文献类型：期刊文献

中文题名：一例婴儿B型肉毒中毒的实验室诊断

英文题名：Laboratory identification of an infant botulism B case

作者：张爱佳[1];孙启杰[1];黄英[2];高鹏亚[3];刘志楠[4];彭筱婧[2];刘洋[1];徐雪芳[2];卓秀伟[5];张炜华[5];吴长德[3];卢选成[6]

第一作者：张爱佳

机构：[1]北京联合大学生物化学工程学院,北京100023;[2]中国疾病预防控制中心传染病预防控制所,传染病预防控制国家重点实验室,北京102206;[3]沈阳农业大学,辽宁沈阳110866;[4]中国检验检疫科学研究院,北京100176;[5]北京儿童医院,北京100034;[6]中国疾病预防控制中心实验动物中心,北京102206

第一机构：北京联合大学生物化学工程学院

年份：2020

卷号：35

期号：6

起止页码：557-562

中文期刊名：疾病监测

外文期刊名：Disease Surveillance

收录：CSTPCD

基金：国家重点研发计划(No.2018YFC1603800);启明星基金(No.201911417SJ127)。

语种：中文

中文关键词：婴儿肉毒中毒;实验室诊断;基因检测;毒素;荧光定量PCR

外文关键词：Infant botulism;Laboratory testing;Gene detection;Toxin;Real-time PCR

摘要：目的对2019年2月来自北京儿童医院1例疑似肉毒中毒的婴儿粪便样品进行实验室诊断,判定是否为肉毒毒素中毒。方法按照GB 4789.12—2016稀释处理样品,利用动物实验鉴定肉毒毒素、接种增菌培养基观察变化、采用卵黄培养基进行分离纯化,扩增测序16S rRNA基因并进行序列比对,采用荧光定量PCR方法进行毒素基因分型。结果粪便样本稀释后对小鼠进行腹腔注射,小鼠出现呼吸急促、腹式呼吸,并在4~6 h内死亡,呈现肉毒中毒典型症状。在卵黄培养基平板上分离到典型肉毒梭菌,镜检观察到芽孢形态,测得的16S rRNA基因序列与GenBank中公开的肉毒梭菌16S rRNA基因序列进行比对,相似率>99%,荧光定量PCR检测显示B型毒素基因阳性,判定该婴儿为B型肉毒中毒。结论此次实验按照国家标准和本实验室建立的荧光定量PCR方法进行鉴定,检测结果和诊断方案为我国婴儿B型肉毒中毒诊断提供参考,也为分析肉毒中毒地域性提供证据,为临床鉴定婴儿B型肉毒中毒提供参考案例。
Objective A laboratory test of the stool sample collected from a suspected infant botulism case in Beijing Children’s Hospital in February 2019 was conducted for the final confirmation.Methods The sample was diluted according to GB 4789.12—2016 for the botulinum toxin identification in animal experiments.The sample was inoculated in enrichment medium to observe its change and inoculated in yolk medium for strain isolation purification.PCR amplification technology was used to amplify 16 S rRNA,and the sequences of the target fragments were aligned.Results Mice were injected intraperitoneally with dilution of stool sample,and the mice developed shortness of breath,abdominal breathing and died within 4-6 hours,showing typical symptoms of botulism.The typical Clostridium botulinum was isolated on the yolk medium plate,and the spore morphology was observed by gram staining.The gene sequence alignment showed that the sequence of the16 S rRNA shared 99%similarity with that of C.botulinum.Fluorescent quantitative PCR showed that toxin gene B of C.botulinum was positive,and the infant was diagnosed with botulism B.Conclusion The test results and diagnosis protocol of this study provide reference for the diagnosis and prevention as well as of clinical identification of botulism B in infants in China.