조류인플루엔자 H5N1 바이러스 유전자의 신속 검출을 위한 초고속 다중 실시간 PCR법의 개발

Development of Ultra-rapid Multiplex Real-time PCR for the Detection of Genes from Avian Influenza Virus subtype H5N1

  • 심사 : 2007.11.27
  • 발행 : 2007.12.30

초록

Cause of high lethality and dissemination to human being, new development of rapid method for the detection of highly pathogenic Avian Influenza Virus (AIV) is still necessary. For the detection of AIV subtype H5N1, typical pathogenic AIV, new method to confirm sub-typing of this virus is also needed. For the purpose of ultra-rapid detection and sub-typing of hemagglutinin and neuraminidase of AIV, this study was planned. As the results we could demonstrate an ultra-rapid multiplex real-time PCR (URMRT PCR) for the detection of AIV In this study, the URMRT PCR were optimized with synthesized AIV H5- and AIV Nl-specific DNA templates and GenSpector TMC, which is a semiconductor process technology based real-time PCR system with high frequencies of temperature monitoring. Under eight minutes, the amplifications of two AIV subtype-specific PCR products were successfully and independently detected by 30 cycled ultra-rapid PCR, including melting point analysis, from $1{\times}10^3$ copies of mixed template DNA. The URMRT PCR for the detection of AIV H5N 1 developed in this study could be expected to apply not only detections of different AIVs, but also various pathogens. It was also discussed that this kind of the fastest PCR based detection method could be improved by advance of related technology in near future.

