The use of chemicals to fertilize crops incurs economic and environmental costs and it is widely recognized that the current level of chemical fertilizer use is unsustainable in many intensive farming systems. Any methods that can reduce fertilizer input and still maintain acceptable yields would be of great benefit to both the farmer and the environment. The use of beneficial endophytes as crop inoculants may go some way towards improving crop yields beyond that achievable using fertilizer increases alone. Field trials were conducted over two seasons on three contrasting field sites to test the effects of fungal endophytes from a wild barley relative on three barley cultivars (Mickle, Planet and Propino). Seeds were either untreated or dressed with a consortium of four endophyte strains, and three levels of nitrogen (N) were applied to both treatments: full N, 50% N and 0 N. On the field site with the lowest overall N input, the endophyte treatment with 50% N restored yield for ‘Planet’ to that associated with untreated plants receiving the full N input. On the same site and with the same cultivar, endophyte treatment increased yield by 15% under full N, and by a mean 12% for all three cultivars with 50% N input. Over both seasons and all three sites, the endophyte treatment increased yield for the cultivar Planet by a mean of 9%. For the endophyte-associated increase in the variety Planet grain yield over the untreated trials strong correlations were found between increased yield and each of low rainfall, greater evaporation and greater number of degree days above the base. Furthermore, the efficacy of the endophytes was not removed by regular foliar fungicidal treatment. These results suggest that fungal endophytes can contribute to improving barley yield grown in low rainfall areas and under a range of fertilizer input regimes, provided that endophyte treatments are applied to compatible crop cultivars and sites.