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A new human coronavirus (HCoV), HCoV-NL, was recently reported for Dutch patients with acute respiratory tract infections (ARTI). Little information is available on the incidence, clinical manifestations and epidemiologic features of HCoV-NL infections.We performed a prospective study of symptomatic (case subjects with ARTI) and asymptomatic (control subjects undergoing elective surgery) children, ≤3 years of age, hospitalized in Canada during 2 winter seasons (2001–2003), to look at the prevalence of respiratory viruses. Reverse transcription-PCR assays were used to retrospectively detect HCoV-NL and to correlate its presence with clinical symptoms.HCoV-NL was detected in nasopharyngeal aspirates from 3.0% of young children (12 of 396 children) hospitalized for treatment of ARTI (case subjects), compared with 1.7% of asymptomatic control subjects (3 of 177 children) (P = 0.6). Nine (75.0%) of the symptomatic children had mixed viral infections. The mean age and mean duration of hospitalization of case subjects were 10.1 months and 4.9 days, respectively. Final diagnoses consisted of bronchiolitis or bronchitis (9 of 12 cases), pneumonitis (1 of 12 cases) and upper respiratory tract infections (2 of 12 cases), although 2 of 3 subjects with single HCoV-NL infections had upper respiratory tract infections only. Sequence analysis of the 1a and spike genes revealed that multiple HCoV-NL strains circulated in the same geographical area in each of the 2 winter seasons. Variability was more pronounced for the spike gene, with 2 clusters of strains.HCoV-NL was not a major respiratory pathogen in Canada during our study, as shown by its low detection rate in hospitalized children with ARTI, coupled with the high frequency of additional pathogens and its occasional detection in healthy children.