Antibacterial Effect of Silver Nanoparticles along with L-Arginine against P. aeruginosa

Abstract

Background and Objectives: Nowadays, the prevalence of multidrug-resistant pathogens such as Pseudomonas aeruginosa is increasing worldwide. Many studies have been seeking new treatment strategies to treat infections caused by these microorganisms. Silver nanoparticles (AgNPs) along with L-arginine have significant antimicrobial effects and could be used as alternatives for ineffective drugs. Methods: In this study, the antibacterial activity of AgNPs, L-arginine and various concentrations of AgNPs along with L-arginine (12.5 and 25 mg/ml) were investigated against P. aeruginosa PAO1 using the broth macrodilution method. Results: Minimum inhibitory concentration of AgNPs, L-arginine and AgNPs combined with 25 and 12.5 mg/ml L-arginine was 15.6 μg/ml, 25 mg/ml, 1.9 μg/ml and 3.9 μg/ml, respectively. Minimum bactericidal concentration of AgNPs, L-arginine and AgNPs combined with 25 and 12.5 mg/ml L-arginine was 31.2 μg/ml, 50 mg/ml, 3.9 μg/ml and 7.8 μg/ml, respectively. Conclusion: Our study suggests that AgNPs along with L-arginine can be used as an alternative antibacterial agent against P. aeruginosa, and might be useful for treatment of wound infections. L-arginine against P. aeruginosa in an aqueous system. MATERIAL AND METHODS Subsequent dilutions (0.2 to 500 μg/ml) of colloidal AgNPs (average size 20 nm, Pishgaman Nano Arya Co.) were prepared in Muller Hinton broth from stock solution. Stock solution of L-arginine (Merck, Germany) was prepared in distilled water with final concentration of 200 mg/ml, and later used to prepare subsequent dilutions (0.1 to 100 mg/ml) in Muller Hinton broth, using serial twofold dilution. Subsequent dilutions of AgNPs (0.2-500 μg/ml) were prepared in Muller Hinton broth. Two concentrations of L-arginine (12.5 and 25 mg/ml) were separately added to each tube. P. aeroginosa PAO1 was obtained from Urmia University of Medical Sciences. The bacteria were grown in Muller Hinton broth at 37 o C for 24 h. Bacterial suspension equivalent to 0.5 McFarland standard (1.5 × 10 8 CFU/ml) was prepared in sterile normal saline (26). Determination of minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) The MIC of AgNPs, L-arginine and AgNPs with L-arginine against P. aeroginosa was determined by broth macrodilution method according to the Clinical and Laboratory Standards Institute (CLSI, 2014). Determination of MIC and MBC of AgNPs and L-arginine Different concentrations of (0.2-500 μg/ml) of AgNPs were prepared in Muller Hinton broth using two fold serial dilutions. Bacterial suspension containing 1.5 × 10 8 CFU/ml was added to each tube, and the tubes were incubated at 37 o C for 24h. The MIC was defined as the lowest concentration of AgNPs that was able to inhibit bacterial growth. The dilution method was also used to determine MBC values. For this propose, 100 μl of bacterial suspension containing ≥MIC concentrations of AgNPs were cultured on Muller Hinton agar plates. After incubation at 37 o C for 24 h, the MBC value was defined as the lowest concentration of AgNPs that was able to kill 99.9% of bacteria. All experiments were performed in triplicate, on three different days (27). The MIC and MBC values for L-arginine were determined using the method