In vitro antimicrobial activity of silver nanoparticles against selected Gram-negative and Gram-positive pathogens.

Background and aim: Infections caused by pathogenic bacteria increase patient morbidity and mortality and significantly raise treatment costs. The use of silver nanoparticles as an alternative treatment for S aureus, E coli, MRSA, E faecalis, K pneumoniae and P aeruginosa indicates their antibacterial effect and prompts medical research to consider the next generation of antibacterial drugs that could change antibiotic therapy. By combining silver nanoparticles with different classes of antibiotics, the antibacterial effect is evidenced by increased values of the inhibition zone compared to the values obtained for some antibiotics commonly used in the treatment of bacterial infections. This study focuses on comparing the antibacterial activity of antibiotics versus antibiotics combined with silver nanoparticles against various bacteria, by comparing inhibition zones obtained for both. We aim to prove that the size of the inhibition zone for antibiotics combined with silver nanoparticles is greater, thus confirming the improved antibacterial effect.

Metods: In this study we tested the antibacterial activity of solutions of silver nanoparticles alone or in combination with different antibiotics. We used standard bacterial strains, ATCC, both Gram positive bacteria Staphylococcus aureus ATCC 29213, Enterococcus faecalis ATCC 29212, as well as Gram negative bacteria Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, but also on clinical isolates: a strain MRSA (Methicillin Resistant Staphylococcus aureus) and a PDR strain (pan drug resistant) of Klebsiella pneumoniae. Bacterial identification was performed using Vitek MS analyzer (bioMerieux). Antibiotic susceptibility determination was performed with VITEK2 COMPACT SYSTEM (bio Merieux, Inc Durham NC) with ready to use VITEK AST cards. The interpretation of the results was done in compliance with EUCAST 2023-2024 standards. Testing was performed for several classes of antibiotics, silver nanoparticle solutions in 2 concentrations (10 μg/mL and 100 μg/mL) and for combinations of antibiotics with silver nanoparticle solutions. The diameter of the inhibition zone (ZOI) for silver nanoparticles, antibiotics and silver nanoparticles combined with antibiotic against each bacterium was expressed in millimeters. The Kirby-Bauer disk-diffusion method, in accordance with current EUCAST standards, was used to analyze the antibacterial effect of antibiotics, silver nanoparticles, and antibiotics combined with silver nanoparticles at biocompatible doses of 10 and 100 μg/mL. The experiments were conducted in triplicate, and the results were almost identical.

Results: The results of this study show that the silver nanoparticles displayed antibacterial activity, proven by the appearance of the inhibition zone, in various sizes, for all bacteria studied. The antibiotic classes tested were beta-lactamins, first, second, third and fourth generation cephalosporins, macrolides, fluoroquinolones, lincosamides, aminoglycosides, glycopeptides, tetracyclines, oxazolidinones, sulfonamides, rifamycins, amphenicols. Testing S aureus ATCC 29213, the highest zone of inhibition was demonstrated for cephalosporins (32.6667 ± 0.701 mm), macrolides (31.6667 ± 0.701 mm, and lincosamides (29.6667 ± 0.701 mm). Testing MRSA (internal code GR0333), the highest zone of inhibition for combination of silver nanoparticles and antibiotics was demonstrated for fluoroquinolones (36.3333 ± 0.701 mm), lincosamides (32.3333 ± 0.701 mm), Fusid acid (32.3333 ± 0.701 mm) and aminoglicosides (31.3333 ± 0.701 mm). Testing E coli ATCC 25922 the highest zone of inhibition was for Fosfomycine, 39 mm and for E faecalis ATCC 29212 for aminoglicosides was 19 mm. For K pneumoniae (internal code GQ8575) the inhibition zone for silver nanoparticles 100 μg/mL was 12.3333 ± 0.701 mm and for P aeruginosa ATCC 27253 was 16 ± 1.214 mm.

Conclusions: The use of metallic nanoparticles, especially silver ones, as antimicrobial agents with definite bactericidal activity has led medical specialists to consider this new treatment which may change antibacterial therapy. Studies of in vitro combinations between silver nanoparticles and different classes of antibiotics represent a highly efficient and effective new antibacterial treatment against multidrug-resistant bacteria. To avoid the problem of antimicrobial resistance associated with conventional antibiotics, it is necessary to understand the adaptive mechanisms of bacteria under the action of metal nanoparticles, which could be exploited in future studies. Further in vitro and in vivo studies that would assess specify the biocompatibility and toxicity of silver nanoparticles will make these super nanomaterials the medicines of the future.