鸡传染性支气管炎病毒中国分离株LH2/01/10纤突蛋白基因的遗传变异

Genetic Variation of Spike Gene of Infectious Bronchitis Virus LH2/01/10 Isolated in China

对传染性支气管炎病毒LH2/01/10的S1和S2基因分别进行RT-PCR扩增、克隆和序列测定。结果表明,其S基因由3 495个核苷酸组成,编码的多肽由1 164个氨基酸残基组成。S蛋白的切割识别位点氨基酸组成为组氨酸-精氨酸-精氨酸-精氨酸-精氨酸(HRRRR)。推测其裂解后形成的S1和S2亚单位中,S1由539个氨基酸残基组成,S2由625个氨基酸残基组成。序列分析发现:LH2/01/10与Beau、M41、CU-T2、D1466、DE072、Holte、Gray、Ark99、Conn、H52、KB8523、SD/97/02、ZJ971和LX4毒株S基因推导的氨基酸同源性在63%～96%之间。其中与国内分离毒株SD/97/02和LX4氨基酸的同源性均在95%以上,三者切割识别位点也相同。但与另一国内分离株ZJ971氨基酸的同源性仅为75%,而与国外毒株之间在83%以下。切割识别位点也与ZJ971和所选的国外参考毒株代表性的RRF/SRR位点不同。与这些参考毒株相比,LH2/01/10 S基因在76～78位缺失TCT碱基,在67～69位插入TCT碱基,105位和549位氨的基酸发生点突变。总体来看, SD/97/02的S1基因推导的氨基酸序列与国内外已报道的42株参考毒株同源性较低,在52%～96%之间。与国内分离毒株亲缘关系相对较近,其中与国内近年来不同地区分离的A2、LX4、SD/97/02、TJ/96/02、JX/99/01。

According to the published S gene sequence of IBV, two pairs of specific primers were designed and synthesized.Two fragments of S1 and S2 of LH2/01/10 strain were amplified by reverse transcription-polymerase chain reaction (RT-PCR)method, then the two fragments were cloned and sequenced respectively. The results showed that the S gene of LH2/01/10 wascomposed of 3 495 bp encoding a polypeptide of 1 164 amino acids. The cleavage site sequence of S protein of LH2/01/10contains five consecutive basic amino acids, namely, His-Arg-Arg-Arg-Arg (HRRRR). The deduced sequence of S1 and S2protein was composed of 539 and 625 amino acids, respectively. Sequence data demonstrated that the complete S amino acidsequences of LH2/01/10 strain shared 63%-96% similarity with the published S sequences of Beau, M41, CU-T2, D1466,DE072, Holte, Gray, Ark99, Conn, H52, KB8523, SD/97/02, ZJ971 and LX4 strains. The results indicated that LH2/01/10 hascloser relationship with SD/97/02 and LX4 strains which were isolated in China. Moreover, sequence analysis showed thatLH2/01/10 had closer relationship with the IBV strains in China isolated in recent years according to the complete S1 aminoacid sequence, hypervariable region (HVR) and six antigenic epitopes of S1 protein. In addition, there were deletion, insertionand point mutation in the S gene of LH2/01/10 compared with the other virus strains. This study suggested that the lowsequence identity between S1 genes of LH2/01/10 and H120 and the changes of pathogenicity resulted by the deletion, insertionand point mutation in the S gene of LH2/01/10 strain would be the main reasons that caused the infection by IBV-LH2/01/10strain in H120 immunized chickens.