Production of delta-endotoxin and hydrolytic enzymes of Bacillus Thuringiensis rt in two culture media using for Spodoptera Frugiperda control

Abstract

Biopesticides prepared from Bacillus thuringiensis (Bt) are an option for pest control for agriculture, forestry and public health. This bacteria produces delta-endotoxin and hydrolytic enzymes that increase its pathogenicity to target insects and could be exploited industraly. It was evaluated the production of delta-endotoxin and hydrolytic enzymes from Bt RT in two culture media. Were used Bt RT native isolate, grown in Luria Bertani and in the M5 medium (formulated from agro-industrial waste), using an Infors 3L work volume fermenter. Delta-endotoxin production was considered at fifth day by solubilization in alkaline conditions method using Bradford assay. For detecting hydrolytic activities 10 μl of culture supernatant was seeded into wells performed on water agar (1,5%), pH 7, containing the different substrates. Incubated for 24 h at 30 °C. Hydrolysis halos were developed with iodine vapors. The product obtained in both media was tested against Spodoptera frugiperda larvae on artificial diet and maize seedlings. After 5 days it reaches a concentration of delta-endotoxin of 155,67 mg / L in LB and 664,32 mg / L in M5. For the hydrolytic activities, the radius (mm) of halos observed at 5 days were: for amylase (BL: 0,00; M5: 2,232), carboxymethylcellulose (LB: 0,466; M5: 7,97), protease (BL: 1,12; M5: 5,29), chitinase (BL: 0,50; M5: 8,86) and xylanase (BL: 0,00; M5: 6,21). No activity was detected in both lipase and pectinase media. The effectiveness of the formulation was 100% for M5 both diet and seedlings, while effectiveness with LB medium was 61,40% and 58,33% respectively.