Small but significant differences were found in allele frequencies among five populations (overall FST estimate ()=0·004, P=0·006; overall RST estimate (RHO)=0·019, P<0·00001) of the demersal cichlid Copadichromis sp. ‘ virginalis kajose ’, collected from five locations in Lake Malawi. Pairwise FST estimates revealed significant differences between the most southerly population (Cape Maclear), and the three most northerly populations (Mbamba Bay, Metangula and Chilola). Pairwise RST estimates also revealed significant differences between some populations, but no geographical pattern was discernible. There was no evidence of isolation by distance using either the shortest straight-line distance between samples, or the distance around the shoreline following a 50 m depth contour. FST estimates were considerably lower than found in previous studies on the mbuna (rock-dwelling species), but higher than those found in a study of three pelagic cichlid species from Lake Malawi. Substructuring in C. sp. ‘ virginalis kajose ’ appears to be on a similar scale to the Atlantic cod.