Genetic variation in HIV disease: significance for diagn

HIV 疾病的遗传变异：对诊断的意义

• 批准号: 6839823
• 负责人: INDIRA HEWLETT
• 金额: --
• 依托单位: BUREAU OF HEALTH PLANNING AND RESOURCES DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6839823
• 关键词: AIDS /HIV test; African; HIV infections; biohazard detection; blood bank /supply contamination; comorbidity; deltaretrovirus; genetic strain; human herpesvirus 8; human immunodeficiency virus 1; human immunodeficiency virus 2; human tissue; immunogenetics; nanotechnology; patient oriented research; serotyping; smallpox vaccine; virus load; virus receptors

We are investigating the evolution of viral genetic diversity in HIV infection and multidrug resistance in patients on therapy. A repository of specimens from Cameroon that have been screened by a rapid HIV test currently in use in blood banks in Cameroon are being characterized by sequencing in the gag, pol and env region in order to identify their subtype and recombinant forms. Cameroon was selected for this study because of the high prevalence of multiple diverse subtypes in this region. The potential for recombinant forms to evolve would be expected to be high in such a setting. Approximately 150 screened positive and 100 screened negative specimens were tested on a number of licensed antibody, antigen and NAT assays. Most specimens were detected by the antibody and NAT assays although some were discordant and are under investigation by sequence analysis. All but one of the specimens were negative on p24 antigen tests. Sequence analysis revealed that most of the 150 positive specimens were recombinant A/G but novel A/C forms were identified. We are culturing virus from the sera for studies on virus biology including cytokine and chemokine profiles of these variant viruses. The prevalence of HTLV and HHV-8 as coinfections with HIV are also under investigation. HIV negative specimens are being evaluated for the presence of novel retroviral agents by a generic PCR assay. We are developing a microarray for detection of the various HIV-1 and -2 subtypes, HTLV and HHV-8 to expedite analysis of these strains. Future efforts will include application of nanotechnology to the detection of these pathogens. In a separate study, multidrug resistant HIV viruses were isolated from a group of patients and their biologic characteristics in regard to co-receptor usage, and replication capacity were studied. Most viruses could be cultured in vitro but although many of them exhibited an NSI phenotype some viruses showed neither X4 nor R5 properties suggesting that they may have evolved new co-receptor usage subsequent to treatment. We are currently exploring the role of other receptors and the potential for these viruses to establish reservoirs in other cell types. We are also studying the cytokine and chemokine profile and apoptotic potential of these viruses in comparison with naive strains in order to understand the pathogenesis of drug resistant HIV. Mass Smallpox vaccination has been considered as a preventive measure to counter-terrorism involving smallpox. This raises concerns about potential viremia in individuals who present as blood donors. Our study involves testing specimens from individual enrolled in 3 IND trials by Taqman and virus culture assays. Virus-cell interactions and host factors in HIV pathogenesis. We have characterized several new strains of the various HIV subtypes ranging from A-G of group M, isolates of group O and HIV-2. We have also obtained several drug resistant strains of HIV. We are sequencing the regulatory genes tat, nef, vpu and vpr to look for any possible relationships between these sequence s and co-receptor usage. Studies are underway to evaluate the replication of non clade B, O viruses and drug resistant viruses in in vitro cell systems with specific emphasis on their cell trpoism, replication kinetics, chemokine and cytokine usage. We are working with CDC to develop strategies in Cameroon (Africa) to identify new variants and recombinants of HIV and related viruses. These viruses are being investigated for their infectivity, drug susceptibility and pathogenesis in appropriate cell culture systems and animal models. We have examined the impact of HIV variation on apoptosis, chemokine and cytokine production in donor PBLs. We will also examine the role of tat and nef in faciltitating fasL mediated apoptosis in PBLs and dendritic cells. These studies will be also carried out in SCID mice. Effects of tat and nef on the regulation of chemokines in susceptible dendritic cells are being investigated. The infectivity of different HIV subtypes and variants for dendritic cells and macrophages are being studied. Detection, quantitation and characterization of HIV. 1. In-house serologic and genetic assays and Taqman probes are being developed for HIV-1 group O and non clade B subtypes. An in-house HIV group O ELISA and primers and probes for HIV group O and M has been developed. The probes will be used to generate microarray based assays for viral genotype. Oligo Chips are being developed to detect all known HIV group M subtypes, group O subtypes. In addition probes that are subtype specific are also under development. These assays will serve as tools in molecular epidemiologic studies to determine prevalence of non clade B strains particularly HIV-1 group O in endemic areas. We have recently initiated a study to study molecular evolution of HIV in Camerooon a region which harbors several subtypes. In these studies we will be characterizing various strains from specimens collected in Cameroon in order to identify new strains. WE have begun studies to investigate the diagnostic significance of strain diversity by testing sera using FDA licensed antibody and nucleic acid tests. We will also compare the performance of various direct viral marker assays such as PERT, RT-PCR, and p24 assays for their ability to detect viral variants and HIV in the early phase of infection. 2. Panels are being developed for the quantitation of RNA of HIV-1 subtypes. Virus isolates from each subtype have been sequenced to verify the subtype. These isolates are being quantitated in a collaborative study. An HIV-2 panel is also under development. Diagnostic and pathogeneic investigations of vaccinia virus: Implications for blood safety. We have initiated studies to investigate the period of viremia post-vaccination. Cells and plasma from recent vaccinees enrolled in candidate vaccine trials are being collected at various time points 0, 3, 7, 14, 28 and 56 days. Sensitive Taqman and virus culture assays have been developed. Plasma viremia studies indicate that virus was not detected in the vaccinee sera to date. In order to evaluate whether vaccination can result in interference in currently licensed viral marker tests used to screen donors samples from vaccinees are being tested on a variety of licensed tests. These studies are in progress. This project incorporates FY2002 projects 1Z01BP002001-10, 1Z01BP002003-10, and 1Z01BP002020-01.

