Pathogenesis of Tick-Borne Flavivirus Infections

蜱传黄病毒感染的发病机制

基本信息

批准号：
10927795
负责人：
Marshall Elliott Bloom
金额：
$ 159.19万
依托单位：
NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
依托单位国家：
美国
项目类别：
财政年份：
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/10927795
关键词：
Acinus organ component Adult Anaplasma Anaplasma phagocytophilum Animal Model Antiviral Agents Arthropods Artificial Membranes Attenuated Basic Science Binding Biological Assay Biology Black-legged Tick Blood Borrelia Borrelia burgdorferi Cells Cellular biology Central Nervous System Diseases Cessation of life Chronic Chronic Disease Collaborations Coxiella burnetii Digestion Disease Dissociation Entomology Experimental Models Flavivirus Infections Genes Genomics Human Image Imaging Techniques Immune response Immunology Infection Infectious Agent Integration Host Factors Laboratories Lyme Disease Methods Microbe Midgut Molecular Molecular Biology Mus Nymph Organ Culture Techniques Pathogenesis Pathway interactions Play Powassan virus Proliferating Proteomics Protocols documentation Publications Publishing Q Fever Refractory Research Rickettsia Rocky Mountain Spotted Fever Rodent Model Role Salivary Glands Side Supporting Cell Suspensions Symptoms System Testing Ticks Vaccine Design Vaccines Vial device Viral Virginia Virus Virus Diseases Virus Replication Virus-like particle Work acute infection animal model development cell type chronic infection co-infection deep sequencing feeding genome wide screen intestinal epithelium microorganism interaction mouse model nervous system disorder novel permissiveness receptor screening single cell analysis small molecule libraries therapy development tick feeding tick-borne tick-borne flavivirus tick-borne virus tool transmission process vector vector transmission vector-borne virology

项目摘要

Research in our laboratory employs virology, immunology, entomology, advanced imaging techniques, genomics, proteomics, cell biology, molecular biology, and vector biology. We study LGTV at BSL-2 and POWV/DTV at BSL-3. TBFV biology in I. scapularis. Infection in ticks is a critical, but understudied, feature of TBFV biology. Our ex vivo organ culture work has yielded considerable information about TBFV infection in midgut and salivary glands of adult ticks, and is presently under review for publication. In the past year, Dr. Ochwoto and Dr. Stewart employed artificial membrane blood feeding and demonstrated that adult I. scapularis ticks can be infected with relatively high efficiency by feeding them on LGTV infected blood. Studies on the midgut of fed adults revealed the expected differentiation and proliferation of the intestinal epithelium into mature digestive cells, and the results indicated that these cells support virus replication. Initial studies suggest that the salivary glands also become infected while the ticks are feeding and that prior infection of the midgut and subsequent systemic dissemination of virus are not required for infection of the salivary glands. The nymphal stage of the ticks is the most important for human infection, so in the past year we have started to study infection in nymphs. However, when efforts were made to infect them using the artificial membrane system, the results were disappointing and inconsistent. As a result, Dr. Weck is planning to infect nymphs by feeding them on LGTV infected mice. Salivary glands have at least 3 types of acini and each acini contains multiple cell types. It is unclear if all acinar types or all cell types are permissive for virus infection. One way to approach this interesting question would be to infect ex vivo salivary gland cultures, dissociate the infected cultures and subject the cells to single cell analysis. In the past year, Dr. Ochwoto and Ms. Theriault applied cell dissociation methods to dissected salivary glands from adult ticks. The acini were refractory to dissociation using simple enzymatic digestion and we have not yet obtained single cell suspensions satisfactory for analysis. Microbial interactions in Ixodes scapularis ticks. Ixodes scapularis ticks harbor multiple infectious agents, including TBFV, B. burgdorferi and Anaplasma phagocytophilum, but polymicrobial human infections are rare. A possible explanation is that infection with one microbe interferes with infection by a second. In the past year, Dr. Stewart established a collaboration with Dr Eastwood from Virginia Tech to examine co-infections with anaplasma and TBFV. Dr Stewart is also developing tools to identify the TBFV receptor in both mammalian and arthropod systems. The approach is to create virus-like particles with a fluourescent tag that can be used in various binding assays. The expression constructs have been developed and protocols to purify the VLP are being tested. Identification of genes essential for TBFV infections and antivirals. In the past year, our work on genome wide screens for host factors regulating TBFV infection has undergone analysis and should be ready for publication in the coming FY. Also in the past year, we began testing several libraries of small molecules against LGTV. Several promising compounds have been identified and will be evaluated to see if they might be useful antivirals. Development of an animal model for chronic POWV/DTV disease. Understanding how these viruses cause long-term symptoms as well as the possible role of viral persistence is important for developing therapies to treat chronic infection. However, there are not good animal models to study chronic disease. In the past year, we published our mouse model of chronic POWV/DTV disease. The results indicated that the continued presence of detectable virus infection was not required for chronic CNS disease, suggesting that host responses play an important role. In an interesting side project, we supplied a vial of 1940's vintage rocky mountain spotted fever vaccine, prepared from infected ticks, to Dr Taubenberger and his colleagues. Deep sequencing of the material revealed the the presence of Rickettsia ricketsii, demonstrating that that agent against which the vaccine designed was present. In addition, however, sequence reads of Coxiella burnetii, the agent of Q fever, were also identified. This likely indicated that some of the ticks were co-infected with both agents.

