(191aw) Evaluation of Culture Conditions on the Production of Antimicrobial Compounds Against Staphylococcus aureus from Lactobacillus viridescens  

Authors: 
Fernandes de Carvalho, L., Regional University of Blumenau
Bertoli, S., Regional University of Blumenau
Krebs de Souza, C., University of Blumenau
Sidooski, T., FURB
One of the major challenges facing the food industry today is to ensure the long life of food and food safety, due to the growing demands of the consumer market and enforcement agencies. Allied to this, the use of synthetic chemical compounds for food preservation and the consequences of its use has been discussed. Other highlighted topics are the use of chemical compounds in crops or grains to control and combat pests besides the alternatives for the treatment of new resistant bacteria to the usual treatments with known antibiotics. In this context, some natural compounds produced by bacteria which have an inhibitory effect against microorganisms associated with food contamination and diseases, have gained space in research and scientific studies. Most lactic acid bacteria species are capable to produce organic substances that are active against contaminating organisms and help in food preservation. One of the main limiting factors for the use of antimicrobial compounds on a large scale in the food industry or other fields is the need of further studies to identify new strains of bacteria which produce these compounds, together with the characterization and verification of safety for its use. In addition, the determination of ideal parameters for the cultivation and purification of these compounds, could help reduce the high costs involved in obtaining them, justify the need for further studies. In this context, this work evaluated the culture conditions for production of antimicrobial compounds by Lactobacillus viridescens and verified its potential of inhibition against the pathogenic microorganism Staphylococcus aureus. First, a culture of the microorganism Lactobacillus viridescens was carried out to determine its growth curve and to evaluate which phase of the growing curve shows the highest production of antimicrobial compounds. For this, three cultures were made in MRS broth (sharpe, de man and rugosa) at 36 °C for 54 h. During cultivation, samples were taken every 6 h for determination of the cell mass and inhibition generated by the antimicrobial compounds produced. Subsequently, the produced metabolites concentration through a freeze-drying process was performed. The produced cell mass was evaluated by the dry weight method. The evaluation of the potential inhibition against Staphylococcus aureus by the antimicrobial compounds was carried out by the agar plate diffusion method. According to the results, the microorganism Lactobacillus viridescens showed that at 6 h it was already in the exponential growth phase, which lasted 24 h, where the transition to the stationary growth phase occurred. It was verified that at the time of 24 h, largest halos of inhibition against the microorganism Staphylococcus aureus were obtained, indicating that at this time the maximum production of antimicrobial compounds should occur. After the determination of the growth curve, a study was carried out to evaluate the influence of physical and nutritional factors on the productivity of the antimicrobial compounds. This study was conducted according to the fractional factorial experimental design type (26-2) with four replicates at the central point, and the study carried out in duplicate. The selected physical factors for this study were the incubation temperature of cultivation and the initial pH of the culture medium. The selected nutritional factors were glucose concentration, lactose concentration, concentration of the yeast extract and sodium chloride. The culture medium used was MRS. After cultivation, each experimental point had was tested and its inhibition evaluated following the same methods used during the evaluation of the growth curve. As a result of this study, it was possible to verify that the temperature and sodium chloride concentration factors had a negative effect, statistically significant effect at a 95% confidence level. This means that the higher the concentration of sodium chloride and the higher the temperature, the lower inhibition zone generated against Staphylococcus aureus. The factors pH, glucose concentration, lactose concentration and yeast extract concentration had a positive effect on the inhibition halo formed, but none of these factors was statistically significant at a 95% confidence level.These results demonstrate the ability of lactic acid bacteria Lactobacillus viridescens to produce compounds with antimicrobial effect against the pathogen Staphylococcus aureus. The study also showed that the antimicrobial metabolites produced by Lactobacillus viridescens are produced at the end of their exponential growth phase, and the incubation temperature during cultivation as well as the concentration of sodium chloride in the culture medium are the main parameters associated with production of antimicrobial compounds by Lactobacillus viridescens.