Benefits of Usage of Immobilized Silver Nanoparticles as Pseudomonas aeruginosa Antibiofilm Factors

Abstract

The aim of this study was to assess the beneficial inhibitory effect of silver nanoparticles immobilized on SiO₂ or TiO₂ on biofilm formation by Pseudomonas aeruginosa-one of the most dangerous pathogens isolated from urine and bronchoalveolar lavage fluid of patients hospitalized in intensive care units. Pure and silver doped nanoparticles of SiO₂ and TiO₂ were prepared using a novel modified sol-gel method. Ten clinical strains of P. aeruginosa and the reference PAO1 strain were used. The minimal inhibitory concentration (MIC) was determined by the broth microdilution method. The minimal biofilm inhibitory concentration (MBIC) and biofilm formation were assessed by colorimetric assay. Bacterial enumeration was used to assess the viability of bacteria in the biofilm. Silver nanoparticles immobilized on the SiO₂ and TiO₂ indicated high antibacterial efficacy against P. aeruginosa planktonic and biofilm cultures. TiO₂/Ag⁰ showed a better bactericidal effect than SiO₂/Ag⁰. Our results indicate that the inorganic compounds (SiO₂, TiO₂) after nanotechnological modification may be successfully used as antibacterial agents against multidrug-resistant P. aeruginosa strains.

Keywords: MBIC; Pseudomonas aeruginosa; SiO2/Ag0; TiO2/Ag0; biofilm; multidrug-resistant bacteria; nanoparticles.

Figure 1

The various antibacterial targets of nanoparticles.

Figure 2

TEM images of amorphous SiO₂/Ag⁰ (A) and TiO₂/Ag⁰ (B).

Figure 3

Agarose gel electrophoresis of PCR products for presence of silE gene (220 bp) in P. aeruginosa strains. Lanes: 1 and 16 molecular size markers (1000 bp, Fermentas), 2—negative control, 3—positive control, E. coli J53, 4—P. aeruginosa 0013, 5—P. aeruginosa 0024, 6—P. aeruginosa 3, 7—P. aeruginosa 472, 8—P. aeruginosa 669, 9—P. aeruginosa 124, 10—P. aeruginosa 137, 11—P. aeruginosa 300, 12—P. aeruginosa 328, 13—P. aeruginosa 407, 15—P. aeruginosa PAO1.

Figure 4

Effect of SiO₂/Ag⁰ and TiO₂/Ag⁰ on biofilm formation by P. aeruginosa strains isolated from urine. Interpretation of the results: OD ≤ 0.096—non-biofilm producer; 0.096 < OD ≤ 0.192—weak-biofilm producer; 0.192 < OD ≤ 0.384—moderate-biofilm producer; OD > 0.384—strong-biofilm producer.

Figure 5

Effect of SiO₂/Ag⁰ and TiO₂/Ag⁰ on biofilm formation by P. aeruginosa strains isolated from bronchoalveolar lavage fluid. Interpretation of the results: OD ≤ 0.096—non-biofilm producer; 0.096 < OD ≤ 0.192—weak-biofilm producer; 0.192 < OD ≤ 0.384—moderate-biofilm producer; OD > 0.384—strong-biofilm producer.

Figure 6

Effect of SiO₂/Ag⁰ and TiO₂/Ag⁰ on the survival of P. aeruginosa strains isolated from urine in biofilm mass.

Figure 7

Effect of SiO₂/Ag⁰ and TiO₂/Ag⁰ on the survival of P. aeruginosa strains isolated from bronchoalveolar lavage fluid in biofilm mass.

Figure 8

Effect of SiO₂/Ag⁰ and TiO₂/Ag⁰ at a concentration of 0.5 × MBIC on biofilm formation by PAO1 and clinical strains of P. aeruginosa. Values represent the mean ± SD for five strains isolated from urine and five from bronchoalveolar lavage fluid. * p ≤ 0.05 compared with control. Interpretation of the results: OD ≤ 0.096—non-biofilm producer; 0.096 < OD ≤ 0.192—weak-biofilm producer; 0.192 < OD ≤ 0.384—moderate-biofilm producer; OD > 0.384—strong-biofilm producer.

Figure 9

Effect of SiO₂/Ag⁰ and TiO₂/Ag⁰ at a concentration of 0.5 × MBIC on survival PAO1 and clinical strains of P. aeruginosa in biofilms. Values represent the mean ±SD for five strains isolated from urine and five from bronchoalveolar lavage fluid. * p ≤ 0.05 compared with control.