show Abstracthide AbstractHuman rotaviruses replicate poorly in most cell lines. However, through serial passage, some rotavirus specimens can be adapted to replication in cultured cells. We serially passaged human rotavirus stool specimens representing five of the genotypes most frequently associated with severe human disease, each in triplicate, three to five times in primary monkey kidney cells then ten times in a monkey kidney cell line (MA104). We used Illumina next-generation sequencing and analysis to identify variants that arose during passage. A key goal was to identify changes that arose in attachment protein and tropism determinant VP4. The identification of conserved VP4 changes could suggest a conserved mechanism of culture adaptation. In the future, such a conserved mechanism could be exploited to study human rotavirus biology or efficiently manufacture rotavirus vaccines.