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PubMedDate not supplied

[Scanning tunneling and electron microscopy of parasporal crystals in Bacillus thuringiensis].

Comparison of the forms of parasporal inclusions in Bacillus thuringiensis subsp. kurstaki and B. thuringiensis subsp. tenebrionis under a scanning electron microscope showed that, in addition to bipyramidal and cuboid crystals, cells of the subspecies kurstaki are able to form crystals taking the shape of flat parallelopipeds typical of the subspecies tenebrionis. The inclusions in the subspecies tenebrionis vary in size and electron density. Examination of the structure of crystals in B. thuringiensis subsp. tenebrionis under a scanning tunneling microscope showed that delta-endotoxins in the crystals are assembled in globules 5.0 nm in diameter and 1.0 nm in height. The globules are connected by its bases and form chains. The subunits of tetragonal crystals in the subspecies tenebrionis are vertically oriented with respect to the chain axis, as was revealed by other methods for subunits of bipyramidal crystals of the subspecies kurstaki.

Bacillus thuringiensis↗
PubMed1985

Separation of the cytolytic and mosquitocidal proteins of Bacillus thuringiensis subsp. israelensis.

The cytolytic and mosquitocidal proteins of Bacillus thuringiensis subsp. israelensis were isolated from parasporal crystals and subsequently separated from each other. The proteins were separated by gel filtration chromatography and their molecular weights were estimated by both gel filtration chromatography and SDS-polyacrylamide gel electrophoresis. The apparent molecular weights of the mosquitocidal protein and the cytolytic protein were estimated to be 65,000 daltons and 28,000 daltons, respectively.

Aedes↗
PubMed2004

Cloning and localization of vip3A gene of Bacillus thuringiensis.

An insecticidal protein gene, vip3A, was cloned from Bacillus thuringiensis strain WB50. The nucleotide sequence of 2,460 bp (GenBank acc. No. AY295778) showed 99% homology with the known vip3A genes. Using specific primers for vip3A gene, PCR was performed to demonstrate that the gene was not located on the bacterial chromosome and this was confirmed by Southern blotting using an internal fragment (486 bp) from vip3A gene as a probe. The gene was carried on a plasmid of 31.8 kb.

Bacillus thuringiensis↗
PubMed2006

Beyond the spore--past and future developments of Bacillus thuringiensis as a biopesticide.

Formulated and sporulated cultures of Bacillus thuringiensis (Bt) are widely used as foliar sprays as part of integrated pest management strategies against insect pests of agricultural crops. Although in several cases the presence of the spore has been shown to improve the activity of the product, other Bt-based insecticides have been developed in which the spore is absent. The most notable of these are transgenic plants expressing just the insect toxin gene from the bacterium. This paper will discuss these developments, and the advantages and disadvantages of having the spore present.

Bacillus thuringiensis↗
PubMed1979

On the formation of crystal proteins during sporulation in Bacillus thuringiensis var. thuringiensis.

The processes of sporulation and crystal formation in Bacillus thuringiensis could be desynchronized, using short term temperature shift-up conditions or addition of chloramphenicol at the beginning of sporulation. Isolation of a Sp-, Cr+ mutant, blocked at early stage of sporulation further confirmed that crystal formation can occur in the absence of sporulation. The presence of one of the two antigenic subunits of the crystal protein in chloramphenicol treated cultures and both of them in cultures shifted up to 37 degrees C was demonstrated. The insect toxicity was exhibited by both the extracts.

Antigens, Bacterial↗
PubMed2006

Proteomic profiling and identification of immunodominant spore antigens of Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis.

