Human rhinovirus (HRV), a single-stranded, positive-sense RNA virus, is associated with mild upper respiratory tract infections in children. The aim of this study was to carry out a molecular characterization and phylogeny-based classification of the circulating genotypes of HRV in hospitalized children with clinical manifestations of acute lower respiratory infection in Paraguay. Nasopharyngeal aspirates were collected from 101 children under 5 years of age, hospitalized with symptoms of acute lower respiratory infection, between May 2010 and December 2011, at the largest public pediatric hospital in the Central Department of Paraguay. Detection was performed by a real-time polymerase chain reaction, followed by conventional amplification of the VP4/VP2 genomic region, sequencing, and phylogenetic analysis. Rhinovirus was detected in 33.7% of the samples. Amplification of 18 samples showed the presence of all three species (HRV-A, -B, and -C). Different genotypes were found for each species: 11 for HRV-A (-9, -12, -22, -30, -36, -43, -59, -61, -68, -88, and -89), one for HRV-B (-4), and four for HRV-C (-C2, -C3, -C6, and -C9). In South America, information about HRV diversity is scarce. This is the first report on HRV genotype diversity in South America. J. Med. Virol. 85:1645–1651, 2013. © 2013 Wiley Periodicals, Inc.