臺灣博碩士論文加值系統

A型鏈球菌所引起的人類疾病種類相當廣泛，由最常見的上呼吸道感染到最嚴重的敗血性休克症候群都有可能產生，甚至會引起自體免疫的後遺症。雖然A型鏈球菌有其臨床的特殊性，但其致病機轉至今尚未完全釐清；過去的研究顯示熱源性外毒素(streptococcal pyrogenic exotoxins; Spes)和其疾病的嚴重程度有相當密切的關係，而其中的Spe B除了具有熱源性外毒素之性質外，亦為一蛋白水解酶(cysteine protease)； Spe B在A型鏈球菌造成之嚴重侵入性感染的情況下扮演相當重要的角色。因此，我們將著重探討臨床疾病的嚴重程度和菌株的熱源性外毒素基因及蛋白水解酶之間的關係。

本實驗中，我們收集從1999至2005年之間289株A型鏈球菌的臨床菌株，利用聚合酶連鎖反應來偵測六種熱源性外毒素基因(spe A、spe B、spe C、spe F、spe G、sme Z)。此外，我們將菌株的DNA定序之後和基因庫的emm基因序列比對，以建立不同的emm型別。我們還利用酪蛋白平板(Casein plate)水解能力測試蛋白酶的活性(cysteine protease activity)。實驗結果顯示，這289株菌株共有28種emm型別，其中以Sp11014/VT5，M11和M 49 emm type最為常見。不同的emm 型別所各自攜帶的六種外毒素基因，均有其特殊的一或
兩種排列組合方式；外毒素基因的出現頻率在相異之emm型別也有其高低差異。

我們也檢定外毒素B (Spe B)，即所謂的半胱氨酸蛋白酶(cysteine protease)的活性，發現侵入性與非侵入性菌株之間的蛋白酶活性並沒有統計學上顯著的差異。

以上的發現，可以提供我們關於臨床上A型鏈球菌感染的嚴重度和不同emm 型別的外毒素基因排列方式，還有蛋白酶活性之間的相互關係之釐清。

In the present study, we screened the distribution of streptococcal pyrogenic exotoxins (spe) of 289 endemic group A streptococcal (GAS) isolates and clinical features were reviewed within a 7-year period in an medical center. We performed polymerase chain reaction (PCR) to detect 6 streptococcal pyrogenic exotoxins (speA, speB, speC, speF, speG, and smeZ). By use of DNA sequencing-based methods approach, 28 different emm types have been identified. In the study, sp11014, M11, and M49 emm protein types were more frequently to GAS isolates. The predominant emm types were different from other area of the world. Analysis revealed that either invasive or noninvasive GAS isolates do not share a common spe-gene profile. Although characteristic spe-gene profiles were detected in most emm types, a predominance of 1 or 2 spe-gene profile could be observed. Interestingly, individual spe gene showed a nonrandom distribution of key spe genes characteristic of the specific emm type. We further examined streptococcal pyrogenic exotoxin B (Spe B), a streptococcal cysteine protease which is believed to be important in group A streptococcal (GAS) pathogenesis. The protease activity of these isolates was tested by the casein plate method. Of the 289 strains with invasive (94 strains) and noninvasive (195 strains) diseases, there is no statistical difference of protease activity. These results demonstrate that no particular emm strain or level of protease activity had associated with severe clinical condition. These findings may have important implications for the role of spe-gene profiles and cysteine protease activity in host-pathogen interactions via the expression of GAS virulence genes and the severity of GAS disease.

目 錄
頁碼
第一章 中文摘要 1
第二章 英文摘要 3
第三章 緒論
第一節 A型鏈球菌之定義 5
第二節 A型鏈球菌臨床表現 5
第三節 A型鏈球菌流行病學 6
第四節 A型鏈球菌之致病機轉 7
第五節 研究動機與目的 12
第四章 材料與方法
第一節 病人收案與菌體收集標準 14
第二節 A型鏈球菌的培養及保存 14
第三節 菌株型別和熱源性外毒素基因的鑑定 15
第四節 DNA的定序和emm基因序列比對 15
第五節 半胱氨酸蛋白酶活性的定義 16
第六節 統計方法 16
第五章 實驗結果
第一節 A型鏈球菌之臨床表徵 18
第二節 年齡的影響 18
第三節 半胱氨酸蛋白酶活性的影響 19
第四節 病人潛在性的疾病之影響 19
第五節 A型鏈球菌之抗藥性 19
第六節 侵入性與非侵入性菌株的臨床表現及
spe基因的分布和抗藥性 20
第七節 從敗血性休克的角度來探討不同外毒素基因
和emm type 20
第八節 侵入性與非侵入性菌株之間的
半胱氨酸蛋白酶之活性比較 21
第九節 不同的emm type所含外毒素基因種類之異同 21
第六章 討論
第一節 半胱氨酸蛋白酶的活性 22
第二節 抗藥性 24
第三節 六種毒性基因在不同的emm type之間呈現非機率性的分布 24
第七章 結論與展望
第一節 結論 26
第二節 展望 27
第八章 參考文獻 28
第九章 圖表
表一、A型鏈球菌引起的疾病與分布關係 34
表二、A型鏈球菌感染之病患年齡和造成侵入性感染、
敗血性休克或病患有潛在性疾病的關係 35
表三、半胱氨酸蛋白酶的活性和病患侵入性感染、
敗血性休克或病患有潛在性疾病的關係 36
表四、病患潛在性疾病和毒性基因，emm type之間的關係 37
表五、A型鏈球菌造成侵入性感染與毒性基因，emm type之間的關係 38
表六、A型鏈球菌造成敗血性休克與毒性基因，emm type之間的關係 39
表七、侵入性或非侵入性A型鏈球菌之間不同抗生素的抗藥性比較 40
表八、半胱氨酸酶的活性在emm type所造成侵入性
與非侵入性感染的比較 41
表九、八株最常見之emm type的毒性基因排列方式 42
圖一、不同月份侵入性和非侵入性A型鏈球菌的感染率 44
圖二、感染A型鏈球菌的病患年紀分布之百分比 45
圖三、不同emm type對不同抗生素的抗藥性比較 46
圖四、八株最常見之emm type的半胱氨酸蛋白酶活性 47
圖五、八種常見的emm type的毒性基因分布圖 48
圖六、不同的emm type在侵入性與非侵入性菌株之間，毒性基因的分布 49

第八章 參考文獻

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