Non-symbiotic nitrogen (N₂) fixation by diazotrophic bacteria is a potential source for biological N inputs in non-leguminous crops and pastures. Perennial grasses generally add larger quantities of above- and belowground plant residues to soil, and so can support higher levels of soil biological activity than annual crops. In this study, the hypothesis is tested that summer-active perennial grasses can provide suitable microsites with the required carbon supply for N₂ fixation by diazotrophs, in particular during summer, through their rhizosphere contribution. In a field experiment on a Calcarosol at Karoonda, South Australia, during summer 2011, we measured populations of N₂-fixing bacteria by nifH-PCR quantification and the amount of ¹⁵N₂ fixed in the rhizosphere and roots of summer-active perennial grasses. Diazotrophic N₂ fixation estimates for the grass roots ranged between 0.92 and 2.35 mg ¹⁵N kg^–1 root day^–1. Potential rates of N₂ fixation for the rhizosphere soils were 0.84–1.4 mg ¹⁵N kg^–1 soil day^–1 whereas the amount of N₂ fixation in the bulk soil was 0.1–0.58 mg ¹⁵N kg^–1 soil day^–1. Populations of diazotrophic bacteria in the grass rhizosphere soils (2.45 × 10⁶ nifH gene copies g^–1 soil) were similar to populations in the roots (2.20 × 10⁶ nifH gene copies g^–1 roots) but the diversity of diazotrophic bacteria was significantly higher in the rhizosphere than the roots. Different grass species promoted the abundance of specific members of the nifH community, suggesting a plant-based selection from the rhizosphere microbial community. The results show that rhizosphere and root environments of summer-active perennial grasses support significant amounts of non-symbiotic N₂ fixation during summer compared with cropping soils, thus contributing to biological N inputs into the soil N cycle. Some pasture species also maintained N₂ fixation in October (spring), when the grasses were dormant, similar to that found in soils under a cereal crop. Surface soils in the rainfed cropping regions of southern Australia are generally low in soil organic matter and thus have lower N-supply capacity. The greater volume of rhizosphere soil under perennial grasses and carbon inputs belowground can potentially change the balance between N immobilisation and mineralisation processes in the surface soils in favour of immobilisation, which in turn contributes to reduced N losses from leaching.