Abstract Comparative genomics of viruses in evolutionary and phylogenetic studies is well established. Previous nucleic acid sequence analyses have demonstrated that enteroviruses and rhinoviruses of the family Picornaviridae exhibit a similar structure of the 5′-nontranslated region (NTR) differing significantly from the 5′-NTR of cardiovirus, aphthovirus, hepatovirus, and echovirus 22 (provisionally parechovirus 1). Available nucleotide sequence information of the 5′- and 3′-nontranslated regions of more than 70 serotypes of enteroviruses, bovine enteroviruses and rhinoviruses has been compared and correlated with previous findings obtained after analysis of the coding and noncoding genome regions. As a result, the 5′- and 3′-NTRs of all three virus groups are characterized by group-specific nucleotide sequences. Focusinng on bovine enterovirus (BEV) serotypes, unique characteristics in all secondary structures of the NTRs were observed. These features clearly separate the BEVs from the human enteroviruses and rhinoviruses. Concerning the 5′-NTR, the most remarkable property is an insertion of about 110 nucleotides between the putative cloverleaf structure at the very 5′-end of the viral genome and the IRES element. This insertion was demonstrated for BEV 1 and 2 and has a predicted folding pattern which is very similar to the 5′-cloverleaf structure. One stem-loop of this second cloverleaf is almost identical to the 3CD pro-binding domain of rhinoviral 5′-cloverleafs. It was also demonstrated that the IRES elements and the 3′-NTRs of both, enteroviruses and rhinoviruses, have group-specific features which differ significantly from the corresponding genome regions of BEV. These results suggest that bovine enteroviruses hold an exceptional taxonomic position besides the established genera Enterovirus and Rhinovirus. Within the Enterovirus and Rhinovirus genera, the existence of virus clusters representing subgenera was previously proposed. Whereas the 5′-NTRs of the four human enterovirus clusters fall into two groups, all four clusters have characteristic secondary structures at the 3′-NTR supporting the concept of enterovirus clusters. For rhinoviruses, the existence of two virus clusters was confirmed.