- 學位論文
非血檢體分離之甲氧西林敏感金黃色葡萄球菌之抗紅黴素基因分析
Erythromycin resistance genes in methicillin-susceptible Staphylococcus aureus isolated from non-blood specimens
摘要
金黃色葡萄球菌的抗藥性，對世界各地來說都是一項棘手的危機，紅黴素是臨床上常用來治療革蘭氏陽性球菌的藥物，紅黴素抗藥菌株問題也相當嚴重，因此，本研究選擇目前較少被注意到，但仍具有臨床重要性的非血來源MSSA，進行抗紅黴素菌株的相關探討。菌株收集2017年9月到2018年2月，經台大醫院細菌室自非血液檢體培養，並鑑定為抗紅黴素MSSA的分離株，總共有209株。利用PCR分析抗藥基因，以及用抗生素敏感性試驗確認抗藥表現型和紅黴素MIC後，發現帶msrA/B基因的有124株，佔總數的59%，其MIC值普遍低於帶erm基因的菌株，而M表現型則有128株(61%)。另以spa typing、PFGE和MLST分析實驗菌株的演化關係，發現最主要的pulsotype包含104株菌，屬於ST15，他們的spa type則是占最大宗的t084和t085，以上種種皆透露出有clonal spreading的狀況，在試圖尋找造成ST15具備感染優勢的因素後，發現可能與毒力因子相關，但仍有待深究。此外，本研究亦針對ermT基因做進一步鑽研，2017到2018半年間共收集到四株帶有ermT基因的MSSA，皆以染色體DNA為攜帶方式，且四株的sequence types都是ST398，此ST是目前帶有ermT的主流。 其中T17.56、T17.83、T18.05三株攜帶ermT的結構與實驗室於2011年發現的NTUH-8300相比，僅在rep基因周圍有些微差別，整體來說，皆為一段和pUR3912質體高度相似的序列，藉由IS431插入在細菌染色體DNA的transposase基因中；而T18.58帶ermT的完整結構尚未明瞭，只知道與目前已知者有很大的不同，包含一段同時比對到ST398質體和ST9染色體的序列，推測由外來DNA插入。除此之外，也承襲過去實驗室成員未完成的NTUH15-4 ermT結構解析，結果顯示由包含ermT基因的pUR3912質體，和pT45質體相鄰插入染色體DNA的pepT_2基因中，兩質體之間還存在一個抗藥基因aadD，而兩質體外側的序列比對到ST9 CC9金黃色葡萄球菌。特別的是，NTUH15-4為一株MSSA ST834-CC9，目前國內外尚未有ST834攜帶ermT基因的報導。
並列摘要
Drug resistance in Staphylococcus aureus is a major problem worldwide. Erythromycin plays an important therapeutic role in Staphylococcal infections. However, overuse of antibiotics for a long time led to resistant strains development. In consideration of the crisis, we performed an epidemiology investigation of MSSA isolated from non-blood specimens in northern Taiwan. Although the researches on MSSA were rarer than those on MRSA, we should still pay attention to MSSA due to its increasing rate of resistance. In the present study, a total of 209 non-blood isolates of erythromycin-resistant MSSA were collected from National Taiwan University Hospital from September 2017 to February 2018. After screening the erythromycin resistance genes by PCR and determining the MLSB resistance phenotypes by means of double disk diffusion assay, we found 124 (59%) isolates harbored msrA/B gene and M phenotype was predominant (128 of 209 isolates). We used PFGE, spa typing and MLST to analyze the molecular evolution of S.aureus. 104 (50%) isolates were grouped into the same pulsotype and typed as ST15-t084/t085. The results indicated clonal spreading during this period. The advantage of ST15 MSSA may be related to virulence factors. On the other hand, we also focused on one of the erythromycin resistance determinant, ermT. Four MSSA ST398 carried ermT gene. Three of them had the gene elements which were highly similar to NTUH-8300, a MSSA isolated from NTUH in 2011, except the rep gene region. This kind of element consisted of pUR3912 plasmid integrating into a transposon in chromosomal DNA. The other isolate, T18.58, carried ermT gene with another structure. It appeared that a plasmid inserted into chromosome DNA, but we haven’t identified it totally yet. Furthermore, our team found ermT gene in ST834-CC9 NTUH15-4 in 2015. Sequencing of the ermT and flanking regions in NTUH15-4 revealed that the ermT was located in chromosome. The element contained fragments from two plasmids, pUR3912 and pT45, and a tobramycin resistance gene, aadD. The integrated site is comparable to the pepT-2 gene in ST9 S. aureus.
參考文獻
延伸閱讀
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