Estudo da interação e efeito modulador da lectina das sementes de dioclea violacea e o antibiótico Gentamicina em bactérias multirresistentes.

Abstract

The World Health Organization ranked the bacterial resistance to antibiotics as an important public health problem in the 21st century, due to the damage caused both to human and animal health and economically, making it necessary to develop new strategies in order to solve this problem. To this end, in vitro studies reveal that a class of proteins with the ability to recognize and interact with specific carbohydrates, called lectins, when associated with antibiotics have been shown an effective alternative for the treatment of infections caused by multidrug-resistant pathogens, since this combination is able to increase the antibacterial effect of aminoglycosides. This work aimed to evaluate the interaction and the modulating effect of the lectin of Dioclea violacea (DVL) and the antibiotic gentamicin against multiresistant bacterial strains. Initially, seeds were collected, flour was prepared and lectin was extracted using 0.15 M NaCl saline solution, followed by lectin purification by means of sephadex G-75 affinity chromatography. Pure lectin was used in the interaction tests, which were inhibition of hemagglutinating activity and fluorescence spectroscopy in order to measure the interaction constant between lectin and antibiotic. In biological assays, multidrug-resistant strains of Staphylococcus aureus 10, Escherichia coli 06 and Pseudomonas aeruginosa 15 were used, where the minimum inhibitory concentration of lectin capable of inhibiting bacterial growth was observed, a subinhibitory concentration of lectin was used in the modulation test of the lectin antibiotic activity, aiming to observe the ability of lectin to potentiate the effect of gentamicin against bacterial strains, in addition, the DVL Influence assay on the development of the adaptive phenotype for gentamicin was carried out. The chromatogram profile showed two peaks, the first (PI) corresponding to the fraction of unretained protein and the second (PII) corresponding to the fraction of retained protein. In electrophoresis, PII showed three bands corresponding to the α (25.5 kDa), β (14 kDa) and γ (12 kDa) DVL chains. The inhibition of hemagglutinating activity showed that DVL has an affinity for gentamicin, glucose and mannose with minimum inhibitory concentrations of 12.5, 25 and 12.5 mM, respectively. The results revealed a gradual decrease in the fluorescence intensity of DVL leading to extinction in the presence of increasing concentrations of gentamicin. The MIC obtained for DVL against all strains studied was not clinically relevant (MIC ≥ 1024 µg / mL). However, when DVL was combined with gentamicin, a significant increase in antibiotic activity was observed, representing a reduction of 80.1% and 60.3% in the amount of gentamicin needed to have the same effect in S. aureus and E. coli respectively, it was found after 10 days of continuous treatment that DVL reduced the tolerance of S. aureus to gentamicin. In this way it was observed that gentamicin can interact with DVL through the carbohydrate recognition domain (CRD), suggesting that the results obtained in this study may be directly related to the DVL-gentamicin interaction.