广州地区婴幼儿感染Noro病毒基因型的初步研究

目的　了解广州市儿童腹泻中诺瓦克病毒Norovirus 感染的流行病学特点及病毒的基因型。方法 用ClustalW比对GII组诺瓦克病毒后，设计了处于ORF1与ORF2连接点两旁两对简并引物，进行巢式PCR扩增出目标片段，克隆于T载体上，测定序列，对ORF1与ORF2连接点两旁序列和衣壳蛋白N/S区用ClustalW进行同源性分析，用phylip3.65软件、NJ方法构建进化树。结果　两份标本NVgz100、NVgz10成功扩增出ORF1与ORF2连接点两旁1208bp片段（相对于Hawaii virus，U07611位置为4476-5683），包含ORF1的3′端RdRp基因的大部分基因和ORF2基因的5′端的S区。对标本NVgz100、NVgz10与另一实验NVgz01（DQ369797）的1208bp进行同源性分析发现，三个标本与GI组同源性为58-61%，与GII组同源性为74-92%；将这三个标本与GI、GII组构建进化树，可发现NVgz100、NVgz10、NVgz01与GII组Bristol等病毒密切相关，将NVgz100、NVgz10、NVgz01的衣壳蛋白N/S区与GII组各基因型进行同源性比较，发现三个标本与GII-4基因型Camberwell、Bristol、Grimsby同源性较高（92-96%），与其它基因型同源性为70-73%，以N/S区构建进化树可发现三个标本与Camberwell、Bristol、Grimsby、Lordsdale处于同一簇中。结论　本实验病毒株为诺瓦克病毒GII组，初步认为广州地区诺瓦克病毒感染以GII组病毒株为主，病毒流行的基因型为与Camberwell、Bristol、Grimsby、Lordsdale等密切相关的GII-4基因型。

The genotype of Noro virus infection in infants in Guangzhou

Objective 　To explore the epidemiological characteristics of Norovirus infection in children with diarrhea and to study the Norovirus genotype in Guangzhou. Methods Primers were designed on the basis of the sequence of Hawaii virus after ClustalW alignment of six human norovirus genomic sequences, and amplify the fragment of ORF1-ORF2 junction by nest PCR. The PCR-products were cloned to T vector and sequenced, phylogenetic tree was constructed by the neighbor joining method in Phylip3.65. Results　The amplified sequence of orf1/orf2 junction is 1208bp in length, correspond to the regions located between nucleotides 4476 and 5683 in the Hawaii virus genome(GenBank accession number U07611).The sequence contains 3′-RNA polymerase sequence and N-terminus/shell domain in ORF2 gene. The three ORF1-ORF2 fragments are compared with those of Norovirus in GenBank, revealed that the homology with genogroupII ranges between 74%-92%, and genogroupI between 58%-61%.The N/S domains of three samples were analyzed by comparing the sequences of strains representing each genotype of GII, the result revesls that the homology with GII-4 ranges between 92%-96%.phylogenetic trees based on capsid N/S domain and ORF1-ORF2 junction sequence were constructed by the neighbor-joining method with Manchester strain as an outgroup, and find that the three samples are closer to GII-4 stains than to other reference strains. Conclusions Norovirus strains infection in Guangzhou belongs to genogroupII.