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Volume 23 Issue 1
February 2008

    Jing-jing WANG, Hai-lin ZHANG, Yan-chun CHE, Li-chun WANG, Shang-hui MA, Long-ding LIU, Yun LIAO and Qi-han LI. Isolation and Complete Genomic Sequence Analysis of a New Sindbis-like Virus[J]. Virologica Sinica, 2008, 23(1): 31-36. doi: 10.1007/s12250-008-2891-5
    Citation: Jing-jing WANG, Hai-lin ZHANG, Yan-chun CHE, Li-chun WANG, Shang-hui MA, Long-ding LIU, Yun LIAO, Qi-han LI. Isolation and Complete Genomic Sequence Analysis of a New Sindbis-like Virus .VIROLOGICA SINICA, 2008, 23(1) : 31-36.  http://dx.doi.org/10.1007/s12250-008-2891-5
    shu

    一个新的辛德毕斯样病毒的分离与全序列的分析

    cstr: 32224.14.s12250-008-2891-5
    • 1.

      ：中国医学科学院 中国协和医科大学 医学生物学研究所;云南昆明 2: 云南省地方病防治所;云南大理

    • 通讯作者： 李琦涵, qihanlister@gmail.com
    • 收稿日期： 2007-07-23
      录用日期： 2007-10-16
    • 辛德毕斯样病毒于1986年首次在我国分离，该基因序列全长11000多bp，编码3700多个氨基酸。其中包括了非结构区的5′端非转录区（5′－NTR）、非结构蛋白1、2、3、4区（nsP1、nsP2、nsP3、nsP4）、保守区（Conserved）、非保守区（Nonconserved）和结构区的核衣壳蛋白（Capsid）、结构1、2、3区（E1、E2、E3）、6K结构区（6K）、3′端非转录区（3′－NTR）我们从疑似脑炎发热病人的血液中分离病毒，经鉴定并传代后，取感染细胞的病毒上清，提取病毒RNA，用RT－PCR的方法，将其分为12个片断，再将扩增的片断连接到T载体上，通过正反向测序12个片断完成了Sindbis－IMB的全序列的测定：序列长全长11717，编码3773个氨基酸，在与其它辛德毕斯病毒比较后与同为云南分离得到的YN87448病毒株同源性最高变异率仅为1％，其次为SAAR86，变异率为1.2％。新分离到辛德毕斯样病毒的在非结构区上核酸序列变化较大，经比较发现在230、231、443、781、1582、1746位蛋白序列上出现了与其它序列完全不同的氨基酸，分别是K、E、N、R、H、L，且在nsp4末端较YN87448多出了3个氨基酸分别是谷氨酸、丝氨酸、丙氨酸(Q、S、A)。

    Isolation and Complete Genomic Sequence Analysis of a New Sindbis-like Virus

    • 1.

      Institute of Medical Biology, Chinese Academy of Medical Sciences and Peking Union Medical College, Kunming 650118, China

    • 2.

      Yunnan Institute for Endemic Diseases Control and Prevention, Dali 671000, China

    • Corresponding author: Qi-han LI, qihanlister@gmail.com
    • Received Date: 23 July 2007
      Accepted Date: 16 October 2007

      Fund Project: National Nature Science Founds 30670094National Nature Science Founds 30560142

    • The Sindbis-like virus was first discovered in China in 1986. Its complete genomic sequence consists of more than 11 000 bp encoding more than 3 700 amino acids. It contains a 5’ non-transcriptional region (5’-NTR) in a non-structural region, four non-structural proteins (nsP1, nsP2, nsP3, nsP4) regions, capsids in conserved and non-conserved regions and structural E1, E2, E3, 6K regions and a 3’ non-transcriptional region (3’-NTR). The Sindbis-IMB was isolated from the blood of a patient suspected to have encephalitis, and was followed by identification and passage. The virus RNA was extracted from virus supernatant in infected cells and the whole genome was divided into 12 fragments; RT-PCR was then performed to amplify the 12 fragments for complete sequencing. The results showed that the whole genomic sequence of Sindbis-IMB consists of 11 717 bp encoding 3 773 amino acids. Homology comparison with other Sindbis-like isolates demonstrated the highest similarity was the YN87448 with a variation of 1% strain isolated in Yunnan Province and the second highest to the SAAR86 strain with a variation of ~1.2%. The nucleotide sequence variations were present in non-structural regions, resulting in amino acids K, E, N, R, H, and L in protein sequences in positions 230, 231, 443,781, 1 582, and 1746 in the new isolation respectively. Furthermore, three additional amino acids--glutamic acid, serine and alanine--were noted in nsp4 terminus as compared to the YN87448 isolate.
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      1. Dennis A S, Nancy L D, Sheching L, et al. 1996. Complete nucleotide sequence and full-length cDNA clone of S. A. AR86, a South African Alphavirus Related to Sindbis. Virology, 222: 464-469.
          doi: 10.1006/viro.1996.0445

