Atividade antibacteriana e antibiofilme de um complexo de rutênio polipiridínico e seu efeito individual e combinado a antibióticos sobre bactérias de interesse clínico

Abstract

Healthcare-associated infections (HAIs) are one of the main sources of preventable diseases in hospitalized patients, and cause significant damage to healthcare resources. They are often caused by bacteria resistant to several antibiotics such as methicillin-resistant Staphylococcus aureus – MRSA and long periods of hospitalization. The objective of this study was to evaluate the antibacterial and antibiofilm activity of a polypyridine ruthenium complex designated ruthenium(II) tris(4-methyl-4-carboxamide(2-anthracenyl)-2,2-bipyridine) [Ru(Ant)]32+ on bacterial strains S. aureus (ATCC 25923 and ATCC 700698 MRSA), and S. epidermidis (ATCC 12228 and ATCC 35984 MRSA) of clinical interest. To determine antibacterial activity, compounds were diluted to concentrations ranging from 3.9 to 250 μg/mL. The ruthenium complex [Ru(Ant)]32+ was evaluated for minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) in relation to S. aureus ATCC 25923, S. aureus ATCC 700698, S. epidermidis ATCC 12228 and S. epidermidis ATCC 35984, through microdilution tests. The determined MICs were combined with the antibiotics ampicillin and tetracycline, and their effects were evaluated using the checkerboard technique. Antibiofilm activity was evaluated by quantifying biomass using crystal violet staining (CV) and the number of viable biofilm cells using colony forming units (CFU) and measuring metabolic activity using the XTT reduction assay. The effect of the treatment was also analyzed using scanning electron microscopy – SEM and confocal laser microscopy using the Live/Dead cell viability kit. In addition, the cytotoxicity of the complex was evaluated in in vitro tests on murine fibroblast lines. The results obtained showed an MIC value of 31.25 μg/mL for the strains S. aureus ATCC 700698 and S. epidermidis ATCC 35984 and an MIC of 125 μg/mL for S. aureus ATCC 25923 and S. epidermidis ATCC 12228 (MRSE). The combination of the complex with the antibiotics ampicillin and tetracycline showed a synergistic and additive effect, depending on the bacterial strain tested. The complex was also able to reduce the formation of biofilms by between 10 and 99%, with a reduction in the number of viable biofilm cells by up to 99.99% and complete inhibition of metabolism in some strains at concentrations ranging from 3.9 to 250.0 μg/mL. In relation to pre-formed biofilms, a reduction of biofilms of up to 80% was observed, and of viable cells by up to 90%, being capable of inhibiting metabolism by up to 80% at a concentration of 250 μg/mL. In relation to SEM, ruptures in the cell wall were observed in all strains treated with CIM. In confocal laser microscopy, the results on S. aureus ATCC 700698 and S. epidermidis ATCC 35984 treated with CIM demonstrated a reduction in cells and biofilm. Regarding cytotoxicity against the L929 strain, cytotoxicity above 125 μg/mL was observed within 24 hours. The results showed that [Ru(Ant)]32+ has potential as an antimicrobial agent or can even assist in the synthesis of new antibacterial agents of clinical interest for the pharmaceutical industry and research institutions.