一起肉毒梭菌中毒事件的实验室诊断
作者:
作者单位:

1.聊城市疾病预防控制中心,山东 聊城 252000;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.山东省疾病预防控制中心,山东 济南 250014

作者简介:

钱璐 女 主管技师 研究方向为微生物检验检测 E-mail:lccdcql@126.com

通讯作者:

窦继波 男 副主任医师 研究方向为微生物检验检测与诊断 E-mail:lccdcdjb@126.com

中图分类号:

R155

基金项目:

山东省医药卫生科技项目(202412060988)


Laboratory diagnosis of poisoning due to Clostridium botulinum
Author:
Affiliation:

1.Liaocheng Center for Disease Control and Prevention, ShandongLiaocheng252000, China;2.State Key Laboratory of Communicable Disease Prevention and Control, Institute for Communicable Disease Prevention and Control, Chinese Center for Disease Control and Prevention, Beijing102206, China;3.Shangdong Center for Disease Control and Prevention, Shandong Ji’nan250014, China

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    摘要:

    目的 对2023年11月发生的1起疑似肉毒中毒事件的相关样本进行实验室检测,对分离到的一株A型肉毒梭菌(C.botulinum)进行全基因组测序(WGS)。方法 对食品、病例粪便和环境样本应用实时荧光定量聚合酶链式反应(PCR)快速检测肉毒毒素基因。参照GB 4789.12—2016对样本进行前处理,经接种疱肉培养基和TPGYT培养基增菌培养5 d,通过动物试验进行毒素检出、确证和定性实验,采用卵黄琼脂平板分离和纯化菌株,并用API 20A和VITEK MS做菌种鉴定。同时对分离得到的菌株开展WGS,分析菌株的遗传特征。结果 病例粪便样本A型肉毒毒素基因阳性,通过动物试验检出A型肉毒毒素,利用卵黄琼脂平板成功分离A型肉毒梭菌RD1和RD2。其他样品均为阴性。WGS分析显示,该菌为肉毒梭菌,共有12个毒素编码相关基因,染色体上有3个噬菌体序列,wg-SNPs分析显示RD1和RD2与BrDura亲缘关系最近。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,检测方案和结果为临床A型肉毒中毒提供参考。

    Abstract:

    Objective Laboratory analyses were performed for a case of suspected botulinum poisoning incident that occurred in November 2023,and to analyze the whole genome of the strain of C.botulinum type A isolated.Methods Real time fluorescence quantitative polymerase chain reaction is applied to rapidly detect botulinum toxin genes in food, case feces and environmental samples. According to GB 4789.12—2016,the samples were subjected to pre-treatment.After being inoculated with blister meat culture medium and TPGYT culture medium for 5 d of enrichment, toxin detection,confirmation,and qualitative experiments were conducted through animal experiments.The strains were isolated and purified using egg yolk agar plates,and identified using API 20A and VITEK MS.Simultaneously,WGS was performed on the isolated strains to analyze their genetic characteristics.Results The fecal sample of the case was positive for type A botulinum toxin gene, and type A botulinum toxin was detected through animal experiments. C.botulinum type A RD1 and RD2 was successfully isolated using egg yolk agar plates,all other samples were negative.Whole genome sequencing analysis showed that the bacterium is C.botulinum,with a total of 12 toxin coding related genes and 3 phage sequences on the chromosome.Wg-SNPs analysis showed that RD1 and RD2 has the closest genetic relationship with BrDura.Conclusion This experiment was identified according to national standards and the fluorescence quantitative PCR method established in our laboratory. The detection plan and results provide reference for clinical type A botulism.

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钱璐,梁胜楠,董晶,徐雪芳,胡彬,张香媛,崔方元,程利红,窦继波.一起肉毒梭菌中毒事件的实验室诊断[J].中国食品卫生杂志,2024,36(8):916-921.

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  • 收稿日期:2024-01-25
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  • 在线发布日期: 2024-11-13
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