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Sample records for bacillus thuringiensis cry1ab

  1. 40 CFR 174.529 - Bacillus thuringiensis modified Cry1Ab protein as identified under OECD Unique Identifier SYN...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis modified Cry1Ab... Tolerance Exemptions § 174.529 Bacillus thuringiensis modified Cry1Ab protein as identified under OECD... Bacillus thuringiensis modified Cry1Ab protein as identified under OECD Unique Identifier SYN-IR67B-1 are...

  2. Identification of Bacillus thuringiensis Cry1AbMod binding-proteins from Spodoptera frugiperda.

    Science.gov (United States)

    Martínez de Castro, Diana L; García-Gómez, Blanca I; Gómez, Isabel; Bravo, Alejandra; Soberón, Mario

    2017-12-01

    Bacillus thuringiensis Cry toxins are currently used for pest control in transgenic crops but evolution of resistance by the insect pests threatens the use of this technology. The Cry1AbMod toxin was engineered to lack the alpha helix-1 of the parental Cry1Ab toxin and was shown to counter resistance to Cry1Ab and Cry1Ac toxins in different insect species including the fall armyworm Spodoptera frugiperda. In addition, Cry1AbMod showed enhanced toxicity to Cry1Ab-susceptible S. frugiperda populations. To gain insights into the mechanisms of this Cry1AbMod-enhanced toxicity, we isolated the Cry1AbMod toxin binding proteins from S. frugiperda brush border membrane vesicles (BBMV), which were identified by pull-down assay and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The LC-MS/MS results indicated that Cry1AbMod toxin could bind to four classes of aminopeptidase (N1, N3, N4 y N5) and actin, with the highest amino acid sequence coverage acquired for APN 1 and APN4. In addition to these proteins, we found other proteins not previously described as Cry toxin binding proteins. This is the first report that suggests the interaction between Cry1AbMod and APN in S. frugiperda. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Effect of gamma irradiation on nutritional components and Cry1Ab protein in the transgenic rice with a synthetic cry1Ab gene from Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Wu Dianxing; Ye Qingfu; Wang Zhonghua; Xia Yingwu

    2004-01-01

    The effects of gamma irradiation on the transgenic rice containing a synthetic cry1Ab gene from Bacillus thuringiensis were investigated. There was almost no difference in the content of the major nutritional components, i.e. crude protein, crude lipid, eight essential amino acids and total ash between the irradiated grains and the non-irradiated transgenic rice. However, the amounts of Cry1Ab protein and apparent amylose in the irradiated transgenic rice were reduced significantly by the doses higher than 200 Gy. In vivo observation showed that Cry1Ab protein contents also decreased in the fresh leaf tissues of survival seedlings after irradiation with 200 Gy or higher doses and showed inhibition of seedling growth. The results indicate that gamma irradiation might improve the quality of transgenic rice due to removal of the toxic Cry1Ab protein

  4. Fluorescence quenching dynamics and mechanism of cry1ab toxin from bacillus thuringiensis by different metal ions

    International Nuclear Information System (INIS)

    Zhou, X.; Zhang, J.

    2016-01-01

    The reaction dynamics of Cry1Ab toxin from Bacillus thuringiensis with sodium, calcium and lead ions was studied by fluorescence quenching technique. Gradual quenching was observed by titration of Cry1Ab toxin with metal ions (Na+, Ca/sup 2+/ or Pb/sup 2+/). The quenched strength of these ions in the descending order was: lead ion > calcium ion > sodium ion. The quenching equilibrium of Cry1Ab toxin by metal ions reached within 60 min, and the quenching dynamics of Cry1Ab toxin could be expressed by the Elovich model. The toxin concentration, pH and temperature had influence on the quenching dynamics. The interaction between Cry1Ab toxin and metal ions is based on static quenching mechanism. (author)

  5. 40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1Ab protein... PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.511 Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus...

  6. In Silico Modeling and Functional Interpretations of Cry1Ab15 Toxin from Bacillus thuringiensis BtB-Hm-16

    Directory of Open Access Journals (Sweden)

    Sudhanshu Kashyap

    2013-01-01

    Full Text Available The theoretical homology based structural model of Cry1Ab15 δ-endotoxin produced by Bacillus thuringiensis BtB-Hm-16 was predicted using the Cry1Aa template (resolution 2.25 Å. The Cry1Ab15 resembles the template structure by sharing a common three-domain extending conformation structure responsible for pore-forming and specificity determination. The novel structural differences found are the presence of β0 and α3, and the absence of α7b, β1a, α10a, α10b, β12, and α11a while α9 is located spatially downstream. Validation by SUPERPOSE and with the use of PROCHECK program showed folding of 98% of modeled residues in a favourable and stable orientation with a total energy Z-score of −6.56; the constructed model has an RMSD of only 1.15 Å. These increments of 3D structure information will be helpful in the design of domain swapping experiments aimed at improving toxicity and will help in elucidating the common mechanism of toxin action.

  7. Induction of Manduca sexta Larvae Caspases Expression in Midgut Cells by Bacillus thuringiensis Cry1Ab Toxin

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    Helena Porta

    2011-01-01

    Full Text Available Bacillus thuringiensis produces crystal toxins known as Cry that are highly selective against important agricultural and human health-related insect pests. Cry proteins are pore-forming toxins that interact with specific receptors in the midgut cell membrane of susceptible larvae making pores that cause osmotic shock, leading finally to insect death. In the case of pore-forming toxins that are specific to mammalian cells, death responses at low doses may induce apoptosis or pyroptosis, depending on the cell type. The death mechanism induced by Cry toxins in insect midgut cells is poorly understood. Here, we analyze the caspases expression by RT-PCR analysis, showing that the initial response of Manduca sexta midgut cells after low dose of Cry1Ab toxin administration involves a fast and transient accumulation of caspase-1 mRNA, suggesting that pyroptosis was activated by Cry1Ab toxin as an initial response but was repressed later. In contrast, caspase-3 mRNA requires a longer period of time of toxin exposure to be activated but presents a sustained activation, suggesting that apoptosis may be a cell death mechanism induced also at low dose of toxin.

  8. Nanobody-based electrochemical immunoassay for Bacillus thuringiensis Cry1Ab toxin by detecting the enzymatic formation of polyaniline

    International Nuclear Information System (INIS)

    Zhu, Min; Li, Guanghui; Li, Min; Zhou, Zikai; Liu, Hong; Lei, Hongtao; Shen, Yanfei; Wan, Yakun

    2015-01-01

    We describe an electrochemical immunoassay for the Cry1Ab toxin that is produced by Bacillus thuringiensis. It is making use of a nanobody (a heavy-chain only antibody) that was selected from an immune phage displayed library. A biotinylated primary nanobody and a HRP-conjugated secondary nanobody were applied in a sandwich immunoassay where horseradish peroxidase (HRP) is used to produce polyaniline (PANI) from aniline. PANI can be easily detected by differential pulse voltammetry at a working voltage as low as 40 mV (vs. Ag/AgCl) which makes the assay fairly selective. This immunoassay for Cry1Ab has an analytical range from 0.1 to 1000 ng∙mL -1 and a 0.07 ng∙mL -1 lower limit of detection. The average recoveries of the toxin from spiked samples are in the range from 102 to 114 %, with a relative standard deviation of <7.5 %. The results demonstrated that the assay represented an attractive alternative to existing immunoassays in enabling affordable, sensitive, robust and specific determination of this toxin. (author)

  9. Dominant negative phenotype of Bacillus thuringiensis Cry1Ab, Cry11Aa and Cry4Ba mutants suggest hetero-oligomer formation among different Cry toxins.

    NARCIS (Netherlands)

    Carmona, D.; Rodriguez-Almazan, C.; Munoz-Garay, C.; Portugal, L.; Perez, C.; Maagd, de R.A.; Bakker, P.; Soberon, M.; Bravo, A.

