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 ORIGINAL ARTICLE
Year : 2011  |  Volume : 29  |  Issue : 2  |  Page : 110-117

Development of a new method for diagnosis of Group B Coxsackie genome by reverse transcription loop-mediated isothermal amplification


1 Department of Virology, King Institute of Preventive Medicine, Guindy, Chennai- 600 032, India
2 Department of Biotechnology, Adhiyamaan College of Engineering, Hosur - 635 109, India

Correspondence Address:
K Jaianand
Department of Virology, King Institute of Preventive Medicine, Guindy, Chennai- 600 032
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/0255-0857.81780

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Background: Coxsackie B viruses (genus, Enterovirus; family, Picornaviridae) can cause aseptic meningitis, encephalitis, pleurodynia, and fatal myocarditis, and are implicated in the pathogenesis of dilated cardiomyopathy. The differentiation of the group B Coxsackieviruses into their subtypes has potential clinical and epidemiological implications. Objective: In this study, we developed a one-step, single-tube genogroup-specific reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of group B Coxsackie genomes targeting 5′ UTR region. Materials and Methods: The amplification can be obtained in less than 1 hour by incubating all the reagents in a single tube with reverse transcriptase and Bst DNA polymerase at 63°C. Detection of gene amplification could be accomplished by agarose gel electrophoresis and the monitoring of gene amplification can also be visualised with the naked eye by using SYBR green I fluorescent dye. Results: A total of 40 samples comprising 31 positive samples and 9 negative samples were used in this study for comparative evaluation. The results were compared with those from Real-Time Polymerase Chain Reaction (RT-PCR). None of the RT-PCR-positive samples were missed by RT-LAMP, thereby indicating a higher sensitivity of the RT-LAMP assay. Conclusion: Thus, due to easy operation without a requirement of sophisticated equipment and skilled personnel, the RT-LAMP assay reported here is extremely rapid, cost-effective, highly sensitive, and specific and has potential usefulness for rapid detection of non-polio enterovirus (NPEV) not only by well-equipped laboratories but also by peripheral diagnostic laboratories with limited financial resources in developing countries.






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