Skip to main content

Advertisement

Log in

Novel reverse transcription loop-mediated isothermal amplification for rapid detection of foot-and-mouth disease virus

  • Published:
Archives of Virology Aims and scope Submit manuscript

Summary.

Speed is paramount in the diagnosis of foot-and-mouth disease (FMD) and simplicity is required if a test is to be deployed in the field. The development of a one-step, reverse transcription loop-mediated amplification (RT-LAMP) assay enables FMD virus (FMDV) to be detected in under an hour in a single tube without thermal cycling. A fragment of the 3D RNA polymerase gene of the virus is amplified at 65 °C in the presence of a primer mixture and both reverse transcriptase and Bst DNA polymerase. Compared with real-time RT-PCR, RT-LAMP was consistently faster, and ten copies of FMDV transcript were detected in twenty-two minutes. Amplification products were detected by visual inspection, agarose gel electrophoresis, or in real-time by the addition of a fluorescent dye. The specificity of the reaction was demonstrated by the absence of amplification of RNA from other viruses that cause vesicular diseases and from that of genetically related picornaviruses. Diagnostic sensitivity was validated by the amplification of reference FMDV strains and archival material from field cases of FMD. In comparison with the performance of the established diagnostic TaqMan® assay, RT-LAMP appears to be sensitive, rapid, specific, and cost-effective, with the potential for field deployment and use by developing countries for FMDV surveillance.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • S Alexandersen Z Zhang AI Donaldson AJ Garland (2003) ArticleTitleThe pathogenesis and diagnosis of foot-and-mouth disease Comp Pathol 129 1–36 Occurrence Handle1:CAS:528:DC%2BD3sXlt1aqtrY%3D Occurrence Handle10.1016/S0021-9975(03)00041-0

    Article  CAS  Google Scholar 

  • Anonymous (2002) The 2001 outbreak of foot and mouth disease. Report by the Controller and Auditor General. National Audit Office HC 939 Session 2001–2002: 21 June 2002

  • JD Callahan F Brown FA Osorio JH Sur E Kramer GW Long J Lubroth SJ Ellis KS Shoulars KL Gaffney DL Rock WM Nelson (2002) ArticleTitleUse of a portable real-time reverse transcriptase-polymerase chain reaction assay for rapid detection of foot-and-mouth disease virus Am Vet Med Assoc 220 1636–1642 Occurrence Handle1:CAS:528:DC%2BD38Xks1OgtLw%3D Occurrence Handle10.2460/javma.2002.220.1636

    Article  CAS  Google Scholar 

  • JAW Coetzer GR Thompson RC Tustin NPJ Kriek (1994) Foot-and-mouth disease JAW Coetzer GR Thomson RC Tustin NPJ Kriek (Eds) Infectious diseases of livestock with special reference to Southern Africa Oxford University Press Cape Town 825–852

    Google Scholar 

  • E Domingo MG Mateu C Escarmis E Martinez-Salas D Andreu E Giralt N Verdaguer I Fita (1995) ArticleTitleMolecular evolution of aphthoviruses Virus Genes 11 197–207 Occurrence Handle8828146 Occurrence Handle1:CAS:528:DyaK28Xkt1Wnsb4%3D Occurrence Handle10.1007/BF01728659

    Article  PubMed  CAS  Google Scholar 

  • AI Donaldson A Hearps S Alexandersen (2001) ArticleTitleEvaluation of a portable, ‘real-time’ PCR machine for FMD Vet Rec 149 430 Occurrence Handle11678219 Occurrence Handle1:STN:280:DC%2BD3MrntVymuw%3D%3D

    PubMed  CAS  Google Scholar 

  • Ferris NP, King DP, Reid SM, Shaw AE, Hutchings GH (2006) Analysis of original laboratory results and retrospective analysis by real-time RT-PCR of virological samples collected from confirmed cases suggest over-reporting of foot-and-mouth disease in the UK in 2001. Vet Rec (in press)

  • S Fukuta T Iida Y Mizukami A Ishida J Ueda M Kanbe Y Ishimoto (2003) ArticleTitleDetection of Japanese yam mosaic virus by RT-LAMP Arch Virol 148 1713–1720 Occurrence Handle14505084 Occurrence Handle1:CAS:528:DC%2BD3sXptlCqu74%3D Occurrence Handle10.1007/s00705-003-0134-5

    Article  PubMed  CAS  Google Scholar 

  • A Hearps Z Zhang S Alexandersen (2002) ArticleTitleEvaluation of the portable Cepheid SmartCycler real-time PCR machine for the rapid diagnosis of foot-and-mouth disease Vet Rec 150 625–628 Occurrence Handle12046786 Occurrence Handle1:STN:280:DC%2BD38zgsFOksA%3D%3D

    PubMed  CAS  Google Scholar 

  • T Iwamoto T Sonobe K Hayashi (2003) ArticleTitleLoop-mediated isothermal amplification for direct detection of Mycobacterium tuberculosis complex, M. avium, and M. intracellulare in sputum samples J Clin Microbiol 41 2616–2622 Occurrence Handle12791888 Occurrence Handle1:CAS:528:DC%2BD3sXlsValtLw%3D Occurrence Handle10.1128/JCM.41.6.2616-2622.2003

