口罩病毒提取法检测呼吸道病毒的可行性及精确性验证  

Feasibility and accuracy verification of respiratory virus detection by mask virus extraction

作　　者：闫丽平[1] 徐丽娜[2] 冯静[2] 王育慧[2] 丁秀萍 赵雪丽[2] 厉锋[3] YAN Li-ping;XU Li-na;FENG Jing;WANG Yu-hui;DING Xiu-ping;ZHAO Xue-li;LI Feng(Weifang People's Hospital,Weifang,Shandong 261041,China)

机构地区：[1]潍坊市人民医院医院感染管理科,山东潍坊261041 [2]潍坊市人民医院医院呼吸内科,山东潍坊261041 [3]潍坊市人民医院医院脊柱外科,山东潍坊261041

出　　处：《中华医院感染学杂志》2022年第12期1880-1884,共5页Chinese Journal of Nosocomiology

摘　　要：目的 探讨一种通过提取口罩残留病毒来检测呼吸道病毒的新方法,并且验证其可靠性和灵敏度。方法将不同稀释浓度的新型冠状病毒类似物--鸡新城疫病毒La Sota株及传染性支气管炎病毒H120株的稀释液通过呼吸模拟器喷洒至医用外科口罩及N95口罩,在室温环境中分别静置2 h和12 h,利用普通PCR技术及实时荧光定量PCR技术检测鸡新城疫病毒N基因条带和传染性支气管炎病毒NP基因条带及其扩增循环数(CT);对比不同存留时间下口罩中最低可检测病毒浓度及病毒含量。结果 不同时间存留的口罩均可以检出新型冠状病毒类似物--鸡新城疫病毒La Sota株及传染性支气管炎病毒H120株的基因条带,且病毒总RNA在10 pg~10 ng的范围内均有较好的扩增曲线。2 h内提取的病毒残留物,普通医用外科口罩和N95口罩检测N基因、NP基因的CT均值分别为22.55±0.34、23.70±0.50和22.86±0.31、24.04±0.34;但12 h后只有部分可被检测到,且两种口罩在相同时间内CT值无统计学差异(P=0.452、P=0.355)。口罩中最低可检测病毒的稀释浓度为1∶800,可检测残存病毒数量约为6.75×10^(3)个。结论 通过检测2 h内口罩中残留病毒来筛查冠状病毒的方法是可行的,且适用于医用外科口罩及N95口罩,该方法可以用于呼吸道病毒的初步筛查。OBJECTIVE To explore a new method for detecting respiratory viruses by extracting residual virus on mask, and verify its reliability and sensitivity. METHODS The novel coronavirus analogs-s La Sota strains of chicken Newcastle disease virus and H120 strains of infectious bronchitis virus with different diluted concentrations were sprayed onto surgical masks and N95 masks through a respiratory simulator, and they were left standing at room temperature for 2 hours and 12 hours, respectively. The cDNA and its amplification cycle(CT) values of the nucleoocapsid protein(N) of chicken Newcastle disease virus and the nucleoprotein(NP) genes of infectious bronchitis virus were detected by ordinary polymerase chain reaction(PCR) and quantitative real-time PCR(qRT-PCR). The minimum detectable virus concentration and viral content in masks under different retention times were compared. RESULTS The gene bands of the Newcastle disease virus La Sota strains and the infectious bronchitis virus H120 strains were detected on the masks stored for different times, and the total RNA of the virus had good amplification curves in the range of 10 pg-10 ng. The mean CT values of N gene and NP gene of the residual virus on the general medical surgical mask and N95 masks placed for 2 h were 22.547±0.342,23.698±0.501 and 22.855±0.308,24.036±0.338, respectively. However, only part of them could be detected after 12 h. respectively, and there was no significant difference in CT values between the two masks during the same period of time(P=0.452, P=0.355). The minimum detectable concentration of virus in the masks was 1∶800, and the number of residual viruses on the mask that can be detected was 6.75×10^(3). CONCLUSION The method of screening coronavirus by detecting virus residues on masks within 2 hours was feasible and suitable for medical surgical masks and N95 masks, which can be used for preliminary screening of respiratory viruses.

关 键 词：呼吸道病毒 病毒检测方法 N基因 NP基因 

分 类 号：R373.1[医药卫生—病原生物学]