CAJ | 학술논문

目的：调查杭州地区2009—2014年间人博卡病毒1型（ human bocavirus 1， HBoV1）全基因组序列的遗传变异和分子进化特征。方法采集2009—2014年间浙江大学医学院附属儿童医院急性呼吸道感染患儿咽拭子标本，采用实时荧光PCR检测HBoV1，最终挑选15株病毒载量高的阳性样本进行全基因组扩增和测序。测序结果上传至GenBank，利用生物信息学软件分析序列。结果15株HBoV1病毒全基因组序列共检测到48个核苷酸突变，最终导致11个氨基酸变化，其中5个位点处于磷脂酶（ PLA2）活性区。基于全基因组编码序列的进化分析表明HBoV1可以分为3个分支，本研究的15株病毒全部属于分支1，与瑞典代表株ST2属于同一簇。另外，进化树构建结果表明VP1／VP2基因可以替代全基因编码序列来构建HBoV1的进化树。 HBoV1全基因组序列的进化速率为每年3．03×10－4（95％HPD，2．14×10－4～3．92×10－4）突变点／位点，对于单个基因，NS1的进化速率最慢，而NP1的进化速率最快。 HBoV1四个基因的选择压力ω值都小于1，表明所有基因都处于纯化选择下，其中VP2的纯化选择压力最强，而NP1的纯化选择压力最弱。结论杭州地区2009—2014年间流行的HBoV1均属于ST2基因型，PLA2活性区变异率相对较高。全基因组序列虽然保守，但进化速率很快，其中NP1基因进化速率最快。4个基因均处于纯化选择压力下，其中VP2基因的纯化选择压力最强。
목적：조사항주지구2009—2014년간인박잡병독1형（ human bocavirus 1， HBoV1）전기인조서렬적유전변이화분자진화특정。방법채집2009—2014년간절강대학의학원부속인동의원급성호흡도감염환인인식자표본，채용실시형광PCR검측HBoV1，최종도선15주병독재량고적양성양본진행전기인조확증화측서。측서결과상전지GenBank，이용생물신식학연건분석서렬。결과15주HBoV1병독전기인조서렬공검측도48개핵감산돌변，최종도치11개안기산변화，기중5개위점처우린지매（ PLA2）활성구。기우전기인조편마서렬적진화분석표명HBoV1가이분위3개분지，본연구적15주병독전부속우분지1，여서전대표주ST2속우동일족。령외，진화수구건결과표명VP1／VP2기인가이체대전기인편마서렬래구건HBoV1적진화수。 HBoV1전기인조서렬적진화속솔위매년3．03×10－4（95％HPD，2．14×10－4～3．92×10－4）돌변점／위점，대우단개기인，NS1적진화속솔최만，이NP1적진화속솔최쾌。 HBoV1사개기인적선택압력ω치도소우1，표명소유기인도처우순화선택하，기중VP2적순화선택압력최강，이NP1적순화선택압력최약。결론항주지구2009—2014년간류행적HBoV1균속우ST2기인형，PLA2활성구변이솔상대교고。전기인조서렬수연보수，단진화속솔흔쾌，기중NP1기인진화속솔최쾌。4개기인균처우순화선택압력하，기중VP2기인적순화선택압력최강。
Objective To investigate the genetic variation and molecular evolution of human bo-cavirus 1 (HBoV1) strains isolated during 2009 to 2014 in Hangzhou, China.Methods Throat swab sam-ples were collected from children with acute respiratory tract infections in the Children′s Hospital Affiliated to the Zhejiang University School of Medicine from 2009 to 2014.Real-time PCR was performed for the detec-tion of HBoV1 strains.Fifteen HBoV1 strains with high virus load were screened out for the amplification and sequencing of complete genomes.The complete genomes were submitted to GenBank for further analysis with bioinformatics software.Results A total of 48 nucleotide mutations were detected in the complete genomes of 15 HBoV1 strains, resulting in 11 amino acid mutations with 5 of them located in the active region of phospholipase A2 ( PLA2) .The 15 HBoV1 isolates along with 16 HBoV1 strains in GenBank were classified into three clusters as indicated by the phylogenetic analysis based on their complete coding sequences.All of the 15 strains were belonged to clusterⅠ, the representative strain of which was the Sweden prototype strain ST2.The phylogenetic trees constructed using genes encoding the capsid proteins VP1 and VP2 were highly similar to those based on the complete coding sequences.The estimated mean evolutionary rate of HBoV1 with regard to the complete coding sequence was 3.03×10-4(95%HPD, 2.14×10-4-3.92×10-4 ) substitu-tions per site per year.With regard to each gene, the NS1 gene was considered to the most conserved gene while the NP1 gene showed the highest substitution rate.The dN/dS ratios (ω) of the four genes were all less than 1, indicating that all of them were under negative selection.Moreover, the VP2 gene was under the strongest negative selection, while the NP1 gene was under the weakest negative selection.Conclusion All of the HBoV1 isolates circulating in Hangzhou province during 2009 to 2014 were belonged to ST2 genotype with a relatively high mutation in the area of PLA2.Despite the complete genome was conservative, its evo-lutionary rate was high.Among the four genes, the NP1 gene showed the highest substitution rate.All of the four genes were under negative selection, of which the VP2 gene was under the strongest negative selection.