人巨细胞病毒临床株UL133基因序列遗传变异分析

Genetic variation in the UL133 gene in clinical strains of the human cytomegalovirus

目的研究人巨细胞病毒（HCMV）临床感染株的UL133基因序列特征。方法采集HCMV-DNA阳性者的临床标本,PCR扩增UL133基因全序列,阳性扩增产物克隆到pEASYTM载体后进行序列测定,结合来自于NCBI数据库的15条序列进行UL133基因多态性分析。结果获得20例HCMV感染者的UL133全长序列。多态性分析显示HCMV临床株UL133基因核苷酸变异率为0～9.7%,氨基酸变异率为0～40.2%;不同感染者UL133序列5＇端的第32-45位发生了相对集中的非同义突变,其他部分序列较少出现氨基酸缺失及错义突变。其中1例临床感染者UL133序列在163-166位核苷酸出现移码突变。综合NCBI数据库的15株序列分析显示,UL133序列分为G1、G2、G3、G4、G5、G6等6个型,但未发现基因型与HCMV感染的临床表现具有显著关联性。编码蛋白翻译后修饰位点包括酪蛋白激酶磷酸化位点（CKP）,蛋白激酶C位点（PKC）以及NLS_BP核定位信号（NLS_BP）。与Toledo株相比,有1株发生移码突变,其他临床株UL133基因编码产物翻译后修饰位点相对保守。结论 HCMV UL133基因核苷酸序列及其编码的氨基酸序列高度保守,但仍具有一定的多态性。这种多态性与HCMV感染临床症状的关系尚待进一步研究。

Objective To examine the features of the sequence of the UL133 gene in clinical strains of the human cytomegalovirus（HCMV）. Methods A polymerase chain reaction（PCR）was used to amplify the complete sequence of the UL133 gene from clinical samples with HCMV and that sequence was then cloned into a pEASYTM vector for further sequencing.The features of the UL133 gene were further analyzed in conjunction with 15 sequences from the NCBI database. Results Twenty complete sequences of the UL133 gene were obtained from clinical samples with HCMV.Polymorphism analysis indicated that the UL133 gene had mutations in 0-9.7% of its nucleotides and mutations in 0-40.2% of its amino acids.Non-synonymous mutations were mainly located in the region from amino acids 32 to 45in the UL133 ORF.The other sequences had fewer missing amino acids or missense mutations.The UL133 gene in 1clinical isolate had a frame-shift mutation in the region from nt 163 to nt 166.Genotyping of the obtained sequences and the 15 sequences from the NCBI database indicated that UL133 sequences could be divided into 6groups（G1,G2,G3,G4,G5,and G6）.There was no significant relationship between the UL133 genotype and clinical outcome.Sites that were modified after translation of the encoded protein included the casein kinase II phosphorylation site（CKP）,the protein kinase C phosphorylation site（PKC）,and the bipartite nuclear localization signal profile（NLS_BP）.Compared to the Toledo reference strain,the posttranslational modification motifs of the putative UL133 protein were conserved in the obtained sequences except for 1sequence with a frame-shift mutation. Conclusion The nucleotide sequence and amino acid sequence of the ULl33 gene of HCMV were highly conserved and had certain polymorphisms.The relationship between pol-ymorphism of the UL133 gene of HCMV and clinical symptoms of HCMV infection need to be investigated further.