人巨细胞病毒UL136基因在临床低传代分离株中多态性分析

目的 研究人巨细胞病毒(human cytomegalovirus，HCMV)UL136基因在临床低传代分离株中的多态性，探讨其多态性与HCMV先天性感染不同致病性之间的关系。方法 对48株经荧光定量PCR方法检测HCMV DNA为阳性的临床低传代分离株进行HCMV ULl36全序列PCR扩增，对于扩增阳性的12株PCR产物进行ULl36基因全序列测定及结果分析。结果 48株临床低传代分离株ULl36 PCR扩增，12株阳性，阳性率25％，以HCMV Toledo株作为参考株，进行序列比较分析表明，12株临床分离株ULl36开放阅读框架(open reading frame，ORF)长度均与Toledo株相同，为723bp，编码241个氨基酸的蛋白。DNA序列变异均为碱基替换，不同临床分离株ULl36基因与Toledo株进行同源性比较，结果在核苷酸水平为97．7％～99．3％，氨基酸水平为96．6％～99．1％。ULl36编码蛋白的氨基酸变异率为0．83％～3．3％。二级结构预测分为两种构象。大多数HCMV ULl36蛋白翻译后修饰位点在所有分离株中均高度保守，仅几个位点在一些分离株中存在缺失或新增。Toledo株及12株临床分离株核苷酸及氨基酸序列系统进化树分析表明：45J最接近Toledo株。结论 12株临床低传代分离株HCMV ULl36基因DNA及其编码产物的氨基酸序列比较保守，但仍存在一定多态性。未发现不同临床分离株ULl36基因多态性与HCMV先天性感染的表现关系。

Polymorphism of human cytomegalovirus UL136 gene in low passage clinical isolates

Objective To investigate the polymorphism of human cytomegalovirus UL136 gene in low passage clinical isolates and to find the relationship between the polymorphism and different pathogenesis of congenital HCMV infection. Methods PCR amplification products of 12 isolates were directly sequenced and analyzed. Results 12 of 48 isolates were successfully amplified with positive rate of 25%. By comparison with Toledo sequence, the length of UL136 ORF in all 12 clinical isolates was similar to that of Toledo, 723 bp in size. They had the potential to encode 241 amino acid protein. DNA sequence variations were nucleotide substitutions, neither insertions nor deletions were found. Alignment comparison of clinical isolates UL136 sequences with that of Toledo revealed nt and aa sequence homologies of 97.7%-99.3% and 96.6%-99.1%, respectively. Amino acid variability rate of UL136 protein was 0.83%-3.3%. Two types of secondary structure have been found. Most posttranslational modification sites of UL136 protein were highly conserved although several strains had deleted or additional sites. The phylogenetic trees based on UL136 genes of Toledo strain and 12 clinical isolates demonstrated that 45J was the most closely strain related to Toledo. Conclusion All DNA and deduced amino acid sequences of UL136 gene shared similarity among HCMV clinical strains regardless of their polymorphism. No linkage was found between diversities of UL136 gene and the outcomes of congenital HCMV infection.