Detection of rhinoviruses by tissue culture and two independent amplification techniques, nucleic acid sequence-based amplification and reverse transcription-PCR, in children with acute respiratory infections during a winter season

Abstract

Five hundred seventeen consecutive nasopharyngeal aspirates were collected between October 1998 and May 1999 for episodes of acute respiratory tract infections in children presenting at the University Hospital of Antwerp. Culture and nucleic acid amplification techniques--nucleic acid sequence-based amplification (NASBA) and reverse transcription-PCR (RT-PCR)--were applied to detect rhinoviruses (RVs). Other respiratory viruses were detected by immunofluorescence (IF) analysis of the specimens and IF analysis of shell vial cultures. Among the 517 specimens, 219 viral agents were identified. They were, in decreasing order, rhinoviruses (93 [18.0%]), respiratory syncytial virus (76 [14.7%]), adenoviruses (16 [3.1%]), influenza viruses (15 [2.9%]), enteroviruses (15 [2.9%]), and herpes simplex virus (4 [0.8%]). For the evaluation of rhinovirus detection, culture positivity and/or a positive reaction in the two independent amplification methods was used as an expanded "gold standard." Based on this standard, the sensitivity, specificity, positive predictive value, and negative predictive value of culture were 44.7, 100, 100, and 99.8%, and those of NASBA and RT-PCR were 85.1, 98.3, 83.3, and 98.5% and 82.9, 93.4, 55.7, and 98.2%, respectively. NASBA and RT-PCR produced comparable results and were significantly more sensitive than virus culture. RVs showed the highest incidence in acute respiratory tract infections in children.

FIG. 1.

RV 5′ NCR subregion sequences of NASBA-negative specimens. Superscript letters: a, group A RV; b, RV (MW > 380 bp). P1 and P2 are 5′ NCR subregions chosen for amplification by the NASBA primers. Biotin probe and ECL-probe sequences are 5′ NCR subregions chosen for amplicon hybridization with the NASBA ECL probes.

FIG. 2.

Temporal distribution of respiratory virus (RSV, influenza virus [IFL], adenovirus [ADE], HSV, enterovirus [ENT], and RV) cases by month between 1 October 1998 and 31 May 1999. Nr, number.