Isolation and Screening of Antagonistic Bacteria for Inhibition of Pathogenic Fungi in Rice
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Abstract
This study aims to isolate soil bacteria from rice fields and to determine antifungal activity of the isolates. Ten soil samples were collected in Bang Len district areas areas: Lam Pa Ya, Bang Sai Pa, Nil Phet, Sai Ngam and Bua Pak Tha subdistricts, and were cross streaked on Tryptic soy agar (TSA) medium. A total of 112 bacteria was isolated. Dual-culture antagonistic tests were conducted for all bacterial isolates against rice pathogenic fungi on PDA medium at 28°C for 5 days. The findings showed that only 3 isolates, namely N5-5, G4-1 and G5-5, had antifungal activities against Rhizoctonia solani, and only 4 isolates, namely N1-1, N5-5, G4-1 and G5-5, had antifungal activities against Bipolaris oryzae. However, all isolates had no antifungal activity against Pyricularia grisea. Selected bacteria were cultured in nutrient broth medium for 18 hours, and cell turbidity was adjusted OD 600 nm to 0.5 (108cell/ml), added 1% inoculum in NB medium and incubated at room temperature (25-30oC) at 200 rpm shaking rate for 48 hours. The cells were centrifuged at 12000 rpm at 4oC for 2 min. The supernatant was added to the PDA medium at a ratio of 2:18. The antifungal activities were measured as the mean colony diameter for growth inhibition. One of the isolates (G4-1) had a moderate level of antifungal activity against R. solani at 54.29%, whereas the isolated N5-5 had the moderate level of antifungal activity against B. oryzae at 55.55%. Next, the optimization of antifungal production was studied in an NB medium at pH 3, 5, 7 and 9, respectively, and shaked at 200 rpm at 25, 30, 35 and 40°C for 48 hours. Results demonstrated that the isolated G5-5 cultured at pH 5 and 25°C had a very high level of antifungal activity against R. solani at 85.33% and B. oryzae at 66.94%. The morphological study of antagonistic bacteria G5-5 showed that it was a gram-positive rod with single and linear cell arrangement and endospore development. Therefore, we suggest that the antagonistic bacterium belongs to the genus Bacillus.
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