the state of Manipur (91.7%) followed by Gujarat (18.5%), Madhya Pradesh (14.4%), and Uttar Pradesh (11.6%). Evidence of JEV infection was observed in equines in Indore (Madhya Pradesh) where a 4-fold or higher rise in antibody titer was observed in 21 out of 34 horses in November 2007 to October 2006. In March 2008, seven of these horses had a subsequent 4-fold rise in JEV antibody titers while this titer decreased in nine animals. JEV-positive horse sera had a JEV/WNV (West Nile virus) ratio over 2.0 according to the HI and/or VNT. These results indicated that JEV is endemic among equines in India. 相似文献