Climate change-driven extreme events are reducing barley productivity. The high use of mineral fertilizers, combined with low nutrient use efficiency, leads to environmental and economic concerns. Indigenous arbuscular mycorrhizal fungi (AMF) inoculants offer a sustainable alternative, especially in intensive farming systems where AM colonization and diversity are low. However, poor adaptation to local conditions limits inoculant success. Few studies have tested indigenous AMF inoculated on field crops, with limited research on barley. No research has yet explored how barley genotype and environment modulate field inoculation outcomes in terms of crop productivity. Key factors such as AM fungal abundance and community structure shifts remain unidentified. This study evaluated the agroecological effects of an indigenous AM fungal consortium on three barley varieties (Atlante, Atomo, and Concerto) over 2 years. In 2020, Atomo and Concerto responded positively to inoculation in terms of root colonization, with grain yield increases of 64% and 37%, respectively. In 2021, only Concerto showed enhanced root colonization, while grain yield increased by 78% in Concerto and 134% in Atlante. Multivariate analysis revealed a strong impact of environment on barley productivity, with a significant third-order interaction among AMF, genotype, and environment. Inoculation slightly altered AM composition but strongly influenced community structure, particularly at different plant growth stages. Root colonization was strongly correlated with barley productivity, with root length containing arbuscules being the best predictor. Changes in the AM community structure, rather than composition, drove barley response, with Glomus and Septoglomus, present in the inoculum, being main players. These findings support the use of indigenous AMF for sustainable biofertilization and highlight the importance of selecting genotypes with a stable AM response across environments. Our results disclose for the first time the role of barley genotype and plant growth stage on AM host preference with and without indigenous AM fungal inoculants.