Synthesis, characterization and evaluation of antimicrobial and cytotoxic activities of biogenic silver nanoparticles synthesized from Streptomyces xinghaiensis OF1 strain

Abstract

We report synthesis of silver nanoparticles (AgNPs) from Streptomyces xinghaiensis OF1 strain, which were characterised by UV-Vis and Fourier transform infrared spectroscopy, Zeta sizer, Nano tracking analyser, and Transmission electron microscopy. The antimicrobial activity of AgNPs alone, and in combination with antibiotics was evaluated against bacteria, namely Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis, and yeasts viz., Candida albicans and Malassezia furfur by using micro-dilution method. The minimum inhibitory concentration (MIC) and minimum biocidal concentration of AgNPs against bacterial and yeast strains were determined. Synergistic effect of AgNPs in combination with antibacterial and antifungal antibiotics was determined by FIC index. In addition, MTT assay was performed to study cytotoxicity of AgNPs alone and in combination with antibiotics against mouse fibroblasts and HeLa cell line. Biogenic AgNPs were stable, spherical, small, polydispersed and capped with organic compounds. The variable antimicrobial activity of AgNPs was observed against tested bacteria and yeasts. The lowest MIC (16 µg ml^-1) of AgNPs was found against P. aeruginosa, followed by C. albicans and M. furfur (both 32 µg ml^-1), B. subtilis and E. coli (both 64 µg ml^-1), and then S. aureus and Klebsiella pneumoniae (256 µg ml^-1). The high synergistic effect of antibiotics in combination with AgNPs against tested strains was found. The in vitro cytotoxicity of AgNPs against mouse fibroblasts and cancer HeLa cell lines revealed a dose dependent potential. The IC₅₀ value of AgNPs was found in concentrations of 4 and 3.8 µg ml^-1, respectively. Combination of AgNPs and antibiotics significantly decreased concentrations of both antimicrobials used and retained their high antibacterial and antifungal activity. The synthesis of AgNPs using S. xinghaiensis OF1 strain is an eco-friendly, cheap and nontoxic method. The antimicrobial activity of AgNPs could result from their small size. Remarkable synergistic effect of antibiotics and AgNPs offer their valuable potential in nanomedicine for clinical application as a combined therapy in the future.

Keywords: Antibacterial activity; Antifungal activity; Antimicrobials; Biogenic silver nanoparticles; Cytotoxicity; Streptomycetes; Synergism.

Fig. 1

UV–Visible spectrum of silver nanoparticles synthesized from Streptomyces xinghaiensis OF1 strain. AgNO₃ (1), control (2), standard AgNPs (3), and experimental (4)

Fig. 2

Transmission electron micrograph of silver nanoparticles synthesized from Streptomyces xinghaiensis OF1 strain

Fig. 3

Nanotracking analysis of silver nanoparticles synthesized from Streptomyces xinghaiensis OF1 strain

Fig. 4

Fourier transform infrared spectroscopy analysis of AgNPs synthesized from Streptomyces xinghaiensis OF1 strain. Absorbance bands: 3432, 2925, 1631, 1385 and 1033 cm⁻¹

Fig. 5

SEM analysis of Candida albicans and Escherichia coli cells before and after treatment with silver nanoparticles. Upper panel is control and lower panel is microbial cells treated with AgNPs. C. albicans (a, b) and E. coli (c, d)

Fig. 6

Cytotoxic activity of AgNPs from Streptomyces xinghaiensis OF1 strain against mouse fibroblasts (3T3) and HeLa cell line

Fig. 7

Microscopic observations of mouse fibroblasts (A) and HeLa cells (B) after treatment with various concentrations of biosynthesized silver nanoparticles (a 1 µg ml⁻¹, b 5 µg ml⁻¹, c 10 µg ml⁻¹, d 20 µg ml⁻¹). Scale bar 100 µm

Fig. 8

Cytotoxic activity of combined antimicrobial agents (AgNPs and antibacterial antibiotics) against a mouse fibroblasts (3T3) and b HeLa cell line. Both antimicrobials were used at concentrations of their FIC (see Table 2): (1) (4 µg ml⁻¹ of AgNPs + 64 µg ml⁻¹ AMP or 32 µg ml⁻¹ K or 4 µg ml⁻¹ TE), (2) (16 µg ml⁻¹ of AgNPs + 64 µg ml⁻¹ AMP or 12 µg ml⁻¹ K or 2 µg ml⁻¹ TE), (3) (1 µg ml⁻¹ of AgNPs + 768 µg ml⁻¹ AMP or 128 µg ml⁻¹ K or 64 µg ml⁻¹ TE), (4) (16 µg ml⁻¹ of AgNPs + 0.06 µg ml⁻¹ AMP or 0.5 µg ml⁻¹ K or 0.25 µg ml⁻¹ TE), (5) (4 µg ml⁻¹ of AgNPs + 0.004 µg ml⁻¹ AMP or 0.125 µg ml⁻¹ K or 0.008 µg ml⁻¹ TE). AMP ampicillin, K kanamycin, TE tetracycline

Fig. 9

Cytotoxic activity of combined antimicrobial agents (AgNPs and antifungal antibiotics) against a mouse fibroblasts (3T3) and b HeLa cell line. Both antimicrobials were used at concentrations of their FIC (see Table 2): (1) (2 µg ml⁻¹ of AgNPs + 0.016 µg ml⁻¹ AMB or 64 µg ml⁻¹ FLU or 64 µg ml⁻¹ KCA), (2) (32 µg ml⁻¹ of AgNPs + 1024 µg ml⁻¹ AMB or 4 µg ml⁻¹ FLU, and 2 µg ml⁻¹ of AgNPs and 64 µg ml⁻¹ KCA). AMB amphotericin B, FLU fluconazole, KCA ketoconazole