A real-time RT-PCR for rapid detection and quantification of mosquito-borne alphaviruses

Marilia Farignoli Romeiro, William Marciel de Souza, Aline Lavado Tolardo, Luiz Carlos Vieira, Dyana Alves Henriques, Jansen de Araujo, Carlos Eduardo Hassegawa Siqueira, Tatiana Elias Colombo, Victor Hugo Aquino, Benedito Antonio Lopes da Fonseca, Roberta Vieira de Morais Bronzoni, Maurício Lacerda Nogueira, Edison Luiz Durigon, Luiz Tadeu Moraes Figueiredo

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

Abstract

Mosquito-borne alphaviruses are widely distributed throughout the world, causing important human illnesses. Therefore, the development of methods to enable early diagnosis of infections by alphavirus is essential. We show here the development and evaluation of a quantitative real-time RT-PCR using genus-specific primers to the nsP1 viral gene of all mosquito-borne alphaviruses. The specificity and sensitivity of the assay were tested using seven alphaviruses and RNA transcribed from Venezuelan equine encephalitis virus. The detection limits of real-time RT-PCR ranged from 10 to 76 copies per ml. The melting temperature (TM) values for amplification of the alphavirus genomes were 83.05 °C and 85.28 °C. Interestingly, the assay suggested the possibility the arthritogenic alphaviruses with TM peaks of 84.83 to 85.28 °C and encephalitic alphaviruses of 83.34 °C to 84.68 °C could be discriminated both diseases. Real-time RT-PCR may prove very useful for the screening and preliminary diagnosis in outbreaks and surveillance studies as well as for measuring the viral load in pathogenesis studies.

Original languageEnglish
Pages (from-to)3171-3177
Number of pages7
JournalArchives of Virology
Volume161
Issue number11
DOIs
StatePublished - Nov 1 2016

Bibliographical note

Publisher Copyright:
© 2016, Springer-Verlag Wien.

ASJC Scopus subject areas

  • Virology

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