Abstract
<jats:p>Fifty-four soil samples were collected from various sites (agricultural, oil-polluted, riverbank, and animal dung-contaminated soils) in northern Basrah, southern Iraq, between September 18 and November 28, 2022. The specimens were transported to the laboratory within 3 hours, serially diluted, and cultivated on selective media. The screening indicated that 28 samples (51.9 %) exhibited positive bacterial growth. Phenotypic and molecular identification via PCR amplification of the 16S rRNA gene (~1400 bp) revealed the presence of Pseudomonas aeruginosa (8 isolates), Escherichia coli (7 isolates), Klebsiella pneumoniae (4 isolates), Staphylococcus aureus (4 isolates), Bacillus cereus (3 isolates), and Bacillus subtilis (2 isolates). Among these, a dominant P. aeruginosa isolate was successfully utilized for the green biosynthesis of iron nanoparticles (FeNPs). The successful formation of biogenic FeNPs was initially indicated by a rapid color transition from yellow to dark brown. UV-Vis spectroscopy confirmed nanoparticle synthesis with a sharp surface plasmon resonance peak at 304 nm, while FT-IR spectroscopy verified the definitive metal-oxygen (Fe–O) stretching bond at 589.97 cm⁻¹ along with capping bacterial functional groups. Scanning electron microscopy (SEM) demonstrated a predominantly spherical nanoparticle morphology with a well-defined representative particle size of 22.83 nm. The quantitative microtiter plate and Congo red agar assays confirmed that the pathogenic isolates possessed a robust baseline capacity for biofilm formation. The biosynthesized FeNPs exhibited exceptional, concentration-dependent antibiofilm efficacy, achieving characteristically high biofilm inhibition rates of up to 91.64 % at concentrations of 50 and 100 µg/mL. Furthermore, the nanoparticles displayed potent antibacterial activity via the agar well diffusion method, with inhibition zones expanding proportionally with concentration and reaching a maximum diameter of 21.0 mm against P. aeruginosa. These collective findings suggest that biosynthesized iron nanoparticles could serve as highly effective, biocompatible, and environmentally friendly nano-antimicrobials for mitigating multidrug-resistant pathogens and persistent biofilm-associated infections.</jats:p>