摘要
目的对草莓中甲型肝炎病毒(hepatitis A virus, HAV)检测的两种检测方法进行优化比较, 以选出最佳的检测方法, 用于草莓中甲肝病毒的检测。方法向已知阴性冷冻草莓标本表面接种不同浓度的HAV, 优化碱性洗脱-PEG浓缩法中的牛肉浸出粉的浓度以及选择最适核酸提取试剂盒, 优化直接裂解法中的最适裂解缓冲液体积, 对草莓标本进行处理, 采用实时荧光定量RT-PCR检测回收的病毒量, 应用SPSS26.0对数据进行统计分析, 并将优化后的两种方法用于实际标本的检测。结果优化后的碱性洗脱-PEG浓缩法的牛肉浸出粉浓度选择3%, 病毒核酸提取试剂盒选择试剂盒B。优化后的直接裂解法的裂解缓冲液体积选择6 ml。对碱性洗脱-PEG浓缩法、直接裂解法在添加HAV不同浓度水平进行比较, 两种方法的HAV病毒回收率分别为21.50±1.06%、5.82±0.01%, 结果表明差异具有统计学意义。同时对四个地区共60份草莓标本进行检测, 结果均为阴性。结论优化后的碱性洗脱-PEG浓缩法灵敏度更高, 更适用于草莓标本中HAV的检测。
Objective The two detection method for the detection of hepatitis A virus(HAV)in strawberries were optimized and compared to select the best detection method for the detection of hepatitis A virus in strawberries.Methods Different concentrations of HAV were inoculated on the surface of known negative frozen strawberry specimens,the concentration of beef extract powder in alkaline elution-PEG concentration method was optimized,the optimal nucleic acid extraction kit was selected,the optimal lysis buffer volume in direct lysis method was optimized,and the recovered viral load was detected by Real-time fluorescence quantitative RT-PCR.SPSS26.0 was used to statistically analyze the data,and the optimized two method were used for the detection of actual specimens.Results The concentration of beef extract powder by the optimized alkaline elution-PEG concentration method was selected at 3%,and the viral nucleic acid extraction kit was selected as kit B.Six ml of lysis buffer was selected for the optimized direct lysis method.The recovery rates of HAV virus by alkaline elution-PEG concentration and direct lysis were compared with the HAV addition levels,and the HAV virus recovery rates of the two method were 21.50±1.06%and 5.82±0.01%,respectively,and the result showed that the differences were statistically significant.A total of 60 strawberry specimens from four regions were tested at the same time,and the result were all negative.Conclusions The optimized alkaline elution-PEG concentration method has higher sensitivity and is more suitable for the detection of HAV in strawberry specimens.
作者
焦梦琪
尹文娇
曹经瑗
毕胜利
Jiao Mengqi;Yin Wenjiao;Cao Jingyuan;Bi Shengli(National Institute for Viral Disease Control and Prevention,Chinese Center for Disease Control and Prevention,National Health Commission Key Laboratory of Medical Virology and Viral Diseases,Beijing 102206,China)
出处
《中华实验和临床病毒学杂志》
CAS
CSCD
2023年第1期83-88,共6页
Chinese Journal of Experimental and Clinical Virology
基金
国家卫生健康委疾控专项(131031103000200004)。
关键词
草莓
甲型肝炎病毒
前处理方法
实时荧光定量RT-PCR
Strawberry
Hepatitis A virus
Pretreatment method
Fluorescence quantitative real-time polymerase chain reaction
