Legacy waterlogging shapes the associations between barley cultivars and their rhizosphere mycobiomes

Climate change is intensifying rainfall events, leading to increased flood risk and substantial threats to crop productivity. Waterlogging affects both above- and belowground ecosystem processes, yet the response of plant-associated fungi to the long-term (legacy) effects of extreme rain events in agricultural fields remains unclear for many crops, especially traditional cultivars being explored for sustainable production. This study established a field experiment to examine the legacy effects of waterlogging on arbuscular mycorrhizal fungi (AMF) colonization, as well as the structure and composition of the rhizosphere mycobiome of barley cultivars using ITS amplicon sequencing. Plant traits were measured to link physiological changes with shifts in mycobiome structure and composition, while ecological networks were analysed to understand the associations between barley cultivars and rhizosphere fungi. Legacy waterlogging was associated with reduced barley productivity and significantly altered the structure and composition of the rhizosphere mycobiome of barley. Specifically, waterlogging contributed to a significant reduction in fungal richness, an increase in symbiont abundance and AMF colonization, and a decrease in saprotrophic fungal richness. The reduction in rhizosphere fungal diversity was associated with a significant decline in barley grain protein content. Additionally, the abundance of the pathogenic fungus Alternaria was significantly reduced with increased AMF colonization, resulting in lower abundance under legacy waterlogging conditions. Ecological network analysis uncovered a core-periphery and anti-nested structure in the relationships between barley cultivars and the rhizosphere mycobiome. This indicates a core group of generalist fungi that perform similar functions, while peripheral fungi tend to be more specialized and associate with only one cultivar. These network patterns depended on different barley cultivars exhibiting preferential associations with specific rhizosphere fungi, and these associations were altered by legacy waterlogging, resulting in shifts in mycobiome composition and structure within network modules. Overall, the results indicate that legacy waterlogging deeply impacts the fungal community structure in the rhizosphere of barley, suggesting the change is directly associated with barley’s response to waterlogging.