    2011-01-01

    Background - Bacillus thuringiensis Cry toxins are used worldwide in the control of different insect pests important in agriculture or in human health. The Cry proteins are pore-forming toxins that affect the midgut cell of target insects. It was shown that non-toxic Cry1Ab helix a-4 mutants had a

  10. Possibly similar genetic basis of resistance to Bacillus thuringiensis Cry1Ab protein in 3 resistant colonies of the sugarcane borer collected from Louisiana, USA.

    Science.gov (United States)

    Yang, Fei; Chen, Mao; Gowda, Anilkumar; Kerns, David L; Huang, Fangneng

    2018-04-01

    The sugarcane borer, Diatraea saccharalis (F.), is a major maize borer pest and a target of transgenic maize expressing Bacillus thuringiensis (Bt) proteins in South America and the mid-southern region of the United States. Evolution of resistance in target pest populations is a great threat to the long-term efficacy of Bt crops. In this study, we compared the genetic basis of resistance to Cry1Ab protein in 3 resistant colonies of sugarcane borer established from field populations in Louisiana, USA. Responses of larvae to the Cry1Ab protein for the parental and 10 other cross colonies were assayed in a diet-incorporated bioassay. All 3 resistant colonies were highly resistant to the Cry1Ab protein with a resistance ratio of >555.6 fold. No maternal effect or sex linkage was evident for the resistance in the 3 colonies; and the resistance was functionally nonrecessive at the Cry1Ab concentrations of ≤ 3.16 μg/g, but it became recessive at ≥10 μg/g. In an interstrain complementation test for allelism, the F 1 progeny from crosses between any 2 of the 3 resistant colonies exhibited the similar resistance levels as their parental colonies, indicating that the 3 colonies most likely shared a locus of Cry1Ab resistance. Results generated from this study should provide useful information in developing effective strategies for managing Bt resistance in the insect. © 2016 Institute of Zoology, Chinese Academy of Sciences.

  11. Changes in Gene Expression in the Larval Gut of Ostrinia nubilalis in Response to Bacillus thuringiensis Cry1Ab Protoxin Ingestion

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    Jianxiu Yao

    2014-04-01

    Full Text Available We developed a microarray based on 2895 unique transcripts assembled from 15,000 cDNA sequences from the European corn borer (Ostrinia nubilalis larval gut. This microarray was used to monitor gene expression in early third-instar larvae of Bacillus thuringiensis (Bt-susceptible O. nubilalis after 6 h feeding on diet, with or without the Bt Cry1Ab protoxin. We identified 174 transcripts, for which the expression was changed more than two-fold in the gut of the larvae fed Cry1Ab protoxin (p < 0.05, representing 80 down-regulated and 94 up-regulated transcripts. Among 174 differentially expressed transcripts, 13 transcripts putatively encode proteins that are potentially involved in Bt toxicity, and these transcripts include eight serine proteases, three aminopeptidases, one alkaline phosphatase, and one cadherin. The expressions of trypsin-like protease and three aminopeptidase transcripts were variable, but two potential Bt-binding proteins, alkaline phosphatase and cadherin were consistently up-regulated in larvae fed Cry1Ab protoxin. The significantly up and down-regulated transcripts may be involved in Cry1Ab toxicity by activation, degradation, toxin binding, and other related cellular responses. This study is a preliminary survey of Cry1Ab protoxin-induced transcriptional responses in O. nubilalis gut and our results are expected to help with further studies on Bt toxin-insect interactions at the molecular level.

  12. Different Effects of Bacillus thuringiensis Toxin Cry1Ab on Midgut Cell Transmembrane Potential of Mythimna separata and Agrotis ipsilon Larvae.

    Science.gov (United States)

    Wang, Yingying; Hu, Zhaonong; Wu, Wenjun

    2015-12-15

    Bacillus thuringiensis (Bt) Cry toxins from the Cry1A family demonstrate significantly different toxicities against members of the family Noctuidae for unknown reasons. In this study, membrane potential was measured and analyzed in freshly isolated midgut samples from Mythimna separata and Agrotis ipsilon larvae under oral administration and in vitro incubation with Bt toxin Cry1Ab to elucidate the mechanism of action for further control of these pests. Bioassay results showed that the larvae of M. separata achieved a LD50 of 258.84 ng/larva at 24 h after ingestion; M. separata larvae were at least eightfold more sensitive than A. ipsilon larvae to Cry1Ab. Force-feeding showed that the observed midgut apical-membrane potential (V(am)) of M. separata larvae was significantly depolarized from -82.9 ± 6.6 mV to -19.9 ± 7.2 mV at 8 h after ingestion of 1 μg activated Cry1Ab, whereas no obvious changes were detected in A. ipsilon larvae with dosage of 5 μg Cry1Ab. The activated Cry1Ab caused a distinct concentration-dependent depolarization of the apical membrane; V(am) was reduced by 50% after 14.7 ± 0.2, 9.8 ± 0.4, and 7.6 ± 0.6 min of treatment with 1, 5, and 10 μg/mL Cry1Ab, respectively. Cry1Ab showed a minimal effect on A. ipsilon larvae even at 20 μg/mL, and V(am) decreased by 26.3% ± 2.3% after 15 min. The concentrations of Cry1Ab displayed no significant effect on the basolateral side of the epithelium. The V(am) of A. ipsilon (-33.19 ± 6.29 mV, n = 51) was only half that of M. separata (-80.94 ± 6.95 mV, n = 75). The different degrees of sensitivity to Cry1Ab were speculatively associated with various habits, as well as the diverse physiological or biochemical characteristics of the midgut cell membranes.

  13. Different Effects of Bacillus thuringiensis Toxin Cry1Ab on Midgut Cell Transmembrane Potential of Mythimna separata and Agrotis ipsilon Larvae

    Directory of Open Access Journals (Sweden)

    Yingying Wang

    2015-12-01

    Full Text Available Bacillus thuringiensis (Bt Cry toxins from the Cry1A family demonstrate significantly different toxicities against members of the family Noctuidae for unknown reasons. In this study, membrane potential was measured and analyzed in freshly isolated midgut samples from Mythimna separata and Agrotis ipsilon larvae under oral administration and in vitro incubation with Bt toxin Cry1Ab to elucidate the mechanism of action for further control of these pests. Bioassay results showed that the larvae of M. separata achieved a LD50 of 258.84 ng/larva at 24 h after ingestion; M. separata larvae were at least eightfold more sensitive than A. ipsilon larvae to Cry1Ab. Force-feeding showed that the observed midgut apical-membrane potential (Vam of M. separata larvae was significantly depolarized from −82.9 ± 6.6 mV to −19.9 ± 7.2 mV at 8 h after ingestion of 1 μg activated Cry1Ab, whereas no obvious changes were detected in A. ipsilon larvae with dosage of 5 μg Cry1Ab. The activated Cry1Ab caused a distinct concentration-dependent depolarization of the apical membrane; Vam was reduced by 50% after 14.7 ± 0.2, 9.8 ± 0.4, and 7.6 ± 0.6 min of treatment with 1, 5, and 10 μg/mL Cry1Ab, respectively. Cry1Ab showed a minimal effect on A. ipsilon larvae even at 20 μg/mL, and Vam decreased by 26.3% ± 2.3% after 15 min. The concentrations of Cry1Ab displayed no significant effect on the basolateral side of the epithelium. The Vam of A. ipsilon (−33.19 ± 6.29 mV, n = 51 was only half that of M. separata (−80.94 ± 6.95 mV, n = 75. The different degrees of sensitivity to Cry1Ab were speculatively associated with various habits, as well as the diverse physiological or biochemical characteristics of the midgut cell membranes.

  14. Bacillus thuringiensis toxin (Cry1Ab) has no direct effect on larvae of the green lacewing Chrysoperla carnea (Stephens) (Neuroptera: Chrysopidae).

