Establishment of reverse transcription polymerase chain reaction assay for simultaneous detection of human parainfluenza virus types 1 to 4 in children with influenza like illnesses.

OBJECTIVES: The aim of this study was to develop an in-house multiplex reverse transcription polymerase chain reaction (mRT-PCR), which can recognize HPIV1-4 in clinical samples. BACKGROUND: Human parainfluenza virus (HPIV) is one of the major causes of viral respiratory infections and can affect people at any age, especially infants and young children. METHODS: Four sets of specific primers targeting conserved areas of hemagglutinin-neuraminidase (HN) genes of HPIV1-4, were designed and tested with type-related plasmid controls. Specificity and sensitivity of mPCR were tested. One-step mRT-PCR was set up using a viral panel containing 10 respiratory viruses, including HPIVs. One hundred nasopharyngeal samples of respiratory infection patients were tested using the set One-step mRT-PCR. RESULTS: The specificity of set mPCR for HPIV1-4 using plasmid positive controls was proved and reaction sensitivity was measured. The specificity of set mRT-PCR was confirmed and 4 and 5 out of 100 clinical samples were HPIV1 and HPIV2 positive, respectively. CONCLUSION: The developed one-step mRT-PCR in this study is an effective and specific assay for clinical diagnosis of HPIV1 to 4 (Tab. 1, Fig. 6, Ref. 28).