Whole-genome, sequence-independent amplification and 454(®) pyrosequencing of a rotavirus SA11 cell culture sample with an unknown passage history yielded consensus sequences of twelve complete genome segments. Two distinct sequences for genome segment 8 (encoding NSP2) were present, indicating a mixed infection with two rotavirus SA11 strains. The genotypes of the viruses were G3-P[2]-I2-R2-C5-M5-A5-Nx-T5-E2-H5, where x was either 5 or 2. The strains were named RVA/Simian-tc/ZAF/SA11-N5/1958/G3P[2] and RVA/Simian-tc/ZAF/SA11-N2/1958/G3P[2]. The genotype (N2) and sequence of genome segment 8 of RVA/Simian-tc/ZAF/SA11-N2/1958/G3P[2] were identical to that of the bovine rotavirus O agent. Five novel amino acids were detected in minor population variants of three genome segments. Genome segment 1 (VP1) has a high nucleotide substitution rate, but the substitutions are synonymous. Distance matrices and Bayesian molecular clock phylogenetics showed that SA11-N2 is a reassortant containing genome segment 8 from the O agent, whereas SA11-N5 is a very close derivative of the prototype SA11-H96.