    Science.gov (United States)

    Romeis, Jörg; Dutton, Anna; Bigler, Franz

    2004-01-01

    Earlier studies have shown that larvae of the green lacewing predator Chrysoperla carnea are negatively affected when preying on lepidopteran larvae that had been fed with transgenic maize expressing the cry1Ab gene from Bacillus thuringiensis. To test whether the observed effects were directly caused by the Cry1Ab toxin, we have developed a bioassay which allows us to feed high concentrations of the toxin directly to the predator. The results of these feeding studies show no direct toxic effect of Cry1Ab on C. carnea larvae. The amount of toxin ingested by first instar C. carnea in the present study was found to be a factor 10,000 higher than the concentration ingested when feeding on Bt-reared lepidopteran larvae, a treatment that was previously shown to have a negative impact on the predator. In addition, feeding first instar C. carnea with the Cry1Ab toxin did not affect the utilisation of subsequently provided prey. Furthermore, the quality of the prey provided to first instars did not affect the sensitivity of second and third instar C. carnea to the Bt-toxin. The presented results strongly suggest that C. carnea larvae are not sensitive to Cry1Ab and that earlier reported negative effects of Bt-maize were prey-quality mediated rather than direct toxic effects. These results, together with the fact that lepidopteran larvae are not regarded as an important prey for C. carnea in the field, led us to conclude that transgenic maize expressing Cry1Ab poses a negligible risk for this predator.

  15. Computational tridimensional protein modeling of Cry1Ab19 toxin from Bacillus thuringiensis BtX-2.

    Science.gov (United States)

    Kashyap, Sudhanshu; Singh, B D; Amla, D V

    2012-06-01

    We report the computational structural simulation of the Cry1Ab19 toxin molecule from B. thuringiensis BtX-2 based on the structure of Cry1Aa1 deduced by x-ray diffraction. Validation results showed that 93.5% of modeled residues are folded in a favorable orientation with a total energy Z-score of -8.32, and the constructed model has an RMSD of only 1.13. The major differences in the presented model are longer loop lengths and shortened sheet components. The overall result supports the hierarchical three-domain structural hypothesis of Cry toxins and will help in better understanding the structural variation within the Cry toxin family along with facilitating the design of domain-swapping experiments aimed at improving the toxicity of native toxins.

  16. Effects of a diet containing genetically modified rice expressing the Cry1Ab/1Ac protein (Bacillus thuringiensis toxin) on broiler chickens.

    Science.gov (United States)

    Li, Zeyang; Gao, Yang; Zhang, Minhong; Feng, Jinghai; Xiong, Yandan

    2015-01-01

    The aim of this study was to evaluate the effect of feeding Bacillus thuringiensis (Bt) rice expressing the Cry1Ab/1Ac protein on broiler chicken. The genetically modified (GM) Bt rice was compared with the corresponding non-GM rice regarding performance of feeding groups, their health status, relative organ weights, biochemical serum parameters and occurrence of Cry1Ab/1Ac gene fragments. One hundred and eighty day-old Arbor Acres female broilers with the same health condition were randomly allocated to the two treatments (6 replicate cages with 15 broilers in each cage per treatment). They received diets containing GM rice (GM group) or its parental non-GM rice (non-GM group) at 52-57% of the air-dried diet for 42 days. The results show that the transgenic rice had a similar nutrient composition as the non-GM rice and had no adverse effects on chicken growth, biochemical serum parameters and necropsy during the 42-day feeding period. In birds fed the GM rice, no transgenic gene fragments were detected in the samples of blood, liver, kidneys, spleen, jejunum, ileum, duodenum and muscle tissue. In conclusion, the results suggest that Bt rice expressing Cry1Ab/1Ac protein has no adverse effects on broiler chicken. Therefore, it can be considered as safe and used as feed source for broiler chicken.

  17. Is the Cry1Ab protein from Bacillus thuringiensis (Bt) taken up by plants from soils previously planted with Bt corn and by carrot from hydroponic culture?

    Science.gov (United States)

    Icoz, I; Andow, D; Zwahlen, C; Stotzky, G

    2009-07-01

    The uptake of the insecticidal Cry1Ab protein from Bacillus thuringiensis (Bt) by various crops from soils on which Bt corn had previously grown was determined. In 2005, the Cry1Ab protein was detected by Western blot in tissues (leaves plus stems) of basil, carrot, kale, lettuce, okra, parsnip, radish, snap bean, and soybean but not in tissues of beet and spinach and was estimated by enzyme-linked immunosorbent assay (ELISA) to be 0.05 +/- 0.003 ng g(-1) of fresh plant tissue in basil, 0.02 +/- 0.014 ng g(-1) in okra, and 0.34 +/- 0.176 ng g(-1) in snap bean. However, the protein was not detected by ELISA in carrot, kale, lettuce, parsnip, radish, and soybean or in the soils by Western blot. In 2006, the Cry1Ab protein was detected by Western blot in tissues of basil, carrot, kale, radish, snap bean, and soybean from soils on which Bt corn had been grown the previous year and was estimated by ELISA to be 0.02 +/- 0.014 ng g(-1) of fresh plant tissue in basil, 0.19 +/- 0.060 ng g(-1) in carrot, 0.05 +/- 0.018 ng g(-1) in kale, 0.04 +/- 0.022 ng g(-1) in radish, 0.53 +/- 0.170 ng g(-1) in snap bean, and 0.15 +/- 0.071 ng g(-1) in soybean. The Cry1Ab protein was also detected by Western blot in tissues of basil, carrot, kale, radish, and snap bean but not of soybean grown in soil on which Bt corn had not been grown since 2002; the concentration was estimated by ELISA to be 0.03 +/- 0.021 ng g(-1) in basil, 0.02 +/- 0.008 ng g(-1) in carrot, 0.04 +/- 0.017 ng g(-1) in kale, 0.02 +/- 0.012 ng g(-1) in radish, 0.05 +/- 0.004 ng g(-1) in snap bean, and 0.09 +/- 0.015 ng g(-1) in soybean. The protein was detected by Western blot in 2006 in most soils on which Bt corn had or had not been grown since 2002. The Cry1Ab protein was detected by Western blot in leaves plus stems and in roots of carrot after 56 days of growth in sterile hydroponic culture to which purified Cry1Ab protein had been added and was estimated by ELISA to be 0.08 +/- 0.021 and 0.60 +/- 0.148 ng g(-1) of

  18. Layer-by-layer films containing peptides of the Cry1Ab16 toxin from Bacillus thuringiensis for potential biotechnological applications

    International Nuclear Information System (INIS)

    Plácido, Alexandra; Oliveira Farias, Emanuel Airton de; Marani, Mariela M.; Vasconcelos, Andreanne G.; Mafud, Ana C.; Mascarenhas, Yvonne P.; Eiras, Carla

    2016-01-01

    Cry1Ab16 is a toxin of crystalline insecticidal proteins that has been widely used in genetically modified organisms (GMOs) to gain resistance to pests. For the first time, in this study, peptides derived from the immunogenic Cry1Ab16 toxin (from Bacillus thuringiensis) were immobilized as layer-by-layer (LbL) films. Given the concern about food and environmental safety, a peptide with immunogenic potential, PcL342–354C, was selected for characterization of the electrochemical, optical, and morphological properties. The results obtained by cyclic voltammetry (CV) showed that the peptide have an irreversible oxidation process in electrolyte of 0.1 mol·L −1 potassium phosphate buffer (PBS) at pH 7.2. It was also observed that the electrochemical response of the peptide is governed mainly by charge transfer. In an attempt to maximize the electrochemical signal of peptide, it was intercalated with natural (agar, alginate and chitosan) or synthetic polymers (polyethylenimine (PEI) and poly(sodium 4-styrenesulfonate (PSS)). The presence of synthetic polymers on the film increased the electrochemical signal of PcL342–354C up to 100 times. Images by Atomic Force Microscopy (AFM) showed that the immobilized PcL342–354C formed self-assembled nanofibers with diameters ranging from 100 to 200 nm on the polymeric film. By UV–Visible spectroscopy (UV–Vis) it was observed that the ITO/PEI/PSS/PcL342–354C film grows linearly up to the fifth layer, thereafter tending to saturation. X-ray diffraction confirmed the presence on the films of crystalline ITO and amorphous polypeptide phases. In general, the ITO/PEI/PSS/PcL342–354C film characterization proved that this system is an excellent candidate for applications in electrochemical sensors and other biotechnological applications for GMOs and environmental indicators. - Highlights: • Peptides of the Cry1Ab16 toxin for potential biotechnological applications • Optimized LbL film deposition for synergic

