博尔纳病病毒感染致病机制的研究进展

博尔纳病病毒是一种新发神经系统感染病毒,一种非细胞溶解的、单股负链的RNA病毒,是单股负链RNA病毒目博尔纳病毒科的典型代表,有严格的嗜神经性,呈低产量、非溶细胞性复制.感染范围从马、牛,羊、鸟、灵长类等剑人类,造成博尔纳病(Borna disease,BD),一种致死性中枢神经疾病,临床上以精神、行为异常为主要表现.自然感染以马和羊为主,人类的BDV感染可能与部分神经精神疾病的发病有关.其致病机制尚不清楚,给博尔纳病的流行病学调查和诊断、治疗带来很大的影响,现就博尔纳病病毒感染机制综述如下.     

精神分裂症患者血液标本检测博尔纳病病毒核酸

博尔纳病病毒（Boma disease vires，BDV）是一种高度嗜神经的RNA病毒。近年研究发现BDV可能与精神分裂症（schizophrenia，SCH）的发病有关，但对此存在较多争论。本研究通过检测SCH患者外周血单个核细胞（PBMC）中BDV p24核苷酸片段来探讨BDV感染与SCH发病的关系。     

用蛋白印迹试验检测精神分裂症患者血清中博尔纳病病毒-p24抗体的研究

目的：探讨博尔纳病病毒（BDV）感染在我国的流行情况及其与精神分裂症的相关性。方法：在对优化表达的GST－BDV－p24融合蛋白进行特异性鉴定的基础上，通过确定融合蛋白与第一抗体、第二抗体间的最佳反应条件，建立可行的检测BDV－p24特异抗体的蛋白印迹（Western-blot）方法，进而对黑龙江地区精神分裂症患者和正常人对照血清中BDV－p24特异性抗体进行了检测，并通过血清－GST蛋白吸收后W estern-blot实验对阳性血清进行了确认。结果：116例精神分裂症患者中检出BDV－p24阳性血清10例，阳性检出率为8.6%，而正常人血清标本中未检出阳性。结论：我国存在博尔纳病病毒的感染，博尔纳病病毒的感染可能与精神分裂症的发生有关。     

临床病毒性脑炎标本的博尔纳病病毒核酸检测

目的 探讨临床病毒性脑炎(viral encephalitis,VE)患者与博尔纳病病毒(Borna disease virus,BDV)感染的关系,分析BDV感染的病毒性脑炎患者临床特征.方法 用荧光定量巢式逆转录聚合酶链反应(FQ-nRT-PCR)方法检测病毒性脑炎患者及非感染性疾病施行蛛网膜下腔阻滞麻醉的手术患者脑脊液单个核细胞(cerebrospinal fluid mononuclear cell,CSFMC)中BDV p24基因片段,同时用β-肌动蛋白(β-actin)作为内参照,脑脊液(CSF)阳性标本基因测序分析并总结出临床特征.结果 32例病毒性脑炎脑脊液标本BDV p24基因片段检出率为12.5%(4/32),拷贝数＞102/μl.对其中一份CSF阳性标本测序后,与BDV标准病毒株Ⅴ和马源的BDV病毒株H1766序列比较同源性分别为95.35%和98.84%.在4个位点出现基因突变(nt1649 T→C、nt1656 G→A、nt1670 C→T、nt1676 C→T).该目的基因片段与马源的BDV病毒株亲缘关系最近.阳性脑炎患者主要以精神行为异常为临床特征.结论 贵州省遵义市部分病毒性脑炎的发生与BDV感染有关,主要以精神行为异常为临床特征.     

多发性硬化患者博尔纳病病毒p24基因片段的检测

博尔纳病病毒(Boma disease virus,BDV)是引发神经系统感染性疾病病原体的一种.我国重庆、哈尔滨、贵州等地区在病毒性脑炎、抑郁症、精神分裂症、帕金森病等部分疾病患者血液和脑脊液中检出过BDV,但多发性硬化(multiplesclerosis,MS)患者中未见报道.我们在13例MS患者外周血单个核细胞(PBMC)中检测BDV-p24基因片段,2例阳性,报道如下.     

