应用宏基因组测序技术区分黄热病毒野毒株和疫苗株

To distinguish wild type strain or vaccine strain of yellow fever virus using next generation sequencing

目的鉴别2019年11月在广州口岸入境的来自埃塞俄比亚发热病例是感染黄热病毒野毒株,还是接种黄热病毒疫苗。方法对入境发热病例的血清样本进行虫媒病毒荧光RT-PCR检测,使用Illumina宏基因组测序平台对血清样本进行宏基因组测序,分析其与黄热病毒野毒株和疫苗株的平均核苷酸一致性(ANIb)序列同源性和系统发育聚类情况。结果入境发热病例血液样本经实验室检测为黄热病毒核酸阳性,登革病毒、基孔肯雅病毒和寨卡病毒核酸阴性。宏基因组测序检出322条黄热病毒核酸序列,序列组装获得了181 bp DNA序列,非组装序列和组装序列的ANIb分析显示其与黄热病毒疫苗株相似性为96%~100%,而与黄热病毒野毒株序列相似性仅为79%~92%。181 bp组装序列的系统发育树显示其与黄热病毒疫苗株17D聚类,与野毒株不在同一个分支。结论宏基因组测序证实入境发热病例血液样本中没有检测到黄热病毒野毒株序列,该病例为接种黄热病毒疫苗株17D引起的发热。

Objective To identify whether the febrile case from Ethiopia who entered Guangzhou port in November2019 was infected by wild type strain or vaccine strain of yellow fever virus.Methods The serum samples of the imported case were detected for arboviruses by using Real-time RT-PCR.Sequences of yellow fever virus were ob-tained by next generation sequencing(NGS)with Illumina platform from the serum samples.The average nucleotide identity(ANIb)sequence homology and phylogenetic clustering were analyzed with wild type strain and vaccine strain of yellow fever virus.Results The imported case was positive for yellow fever virus nucleic acid,and negative for dengue virus,chikungunya virus and Zika virus.A piece of 322 yellow fever virus sequences were obtained by NGS,and 181 bp DNA sequence was assembled.ANIb sequence of the non-assembled sequences and the assembled sequences showed 96%-100%homology to vaccine strain,while only 79%-92%homology to wild strain of yellow fever virus.The phylogenetic tree based on the 181 bp assembled sequence showed that it clustered with 17 D vac-cine strain of yellow fever virus,and was not in the same branch with the wild type strain.Conclusion No wild type strain sequences of yellow fever virus was detected in the serum samples of imported case,his fever was caused by vaccination with 17 D vaccine of yellow fever virus.