The chromosomal complements of nine species of Blenniidae (Aidablennius sphylnx, Blennius ocellaris, Lypophris adriaticus, L. pavo, L. trigloides, Parcablennius gattorugine, P. ponticus, P. sanguinolentus, P. tentacularis) from the Adriatic Sea were analysed with several banding methods and in-situ hybridization. In all species, the diploid set consists of 48 mostly acrocentric chromosomes and has a similar location (terminal centromeric) of NORs, except for L. pavo (interstitial pericentric) and P. ponticus (terminal on the long arm). There are major differences in karyotype with regard to the amount and distribution of heterochromatin. Parablennius tentacularis shows a distinctive sex-chromosome system involving 2n = 48 males with a large totally heterochromatic Y chromosome, and males with 2n = 47. This difference is likely to be the consequence of a translocation of an autosome on the original Y. This finding constitutes an additional instance of the great variability in origins of multiple sex chromosome systems in vertebrates.