牛疱疹病毒Ⅰ型VP22和猪繁殖与呼吸综合征病毒GP5融合基因DNA疫苗的免疫效应

为了提高表达GP5的猪繁殖与呼吸综合征病毒（PRRSV）DNA疫苗的免疫效应,将具有蛋白转导功能的牛疱疹病毒1型（BHV-1）VP22基因插入到经过修饰具有更好免疫原性的PRRSV修饰型ORF5基因（ORF5M）上游,构建VP22和ORF5M融合表达的真核表达质粒pCI-VP22-ORF5M。经间接免疫荧光试验（IFA）和Westernblot检测证实体外表达后,免疫BALB/c小鼠,检测小鼠免疫后的GP5特异性ELISA抗体、抗PRRSV中和抗体和脾淋巴细胞增殖反应,并与非融合的真核表达质粒pCI-ORF5M进行比较。结果显示,融合表达VP22-GP5的DNA疫苗 pCI-VP22ORF5M诱导的体液免疫和细胞免疫反应均明显高于非融合表达的DNA疫苗pCI-ORF5M,表明蛋白转导相关蛋白BHV-1 VP22能显著增强表达GP5的PRRSV DNA 疫苗的免疫效应,有效发挥了基因免疫佐剂效应;这为研制PRRSV高效DNA疫苗奠定了基础,同时也为其它疾病的高效新型疫苗研究提供了思路。     

Unique structures in a tumor herpesvirus revealed by cryo-electron tomography and microscopy

Gammaherpesviruses, including the human pathogens Epstein–Barr virus and Kaposi’s sarcoma-associated herpesvirus, are causative agents of lymphomas and other malignancies. The structural characterization of these viruses has been limited due to difficulties in obtaining adequate amount of virion particles. Here we report the first three-dimensional structural characterization of a whole gammaherpesvirus virion by an emerging integrated approach of cryo-electron tomography combined with single-particle cryo-electron microscopy, using murine gammaherpesvirus-68 (MHV-68) as a model system. We found that the MHV-68 virion consists of distinctive envelope and tegument compartments, and a highly conserved nucleocapsid. Two layers of tegument are identified: an inner tegument layer tethered to the underlying capsid and an outer, flexible tegument layer conforming to the overlying, pleomorphic envelope, consistent with the sequential viral tegumentation process inside host cells. Surprisingly, comparison of the MHV-68 virion and capsid reconstructions shows that the interactions between the capsid and inner tegument proteins are completely different from those observed in alpha and betaherpesviruses. These observations support the notion that the inner layer tegument across different subfamilies of herpesviruses has evolved significantly to confer specific characteristics related to viral–host interactions, in contrast to a highly conserved capsid for genome encapsidation and protection.     

Cryo-electron tomography of Kaposi's sarcoma-associated herpesvirus capsids reveals dynamic scaffolding structures essential to capsid assembly and maturation

Kaposi’s sarcoma-associated herpesvirus (KSHV) is a recently discovered DNA tumor virus that belongs to the γ-herpesvirus subfamily. Though numerous studies on KSHV and other herpesviruses, in general, have revealed much about their multilayered organization and capsid structure, the herpesvirus capsid assembly and maturation pathway remains poorly understood. Structural variability or irregularity of the capsid internal scaffolding core and the lack of adequate tools to study such structures have presented major hurdles to earlier investigations employing more traditional cryo-electron microscopy (cryoEM) single particle reconstruction. In this study, we used cryo-electron tomography (cryoET) to obtain 3D reconstructions of individual KSHV capsids, allowing direct visualization of the capsid internal structures and systematic comparison of the scaffolding cores for the first time. We show that B-capsids are not a structurally homogenous group; rather, they represent an ensemble of “B-capsid-like” particles whose inner scaffolding is highly variable, possibly representing different intermediates existing during the KSHV capsid assembly and maturation. This information, taken together with previous observations, has allowed us to propose a detailed pathway of herpesvirus capsid assembly and maturation.     

