| 多重荧光定量PCR检测婴幼儿腹泻病毒感染及其临床应用 |
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| 引用本文： | 张蝶1,2,卢晋英2,唐雪峰2,刘树业2,李会强1. 多重荧光定量PCR检测婴幼儿腹泻病毒感染及其临床应用[J]. 天津医科大学学报, 2021, 0(1): 83-87 |
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| 作者姓名： | 张蝶1 2 卢晋英2 唐雪峰2 刘树业2 李会强1 |
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| 作者单位： | 1.天津医科大学医学检验学院，天津 300203；2.天津市第三中心医院检验科，天津 300170 |
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| 摘 要： | 目的:建立一种能同时检测婴幼儿腹泻粪便中常见病毒的多重荧光定量 PCR技术。方法:根据GenBank上几种病毒基因组保守序列设计引物序列，建立多重荧光定量PCR方法，对所建立的多重荧光定量PCR方法的特异性、灵敏性及重复性进行验证；并以所建立的方法对150例婴幼儿病毒性腹泻患者粪便标本进行检测。结果:所建立的多重荧光定量PCR检测方法具有很好的特异性，灵敏性检测可达102拷贝/mL，检测不同病毒核酸浓度各自的检测 Ct 值标准差均较小，变异系数均低于1.0%，具有较好的重复性；检测150 份粪便标本，多重荧光定量 PCR 的检出率为36%，胶体金方法检出率为38.67%，两者比较差别无统计学意义（χ2= 6.91，P > 0.05）。多重荧光定量 PCR方法中轮状病毒、腺病毒、诺如病毒、星状病毒的检出率分别为12.67%、6.00%、13.33%和4.00%，测序结果与已知病毒株基因都具有高度同源性。结论:所建立的多重荧光定量PCR方法快速、特异、灵敏，可以作为临床病原诊断的一个重要工具且适用于流行病学调查研究。
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| 关 键 词： | 腹泻 病毒 轮状病毒 腺病毒 诺如病毒 星状病毒 多重荧光定量PCR |
The detection of viral infection in infants with diarrhea with multiplex fluorescent quantitative PCR and its clinical application |
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| ZHANG Die1,2,LU Jin-ying2,TANG Xue-feng2,LIU Shu-ye2,LI Hui-qiang1. The detection of viral infection in infants with diarrhea with multiplex fluorescent quantitative PCR and its clinical application[J]. Journal of Tianjin Medical University, 2021, 0(1): 83-87 |
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| Authors: | ZHANG Die1 2 LU Jin-ying2 TANG Xue-feng2 LIU Shu-ye2 LI Hui-qiang1 |
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| Affiliation: | 1. School of Medical Laboratory，Tianjin Medical University，Tianjin 300203，China；2. Clinical Laboratory，Tianjin Third Central Hospital，Tianjin 300170，China |
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| Abstract: | Objective: To develop and evaluate a multiplex fluorescence quantitative PCR assay for the simultaneous detection of common virus of faeces in infants with diarrhea. Methods: Genbank sequences of Rotavirus，Adenovirus，Norovirus and Astrovirus were included as reference sequences． A multiplex fluorescence quantitative PCR assay was developed and the primers and probes were designed based on the reference sequences，and the specificity，sensitivity and reproducibility of the assay were evaluated. Fecal samples from 150 patients with viral diarrhea were detected and verified by gene sequencing. Results: There were high specificity of the multiplex real-time PCR assay for detecting Rotavirus，Adenovirus，Norovirus and Astrovirus. The sensitivity of the method was 102 copies/mL. The standard deviations of CT values of different viral nucleic acid concentrations were small，and the coefficient of variation was less than 1.0%，which showed good repeatability. The detection rate of multiplex quantitative PCR was 36.00% in the 150 stool samples of infants with diarrhea，and that of colloidal gold method was 38.67%. There was no significant difference between the two methods. The detection rates of Rotavirus，Adenovirus，Norovirus and Astrovirus were 12.67%，6.00%，13.33% and 4.00%，respectively with multiplex fluorescent quantitative PCR. The sequencing results were highly homologous with the genes of known virus strains. Conclusion: Rotavirus，Adenovirus，Norovirus and Astrovirus can be detected and identified rapidly by the multiplex fluorescence quantitative PCR assay with high specificity and sensitivity. The assay developed in this study can be applied to the clinical diagnosis and epidemiological investigation. |
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| Keywords: | diarrhea virus rotavirus adenovirus norovirus astrovirus multiplex fluorescent quantitative PCR |
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