Selection and evolutionary analysis in the nonstructural protein NSP2 of rotavirus A

► Mechanisms of evolution of the rotavirus A nonstructural protein NSP2 were examined. ► Rates of nucleotide substitution in NSP2 gene sequences were high. ► Strong negative selection was found to act on NSP2 protein sequences. ► Three sites of positive selection were associated with an antibody binding epitope. Rotavirus A is the leading cause of acute gastroenteritis in infants and young children worldwide. The nonstructural protein 2 (NSP2) plays essential roles in the replication cycle of rotavirus and may play a role in protective immunity against rotavirus disease. Using a Bayesian approach, we measured the mutation rate of genotype N1 NSP2 gene sequences. The N1 genotype is the main NSP2 genotype associated with rotavirus strains causing severe disease, and was found to have a high mutation rate (8.7×10−4 substitutions/site/year) in comparison to the rotavirus VP4 gene and rates of mutation in other RNA viruses. NSP2 has traditionally been considered as a conserved rotavirus protein and selection analysis indicated that the NSP2 protein was under strong negative selection, suggesting that most nucleotide substitutions were synonymous. This conservation is likely a result of functional constraints of NSP2 in the rotavirus replication cycle. Four sites of positive selection were identified; two of these (positions 249 and 255) were located in a previously characterised antibody binding epitope. The remaining sites were not located in known functional regions, and the reason for variation at these sites remains to be elucidated.