Cuminaldehyde enhances the efficacy of ciprofloxacin and gentamicin against the biofilm of Escherichia coli: A Response Surface Methodology (RSM) approach

Uropathogenic Escherichia coli (UPEC) significantly complicates the management of urinary tract infections (UTIs) through its ability to form biofilms, which contributes to the emergence of antibiotic resistance and frequent therapeutic failures. Conventional monotherapy has often been proved ineffective against biofilm-embedded bacteria. To address this challenge, the present study evaluated the antibiofilm efficacy of a novel triple combination of cuminaldehyde (CA), gentamicin (GN), and ciprofloxacin (CF) to overcome biofilm-associated threats. Dose optimization of the triple-compound combination was performed using Response Surface Methodology (RSM) coupled with a Central Composite Design (CCD). The CCD generated 19 experimental runs for three-factor combination, which were evaluated against five (5) UPEC isolates. Biofilm formation was quantified using the crystal violet assay in 96-well microtiter plates. Analysis of variance (ANOVA) confirmed the accuracy and statistical significance of the quadratic model ( p < 0.05). Significant synergistic interactions among CA, GN, and CF were observed across all five UPEC strains strains. While individual compounds and double-compound combinations failed to achieve substantial biofilm inhibition, the optimized doses of triple compound combination markedly inhibited biofilm formation. These predicted outcomes were experimentally validated and further corroborated by light microscopic analysis. The findings demonstrated that the triple compound combination (CA-GN-CF) exhibits remarkable antibiofilm potential against UPEC, outperforming individual and dual-compound combination treatments. Moreover, this study emphasized on the utility of RSM as a robust, statistically driven tool for dose optimization, enabling enhanced antibiofilm efficacy. Such combination strategies hold promise for improving therapeutic outcomes in biofilm-linked multidrug-resistant UTIs.