Cover cropping enhances fruit quality in protected citrus cultivation by modulating rhizosphere microbiome and iron availability

Introduction Citrus is one of the most widely cultivated fruit trees worldwide, and protected cultivation has become increasingly prevalent in recent years. Cover cropping improves orchard soil health, yet its mechanisms in protected citrus cultivation remain unclear. This study investigated how white clover ( Trifolium repens L.) and ryegrass ( Lolium perenne L.) affect soil properties, rhizosphere microbiota, and fruit quality in greenhouse-grown ‘Kanpei’ citrus through integrated analyses of soil physicochemical properties, high-throughput amplicon sequencing, and microbial isolation. Results Both cover crops significantly increased total soluble solids (TSS) and vitamin C levels in mature fruits. Ryegrass enhanced the availability of nitrogen, phosphorus, calcium, magnesium, and manganese, whereas white clover more effectively acidified the soil and increased iron (Fe) availability. Each cover crop distinctively altered the rhizosphere microbial community. Notably, white clover specifically enriched Pseudomonas, which strongly correlated with elevated soil available Fe, TSS, and vitamin C. Screening with Chrome Azurol S (CAS) agar identified Pseudomonas as the dominant siderophore-producing genus. Inoculation with a representative strain, Pseudomonas sp. PA9, significantly enhanced Fe uptake, chlorophyll content, and fruit quality, offering insights into its potential role in promoting fruit quality under protected cultivation. Conclusions This work provides a comprehensive understanding of how white clover promotes fruit quality via fostering siderophore-producing Pseudomonas that enhance Fe mobilization, suggesting new avenues for developing microbiome-based management strategies in protected citrus cultivation. These findings underscore the potential of cover crop-mediated microbial recruitment in advancing sustainable citrus production and soil health improvement.