The present study was carried out to evaluate the larvicidal potential of methanol, hexane, acetone, chloroform, and aqueous bark extracts of Holarrhena antidysenterica (L.) Wall. and silver nanoparticles (AgNPs) synthesized using aqueous bark extract against the third instar larvae of Aedes aegypti L. and Culex quinquefasciatus Say. AgNPs were prepared by adding 10 ml of aqueous bark extract in 90 ml of 1 mM silver nitrate (AgNO) solution. After 5 min of mixing, a change in color from yellow to dark brown occurred indicating the synthesis of AgNPs. Their further characterization was done through ultraviolet-visible spectroscopy (UV-Vis), X-ray diffraction analysis (XRD), field emission scanning electron microscope (FE-SEM), transmission electron microscopy (TEM), and Fourier transform infrared spectroscopy (FT-IR). UV-Vis spectrum of synthesized AgNPs showed a maximum absorption peak at 420 nm wavelength. Crystalline nature of AgNPs was confirmed by the presence of characteristic Bragg reflection peaks in XRD pattern. TEM images have shown that most of the AgNPs were spherical in shape with an average size of 32 nm. FT-IR spectrum of AgNPs showed prominent absorbance peaks at 1012.2 (C-O) and 3439.44 cm (O-H) which represent the major constituents of phenolics, terpenoids, and flavonoids compounds. LC-MS analysis of the bark extract confirmed the presence of carbonyl and hydroxyl functional groups which were directly correlated with FT-IR results. These AgNPs were assayed against different mosquito vectors, and the maximum mortality was recorded against the larvae of A. aegypti with LC and LC values being 5.53 and 12.01 ppm, respectively. For C. quinquefasciatus, LC and LC values were 9.3 and 19.24 ppm, respectively, after 72 h of exposure. Bark extracts prepared in different solvents such as methanol, chloroform, hexane, acetone, and water showed moderate larvicidal activity against A. aegypti their respective LC values being 71.74, 94.25, 102.25, 618.82, and 353.65 ppm and LC values being 217.36, 222.24, 277.82, 1056.36, and 609.37 ppm. For C. quinquefasciatus, their LC values were 69.43, 112.39, 73.73, 597.74, and 334.75 ppm and LC values of 170.58, 299.76, 227.48, 1576.98, and 861.45 ppm, respectively, after 72 h of treatment. AgNPs proved to be nontoxic against the non-target aquatic organism, Mesocyclops thermocyclopoides Harada when exposed for 24, 48, and 72 h. The results showed that bark extract-derived AgNPs have extremely high larvicidal potential compared to other organic solvents as well as aqueous bark extract alone. These AgNPs, therefore, can be used safely for the control of dengue and filarial vectors that cause severe human health hazards.