Differentiation between two samples of the polychaete Perinereis cultrifera collected in a brackish-water habitat and in an adjacent marine site at Elba Island (Italy) was assessed by morphological (number of paragnaths on the zones of the pharynx) and genetic (allozymes) approaches. Mean number of paragnaths differed significantly in zones IIl, IIr, III and V by t-test (P<0·05). Multidimensional scaling performed on Manhattan distances separated the individuals in two groups with a narrow zone of juxtaposition. Electrophoretic analyses of 22 allozyme loci revealed high levels of both within- and between-sample genetic heterogeneity, with a large number of private alleles. A general deficit of heterozygotes accounted for deviations from the Hardy–Weinberg equilibrium. Nei's genetic distance (D=0·185±0·018) and the mean value of the coancestry coefficient (Θ=0·381± 0·123, P<0·001 by permutation test) indicated considerable genetic divergence between the two samples. We concluded that the two groups can be assigned to sibling species. The observed divergence between the two samples may results from absence of gene flow and subsequent microallopatric differentiation through genetic drift, or, assuming migration between populations of the two sites, the model of divergence with gene flow may be compatible with this system.