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从内源性绵羊肺腺瘤反转录病毒（enJSRV）与外源性绵羊肺腺瘤反转录病毒（exJSRV）基因组结构的比较、enJSRV与绵羊肺腺瘤病（OPA）的关系、enJSRV在羊体内的表达及其系统发育分析等方面综述了enJSRV的研究进展，旨在为早期诊断及预防OPA提出新方法，为揭示该病的分子发病机理及探索其基因治疗提供新思路。 相似文献
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为深入研究绵羊肺腺瘤病毒(JSRV)与绵羊肺腺瘤病(OPA)发病关系,本研究采用PCR方法从pGEX-4T-1-TM重组质粒中扩增编码JSRV跨膜蛋白(TM)的基因序列,并引入标签多肽HA序列和限制性内切酶位点,将其重组至真核表达载体pcDNA3.1(+)中,构建了重组质粒pcDNA-TM-HA.通过转染HepG2细胞并用G418筛选,对稳定表达TM的阳性细胞进行纯化,获得了稳定表达JSRV tm基因的HepG2细胞系.间接免疫荧光及western blot检测结果表明,重组蛋白TM-HA在HepG2细胞中得到正确表达.JSRV TM蛋白稳定表达细胞系的建立为进一步研究该蛋白与JSRV诱导OPA发病关系提供了重要的实验平台. 相似文献
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绵羊肺腺瘤病(OPA)是由绵羊肺腺瘤病毒(JSRV)引起的绵羊传染性肺肿瘤疾病。为探究JSRV对绵羊肺脏的致瘤机制,本研究从自然感染JSRV的羊(OPA患羊)肺肿瘤组织和健康羊肺组织中提取总RNA,构建二者的cDNA文库后采用Illumina Hi Seq 4000高通量测序平台进行转录组学测序(RNA-Seq),采用DESeqR筛选健康绵羊与患病绵羊肺组织中的转录差异基因,并以P<0.05和log2(Fold change)≥1筛选转录显著差异基因。通过GO和KEGG数据库对转录显著差异基因进行GO功能和KEGG信号通路的富集分析,并采用RT-qPCR对随机选择的10个转录显著差异基因进行验证。结果显示,与对照组相比,OPA羊肺肿瘤组织中共筛选到1 360个转录上调基因和783个转录下调基因,其中154个转录显著上调基因,212个转录显著下调基因。GO功能分析显示,转录显著差异基因显著富集在178个GO条目中,包括114个生物过程(BP)、19个细胞成分(CC)和45个分子功能(MF),主要涉及生长因子活性、复制后修复、NAD+二磷酸酶活性、核苷代谢过程和... 相似文献
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绵羊肺腺瘤病研究进展 总被引：15，自引：1，他引：14
绵羊肺腺瘤病 ( OPA)是由外源性的 D型和 B型绵羊肺腺瘤反转录病毒 ( JSRV)引起的肺脏肿瘤性疾病 ,该病毒主要在肺肿瘤上皮细胞内高水平表达 ,病毒基因组内 L TR的活化和病毒囊膜蛋白的表达是引起体外细胞转化癌变的主要因素 ,其致瘤的可能机制是通过激活磷酯酰肌醇 3 -激酶 ( PI-3 K)和蛋白酶 K( AKT)信号通路来启动细胞内抑制细胞凋亡和刺激细胞增生的信号转导系统 ,从而引起细胞增生癌变。该病与人的细支气管 -肺泡癌 ( BAC)极相似 ,通过对 OPA的研究 ,不仅为早期诊断及预防该病提出新方法 ,而且为揭示 BAC的分子发病机制及探索其基因治疗提供新思路。 相似文献
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绵羊基因组中内源性绵羊肺腺瘤病毒相关序列的确定 总被引：1，自引：0，他引：1
绵羊都含有与绵羊肺腺瘤病毒(Jaagsiekte sheep retrovirus,JSRV)密切相关的15~20拷贝内源性绵羊肺腺瘤病毒(enJSRV)相关序列。宿主可利用内源性病毒来预防致病性反转录病毒的感染,一些内源性病毒可以有效地干扰相关外源性病毒的复制。本试验通过分子生物学手段确定了蒙古绵羊基因组中含有enJSRV6和enJSRV10两个内源性病毒基因而内蒙古白绒山羊中未发现。通过比较内、外源病毒LTR序列的酶切图谱,获得专一作用外源性病毒的核酸内切酶M spⅠ、TfiⅠ、BsaWⅠ,如果将酶切与聚合酶链式反应(PCR)相结合,不需要经过测序就可分辨enJS-RV和外源性绵羊肺腺瘤病毒(exJSRV),形成"酶切-PCR"检测技术,将为绵羊肺腺瘤病的快速诊断提供了新的手段。 相似文献
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绵羊肺腺瘤病毒内蒙古株的分离与观察 总被引：1，自引：0，他引：1
以内蒙古某羊场自然感染绵羊肺腺瘤病的病肺组织4例和3月龄绵羊胎肺2例作为材料，对胎肺细胞进行了原代培养并接种绵羊肺腺瘤病毒悬液，进行了绵羊肺腺瘤病毒(JSRV)的分离，同时采用电镜负染色技术和超薄切片透射电镜技术对上述样品进行了观察。结果表明，自然感染绵羊肺腺瘤病的4例病肺组织中都有散在的病毒粒子，其直径为100～125nm，接种病毒悬液的胎肺细胞从第2代到第9代均出现细胞病变，但对绵羊胎肺培养的病毒悬液电镜负染色观察未发现典型的病毒粒子，而超薄切片透射电镜观察发现了病毒样粒子。 相似文献
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为建立检测绵羊肺腺瘤病毒(Jaagsiekte sheep retrovirus,JSRV)实时荧光定量PCR(Real-time qPCR)方法,根据外源性JSRV-NM株env基因序列,选其保守序列作为目的片段,设计引物和TaqMan探针,以自然病例的肺肿瘤组织基因组DNA为模板,经PCR扩增目的基因、克隆,重组质粒鉴定,并严格定量后,梯度稀释作为阳性标准品,优化反应条件进行Real-time qPCR扩增,获得的标准曲线为:Y=-3.308X+47.848,线性相关系数为0.991;Ct值变异系数小,并且灵敏度高,初步建立了检测JSRV前病毒DNA的Real-time qPCR方法。应用该方法对不同来源(A、B、C、D、E组)的绵羊外周血及其他组织样品进行测定其前病毒载量。结果显示B组和C组外周血白细胞、肺脏、肺门淋巴结以及鼻液中检测均为阳性,并发现前病毒DNA的载量在肺脏中明显高于外周血白细胞;D组虽未发现有绵羊肺腺瘤(SPA)临床症状,但在肺门淋巴结里可以检测到;E组中1只绵羊的肺脏也检出低拷贝数的前病毒DNA,而在A组中检测结果均为阴性。本研究对检测未知羊群JSRV感染程度及研究SPA流行病学等均有重要意义。 相似文献
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为建立绵羊肺腺瘤病毒(JSRV)实时荧光定量PCR检测方法,本研究根据JSRV的ag基因,选取其保守序列作为检测目的片段,设计并合成相应的引物和TaqMan探针,以自然病例的肺肿瘤组织基因组DNA为模板,经PCR扩增目的基因构建重组质粒pMD-gag,并将其作为阳性标准品,梯度稀释建立标准曲线,得到扩增方程为:y=-0.304x+38.4,扩增效率为100％,线性相关系数为0.999,该方法最低检出量为103拷贝,与其它病原核酸样品无交叉反应,其变异系数在1.374％以内.本研究为快速检测JSRV以及组装检测试剂盒奠定了基础. 相似文献
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De Las Heras M Ortín A Salvatori D Pérez de Villareal M Cousens C Miguel Ferrer L Miguel Cebrián L García de Jalón JA Gonzalez L Michael Sharp J 《Research in veterinary science》2005,79(3):201-264
