A multiplex RT-PCR for the detection of parainfluenza viruses 1-3 in clinical samples

Parainfluenza viruses (PIV) are an important cause of respiratory morbidity . Conventional diagnostic methods for detection of PTV are time consuming o r lack sensitivity. A multiplex PCR that detects PIV 1-3 was developed usin g novel primers for PIV viruses 1 and 2 and primers for PIV 3 described pre viously. Following RNA extraction a single multiplex reverse transcription was undertaken using antisense primers specific for each virus type. This w as followed by a 40-cycle multiplex PCR using primers directed towards the haemagglutinin-neuraminidase coding region of each virus type. Products wer e probed with type-specific fluorescein labelled internal probes and detect ed by chemiluminescence. Cultured PIV viruses were detectable to a sensitiv ity of 1 TCID50. The technique was applied to 57 nasal aspirates taken from children presenting with various acute respiratory conditions and analysed previously by culture, immunofluorescence and/or serology. It was possible to detect PIV 1, 2 or 3 in 13/13 samples found previously positive for PIV by tissue culture, 13/15 found previously positive by immunofluorescence a nd 6/10 that coincided with positive serology. None of the samples found pr eviously positive for other viruses (26) or negative to virus detection (6) were found positive by RT-PCR. It is concluded that this method is as sens itive as combined immunofluorescence and tissue culture for the detection o f the PIV viruses 1-3 and should be useful for rapid diagnosis of PIV 1-3 i nfections. (C) 1999 Elsevier Science B.V. All rights reserved.