Showing papers on "Bacillus thuringiensis published in 1984"
Patent•
24 Sep 1984
TL;DR: In this article, a method for inserting expressible insecticidal protein structural genes into plant genomes is provided. In the preferred embodiments this invention comprises placing a structural gene for the Bacillus thuringiensis crystal protein under control of a plant or a T-DNA promoter and ahead of a poly-adenylation site followed by insertion of said promoter/ structural gene combination into a plant genome by utilizing a Agrobacterium tumefaciens Ti plasmid-based transformation system.
Abstract: A method for introducing expressible insecticidal protein structural genes into plant genomes is provided. In the preferred embodiments this invention comprises placing a structural gene for the Bacillus thuringiensis crystal protein under control of a plant or a T-DNA promoter and ahead of a poly-adenylation site followed by insertion of said promoter/ structural gene combination into a plant genome by utilizing a Agrobacterium tumefaciens Ti plasmid-based transformation system. The modified Ti plasmid is then used to transform recipient plant cells. Also provided are the plants and tissues produced by this method and bacterial strains, plasmids, and vectors useful for execution of this invention.
414 citations
TL;DR: The enzyme did not show a stringent requirement for a specific amino acid sequence at the cleavage site but prefers a hydrophobic residue on the C-terminal side and is explained in terms of the open structure of the cecropins and a pronounced inability of inhibitor A to attack globular proteins.
Abstract: The insect pathogen Bacillus thuringiensis produces an exoprotease, inhibitor A, at the beginning of the stationary growth phase. In vitro, the enzyme selectively destroys cecropins and attacins, two antibacterial proteins found in immune hemolymph from Hyalophora cecropia. The specificity of this enzyme was investigated using cecropin A(1–33) and HPLC for separation and characterization of the fragments obtained.
A maximum of 12 different peptides were produced and their positions in the known sequence of cecropin A(1–33) were deduced from their amino acid compositions. The enzyme did not show a stringent requirement for a specific amino acid sequence at the cleavage site but prefers a hydrophobic residue on the C-terminal side. The specificity of the enzyme is explained in terms of the open structure of the cecropins and a pronounced inability of inhibitor A to attack globular proteins.
151 citations
TL;DR: The presence or absence of mosquito-toxic activity in all Cry+ and Cry- variants of BTI was confirmed by bioassay of sporulated cultures against larvae of Aedes aegypti.
151 citations
TL;DR: The results indicate the isolation of a B. thuringiensis strain which shows the same toxicity as that of subsp.
105 citations
TL;DR: Results suggest that the Bacillus thuringiensis var.
104 citations
TL;DR: Antibiotic-resistant transductants derived from any of the three species were effective donors of plasmids to recipients from all three species.
Abstract: Bacteriophage CP-51, a generalized transducing phage for Bacillus anthracis, B. cereus, and B. thuringiensis, mediates transduction of plasmid DNA. B. cereus GP7 harbors the 2.8-megadalton multicopy tetracycline resistance plasmid, pBC16. B. thuringiensis 4D11A carries pC194, the 1.8-megadalton multicopy chloramphenicol resistance plasmid. When phage CP-51 was propagated on these strains, it transferred the plasmid-encoded antibiotic resistances to the nonvirulent Weybridge (Sterne) strain of B. anthracis, to B. cereus 569, and to strains of several B. thuringiensis subspecies. The frequency of transfer was as high as 10(-5) transductants per PFU. Tetracycline-resistant and chloramphenicol-resistant transductants contained newly acquired plasmid DNA having the same molecular weight as that contained in the donor strain. Antibiotic-resistant transductants derived from any of the three species were effective donors of plasmids to recipients from all three species.
96 citations
TL;DR: Two sets of inverted repeat DNA sequences, IR2150 and IR1750, were discovered flanking the crystal protein gene on the 75kilobase plasmid of Bacillus thuringiensis subsp. kurstaki HD73 as discussed by the authors.
