Background/Question/Methods This project attempts to quantify the resilience of prairie ecosystems to climate change in the Pacific Northwest (PNW). In this region, prairie ecosystems currently sustain ~1.3 million beef cows and calf production costs are expected to increase to offset drought-induced plant productivity loss. Here, we investigate patterns of asymbiotic nitrogen fixation (ANF) and biogeochemical controls, that also influence plant community composition and prairie productivity, under experimental drought to address a major challenge for sustainable agriculture in the region. We hypothesize that the effect of drought on prairie vegetation cover increases soil asymbiotic N inputs by diminishing the dominance of symbiotic root-fungal networks. To test this hypothesis, we quantified the impacts of decadal drought stress on soil ANF using 15N-labeled dinitrogen (15N2) incubations of soils from high- and low-diversity prairies across a 520-km latitudinal gradient (i.e., southern Oregon-SOR, central Oregon-COR, and central Washington-CWA) representing increasingly severe Mediterranean conditions. We also quantified total soil organic carbon-C, total, and available N, and available phosphorus-P and iron-Fe pools to better understand underlying mechanisms governing drought-induced changes in ANF. At each site, composite soil samples (n = 3) were collected from five co-located high- and low-diversity prairie plots under control (ambient) and drought (-40% precipitation) conditions. Results/Conclusions We found that soil ANF response to drought increased with the PNW Mediterranean drought intensity gradient; while ANF rates increased nearly two-fold in the southernmost site (SOR), a significant decrease in ANF was verified in the northernmost site (CWA). ANF response to drought also varied depending on plant diversity, where low-diversity prairies had a more predictable response to drought than high-diversity prairies. For instance, ANF in SOR high-diversity prairies was suppressed but no change was verified in COR high diversity prairies. Soil C and N contents were generally higher in high-diversity prairies whereas treatment had no significant effect across sites. Soil P availability, also affected by drought, and pH were the most important variables explaining ANF variability across vegetation types and sites. Based on our findings, low-diversity prairies in central WA may be those most severely impacted by increased climate change-induced drought stress. Our study highlights the importance of using soil-plant-atmosphere interactions to assess prairie ecosystem resilience to drought in the PNW.