Abstract

The present study focused on the identification and characterization of free-living nitrogen-fixing bacteria isolated from the rhizosphere of Balanites aegyptiaca and Adansonia digitata trees as potential biofertilizer candidates. Nitrogen-fixing bacterial strains were isolated using Ashby’s mannitol agar medium and characterized through morphological, biochemical and molecular analyses, including amplification and sequencing of the 16S rRNA gene. A total of thirty Composite soil samples were collected, yielding seven isolates on Ashby’s medium. Morphological and biochemical evaluations identified two promising free-living nitrogen-fixing strains, one from each tree species, with potential as biofertilizer candidates. The selected strains exhibited circular morphology, milky-white pigmentation, smooth margins, motile rod-shaped structures and Gram-negative characteristics. Biochemical tests revealed positive results for catalase, urease, methyl red and nitrate reduction. Molecular characterization further identified the isolates as Paenibacillus timonensis and Enterobacter hormaechei. Experimental inoculation of millet plants with the isolates resulted in significant nutrient enhancements, with increases of 114.86% nitrogen, 165% phosphorus and 73.58% potassium in the millet shoots compared to the untreated control. Nitrogen uptake by millet was elevated by 208%, 108% and 420% for the experimental isolates and NPK fertilizer treatment respectively, relative to the negative control. Similarly, phosphorus uptake showed an increase of 257%, 100% and 171% for the isolates and NPK treatment, respectively. These findings suggest that the rhizospheres of trees found in the semi-arid and arid regions of Nigeria harbor nitrogen-fixing bacteria with considerable potential for biofertilizer development, thereby supporting sustainable agricultural practices.