      2. Dubuisson J, Shlomo L, Nicolas Rug, et al. 1997. Genetic determinants of Sindbis virus neuroinvasiveness. J Virol, 71: 2636-3646.

      3. Laine M, Luukkainen R, Toivanen A. 2004. Sindbis virus and other alphavirus as cause of human arthritic disease. J Internal Med, 256: 457-471.
          doi: 10.1111/jim.2004.256.issue-6

      4. Malherbe H S, Cholmley M, Jackson A.L. 1963. Sindbis virus infection in man. South Afr Med J, 37: 547-552.

      5. Martin P, Richard M K, Oskar R K. 1998. The alphavirus 3'-nontranslated region:size heterogeneity and ar2rangement of repeated sequence elements. Virology, 240 (1): 100-110.
          doi: 10.1006/viro.1997.8907

      6. Shailly T, Richard W, Janet L S, et al. 2006. Catalytic Core of Alphavirus Nonstructural Protein Nsp4 Possesses Terminal Adenylyltransferase Activity. J Virol, 10: 9962-9969.

      7. Zhou G L, Lian G D, Li L, et al. 1999. Complete nucleotide sequence of the structural region gene of Alphavirus YN87448 strain isolated in China and its relationship to other Sindbis virus. Virologica Sinica, 15: 205-210. (in Chinese).

      8. Zhou G L, Lian G D, Li L, et al. 1999. Complete nucleotide sequence of the nonstructural gene of Alphavirus YN87448 strain isolated in China and its relationship to other Sindbis virus. Chin J Exp Clin Virol, 13: 314-320. (in Chinese).

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      Isolation and Complete Genomic Sequence Analysis of a New Sindbis-like Virus

        Corresponding author: Qi-han LI, qihanlister@gmail.com
      • 1. Institute of Medical Biology, Chinese Academy of Medical Sciences and Peking Union Medical College, Kunming 650118, China
      • 2. Yunnan Institute for Endemic Diseases Control and Prevention, Dali 671000, China
      • Received Date: 23 July 2007
      • Accepted Date: 16 October 2007
      Fund Project:  National Nature Science Founds 30670094National Nature Science Founds 30560142

      Abstract: The Sindbis-like virus was first discovered in China in 1986. Its complete genomic sequence consists of more than 11 000 bp encoding more than 3 700 amino acids. It contains a 5’ non-transcriptional region (5’-NTR) in a non-structural region, four non-structural proteins (nsP1, nsP2, nsP3, nsP4) regions, capsids in conserved and non-conserved regions and structural E1, E2, E3, 6K regions and a 3’ non-transcriptional region (3’-NTR). The Sindbis-IMB was isolated from the blood of a patient suspected to have encephalitis, and was followed by identification and passage. The virus RNA was extracted from virus supernatant in infected cells and the whole genome was divided into 12 fragments; RT-PCR was then performed to amplify the 12 fragments for complete sequencing. The results showed that the whole genomic sequence of Sindbis-IMB consists of 11 717 bp encoding 3 773 amino acids. Homology comparison with other Sindbis-like isolates demonstrated the highest similarity was the YN87448 with a variation of 1% strain isolated in Yunnan Province and the second highest to the SAAR86 strain with a variation of ~1.2%. The nucleotide sequence variations were present in non-structural regions, resulting in amino acids K, E, N, R, H, and L in protein sequences in positions 230, 231, 443,781, 1 582, and 1746 in the new isolation respectively. Furthermore, three additional amino acids--glutamic acid, serine and alanine--were noted in nsp4 terminus as compared to the YN87448 isolate.

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