Fabrication and antimicrobial properties of novel meropenem-honey encapsulated chitosan nanoparticles against multiresistant and biofilm-forming Staphylococcus aureus as a new antimicrobial agent

Abstract

Background: Honey exhibits a broad spectrum of antibacterial activity against Gram-positive and Gram-negative bacteria, including methicillin-resistant Staphylococcus aureus (MRSA) ones. Chitosan (Cs) is a mucoadhesive polymer that also has antibacterial properties. Special attention has been paid to the design of polymeric nanoparticles (NPs) as new nano drug delivery systems to overcome bacterial resistance and its problems.

Objectives: The aim of the present study is to synthesize Cs-meropenem NPs with/without honey as an antibiofilm and antibacterial agent to inhibit Staphylococcus aureus.

Methods: This study synthesized meropenem and honey-loaded Cs nanogels and subsequently characterized them by Field Emission Scanning Electron Microscopy (FESEM), Fourier Transform Infrared Spectroscopy (FTIR), and DLS-zeta potential. Using the broth microdilution and crystal violet assays, the antibacterial and antibiofilm activity of meropenem and honey-loaded Cs nanogel, free meropenem, free honey, and free Cs NPs were investigated in vitro against MRSA strains. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) was also used to test the cytotoxicity of several Cs-NPs compound against the HEK-293 regular cell line.

Results: The average size of meropenem and honey-Cs-NPs was reported to be 119.885 nm, and encapsulation efficiency was 88.33 ± 0.97 with stability up to 60 days at 4°C. The NPs showed enhanced antibiofilm efficacy against S. aureus at sub-minimum inhibitory concentrations. Additionally, the cytotoxicity of meropenem and honey-encapsulated Cs against the HEK-293 normal cell line was insignificant.

Conclusions: Our findings suggested that meropenem and honey-Cs-NPs might be potential antibacterial and antibiofilm materials.

Keywords: chitosan; drug delivery; honey; multidrug resistance; nanobiotechnology.

FIGURE 1

Field‐emission scanning electron microscopes of the prepared meropenem and honey chitosan‐nanoparticles (Cs‐NPs). The morphological features and surface appearance of Cs‐NPs were revealed by ESEM pictures. Meropenem‐honey Cs‐NPs have an almost spherical form, a smooth surface, and a size range of 119.885 nanometers. FSEM image of meropenem (A), FSEM image of meropenem and honey (B), FSEM image of Cs containing honey (C), FSEM image of Cs containing meropenem (D), and FSEM image of Cs encapsulated with meropenem and honey (E and F). ESEM, Environmental Scanning Electron Microscopy.

FIGURE 2

Rates of drug release, and loading efficiency of different combinations of nanoparticles (NPs). (A) Data are shown as mean standard deviation (SD) for the in vitro release rate of meropenem and honey from chitosan (Cs)‐NPs at 37°C and pH 7.4; (B) effectiveness of C1–C6 combination of meropenem, meropenem + Cs, honey + Cs, and meropenem + honey Cs‐NPs. ***p < 0.001, **p < 0.01, *p < 0.05. ns, not significant. EE, encapsulation efficiency.

FIGURE 3

The micro‐pattern biofilm formation of MRSA and MDR strains‐isolated pathogens, when compared to controls (ATCC). The results are mentioned as the average of the OD value ± standard deviation (SD) from three independent experiments. ATCC, American Type Culture Collection; OD, Optical Density.

FIGURE 4

Anti‐biofilm properties of various chitosan‐nanoparticles (Cs‐NPs). Biofilm growth suppression of different combinations Cs‐encapsulated. Data are mean ± SD. ***p < 0.000, ***p < 0.001, **p < 0.01, *p < 0.05. ns, not significant; OD, Optical Density.

FIGURE 5

In response to C1–C6 chitosan formulations, mRNA expression of three important genes involved in biofilm formation (i.e., ompA, pgaB, and Bap) in all four isolates. Before (before the treatment of selected clinical strains with formulations C1 to C6) and after (after the treatment of selected clinical strains with formulations C1 to C6). Data are represented as mean ± SD. ***p < 0.000, ***p < 0.001, **p < 0.01, *p < 0.05. ns, not significant.

FIGURE 6

Over the course of 24 h, cell viability percentages of HEK 293 cells treated with various combinations of chitosan (Cs)‐encapsulated meropenem and honey were compared to free meropenem. The results reflect the mean and standard deviation of three separate experiments. ***p < 0.001, **p < 0.01, *p < 0.0.5 were used as the significant levels.