我们实验室的研究采用病毒学，免疫学，昆虫学，高级成像技术，基因组学，蛋白质组学，细胞生物学，分子生物学和载体生物学。我们在BSL-2和BSL-3的POWV/DTV上研究LGTV。 I. capularis中的TBFV生物学。 TICK中的感染是TBFV生物学的关键但研究的特征。我们的离体器官文化工作已经提供了有关成人tick虫的Midgut和唾液腺感染的大量信息，目前正在审查出版。在过去的一年中，Ochwoto博士和Stewart博士采用了人造膜血液喂养，并证明了成年的I.肩cap虫可以通过在LGTV感染的血液上喂食，以相对较高的效率感染。关于美联储成年人中肠的研究揭示了肠上皮的预期分化和扩散到成熟的消化细胞中，结果表明这些细胞支持病毒复制。初步研究表明，唾液腺在进食时也被感染，并且在感染唾液腺的感染并不需要中肠感染和随后的全身传播病毒。壁虱的若虫阶段对于人类感染最为重要，因此在过去的一年中，我们开始研究若虫的感染。但是，当努力使用人造膜系统感染它们时，结果令人失望和不一致。结果，Weck博士计划通过将若虫喂在LGTV感染的小鼠上来感染若虫。唾液腺至少具有3种类型的acini，每种acini包含多种细胞类型。目前尚不清楚所有腺泡类型或所有细胞类型是否允许病毒感染。解决这个有趣的问题的一种方法是感染离体唾液腺培养物，解离感染培养物，并使细胞接受单细胞分析。在过去的一年中，Ochwoto博士和Theriault女士将细胞解离方法应用于成人tick虫的唾液腺。使用简单的酶消化是难治性解离的难治性，我们尚未获得单个细胞悬浮液以进行分析。 ixodes肩cap虫中的微生物相互作用。 ixodes capularis tick虫具有多种传染剂，包括TBFV，B。Brgdorferi和Anaplasma phagocytophilum，但多数菌种人类感染很少见。一个可能的解释是，一种微生物感染会干扰感染一秒钟。在过去的一年中，斯图尔特博士与弗吉尼亚理工大学的伊斯特伍德博士建立了合作，以检查Anaplasma和TBFV的共同感染。 Stewart博士还正在开发识别哺乳动物和节肢动物系统中TBFV受体的工具。该方法是创建具有荧光标签的病毒样颗粒，可用于各种结合测定。已经开发了表达构造，并正在测试净化VLP的协议。鉴定对TBFV感染和抗病毒药所必需的基因。在过去的一年中，我们在基因组宽屏幕上为调节TBFV感染的宿主因素的工作已经进行了分析，应该准备在即将到来的FY中发表。同样在过去的一年中，我们开始测试几个针对LGTV的小分子库。已经确定了几种有希望的化合物，并将评估它们是否可能是有用的抗病毒药。开发用于慢性POWV/DTV疾病的动物模型。了解这些病毒如何引起长期症状以及病毒持久性的可能作用对于开发治疗慢性感染的疗法很重要。但是，没有很好的动物模型来研究慢性疾病。在过去的一年中，我们发表了慢性POWV/DTV疾病的小鼠模型。结果表明，慢性中枢神经系统疾病不需要持续存在可检测的病毒感染，这表明宿主反应起着重要作用。在一个有趣的方面项目中，我们向Taubenberger博士及其同事提供了1940年的1940年Vintage Rocky Mountain Spotted发烧疫苗。对材料的深度测序揭示了立克西氏菌的存在，表明存在疫苗设计的那种药物。然而，还确定了Q发烧的药物Coxiella burnetii的序列读数。这可能表明，某些壁虱已与两个代理共同感染。

项目成果

期刊论文数量（32）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Genetic sequencing of a 1944 Rocky Mountain spotted fever vaccine.

DOI：
10.1038/s41598-023-31894-0
发表时间：
2023-03-22
期刊：
SCIENTIFIC REPORTS
影响因子：
4.6
作者：
Xiao, Yongli;Beare, Paul A.;Best, Sonja M.;Morens, David M.;Bloom, Marshall E.;Taubenberger, Jeffery K.
通讯作者：
Taubenberger, Jeffery K.

A three-dimensional comparison of tick-borne flavivirus infection in mammalian and tick cell lines.

DOI：
10.1371/journal.pone.0047912
发表时间：
2012
期刊：
PloS one
影响因子：
3.7
作者：
Offerdahl DK;Dorward DW;Hansen BT;Bloom ME
通讯作者：
Bloom ME

Cytoarchitecture of Zika virus infection in human neuroblastoma and Aedes albopictus cell lines.

DOI：
10.1016/j.virol.2016.11.002
发表时间：
2017-01-15
期刊：
VIROLOGY
影响因子：
3.7
作者：
Offerdahl, Danielle K.;Dorward, David W.;Hansen, Bryan T.;Bloom, Marshall E.
通讯作者：
Bloom, Marshall E.

Characterization of flavivirus infection in salivary gland cultures from male Ixodes scapularis ticks.

DOI：
10.1371/journal.pntd.0008683
发表时间：
2020-10
期刊：
PLoS neglected tropical diseases
影响因子：
3.8
作者：
Kendall BL;Grabowski JM;Rosenke R;Pulliam M;Long DR;Scott DP;Offerdahl DK;Bloom ME
通讯作者：
Bloom ME

Transcriptome Analysis Reveals a Signature Profile for Tick-Borne Flavivirus Persistence in HEK 293T Cells.

DOI：
10.1128/mbio.00314-16
发表时间：
2016-05-24
期刊：
mBio
影响因子：
6.4
作者：
Mlera L;Lam J;Offerdahl DK;Martens C;Sturdevant D;Turner CV;Porcella SF;Bloom ME
通讯作者：
Bloom ME