Antimicrobial and antibiotic resistance is a major threat to mankind due to increasing resistance towards existing medicinal drugs. Plant-based antimicrobials are promising alternatives to conventional drugs. Cress (Lepidium sativum L.) is known as garden cress, garden cress pepper weed or garden pepperwort and is a member of the Brassicaceae family. Lepidium satvium L. (LS) is widely spread throughout the world as a fast-growing annual herb. LS seed oil has antimicrobial, antioxidant, and anti-inflammatory activities. Silver nanoparticles (AgNP) gained attention due to their antimicrobial properties. We aimed to synthesize LS encapsulated AgNP to enhance the microbicidal activities of the nanoparticles and prevent microbial resistance by plant-based synergistic mechanisms. AgNP were prepared in a one-pot synthesis by plant-biomolecules-induced reduction of silver nitrate via green method. The biomolecules and metabolites in the aqueous LS extract act as reducing, capping and stabilizing agents of AgNP. Fourier transform infrared spectroscopy (FT-IR), ultraviolet-visible spectroscopy (UV-Vis) and dynamic light scattering (DLS) confirmed the composition of the LS-AgNP biohybrids. Antimicrobial testing by disc dilution method against a total of 10 reference strains of microorganisms verified excellent to intermediate antimicrobial activity. The Gram-negative pathogens E. coli WDCM 00013 and P. aeruginosa WDCM 00026 were highly inhibited, followed by intermediate results for the Gram-positive bacteria B. subtilis WDCM 00003, S. pneumoniae ATCC 49619, S. aureus ATCC 25923, S. pyogenes ATCC 19615, E. faecalis ATCC 29212, and the fungus C. albicans WDCM 00054. Our biohybrid LS-AgNP showed increased antimicrobial activity with potential uses as disinfectant and wound care product.