利用探针捕获宏基因组测序法从咽拭子样本测定基孔肯雅病毒全基因组序列的个案报道

Case report on the identification of whole genome sequence of chikungunya virus using probe-capture metagenomics sequencing from throat swab sample

  • 摘要:
    目的 探索从咽拭子样本中检测基孔肯雅病毒(Chikungunya virus, CHIKV)的可行性。方法选取2025年5月北京口岸1例输入性基孔肯雅病毒感染者的血液和咽拭子样本,进行荧光定量逆转录聚合酶链式反应(quantitative reverse transcription polymerase chain reaction, RT-qPCR)检测,通过液相探针捕获结合高通量测序技术进行宏基因组测序,利用生物信息学方法组装并与公共数据库比对,分析遗传特征。结果采用RT-qPCR方法在咽拭子和血液样本均检出CHIKV核酸,CT值分别为28和26。咽拭子和血液样本宏基因组测序均检出CHIKV核酸序列,其中咽拭子样本检出CHIKV序列6757条,基因组覆盖度94.31%,血液样本检出CHIKV序列9558条,基因组覆盖度98.13%。两者与美国国家生物技术信息中心(National Center for Biotechnology Information, NCBI)数据库登录号PP896918.1和MH124579.1的CHIKV基因序列核苷酸同源性均大于99%。最大似然法系统进化树分析显示,两者与NCBI数据库中PP896921.1和MW281311.1聚类在同一分支,均属东/中/南非洲遗传谱系(ESCA基因型)。
    结论 咽拭子样本可检出CHIKV全基因组序列,为婴幼儿等采血困难人群中疑似病例实验室诊断提供了基于无创性样本检测的证据。

     

    Abstract: Objective Exploring the feasibility of detecting Chikungunya virus (CHIKV) from throat swab samples. Methods Blood and throat swab samples were collected from an imported case of Chikungunya virus infection at Beijing port in May 2025. Both samples were tested using quantitative real-time RT-PCR. Additionally, liquid-phase probe capture combined with high-throughput sequencing technology was employed for metagenomic sequencing. Bioinformatics methods were used for sequence assembly and comparison with public databases to analyze genetic characteristics. Results CHIKV nucleic acid was detected in both throat swab and blood samples by fluorescence quantitative RT-PCR, with CT values of 28 and 26 respectively. CHIKV nucleic acid sequences were also detected in both throat swab and blood samples through metagenomic sequencing..The throat swab sample yielded 6,757 CHIKV sequences with a genome coverage of 94.31%, while the blood sample yielded 9,558 CHIKV sequences with a genome coverage of 98.13%. Both samples showed nucleotide sequence homology greater than 99% with CHIKV metagenomic sequences from the NCBI database (accession numbers PP896918.1 and MH124579.1). Maximum likelihood phylogenetic tree analysis revealed that both samples clustered in the same branch as NCBI database sequences PP896921.1 and MW281311.1, belonging to the East/Central/South African genetic lineage (ESCA genotype). Conclusion The detection of the whole genome sequence of CHIKV in nasopharyngeal swab samples provides evidence for the laboratory diagnosis of suspected cases in populations where blood collection is challenging, such as infants and young children, using non-invasive sampling methods.

     

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