Inhibition of Acinetobacter baumannii Biofilm Formation Using Different Treatments of Silica Nanoparticles.

Abstract

There exists a multitude of pathogens that pose a threat to human and public healthcare, collectively referred to as ESKAPE pathogens. These pathogens are capable of producing biofilm, which proves to be quite resistant to elimination. Strains of A. baumannii, identified by the "A" in the acronym ESKAPE, exhibit significant resistance to amoxicillin in vivo due to their ability to form biofilm. This study aims to inhibit bacterial biofilm formation, evaluate novel silica nanoparticles' effectiveness in inhibiting biofilm, and compare their effectiveness. Amoxicillin was utilized as a positive control, with a concentration exceeding twice that when combined with silica NPs. Treatments included pure silica NPs, silica NPs modified with copper oxide (CuO.SiO2), sodium hydroxide (NaOH.SiO2), and phosphoric acid (H3PO4.SiO2). The characterization of NPs was conducted using scanning electron microscopy (SEM), while safety testing against normal fibroblast cells was employed by MTT assay. The microtiter plate biofilm formation assay was utilized to construct biofilm, with evaluations conducted using three broth media types: brain heart infusion (BHI) with 2% glucose and 2% sucrose, Loria broth (LB) with and without glucose and sucrose, and Dulbecco's modified eagle medium/nutrient (DMEN/M). Concentrations ranging from 1.0 mg/mL to 0.06 µg/mL were tested using a microdilution assay. Results from SEM showed that pure silica NPs were mesoporous, but in the amorphous shape of the CuO and NaOH treatments, these pores were disrupted, while H3PO4 was composed of sheets. Silica NPs were able to target Acinetobacter biofilms without harming normal cells, with viability rates ranging from 61-73%. The best biofilm formation was achieved using a BHI medium with sugar supplementation, with an absorbance value of 0.35. Biofilms treated with 5.0 mg/mL of amoxicillin as a positive control alongside 1.0 mg/mL of each of the four silica treatments in isolation, resulting in the inhibition of absorbance values of 0.04, 0.13, 0.07, 0.09, and 0.08, for SiO2, CuO.SiO2, NaOH.SiO2 and H3PO4.SiO2, respectively. When amoxicillin was combined, inhibition increased from 0.3 to 0.04; NaOH with amoxicillin resulted in the lowest minimum biofilm inhibitory concentration (MBIC), 0.25 µg/mL, compared to all treatments and amoxicillin, whereas pure silica and composite had the highest MBIC, even when combined with amoxicillin, compared to all treatments, but performed better than that of the amoxicillin alone which gave the MBIC at 625 µg/mL. The absorbance values of MBIC of each treatment showed no significant differences in relation to amoxicillin absorbance value and relation to each other. Our study showed that smaller amoxicillin doses combined with the novel silica nanoparticles may reduce toxic side effects and inhibit biofilm formation, making them viable alternatives to high-concentration dosages. Further investigation is needed to evaluate in vivo activity.