  19. Layer-by-layer films containing peptides of the Cry1Ab16 toxin from Bacillus thuringiensis for potential biotechnological applications

    Energy Technology Data Exchange (ETDEWEB)

    Plácido, Alexandra [REQUIMTE/LAQV, Instituto Superior de Engenharia do Porto, Instituto Politécnico do Porto, Rua Dr. António Bernardino de Almeida, 431, 4200-072 Porto (Portugal); Oliveira Farias, Emanuel Airton de [Núcleo de Pesquisa em Biodiversidade e Biotecnologia, BIOTEC, Campus Ministro Reis Velloso, CMRV, Universidade Federal do Piauí, UFPI, 64202020 Parnaíba, Piaui (Brazil); Marani, Mariela M. [IPEEC-CENPAT-CONICET, Centro Nacional Patagónico, Consejo Nacional de Investigaciones Científicas y Técnicas, 9120 Puerto Madryn, Chubut (Argentina); Vasconcelos, Andreanne G. [Núcleo de Pesquisa em Biodiversidade e Biotecnologia, BIOTEC, Campus Ministro Reis Velloso, CMRV, Universidade Federal do Piauí, UFPI, 64202020 Parnaíba, Piaui (Brazil); Mafud, Ana C.; Mascarenhas, Yvonne P. [Instituto de Física de São Carlos, Universidade de São Paulo, USP, 13566-590 São Carlos, SP (Brazil); Eiras, Carla [Núcleo de Pesquisa em Biodiversidade e Biotecnologia, BIOTEC, Campus Ministro Reis Velloso, CMRV, Universidade Federal do Piauí, UFPI, 64202020 Parnaíba, Piaui (Brazil); Laboratório de Materiais Avançados, LIMAV, Engenharia de Materiais, Centro de Tecnologia, CT, Universidade Federal do Piauí, UFPI, 64049550 Teresina, Piaui (Brazil); and others

    2016-04-01

    Cry1Ab16 is a toxin of crystalline insecticidal proteins that has been widely used in genetically modified organisms (GMOs) to gain resistance to pests. For the first time, in this study, peptides derived from the immunogenic Cry1Ab16 toxin (from Bacillus thuringiensis) were immobilized as layer-by-layer (LbL) films. Given the concern about food and environmental safety, a peptide with immunogenic potential, PcL342–354C, was selected for characterization of the electrochemical, optical, and morphological properties. The results obtained by cyclic voltammetry (CV) showed that the peptide have an irreversible oxidation process in electrolyte of 0.1 mol·L{sup −1} potassium phosphate buffer (PBS) at pH 7.2. It was also observed that the electrochemical response of the peptide is governed mainly by charge transfer. In an attempt to maximize the electrochemical signal of peptide, it was intercalated with natural (agar, alginate and chitosan) or synthetic polymers (polyethylenimine (PEI) and poly(sodium 4-styrenesulfonate (PSS)). The presence of synthetic polymers on the film increased the electrochemical signal of PcL342–354C up to 100 times. Images by Atomic Force Microscopy (AFM) showed that the immobilized PcL342–354C formed self-assembled nanofibers with diameters ranging from 100 to 200 nm on the polymeric film. By UV–Visible spectroscopy (UV–Vis) it was observed that the ITO/PEI/PSS/PcL342–354C film grows linearly up to the fifth layer, thereafter tending to saturation. X-ray diffraction confirmed the presence on the films of crystalline ITO and amorphous polypeptide phases. In general, the ITO/PEI/PSS/PcL342–354C film characterization proved that this system is an excellent candidate for applications in electrochemical sensors and other biotechnological applications for GMOs and environmental indicators. - Highlights: • Peptides of the Cry1Ab16 toxin for potential biotechnological applications • Optimized LbL film deposition for synergic

  20. Construction of occluded recombinant baculoviruses containing the full-length cry1Ab and cry1Ac genes from Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    B.M. Ribeiro

    1998-06-01

    Full Text Available The administration of baculoviruses to insects for bioassay purposes is carried out, in most cases, by contamination of food surfaces with a known amount of occlusion bodies (OBs. Since per os infection is the natural route of infection, occluded recombinant viruses containing crystal protein genes (cry1Ab and cry1Ac from Bacillus thuringiensis were constructed for comparison with the baculovirus prototype Autographa californica nucleopolyhedrovirus (AcNPV. The transfer vector pAcUW2B was used for construction of occluded recombinant viruses. The transfer vector containing the crystal protein genes was cotransfected with linearized DNA from a non-occluded recombinant virus. The isolation of recombinant viruses was greatly facilitated by the reduction of background "wild type" virus and the increased proportion of recombinant viruses. Since the recombinant viruses containing full-length and truncated forms of the crystal protein genes did not seem to improve the pathogenicity of the recombinant viruses when compared with the wild type AcNPV, and in order to compare expression levels of the full-length crystal proteins produced by non-occluded and occluded recombinant viruses the full-length cry1Ab and cry1Ac genes were chosen for construction of occluded recombinant viruses. The recombinant viruses containing full-length and truncated forms of the crystal protein genes did not seem to improve its pathogenicity but the size of the larvae infected with the recombinant viruses was significantly smaller than that of larvae infected with the wild type virus.

  1. A 90-day safety study of genetically modified rice expressing Cry1Ab protein (Bacillus thuringiensis toxin) in Wistar rats

    DEFF Research Database (Denmark)

    Schrøder, Malene; Poulsen, Morten; Wilcks, Andrea

    2007-01-01

    , macroscopic and histopathological examinations were performed with only minor changes to report. The aim of the study was to use a known animal model in performance of safety assessment of a GM crop, in this case KMD1 rice. The results show no adverse or toxic effects of KMD1 rice when tested in the design...... used in this 90-day study. Nevertheless the experiences from this study lead to the overall conclusion that safety assessment for unintended effects of a GM crop cannot be done without additional test group(s).......An animal model for safety assessment of genetically modified foods was tested as part of the SAFOTEST project. In a 90-day feeding study on Wistar rats, the transgenic KMD1 rice expressing Cry1Ab protein was compared to its non-transgenic parental wild type, Xiushui 11. The KMD1 rice contained 15...

  2. Dominant negative mutants of Bacillus thuringiensis Cry1Ab toxin function as anti-toxins: demonstration of the role of oligomerization in toxicity.

    Directory of Open Access Journals (Sweden)

    Claudia Rodríguez-Almazán

    Full Text Available Bacillus thuringiensis Cry toxins, that are used worldwide in insect control, kill insects by a mechanism that depends on their ability to form oligomeric pores that insert into the insect-midgut cells. These toxins are being used worldwide in transgenic plants or spray to control insect pests in agriculture. However, a major concern has been the possible effects of these insecticidal proteins on non-target organisms mainly in ecosystems adjacent to agricultural fields.We isolated and characterized 11 non-toxic mutants of Cry1Ab toxin affected in different steps of the mechanism of action namely binding to receptors, oligomerization and pore-formation. These mutant toxins were analyzed for their capacity to block wild type toxin activity, presenting a dominant negative phenotype. The dominant negative phenotype was analyzed at two levels, in vivo by toxicity bioassays against susceptible Manduca sexta larvae and in vitro by pore formation activity in black lipid bilayers. We demonstrate that some mutations located in helix alpha-4 completely block the wild type toxin activity at sub-stoichiometric level confirming a dominant negative phenotype, thereby functioning as potent antitoxins.This is the first reported case of a Cry toxin dominant inhibitor. These data demonstrate that oligomerization is a fundamental step in Cry toxin action and represent a potential mechanism to protect special ecosystems from the possible effect of Cry toxins on non-target organisms.

