[Objective] Newcastle disease virus (NDV) is one of the most serious infection problems in China in the last two decades,resulting in huge economic losses for the poultry industry.The genetic evolution of NDV strain is a very important topic for controlling ND.In this study,the evolution of NDV under the immune pressure of vaccine was further assessed by comparing the molecular characteristics and genetic variation frequency of F and HN genes of different genotypes of NDV.[Methods] NDV sequences of F gene and HN gene including 89 NDV isolates in our laboratory,364 NDV pandemic strains worldwide and 15 classical strains,were collected and downloaded from the GenBank,and further genetically analyzed to determine their evolution,molecular characteristic and substitution rate by using Lasergene 7.1 and MEGA 5.1 software.[Results] Phylogenetic tree analysis suggested that NDV evolved into 15 distinct genotypes from genotype I to XV.Most NDV pandemic isolates during 1997-2015 belonged to sub-genotype Ⅶ d,suggesting that this genotype was the dominate isolate in China in the last two decades.Furthermore,comparison based on homologies of the nucleotide and amino acid sequence of F and HN gene of NDV showed that different genotypes had their own distinct characteristics,and significant accumulation of amino acid variation was also found.In addition,in comparing F and HN gene with reference Go/GD/QY/1997,which was the first genotype Ⅶ in China,average annual substitution frequencies of NDV pandemic strain nucleotide (amino acid) were 2.31 × 10-3 (2.26 × 10 3) and 3.37 × 10-3 (2.35×10-3),respectively.Substitution rates ofF and HN during 1997-2001 were 4.72× 10-3 and 8.28× 10-3,higher than that during 2002-2015 (1.6× 10-3 and 1.84× 10 3),when the genotype Ⅶ vaccine was initially applied in the field.[Conclusion] Bioinformatics analysis proved that matching NDV vaccine strain with epidemic strains in genotype is an important factor in mitigating NDV variability.%[目的]新城疫(ND)是中国流行最严重的疫病之一,对家禽业可造成巨大的经济损失,疫苗防控是控制ND的重要措施.新城疫病毒(NDV)流行株的遗传演化一直是研究NDV的焦点.本文利用分子信息学手段,通过比较近20年间NDV流行株不同基因型F和HN基因的分子特征和遗传变异频率,解析免疫压力下NDV的演化规律.[方法]利用Lasergene 7.1和MEGA5.1软件,选取本实验室89株NDV分离株,结合从GenBank下载的364株NDV流行株以及15株NDV经典毒株的基因序列,对其进行系统发育、分子特征和替代频率分析.[结果]系统发育表明,NDV已经演化为15个基因型.一致性比较显示,NDV流行株相同基因型之间核苷酸(氨基酸)高度同源,而不同基因型之间差异较大且存在明显的氨基酸变异积累.NDV基因型的分布与时间、地域密切相关,Ⅶd亚型为中国NDV优势流行株.为评估NDV变异的频率,以Go/GD/QY/1997株(中国较早发生的基因Ⅶ亚型)为参照,1997-2015年间NDV的F/HN基因的年平均核苷酸(氨基酸)替代率为2.31×10 3 (2.26×10 3)/3.37×10 3 (2.35×10-3).其中,1997-2001年(未使用基因Ⅶ型疫苗)F/HN基因核苷酸年平均替代率为4.72× 10-3/8.28× 10-3;2002-2015年(疫苗使用后)为1.6×10-3/1.84×10-3,显示出基因Ⅶ型疫苗在控制NDV变异速度方面具有明显的效果.[结论]生物信息学分析证实:研制出与NDV流行毒株相匹配的新型疫苗是控制当前NDV变异的关键.
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