The in vitro antiviral activity of an aliphatic nitro compound from Heteropteris aphrodisiaca

We investigated the antiviral activity of an aliphatic nitro compound (NC) isolated from Heteropteris aphrodisiaca O. Mach. (Malpighiaceae), a Brazilian medicinal plant. The NC was tested for its antiviral activity against poliovirus type 1 (PV-1) and bovine herpes virus type 1 (BHV-1) by plaque reduction assay in cell culture. The NC showed a moderate antiviral activity against PV-1 and BHV-1 in HEp-2 cells, and the 50% inhibitory concentration (IC50) were 22.01 microg/ml (selectivity index (SI)=2.83) and 21.10 microg/ml (SI=2.95), respectively. At the highest concentration of the drug (40 microg/ml) a reduction of approximately 80% in plaque assay was observed for both viruses. The treatment of cells or virus prior to infection did not inhibit the replication of virus strains.     

Prevalence of Kaposi’s sarcoma-associated herpesvirus among intravenous drug users: a systematic review and meta-analysis

Intravenous drug users(IDUs) have been demonstrated to be highly vulnerable to HIV/AIDS.Nevertheless, the prevalence of Kaposi's sarcoma associated herpesvirus(KSHV), an important co-infected agent with HIV, among this population remained obscure. We conducted a systematic review on the epidemiological features of KSHV among IDUs worldwide. Eligible studies were retrieved from 6 electronic databases(Pub Med, EMBASE, Web of Science, CBM, CNKI and Wanfang).We calculated the pooled prevalence and 95% confidence interval(CI) overall and among subgroups using either random-effects model or fixed-effects model depending on between-study heterogeneity. The potential publication bias was assessed by the Egger's test. A meta-regression analysis was performed to explore the sources of heterogeneity. Finally, twenty-two studies with a total sample of 7881 IDUs were included in the analysis. The pooled prevalence of KSHV was14.71%(95% CI 11.12%–19.46%) among IDUs. Specifically, KSHV prevalence was 10.86%(95% CI6.95%–16.96%) in HIV-negative IDUs, and 13.56%(95% CI 10.57%–17.38%) in HIV-positive IDUs.Moreover, prevalence among IDUs from the three continents involved in the current study was similar:16.10%(95%CI 7.73%–33.54%) in Asia; 14.22%(95%CI 8.96%–22.57%) in Europe and 14.06%(95%CI11.38%–17.37%) in America. Globally, IDUs are at higher risk of the KSHV infection when compared with the general population, regardless of geographical region or HIV-infection status.     

Detection of congenital cytomegalovirus in newborns using nucleic acid amplification techniques and its public health implications

Human cytomegalovirus (HCMV), a herpesvirus, is an important human pathogen that causes asymptomatic infections in healthy or immunocompetent individuals but can lead to severe and potentially life-threatening complications in immune-immature individuals such as neonates or immune-compromised patients such as organ-transplant recipients and HIV-positive individuals. Congenital HCMV infection represents a significant public health issue and poses substantial healthcare and economic burden to society. This virus causes the most common viral congenital infection worldwide, and is the leading non-genetic cause of sensorineural hearing loss in children in developed countries. Congenital HCMV infection is believed to fulfill the criteria of the American College of Medical Genetics to be considered as a condition targeted for a newborn screening program. This is because congenital HCMV infection can be identified during a time (within 2 days after birth) at which it would not ordinarily be detected clinically, and there are demonstrated benefits of early detection, timely intervention, and efficacious treatment of the condition. Recent progresses in developing polymerase chain reaction-based approaches to detect HCMV in samples obtained from newborns have generated much excitement in the field. In this review, we highlight the recent progress in diagnostic techniques that could potentially be used for the detection of HCMV infection in neonates and its direct implications in public health settings for diagnosing congenital HCMV infection.

