摘要
目的了解北京市甲型H1N1流行性感冒（简称流感）轻症与重症患者流感病毒的全基因组特性。方法选取北京市甲型H1Nl流感检测网络实验室于2009年6—12月在北京市定点医院采集的流感病例咽拭子标本作为研究对象，共21份，包括重症病例标本10份（其中有4份标本来源于死亡病例），轻症病例标本11份。由咽拭子标本中提取病毒核酸，设计基因组全长扩增引物，病毒核酸逆转录后进行PCR，对PCR产物测序，通过序列分析软件，分析病毒各基因进化和氨基酸变异情况。结果2009年北京市流行的甲型川Nl流感病毒8个基因节段与疫苗株A／California／07／2009（HlNl）相比，核苷酸同源性高达99％以上，未发生较大变异，其中，血凝素蛋白（HA）、神经氨酸酶（NA）、核蛋白（NP）基因的遗传距离相对较长，分别为0．0050，0．0040，0．0040；HA基因的P83S、1321V，NA基因的N248D，多聚酶（PA）基因的P224S，NP基因的V100I、L122Q变异在所有样本中均发生。重症病例的HA、NA、M、朋、多聚酶（PB2）、非结构蛋白（NS）基因具有明显基因进化聚集性；HA基因的S128P和$203T、PA基因的R269K和D547E、PB2基因的T588I、NS基因的1123V变异主要发生在重症病例中，分别有6、9、6、7、9、6例；其中HA基因的S203T、PA基因的R269K及D547E变异与重症病例的相关性具有统计学意义（P〈0．05）。HA基因变异主要分布在ca与cb抗原决定簇上，NA基因上无耐药突变，膜蛋白（M2）基因均发生耐药性突变，其余毒力相关基因未见突变。结论2009年北京市甲型HlNl流感病毒与疫苗株A／California／07／2009（H1N1）同源性较高，发现与重症及死亡病例相关的突变位点，未发现毒力基因突变。
Objective To explore the characteristics of the whole genome of the influenza H1N1 virus of the mild and severe cases in Beijing. Methods A total of 21 samples of throat swabs were collected from surveillance-designated hospitals between June and December in 2009, including 10 severe cases （4 death cases） and 11 mild cases. RNA of the virus were extracted,and the amplified primers of the whole genome were designed. Reverse transcription and PCR were performed to the RNA and then the PCR product was sequenced by software to analyze the evolution of the viral genes and the variation of the amino acids. Results Compared with the reference vaccine strain A/Califoruia/07/2009 （HIN1）, the genetic nucleotide homology in the eight segments of the pandemic H1N1 virus in Beijing in 2009 was higher than 99% ,without significant variation. Among them, the genetic distance of hemagglutinin （ HA）, neuraminida （NA） and nucleoprotein （NP） was comparatively far, separately 0. 0050,0. 0040 and 0. 0040. The gene of HA,P83S,the gene of NA, N248D, the gene of polymerase （PA）, P224S and the gene of NP, V100I and L122Q were found to mutate in all the samples. Genes of HA,NA,NP,PA,PB2 and nonstructural protein （NS1） in severe cases showed obviously clustered evolution. The mutation of gene S128P and $203T of HA, gene R269R and D547E of PA ,gene T588I of PB2 and gene I123V of NS mainly happened in severe cases,separately counting 6,9,6,7,9 and 6 cases. The relevance between the mutation happened in $203T of HA, R269K and D547E of PA and the severeness of the cases showed statistical significance （ P 〈 0. 05 ）. The mutations of HA gene were mainly on the Ca and Cb antigene domains. No drug resistant mutation was found on NA gene but happened on matrix protein 2 （ M2 gene）. None of the mutations were found on the virulence related genes. Conclusion A high homology was found between the pandemic H1 N1 virus in Beijing in 2009 and the reference vaccine strain A/California/07/2009 （ H1N1 ）. Mutational sites related with the severe and fatal cases were found, but not the virulence related mutation.
出处
《中华预防医学杂志》
CAS
CSCD
北大核心
2013年第5期420-426,共7页
Chinese Journal of Preventive Medicine
