| Congresso Brasileiro de Microbiologia 2023 | Resumo: 605-2 | ||||
Resumo:Considering the vast diversity of existing microorganisms, microbial metabolites are a potential source of bioactive molecules used in many industrial processes and for developing biofungicides for agricultural use. Cyclic Lipopeptides (CLPs) produced by many Bacillus species have potent antifungal activity by forming ion-conducting pores in the cytoplasmic membrane, increasing the membrane permeability, and even causing cell death. The objective of this work was to evaluate and identify antifungal compounds produced by three strains of Bacillus velezensis (CT02, LIS05, and IM14) against Fusarium verticillioides. First, the strains were grown in four media to determine the best medium for metabolites induction. They were TSB Soy Tryptone Broth (17 g/L casein peptone, 3 g/L soy peptone, 2.5 g/L glucose, 5 g/L sodium chloride, 2.5 g/L dipotassium phosphate), Potato Dextrose Broth (200 g/L potato infusion, 20 g/L glucose), Luria Bertani LB with glucose (20 g/L glucose, 10 g/L tryptone, 5 g/L yeast extract, 5 g/L NaCl), and Landy (20 g/L glucose, 5 g/L L-glutamic acid, 0.5 g/L MgSO4, 0.5 g/L KCl, 1.0 g/L KH2PO4, 0.0012 g/L Fe2SO3, 0.0014 g/L MnSO4, 0.0016 g/L CuSO4). After incubation at 28°C and 160 rpm for 72 h, the supernatants were centrifuged at 12,000 g for 10 minutes and filtered through 0.22 μm membranes. The bacterial extracts were tested for antifungal activity against F. verticillioides, using broth dilution to determine the susceptibility of filamentous fungi. For micro dilutions, concentrations ranging from 0.3% to 100% of the crude extract and positive and negative controls were used. The microplates were incubated at 28°C without agitation for 96 h. After this period, the absorbance was read at 490 nm in a digital spectrophotometer to determine the reduction in fungal growth. The strains were grown in the culture medium where the highest production of antifungal compounds occurred to identify the metabolites. After incubation for 72 hours at 28 °C and 160 rpm, the released supernatant was collected by centrifugation and extraction performed by acid precipitation. Using the TSB medium, the CT02 strain extract showed the highest percentage of inhibition (45.3%) at the lowest concentration (0.3%). The Landy medium showed better results for IM14 and LIS05, which inhibited the growth of F. verticillioides by 100% and 98.7%, respectively, at a concentration of 50%. The results showed that the inhibition percentages in different concentrations of crude extracts varied with the medium. Second, UHPLC Ultra Performance Liquid Chromatograph coupled with Quadrupole/Time of Flight (UPLC-QTOF-MSE) Xevo System (Waters Corp, Milford, MA, USA) was used for identifying the antifungal compounds. Different isoforms of CLPs, such as Iturin, fengycin, and surfactin were detected in IM14 and LIS05, and fengycin and surfactin in CT02. The strains from this work exhibited a directly antagonistic effect against F. verticillioides, and CLPs possibly played an important role. Palavras-chave: antagonistic activity, biofungicides, biosurfactants, lipopeptides Agência de fomento:Embrapa Milho e Sorgo/CNPq/Simbiose | |||||