OBJECTIVE To investigate the molecular epideiological and clinical feature of human metapneumovirus in children with acute respiratory tract infection in Nanjing city, China. METHOD Nasopharyngeal aspirates and nasopharyngeal swab were taken from 642 outpatients or hospitalized pediatric patients with acute at the Children Hospital of Nanjing, Jiangsu Province, China, between August 2009 and July 2010. Respiratory speciments were tested for the M gene of hMPV by reverse-transcription polymerase chain reaction (RT-PCR). All RT-PCR positive products were sequenced and phlogenetic analysis was conducted. RESULT hMPV was detected in 35 (5.5%) of the 642 children. Phylogenetic analysis revealed that 51.4% of the hMPV were B1, 31.4% were A2b. The peak of the positive rate was in April. The majority of the hMPV-positive patients(71.4%) were 0-1 years old. Of the 35 hMPV-positive patients, 15 (42.8%) were co-infected with other respiratory viruses, and human rhinovirus (HRV) were the most common additional respiratory virus. The most common clinical diagnosis was pneumonia (48.6%). CONCLUSION Human metapneumovirus is an important pathogen of acute respiratory tract infection in children in Nanjing city. The subtype B1 was the predominating lineage in 2009-2010 in Nanjing city. No significant differences were found for clinical characteristics between genotype A and genotype B human metapneumovirus infection in children in Nanjing.