低温保存对鼻咽拭子标本中百日咳博德特菌核酸检测和细菌培养结果的影响  

Effects of cryopreservation on the results of nucleic acid detection and culture of Bordetella pertussis in nasopharyngeal swab specimens

作　　者：刘志云 华春珍[2] 周瑾思 杨颖[2] 米芋枚[2] 周明明[3] 李建平[3] 李伟[3] 张学潮 Liu Zhiyun;Hua Chunzhen;Zhou Jinsi;Yang Ying;Mi Yumei;Zhou Mingming;Li Jianping;Li Wei;Zhang Xuechao(Department of Pediatrics,Hangzhou Fuyang District Hospital of Traditional Chinese Medicine,Fuyang 311400,China;Department of Infectious Disease,Children′s Hospital,Zhejiang University School of Medicine,Hangzhou 310052,China;Experimental Laboratory Center,Children′s Hospital,Zhejiang University School of Medicine,Hangzhou 310052,China;Hangzhou Center for Disease Control and Prevention,Hangzhou 310000,China)

机构地区：[1]杭州市富阳区中医院儿科,富阳311400 [2]浙江大学医学院附属儿童医院感染科,杭州310052 [3]浙江大学医学院附属儿童医院实验检验中心,杭州310052 [4]杭州市疾病预防与控制中心,杭州310000

出　　处：《中华检验医学杂志》2025年第2期265-269,共5页Chinese Journal of Laboratory Medicine

基　　金：浙江省基础公益研究计划(LGF18H010001);浙江省卫生健康科技计划(2021KY267)。

摘　　要：目的了解鼻咽拭子标本低温保存对百日咳博德特菌培养和核酸检测结果的影响,为标本集中转运检测提供依据。方法本研究为横断面研究,对2022年1—8月浙江大学医学院附属儿童医院门诊百日咳博德特菌培养阳性的鼻咽拭子标本,采用随机数字表法分别置-20℃和-70℃,冻存(114.3±31.9)d后复温集中进行培养和百日咳DNA的PCR检测,根据冻存温度和初始标本菌量进行分组分层,采用χ^(2)检验比较组间的阳性率。结果244份鼻咽拭子冻存后细菌培养阳性166例,阳性率较冻存前下降32%。其中冻存前菌量"少量"的84份鼻咽拭子冻存后培养阳性率下降了56%。但-70℃和-20℃冻存后培养阳性率差异无统计学意义(χ^(2)=1.65,P=0.20)。冻存前DNA检测阳性217例,阳性率88.9%,冻存后阳性率较冻存前下降10.6%。-70℃冻存组的阳性率高于-20℃组(χ^(2)=5.11,P=0.02)。菌量少量的标本?70℃冻存后的阳性率高于-20℃组(χ^(2)=4.86,P=0.03)。菌量一个"+"以上的标本-20℃和-70℃冻存后DNA检测阳性率差异无统计学意义(χ^(2)=1.25,P=0.26)。冻存后鼻咽拭子培养阳性率[68.0%(166/244)]低于DNA检测阳性率[78.3%(191/244),χ^(2)=6.52,P=0.01]。结论低温冷冻保存的鼻咽拭子可用于百日咳博德特菌核酸检测和细菌培养,原始标本菌量影响低温保存后的阳性检出率,核酸检测结果受影响更小,-70℃保存优于-20℃。Objective To investigate the effects of cryopreservation on the results of nucleic acid detection and culture for Bordetella pertussis in residual culture-positive nasopharyngeal swab specimens,aiming to provide the basis for specimens preservation,transport and centralized detection.MethodsIn this cross-sectional study,the residual nasopharyngeal swab specimens which were culture-positive for Bordetella pertussis were collected in the Children′s Hospital,Zhejiang University School of Medicine from January to August in 2022.The specimens were placed at‒20℃and‒70℃by random number table method,respectively.Re-detection by culture and PCR for Bordetella pertussis were conducted after these specimens were frozen for 114.3±31.9 days.The specimens were grouped according to the cryopreservation temperature and the semi-quantitative results by bacteria culture.The positive rates of the results were compared withχ^(2) test between groups.ResultsA total of 244 nasopharyngeal swabs specimens were included and 166 were culture-positive after cryopreservation,the positive rate decreased by 32%.Among them,the positive rate of re-culture of specimens containing low bacterial loads decreased by 56%after cryopreservation.However,there was no significant difference in the positive rate of culture between the specimens freezing at‒70℃and‒20℃(χ^(2)=1.65,P=0.20).The positive rate of DNA detection decreased by 10.6%(88.9%vs 78.3%)after cryopreservation.The positive rate of the‒70℃storage group was significantly higher than that of the‒20℃group(χ^(2)=5.11,P=0.02).The positive rate of the re-detection of DNA of nasopharyngeal swabs with low bacteria loads in‒70℃storage group was significantly higher than that of the‒20℃group(χ^(2)=4.86,P=0.03).While for the samples with a bacterial load of"+"or more,there was no significant difference in the positive rate of DNA detection after cryopreservation between the‒20℃and-70℃(χ^(2)=1.25,P=0.26)groups.The positive rate of nasopharyngeal swab culture a

关 键 词：博德特菌 百日咳 聚合酶链反应 培养 低温保存 

分 类 号：R725.1[医药卫生—儿科] R446.5[医药卫生—临床医学]