八种灭活型病毒保存液对病毒核酸稳定性的影响

Effect of eight kinds of inactivated virus preservation solutions on the stability of virus nucleic acid

目的评价八种不同的灭活型病毒保存液对病毒核酸的保护效果。方法收集八种商业化的灭活型病毒保存液,分别将新冠病毒假病毒标准物质和禽流感病毒样本加入保存液中,同时设置2个对照组(生理盐水和磷酸盐缓冲液),在保存温度为24℃和37℃条件下,保存时间为0 h、6 h、12 h、24 h、48 h、72 h,分别提取各保存液中的病毒核酸,采用逆转录荧光定量PCR(RT-qPCR)和逆转录数字PCR方法(RT-dPCR)检测病毒核酸的含量变化。结果四种商业化病毒保存液A、E、G、H在不同温度和时间条件下对病毒核酸的保存效果明显。另外四种保存液保护效果较差:保存液B和C在37℃条件下12 h后保存效果变差,保存液D的保存效果在24℃和37℃下6 h后均明显减弱;保存液F对病毒核酸没有任何保护作用。结论病毒保存液的保护效果差会导致核酸检测的假阴性,建议开展核酸检测时对使用的病毒保护液进行质量控制。

Objective To evaluate the protective effects of 8 different inactivated virus preservation solutions on viral nucleic acids.Methods Eight commercial inactivated virus preservation solutions were evaluated by adding the pseudovirus standard material for SARS-CoV-2 and avian influenza virus,respectively.Two control groups(physiological saline and phosphate buffer)were set up at the same time.Reverse transcription real time quantitative PCR(RT-qPCR)and reverse transcription digital PCR(RT-dPCR)were used to quantify the nucleic acids extracted from the above treatment at 0 h,6 h,12 h,24 h,48 h,72 h at 24℃and 37℃.Results Four commercial virus preservation solutions(A,E,G,and H)could effectively protect the viral nucleic acids under different temperature and time conditions.However,the preservation solution B and C could not provide good protection after 12 h at 37℃.The protection effect of preservation solution D significantly decreased after 6 h at both 24℃and 37℃.Preservation solution F could not provide any protection on viral nucleic acid.Conclusions The poor protective effect of virus preservation solution can lead to false negatives in nucleic acid testing.It is recommended to control the quality of the virus preservation solution during nucleic acid testing.