The role of phytochemicals in Arcobacter butzleri resistance

• Autor(a) principal: Sousa, Vanessa Cristina Gouveia
• Data de Publicação: 2017
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10400.6/6392
• topic: Additive Interaction; Arcobacter Butzleri; Efflux Pump Inhibitors; Phytochemicals; Resistance; Domínio/Área Científica::Engenharia e Tecnologia::Engenharia Química

Arcobacter butzleri is an emergent pathogen found in a wide range of habitats and hosts, which has developed resistance to several antibiotics. Efflux pumps are an important mechanism of antimicrobial resistance, therefore, the use of efflux pump inhibitors (EPIs) may have the potential to restore A. butzleri susceptibility to old antibiotics. Plants have shown the ability to fight off infections despite the moderate antimicrobial action of some phytochemicals, so we aimed to test several bioactive compounds as putative EPIs, evaluating their role in the improvement of antibiotics’ performance against A. butzleri. To achieve this goal, the tolerance or resistance profile of A. butzleri strains regarding phytochemicals and antibiotics was traced through the determination of the minimum inhibitory concentration (MIC); assays of ethidium bromide accumulation were performed to assess the inhibition of the efflux pumps; the MIC of the phytochemicals in the presence of known EPIs was determined to examine the potential role of efflux pumps as resistance mechanism to the phytochemicals; checkerboard assays were made to investigate if the phytochemicals had a synergic interaction with the antibiotics; and finally, quorum sensing inhibition tests were carried out, since this mechanism is a promisor target to fight off bacterial infection. The determination of the MIC of the phytochemicals demonstrated that none of the compounds had antimicrobial activity at the concentrations tested, except for stilbenes, which MIC ranged from 64 to 512 µg/mL. Ethidium bromide accumulation assays showed that some of the tested phytochemicals presented a fluorescence folding increase higher than the controls, indicating that they may inhibit efflux pumps; however only the stilbenes presented a typical EPI profile. The assessment of the MIC of the phytochemicals in the presence of a sub-inhibitory concentration of EPIs, revealed that the importance of efflux pumps in the bacteria resistance to phytochemicals is dependent on the strain. Several phytochemicals were selected for checkerboard titration assays revealing no synergism with antibiotics, however, several cases of additivity were detected. Quorum sensing assays revealed that resveratrol and pinosylvin were able to inhibit this mechanism. In conclusion, some of the phytochemicals tested presented potential to reduce A. butzleri resistance to antibiotics as demonstrated by the results obtained to resveratrol, pinosylvin and gallic acid, which have shown an additive effect when combined with the antibiotics. According to the ethidium bromide accumulation assay, the additive action of resveratrol and pinosylvin may be associated with efflux pump inhibition. Furthermore, these two stilbenes also possess the capacity to inhibit quorums sensing, suggesting that they may be able to inhibit A. butzleri virulence traits.