Human coronaviruses (HCoVs) are a common etiological agent of acute respiratory tract infections. HCoV infections, especially those caused by the two HCoVs identified most recently, NL63 and HKU-1, have not been characterized fully. To evaluate the prevalence and clinical presentations of HKU1 and NL63 in adults with acute respiratory tract infections, an investigation of HCoV infections in Beijing, China from 2005 to 2009 was performed by using reverse transcriptase PCR assays and sequencing analysis. Among 8,396 respiratory specimens studied, 87 (1%) clinical samples were positive for HCoVs, of which 50 samples (0.6% of the total) were positive for HCoV-OC43, 15 (0.2%) for HCoV-229E, 14 (0.2%) for HCoV-HKU1, and 8 (0.1%) for HCoV-NL63. The prevalence of HCoV infection in adults exhibited distinct seasonal fluctuations during the study period. In addition, patients positive for HCoV-229E infections were more likely to be co-infected with other respiratory viruses. Enterovirus, rhinovirus, and parainfluenza virus type 3 were the most common viruses found in patients with HCoV infections. The demographic and clinical data present in this study of HCoV infections in adults with acute respiratory tract infections should improve our understanding of the pathogenesis of HCoVs.