Serotype-specific identification of enterovirus 71 by PCR

Background: Enterovirus 71 and coxsackievirus A16 are closely related genetically and are causative agents of hand foot and mouth disease. Because enterovirus 71 is more often associated with severe neurological disease, there is a need to rapidly discriminate between enterovirus 71 and coxsackievirus A16 during hand, foot, and mouth disease outbreaks. Objectives: Our goal was to develop and evaluate a serotype-specific reverse transcription-polymerase chain reaction (RT/PCR)-based typing method for enterovirus 71. Study design: Two sets of PCR primers were designed to match conserved amino acid intervals of enterovirus 71. One diagnostic primer pair contains deoxyinosine at sites of 4-fold codon degeneracy. A second primer pair was designed for use in sequencing and molecular epidemiology studies. Primer pairs were tested on strains encountered in routine diagnostic samples. Results: Using both sets of primers on a panel of 61 prototype enteroviral strains, both primer pairs gave strong positive signals for only enterovirus 71. These primers amplified all enterovirus 71 isolates tested and discriminated between enterovirus 71 and the most closely related enterovirus, coxsackievirus A16. Conclusions: Our RT-PCR assay can be used for specific identification of enterovirus 71 clinical isolates. Furthermore, the 484-bp product of one primer pair has proven useful in sequencing studies to identify distinct genotypes of enterovirus 71. Published by Elsevier Science B.V.