The purpose of this study was to examine population structure based on the polymorphisms of biallelic (M8, YAP, SRY-1532, 92R7, SRY-8299/4064, 12f2, M20, M52, M70, M89, M124, M170, M201, M173, M175) and eight microsatelitte markers (DYS19, DYS388, DYS389I, DYS389II, DYS390, DYS391, DYS392 and DYS393) in 457 Croatian Y chromosomes. Genetic composition was assessed based on the allelic frequencies, whereas genetic variability was based on haplotype and locus diversity. Phylogenetic relationships among microsatellite haplotypes within haplogroups were determined by using the reduced median network and median joining network, as suggested by Bandelt et al. (1999). Analysis of the biallelic markers allowed characterization of nine different haplogroups. Haplogroup I was found at the highest frequency (48.4%) followed by haplogroup R1a (24.7%). Its higher frequency in the southern island population is contrasted with higher frequency of group R1a chromosomes in the northern island of Krk and in the mainland population. Since Croats belong to a Slavic linguistic family, R1a speaks in favor of possible Slavic genetic impact into their Y-chromosomal pool. Haplogroups J, G and E that can be related to the spread of farming characterize the minor part (12.5%) of the Croatian paternal lineages, which corresponds to that observed among the western European populations. In one of the southern island (Hvar) populations, we found relatively high frequency (14%) of lineages belonging to P* (xM173) cluster which is unusual for European populations. Interestingly, the same population also harbored mitochondrial haplogroup F that is virtually absent in European populations.