Ovine pulmonary adenocarcinoma (OPA) is a naturally occurring contagious lung neoplasia caused by jaagsiekte sheep retrovirus (JSRV). Although no specific circulating antibodies against the virus can be detected in infected sheep, JSRV proviral DNA sequences can be found in peripheral blood leukocytes (PBLs) in clinically affected and in a proportion of in contact animals. In this study, existing hemi-nested PCR procedure is compared with a new one-step PCR technique that was developed to minimise potential DNA contamination and reduce sample and reagent handling. Different blood preparations were assessed and the best results were achieved on DNA prepared from buffy coat. The sensitivity of this PCR was lower in JSRV infected sheep without lesions of OPA than in clinically affected sheep, which indicate that this PCR may not be not fully appropriate for screening of individual sheep, but rather to provide results at flock level. This PCR is the only currently available blood test for detection of JSRV infected sheep and may be useful in epidemiological studies and in control programmes of OPA. 相似文献
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Marta Borobia Aurora Ortín Luis M. Ferrer Juán J. Ramos Delia Lacasta Marcelo De las Heras 《Canadian journal of veterinary research》2014,78(3):237-240
Ovine pulmonary adenocarcinoma (OPA) is a transmissible lung cancer caused by Jaggsiekte sheep retrovirus (JSRV). It is difficult to identify animals infected with JSRV but are clinically healthy. The virus does not induce a specific antibody response and, although proviral DNA sequences of JSRV can be found in mononuclear blood cells, the detection is inconsistent. The aim of this study was to investigate the presence of JSRV in the bone marrow of infected sheep and develop a more consistent screening method. Immunohistochemical examination of bone marrow samples from 8 asymptomatic JSRV-infected sheep revealed the presence of positively labelled cells. However, JSRV could not be detected by a highly sensitive polymerase chain reaction (PCR) in bone marrow aspirates periodically collected from these animals. Results suggest that JSRV-infected cells may be present in the bone marrow of symptomless animals, but the number is below the detectable level for PCR. Therefore, this technique does not seem to be helpful for preclinical diagnosis of OPA. 相似文献
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Christina Cousens Leenadevi Thonur Joanne Crawford David J. Griffiths 《Research in veterinary science》2009,87(1):154-156
Jaagsiekte sheep retrovirus (JSRV) causes a fatal lung cancer of sheep known as ovine pulmonary adenocarcinoma (OPA). OPA is a significant disease in many sheep-rearing countries and there is no effective method of control. A unique feature of OPA is the overproduction of fluid in the lung of affected animals. This lung fluid contains JSRV and provides a means of transmission through the inhalation of virus. In this study we demonstrated that lung fluid from different OPA cases contained between 107 and 1010 copies of JSRV RNA per ml. Examination of JSRV RNA survival under conditions that mimic natural conditions suggested that intact JSRV virions may persist for several weeks in the environment. These are the first quantitative data on JSRV in lung fluid and provide valuable information for implementing appropriate biosecurity measures to control the spread of JSRV in the field. 相似文献
