Purification and characterization of a lipopeptide produced by Bacillus thuringiensis CMB26

• Published in: Journal of applied microbiology Vol. 97; no. 5; pp. 942 - 949
• Main Authors: Kim, P.I, Bai, H, Bai, D, Chae, H, Chung, S, Kim, Y, Park, R, Chi, Y.T
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Science Ltd 01-01-2004; Blackwell Science; Oxford University Press
• Subjects: Amino Acids - analysis; Anti-Infective Agents - chemistry; Anti-Infective Agents - isolation & purification; Anti-Infective Agents - pharmacology; Antibiosis; Bacillus thuringiensis; Bacillus thuringiensis - classification; Bacillus thuringiensis - genetics; Bacillus thuringiensis - metabolism; Bacillus thuringiensis CMB26; Bacterial Proteins - biosynthesis; Bacterial Proteins - chemistry; Bacterial Proteins - isolation & purification; Bacterial Proteins - pharmacology; bioactive properties; biocontrol agent; Biological and medical sciences; biological control; biological control agents; biosurfactants; Colletotrichum - drug effects; Colletotrichum gloeosporioides; cyclic lipopeptides; Escherichia coli; Fatty Acids - analysis; fengycin; Fundamental and applied biological sciences. Psychology; fungal antagonists; Glomerella cingulata; Insecticides - chemistry; Insecticides - isolation & purification; Insecticides - pharmacology; lipopeptide; Microbiology; Microscopy, Electron, Scanning; Mitosporic Fungi - drug effects; Mitosporic Fungi - ultrastructure; Pest Control, Biological - methods; Pieris rapae crucivora; plant pathogenic fungi; Soil Microbiology; surfactants; fengycin; Purification; Applied microbiology; Bacillus thuringiensis; Biological control; biocontrol agent; Bacillaceae; Fungi; Characterization; Bacillales; Bacteria; Fungi Imperfecti; Colletotrichum gloeosporioides; Bacillus thuringiensis CMB26; Lipopeptide; Thallophyta
• ISSN: 1364-5072; 1365-2672
• DOI: 10.1111/j.1365-2672.2004.02356.x

Aims: To isolate an antagonist for use in the biological control of phytopathogenic fungi including Colletotrichum gloeosporioides, then to purify and characterize the biocontrol agent produced by the antagonist. Methods and Results: Bacteria that exhibited antifungal activity against the causative agent pepper anthracnose were isolated from soil, with Bacillus thuringiensis CMB26 showing the strongest activity. A lipopeptide produced by B. thuringiensis CMB26 was precipitated by adjusting the pH 2 with 3 n HCl and extracted using chloroform/methanol (2:1, v/v) and reversed-phase HPLC. The molecular weight was estimated as 1447 Da by MALDI-TOF mass spectrometry. Scanning electron and optical microscopies showed that the lipopeptide has activity against Escherichia coli O157:ac88, larvae of the cabbage white butterfly (Pieris rapae crucivora) and phytopathogenic fungi. The lipopeptide had cyclic structure and the amino acid composition was L-Glu, D-Orn, L-Tyr, D-allo-Thr, D-Ala, D-Val, L-Pro, and L-Ile in a molar ratio of 3:1: 2:1:1: 2:1:1. The purified lipopeptide showed the same amino acid composition as fengycin, but differed slightly in fatty acid composition, in which the double bond was at carbons 13-14 (m/z 303, 316) and there was no methyl group. Conclusion: A lipopeptide was purified and characterized from B. thuringiensis CMB26 and found to be similar to the lipopeptide fengycin. This lipopeptide can function as a biocontrol agent, and exhibits fungicidal, bactericidal, and insecticidal activity. Significance and Impact of the Study: Compared with surfactin and iturin, the lipopeptide from B. thuringiensis CMB26 showed stronger antifungal activity against phytopathogenic fungi. This lipopeptide is a candidate for the biocontrol of pathogens in agriculture.