The Australian sea lion, Neophoca cinerea, one of the world's rarest otariids, is notable for an asynchronous, aseasonal breeding chronology. Determining the ecological features that shape the genetic structure of marine predators such as Australian sea lions is challenging because their demersal foraging habitat is difficult to observe and quantify. Recent developments in stable isotope screening techniques using milk-dependent pups as proxies for maternal isotope signatures identified temporally stable, alternative (inshore and offshore) foraging ecotypes in adult female Australian sea lions. We combined this technique with mitochondrial DNA (mtDNA) analysis of samples of 40-60% of all pups produced at 17 of the largest South Australian colonies to determine whether ecological specialization in foraging ecotype within and between colonies has shaped maternal population structure within the species. Genetic isolation by distance was apparent at very fine geographical scales with three distinct clusters of colonies that share multiple haplotypes being interspersed with isolated breeding sites. There was no congruence between mtDNA haplotype distribution and foraging ecotypes suggesting that observed behavioural specialization was not maintained along matrilines. We propose that foraging specialization within discrete fine-scale foraging areas and habitats at the individual level limits the dispersive capacity of adult female Australian sea lions which in turn drives population structure. Given this species' vulnerability to anthropogenic impacts and the high degree of female population structure, determining the extent of male-mediated gene flow in this species is critical. Only then can breeding colony connectivity be established and appropriate management units identified for the species.