Diagnosis of enterovirus and rhinovirus infections by RT-PCR and time-resolved fluorometry with lanthanide chelate labeled probes

Detection of enteroviruses and rhinoviruses has traditionally been based on laborious and time-consuming virus isolation. Recently, rapid and sensitiv e assays for detecting enterovirus and rhinovirus genomic sequences by reve rse transcription-polymerase chain reaction (RT-PCR) have been introduced. An RT-PCR assay is described that amplifies both enteroviral and rhinoviral sequences, followed by liquid-phase hybridization carried out in a microti ter plate format. In the hybridization assay, amplicons are identified by e nterovirus- or rhinovirus-specific probes carrying lanthanide chelate label s, which can be detected simultaneously by time-resolved fluorometry. The s ensitivity and specificity of the RT-PCR-hybridization method were evaluate d with a representative collection of enteroviruses and rhinoviruses and te sted further its applicability to the clinical setting with cerebrospinal f luid samples and nasopharyngeal aspirates. The RT-PCR assay amplified all e nteroviruses and rhinoviruses tested, and all but one amplicon gave a posit ive result in the subsequent hybridization assay. The RT-PCR-hybridization method was more sensitive than virus isolation for the detection of enterov iruses and rhinoviruses in the clinical samples. High sensitivity, rapidity , and easy performance make the assay suitable for the routine diagnosis of enterovirus and rhinovirus infections. (C) 1999 Wiley-Liss, Inc.