Rapid detection of parainfluenza virus type 3 RNA in respiratory specimens: Use of reverse transcription-PCR-enzyme immunoassay

Parainfluenza virus type 3 (PIV-3), an important lower respiratory tract pathogen in young children and immunocompromised individuals, may be underdiagnosed because of the insensitivity of available culturing systems and delay in identification of virus in cell culture. We developed a reverse transcription-PCR-enzyme immunoassay (RT-PCR-EIA) for PIV-3, using primers specific for a highly conserved region of the hemagglutinin-neuraminidase gene. Testing of nasal washes spiked with PIV-3 or other respiratory viruses showed that this assay detected seven strains of PIV-3 but not other respiratory viruses. Of 103 respiratory tract samples obtained from children experimentally infected with a live PIV-3 vaccine or naturally infected with wild-type PIV-3, 51 were positive by culture and 48 were positive by RT-PCR- EIA. Eleven of the culture-positive samples were negative by RT-PCR-EIA; however, none of these grew virus upon reinoculation into cell culture, indicating that virus was lost or was present at a very low titer. Eight of the culture-negative samples were positive by RT-PCR-EIA: two were obtained from a subject who was culture negative but had a serologic response to PIV- 3, four were obtained 7 to 9 days after the first positive culture, and two were obtained 1 day prior to the first positive culture. Thus, this RT-PCR- EIA for PIV-3 is sensitive and specific and can detect viral RNA in samples from which virus cannot be cultivated. This assay could be used for diagnosis late in the course of PIV-3 infection and for accurate detection of disease outbreaks.