  3. A Comparison of Soil microbial community structure, protozoa and nematodes in field plots of conventional and genetically modified maize expressing the Bacillus thuringiensis Cry1Ab toxin

    DEFF Research Database (Denmark)

    Griffiths, B. S.; Caul, S.; Thompson, J.

    2005-01-01

    Field trials were established at three European sites (Denmark, Eastern France, South-West France) of genetically modified maize (Zea mays L.) expressing the CryIAb Bacillus thuringiensis toxin (Bt), the near-isogenic non-Bt cultivar, another conventional maize cultivar and grass. Soil from Denmark...... was sampled at sowing (May) and harvest (October) over two years (2002, 2003); from E France at harvest 2002, sowing and harvest 2003; and from SW France at sowing and harvest 2003. Samples were analysed for microbial community structure (2003 samples only) by community-level physiological-profiling (CLPP...... there was a reduced protozoan population under Bt maize compared to non-Bt maize. Microbial community structure within the sites only varied with grass compared to maize, with one occurrence of CLPP varying between maize cultivars (Bt versus a conventional cultivar). An overall comparison of Bt versus non-Bt maize...

  4. Testing potential effects of maize expressing the Bacillus thuringiensis Cry1Ab endotoxin (Bt maize) on mycorrhizal fungal communities via DNA- and RNA-based pyrosequencing and molecular fingerprinting.

    Science.gov (United States)

    Verbruggen, Erik; Kuramae, Eiko E; Hillekens, Remy; de Hollander, Mattias; Kiers, E Toby; Röling, Wilfred F M; Kowalchuk, George A; van der Heijden, Marcel G A

    2012-10-01

    The cultivation of genetically modified (GM) crops has increased significantly over the last decades. However, concerns have been raised that some GM traits may negatively affect beneficial soil biota, such as arbuscular mycorrhizal fungi (AMF), potentially leading to alterations in soil functioning. Here, we test two maize varieties expressing the Bacillus thuringiensis Cry1Ab endotoxin (Bt maize) for their effects on soil AM fungal communities. We target both fungal DNA and RNA, which is new for AM fungi, and we use two strategies as an inclusive and robust way of detecting community differences: (i) 454 pyrosequencing using general fungal rRNA gene-directed primers and (ii) terminal restriction fragment length polymorphism (T-RFLP) profiling using AM fungus-specific markers. Potential GM-induced effects were compared to the normal natural variation of AM fungal communities across 15 different agricultural fields. AM fungi were found to be abundant in the experiment, accounting for 8% and 21% of total recovered DNA- and RNA-derived fungal sequences, respectively, after 104 days of plant growth. RNA- and DNA-based sequence analyses yielded most of the same AM fungal lineages. Our research yielded three major conclusions. First, no consistent differences were detected between AM fungal communities associated with GM plants and non-GM plants. Second, temporal variation in AMF community composition (between two measured time points) was bigger than GM trait-induced variation. Third, natural variation of AMF communities across 15 agricultural fields in The Netherlands, as well as within-field temporal variation, was much higher than GM-induced variation. In conclusion, we found no indication that Bt maize cultivation poses a risk for AMF.

  5. Single amino acid insertions in extracellular loop 2 of Bombyx mori ABCC2 disrupt its receptor function for Bacillus thuringiensis Cry1Ab and Cry1Ac but not Cry1Aa toxins.

    Science.gov (United States)

    Tanaka, Shiho; Miyamoto, Kazuhisa; Noda, Hiroaki; Endo, Haruka; Kikuta, Shingo; Sato, Ryoichi

    2016-04-01

    In a previous report, seven Cry1Ab-resistant strains were identified in the silkworm, Bombyx mori; these strains were shown to have a tyrosine insertion at position 234 in extracellular loop 2 of the ABC transporter C2 (BmABCC2). This insertion was confirmed to destroy the receptor function of BmABCC2 and confer the strains resistance against Cry1Ab and Cry1Ac. However, these strains were susceptible to Cry1Aa. In this report, we examined the mechanisms of the loss of receptor function of the transporter by expressing mutations in Sf9 cells. After replacement of one or two of the five amino acid residues in loop 2 of the susceptible BmABCC2 gene [BmABCC2_S] with alanine, cells still showed susceptibility, retaining the receptor function. Five mutants with single amino acid insertions at position 234 in BmABCC2 were also generated, resulting in loop 2 having six amino acids, which corresponds to replacing the tyrosine insertion in the resistant BmABCC2 gene [BmABCC2_R(+(234)Y)] with another amino acid. All five mutants exhibited loss of function against Cry1Ab and Cry1Ac. These results suggest that the amino acid sequence in loop 2 is less important than the loop size (five vs. six amino acids) or loop structure for Cry1Ab and Cry1Ac activity. Several domain-swapped mutant toxins were then generated among Cry1Aa, Cry1Ab, and Cry1Ac, which are composed of three domains. Swapped mutants containing domain II of Cry1Ab or Cry1Ac did not kill Sf9 cells expressing BmABCC2_R(+(234)Y), suggesting that domain II of the Cry toxin is related to the interaction with the receptor function of BmABCC2. This also suggests that different reactions against Bt-toxins in some B. mori strains, that is, Cry1Ab resistance or Cry1Aa susceptibility, are attributable to structural differences in domain II of Cry1A toxins. Copyright © 2016. Published by Elsevier Inc.

  6. Shared midgut binding sites for Cry1A.105, Cry1Aa, Cry1Ab, Cry1Ac and Cry1Fa proteins from Bacillus thuringiensis in two important corn pests, Ostrinia nubilalis and Spodoptera frugiperda.

    Science.gov (United States)

    Hernández-Rodríguez, Carmen Sara; Hernández-Martínez, Patricia; Van Rie, Jeroen; Escriche, Baltasar; Ferré, Juan

    2013-01-01

    First generation of insect-protected transgenic corn (Bt-corn) was based on the expression of Cry1Ab or Cry1Fa proteins. Currently, the trend is the combination of two or more genes expressing proteins that bind to different targets. In addition to broadening the spectrum of action, this strategy helps to delay the evolution of resistance in exposed insect populations. One of such examples is the combination of Cry1A.105 with Cry1Fa and Cry2Ab to control O. nubilalis and S. frugiperda. Cry1A.105 is a chimeric protein with domains I and II and the C-terminal half of the protein from Cry1Ac, and domain III almost identical to Cry1Fa. The aim of the present study was to determine whether the chimeric Cry1A.105 has shared binding sites either with Cry1A proteins, with Cry1Fa, or with both, in O. nubilalis and in S. frugiperda. Brush-border membrane vesicles (BBMV) from last instar larval midguts were used in competition binding assays with (125)I-labeled Cry1A.105, Cry1Ab, and Cry1Fa, and unlabeled Cry1A.105, Cry1Aa, Cry1Ab, Cry1Ac, Cry1Fa, Cry2Ab and Cry2Ae. The results showed that Cry1A.105, Cry1Ab, Cry1Ac and Cry1Fa competed with high affinity for the same binding sites in both insect species. However, Cry2Ab and Cry2Ae did not compete for the binding sites of Cry1 proteins. Therefore, according to our results, the development of cross-resistance among Cry1Ab/Ac, Cry1A.105, and Cry1Fa proteins is possible in these two insect species if the alteration of shared binding sites occurs. Conversely, cross-resistance between these proteins and Cry2A proteins is very unlikely in such case.

  7. Shared midgut binding sites for Cry1A.105, Cry1Aa, Cry1Ab, Cry1Ac and Cry1Fa proteins from Bacillus thuringiensis in two important corn pests, Ostrinia nubilalis and Spodoptera frugiperda.

