Attributing the source of antimicrobial resistant diarrheal pathogens in African children

空肠弯曲菌是一种人畜共患病的肠道病原体，是世界范围内引起胃肠炎的最常见原因之一。虽然空肠弯曲菌感染在较富裕的国家通常是自我限制的，但由于慢性再感染，它在低收入到中等收入国家(LMIC)施加了更大的负担。空肠弯曲菌在包括鸡、牛和野鸟在内的动物的肠道中定居；并通过食用未煮熟的肉类和受污染的水源传播。由于空肠弯曲菌占据着大量的宿主和宿主，了解空肠弯曲菌是如何在群落中传播的，更具体地说，了解这些传播网络中的生存和对环境应激源的耐受性是重要的。这项工作的总体目标是了解空肠弯曲菌对环境应激源的反应，环境应激源可能导致腹泻疾病在LMICs中的传播。该项目将利用冈比亚、布基纳法索和加纳的样本分离株，这是一项名为“GetCampy Africa”的多所大学研究的一部分。这将通过开发高通量方案来评估空肠弯曲菌对诸如氧气、抗生素和洗涤剂等应激源的耐受性的表型和基因类型来实现。这些结果将为我在有氧压力下进化分离株的实验提供依据，并识别赋予有氧耐受性的适应性突变。最终目标是提供更好的分辨率和对增加空肠弯曲菌传播性的适应特征的理解，这最终将为从更大的‘GetCampy Africa’项目绘制的传播网络提供信息。目标1：使用实验室参考和欧洲空肠弯曲菌菌株优化高通量生长和表型分析。该方法将构成评估环境应激源对更多空肠弯曲菌影响的其他方案的基础。目标2：从“GetCampy Africa”项目分离的空肠弯曲菌菌株的表型和测序。使用目标1中开发的方法评估有氧耐力和有氧生长的差异，以确定宿主和/或谱系特定的差异和相似之处。目的：利用实验进化方法筛选耐氧性基因。在整个项目中，进化实验将与暴露于不同环境应激源的控制线和实验线一起进行--最初集中在有氧应激上。祖先和进化的株系将被测序，以识别赋予耐氧性的基因，并将它们与全基因组关联研究中在自然种群中发现的突变进行比较。目标4：实验室参考菌株和自然分离株的分子工程。将从实验室菌株中剔除目标3中确定的候选基因，以便进行表型分析以量化对空气耐受性的影响。

Campylobacter jejuni (C. jejuni) is an enteric zoonotic pathogen that is one of the most common causes of gastroenteritis worldwide. Whilst C. jejuni infection is typically self-limiting in more affluent countries, it exerts a more significant burden in lower to middle-income countries (LMICs) due to chronic reinfection. C. jejuni colonises the guts of animals including chickens, cattle, and wild birds; and is transmitted via the consumption of undercooked meats and contaminated water sources. Due to the vast array of hosts and reservoirs C. jejuni occupies it is important to understand how C. jejuni spreads through a community and more specifically to understand survival and tolerance to environmental stressors within these transmission networks. The overall aim of this work is to understand C. jejuni response to environmental stressors that may contribute to the transmission of diarrhoeal disease in LMICs. This project will utilise isolates from sampling in The Gambia, Burkina Faso, and Ghana as part of a multi-university study called 'GetCampy Africa'. This will be achieved by the development of high-throughput protocols to assess C. jejuni phenotypes and genotypes responsible for tolerance to stressors, such as oxygen, antibiotics, and detergents. Results from these will inform experiments where I will evolve isolates under aerobic stress and identify adaptive mutations that confer aerotolerance. With the end goal of providing better resolution to, and understanding of, adaptive features that increase the transmissibility of C. jejuni that will ultimately inform transmission networks mapped from the larger 'GetCampy Africa' project.Objective 1: Optimise high-throughput growth and phenotypic characterization assays using lab reference and European C. jejuni strains. This method will form the basis of other protocols to assess the impact of environmental stressors on larger sets of C. jejuni Objective 2: Phenotyping and sequencing of C. jejuni strains isolated from the 'GetCampy Africa' Project. Using the methods developed in objective 1 assess variance in aerotolerance and aerobic growth to identify host and/or lineage-specific differences and similarities. Objective 3: Using experimental evolution to identify genes under selection for aerotolerance. Throughout the project evolution experiments will be run with control lines and experimental lines exposed to different environmental stressors- initially focussing on aerobic stress. Ancestral and evolved lines will be sequenced to identify genes that confer aerotolerance and compare them to mutations seen in natural populations identified from genome-wide association studies.Objective 4: Molecular engineering of lab reference strain and natural isolates. Candidate genes identified in objective 3 will be knocked 'in' or 'out' from lab strains to allow phenotypic analysis to quantify the impacts on aerotolerance.