博尔纳病病毒ORFⅠ基因中部片段的检测

目的　通过对中国神经精神病人外周血单个核细胞 (PBMC)中博尔纳病病毒 (BDV)ORFⅠ中部片段的检测 ,了解BDVORFⅠ基因中部片段是否为检测BDV感染敏感的标记物。方法　用套式逆转录聚合酶链反应 (nestedRT PCR)同时检测 60例BDV感染阳性或阴性的神经精神病人以及 60例健康献血者的PBMC中BDVORFⅠ (P40 )基因中部片段。结果　 4例BDV感染阳性的神经精神病人的检测结果均为阳性。结论　该研究结果提示 ,感染人体的BDV在病毒ORFⅠ基因片段的中间部分存在着高度的保守性 ,该片段可能是证实BDV感染的另一敏感性的标记物     

多发性硬化患者博尔纳病病毒p24基因片段的检测

博尔纳病病毒(Boma disease virus,BDV)是引发神经系统感染性疾病病原体的一种.我国重庆、哈尔滨、贵州等地区在病毒性脑炎、抑郁症、精神分裂症、帕金森病等部分疾病患者血液和脑脊液中检出过BDV,但多发性硬化(multiplesclerosis,MS)患者中未见报道.我们在13例MS患者外周血单个核细胞(PBMC)中检测BDV-p24基因片段,2例阳性,报道如下.     

多发性硬化患者博尔纳病病毒p24基因片段的检测

博尔纳病病毒(Boma disease virus,BDV)是引发神经系统感染性疾病病原体的一种.我国重庆、哈尔滨、贵州等地区在病毒性脑炎、抑郁症、精神分裂症、帕金森病等部分疾病患者血液和脑脊液中检出过BDV,但多发性硬化(multiplesclerosis,MS)患者中未见报道.我们在13例MS患者外周血单个核细胞(PBMC)中检测BDV-p24基因片段,2例阳性,报道如下.     

多发性硬化患者博尔纳病病毒p24基因片段的检测

博尔纳病病毒(Boma disease virus,BDV)是引发神经系统感染性疾病病原体的一种.我国重庆、哈尔滨、贵州等地区在病毒性脑炎、抑郁症、精神分裂症、帕金森病等部分疾病患者血液和脑脊液中检出过BDV,但多发性硬化(multiplesclerosis,MS)患者中未见报道.我们在13例MS患者外周血单个核细胞(PBMC)中检测BDV-p24基因片段,2例阳性,报道如下.     

多发性硬化患者博尔纳病病毒p24基因片段的检测

博尔纳病病毒(Boma disease virus,BDV)是引发神经系统感染性疾病病原体的一种.我国重庆、哈尔滨、贵州等地区在病毒性脑炎、抑郁症、精神分裂症、帕金森病等部分疾病患者血液和脑脊液中检出过BDV,但多发性硬化(multiplesclerosis,MS)患者中未见报道.我们在13例MS患者外周血单个核细胞(PBMC)中检测BDV-p24基因片段,2例阳性,报道如下.     

Borna disease virus RNA in peripheral blood mononuclear cells obtained from healthy dairy cattle

We examined natural infection of Borna disease virus (BDV) in 74 healthy dairy cattle in Japan. The seroprevalence determined using recombinant BDV p24 protein was 20.3%. The prevalence of BDV RNA in peripheral blood mononuclear cells (PBMCs) determined by polymerase chain reaction for p24 was 10.8%. Cell fraction of the PBMCs revealed the presence of BDV RNA in the T cell-enriched fraction. Thus, BDV infection is wide-spread even among healthy cattle. Received: 12 August 1996     

Development of a real-time RT-PCR assay for improved detection of Borna disease virus

Borna disease virus (BDV) is a non-segmented, negative-stranded RNA virus, which infects cells of the central nervous system (CNS) in many different species. BDV is the causative agent of the neurological disorders in horses and sheep termed classical Borna disease (BD), as well as staggering disease in cats. At present, the diagnosis staggering disease or feline BD is made by histopathology or immunohistochemistry of the CNS. In order to obtain a better clinical diagnostic tool, a duplex real-time RT-PCR assay (rRT-PCR) was developed. TaqMan probes and primers specific for the BDV P and BDV L genes were designed by aligning the sequences of known BDV strains. After optimisation, the sensitivity and specificity of the rRT-PCR were established. The detection limit was set to 10-100 viral genomic copies per reaction and the assay detects the BDV strains V and He/80, as well as the most divergent BDV strain known so far, No/98. Furthermore, the system detected feline BDV variants in five naturally infected cats and a feline isolate used in experimental infection of cats. This rRT-PCR assay will be a powerful tool in further studies of BDV, including epidemiological screening and diagnosis.     

