Microbial diversity loss affects old and modern barley cultivars differently under varying nitrogen sources

Soil microbial diversity is crucial for plant nutrition and health, yet how its loss affects plant performance remains unclear. We used a dilution-to-extinction approach to test how declining rhizo-microbiome diversity influences two barley cultivars: the modern RGT Planet and the older Babushka. Plants were grown in sterilized systems amended with mineral or organic nitrogen and inoculated with microbiome treatments (10^-¹, 10^-³, 10^-⁵, and 10^-⁷ dilutions), plus a no-inoculum treatment. We used amplicon sequencing (16S, ITS, 18S) to profile rhizosphere communities, and quantified plant biomass, shoot nitrogen, and chitin mineralization. Protists and fungi were present in 10^-¹ and 10^-³ but absent in all others. Microbiome inoculum and nitrogen source explained most variation in rhizo-microbiome composition, with cultivar having a smaller effect. Under organic nitrogen, Babushka showed a marked decline in biomass with decreasing diversity, whereas RGT was largely unaffected, indicating that the older cultivar relied more on a diverse microbiome to maintain growth. At intermediate diversity, when protists and fungi were lost, both cultivars showed improved growth and shoot nitrogen, coinciding with shifts in bacterial composition and loss of potential pathogens. Hence, reduced diversity did not always impair growth, suggesting functional compensation. Under mineral nitrogen, both cultivars were less sensitive to diversity loss. Overall, nitrogen source and cultivar identity modulated plant responses to microbial diversity loss. Diverse microbiomes promoted efficient use of organic nitrogen, particularly for the older cultivar, while the modern cultivar maintained growth at lower diversity. Our results demonstrate that the consequences of diversity loss are context-dependent and cultivar-specific.