Big Chico Creek (Northern California, USA) is one of the region’s few remaining undammed streams, maintaining a natural flow regime while traversing steep elevational and urbanization gradients. Decades of benthic macroinvertebrate (BMI) sampling show strong community turnover along this gradient, from sensitive specialist taxa at higher elevations to more generalist assemblages downstream. We tested the relative influence of elevation, urbanization, and other environmental predictors on BMI community composition. Ordination analyses show elevation as the dominant driver of community separation among sites. We further evaluate associations between community patterns and additional predictors (e.g., habitat and water-quality variables) to assess how correlated environmental factors may interact with elevation and urbanization to shape BMI assemblages. These findings clarify which gradients most strongly structure communities in Big Chico Creek and inform expectations for how BMI assemblages may respond to environmental change across mixed elevational–urban stream networks.