RESUMO
Citral is a flavor component that is commonly used in food, beverage and fragrance industries. Cronobacter sakazakii is a food-borne pathogen associated with severe illness and high mortality in neonates and infants. The objective of the present study was to evaluate antimicrobial effect of citral against C. sakazakii strains. The minimum inhibitory concentration (MIC) of citral against C. sakazakii was determined via agar dilution method, then Gompertz models were used to quantitate the effect of citral on microbial growth kinetics. Changes in intracellular pH (pHin), membrane potential, intracellular ATP concentration, and membrane integrity were measured to elucidate the possible antimicrobial mechanism. Cell morphology changes were also examined using a field emission scanning electron microscope. The MICs of citral against C. sakazakii strains ranged from 0.27 to 0.54 mg/mL, and citral resulted in a longer lag phase and lower growth rate of C. sakazakii compared to the control. Citral affected the cell membrane of C. sakazakii, as evidenced by decreased intracellular ATP concentration, reduced pHin, and cell membrane hyperpolarization. Scanning electron microscopy analysis further confirmed that C. sakazakii cell membranes were damaged by citral. These findings suggest that citral exhibits antimicrobial effect against C. sakazakii strains and could be potentially used to control C. sakazakii in foods. However, how it works in food systems where many other components may interfere with its efficacy should be tested in future research before its real application.
Assuntos
Antibacterianos/farmacologia , Cronobacter sakazakii/efeitos dos fármacos , Monoterpenos/farmacologia , Monoterpenos Acíclicos , Trifosfato de Adenosina/análise , Cronobacter sakazakii/química , Concentração de Íons de Hidrogênio , Potenciais da Membrana/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Microscopia Eletrônica de VarreduraRESUMO
Cronobacter sakazakii is an opportunistic pathogen transmitted by food that affects mainly newborns, infants, and immune-compromised adults. In this study, the antibacterial activity of ferulic acid was tested against C. sakazakii strains. Minimum inhibitory concentration of ferulic acid against C. sakazakii strains was determined using the agar dilution method. Changes in intracellular pH, membrane potential and intracellular ATP concentration were measured to elucidate the possible antibacterial mechanism. Moreover, SYTO 9 nucleic acid staining was used to assess the effect of ferulic acid on bacterial membrane integrity. Cell morphology changes were observed under a field emission scanning electron microscope. The minimum inhibitory concentrations of ferulic acid against C. sakazakii strains ranged from 2.5 to 5.0 mg/mL. Addition of ferulic acid exerted an immediate and sustained inhibition of C. sakazakii proliferation. Ferulic acid affected the membrane integrity of C. sakazakii, as evidenced by intracellular ATP concentration decrease. Moreover, reduction of intracellular pH and cell membrane hyperpolarization were detected in C. sakazakii after exposure to ferulic acid. Reduction of green fluorescence indicated the injury of cell membrane. Electronic microscopy confirmed that cell membrane of C. sakazakii was damaged by ferulic acid. Our results demonstrate that ferulic acid has moderate antimicrobial activity against C. sakazakii. It exerts its antimicrobial action partly through causing cell membrane dysfunction and changes in cellular morphology. Considering its antimicrobial properties, together with its well-known nutritional functions, ferulic acid has potential to be developed as a supplement in infant formula or other foods to control C. sakazakii.
Assuntos
Antibacterianos/metabolismo , Ácidos Cumáricos/metabolismo , Cronobacter sakazakii/metabolismo , Suplementos Nutricionais , Conservantes de Alimentos/metabolismo , Trifosfato de Adenosina/metabolismo , Antibacterianos/farmacologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Proliferação de Células/efeitos dos fármacos , China , Contagem de Colônia Microbiana , Ácidos Cumáricos/farmacologia , Cronobacter sakazakii/efeitos dos fármacos , Cronobacter sakazakii/crescimento & desenvolvimento , Cronobacter sakazakii/ultraestrutura , Farmacorresistência Bacteriana , Conservantes de Alimentos/farmacologia , Humanos , Concentração de Íons de Hidrogênio , Lactente , Alimentos Infantis/microbiologia , Fórmulas Infantis/microbiologia , Líquido Intracelular/efeitos dos fármacos , Líquido Intracelular/metabolismo , Potenciais da Membrana/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Microscopia Eletrônica de Varredura , Especificidade da EspécieRESUMO
Syringic acid (SA) has been reported to exhibit antibacterial ability against various microorganisms, but little work has been done on its effect on Cronobacter sakazakii. In this study, minimum inhibitory concentrations (MICs) of SA against various C. sakazakii strains were determined. Moreover, changes in intracellular ATP concentration, intracellular pH (pHin), membrane potential and membrane integrity were measured to evaluate the influence of SA on cell membrane. Finally, field emission scanning electron microscope (FESEM) was used to assess the morphological changes of bacterial cells caused by SA. It was shown that the MICs of SA against all tested C. sakazakii strains were 5mg/mL. SA retarded bacterial growth, and caused cell membrane dysfunction, which was evidenced by intracellular ATP concentration decrease, pHin reduction, cell membrane hyperpolarization and changes in cellular morphology. These findings indicated that SA has potential to be developed as a natural preservative to control C. sakazakii in foods associated with this pathogen and prevent related infections.
