Molecular basis of bacterial small-colony variants / Base moléculaire des variants à petites colonies chez les bactéries

金黄色葡萄球菌（Staphylococcus aureus，SA）和耐甲氧西林金黄色葡萄球菌（Antibiotic resistant methicillin-resistant SA，MRSA）常常引起慢性和难以治疗的感染。SA/MRSA能够产生小菌落变体（SCV），这与持续感染有关。金黄色葡萄球菌/耐甲氧西林金黄色葡萄球菌转换为SCV的能力，反之亦然，似乎是导致持续性感染过程的一个组成部分。与它们的正常对应物相比，细菌SCV产生大量的保护性生物膜（细菌嵌入其中的细胞外基质）。SCV还具有在宿主细胞内存活和持续存在的强大能力，这是一种保护它们免受宿主免疫反应的特性。转向SCV也促进了抗生素耐药性的发展，使SA/MRSA感染更加难以治疗。本研究计划旨在研究SCV形成的分子基础。*具体目标是：1）通过全基因组测序研究SCV形成的分子基础（DNA和RNA）; 2）研究通过SCV发生的抗生素抗性发展的分子基础; 3）研究SCV形成对哺乳动物宿主中的毒力和持久性的影响;和4）建立针对SCV的未来策略的分子基础。在目标1和2中，将鉴定导致SCV表型的基因组和转录组改变。我们希望找到不同的途径导致SCV的形成，因为存在几种表型表达。然后，我们将使用各种体外，细胞和动物模型来研究SCV形成的不同分子机制如何影响宿主中的毒力和持久性（目标3）。 目标4将建立在这些结果的基础上，以研究和建立控制SCV的新方法的分子基础，例如开发专门针对细胞内SA的疫苗。慢性肺部感染是囊性纤维化（CF）患者发病和死亡的最常见原因。SA/MRSA总体上是CF患者中最常分离的病原体，SCV有助于感染的持续性。研究已经证明了SCV和MRSA分别对儿童和成人CF的有害影响。 SCV还涉及其他疾病如牛乳腺炎的持续存在。 牛奶生产是加拿大的重要产业。奶牛群中与乳腺炎相关的总损失估计每年超过4亿美元，SA是加拿大牛乳腺炎的主要原因。 迄今为止，没有SA疫苗显示出保护性功效，部分原因是免疫应答不足以对抗细胞内形式的SA。 随着对解释SCV形成的基本机制的理解，这项研究计划将有助于未来开发预防SCV形成和SA/MRSA持久性的特定抗生素或疫苗。

Staphylococcus aureus (SA) and antibiotic resistant methicillin-resistant SA (MRSA) often cause chronic and difficult-to-treat infections. SA/MRSA are able to generate Small-Colony Variants (SCVs), which have been linked to persistent infections. The ability of SA/MRSA to switch to SCVs and vice versa seems to be an integral part the infection process leading to persistence. Compared to their normal counterparts, bacterial SCVs produce high amounts of protective biofilms (extracellular matrix in which bacteria are embedded). SCVs also have a great ability to survive and persist within host cells, a property that protects them from the host immune response. The switch to SCVs also facilitates the development of antibiotic resistance making SA/MRSA infections even more difficult to treat.*******This research program aims to study the molecular basis of SCV formation.*******The specific objectives are to: 1) Investigate the molecular basis of SCV formation through whole-genome sequencing (DNA and RNA) of genetically-related "SA-SCV" pairs; 2) Investigate the molecular basis of antibiotic resistance development occurring through SCVs; 3) Study the impact of SCV formation on virulence and persistence in the mammalian host; and 4) Establish the molecular basis of future strategies targeting SCVs.*******In Objectives 1 and 2, genome and transcriptome alterations responsible for the SCV phenotype will be identified. We expect finding different paths leading to SCV formation since several phenotypic expressions exist. We will then use a variety of in vitro, cellular and animal models to investigate how the different molecular mechanisms of SCV formation impact on virulence and persistence in the host (Objective 3). Objective 4 will built on these results to investigate and establish the molecular basis of new approaches to control SCVs such as the development of vaccines specifically tailored to target intracellular SA.*******Chronic lung infection is the most common cause of morbidity and mortality in cystic fibrosis (CF) patients. SA/MRSA is overall the most frequently isolated pathogen from CF patients and SCVs contribute to the persistence of infection. Studies have demonstrated the deleterious effects of SCVs and of MRSA on CF in children and adults, respectively. SCVs are also involved in the persistence of other diseases like bovine mastitis. Milk production is an important industry in Canada. Total losses linked to mastitis in dairy herds are estimated at over $400 million per year and SA is the leading cause of bovine mastitis in Canada. No SA vaccine has shown protective efficacy to date, in part because of the inadequacy of the immune response to counteract intracellular forms of SA. With the understanding of the fundamental mechanisms explaining the formation of SCVs, this research program will help the future development of specific antibiotics or vaccines that prevent SCV formation and SA/MRSA persistence.******