Differentially expressed and immunogenic spore proteins of the Bacillus cereus group of bacteria, which includes Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis, were identified. Comparative proteomic profiling of their spore proteins distinguished the three species from each other as well as the virulent from the avirulent strains. A total of 458 proteins encoded by 232 open reading frames were identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry analysis for all the species. A number of highly expressed proteins, including elongation factor Tu (EF-Tu), elongation factor G, 60-kDa chaperonin, enolase, pyruvate dehydrogenase complex, and others exist as charge variants on two-dimensional gels. These charge variants have similar masses but different isoelectric points. The majority of identified proteins have cellular roles associated with energy production, carbohydrate transport and metabolism, amino acid transport and metabolism, posttranslational modifications, and translation. Novel vaccine candidate proteins were identified using B. anthracis polyclonal antisera from humans postinfected with cutaneous anthrax. Fifteen immunoreactive proteins were identified in B. anthracis spores, whereas 7, 14, and 7 immunoreactive proteins were identified for B. cereus and in the virulent and avirulent strains of B. thuringiensis spores, respectively. Some of the immunodominant antigens include charge variants of EF-Tu, glyceraldehyde-3-phosphate dehydrogenase, dihydrolipoamide acetyltransferase, Delta-1-pyrroline-5-carboxylate dehydrogenase, and a dihydrolipoamide dehydrogenase. Alanine racemase and neutral protease were uniquely immunogenic to B. anthracis. Comparative analysis of the spore immunome will be of significance for further nucleic acid- and immuno-based detection systems as well as next-generation vaccine development.

Antigens, Bacterial↗
PubMedDate not supplied

[Analysis of the changes in the electrophysical and morphometric parameters in the growth and sporulation periods of Bacillus thuringiensis].

Changes in the electrophysical and morphometrical parameters of Bacillus thuringiensis cells were studied during their growth and differentiation using the electrophysical procedure with the frequency of an alternating electric field varying within a wide range. The appearing variations were interpreted on the basis of the data about changes in the concentration of vegetative cells and spores, the optical density and pH of the cultural broth, and the assimilation of glucose by the culture. The changes found in the electrophysical and morphometrical parameters were shown to accompany the main physiological processes in the cells. In principle, they can be used to control the process of cultivation.

Bacillus thuringiensis↗
PubMed2000

Kurstakins: a new class of lipopeptides isolated from Bacillus thuringiensis.

A novel class of lipopeptides was isolated from Bacillus thuringiensis kurstaki HD-1. Four compounds (1-4) were separated by high-performance liquid chromatography and their primary structures determined using a combination of chemical reactions and mass spectrometry. The four lipopeptides were found to have the same amino acid sequence, Thr-Gly-Ala-Ser-His-Gln-Gln, but different fatty acids. The fatty acyl chain is linked to the N-terminal amino acid residue via an amide bond. Each lipopeptide has a lactone linkage between the carboxyl terminal amino acid and the hydroxyl group in the side chain of the serine residue. Antifungal activity was demonstrated against Stachybotrys charatum.

Amino Acids↗
PubMed2007

Diversity of Colombian strains of Bacillus thuringiensis with insecticidal activity against dipteran and lepidopteran insects.

AIM: To evaluate the genetic and molecular diversity and insecticidal activity of Bacillus thuringiensis isolates from all the natural regions of Colombia. METHODS AND RESULTS: A total of 445 isolates from a collection of B. thuringiensis were characterized. The parasporal crystal morphology that was most abundant was bipyramidal (60%). Almost 10% of the isolates were toxic to Spodoptera frugiperda and 5.6% against Culex quinquefasciatus larvae. cry gene content determined by PCR indicated that 10.6% of the isolates contained cry1 genes and 1.1% contained cry2, cry4 or cry11 genes. Protein content of the parasporal crystal was determined by SDS-PAGE; 25 and 18 different protein profiles were found in isolates active against S. frugiperda and C. quinquefasciatus, respectively. CONCLUSIONS: Bacillus thuringiensis presents great genetic and molecular diversity even in isolates from the same soil sample. Moreover, the diversity and activity of the isolates might have a relationship with the geographical origin of the samples. SIGNIFICANCE AND IMPACT OF THE STUDY: The results obtained here indicate that some of the B. thuringiensis isolates characterized in this study are potential control agents that could be used in programmes against mosquitoes and S. frugiperda.

Adenosine↗
PubMed1998

Development of Bacillus thuringiensis fermentation and process control from a practical perspective.