Abstract: Two sets of inverted repeat DNA sequences, IR2150 and IR1750, were discovered flanking the crystal protein gene on the 75-kilobase plasmid of Bacillus thuringiensis subsp. kurstaki HD73. A restriction map of ca. 40 kilobases around the crystal protein gene was constructed, and the positions of the copies of IR2150 and IR1750 were determined. Three copies of IR2150 were found flanking the crystal protein gene in an inverted orientation, and one partial and three intact copies of IR1750 were found in both inverted and direct orientations around the gene. Hybridization experiments with fragments from within IR2150 and IR1750 demonstrated the presence of multiple copies of these sequences on the chromosome of B. thuringiensis subsp. kurstaki HD73 and also revealed a strong correlation between the presence of these sequences and the presence of the crystal protein gene on plasmids from 14 strains of B. thuringiensis.
88 citations
TL;DR: The biological authenticity of the cloned product was further confirmed by demonstrating that the cytotoxicity of the polypeptide was neutralised by antiserum to the authentic δ‐endotoxin or by preincubation with excess toxin receptor.
72 citations
TL;DR: This investigation shows that the medium containing V. subterranean can be used for the production of B. thuringiensis subsp.
Abstract: Five media, formulated from the seeds of five legume varieties, dried cow blood, and mineral salts, were assessed for the growth and production of insecticidal properties of Bacillus thuringiensis subsp. israelensis. Bacterial powders prepared from the broth cultures were assayed against the larvae of Aedes aegypti, Culex quinquefasciatus, and Anopheles gambiae. A standard primary powder of B. thuringiensis subsp. israelensis (IPS78) was included in the assay for comparison. Good growth was obtained in all the media, and all powders were effective against the three types of mosquito larvae. The powder containing ground seeds of Voandzeia subterranean was the most effective and compared favorably with the standard (IPS78). The concentrations required to kill 50% of the larvae of Aedes aegypti, C. quinquefasciatus, and Anopheles gambiae were 1.13 X 10(-2) +/- 1.79 X 10(-3), 1.83 X 10(-2) +/- 2.55 X 10(-3), and 2.25 X 10(-2) +/- 1.88 X 10(-3) micrograms/ml, respectively. This investigation shows that the medium containing V. subterranean can be used for the production of B. thuringiensis subsp. israelensis primary powder.
62 citations
TL;DR: Biochimie et genetique des endotoxines delta de B-thuringiensis, Perspectives d'utilisation du genie genetiques pour augmenter the production of the toxine.
Abstract: Biochimie et genetique des endotoxines delta de B-thuringiensis. Perspectives d'utilisation du genie genetique pour augmenter la production de la toxine
60 citations
TL;DR: The mild effect of pyrethroids on sporulation processes of B. thuringiensis, compared to effects of other classes of chemical insecticides, suggested little or no interference with the ecology and perpetuation of this useful bacterium at the site of application.
Abstract: We determined the effects of insecticides of different chemical groups on sporulation yields of Bacillus thuringiensis var. entomocidus . Among the carbamates tested, carbaryl had a more deleterious effect on the sporulation process of B. thuringiensis than methomyl. Within the organophosphorous group, phoxim inhibited sporulation less than profenofos. The pyrethroid group, represented by fenvalerate, cypermethrin, and permethrin, generally had less deleterious effects on sporulation of B. thuringiensis than did the carbamate and organophosphorous compounds. The pyrethroids and most organophosphorous compounds tested potentiated the activity of B. thuringiensis applied against the cotton leafworm, Spodoptera littoralis (Boisduval). The carbamates, diflubenzuron, and a combination of methomyl and diflubenzuron (disa) showed an additive effect when applied jointly with B. thuringiensis varieties. The mild effect of pyrethroids on sporulation processes of B. thuringiensis , compared to effects of other classes of chemical insecticides, suggested little or no interference with the ecology and perpetuation of this useful bacterium at the site of application. Synergistic interactions suggest that application of pyrethroids with B. thuringiensis may be a safe and effective means for controlling S. littoralis .
TL;DR: A procedure is described whereby soluble toxins are adsorbed to 0.8-micrometer latex beads, with retention of toxicity, and the latex bead assay should make it possible to analyze the structure and mode of action of the mosquito toxin.
Abstract: Solubilized crystals of Bacillus thuringiensis var. israelensis were 7000 times less toxic to Aedes aegypti larvae than intact crystals, presumably because mosquito larvae are filter feeders and selectively concentrate particles while excluding water and soluble molecules. A procedure is described whereby soluble toxins are adsorbed to 0.8-micrometer latex beads, with retention of toxicity. The latex bead assay should make it possible to analyze the structure and mode of action of the mosquito toxin.