    Directory of Open Access Journals (Sweden)

    Carmen Sara Hernández-Rodríguez

    Full Text Available First generation of insect-protected transgenic corn (Bt-corn was based on the expression of Cry1Ab or Cry1Fa proteins. Currently, the trend is the combination of two or more genes expressing proteins that bind to different targets. In addition to broadening the spectrum of action, this strategy helps to delay the evolution of resistance in exposed insect populations. One of such examples is the combination of Cry1A.105 with Cry1Fa and Cry2Ab to control O. nubilalis and S. frugiperda. Cry1A.105 is a chimeric protein with domains I and II and the C-terminal half of the protein from Cry1Ac, and domain III almost identical to Cry1Fa. The aim of the present study was to determine whether the chimeric Cry1A.105 has shared binding sites either with Cry1A proteins, with Cry1Fa, or with both, in O. nubilalis and in S. frugiperda. Brush-border membrane vesicles (BBMV from last instar larval midguts were used in competition binding assays with (125I-labeled Cry1A.105, Cry1Ab, and Cry1Fa, and unlabeled Cry1A.105, Cry1Aa, Cry1Ab, Cry1Ac, Cry1Fa, Cry2Ab and Cry2Ae. The results showed that Cry1A.105, Cry1Ab, Cry1Ac and Cry1Fa competed with high affinity for the same binding sites in both insect species. However, Cry2Ab and Cry2Ae did not compete for the binding sites of Cry1 proteins. Therefore, according to our results, the development of cross-resistance among Cry1Ab/Ac, Cry1A.105, and Cry1Fa proteins is possible in these two insect species if the alteration of shared binding sites occurs. Conversely, cross-resistance between these proteins and Cry2A proteins is very unlikely in such case.

  8. Characteristics of Cry1Ab Protein from Bioinsecticides and Insect Resistant GM Crops

    Directory of Open Access Journals (Sweden)

    Żmijewska Ewelina

    2016-06-01

    Full Text Available Biological insecticides are an effective method used in plant protection. One of the most widely used active substances in biological insecticides is Cry1Ab protein, which is toxic for lepidopteran insects. This protein is produced during bacterial sporulation by Bacillus thuringiensis. Other sources of Cry1Ab protein are genetically modified plants (GM with expression of cry1Ab gene. Cry1Ab protein in both bioinsecticides and GM plants is present in the form of protoxin, which requires activation by enzymatic treatment in the gut of susceptible insects. So far, Cry1Ab mode of action is not fully understood, but there are 3 main concepts describing it. Two of them assume that a toxic protein after binding to receptors in the insect gut penetrates into the cells, causing pore formation in the gut, which leads to the death of the sensitive insect. In the third model Cry1Ab toxic action is a result of toxin-induced chemical processes initiating a cell death pathway. This work describes the structure and mode of action of Cry1Ab protein, present in biological insecticides and genetically modified plants.

  9. Down regulation of a gene for cadherin, but not alkaline phosphatase, associated with Cry1Ab resistance in the sugarcane borer Diatraea saccharalis.

    Directory of Open Access Journals (Sweden)

    Yunlong Yang

    Full Text Available The sugarcane borer, Diatraea saccharalis, is a major target pest of transgenic corn expressing Bacillus thuringiensis (Bt proteins (i.e., Cry1Ab in South America and the mid-southern region of the United States. Evolution of insecticide resistance in such target pests is a major threat to the durability of transgenic Bt crops. Understanding the pests' resistance mechanisms will facilitate development of effective strategies for delaying or countering resistance. Alterations in expression of cadherin- and alkaline phosphatase (ALP have been associated with Bt resistance in several species of pest insects. In this study, neither the activity nor gene regulation of ALP was associated with Cry1Ab resistance in D. saccharalis. Total ALP enzymatic activity was similar between Cry1Ab-susceptible (Cry1Ab-SS and -resistant (Cry1Ab-RR strains of D. saccharalis. In addition, expression levels of three ALP genes were also similar between Cry1Ab-SS and -RR, and cDNA sequences did not differ between susceptible and resistant larvae. In contrast, altered expression of a midgut cadherin (DsCAD1 was associated with the Cry1Ab resistance. Whereas cDNA sequences of DsCAD1 were identical between the two strains, the transcript abundance of DsCAD1 was significantly lower in Cry1Ab-RR. To verify the involvement of DsCAD1 in susceptibility to Cry1Ab, RNA interference (RNAi was employed to knock-down DsCAD1 expression in the susceptible larvae. Down-regulation of DsCAD1 expression by RNAi was functionally correlated with a decrease in Cry1Ab susceptibility. These results suggest that down-regulation of DsCAD1 is associated with resistance to Cry1Ab in D. saccharalis.

  10. The fate and transport of the Cry1Ab protein in an agricultural field and laboratory aquatic microcosms.

    Science.gov (United States)

    Strain, Katherine E; Lydy, Michael J

    2015-08-01

    Genetically engineered crops expressing insecticidal crystalline proteins derived from Bacillus thuringiensis (Bt), were commercialized almost two decades ago as a means to manage agricultural pests. The Bt proteins are highly specific and only lethal upon ingestion, limiting the scope of toxicity to target insects. However, concern of exposure to non-target organisms and negative public perceptions regarding Bt crops has caused controversy surrounding their use. The objective of this research was to monitor the fate and transport of a Bt protein, Cry1Ab, in a large-scale agricultural field containing maize expressing the Cry1Ab protein and a non-Bt near isoline, and in aquatic microcosms. The highest environmental concentrations of the Cry1Ab protein were found in runoff water and sediment, up to 130ngL(-1) and 143ngg(-1) dry weight, respectively, with the Cry1Ab protein detected in both Bt and non-Bt maize fields. As surface runoff and residual crop debris can transport Bt proteins to waterways adjacent to agricultural fields, a series of laboratory experiments were conducted to determine the potential fate of the Cry1Ab protein under different conditions. The results showed that sediment type and temperature can influence the degradation of the Cry1Ab protein in an aquatic system and that the Cry1Ab protein can persist for up to two months. Although Cry1Ab protein concentrations measured in the field soil indicate little exposure to terrestrial organisms, the consistent input of Bt-contaminated runoff and crop debris into agricultural waterways is relevant to understanding potential consequences to aquatic species. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Transgenic Cry1Ab rice does not impact ecological fitness and predation of a generalist spider.

    Directory of Open Access Journals (Sweden)

    Jun-Ce Tian

    Full Text Available BACKGROUND: The commercial release of rice genetically engineered to express a Cry1Ab protein from Bacillus thuringiensis (Bt for control of Lepidoptera in China is a subject of debate. One major point of the debate has focused on the ecological safety of Bt rice on nontarget organisms, especially predators and parasitoids that help control populations of insect pests. METHODOLOGY/PRINCIPAL FINDINGS: A tritrophic bioassay was conducted to evaluate the potential impact of Cry1Ab-expressing rice on fitness parameters of a predaceous ground spider (Pardosa pseudoannulata (Bösenberg et Strand that had fed on Bt rice-fed brown planthopper (Nilaparvata lugens (Stål nymphs. Survival, development time and fecundity of this spider were not different when they were fed with Bt rice-fed or non-Bt rice-fed prey. Furthermore, ELISA and PCR gut assays, as well as a functional response trial, indicated that predation by P. pseudoannulata was not significantly different in Bt rice or non-Bt rice fields. CONCLUSIONS/SIGNIFICANCE: The transgenic Cry1Ab rice lines tested in this study had no adverse effects on the survival, developmental time and fecundity of P. pseudoannulata in the laboratory or on predation under field conditions. This suggests that this important predator would not be harmed if transgenic Cry1Ab rice were commercialized.

