The study was undertaken for antigenic characterization of rotavirus strains isolated from South African children. During July 1996-July 1997, an epidemiological surveillance of rotavirus-associated gastroenteritis was carried out in Pretoria and Gauteng provinces. In total, 1,229 diarrhoeal faecal samples were collected from a pathology laboratory. Of 389 (32%) rotavirus strains detected, most (97%) were group A rotaviruses by serological assay. However, 12 rotavirus strains identified did not apparently carry the common group A-specific antigen as determined by both commercial and monoclonal antibody-based ELISAs. Electrophoretic analysis of the viral RNA genome revealed that these strains shared a common electropherotype and that this resembled the group A rotavirus constellation of RNA segments. Furthermore, the conserved terminal sequences of the group A VP6 and VP7 genes could not be targeted by standard reverse transcriptase-polymerase chain reaction with a routine set of primers to the VP6 or VP7 genes. These strains present an interesting phenotypic variation of the recognized rotaviruses and warrant further characterization.