Characterisation of mucoid Staphylococcus aureus recovered from the airways of cystic fibrosis patients: prevalence, impact on clinical course and detailed analysis by molecular and imaging techniques

金黄色葡萄球菌是最早从CF患者呼吸道中分离到的病原体之一。在许多患者中，尽管抗生素治疗和宿主防御，金黄色葡萄球菌仍持续存在数十年。在呼吸道持续生存期间，金黄色葡萄球菌需要适应这种敌对的生态位。就在最近，我们描述了粘液样金黄色葡萄球菌分离物，作为一种迄今尚未报道的表型，从CF患者的气道中恢复。这些黏液分离株在ica-操纵子的icaR-icaA基因间区携带5bp缺失，导致生物膜的高表达。到目前为止，粘液样金黄色葡萄球菌的患病率和对CF患者肺部疾病的影响都不清楚。此外，对气道内粘液样金黄色葡萄球菌表现的详细分析（例如厌氧条件、与铜绿假单胞菌粘液样/非粘液样的相互作用、痰液作为离体标本的空间分析）尚不清楚。此外，最近描述的没有5bp缺失的粘液样金黄色葡萄球菌分离株的潜在机制尚不清楚，将进一步表征，这些分离株也来自我们的CF患者。因此,在我们的提议,我们计划执行(1)临床项目的一部分来确定黏液状的金黄色葡萄球菌的流行隔离在一个横断面研究和识别的影响黏液状的金黄色葡萄球菌的临床病程肺病患者通过观察黏液状的金黄色葡萄球菌12个月相比,一群年龄、性别匹配组s和p . aeruginosa-positive /负面aureus-positive病人没有黏液状的隔离。在我们项目的第二部分，我们计划描述(2)在体外不同环境条件下（有氧与厌氧）气道内粘膜金黄色葡萄球菌分离物的性能，这些环境条件在生物膜形成、与宿主细胞相互作用和宿主防御方面模拟CF肺栖息地的体内情况。此外，(3)我们计划研究黏液分离株的潜在机制，因为我们无法使用下一代测序和RNA-seq分析在icaR-icaA基因间区检测到5bp缺失。最后，我们计划使用一种名为MiPACT的新型可视化技术，该技术可以表征黏液样/非黏液样金黄色葡萄球菌与宿主细胞和其他重要的CF病原体（如CF患者的离体标本中的铜绿假单胞菌）的相互作用。

S. aureus is one of the first isolated pathogens recovered from the airways of CF patients. In many patients, S. aureus persists for decades in spite of antibiotic treatment and host defence. During persistence in the airways, S. aureus needs to adapt to this hostile niche. Just recently we described mucoid S. aureus isolates as a so far not reported phenotype recovered from the airways of CF patients. These mucoid isolates carried a 5bp-deletion in the intergenic icaR-icaA region of the ica-operon leading to hyper-expression of biofilm. So far, neither the prevalence of mucoid S. aureus nor the impact on lung disease in CF patients is known. Furthermore, a detailed analysis of the performance of mucoid S. aureus within the airways (e.g. anaerobic conditions, the interaction with P. aeruginosa mucoid/non-mucoid, spatial analysis in sputum as an ex vivo specimen) is not clear. Moreover, the underlying mechanism of mucoid S. aureus isolates without a 5bp-deletion as recently described, which were also isolated from our CF patients, is not clear and will be further characterised.Therefore, in our proposal we plan to perform (1.) a clinical part of the project to determine the prevalence of mucoid S. aureus isolates in a cross-sectional study and the impact of identified mucoid S. aureus on the clinical course of the lung disease by observing patients with mucoid S. aureus for 12 months in comparison with a group of age-, gender- and P. aeruginosa-positive/negative matched group of S. aureus-positive patients without mucoid isolates. In the second part of our project we plan to characterise (2.) the performance of mucoid S. aureus isolates within the airways under different environmental conditions in vitro (aerobic versus anaerobic), which mimic the in vivo situation in the CF lung habitat in terms of biofilm formation, interaction with host cells and host defence. Furthermore, (3.) we plan to investigate the underlying mechanisms of mucoid isolates, for which we could not detect a 5bp-deletion in the icaR-icaA intergenic region using next generation sequencing and RNA-seq analysis. Lastly, (4.) we plan to use a new visualisation technique named MiPACT, which allows to characterise the interaction of mucoid/non-mucoid S. aureus with host cells and other important CF pathogens such as P. aeruginosa in sputa from CF patients as ex vivo specimens.