TL;DR: This solubilization procedure gave protein gel profiles identical with those for intact crystals while maintaining full biological activity in the form of erythrocyte lysis capability.
Abstract: A procedure is described in which the protein crystals produced by Bacillus thuringiensis var. israelensis were solubilized in 50 mM NaOH with 10 mM EDTA at pH 11.7. This solubilization procedure gave protein gel profiles identical with those for intact crystals while maintaining full biological activity in the form of erythrocyte lysis capability. Crystals with and without protease activity were equally toxic to Aedes aegypti larvae.
TL;DR: Characterization of 10 mosquitocidal strains of Bacillus sphaericus indicated that the traits employed in this study readily distinguished these highly related organisms from strains of B. thuringiensis and B. cereus, suggesting that the properties reported to differentiate serovars can be attributed to the internal phenotypic diversity of the species.
TL;DR: The Staphylococcus aureus plasmid pC194 which codes for resistance to chloramphenicol was introduced into six Bacillus thuringiensis strains representing five varieties by protoplast transformation, introducing a new extrachromosomal element of somewhat lower electrophoretic mobility hybridizing with pC 194.
Abstract: The Staphylococcus aureus plasmid pC194 which codes for resistance to chloramphenicol was introduced into six Bacillus thuringiensis strains representing five varieties by protoplast transformation. Six other varieties could not be transformed. pC194 could be identified in transformed strains as autonomous plasmid. The transformed clones contained in addition a new extrachromosomal element of somewhat lower electrophoretic mobility hybridizing with pC194, and pC194 in multimeric forms. pC194 was also transferred from one B. thuringiensis variety to another and from Bacillus thuringiensis to Bacillus subtilis and vice versa by a conjugation-like process, requiring close cell-to-cell contact.
TL;DR: The amount of protoxin produced in these B. cereus transcipients , however, was somewhat less than that produced in the parental strain HD73 , implying that catabolic differences, gene dosage, or the presence of a chromosomal gene (or a combination of these) may be necessary for maximum production.
Abstract: A derivative of Bacillus thuringiensis subsp. kurstaki (HD-1) formed parasporal inclusions at 25 degrees C, but not at 32 degrees C. This strain differed from the parent only in the loss of a 110-megadalton (Md) plasmid, but plasmid and chromosomal copies of protoxin genes were present in both strains. On the basis of temperature shift experiments, the sensitive period appeared to be during midexponential growth, long before the time of protoxin synthesis at 3 to 4 h after the end of exponential growth. The conditional phenotype could be transferred by cell mating to naturally acrystalliferous Bacillus cereus. In all such cases, a 29-Md protoxin -encoding plasmid was transferred, but this plasmid alone was barely sufficient for protoxin synthesis. Protoxin production increased to detectable levels, but well below those of the parental donor strain, by simultaneous transfer of a 44-Md protoxin -encoding plasmid. Transfer of a 5-Md plasmid with the two larger protoxin -coding plasmids resulted in a protoxin synthesis level approaching that of the donor strain. A role for some of the cryptic plasmids of kurstaki in parasporal body formation was implied. In contrast, a closely related B. thuringiensis strain, HD73 , produced crystals at both 25 and 32 degrees C even when the capacity was transferred on a 50-Md plasmid to B. cereus. The amount of protoxin produced in these B. cereus transcipients , however, was somewhat less than that produced in the parental strain HD73 , implying that catabolic differences, gene dosage, or the presence of a chromosomal gene (or a combination of these) may be necessary for maximum production. A regulatory component of the 29-Md plasmid appeared to be trans-acting and dominant since B. cereus transcipients containing the 29-Md plasmid from kurstaki and the 50-Md plasmid from HD73 produced more protoxin at 25 degrees C than at 30 degrees C. Similar results were obtained when protoxin synthetic capacity was transferred from B. thuringiensis subsp. israelensis to the conditional B. thuringiensis subsp. kurstaki strain.
TL;DR: The rate of loss of crystal activity in natural soil correlated well with assay data reported in the literature using Galleria mellonella, which measures the combined activity of spore and crystal, but in autoclaved soil correlation was poor, probably due to variability in the bioassay data.
TL;DR: This paper summarizes the work of several laboratories engaged in determining how, at the molecular level, such regulation is accomplished with respect to sporulation in gram-positive bacteria.
