Delta-endotoxins

Different subspecies have been shown to be active against different orders of insect. Although the method of insect death is the same, the different endotoxins either have different binding regimes or are activated by the gut enzymes of different insect species and this leads to the differences in activity spectrum. There is evidence, however, that different insect orders act upon the delta-endotoxins in different ways. Treatment of a pro-toxin with lepidopteran gut juice leads to a toxin that is active against Lepidoptera whereas treatment with dipteran gut juice produces... 

The key insect-specific bacterium is Bacillus thuringiensis and this is through the production of delta-endotoxins. It has been shown that if the endotoxins are applied to crops together with living spores of the bacterium, insect control is improved. It is common for living formulations to be sold. 

Li JD, Carroll J, Ellar DJ (1991) Crystal structure of insecticidal delta-endotoxin from Bacillus thuringiensis at 2.5 A resolution. In Nature 353 815-821. 

English L and Slatin SL (1992) Mode of action of delta-endotoxins from Bacillus thuringiensis A comparison with other bacterial toxins. Insect Biochemistry and Molecular Biology 22 1-7. 

The use of this bacterial species has been deemed to be ideally suited for GMO use due to possession of toxins known as delta endotoxins. The structure of the delta endotoxin, is complex, containing three major regions or sections that each connote differential characteristics to the ultimate toxin (Figure 1). 

Figure 1 Molecular structure of Bacillus thuringienses delta endotoxin. (Reproduced from Li J, Carroll J, and Ellar DJ (1991) Nature 353 815-821, with permission of the Nature Publishing Group.)...

Coded genes for the Bt delta endotoxin production have been isolated for insertion into a variety of vegetables including potatoes, field corn, sweet corn, and soybeans. 

Structural and functional studies of a synthetic peptide that mimics a proposed membrane inserting segment of a Bacillus thuringiensis delta-endotoxin have been conducted. An NMR study of a methanol solution of a synthetic 31-mer peptide corresponding to the sequence of a putative pore-forming segment of the CrylA(c) toxin showed that the peptide exists as an a-helix. Hie peptide forms discrete, characterizable channels in planar lipid bilayers. It is possible that this helix is a component of the transmembrane pore formed by Bacillus thuringiensis delta-endotoxins in vivo. 

Commercial preparations of B.t.. obtained through conventional fermentation techniques, have been used for more than two decades as biological insecticides (I). They exhibit desirable properties such as high insect toxicity and environmental safety. B.t. does not affect non-target insects and is completely nontoxic to vertebrates. Nevertheless its use has been limited due to high production costs, limited stability in field conditions and a too narrow insecticidal spectrum. The insecticidal activity of B.t. relies in the crystalline inclusions which are produced upon sporulation. The crystals contain insecticidal proteins, delta-endotoxins, which affect the midgut epithelium of sensitive insects. The exact mechanism of their toxic activity is still unknown. 

Bt2 is an example of a type 1 delta-endotoxin. It has a relatively broad spectrum of insecticidal activity, but is nearly nontoxic to S. littoralis. A type 2 toxin is found in pure form in the crystals of B.t. thuringiensis strain 4412 and is toxic to Pieris brassicae but not to M. sexta and S. littoralis. An example of a type 3 toxin is present together with a type 1 toxin in B.t. aizawai and is characterized by high toxicity to S. littoralis. 

Specific monoclonal antibodies were generated against these three types of delta-endotoxins. These antibodies provide an efficient tool for very rapid immunological screening of large numbers of B.t. strains. 

Mode of Action. Despite the wide spectrum of insect host range activities displayed by Bt varieties, the mode of action of the insecticidal delta endotoxins is similar. When susceptible insects ingest Bt protein crystals, the first gross symptom observed is feeding inhibition due to paralysis of the digestive tract, including mouth parts. This usually occurs within one hour of ingestion ( ). 

Figure 1. Foliar persistence of microbial insecticides on cabbage. Arlington, Wisconsin 1988. Dipel and Javelin are commercial products based on Bt var. kurstaki. Persistence was measured by bioassay of field treated foliage, removed at various times post-application against 3rd instar DBM larvae. Rates of product applied are expressed in terms of grams of delta endotoxin (as measured by polyacrilamide gel electrophoresis) applied per acre.

Unique Mode of Action. As described above, the Bt delta endotoxin is a stomach poison which acts on the midgut epithelial cells of susceptible insects. This is in contrast to the typical mode of action for synthetic chemical insecticides, which act as nerve poisons. Because the Bt site of action is so different from that of synthetic insecticides, Bt has been successfully utilized for control of otherwise resistant insects such as mosquitoes, the diamondback moth and the Colorado potato beetle. 

Flexibility of Use. Products based on Bt have been successfully incorporated into pest management programs in agriculture, forestry and vector control. This is largely due to the fact that the Bt delta endotoxin and spores are relatively stable when tank mixed with conventionally used fungicides, insecticides and spray adjuvants. In addition, most commercial Bt formulations have been designed to optimize shelf life, as well as physical compatibility with other agricultural chemicals and application systems. 

The dead bacterial cell wall now serves as a protective "microcapsule" for the enclosed delta endotoxin. 

Knowles, B.H. and Dow, J.A.T. (1993). The crystal delta-endotoxins of Bacillus thuringiensis - models for their mechanism of action in the insect gut. Bio Essays 15,469-476. 

Hofmann, C. and Luthy, P. (1986). Binding and activity of Bacillus thuringien-sis delta-endotoxin to invertebrate cells. Arch. Microbiol. 146,7-11. 

Hofmann, C., Luthy, P., Hutter, R. and Pliska, V. (1988). Binding of the delta-endotoxin from Bacillus thuringiensis to brush-border membrane vesicles of the cabbage butterfly (Pieris brassicae). Eur. J. Biochem. 173, 85-91. 

Simpson, R.M., Burgess, E.P.J. and Markwick, N.P. (1997). Bacillus thuringiensis delta-endotoxin binding sites in two Lepidoptera, Wiseana spp. and Epiphyas postvittana. J. Invertebr. Pathol. 70,136-142. 

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