丁型肝炎病毒中国湖南株高变区部分序列分析

SEQUENCING PART OF HYPERVARIABLE REGION OF HEPATITIS D VIRUS GENOME DERIVED FROM HUNAN CHINA

用耐高温逆转录以及套式聚合酶链反应方法,对HDV中国湖南株基因组高变区1642～417片段(D1),以及保守区681～895片段(D3)进行扩增,用荧光法直接测序,对其进行序列分析。D1片段除去引物外可判读222nt(1662～197),D3片段除去引物还有176nt(701～875)。两段序列分别与美国株、意大利株、日本株以及中国河南株进行了比较。结果显示,中国湖南株高变区1662～197片段(D1)与日本株的同源性仅为44.06％,与意大利株的同源性相对较高,但也仅为72.57％。日本株在该片段的同源性特别低,而中国河南株在该片段的同源性并不特别低。各株之间在该片段的同源性仅为44.06％～72.56％,说明HDV基因组1662～197片段是高变区。中国湖南株D1片段与其它各株的同源性为44.06％～72.57％,且与中国河南株同源性也仅为62.61％,提示中国湖南株可能系一新的亚型。各株之间在701～875片段(D3)的同源性较高,中国湖南株与意大利株更为接近(93.03％),而与中国河南株(91.82％)、美国株(89.54％)和日本株(89.54％)的同源性相对较低。各株之间的同源性比较接近,提示HDV基因组701～875片段是相对保守区。同源性的高低与地理位置分布不一致,其原因有待进一步研究。

With themethod of thermal resisting reverse transcription and nest polymerase chain reaction (PCR).we amplified the Dl segment (1642-417) in hypervariable region and D3 segment (681~895) in conservative region of HDV genome Hunan isolate. The PCR products were directly sequenced using fluorescencemethod, and the sequences were analyzed. Except primers,222nt in segment Dl(1662~197) and 176nt in segment D3 (701~875) were analyzed. The two segments were compared with the isolates from USA, Italy ,Japan and Henan China, respectively. Results showed that,the homogeneity of segment 1662-197 (Dl) in hypervariable region of HDV genome Hunan isolate was as low as 44. 06% compared with that of Japan,and was relatively high with that of Italy(72. 57%). The homogeneity of Japan isolate in this segment was significantly low among the 5 isolates,and that of Hunan isolate was not. The fact that the homogeneity in this segment was low (44. 06%-74. 56%) among the 5 isolates reveals that segment 1662-197 of HDV genome is a hypervariable region. And that the homogeneity of segment Dl of Hunan isolate was 44. 06%~72. 57% compared with that of the other 4 isolated and was only 62. 61% with that of Henan China,reveals that the isolate from Hunan China is probably a new subtype. The homogeneity was high in segment 701-875 (D3) among the 5 isolates. The Hunan isolate wasmore close to Italy (93.03%) than Henan (91. 82%),USA(89. 54%) and Japan (89. 54%). That the homogeneity was close in. different isolates reveals that segment 701-875 of HDV genome is a relatively conservative region. The difference between the homogeneity and geographic distribution needs to be further studied.