Abstract: INTRODUCTION Several species of gram-positive bacteria are noted for their elaborate adaptive responses to conditions of nutrient depletion. These developmental changes are sometimes transient and reversible, as in the attainment of “competence” for DNA uptake and transformation among some Streptococcus , Staphylococcus , and Bacillus species, but also include examples of true cellular differentiation, as in the case of spore formation in most Bacillus and Streptomyces species. In both Bacillus and Streptomyces , sporulation is actually the culmination of a complex sequence of physiological and morphological changes, not all of which are directly connected with the generation of a spore. Many Bacillus species, for example, produce and secrete an abundance of exoenzymes and small molecules, such as proteases and antibiotics, as they sporulate (most of these substances having no apparent role in spore formation), and Streptomyces species are well known for the diversity of antibiotics and other secondary metabolites they produce as they reach stationary phase (whether or not spores are formed). A very striking sporulation-associated event (irrelevant to the formation of a viable spore) peculiar to Bacillus thuringiensis is the production of an intracellular crystalline protein aggregate (the “crystal toxin”), which has extremely potent insecticidal properties (Bechtel and Bulla 1976). Not surprisingly, the complex changes that accompany and comprise the process of Sporulation in these organisms are dependent on the temporally regulated expression of many sets of genes at multiple chromosomal locations. This paper summarizes the work of several laboratories engaged in determining how, at the molecular level, such regulation is accomplished...
TL;DR: It is suggested that canavanine enhances the effect of B. thuringiensis on gut permeability and active transport, as evidenced by lower LD 50 and LT 50 values, observed when M. sexta were reared on an artificial diet supplemented with a sublethal concentration of l -canavanine.
TL;DR: A strain of Bacillus thuringiensis subsp.
Abstract: A strain of Bacillus thuringiensis subsp. israelensis was found to be larvicidal to horn flies, Haematobia irritans (L. [Diptera:Muscidae]). The toxic activity was particulate, appeared during sporulation, and could be prevented by the addition of streptomycin before sporulation. Density gradient centrifugation in Renografin was used to separate endospores, crystals, and low-density particulate matter (fraction 3) from sporulated preparations. Larvicidal activity was restricted to purified crystals and fraction 3, indicating that delta-endotoxin of B. thuringiensis subsp. israelensis was active against horn fly larvae. Purified crystals produced mortality during larval feeding stages, but not pupal stages. Fraction 3 produced significant mortality during both larval and pupal stages. The mortality data indicated the presence of at least two dipteran-active toxins.
TL;DR: Three groups of linked markers were mapped in Bacillus thuringiensis 4042B by using two-, three-, and four-factor crosses mediated by the temperate bacteriophages TP-13 and TP-18, and Electron microscopic measurements of head sizes suggested that the volume of the TP- 13 phage head is seven times greater than that of theTP-18 phageHead.
Abstract: Three groups of linked markers were mapped in Bacillus thuringiensis 4042B by using two-, three-, and four-factor crosses mediated by the temperate bacteriophages TP-13 and TP-18. The order of markers was (trp-11, trp-2)-(leu-1, leu-2)-his-1-(lys-1, lys-2)-cys-1 in the first group; met-1-(argCl, argOl)-met-2-(pyr-1, pyrA2) in the second group; and met-3-pur-1-(nal-1, nal-2)-str-1-(pur-2, pur-4)-pur-3 in the third group. Electron microscopic measurements of head sizes suggested that the volume of the TP-13 phage head is seven times greater than that of the TP-18 phage head. The TP-18 genome was shown by DNA restriction analysis to have a molecular mass of 36 megadaltons. TP-13 was useful for scanning large segments of the B. thuringiensis chromosome, and TP-18 was effective for ordering markers too closely linked for simple resolution with TP-13.
TL;DR: The protein demonstrating larvicidal activity to the mosquito Aedes aegypti was purified from the alkali extract of the spore-parasporal inclusion complex of the isolate, 73-E-10-2, belonging to Bacillus thuringiensis serotype 10.
TL;DR: The moist micro habitat and moderate temperatures during most of the artichoke-growing season in the fog belt region of the central coast of California, together with the tunneling habit of the host larvae, suggest that the nematode could be successfully used as a biological control agent against the artICHoke plume moth larvae infesting the vegetative shoots.