  12. Field evidence for the exposure of ground beetles to Cry1Ab from transgenic corn.

    Science.gov (United States)

    Zwahlen, Claudia; Andow, David A

    2005-01-01

    Non-target organisms associated with the soil might be adversely affected by exposure to the CrylAb protein from Bacillus thuringiensis (Bt) in transgenic corn (Zea mays L.). To check for such exposure, we used ELISA to test for Cry1Ab in ground beetles collected live from fields with Bt corn residues and Bt corn (Bt/Bt), Bt corn residues and non-Bt crops (Bt/non-Bt), or non-Bt corn residues and non-Bt crops (non-Bt/non-Bt). In fields with Bt corn residues (Bt/Bt and Bt/non-Bt), Cry1Ab was present in all seven species of ground beetles examined (Agonum placidum, Bembidion rupicola, Clivina impressefrons, Cyclotrachelus iowensis, Harpalus pensylvanicus, Poecilus chalcites, and Poecilus lucublandus). For the two most abundant species, P. chalcites and P. lucublandus, the proportion of beetles with Cry1Ab was significantly higher in Bt/Bt fields (0.50-1.0) and Bt/non-Bt fields (0.41-0.50) than in non-Bt/non-Bt fields (0.0). This is the first field evidence that some ground beetle species are exposed to Cry1Ab. The implications of exposure on the performance of these non-target organisms are unclear.

  13. VERTICAL MOVEMENT IN SOIL OF INSECTICIDAL CRY1AB PROTEIN FROM BACILLUS THURINGIENSIS. (R826107)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  14. Application of Cry1Ab/Ac Bt strip for screening of resistance for ...

    African Journals Online (AJOL)

    In this study, the efficacy of using Cry1Ab/Ac Bt strip for detecting Maruca resistant transgene in transgenic cowpea was systematically investigated for the first time through field derived progenies. The results show ... Keywords: Bacillus thuriengiensis, Cry1Ab/Ac Bt strips, transgenic cowpea, Maruca vitrata. African Journal of ...

  15. Screen of Bacillus thuringiensis toxins for transgenic rice to control Sesamia inferens and Chilo suppressalis

    Science.gov (United States)

    Transgenic rice to control stem borer damage is under development in China. To assess the potential of Bacillus thuringiensis (Bt) transgenes in stem borer control, the toxicity of five Bt protoxins (Cry1Aa, Cry1Ab, Cry1Ac, Cry1Ba and Cry1Ca) against two rice stem borers, Sesamia inferens (pink stem...

  16. Effects of ensiling of Bacillus thuringiensis (Bt) maize (MON810) on ...

    African Journals Online (AJOL)

    The study investigated the degradation of the Bt protein (Cry1Ab) in Bt maize during ensiling and chemical composition of the silage. Two laboratory studies were conducted at the University of Fort Hare. One Bacillus thuringiensis (Bt) maize cultivar (DKC80-12B) and its isoline (DKC80-10) in the 2008/2009 study and two Bt ...

  17. Aquatic degradation of Cry1Ab protein and decomposition dynamics of transgenic corn leaves under controlled conditions.

    Science.gov (United States)

    Böttger, Rita; Schaller, Jörg; Lintow, Sven; Gert Dudel, E

    2015-03-01

    The increasing cultivation of genetically modified corn plants (Zea mays) during the last decades is suggested as a potential risk to the environment. One of these genetically modified variety expressed the insecticidal Cry1Ab protein originating from Bacillus thuringiensis (Bt), resulting in resistance against Ostrinia nubilalis, the European corn borer. Transgenic litter material is extensively studied regarding the decomposition in soils. However, only a few field studies analyzed the fate of the Cry1Ab protein and the impact of green and senescent leaf litter from corn on the decomposition rate and related ecosystem functions in aquatic environments. Consequently, a microbial litter decomposition experiment was conducted under controlled semi-natural conditions in batch culture using two maize varieties: one variety with Cry1Ab and another one with the appertaining Iso-line as control treatment. The results showed no significant differences between the treatment with Cry1Ab and the Iso-line regarding loss of total mass in dry weight of 43% for Iso-line and 45% for Bt-corn litter, lignin content increased to 137.5% (Iso-line) and 115.7% (Bt-corn), and phenol loss decreased by 53.6% (Iso-line), 62.2% (Bt-corn) during three weeks of the experiment. At the end of the experiment Cry1Ab protein was still detected with 6% of the initial concentration. A slightly but significant lower cellulose content was found for the Cry1Ab treatment compared to the Iso-line litter at the end of the experiment. The significant higher total protein (25%) and nitrogen (25%) content in Bt corn, most likely due to the additionally expression of the transgenic protein, may increase the microbial cellulose degradation and decrease microbial lignin degradation. In conclusion a relevant year by year input of protein and therefore nitrogen rich Bt corn litter into aquatic environments may affect the balanced nutrient turnover in aquatic ecosystems. Copyright © 2014 Elsevier Inc. All rights

  18. INSECTICIDAL TOXIN FROM BACILLUS THURINGIENSIS IS RELEASED FROM ROOTS OF TRANSGENIC BT CORN IN VITRO AND IN SITU. (R826107)

    Science.gov (United States)

    AbstractThe insecticidal toxin encoded by the cry1Ab gene from Bacillus thuringiensis was released in root exudates from transgenic Bt corn during 40 days of growth in soil amended to 0, 3, 6, 9, or 12% (v/v) with montmorillonite or kaolinite in a...

  19. Bacillus thuringiensis

    Science.gov (United States)

    Hollensteiner, Jacqueline; Wemheuer, Franziska; Harting, Rebekka; Kolarzyk, Anna M; Diaz Valerio, Stefani M; Poehlein, Anja; Brzuszkiewicz, Elzbieta B; Nesemann, Kai; Braus-Stromeyer, Susanna A; Braus, Gerhard H; Daniel, Rolf; Liesegang, Heiko

    2016-01-01

    Verticillium wilt causes severe yield losses in a broad range of economically important crops worldwide. As many soil fumigants have a severe environmental impact, new biocontrol strategies are needed. Members of the genus Bacillus are known as plant growth-promoting bacteria (PGPB) as well as biocontrol agents of pests and diseases. In this study, we isolated 267 Bacillus strains from root-associated soil of field-grown tomato plants. We evaluated the antifungal potential of 20 phenotypically diverse strains according to their antagonistic activity against the two phytopathogenic fungi Verticillium dahliae and Verticillium longisporum . In addition, the 20 strains were sequenced and phylogenetically characterized by multi-locus sequence typing (MLST) resulting in 7 different Bacillus thuringiensis and 13 Bacillus weihenstephanensis strains. All B. thuringiensis isolates inhibited in vitro the tomato pathogen V. dahliae JR2, but had only low efficacy against the tomato-foreign pathogen V. longisporum 43. All B. weihenstephanensis isolates exhibited no fungicidal activity whereas three B. weihenstephanensis isolates showed antagonistic effects on both phytopathogens. These strains had a rhizoid colony morphology, which has not been described for B. weihenstephanensis strains previously. Genome analysis of all isolates revealed putative genes encoding fungicidal substances and resulted in identification of 304 secondary metabolite gene clusters including 101 non-ribosomal polypeptide synthetases and 203 ribosomal-synthesized and post-translationally modified peptides. All genomes encoded genes for the synthesis of the antifungal siderophore bacillibactin. In the genome of one B. thuringiensis strain, a gene cluster for zwittermicin A was detected. Isolates which either exhibited an inhibitory or an interfering effect on the growth of the phytopathogens carried one or two genes encoding putative mycolitic chitinases, which might contribute to antifungal activities

  20. Production of the {sup 14}C-labeled insecticidal protein Cry1Ab for soil metabolic studies using a recombinant Escherichia coli in small-scale batch fermentations

    Energy Technology Data Exchange (ETDEWEB)

    Valldor, Petra; Miethling-Graff, Rona; Dockhorn, Susanne; Martens, Rainer; Tebbe, Christoph C. [Federal Research Institute for Rural Areas, Forestry and Fisheries, Braunschweig (Germany). Thuenen Institute (vTI) for Biodiversity

