Phenotypic high-content screening identifies plant-derived extracts targeting microbial growth, biofilms and swarming activity

The rise of antimicrobial resistance highlights the urgent need for new antibacterial agents, particularly from natural sources. In this study, 200 aqueous plant extracts were screened for antimicrobial activity against six clinically relevant pathogens. The test organisms included: Escherichia coli ATCC35401, Pseudomonas aeruginosa DSM19882, Salmonella enterica subsp. enterica ser. Typhimurium ATCC14028, Staphylococcus aureus NCCB32030, Streptococcus mutans DSM20523 and Enterococcus faecalis DSM2570. Three probiotic lactobacilli ( Lacticaseibacillus paracasei , Lactiplantibacillus plantarum and Lacticaseibacillus rhamnosus ) were used to differentiate pathogen-specific activity from broad-spectrum inhibition. Antimicrobial activities were initially assessed using the agar disk diffusion method, followed by broth microdilution to determine minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) for active plant extracts. Additionally, the effects on biofilm formation by E. coli , S. Typhimurium, S. aureus and S. mutans , and on swarming motility of P. aeruginosa and S. Typhimurium, were evaluated. 21 extracts inhibited at least one pathogen in the disk diffusion assay. Defined MIC values were detected for 19 extracts, while only 5 demonstrated bactericidal activity. In addition to the known antibacterial plants Allium sativum and Allium ursinum , previously unreported leaf extracts from Castanea sativa , Ribes × nidigrolaria and Rubus laciniatus showed promising activity against the tested pathogens. These extracts not only inhibited bacterial growth but also reduced biofilm formation and swarming motility of some pathogenic species at subinhibitory concentrations. Notably, no inhibition of Lactobacillus growth was detected in the disk diffusion assay. Overall, our findings highlight the potential of aqueous plant extracts as sources of antimicrobial and anti-virulence activities and demonstrate the importance of pathogen-specific effects in the discovery of plant-derived antimicrobials.