The functional avidity of virus-specific CD8+ T cells is down-modulated in Borna disease virus-induced immunopathology of the central nervous system

Borna disease virus (BDV) infection of the central nervous system (CNS) leads to severe neurological symptoms in susceptible MRL mice. The disease is mainly mediated by CD8+ T cells specific for the immunodominant epitope TELEISSI in the BDV nucleoprotein. In this study, TELEISSI/MHC class I tetramers were used to directly visualize antigen-specific CD8+ T cells. We found that on average approximately 30% of the ex vivo analyzed CD8+ T cells in the CNS of diseased mice were specific for TELEISSI. Unexpectedly, the frequency of tetramer-reactive brain-derived CD8+ T cells doubled following overnight culture in the absence of antigen. The majority of CD8+ T cells showed enhanced tetramer binding without up-regulation of T cell receptor surface expression. The frequency of IFN-gamma-secreting CD8+ T cells after antigen-specific stimulation was higher in overnight cultures than in freshly isolated BDV-specific brain lymphocytes, and enhanced tetramer binding correlated with elevated sensitivity to lower levels of peptide antigen in cytotoxicity assays. These results indicate that the functional avidity of virus-specific CD8+ T cells was down-modulated in vivo. Thus, quantification of tissue-infiltrating CD8+ T cells by the tetramer technique must be interpreted with caution as it may underestimate the real frequency of antigen-specific CD8+ T cells.     

Neurological disorder after Borna disease virus infection in the absence of either interferon-gamma, Fas, inducible NO synthase, or chemokine receptor CXCR3

Borna disease virus (BDV) can induce severe neurological disorder in Lewis rats and MRL mice. Antiviral CD8 T cells have been shown to be the mediators of disease in these animals. To define molecules involved in the disease process, we performed infection studies in MRL mice lacking either interferon-gamma, a functional Fas/FasL system, chemokine receptor CXCR3, or inducible NO synthase. We further used transgenic MRL mice expressing interferon-gamma-inducible, T cell-attracting chemokine CXCL10 in brain astrocytes. After intracerebral infection with BDV, wild-type and mutant mice developed CD8 T cell responses and neurological disease at similar frequency and with similar kinetics, suggesting that these factors are not required for initiation and maintenance of the immunopathological process. Similarly, the course of disease could not be altered by treating infected MRL mice or Lewis rats with the drug L-N(6)-(1-iminoethyl)-lysine (L-NIL) that specifically blocks the activity of the inducible NO synthase. We therefore have excluded a number of important factors that have been demonstrated to be crucial in the pathogenesis of a broad number of pathologic conditions. Thus, BDV-induced disease may not result from the action of a single dominant T cell-dependent effector molecule. Disease rather reflects a combined influence of several as yet undefined factors from CD8 T cells.     

新疆伊犁地区动物脑组织博尔纳病病毒p24基因片段的检测

目的 了解新疆伊犁地区动物脑博尔纳病病毒(Borna disease virus,BDV)感染现状,分析该病毒的种系来源,预防我国BDV大规模暴发流行.方法 采用巢式逆转录实时荧光定量聚合酶链反应(FQ-RT-PCR)的方法,对新疆地区200匹马、75头驴和100只牧羊犬脑组织标本进行BDVp24基因片段的检测.并对阳性标本进行BDV p40基因片段扩增电泳,同时排除GFP-p24和pMD-19质粒污染,最后克隆测序,进行基因同源性和氨基酸序列分析并分析BDV种系发生.结果 3例伊犁马、4例伊犁驴和9例牧羊犬的脑组织标本BDV p24检测阳性,阳性率分别为1.5%、5.3%、9.0%;BDV p40基因片段检测和质粒标准品GFP-p24、pMD-19检测均为阴性;BDV p24扩增产物序列与He/80株的同源性为100%.结论 新疆伊犁地区马、驴和狗可能存在BDV脑内的自然感染,该地区BDV流行株与He/80株存在同源性.     