키워드

참고문헌

  1. Bustin SA. Absolute quantification of mRNA using real-time reverse transcription polymerase chain reaction assays. J Mol Endocrinol 2000, 25, 169-193 https://doi.org/10.1677/jme.0.0250169
  2. Bustin SA, Mueller R. Real-time reverse transcription PCR (qRT-PCR) and its potential use in clinical diagnosis. Clin sci 2005, 109, 365-379 https://doi.org/10.1042/CS20050086
  3. Cho YK, Kim J, Lee Y, Kim YA, Namkoong K, Lim H, Oh KW, Kim S, Han J, Park C, Pak YE, Ki CS, Choi JR, Myeong HK, Ko C. Clinical evaluation of micro-scale chip-based PCR system for rapid detection of hepatitis B virus. Biosens Bioelectron 2006, 21, 2161-2169 https://doi.org/10.1016/j.bios.2005.10.005
  4. Ellis JS, Zambon MC. Molecular diagnosis of influenza. Rev Med Virol 2002, 12, 375-389 https://doi.org/10.1002/rmv.370
  5. Fouchier RA, Munster V, Wallensten A, Bestebroer TM, Herfst S, Smith D, Rimmelzwaan GF, Olsen B, Osterhaus AD. Characterization of a Novel Influenza A Virus Hemagglutinin Subtype (H16) Obtained from Black-Headed GulIs. J Virol 2005, 79, 2814-2822 https://doi.org/10.1128/JVI.79.5.2814-2822.2005
  6. Gibson UE, Heid CA, Williams PM. A novel method for real time quantitative RT-PCR. Genome Res 1996, 6, 995-1001 https://doi.org/10.1101/gr.6.10.995
  7. Han SH, Lim YK, Yoon BS. Manual of Method for Molecular Biology III. 3rd ed. pp. 82-84, Kyonggi university Press, Suwon, 2006
  8. Hoffmann B, Harder T, Stariek E, Depner K, Werner O, Beer M. Rapid and highly sensitive pathotyping of avian influenza A H5NI virus by using real-time reverse transcription-PCR. J Clin Microbiol 2007, 45, 600-603 https://doi.org/10.1128/JCM.01681-06
  9. Horimoto T, Kawaoka Y. Pandemic threat posed by avian influenza a viruses. Clin Microbiol Rev 2001, 14, 129-149 https://doi.org/10.1128/CMR.14.1.129-149.2001
  10. Kim EH, Lee DW, Han SH, Kwon SH, Yoon BS. Rapid detection of avian influenza subtype H5N1 using quick real-time PCR. Kor J Microbiol 2007, 43, 23-30
  11. Lau LT, Banks J, Aherne R, Brown IH, Dillon N, Collins RA, Chan KY, Fung YW, Xing J, Yu AC. Nucleic acid sequence-based amplification methods to detect avian influenza virus. Biochem Biophys Res Commun 2004, 313, 336-342 https://doi.org/10.1016/j.bbrc.2003.11.131
  12. Lee DW, Kim EH, Yoo MS, Han SH, Yoon HS. Ultra-rapid real-time PCR for the detection of human immunodeficiency virus (HIV). Kor J Microbiol 2007, 43, 91-99
  13. Munch M, Nielsen LP, Handberg KJ, Jorgensen PH. Detection and sub typing (H5 and B7) of avian type A influenza virus by reverse transcription-PCR and PCR-ELISA. Arch Virol 2001, 146, 87-97 https://doi.org/10.1007/s007050170193
  14. Murphy BR, Webster RG. Orythomyxoviruses. In: Fields BN, Knipe DM, Howley PM (eds.). Fields Virology, 3rd ed. vol. 1. pp. 1397-1445, LippincottRaven Publishers, Philadelphia, 1996
  15. Nicholson KG, Wood JM, Zambon M. Influenza. Lancet 2003, 362, 1733-1745 https://doi.org/10.1016/S0140-6736(03)14854-4
  16. Payungporn S, Chutinimitkul S, Chaisingh A, Damrongwantanapokin S, Buranathai C, Amonsin A, Theamboonlers A, Poovorawan Y. Single step multiplex real-time RT-PCR for H5N1 influenza A virus detection. J Virol Methods 2006, 131, 143-147 https://doi.org/10.1016/j.jviromet.2005.08.004
  17. Poddar SK. Influenza virus types and subtypes detection by single step single tube multiplex reverse transcription-polymerase chain reaction (RT-PCR) and agarose gel electrophoresis. J Virol Metheds 2002, 99, 63-70 https://doi.org/10.1016/S0166-0934(01)00380-9
  18. Reina J, Munar M, Blanco I. Evaluation of a direct immunofluorescence assay, dot-blot enzyme immunoassay, and shell vial culture in the diagnosis of lower respiratory tract infections caused by influenza A virus. Diagn Microbiol Infect Dis 1996, 25, 143-145 https://doi.org/10.1016/S0732-8893(96)00131-9
  19. Spackman E, Senne DA, Myers TJ, Bulaga LL, Garber LP, Perdue ML, Lohman K, Daum LT, Suarez DL. Development of a real-time reverse transcriptase PCR assay for type a influenza virus and the avian H5 and H7 hemagglutinin subtypes. J Clin Microbiol 2002, 40, 3256-3260 https://doi.org/10.1128/JCM.40.9.3256-3260.2002
  20. Starick E, Rmer-Oberdrfer A, Werner O. Type- and subtype-specific RT-PCR assays for avian influenza A viruses (AIV). J Vet Med B Infect Dis Vet Public Health 2000, 47, 295-301 https://doi.org/10.1046/j.1439-0450.2000.00386.x
  21. Steininger C, Kundi M, Aberle SW, Aberle JR, Popow-Kraupp T. Effectiveness of Reverse Transcription-PCR, Virus Isolation, and EnzymeLinked Immunosorbent Assay for Diagnosis of Influenza A Virus Infection in Different Age Groups. J Clin Microbiol 2002, 40, 2051-2056 https://doi.org/10.1128/JCM.40.6.2051-2056.2002
  22. van Elden LJR, Nijhuis M, Schipper P, Schuurman R, van Loon AM. Simultaneous detection of influenza viruses A and B using real-time quantitative PCR. J Clin Microbiol 2001, 39, 196-200 https://doi.org/10.1128/JCM.39.1.196-200.2001
  23. Wang S, Levin RE. Thermal factors influencing detection of Vibrio vulnificus using real-time PCR. J Microbiol Methods 2007, 69, 358-363 https://doi.org/10.1016/j.mimet.2007.02.003
  24. Webster RG, Kawaoka Y. Avian influenza. Rev Poultry Biol 1987, 1, 212-246
  25. Wei HL, Bai GR, Mweene AS, Zhou YC, Cong YL, Pu J, Wang S, Kida H, Liu JR. Rapid detection of avian influenza virus A and subtype H5N1 by single step multiplex reverse transcription-polymerase chain reaction. Virus Genes 2006, 32, 261-267 https://doi.org/10.1007/s11262-005-6910-4
  26. Xie Z, Pang YS, Liu J, Deng X, Tang X, Sun J, Khan MI. A multiplex RT-PCR for detection of type A influenza virus and differentiation of avian H5, H7, and H9 hemagglutinin subtypes. Mol Cell Probes 2006, 20, 245-249 https://doi.org/10.1016/j.mcp.2006.01.003
  27. Yuen KY, Chan PKS, Peiris M, Tsang DN, Que TL, Shortridge KF, Cheung PT, To WK, Ho ET, Sung R, Cheng AF. Clinical features and rapid viral diagnosis of human disease associated with avian influenza A H5N1 virus. Lancet 1998, 351, 467-471 https://doi.org/10.1016/S0140-6736(98)01182-9