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目的随着加入世界贸易组织,我国海关检疫压力增大,为了维护国家主权和国际声誉,加强出入境检验检疫势在必行。衣原体病是一种重要的人畜共患病,也给养禽业带来巨大的经济损失,必须给予足够的重视,鹦鹉热衣原体对多种动物宿主有广泛的寄生性引起不同动物的多症候群疾病,是世界动物卫生组织动物名录疾病。方法对入境的一批肉种鸡进行抽样检验,按照中华人民共和国出入境检验检疫行业标准用间接血凝检测鹦鹉热衣原体病,做出试验报告。结果经检测出现阳性,肉鸡携带有衣原体病毒。结论根据中华人民共和国进出境动植物检疫法第二章进境检疫第十六条检出二类传染病、寄生虫病的动物,退回或者扑杀,同群其他动物在隔离场或者其他指定地点隔离观察。 相似文献
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Grego E De Meneghi D Alvarez V Benito AA Minguijón E Ortín A Mattoni M Moreno B Pérez de Villarreal M Alberti A Capucchio MT Caporale M Juste R Rosati S De las Heras M 《Veterinary microbiology》2008,130(3-4):247-257
Ovine pulmonary adenocarcinoma (OPA) is a contagious disease caused by jaagsiekte sheep retrovirus (JSRV). In the three studies performed, we have obtained data of the importance of colostrum/milk (C/M) in the transmission of JSRV. In the first study, a group of sheep from a flock with a long history of OPA, samples from colostrum and peripheral blood leucocytes (PBLs) were collected. Two specific PCRs (U3-LTR and env of the JSRV) were carried out. Using U3PCR 8/34 sheep were positive in colostrum whereas with envPCR 7/34 were positive. From these animals only one was positive with U3PCR in the PBLs. Evidence of the transmission of JSRV infection by C/M was obtained in two more separate studies. In the second study, PBLs from five lambs from JSRV+ ewes and two from JSRV-ewes were tested by the U3PCR. They were fed C/M by their mothers during 3 months and slaughtered 7 months after birth. Three out of five lambs from the JSRV+ sheep become PBL positive at 3-4 months old and the other two were also positive at 4-6 months of age. One lamb of the JSRV-sheep became also PBL positive at an age of 3 months. In the third study, a group of lambs from JSRV negative mothers were fed with C/M from JSRV+ sheep and housed in separate unit. For comparison, another group of the same origin and maintained in another different unit, were fed with C/M containing a JSRV virus preparation. All lambs were blood sampled monthly and JSRV infection was detected as early as 15 days and several times onwards in both groups. Control groups fed with C/M from JSRV free flock and JSRV blood test negative sheep were always negative. Together these results indicate that suckling is an important natural transmission route for JSRV. 相似文献
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Voigt K Brügmann M Huber K Dewar P Cousens C Hall M Sharp JM Ganter M 《Research in veterinary science》2007,83(3):419-427
Ovine pulmonary adenocarcinoma (OPA) is a contagious lung tumour of sheep caused by Jaagsiekte sheep retrovirus (JSRV). The disease is a particular problem in flocks in many parts of the world. The aim of the study was to assess screening methods for individual animals as a prelude to future eradication trials. Results of histological examination were used as the standard to evaluate the relative sensitivity and specificity of an established heminested polymerase chain reaction (PCR) test for JSRV proviral DNA from blood and bronchoalveolar lavage (BAL) samples. PCR results from tissue samples are included as control data. PCR testing of blood samples was found to have an estimated sensitivity of only 10% (95% confidence interval (CI) 3-20) while the sensitivity of the PCR test on BAL samples was 89% (CI 79-96) in comparison to the results of histological examination. We conclude that PCR testing of BAL samples is an effective confirmatory test for sheep with suspected clinical OPA. It is also a useful tool for the pre-clinical identification of individual infected sheep within an infected flock and therefore may prove beneficial in future control or eradication programmes. 相似文献
