Antibacterial activity of plant-mediated synthesized CuS nanoparticles against gram-positive Bacillus subtilis and gram negative Pseudomonas putida is studied in this presented work. A facile hydrothermal approach was followed to synthesize spherical shaped CuS nanoparticles using flowers extract of Tagetes patula (S1), leaves extract of Azadirachta indica (S2), and bark extract of Terminalia arjuna (S3). Crystal phase identification, average crystallites size, surface morphology, absorbance spectra and energy band gap analyses of samples were further determined using XRD, SEM, and UV-DRS spectroscopy respectively. Furthermore, under irradiation of visible light, antibacterial activity of marigold flowers mediated (S1), neem leaves mediated (S2), and arjuna bark mediated (S3) CuS samples were studied. XRD data confirmed the marigold mediated CuS sample has relatively smaller crystallites size of 10.67 nm. The estimated band gap energies for respective S1, S2, and S3 samples are 174 eV, 175 eV, and 177 eV. The antibacterial analysis of S1 sample displays its excellent antibacterial activity with the formation of relatively large inhibition zones of 29 mm and 30 mm diameters, against Bacillus subtilis and Pseudomonas putida respectively. The demonstrated antibacterial activity of biocompatible CuS nanoparticles suggests their potential use in biomedical applications.