云南省昆明地区2022年柯萨奇病毒A16型的VP1区分子特征  

Molecular Characterization of the VP1 Region of Coxsackievirus 16 in Kunming,Yunnan in 2022

作　　者：张磊 沈茹 楚昭阳 马绍辉[2] 李丽[1] ZHANG Lei;SHEN Ru;CHU Zhaoyang;MA Shaohui;LI Li(Dept.of Clinical Laboratory,Kunming Maternal and Child Health hospital,Kunming Yunnan 650031;Institute of Medical Biology,Chinese Academy of Medical Sciences and Peking Union Medical College,Key Laboratory for Research and Development of Major Infectious Diseases Vaccine in Yunnan,Kunming Yunnan 650118,China)

机构地区：[1]昆明市妇幼保健院检验科,云南昆明650031 [2]中国医学科学院/北京协和医学院医学生物学研究所/云南省重大传染病疫苗研发重点实验室,云南昆明650118

出　　处：《昆明医科大学学报》2024年第7期73-78,共6页Journal of Kunming Medical University

基　　金：云南省科技计划项目(202202AA100016);昆明市卫生健康委科研项目(2022-11-01-001);科技人才与平台计划(202005AD160025)。

摘　　要：目的对昆明市某医院2022年从手足口病患儿分离的柯萨奇病毒A组16型(coxsackievirus A16,CVA16)的全VP1基因区分子特征分析。方法对昆明市某医院2022年从手足口病(hand foot mouth disease,HFMD)患儿200份临床样品粪便标本进行处理,分别通过RD细胞和Vero细胞分离,对经3次培养后仍产生细胞效应(cytopathogenic effect,CPE)的细胞培养液提取病毒核酸,然后用通用引物222/224经RT-PCR扩增并测序,序列经BLAST比对确定其血清型,对鉴定为柯萨奇病毒A组16型(coxsackievirus A16,CVA16)的再用特异引物CA16VP1F/CA16VP1R扩增其全VP1区并测序和拼接。下载CVA16病毒标准序列,采用Geneious 5.4.1和MEGA 6.05等软件对本研究CVA16的全VP1区的核苷酸和氨基酸序列分析。结果经鉴定后共检出22株从Vero细胞分离CVA16和6株RD细胞分离的CVA16毒株,进一步通过特异引物扩增获得21株可用的CVA16VP1全长序列,基于全VP1基因系统进化分析,这21株CVA16全属于B1a基因型。21株CVA16全长VP1基因之间的核苷酸和氨基酸序列同源性均较高,分别为91.9%~99.6%和89.2%~99.3%;与中国CVA16 B1a基因型的其他株的核苷酸和氨基酸序列同源性分别为89.7%~96.3%和89.2%~97.0%。与CVA16 B1b其他参考株对比,其核苷酸和氨基酸序列同源性分别在84.3%~87.2%和89.2%~97.0%。与其它CVA16的B1a基因亚型相比较发现21个氨基酸位点出现变异。结论2022年中国云南省昆明某医院引起HFMD的CVA16毒株属于B1a基因型,应继续监测以明确其流行特征。Objective To analyze the molecular characteristics of the whole VP1 gene region of coxsackievirus A16(CVA16)isolated from children with hand,foot and mouth disease(HFMD)in 2022 from a hospital in Kunming,Yunnan.Methods Two hundreds fecal specimens of HFMD clinical samples from 2022 in a hospital in kunming,were processed and isolated by RD cells and Vero cells,respectively,and viral nucleic acids were extracted from the cell cultures that still produced cytopathogenic effect(CPE)after three times of incubation,then amplified and sequenced by RT-PCR using universal primers 222/224,and sequenced,and the sequences were compared by BLAST to determine the serotypes,and then the full VP1 region of CVA16 was amplified with specific primers CA16VP1F/CA16VP1R,then sequenced and spliced.The standard sequences of CVA16 virus were downloaded,and the nucleotide and amino acid sequences of the whole VP1 region of CVA16 in this study were analyzed using software such as Geneious 5.4.1 and Mega 6.05.Results 22 strains of CVA16 isolated from Vero cells and 6 strains isolated from RD cells were detected by RT-PCR using universal primers,and the available sequences of 21 strains of full-length CVA16 VP1 region were further amplified by the specific primers of CVA16,and the nucleotide and amino acid sequences of the 21 strains of full-length CVA16 VP1 genes were highly homologous to each other,with 91.9%~99.6%and 89.2%~99.3%,respectively.The nucleotide and amino acid sequence homology with other strains of the chinese B1a genotype ranged from 89.7%~96.3%and 94.6%~99.0%,respectively;and nucleotide and amino acid sequence homology was in the range of 84.3%~87.2%and 89.2%~97.0%,respectively,when compared with the other reference strains of B1b.The phylogenetic analysis showed that all 21 CVA16 strains belonged to the B1a genotype.The 21 mutations were found by compared with that of other B1a strains.Conclusions The CVA16 strain that caused HFMD in a hospital in Kunming,Yunnan Province,China in 2022 belongs to the B1a genotype.Surveilla

关 键 词：柯萨奇病毒A组16型 全VP1基因 序列分析 

分 类 号：R373.2[医药卫生—病原生物学]