    Article  PubMed  CAS  Google Scholar 

  • MY Iwasaki T Yonekawa K Otsuka W Suzuki K Nagamine T Hase K-I Tatsumi T Horigome T Notomi H Kanda (2003) ArticleTitleValidation of the loop-mediated isothermal amplification method for single nucleotide polymorphism genotyping with whole blood Genome Lett 2 119–126 Occurrence Handle1:CAS:528:DC%2BD3sXps1ent7o%3D

    CAS  Google Scholar 

  • DP King NP Ferris AE Shaw SM Reid GH Hutchings AC Giuffre JM Robida JD Callahan WM Nelson TR Beckham (2005) ArticleTitleDetection of foot-and-mouth disease virus: comparative diagnostic sensitivity of two independent real-time RT-PCR assays J Vet Diagn Invest 18 92–96

    Google Scholar 

  • NJ Knowles AR Samuel (2003) ArticleTitleMolecular epidemiology of foot-and-mouth disease virus Virus Res 91 65–80 Occurrence Handle12527438 Occurrence Handle1:CAS:528:DC%2BD3sXjtF2msg%3D%3D Occurrence Handle10.1016/S0168-1702(02)00260-5

    Article  PubMed  CAS  Google Scholar 

  • Y Mori K Nagamine N Tomita T Notomi (2001) ArticleTitleDetection of loop-mediated isothermal amplification reaction by turbidity derived from magnesium pyrophosphate formation Biochem Biophys Res Commun 289 150–154 Occurrence Handle11708792 Occurrence Handle1:CAS:528:DC%2BD3MXot12ht7Y%3D Occurrence Handle10.1006/bbrc.2001.5921

    Article  PubMed  CAS  Google Scholar 

  • K Nagamine K Watanabe K Ohtsuka T Hase T Notomi (2001) ArticleTitleLoop-mediated isothermal amplification reaction using a nondenatured template Clin Chem 47 1742–1743 Occurrence Handle11514425 Occurrence Handle1:CAS:528:DC%2BD3MXmtlShtr4%3D

    PubMed  CAS  Google Scholar 

  • K Nagamine T Hase T Notomi (2002) ArticleTitleAccelerated reaction by loop-mediated isothermal amplification using loop primers Mol Cell Probes 16 223–229 Occurrence Handle12144774 Occurrence Handle1:CAS:528:DC%2BD38XlsFWqsr4%3D Occurrence Handle10.1006/mcpr.2002.0415

    Article  PubMed  CAS  Google Scholar 

  • T Notomi H Okayama H Masubuchi T Yonekawa K Watanabe N Amino T Hase (2000) ArticleTitleLoop-mediated isothermal amplification of DNA Nucleic Acids Res 28 E63 Occurrence Handle10871386 Occurrence Handle1:CAS:528:DC%2BD3cXmtlKqsr8%3D Occurrence Handle10.1093/nar/28.12.e63

    Article  PubMed  CAS  Google Scholar 

  • M Parida G Posadas S Inoue F Hasebe K Morita (2004) ArticleTitleReal-time reverse transcription loop-mediated isothermal amplification for rapid detection of West Nile virus J Clin Microbiol 42 257–263 Occurrence Handle14715762 Occurrence Handle1:CAS:528:DC%2BD2cXht1Wnt7w%3D Occurrence Handle10.1128/JCM.42.1.257-263.2004

    Article  PubMed  CAS  Google Scholar 

  • S Reid N Ferris G Hutchings Z Zhang G Belsham S Alexandersen (2002) ArticleTitleDetection of all seven serotypes of foot-and-mouth disease virus by real-time, fluorogenic reverse transcription polymerase chain reaction assay J Virol Methods 105 67–80 Occurrence Handle12176143 Occurrence Handle1:CAS:528:DC%2BD38XmtlGrt70%3D Occurrence Handle10.1016/S0166-0934(02)00081-2

    Article  PubMed  CAS  Google Scholar 

  • SM Reid MA Forsyth GH Hutchings NP Ferris (1998) ArticleTitleComparison of reverse transcription polymerase chain reaction, enzyme linked immunosorbent assay and virus isolation for the routine diagnosis of foot-and-mouth disease J Virol Methods 70 213–217 Occurrence Handle9562415 Occurrence Handle1:CAS:528:DyaK1cXitVKhurs%3D Occurrence Handle10.1016/S0166-0934(97)00181-X

    Article  PubMed  CAS  Google Scholar 

  • SM Reid SS Grierson NP Ferris GH Hutchings S Alexandersen (2003) ArticleTitleEvaluation of automated RT-PCR to accelerate the laboratory diagnosis of foot-and-mouth disease virus J Virol Methods 107 129–139 Occurrence Handle12505626 Occurrence Handle1:CAS:528:DC%2BD38Xpslansb4%3D Occurrence Handle10.1016/S0166-0934(02)00210-0

    Article  PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Dukes, J., King, D. & Alexandersen, S. Novel reverse transcription loop-mediated isothermal amplification for rapid detection of foot-and-mouth disease virus. Arch Virol 151, 1093–1106 (2006). https://doi.org/10.1007/s00705-005-0708-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00705-005-0708-5

Keywords

Navigation