甲型肝炎病毒四种核酸检测方法的比较  

Comparison of four nucleic acid detection methods for hepatitis A virus

作　　者：石峰 曹经瑗[1] 邱丰[1] 尹文娇[1] 周文亭[1] 毕胜利[1] Shi Feng;Cao Jingyuan;Qiu Feng;Yin Wenjiao;Zhou Wenting;Bi Shengli(National Institute for Viral Disease Control and Prevention,Chinese Center for Disease Control and Prevention,Beijing 102206,China)

机构地区：[1]中国疾病预防控制中心病毒病预防控制所,北京102206

出　　处：《中华实验和临床病毒学杂志》2021年第2期213-217,共5页Chinese Journal of Experimental and Clinical Virology

基　　金：国家科技重大专项(2018ZX10201002-004-003)。

摘　　要：目的比较甲型肝炎病毒(hepatitis A virus,HAV)四种核酸检测方法。方法采用A、B、C实时荧光定量RT-PCR(RT-qPCR)及D微滴芯片数字RT-PCR(RT-dPCR)分别对HAV质粒标准品、梯度稀释的HAV疫苗进行灵敏度检测;对相关病毒核酸进行特异性检测;用A、B、C方法对40份人工污染HAV牡蛎、市售牡蛎及血清标本、HAV疫苗标本进行检测,比较检出率;比较A、D方法对低浓度HAV人工污染牡蛎的回收率。结果 A、B方法对质粒标准品均可检测至10拷贝/μL;检测梯度稀释的HAV疫苗,A、B、C方法的斜率、R^(2)值、扩增效率(-3.446~-3.297、0.991~0.998、95.07%~101.051%)均在可接受范围;40份不同来源的标本,A、B、C方法的阳性检出率分别是50%(20/40)、47.5%(19/40)、55%(22/40),差异无统计学意义(χ^(2)=0.467,P=0.792);A、D方法对梯度稀释疫苗检测灵敏度无明显差别,对低浓度HAV人工污染牡蛎检测,D方法回收率高于A方法,但差异无统计学意义(F=0.294,P=0.642)。结论 A、B、C方法无明显差异,较方便快捷;在检测低浓度HAV污染的食品时,D方法略有优势,但检测成本稍高,选取检测方法可根据实际情况选择。Objective To compare the four nucleic acid detection method of hepatitis A virus.Methods Using method A,B,and C real-time fluorescent quantitative RT-PCR(RT-qPCR)and method D droplet chip digital PCR(RT-dPCR)to detect the sensitivity of HAV plasmid and gradient dilution HAV vaccine respectively.Specific detection of related viral nucleic acid was performed.Methods A,B,and C were used to detect 40 artificially contaminated HAV oysters,commercially available oysters and serum samples,and HAV vaccine samples,and compare the detection rates.The recovery rates of method A and D on artificially contaminated oysters were compared with low concentration of HAV.Results Both method A and B could detect HAV plasmids up to 10 copies/μL.In the detection of HAV vaccine with gradient dilution,the slope,R2 value and amplification efficiency of method A,B,and C were all within the acceptable range(-3.446~-3.297,0.991-0.998,-95.07%-101.051%).For 40 specimens from different sources,the positive detection rates of method A,B,and C were 50%(20/40),47.5%(19/40),55%(22/40),and the difference was not statistically significant(χ^(2)=0.467,P=0.792).Methods A and D have no significant difference in the detection sensitivity of gradient dilution vaccines.For the detection of artificially contaminated oysters with low concentration of HAV,the recovery rate of method D was higher than that of method A,but the difference was not statistically significant(F=0.294,P=0.642).Conclusions There is no significant difference between method A,B,and C,which is more convenient and fast.When detecting low concentrations of HAV in food,Methods D had a slight advantage,but the detection cost is slightly higher.The detection method can be selected according to the actual situation.

关 键 词：肝炎病毒 甲型 实时荧光定量RT-PCR 微滴芯片数字RT-PCR 

分 类 号：R373.21[医药卫生—病原生物学]