Tracking antigen-specific CD8+ T cells in the rat using MHC class I multimers

Studies of the quantitative and qualitative aspects of anti-microbial, anti-tumoral or autoreactive immune responses have been greatly facilitated by the possibility to stain antigen-specific CD8(+) T cells using fluorescently labeled multimeric major histocompatibility complex (MHC) class I/peptide complexes. So far, this technology has been developed for human and mouse, but not yet in the rat. Here, we describe the generation of the first rat MHC multimer. We produced a rat RT1(l) Pro5 MHC Pentamer combined with the immunodominant peptide for Borna disease virus (BDV), in order to study the characteristics of the antiviral CD8(+) T cell response. BDV is an RNA virus that can cause persistent infections of the central nervous system (CNS), often associated with prominent brain inflammation. In adult Lewis rats, of the RT1(l) MHC haplotype, BDV infection leads to severe immune-mediated neurological symptoms. The pathogenic role of the immune response is due primarily to antiviral CD8(+) T cells, many of them being specific for an immunodominant epitope located in the BDV nucleoprotein (N(230-238)). Ex vivo flow cytometry analyses revealed that 3 to 12% of CD8(+) T cells found in the brains of BDV-infected rats stained positively with the BDV-Pentamer. Interestingly, the frequency of Pentamer-positive cells increased up to 3.3 fold after a short resting period in culture. Virus-specific CD8(+) T cells were mainly detected in the brain and were virtually undetectable in peripheral lymphoid organs. This novel rat Pro5 MHC Pentamer represents an attractive tool for the detection, isolation and characterization of antigen-specific CD8(+) T cell responses in the rat.     

Amantadine does not have antiviral activity against Borna disease virus

Summary.  We have investigated the antiviral activity of amantadine (AD) against Borna disease virus (BDV) in several culture cell systems. We present evidence that AD, in the range 5 to 10 μM, does not have antiviral activity against BDV. Treatment of BDV infected cells with AD for six days caused neither a reduction in the number of infected cells, nor a decrease in steady state levels of BDV RNA or proteins. Moreover, treatment of cells with AD prior infection did not affect BDV multiplication, whereas influenza A virus yield was less than 1% with respect to that obtained in untreated control cells. Received May 20, 1997 Accepted June 19, 1997     

Demonstration of Borna disease virus (BDV) in specific regions of the brain from horses positive for serum antibodies to BDV but negative for BDV RNA in the blood and internal organs

Sero- and molecular-epidemiological studies on Borna disease virus (BDV) infection show that BDV RNA is not always detected in the peripheral blood mononuclear cells (PBMCs) from serum anti-BDV antibody-positive individuals such as horses, sheep, cattle, cats, and humans. In this study we demonstrated BDV RNA signals by polymerase chain reaction only in restricted regions of the brain from horses with locomotor disease. Four of six horses examined showed apparently positive reactions for anti-BDV antibodies. Specific regions of the brain of these four horses were positive for BDV RNA but the internal organs, lymph nodes, and PBMCs were negative. Histological studies of their brains revealed no apparent histological abnormalities such as inflammatory reactions. These results suggest that BDV chronically infects certain restricted regions of brain in seropositive horses. Received: 6 January 1997     

Borna disease virus

Borna disease virus (BDV) is unique amongst animal RNA viruses in its molecular biology and capacity to cause persistent, noncytolytic CNS-infection in a wide variety of host species. Unlike other non-segmented negative-strand RNA animal viruses, BDV replicates in the nucleus of the host cell where splicing is employed for expression of a very compact genome. Epidemiological studies indicate a broad host range and geographical distribution, and some investigators have proposed that human infection may result in neuropsychiatric disorders. Experimental Borna disease in neonatal and adult rats provides an intriguing model for immune-mediated disturbances of brain development and function.