我们正在研究HIV感染中病毒遗传多样性的演变和治疗患者的多药耐药性。来自喀麦隆的标本的存储库已通过当前在喀麦隆血库中使用的快速HIV测试进行筛选，这是通过在GAG，POL和ENV区域进行测序的特征，以识别其亚型和重组形式。由于该区域多种多样的亚型的高患病率很高，因此选择了喀麦隆。在这种情况下，重组形式进化的可能性将很高。在许多许可抗体，抗原和NAT测定法上测试了大约150个筛选的阳性和100个筛选的负标本。大多数样品是通过抗体和NAT分析检测到的，尽管有些是不一致的，并且通过序列分析进行了研究。除一个标本外，所有样品均为P24抗原测试。序列分析表明，150个阳性标本中的大多数是重组A/G，但已鉴定出新型的A/C形式。我们正在从血清中培养病毒，以研究这些变体病毒的细胞因子和趋化因子谱。 HTLV和HHV-8作为HIV共感染的患病率也正在研究中。通过通用PCR分析，正在评估HIV阴性标本的新型逆转录病毒药物。 我们正在开发一个微阵列，用于检测各种HIV-1和-2亚型HTLV和HHV-8，以加快对这些菌株的分析。未来的努力将包括将纳米技术应用于这些病原体的检测。 在另一项研究中，从一组患者中分离了多药抗HIV病毒及其在共受体使用方面的生物学特征，并研究了复制能力。大多数病毒可以在体外培养，但是尽管其中许多病毒表现出NSI表型，但某些病毒既没有X4也没有R5特性，表明它们可能在治疗后进化了新的共受体用法。我们目前正在探索其他受体的作用，以及这些病毒在其他细胞类型中建立储层的潜力。与天真菌株相比，我们还研究了这些病毒的细胞因子和趋化因子谱以及凋亡潜力，以了解耐药性HIV的发病机理。 质量天花疫苗接种被认为是涉及天花的反恐的预防措施。这引起了人们对作为献血者表现的个体潜在病毒血症的担忧。我们的研究涉及从塔克曼和病毒培养试验中参加3项IND试验的个体测试标本。 HIV发病机理中的病毒 - 细胞相互作用和宿主因子。我们已经表征了各种HIV亚型的几种新菌株，其范围为M组M和HIV-2组分离株。 我们还获得了几种抗药性HIV菌株。我们正在测序调节基因TAT，NEF，VPU和VPR，以寻找这些序列S和共受体使用之间的任何可能关系。正在进行研究以评估体外细胞系统中非进化枝B，O病毒和耐药性病毒的复制，并特别强调其细胞TRPOISM，复制动力学，趋化因子和细胞因子使用。 我们正在与CDC合作，在喀麦隆（非洲）制定策略，以识别艾滋病毒和相关病毒的新变异和重组。 这些病毒正在研究适当的细胞培养系统和动物模型中的感染性，药物敏感性和发病机理。我们已经检查了HIV变异对供体PBL中凋亡，趋化因子和细胞因子产生的影响。我们还将研究TAT和NEF在PBL和树突状细胞中促进FASL介导的凋亡中的作用。这些研究也将在SCID小鼠中进行。正在研究TAT和NEF对易感树突状细胞中趋化因子调节的影响。 正在研究不同HIV亚型和树突状细胞和巨噬细胞变异的感染性。 HIV的检测，定量和表征。 1。正在为HIV-1组O和非进化枝B亚型开发内部血清学和遗传测定法和Taqman探针。 已经开发了一个内部HIV组O ELISA以及HIV组O和M的引物和探针。这些探针将用于生成基于微阵列的病毒基因型测定法。正在开发寡芯片，以检测所有已知的HIV组M亚型，O组亚型。此外，特定于亚型的探针也正在开发中。这些测定将用作分子流行病学研究的工具，以确定非进化核B菌株的患病率，尤其是在流行地区的HIV-1组O。我们最近开始了一项研究，研究了摄影室中HIV的分子进化，该区域具有多种亚型。在这些研究中，我们将表征喀麦隆收集的标本的各种菌株，以识别新菌株。我们已经开始研究通过使用FDA许可抗体和核酸测试来研究血清的诊断意义。我们还将比较各种直接病毒标记测定的性能，例如PERT，RT-PCR和P24分析，以在感染的早期检测病毒变异和HIV的能力。 2。正在开发用于定量HIV-1亚型RNA的面板。已经对每个亚型的病毒分离株进行了测序以验证亚型。这些分离株正在协作研究中进行定量。 HIV-2面板也正在开发中。 疫苗病毒的诊断和病原体研究：对血液安全的影响。我们已经开始研究疫苗接种后的病毒血症期。在不同时间点0、3、7、14、28和56天，正在收集参加候选疫苗试验的近期疫苗的细胞和血浆。 敏感的Taqman和病毒培养试验已经开发出来。血浆病毒血症研究表明，迄今为止，在疫苗血清中未发现病毒。 为了评估疫苗接种是否会导致对目前有执照的病毒标记测试的干扰，用于从疫苗中筛查供体样品的样本正在接受各种许可测试测试。这些研究正在进行中。 该项目包含2002财年项目1Z01BP002001-10、1Z01BP002003-10和1Z01BP0020202020202020-01。