Biological properties of two mengovirus mutants, 205 and 280, were compared to those of wild-type virus. The mutants exhibited alterations in plaque morphology, hemagglutination, and virulence in mice, but were not temperature-sensitive. Agglutination of human erythrocytes by mengovirus was dependent on the presence of sialic acid on the erythrocyte surface; however, free sialic acid failed to inhibit hemagglutination. Glycophorin, the major sialoglycoprotein of human erythrocyte membranes, exhibited receptor specificity for wild-type virus, but not for mutants 205 or 280. Cross-linking studies indicated that glycophorin exhibited binding specificity for the alpha (1 D) structural protein. The LD50 titers for wild-type mengovirus were 7 and 1500 plaque forming units (PFU) in mice infected intracranially (IC) and intraperitoneally (IP), respectively. However, mice infected IC or IP with 10(6) or 10(7) PFU of mutant 205 or 280 did not exhibit symptoms indicative of virus infection. Revertants were isolated from the brains of mice infected with mutant 205, but not from the brains of mice infected with mutant 280. The biological characterization of the revertants indicated that hemagglutination and virulence may be phenotypically-linked traits.