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Applied and Environmental Microbiology, August 2009, p. 5237-5243, Vol. 75, No. 16
0099-2240/09/$08.00+0 doi:10.1128/AEM.00532-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Bacillus thuringiensis Bel Protein Enhances the Toxicity of Cry1Ac Protein to Helicoverpa armigera Larvae by Degrading Insect Intestinal Mucin
Shangling Fang,1,3
Li Wang,1
Wei Guo,2*
Xia Zhang,2
Donghai Peng,1
Chunping Luo,1
Ziniu Yu,1 and
Ming Sun1*
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, People's Republic of China,1 Biological Control Center of Plant Diseases and Plant Pests of Hebei Province, College of Life Sciences, Agricultural University of Hebei, Baoding 071001, People's Republic of China,2 Key Laboratory of Fermentation Engineering (Ministry of Education), College of Bioengineering, Hubei University of Technology, Wuhan 430064, People's Republic of China3
Received 1 March 2009/ Accepted 8 June 2009
Bacillus thuringiensis has been used as a bioinsecticide to control agricultural insects. Bacillus cereus group genomes were found to have a Bacillus enhancin-like (bel) gene, encoding a peptide with 20 to 30% identity to viral enhancin protein, which can enhance viral infection by degradation of the peritrophic matrix (PM) of the insect midgut. In this study, the bel gene was found to have an activity similar to that of the viral enhancin gene. A bel knockout mutant was constructed by using a plasmid-free B. thuringiensis derivative, BMB171. The 50% lethal concentrations of this mutant plus the cry1Ac insecticidal protein gene were about 5.8-fold higher than those of the BMB171 strain. When purified Bel was mixed with the Cry1Ac protein and fed to Helicoverpa armigera larvae, 3 µg/ml Cry1Ac alone induced 34.2% mortality. Meanwhile, the mortality rate rose to 74.4% when the same amount of Cry1Ac was mixed with 0.8 µg/ml of Bel. Microscopic observation showed a significant disruption detected on the midgut PM of H. armigera larvae after they were fed Bel. In vitro degradation assays showed that Bel digested the intestinal mucin (IIM) of Trichoplusia ni and H. armigera larvae to various degrading products, similar to findings for viral enhancin. These results imply Bel toxicity enhancement depends on the destruction of midgut PM and IIM, similar to the case with viral enhancin. This discovery showed that Bel has the potential to enhance insecticidal activity of B. thuringiensis-based biopesticides and transgenic crops.
* Corresponding author. Mailing address for Ming Sun: College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, People's Republic of China. Phone: 86-27-87283455. Fax: 86-27-87280670. E-mail: m98sun{at}mail.hzau.edu.cn. Mailing address for Wei Guo: College of Life Sciences, Agricultural University of Hebei, Baoding 071001, People's Republic of China. Phone and fax: 86-312-7528-178. E-mail: guowei{at}hebau.edu.cn
Published ahead of print on 19 June 2009.
Applied and Environmental Microbiology, August 2009, p. 5237-5243, Vol. 75, No. 16
0099-2240/09/$08.00+0 doi:10.1128/AEM.00532-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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