Bacillus thuringiensis and its toxins

The genes responsible for the crystal proteins are called cry genes, and the crystal toxin is called 5-endotoxin or Cry toxin, whereas the genes responsible for toxic cytosolic proteins (Cyt toxins) are called cyt genes. Many Bt strains are able to produce a number of such smaller cytosolic endotoxins in addition to the 8-endotoxins, but these are often deposited in inclusion bodies inside the crystals where they can comprise a considerable part of the crystal. Unlike the Cry toxins, the other toxins display a broader unspecific activity and may have some mammalian toxicity. They include the P-exotoxins, hemolysins, and enterotoxins. 

Hemolysins, which lyse vertebrate erythrocytes, are important virulence factors in vertebrate bacterial pathogens. Such toxins are also found in some Bt strains and seem to be identical to the hemolysin found in B. cereus. Some Bt isolates have been found to produce the same type of diarrhea-producing enterotoxins as B. cereus. Bt may therefore have implications in causing gastroenteritis. 

Bt produces and secretes a number of enzymes, e.g., chitinases, proteases, and phospholipases, of importance to the pathogenicity. They disrupt the per-itrophic membrane, providing access for the true toxins to the gut epithelium. 

A new class of insecticidal toxins called vegetative insecticidal proteins has recently been isolated from Bt. They are produced during the vegetative growth stage. The proteins are different from other known proteins, and their function and versatility for insect control have yet to be elucidated. 

As mentioned, the 8-endotoxin (Cry protein) that makes up most of the conspicuous crystals is the main insecticidal component of B. thuringiensis. At sporulation, the majority of Bt strains produce crystalline inclusions that contain this insecticidal 8-endotoxin. The crystals account for 20% or more of the total bacterial protein at sporulation and may contain one or several endotoxins, which differ in activity. Many Bt toxin genes and genes for some 

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The insecticidal activity of Bacillus thuringiensis and its toxin is considerably increased when combined with lecithin (381). 

Nakanishi, K., K. Yaoi, N. Shimada, T. Kadotani, and R. Sato. 1999. Bacillus thuringiensis insecticidal CrylAa toxin binds to a highly conserved region of animopeptidase N in the host insect leading to its evolutionary success. Biochim. Biophys. Acta. 1 57-63. 

A possible example of this thesis is the crystalline insect toxin found in Bacillus thuringiensis spores and discussed here by Dr. Anderson. Although neither the bacillus nor its spores exhibit useful antibiotic activity against other microorganisms, the very specific toxicity to insects has become of major commercial interest. The enormous number and variety of fungal species available for further examination must lead inevitably to one or more which produces pesticidal metabolites. 

Bacillus thuringiensis produces a variety of organic compounds which are toxic to the larvae of certain susceptible insect hosts. Among the toxic entities are proteinaceous crystals, probably three soluble toxins, and certain enzymes. The protein material is the major toxin active in killing lepidopterous larvae. The protein is formed by the cells apparently in close synchrony with sporulation, and its nature is a constant function of bacterial strain. The mode of action of the protein is under study. The sequence of events in the pathology observed is probably solubilization of the crystal (enzymatic or physical)—>liberation of toxic unit—>alteration of permeability of larval gut wall— change in hemolymph pH—>invasion of hemolymph by spores or vegetative cells of the bacterium. 

The varieties of Bacillus thuringiensis used commercially survive when injected into mice, and at least one of the purified insecticidal toxins is toxic to mice. Infections of humans have been extremely rare (two recognized cases) and no occurrences of human toxicosis have been reported. From studies involving deliberate ingestion by human subjects, it appears possible, but not likely, that the organism can cause gastroenteritis. Bacillus thuringiensis... 

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. 

Applications of immunoassay to pesticide chemistry have been described which address some difficult problems in analysis by classical methods. These include stereospecific analysis of optically active compounds such as pyrethroids (38), analysis of protein toxins from Bacillus thuringiensis (5,37), and compounds difficult to analyze by existing methods, such as diflubenzuron (35) and maleic hydrazide (15 also Harrison, R.O. Brimfield, A.A. Hunter, K.W.,Jr. Nelson, J.O. J. Agric. Food Chem. submitted). An example of the excellent specificity possible is seen in assays for parathion (10) and its active form paraoxon (3). Some immunoassays can be used directly for analysis without extensive sample extraction or cleanup, dramatically reducing the work needed in typical residue analysis. An example of this is given in Figures 2 and 3, comparing the direct ELISA analysis of molinate in rice paddy water to the extraction required before GC analysis. 

Bacillus thuringiensis (Bt) is a microbe that produces a toxin deadly to several kinds of caterpillars that ingest it. Bt toxin causes mature caterpillar gut cells to swell, burst, and die. Normally, if the Bt concentration is high enough, the caterpillar dies. However, a lower Bt concentration allows surviving cells to emit cytokines (see Section 6.20), which signal gut stem cells to multiply and rapidly form new mature gut cells. If more new cells can be produced than are killed, the caterpillar survives (ASAE, 2001). 

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