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人类疱疹病毒7开放读码框U41功能的初步探讨

应用免疫荧光检测U41蛋白的细胞内定位,RT-PCR检测U41RNA的表达时相,DNA结合试验检测U41蛋白对单、双链DNA的不同结合能力,对人类疱疹病毒7(human herpesvirus 7,HHV-7)U41的部分功能进行分析鉴定.免疫荧光检测显示,无论在HHV-7感染细胞或HHV-7 U41转染细胞,U41蛋白都是仅在其胞核中呈弥漫性分布.在分别加入环己酰亚胺(cycloheximide)和磷甲酸(phosphonoformic acid)作为蛋白合成和核酸合成抑制剂的感染细胞中,U41的表达时相检测结果显示,U41蛋白在HHV-7感染细胞中是一早期表达蛋白.通过DNA结合试验发现,U41蛋白能结合单链DNA而不能结合双链DNA.提示U41的主要功能是保持DNA模板的合适构型,以便DNA的延伸和病毒DNA的合成.     

The Biology of Kaposi＇s Sarcoma-Associated Herpesvirus and the Infection of Human Immunodeficiency Virus

Kaposi sarcoma-associated herpesvirus (KSHV),also known as human herpesvirus 8 (HHV-8),is discovered in 1994 from Kaposi's sarcoma (KS) lesion of an acquired immunodeficiency syndrome (AIDS)patient.In addition to its association with KS,KSHV has also been implicated as the causative agent of two other AIDS-associated malignancies:primary effusion lymphoma (PEL) and multicentric Castleman's disease (MCD).KSHV is a complex DNA virus that not only has the ability to promote cellular growth and survival for tumor development,but also can provoke deregulated angiogenesis,inflammation,and modulate the patient's immune system in favor of tumor growth.As KSHV is a necessary but not sufficient etiological factor for KS,human immunodeficiency virus (HIV) is a very important cofactor.Here we review the basic information about the biology of KSHV,development of pathogenesis and interaction between KSHV and HIV.     

人疱疹病毒8型KS330基因片段的检出与Kaposi肉瘤的关系

HHV-8 sequences were recently identified in 100% of the amplifiable samples from AIDS patients with Kaposi's sarcoma(KS)and in 15% of the non-KS tissue samples from AIDS patients, so there is a strong correlation of Kaposi's sarcoma with HHV-8. Serum and DNA samples from a clinically diagnosed Kaposi's sarcoma Chinese patient were tested. HHV-8 antibody was tested positive by IFA and HIV-I antibody was negative by Western blot. The KS330 PCR product was found both in peripheral blood mononuclear cells and in KS tumor cells from this Chinese patient. This supports the hypothesis that Kaposi's sarcoma results from infection of HHV-8.     

Efficient propagation of human herpesvirus 6B in a T-cell line derived from a patient with adult T-cell leukemia/lymphoma.

The efficient propagation of the OK strain of the B variant of human herpesvirus 6 (HHV-6B) was demonstrated in a line of T cells, TaY, established from the peripheral blood lymphocytes of a patient with adult T-cell leukemia/lymphoma (ATL). Growth of TaY cells depends on the presence of IL-2 and the cells harbor HTLV-I genomes. A severe cytopathic effect (CPE) was observed in many HHV-6B(OK)-infected TaY cells one week after infection. The release of virus from HHV-6B(OK)-infected TaY cells [TaY(OK)] was first detected after three days and increased rapidly for up to seven days after infection, as demonstrated by PCR. The titer of HHV-6B(OK) in the supernatant was comparable to the value of 10(3.5) TCID50/ml obtained with PHA-activated cord blood lymphocytes (CBL) that had been infected with HHV-6B(OK). The replication of the virus was shown to depend to a considerable extent on cell viability. Electron microscopy revealed many herpesvirus-type capsid- and enveloped-viruses in the nuclei and cytoplasm of degenerated cells in TaY(OK) cultures. The U1102 strain of HHV-6A and the Z29 strain of HHV-6B also infected TaY cells productively, as detected by PCR and an immunofluorescence test. These results suggest that the activation of CD4+ T lymphocytes with mitogens such as PHA or IL-2 and the expression of some cellular gene or the HTLV-I gene might be essential for efficient propagation of HHV-6B. TaY cells should play an important role in future investigations of cell-virus interactions and genetic variations or cell tropism of HHV-6 isolates since no cell line that shows propagation of both HHV-6A and HHV-6B has been reported to date.