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Dasycercus cristicauda and Dasycercus blythi are significant mesopredators in arid Australia and both species are threatened nationally. We investigated size dimorphism, habitat use, and population dynamics of sympatric populations during a low rainfall period in central Australia and examined congruence between morphological and molecular methods of species identification. Our molecular analyses confirmed the presence of two Dasycercus species in sympatry and showed that the descriptions of Woolley (2005) allowed confident identification of the two species in the field. We captured 23 D. cristicauda and 55 D. blythi over 39 months on 20 monitoring plots and another 49 survey sites stratified across five habitat types within a 7000 km(2) area. Both species showed sexual dimorphism in body mass with males being significantly heavier. The species typically occupied non-overlapping habitat; D. cristicauda was restricted to sandridges (n = 4 sites) with an understorey dominated by spinifex (Triodia), whereas D. blythi occupied sand plain and gibber plain, the latter being a new habitat for the species. Capture rate of D. cristicauda peaked in autumn-winter 2008 and winter 2009. D. blythi exhibited similar variability with peaks in winter 2009 and summer-autumn 2010. We consider that the relatively low abundance of each species during low rainfall periods makes them vulnerable to predation by feral house cats and European red foxes.