Abstract: The entomogenous nematode Neoaplectana carpocapsae Weiser and the bacterium Bacillus thuringiensis var. kurstaki were evaluated in the field against the artichoke plume moth infesting artichoke in its vegetative growth phase. In general, nematode concentrations ≥ 1,000 nematodes/ml were highly effective against older larvae when applied to artichoke plants to runoff. Used at 2,000 nematodes per ml, the nematode gave 100% control of 3rd and 4th instars 15 days after treatment, and its residual effect, evaluated 24 days after treatment, was greater than that of a commonly used insecticide, methidathion. Although B. thuringiensis var. kurstaki used at the rate of 19.6 billion International Units/ ha gave significant larval control, its efficacy was numerically but not significantly lower than that of methidathion. Combination of the nematode and B. thuringiensis var. kurstaki did not result in significantly greater control than that achieved by the nematode used alone. The moist micro habitat and moderate temperatures during most of the artichoke-growing season in the fog belt region of the central coast of California, together with the tunneling habit of the host larvae, suggest that the nematode could be successfully used as a biological control agent against the artichoke plume moth larvae infesting the vegetative shoots.
TL;DR: Inhibition of nerve conduction in the sixth abdominal ganglion of the American cockroach was observed following application of protein A, which was found to be haemolytic to human and rabbit erythrocytes.
Abstract: Two proteins (protein A and B) were isolated from Bacillus thuringiensis var. israelensis crystals using ion-exchange chromatography. Protein A was found to have haemolytic and neurotoxic activities. Inhibition of nerve conduction in the sixth abdominal ganglion of the American cockroach was observed following application of protein A. Protein A was found to be haemolytic to human and rabbit erythrocytes. Protein B did not exhibit any of these activities.
TL;DR: In this article, Bacillus thuringiensis var. aizawai H-serotype 7, strain HD137, streptomycin-resistant mutant, were added to normal and autoclaved aliquots of p H 5 soil incubated at 25°C and −0.10 MPa water availability.
TL;DR: The solubilized entomotoxin of Bacillus thuringiensis subsp.
Abstract: The solubilized entomotoxin of Bacillus thuringiensis subsp. israelensis killed adult male and female mosquitoes of several genera and of various physiological states when it was administered orally. Adult mosquito mortality was further influenced when the preparation was contained in sucrose solution. The potential implication for the control of adult mosquitoes is discussed. Images
TL;DR: An experimental preparation of Bacillus thuringiensis Berliner var.
Abstract: An experimental preparation of Bacillus thuringiensis Berliner var. israelensis (Goldberg) tested against a sciarid fly Lycoriella mali (Fitch) in mushroom compost gave over 90% control. The LC95 was one part B. thuringiensis var. israelensis to 696 parts water (wt/wt); the L95 was one part B. thuringiensis var. israelensis to 64 parts water. With a 1:60 dilution, the mortality exceeded 99.5%. Two commercial preparations containing B. thuringiensis var. israelensis tested at a dilution of 1:100 produced mortality rates of either 79.2 or 94.4%.
TL;DR: Protein isolated from crystals ofB.
Abstract: Cultured tissue cells from lepidopteran and dipteran sources displayed an order-specific response to entomocidal protein from crystals of Bacillus thuringiensis. Protein isolated from crystals of B. thuringiensis subsp. kurstaki was effective against cells of the spruce budworm (Choristoneura fumiferana) and the tobacco hornworm (Manduca sexta), but was inactive against both mosquito cell lines tested (Aedes aegypti and Anopheles gambiae). Conversely, protein from inclusion bodies of B. thuringiensis subsp. israelensis was fully active only against the mosquito cell lines but displayed reduced (four- to seven-fold) toxicity for the lepidopteran cell lines. One exception to this pattern of specificity was observed with a Plodia interpunctella cell line, which failed to respond to either crystal protein preparation. The moth toxin was stable at 4 degrees C for months, whereas the mosquito toxin was susceptible to proteolytic degradation and was unstable for periods longer than 2 wk.
TL;DR: Encapsulation of Bacillus thuringiensis var.
Abstract: Encapsulation of Bacillus thuringiensis var. israelensis Serotype H-14 (B.t.i.) in polyethylene strongly increased its persistence against second stage Aedes aegypti larvae in laboratory conditions simulating natural field conditions. B.t.i. encapsulated in palmitic acid was less persistent. The reasons for the enhanced efficacy of encapsulated B.t.i. are discussed.