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Ovine pulmonary adenocarcinoma (OPA) is a naturally occurring retrovirus-induced transmissible lung cancer in sheep. Lungs
and associated (bronchial and mediastinal) lymph nodes of seven sheep with OPA were examined. Lungs had few multifocal consolidated
slightly elevated gray to white masses ranging from 0.5 to 3 cm in diameter. Histopathologically, these masses appeared as
well-differentiated acinar adenocarcinoma with little evidence of anaplasia. The acini composed of well-differentiated cuboidal
to low columnar epithelium with clear or vacuolated cytoplasm and low mitotic index. No metastases were observed in the bronchial
and mediastinal lymph nodes of any animal. The presence of Jaagsiekte sheep retrovirus (JSRV) was demonstrated in the lungs by immunohistochemistry. JSRV protein was detected in all tumor epithelial cells, histologically
normal alveolar type II cells, and few bronchiolar epithelial cells, alveolar macrophages, lymphocytes, and plasma cells.
This study is the first to confirm the presence of natural OPA in Egypt. 相似文献
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牛结核病检疫方法研究进展 总被引：3，自引：2，他引：1
牛结核病主要是由牛分枝杆菌引起的一种慢性消耗性疾病,在许多国家尤其是发展中国家仍然广泛流行,该病不仅给畜牧业造成巨大的经济损失,而且严重威胁着人类的健康。因此,牛结核病的检疫意义重大。细菌学检测、免疫学检测及分子生物学诊断为牛结核病检疫的研究奠定了基础。然而,目前迫切需要研究快速、灵敏、特异的牛结核病诊断方法。牛结核病血清学诊断技术研究一直是研究热点之一,但至今仍存在许多问题。作者就这些诊断检疫方法进展作一简要介绍。 相似文献
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Summers C Norval M De Las Heras M Gonzalez L Sharp JM Woods GM 《Veterinary immunology and immunopathology》2005,106(3-4):285-294
Infection with a retrovirus, Jaagsiekte sheep retrovirus (JSRV), causes ovine pulmonary adenocarcinoma (OPA). The excess production of surfactant proteins by alveolar tumour cells results in increased production of pulmonary fluid, which is characteristically expelled through the nostrils of affected sheep. The immune response to JSRV and the tumour is poorly understood: no JSRV-specific circulating antibodies or T cells have been detected to date. The aim of the present study was to obtain phenotypic evidence for a local immune response in OPA lungs. Specific-pathogen free lambs were infected intratracheally with JSRV. When clinical signs of OPA were apparent, the lungs were removed at necropsy and immunohistochemistry (IHC) was performed on lung sections using a panel of mouse anti-sheep mAbs. No influx of dendritic cells, B cells, CD4, CD8 or gammadelta T cells was seen in the neoplastic nodules or in their periphery. MHC Class II-positive cells were found intratumourally, peritumourally and in the surrounding alveolar lumina. In the tumours, many of these cells were shown to be fibroblasts and the remainder were likely to be mature macrophages. In the alveolar lumen, the MHC Class II-positive cells were CD14-positive and expressed high levels of IFN-gamma. They appeared to be immature monocytes or macrophages which then differentiated to become CD14-negative as they reached the periphery of the tumours. A high level of MHC Class I expression was detected on a range of cells in the OPA lungs but the tumour nodules themselves contained no MHC Class I-positive cells. On the basis of these findings, it is proposed that the lack of an effective immune response in OPA could result from a mechanism of peripheral tolerance in which the activity of the invading macrophages is suppressed by the local environment, possibly as a consequence of the inhibitory properties of the surfactant proteins. 相似文献