Phosphate Solubilisation, Organic Acid Production, and Nitrogen Fixation Potential of Free-Living Diazotroph (Azotobacter vinelandii FB2)

Abstract

Phosphorus and nitrogen are two major macronutrients that commonly limit crop productivity in agricultural soils. The excessive use of chemical fertilizers to overcome these limitations has resulted in environmental degradation and increased production costs, highlighting the need for sustainable alternatives. Microorganisms capable of simultaneously solubilising insoluble phosphates and fixing atmospheric nitrogen offer a promising eco-friendly solution. In the present study, Azotobacter vinelandii FB2, isolated from agricultural rhizosphere soil of Maharashtra, India, was evaluated for its phosphate solubilisation, organic acid production, and nitrogen fixation potential. The isolate was identified based on morphological and biochemical characteristics and confirmed by 16S rRNA gene sequencing. Qualitative screening on Pikovskaya’s agar medium revealed prominent halo zone formation with a mean solubilisation index of 3.40 ± 0.01. Quantitative estimation in Pikovskaya’s broth showed a significant and progressive increase in soluble phosphorus, reaching a maximum of 285.2 ± 2.1 µg mL⁻¹ on the 7th day of incubation (p ≤ 0.05), accompanied by a corresponding decline in pH from 7.0 to 4.3. High-performance liquid chromatography analysis confirmed gluconic acid as the predominant organic acid, followed by citric and oxalic acids, indicating an acidification-mediated phosphate solubilisation mechanism. Nitrogen fixation activity assessed using the acetylene reduction assay recorded a nitrogenase activity of 150.5 ± 1.3 nmol C₂H₄ mL⁻¹ h⁻¹, confirming the diazotrophic competence of the strain. The results demonstrate that A. vinelandii FB2 exhibits efficient dual functionality in mobilising phosphorus and fixing nitrogen, highlighting its strong potential as a multifunctional biofertiliser for sustainable agricultural applications.