比較雙基因位點序列分型及脈衝電場膠體電泳技術研究中臺灣抗甲氧苯青黴素金黃色葡萄球菌之分子流行病學調查

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Title:	比較雙基因位點序列分型及脈衝電場膠體電泳技術研究中臺灣抗甲氧苯青黴素金黃色葡萄球菌之分子流行病學調查 Comparison of Double-Locus（clfB and spa）Sequence Typing with Pulsed-Field Gel Electrophoresis in Molecular Epidemiology of Methicillin-Resistant Staphylococcus aureus in Central Taiwan
Authors:	黃丞正;Huang, Chen-Cheng
Contributors:	臨床醫學研究所碩士班
Keywords:	抗甲氧苯青黴素金黃色葡萄球菌;雙基因位點序列分型;脈衝電場膠體電泳技術;Methicillin-Resistant Staphylococcus aureus;Double-Locus Sequence Typing
Date:	2012-07-18
Issue Date:	2012-08-31 16:35:32 (UTC+8)
Publisher:	中國醫藥大學
Abstract:	金黃色葡萄球菌(Staphylococcus aureus)常引發許多嚴重的疾病，如敗血症及血流的感染。幾個以DNA基因序列為主的方法被發展出來研究抗甲氧苯青黴素金黃色葡萄球菌之流行病學調查。我們從2008年7月至12月於中台灣一家醫學中心收集了91株來自不同病人之血液培養抗甲氧苯青黴素金黃色葡萄球菌菌株。每株菌株皆進行分子方法鑑定進行比較，包括spa分型(protein A gene, spa) 、clfB序列分型 (Clumping Factor B Gene , clfB)、雙基因位點分型(DLST，spa和clfB)、多基因位點序列分型(multi-locus sequence typing, MLST)、葡萄球菌夾式具抗藥基因染色體分型(staphylococcus cassette chromosome mec, SCCmec)及脈衝電場膠體電泳技術(pulse field gel electrophoresis, PFGE)。研究結果發現一共有5種主要clfB 型別(型別 A to E)，18種spa 型別，33種DLST基因型，5種SCCmec型別，17種pulsotypes(以80%相似度作分類)。其中有三個主要的DLST基因型 A1-t002 (佔29.7%), C0-t037 (佔16.5%), 及B1-t437 (佔14.3%)，兩個主要的pulsotype：types 6和8，和三個主要的clfB 型別：A、B和C。所有的clfB次型別A1及A2菌株皆為SCCmec type II, ST5。所有的clfB 型別C菌株皆為SCCmec type III, ST239。 88.9%的clfB 型別 B菌株為SCCmec type IV及VT，而所有的clfB 型別 B菌株為ST59。所有的clfB 次型別A1及A2（2-10型）及型別C菌株(3-50型)皆屬於院內型MRSA，而僅約30% clfB 型別B菌株屬於社區型MRSA。而攜帶clfB型別A或C的病人相較於clfB 型別B有較高的死亡率。另外，雙基因位點(DLST)鑑別度指數(ID)為0.8676,等同於脈衝電場膠體電泳分型(85%相似度)的鑑別度指數。結論：在區域性進行抗甲氧苯青黴素金黃色葡萄球菌之分子流行病學調查時，雙基因位點分型方法可達到和脈衝電場膠體電泳分型一樣的鑑別力，同時可以節省時間、成本、人力，是一個適合的PFGE替代方法甚至和死亡率有相關性。 Several DNA sequence-based methods have been developed for the epidemiological investigation of methicillin-resistant Staphylococcus aureus (MRSA). Ninety-one MRSA isolates were collected from the blood cultures of different patients from July to December of 2008 in central Taiwan. The molecular characteristics of each isolate, double locus sequencing typing (DLST, including spa and clfB typing) , multilocus sequencing typing (MLST), Staphylococcus cassette chromosome mec (SCCmec) and pulsed-ﬁeld gel electrophoresis (PFGE) were determined for comparison. There are 5 major clfB types (types A to E), 18 spa types, and 33 DLST genotypes, 5 SCCmec types, 17 pulsotypes with 80% similarity observed. Three major DLST (clfB and spa) genotypes A1-t002, C0-t037, and B1-t437, two major pulsotye：types 6 and 8, were identified. In the 3 major clfB types A, B and C, clfB subtype A1 and A2 isolates were all SCCmec type II (100%), ST5, and all clfB type C isolates were SCCmec type III (100%), ST239. Most clfB type B isolates（88.9%） were SCCmec type IV (59.3%) and VT (29.6%) and all belonged to ST59. All clfB subtype A1, A2 and type C isolates (100%) belonged to HA-MRSA, and about 30% clfB type B isolates also belonged to CA-MRSA. Patients carrying clfB type A or C have higher mortality than those with clfB type B, the index of discrimination (ID) of double-locus (spa and clfB) sequence typing was 0.8676, which has a comparable discriminatory power to 85% similarity in PFGE. In conclusion, this DLST method can give a compatible discriminatory power to PFGE with saving time, cost, and labor in local investigation of molecular epidemiology of MRSA. The method is a promising alternative to PFGE and even had a good asociation with mortality.
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