The epidemiology and host-fungal neuroimmune interaction of paediatric cryptococcal infection in South Africa.

南非儿童隐球菌感染的流行病学和宿主-真菌神经免疫相互作用。

• 批准号: 2720640
• 金额: --
• 依托单位: UNIVERSITY OF EXETER
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2720640
• 关键词: epidemiology; host; fungal; neuroimmune; interaction

The fungus Cryptococcus is the leading cause of meningitis in adults in Sub-Saharan Africa and has very poor outcomes with most patients dying or developing brain damage. It predominantly affects patients with HIV infection. Although childhood infection is recognised to be rarer than in adults, the resulting damage to the developing brain is significant and the number of infections in children is largely unknown. The World Health Organization recommends testing every child over 10 years old with advanced HIV disease for evidence of cryptococcal infection as pre-emptive treatment saves lives.The aims of the project are:a) Define the prevalence and clinical management of cryptococcal antigenaemia and incidence of CM in children in SA.A retrospective cohort study will be performed across all public healthcare facilities in SA including all CrAg positive results from children <18 yrs between 2017 and 2022 identified by the Surveillance Data Warehouse of the National Institute for Communicable Diseases (NICD). Prevalence will be calculated using mid-year population denominators from Statistics South Africa and the Thembisa model. A prospective study will be set-up using GERMS-SA, an active NICD surveillance system. Clinical isolates and CSF will be stored for host-fungus interaction studies for aim 2 and 3. Already collected and stored paediatric cryptococcal isolates are also available for use in aim 2.b) Compare the Cryptococcal neoformans cell wall composition of adult and paediatric clinical isolates.Five clinical isolates from adult and five from paediatric infections and wild-type C. neoformans will be grown in host-mimicking conditions. Stored isolates (NICD) will be used, with fresh samples from the prospective study added when available. Cell wall and capsule components will be compared by staining for chitin, chitosan, chito-oligomers, glucan and mannans and measuring fluorescence intensity on flow cytometry. Morphology will be compared by staining cell surface components and visualisation with confocal and electron microscopy. This work will inform the isolates used in aim 3.c) Describe the neuroinflammatory effects of paediatric C. neoformans isolates using in vivo and ex vivo models.The paediatric CSF samples (aim 1) will be analysed by ELISA for their cytokine profile (e.g. TNF, IL-6, INF-a) and compared to adult samplesWe will characterize the paediatric neuroimmune response to Cryptococcus infection using an established mouse model. To delineate molecular mechanisms underlying paediatric neurocryptococcosis at the cellular level, ex vivo, organotypic murine brain slice cultures will be used. Organotypic brain slices obtained from six-day-old mouse pups will be cultured on a semi-permeable membrane in artificial CSF then stimulated with clinical Cryptococcal isolates. Neuroinflammatory activation will be measured using immunohistochemistry and cytokine assays as described above. Dr Dangarembizi's laboratory has access to paediatric brain tissue from epilepsy surgery so there is a possibility of advancing to human organotypic slice cultures.This project will provide important epidemiological information about paediatric cryptococcal antigenaemia and cryptococcosis in South Africa. It will help fill the existing gap in knowledge of the pathogenesis of cryptococcal meningitis and the differences observed in the adult and paediatric population.

隐球菌真菌是撒哈拉以南非洲成年人脑膜炎的主要原因，其结果非常差，大多数患者死亡或出现脑损伤。它主要影响艾滋病毒感染者。虽然儿童感染被认为比成人罕见，但对发育中的大脑造成的损害是显著的，儿童感染的数量在很大程度上是未知的。世界卫生组织建议对每一名10岁以上的晚期艾滋病儿童进行隐球菌感染的检测，以获得隐球菌感染的证据，作为预防性治疗可以挽救生命。该项目的目的是：a）确定南非儿童隐球菌抗原血症的患病率和临床管理以及CM的发病率。将在南非所有公共医疗机构进行回顾性队列研究，包括2017年至2022年期间由国家传染病研究所监测数据仓库（NICD）确定的18岁以下儿童的所有CrAg阳性结果。将使用南非统计局的年中人口乘数和Thembisa模型计算流行率。一项前瞻性研究将使用GERMS-SA，一个积极的NICD监测系统。将储存临床分离株和CSF，用于目标2和3的宿主-真菌相互作用研究。已经收集和储存的儿科隐球菌分离株也可用于目标2。b）比较成人和儿科临床分离株的新型隐球菌细胞壁组成。新型人将在模拟宿主的条件下生长。将使用储存的分离株（NICD），并在可用时添加来自前瞻性研究的新鲜样本。将通过对几丁质、壳聚糖、壳聚糖低聚物、葡聚糖和甘露聚糖染色并在流式细胞术上测量荧光强度来比较细胞壁和囊组分。将通过染色细胞表面组分并使用共聚焦和电子显微镜观察来比较形态学。这项工作将为目标3.c）中使用的分离株提供信息，描述儿童C。将通过ELISA分析儿科CSF样品（目的1）的细胞因子谱（例如TNF、IL-6、INF-α），并与成人样品进行比较。我们将使用建立的小鼠模型表征儿科对隐球菌感染的神经免疫应答。为了在细胞水平上描述儿科神经隐球菌病的分子机制，将使用离体器官型小鼠脑切片培养物。将从6日龄小鼠幼仔获得的器官型脑切片在人工CSF中的半透膜上培养，然后用临床隐球菌分离株刺激。将使用如上所述的免疫组织化学和细胞因子测定来测量神经炎性活化。Dangarembizi博士的实验室可以从癫痫手术中获得儿科脑组织，因此有可能推进人类器官型切片培养。该项目将提供有关南非儿科隐球菌抗原血症和隐球菌病的重要流行病学信息。这将有助于填补隐球菌性脑膜炎发病机制以及在成人和儿童人群中观察到的差异方面的现有知识空白。