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題名 | Polymerase Chain Reaction to Detect Four Species of Herpesviral Infection by Using a Pair of Common Primers=以一對共同引子行聚合酶連鎖反應同時偵測四種疱疹病毒之感染 |
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作者 | 陳伯彥; 蔡啟仁; 劉武哲; 湯仁彬; 余國煥; | 書刊名 | 中華民國感染症醫學會雜誌 |
卷期 | 8:2 1997.12[民86.12] |
頁次 | 頁87-95 |
分類號 | 414.84 |
關鍵詞 | 共同引子; 聚合酶連鎖反應; 疱疹病毒; HSV-1; HSV-2; CMV; EBV; CHV-PCR; |
語文 | 英文(English) |
中文摘要 | 對於有免疫功能缺陷的病人來說,發展快速、簡單、有效、非侵犯性的□疹病毒群(herpesviruses)檢測方法,以提供感染時之早期診斷、治療及追蹤是非常需要的。本研乃以一對共同□疹病毒引子行聚合□連鎖反應(CHV-PCR, common herpesvirus-primer polymerase chain reaction)來同時偵測HSV-1, HSV-2, EBV, CMV四種病毒之DNA;並可藉由簡單之限制□-Sma I, Bam H I及Hinf I切割後,在洋菜膠電泳上進一判定是何種□疹病毒之感染。HSV-1的參考株KOS, CMV的參考株AD169, EBV的參考株B95-8及P3HR1之反應產物大小分別為518 bp, 589 bp及524 bp。而VZV的參考株WR; HHV-6的參考株U1102, HP及CTW; HHV-7的參考株YK則皆無產物出現。臺北榮民總醫院臨床病毒室由臨床病人所分離保存之三種□疹病毒中,22/23株HSV-1,21/21株HSV-2,20/21株CMV呈陽性反應。3株腺病毒,1株vaccina病毒,Hep-2及HEL細胞株則皆無產物。經三種限制□切割後,除3株CMV在Hinf I切割時因點突變而多一切位外,皆呈現一致而穩定切割結果。此共同□疹病毒引子聚合□連鎖反應並具有高度之敏感性可偵測少至1.3到4隻之病毒(病毒斑分析法)或100個□疹病毒基因。75個正常之腦脊髓液(CSF)經反應後,皆無產物出現。102個臨床病人的CSF經此PCR反應後,則有8個CSF有陽性產物。本研究證明此□疹病毒共同引子聚合□連鎖反應(CHV-PCR)具有很好的專一性及敏感性,將可提供臨床應用上快速而簡便的診斷方法。 |
英文摘要 | Facing a high morbidity and mortality rate of herpesviral infection, it is imperative to develop a simple, rapid, effective and non-invasive method to diagnose and treat patients as early as possible, especially for immunocompromised subjects. A pair of common primers, which is specific for 4 species of herpesvirus (namely HSV-1, HSV-2, CMV, EBV), derived from a highly conservative DNA polymerase gene region, was used in this common herpesvirus-primer polymerase chain reaction (CHV-PCR). Amplification products were analyzed by the digestion patterns of Sma I, Barn H I and Hinf I restriction enzymes. HSV-1, HSV-2, CMV and EBV could be then easily discriminated on a simple agarose mini-gel electrophoresis. All the herpesviral reference strains KOS (HSV-1), AD169 (CMV), B95-8 and P3HR1 (EBV) showed a positive amplification product on 518 bp, 589 bp and 524 bp separately, while U 1102 (HHV-6A), HP (HHV-6B), CTW (HHV-6), YK (HHV-7), WR (VZV) strains had a negative result. Twenty-two of 23 HSV-1, all of 21 HSV-2 and 20 of 21 CMV pooled clinical viral strains showed positive amplification results; while all of the three adenoviruses, one vaccinia virus and 2 cell lines (Hep-2 and HEL cell) yielded a negative amplification product. All of the amplification products showed a steady digestive pattern except for three strains of CMV, which showed an addititonal digestion site due to a point mutation. We also demonstrated a good sensitivity of this CHV-PCR to detect 1.3 to 4 viruses by using a viral plaque assay, and to detect about 100 viral copy numbers by using a semiquantitative method. Cerebrospinal fluid (CSF) samples from 79 patients with no known central nervous system (CNS) disease were used as the clinically negative control group, and all of them showed a negative result for the presence of herpesvirus. Eight CSF samples from 102 patients with known or unknown etiology of CNS disease showed a positive amplification product by using this CHV-PCR method. This CHV-PCR provided a rapid method to diagnose herpesviral infection with a high sensitivity and specificity. It was also a much simpler method to detect a group of microorganisms within a reaction as compared with other complicated PCR methods. |
本系統之摘要資訊系依該期刊論文摘要之資訊為主。