Detection and identification of human parainfluenza viruses 1, 2, 3, and 4 in clinical samples of pediatric patients by multiplex reverse transcription-PCR.
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 (TCID(50)) for HPIV type 4B (HPIV-4B) to 32 TCID(50)s 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.
Aguilar
J C
JC
Servicio de Virología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Carretera de Majadahonda Pozuelo s/n, 28220 Majadahonda, Madrid, Spain. jaguilar@isciii.es
Pérez-Breña
M P
MP
García
M L
ML
Cruz
N
N
Erdman
D D
DD
Echevarría
J E
JE
D000818:Animals; D002460:Cell Line; D002648:Child; D017931:DNA Primers; D004279:DNA, Viral / Q000302:isolation & purification; D004285:Dogs; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D001239:Inhalation; D010222:Parainfluenza Virus 1, Human / Q000302:isolation & purification*; D010223:Parainfluenza Virus 2, Human / Q000302:isolation & purification*; D010224:Parainfluenza Virus 3, Human / Q000302:isolation & purification*; D010253:Respirovirus Infections / Q000175:diagnosis*; D012189:Retrospective Studies; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D019213:Rubulavirus / Q000302:isolation & purification*; D019351:Rubulavirus Infections / Q000175:diagnosis*; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D004279:DNA, Viral