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Journal of Clinical Microbiology, March 2000, p. 1191-1195, Vol. 38, No. 3
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Detection and Identification of Human Parainfluenza Viruses 1, 2, 3, and 4 in Clinical Samples of Pediatric Patients by Multiplex Reverse Transcription-PCR

Jose C. Aguilar,1,* María P. Pérez-Breña,1 María L. García,2 Nieves Cruz,1 Dean D. Erdman,3 and Juan Emilio Echevarría4

Servicio de Virología1 and Servicio de Microbiología Diagnóstica,4 Centro Nacional de Microbiología, Instituto de Salud Carlos III, Carretera de Majadahonda Pozuelo s/n, 28220 Majadahonda, and Servicio de Pediatría, Hospital Severo Ochoa, Leganes,2 Madrid, Spain, and Respiratory and Enteric Viruses Branch, Division of Viral and Rickettsial Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 303333

Received 12 July 1999/Returned for modification 21 September 1999/Accepted 8 December 1999

We describe a multiplex reverse transcription-PCR (m-RT-PCR) assay that is able to detect and differentiate all known human parainfluenza viruses (HPIVs). Serial dilution experiments with reference strains that compared cell culture isolation and m-RT-PCR showed sensitivities ranging from 0.0004 50% tissue culture infective dose (TCID50) for HPIV type 4B (HPIV-4B) to 32 TCID50s for HPIV-3. As few as 10 plasmids containing HPIV PCR products could be detected in all cases. When 201 nasopharyngeal aspirate specimens from pediatric patients hospitalized for lower respiratory illness were tested, m-RT-PCR assay detected 64 HPIVs (24 HPIV-3, 23 HPIV-1, 10 HPIV-4, and 7 HPIV-2), while only 42 of them (21 HPIV-1, 14 HPIV-3, 6 HPIV-2, and 1 HPIV-4 isolates) grew in cell culture. Our m-RT-PCR assay was more sensitive than either cell culture isolation or indirect immunofluorescence with monoclonal antibodies for the detection of HPIV infections. Also, HPIV-4 was more frequently detected than HPIV-2 in this study, suggesting that it may have been underestimated as a lower respiratory tract pathogen because of the insensitivity of cell culture.

* Corresponding author. Mailing address: Centro Nacional de Microbiología, Instituto de Salud Carlos III, Carretera de Majadahonda Pozuelo s/n, 28220 Majadahonda, Madrid, Spain. Phone: 34-91-5097901. Fax: 34-91-5097966. E-mail: jaguilar{at}isciii.es.

Journal of Clinical Microbiology, March 2000, p. 1191-1195, Vol. 38, No. 3
0095-1137/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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