Bacillus thuringiensis (Bt) is the most widely used biopesticide producer in the biological control market. It is very critical for the Bt pesticide industry to be able to achieve a high yield in the Bt fermentation process in order to reduce its cost and compete with chemical pesticides in the market. We review the overall development of Bt fermentation process research and provide our point of view for the future research opportunities and potential improvements. This minireview covers the areas of fermentation physiology, growth dynamics and high-yield process control. It is pointed out that many studies aimed to improve spore count and process research focusing on toxin protein yield is lacking. In addition, significant development opportunities reside in the process development for the genetically engineered Bt strains expressing multiple toxin proteins.

Bacillus thuringiensis↗
PubMed1998

Larvicidal toxicity of Japanese Bacillus thuringiensis against the mosquito Anopheles stephensi.

Japanese isolates of Bacillus thuringiensis were screened for larvicidal activity against the mosquito Anopheles stephensi, the urban malaria vector of the Indian subcontinent. Among more than 30 strains identified, larvicidal activity causing > 80% mortality in 72 h was demonstrated for 41/1449 (2.8%) isolates. The majority of strains and isolates (97.2%) exhibited little or no larvicidal activity. Anopheles-active strains belonged to more than 12 H serotypes, especially H3ade (serovar fukuokaensis) and H44 (serovar higo). SDS-PAGE profiles of inclusion proteins showed 4 distinct types among 6 active strains examined. The most active Japanese isolates were H20 strain 89-T-34-14 (LC50 4.4 micrograms/ml) and H44 serovar higo strain 74-E-45-24 (LC50 7.6 micrograms/ml), respectively, 13-fold and 23-fold less active than the international standard H14 serovar israelensis (LC50 0.33 microgram/ml).

Animals↗
PubMed1979

Mechanism of inhibition of active potassium transport in isolated midgut of Manduca sexta by Bacillus thuringiensis endotoxin.

After incubation at pH 10 or higher, Bacillus thuringiensis spores and endotoxin, at concentrations above 0.1 IU/ml, affected transport parameters in the isolated midgut of Manduca sexta larvae. (Toxic activity was lost during roughly 1 week at pH 11.) About 60% of the short-circuit current was inhibited, and the remainder was reversibly inhibited by anoxia. Electrical resistance was reduced by about 55% and oxygen uptake stimulated by about 30%. Influx of potassium from blood-side to lumen-side ('active' flux) was unaffected but flux in the reverse direction was nearly tripled. These results suggest that hydrolysis of the toxin yields an inhibitor of potassium transport, presumably a polypeptide. It is argued that inhibition is not primarily by uncoupling of oxidative phosphorylation, but instead by interference with an active depression of the efflux of potassium from lumen-side to blood-side.

Animals↗
PubMedDate not supplied

[Features of the kinetics of accumulating entomocidal exotoxins of Bacillus thuringiensis subsp. thuringiensis during fermentation stage of bitoxibacillin production].

Accumulation of Bacillus thuringiensis subsp. thuringiensis beta-exotoxin (BET) in the course of industrial fermentation (a stage in the production of the entomocidal biopreparation bitoxibacillin) has been studied. It has been demonstrated in model experiments that the decrease in the content of BET in the culture fluid is accounted for by the toxin interaction with an attendant product, the exogenous metabolite (EM). EM has been isolated from the culture fluid and characterized. EM causes alkalization of the medium, exerts entomocidal effects (in Musca domestica) and fails to form salts on treatment with BaCl2. The absorption spectrum of EM is similar to that of BET, showing a maximum at lambda = 259 nm. The light-absorbing chromophore is a pyrimidine or purine base. A method for quantitative determination of both exotoxins (BET and EM) in bacterial preparations has been developed.

Animals↗
PubMed1987

Purification of the mosquitocidal and cytolytic proteins of Bacillus thuringiensis subsp. israelensis.

