Divergent functional strategies of three fungal symbionts on iron/magnesium transport, photosynthetic carbon metabolism, and growth in citrus

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The cultivation of high-quality citrus cultivars such as ‘Beni-Madonna’ is often limited by suboptimal soil conditions. Root symbiotic fungi, including arbuscular mycorrhizal fungi (AMF) and the endophyte Serendipita indica, represent promising tools for sustainable crop improvement. However, the species-specific mechanisms by which these fungi regulate key physiological processes like photosynthesis and sugar metabolism remain unclear, limiting their targeted application. This study investigated the differential effects of two AMF species (Funneliformis mosseae and Diversispora versiformis) and Serendipita indica on root colonization, plant growth, leaf gas exchange, chlorophyll and nitrogen balance indices, iron (Fe) and magnesium (Mg) concentrations, soluble sugar levels, and the expression of genes related to sugar metabolism and Fe/Mg transport in ‘Beni-Madonna’ grafted onto trifoliate orange rootstock. All three fungi successfully colonized the roots, with S. indica showing the highest colonization rate. Fungal inoculation significantly enhanced leaf photosynthetic performance, chlorophyll index, Fe and Mg concentrations, and soluble sugar accumulation compared to non-inoculated controls. Among the treatments, F. mosseae most comprehensively induced the upregulation of Fe (CsFRO1 and CsIRT1) and Mg (CsMGT2 and CsMGT6) transporter genes, which was associated with improved chlorophyll index and photosynthetic demand, thereby exhibiting the best performance in enhancing net photosynthetic rate and other photosynthetic efficiency parameters. In contrast, S. indica was most effective in promoting the accumulation of sucrose, glucose, and fructose in leaves. Gene expression analysis revealed a common downregulation of sucrose synthesis genes (CsSPS1/4) across all treatments. The fungi differentially regulated sucrose hydrolysis and transport: D. versiformis and S. indica strongly upregulated invertase genes (CsVINV1 and CsA/NINV4), while both AMF species upregulated the sugar transporter gene CsSWEET. S. indica conferred the greatest promotion of plant height and stem diameter, indicating that its carbon demand strategy effectively channels carbon toward vegetative growth. In conclusion, all three symbiotic fungi enhanced carbon assimilation and accumulation in ‘Beni-Madonna’ leaves, with F. mosseae primarily optimizing the photosynthetic machinery via improved nutrient homeostasis, whereas D. versiformis and S. indica prominently activated sucrose hydrolytic pathways. These findings provide a basis for species-specific application of symbiotic fungi in citrus cultivation.