Cytogenetical analyses based on conventional and differential staining were performed for the first time on five species of the Brazilian lizard genus Enyalius: E. bibronii, E. bilineatus, E. iheringii, E. leechii, and E. perditus. The species share a similar 2n = 36 (12M + 24m) karyotype, comprised of 12 metacentric or submetacentric macrochromosomes, except for an acrocentric pair 6 that characterizes E. bibronii. The 24 microchromosomes were acrocentrics, but in E. perditus two meta/submetacentric microchromosome pairs were unambiguously identified. Karyotypes with 2n = 37 and 2n = 37/38 chromosomes were also observed in some specimens of E. bilineatus as a result of the presence of supernumerary chromosomes (Bs). Ag-NORs were always located at the distal region of the long arm of the submetacentric pair 2. The constitutive heterochromatin was mostly restricted to the pericentromeric regions of some macrochromosomes and microchromosomes. A XX:XY mechanism of sex determination with a dot-like Y microchromosome occurs in E. bilineatus, E. leechii, and E. perditus.