Ethanolic Chromolaena odorata (Siam weed) leaf extract exhibits broad-spectrum antimicrobial, antibiofilm, antioxidant, and cell-disruptive activities against clinically relevant bacteria

Background and Aim: The rapid rise of antimicrobial resistance threatens effective infection control and reinforces the need for alternative therapeutics. Chromolaena odorata (Siam weed), a traditionally used medicinal plant rich in phenolic and flavonoid compounds, has been reported to possess antimicrobial properties. This study evaluated the antimicrobial, antibiofilm, antioxidant, and ultrastructural effects of ethanolic C. odorata leaf extract against a diverse panel of Gram-positive and Gram-negative bacteria. Materials and Methods: Ethanolic crude extract was prepared from dried C. odorata leaves, and its antimicrobial activity was assessed against 46 bacterial isolates using disk diffusion, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) assays. Total phenolic and flavonoid contents were quantified using Folin–Ciocalteu and aluminum chloride methods. Antioxidant activity was measured using the 2,2-diphenyl picrylhydrazyl (DPPH) radical scavenging assay. Antibiofilm efficacy against Bacillus cereus was determined using crystal violet staining at sub-MIC levels. Ultrastructural alterations in B. cereus were examined via scanning electron microscopy (SEM). Results: The crude extract inhibited 78.26% (36/46) of tested isolates, with strong activity against nine species, including B. cereus, Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus luteus, Aeromonas hydrophila, Stenotrophomonas maltophilia, Citrobacter freundii, and Shigella sonnei. MIC values ranged from 31.25–125 mg/mL, with B. cereus showing the lowest MIC and MBC (31.25 mg/mL). The extract exhibited high phenolic (96.82 ± 2.07 μg Gallat-equivalents/mg) and flavonoid (62.98 ± 2.64 μg Quercetin equivalent /mg) content, and moderate antioxidant activity (IC₅₀ = 120.02 ± 16.31 μg/mL). Sub-MIC concentrations significantly inhibited B. cereus biofilm formation in a dose- and time-dependent manner, achieving up to 66.16% inhibition at 1/2 MIC after 72 h (p < 0.001). SEM analysis revealed cell shrinkage, wall collapse, and surface roughening in treated B. cereus, indicating disrupted cell integrity. Conclusion: Ethanolic C. odorata extract demonstrates broad-spectrum antibacterial, antibiofilm, antioxidant, and cell-disruptive activities, with pronounced effects against B. cereus. These findings highlight its potential as a natural antimicrobial or disinfectant candidate and support future development of plant-based agents to mitigate resistant bacterial infections. Keywords: antimicrobial activity, ethanolic Chromolaena odorata extract, biofilm inhibition, antioxidant activity, phytochemical profiling, Bacillus cereus, scanning electron microscopy, natural antibacterial agents.