Biofilm Formation by Rice Rhizosphere Nitrogen-Fixing Microorganisms and Its Effect on Rice Growth Promotion

Excessive nitrogen fertilizer use contributes to environmental pollution and undermines agricultural sustainability. Enhancing symbiotic interactions between rice and nitrogen-fixing microorganisms offers a promising strategy to potentially improve nitrogen use efficiency (NUE). This study investigates the role of rice root exudates in promoting biofilm formation by nitrogen-fixing microbes to enhance nitrogen fixation. Nine nitrogen-fixing microbial strains were evaluated for biofilm formation in response to flavone and apigenin treatments, with Gluconacetobacter diazotrophicus KACC 12358 serving as the reference strain. The most responsive strain was selected, and a library of 1597 natural compounds was screened to identify those that promote biofilm formation in both the selected and reference strains. A. indigens KACC 11682 exhibited the highest biofilm-forming capacity, with apigenin treatment showing an OD595 value approximately 1.4 times higher than the DMSO control. Screening identified 68 compounds that enhanced biofilm formation by more than 500% compared to the control. Among them, eight compounds induced strong biofilm formation (O.D. > 2.0) in A. indigens. Cardamomin, a chalconoid flavonoid, emerged as one of the most effective compounds, showing a 245% increase in biofilm formation. Growth promotion assays showed that A. indigens increased rice fresh weight by approximately 128% compared to untreated controls. This study demonstrates the potential of rice root exudate-derived compounds to promote beneficial symbiosis with nitrogen-fixing microbes. These findings offer a novel approach that may contribute to enhancing rice NUE. Future research will focus on evaluating the long-term effects of these compounds and microorganisms, assessing their applicability in real agricultural settings, and conducting further validation across various rice cultivars.