探討M4 A型鏈球菌之線毛在表皮感染中所扮演的角色

Investigate the Role of M4 Streptococcal Pili in Superficial Skin Infection

李紹暳(Shao-Hui Li)

指導教授 ： 張永祺

國立臺灣大學/醫學院/微生物學研究所/碩士(2017年)

https://doi.org/10.6342/NTU201702615

摘要

A型鏈球菌為常見的人類致病菌之一，能造成不同程度的感染，小至輕微的咽喉炎、皮疹，大至侵入性的壞死性筋膜炎、鏈球菌中毒性休克症候群。成功的貼附並侵入宿主表皮細胞是細菌得以造成後續的侵襲感染的重要步驟，目前已經知道A型鏈球菌可以表現M蛋白、細胞外基質結合蛋白、線毛等多項致病因子來增強其與宿主細胞結合的能力。細菌線毛於1949年首次在革蘭氏陰性菌被發現，為由非共價連結的蛋白次單位所組成的長鏈狀聚合體，具有貼附宿主細胞、形成生物膜、電子轉移、接合、運動，還有免疫調節的功能。然而在革蘭氏陽性菌上，則是近十年才開始蓬勃研究其線毛的作用。
組成A型鏈球菌線毛所需的基因皆聚在一個稱為FCT區域的基因座上，可轉錄出線毛的蛋白次單元及使它們之間產生共價鍵的轉肽酶。不同於革蘭氏陰性菌，革蘭氏陽性菌的蛋白次單位間為共價鍵結，形成的線毛更加纖細。實驗室之前所建構的A型鏈球菌M4血清型基因剔除株 (M4 Δspy0116) 係剔除一其具嵌入細胞壁功能的蛋白Spy0116，因此基因位於A型鏈球菌M4血清型的FCT區域上，故我們猜測突變株的線毛形成應會大受影響。我們的實驗結果發現，spy0116基因與其上下游的基因有被共轉錄的現象，M4 Δspy0116突變株確實缺少表面的長鏈狀構造，其形成生物膜與貼附細胞的能力亦較野生株遜色不少，在小鼠體內實驗中也觀察到此突變株較無法在小鼠皮下存活及造成傷口。綜合以上結果，我們成功的證明線毛對A型鏈球菌M4血清型要成功的造成表皮感染相關的各項機制均扮演重要角色。未來將繼續研究線毛的表現是否有可能會在其與宿主交互作用時受到影響。

關鍵字

A型鏈球菌 ； 線毛 ； 生物膜 ； 貼附 ； 表皮感染

並列摘要

Group A Streptococcus (Streptococcus pyogenes, GAS) is a severe and widespread human pathogen that cause mild diseases such as pharyngitis, impetigo as well as life-threatening necrotizing fasciitis and streptococcal septic shock syndrome. Successful colonization and invasion to host epithelial cell is a critical first step for succeeding invasive diseases. Multiple virulence factors have been reported in group A Streptococcus to strengthen its interaction with host cells, such as M protein, extracellular matrix-binding proteins and pili. Pili was a long non-covalent proteinaceous structure assembled on bacterial surface first observed in Gram-negative bacteria in 1949 and it is responsible for host cell adherence, biofilm formation, electron transport, conjugation, mobility and immunomodulation. However, functional characterization of pili in Gram-positive bacteria was just began in the past decade.
The genes encoding the streptococcal pilus proteins all cluster in a highly variable pilus island, the fibronectin-binding, collagen-binding, T antigen (FCT) region, which would translate into three pilus subunits and assembly enzymes, sortase. Unlike the non-covalent linkage of pili observed in Gram-negative bacterium, pili in Gram-positive bacteria are composed of covalently linked pilins which polymerize into thin rods with various length. The previously constructed serotype M4 streptococcal mutant, M4 Δspy0116, lacks the cell wall anchor protein encoded by a gene located in the FCT region, therefore, we speculated that the formation of pili in this M4 Δspy0116 mutant should be affected. We found that the M4 streptococcal pilus subunits and sortase in the FCT region can be co-transcribed into a polycistronic mRNA. In addition, our data demonstrated that the M4 Δspy0116 mutant lacks the long flexible polymers protruding from the bacteria, and are less capable to form biofilm and to adhere to human nasal septum cells. Moreover, in the in vivo murine subcutaneous infection model, the pilus-absent mutant resulted in mild skin infection with reduced bacterium recovery in the lesion compared to the parental wild-type bacteria. In summary, our data suggest that M4 streptococcal pili play a crucial role in all steps responsible for a successful streptococcal skin infection. In the future, we will analyze whether the expression level of pilus components can be modulated during its interplaying with host cells.

並列關鍵字

group A Streptococcus ； pilus ； biofilm ； adhesion ； skin infection

參考文獻

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