Two proteins from parasporal crystals of Bacillus thuringiensis subsp. israelensis were purified to electrophoretic homogeneity by gel filtration and anion-exchange chromatography. The larger of the two proteins (molecular weight, 68,000) was not cytolytic, whereas the smaller protein (molecular weight, 28,000) was highly cytolytic when assayed against rat erythrocytes. When these proteins were assayed against larvae of the yellow fever mosquito, Aedes aegypti, the larger protein was at least 100-fold more toxic than the smaller protein. Although proteolytic activity was not detected in solubilized crystals nor in purified protein preparations, the toxin (molecular weight, 68,000) was readily degraded to smaller, nontoxic molecules, even when maintained at 4 degrees C. Mixtures of the two purified proteins were significantly more toxic to mosquito larvae than was either protein alone. Thus, it is likely that both the mosquitocidal and the cytolytic protein play roles in the overall insecticidal action of the parasporal crystal produced by this bacterium.

Aedes↗
PubMedDate not supplied

[Correlation between insecticidal and antibiotic activities of Bacillus thuringiensis parasporal crystals].

The work was concerned with studying the insecticide and antibiotic activities of solutions containing parasporal crystals of several Bacillus thuringiensis subspecies. The correlation coefficients of these values were determined. A direct correlation was established between the insecticide and antibiotic activities of solutions of parasporal crystals from several B. thuringiensis subspecies. Therefore, the quality of bacterial insecticides can be controlled by assaying the antibiotic activity of polypeptide crystals in biotechnology.

Anti-Bacterial Agents↗
PubMed2004

Response of larval Chironomus tepperi (Diptera: Chironomidae) to individual Bacillus thuringiensis var. israelensis toxins and toxin mixtures.

The biopesticide Bacillus thuringiensis israelensis (B.t.i.) is highly toxic to the larvae of Chironomus tepperi, an important pest of aerially sown rice in southern Australia. In this study, all of the known Cry genes and the Cyt1A gene from B.t.i. were expressed and tested for individual toxicity against fourth instar C. tepperi larvae. Possible synergism between toxins in two component mixtures involving all toxins except Cry10A was also evaluated. Of the Cry toxins, only Cry11A and Cry4B displayed substantial toxicity; however, both were 10- to 20-fold less toxic than the parental B.t.i. strain. The only detected synergy was between the mildly toxic Cry4A and Cyt1A toxins. In direct contrast to previous studies with mosquitoes, mixtures of Cry11A/Cry4B and Cry11A/Cyt1A were mildly antagonistic. The activity of Cry11A and Cry4B is sufficient to justify investigation as to whether their expression in transgenic rice plants could provide control of C. tepperi larvae.

Animals↗
PubMed1996

Inconsistencies in determining Bacillus thuringiensis toxin binding sites relationship by comparing competition assays with ligand blotting.

Receptor binding properties of Cry1Aa, Cry1Ab, and Cry1Ac Bacillus thuringiensis toxins to Lymantria dispar brush border membrane vesicles (BBMV) were investigated by competition assays and BBMV ligand blotting. Homologous competition binding assays demonstrated that all Cry IA toxins bound to L. dispar BBMV with high binding affinities. Heterologous competition assays suggested that all three toxins share the same binding sites. However, our ligand blotting experiments were not consistent with the heterologous competition assays. We identified a 120 kDa peptide as a Cry1Ac binding protein and a 210 kDa peptide as a Cry1Aa and Cry1Ab binding protein. These results indicated that determining the relationships between toxin binding sites by either competition assays or ligand blotting alone may not be conclusive.

Animals↗
PubMed1986

Integration of the delta-endotoxin gene of Bacillus thuringiensis into the chromosome of root-colonizing strains of pseudomonads using Tn5.

The delta-endotoxin gene (tox) from Bacillus thuringiensis subsp. kurstaki HD-1 was cloned into Tn5 and the resulting Tn5-tox element transposed from a vector plasmid into the chromosome of six corn-root-colonizing strains of Pseudomonas fluorescens and Agrobacterium radiobacter. Chromosomal integration of the tox gene maximized stability and minimized the potential for horizontal transfer of the tox gene to other bacterial species. Expression of the tox gene was demonstrated by Western blot analysis and by toxicity against larvae of the tobacco hornworm (Manduca sexta). The method described illustrates how a given gene can be stably integrated into the chromosome of diverse bacterial species.

Bacillus thuringiensis↗