    2012-10-15

    Insecticidal Cry proteins naturally produced by Bacillus thuringiensis are a major recombinant trait expressed by genetically modified crops. They are released into the soil during and after cropping. The objective of this study was to produce {sup 14}C-labeled Cry1Ab proteins for soil metabolic studies in scope of their environmental risk assessment. Cry1Ab was synthesized as a protoxin by Escherichia coli HB101 pMP in 200-mL liquid batch culture fermentations and purified from inclusion bodies after trypsin digestion. For cultivation, U-{sup 14}C-glycerol was the main carbon source. Inclusion bodies were smaller and Cry1Ab yield was lower when the initial amount of total organic carbon in the cultivation broth was below 6.4 mg C L{sup -1}. Concentrations of 12.6 g {sup 14}C-labeled glycerol L{sup -1} (1 % v/v) resulted in the production of 17.1 mg {sup 14}C-Cry1Ab L{sup -1} cultivation medium. {sup 14}C mass balances showed that approx. 50 % of the label was lost by respiration and 20 % remained in the growth media, while the residual activity was associated with biomass. Depending on the production batch, 0.01 to 0.05 % of the total {sup 14}C originated from Cry1Ab. In the presence of 2.04 MBq {sup 14}C-labeled carbon sources, a specific activity of up to 268 Bq mg{sup -1} {sup 14}C-Cry1Ab was obtained. A more than threefold higher specific activity was achieved with 4.63 MBq and an extended cultivation period of 144 h. This study demonstrates that {sup 14}C-labeled Cry1Ab can be obtained from batch fermentations with E. coli in the presence of a simple {sup 14}C-labeled carbon source. It also provides a general strategy to produce {sup 14}C-labeled proteins useful for soil metabolic studies. (orig.)

  1. Activity of Bacillus thuringiensis D(delta)-endotoxins against codling moth (Cydia pomonella L.) larvae

    NARCIS (Netherlands)

    Boncheva, R.; Dukiandjiev, S.; Minkov, I.; Maagd, de R.A.; Naimov, S.

    2006-01-01

    Solubilized protoxins of nine Cry1 and one hybrid Cry1 ¿-endotoxin from Bacillus thuringiensis were tested for their activity against larvae of the codling moth (Cydia pomonella L). Cry1Da was the most toxic, followed by Cry1Ab, Cry1Ba, and Cry1Ac, while Cry1Aa, Cry1Fa, Cry1Ia, and SN19 were still

  2. Bt Jute Expressing Fused δ-Endotoxin Cry1Ab/Ac for Resistance to Lepidopteran Pests

    Science.gov (United States)

    Majumder, Shuvobrata; Sarkar, Chirabrata; Saha, Prosanta; Gotyal, Bheemanna S.; Satpathy, Subrata; Datta, Karabi; Datta, Swapan K.

    2018-01-01

    Jute (Corchorus sp.) is naturally occurring, biodegradable, lignocellulosic-long, silky, golden shiny fiber producing plant that has great demands globally. Paper and textile industries are interested in jute because of the easy availability, non-toxicity and high yield of cellulosic biomass produced per acre in cultivation. Jute is the major and most industrially used bast fiber-producing crop in the world and it needs protection from insect pest infestation that decreases its yield and quality. Single locus integration of the synthetically fused cry1Ab/Ac gene of Bacillus thuringiensis (Bt) in Corchorus capsularis (JRC 321) by Agrobacterium tumefaciens-mediated shoot tip transformation provided 5 potent Bt jute lines BT1, BT2, BT4, BT7 and BT8. These lines consistently expressed the Cry1Ab/Ac endotoxin ranging from 0.16 to 0.35 ng/mg of leaf, in the following generations (analyzed upto T4). The effect of Cry1Ab/Ac endotoxin was studied against 3 major Lepidopteran pests of jute- semilooper (Anomis sabulifera Guenee), hairy caterpillar (Spilarctia obliqua Walker) and indigo caterpillar (Spodoptera exigua Hubner) by detached leaf and whole plant insect bioassay on greenhouse-grown transgenic plants. Results confirm that larvae feeding on transgenic plants had lower food consumption, body size, body weight and dry weight of excreta compared to non-transgenic controls. Insect mortality range among transgenic feeders was 66–100% for semilooper and hairy caterpillar and 87.50% for indigo caterpillar. Apart from insect resistance, the transgenic plants were at par with control plants in terms of agronomic parameters and fiber quality. Hence, these Bt jutes in the field would survive Lepidopteran pest infestation, minimize harmful pesticide usage and yield good quality fiber. PMID:29354143

  3. Bt Jute Expressing Fused δ-Endotoxin Cry1Ab/Ac for Resistance to Lepidopteran Pests

    Directory of Open Access Journals (Sweden)

    Shuvobrata Majumder

    2018-01-01

    Full Text Available Jute (Corchorus sp. is naturally occurring, biodegradable, lignocellulosic-long, silky, golden shiny fiber producing plant that has great demands globally. Paper and textile industries are interested in jute because of the easy availability, non-toxicity and high yield of cellulosic biomass produced per acre in cultivation. Jute is the major and most industrially used bast fiber-producing crop in the world and it needs protection from insect pest infestation that decreases its yield and quality. Single locus integration of the synthetically fused cry1Ab/Ac gene of Bacillus thuringiensis (Bt in Corchorus capsularis (JRC 321 by Agrobacterium tumefaciens-mediated shoot tip transformation provided 5 potent Bt jute lines BT1, BT2, BT4, BT7 and BT8. These lines consistently expressed the Cry1Ab/Ac endotoxin ranging from 0.16 to 0.35 ng/mg of leaf, in the following generations (analyzed upto T4. The effect of Cry1Ab/Ac endotoxin was studied against 3 major Lepidopteran pests of jute- semilooper (Anomis sabulifera Guenee, hairy caterpillar (Spilarctia obliqua Walker and indigo caterpillar (Spodoptera exigua Hubner by detached leaf and whole plant insect bioassay on greenhouse-grown transgenic plants. Results confirm that larvae feeding on transgenic plants had lower food consumption, body size, body weight and dry weight of excreta compared to non-transgenic controls. Insect mortality range among transgenic feeders was 66–100% for semilooper and hairy caterpillar and 87.50% for indigo caterpillar. Apart from insect resistance, the transgenic plants were at par with control plants in terms of agronomic parameters and fiber quality. Hence, these Bt jutes in the field would survive Lepidopteran pest infestation, minimize harmful pesticide usage and yield good quality fiber.

  4. Expression of Cry1Ab and Cry2Ab by a polycistronic transgene with a self-cleavage peptide in rice.

    Directory of Open Access Journals (Sweden)

    Qichao Zhao

    Full Text Available Insect resistance to Bacillus thuringiensis (Bt crystal protein is a major threat to the long-term use of transgenic Bt crops. Gene stacking is a readily deployable strategy to delay the development of insect resistance while it may also broaden insecticidal spectrum. Here, we report the creation of transgenic rice expressing discrete Cry1Ab and Cry2Ab simultaneously from a single expression cassette using 2A self-cleaving peptides, which are autonomous elements from virus guiding the polycistronic viral gene expression in eukaryotes. The synthetic coding sequences of Cry1Ab and Cry2Ab, linked by the coding sequence of a 2A peptide from either foot and mouth disease virus or porcine teschovirus-1, regardless of order, were all expressed as discrete Cry1Ab and Cry2Ab at high levels in the transgenic rice. Insect bioassays demonstrated that the transgenic plants were highly resistant to lepidopteran pests. This study suggested that 2A peptide can be utilized to express multiple Bt genes at high levels in transgenic crops.

  5. Ingestion of Bt corn pollen containing Cry1Ab/2Aj or Cry1Ac does not harm Propylea japonica larvae.

    Science.gov (United States)

    Liu, Yanmin; Liu, Qingsong; Wang, Yanan; Chen, Xiuping; Song, Xinyuan; Romeis, Jörg; Li, Yunhe; Peng, Yufa

    2016-03-23

    Propylea japonica (Thunberg) (Coleoptera: Coccinellidae) is a prevalent pollen consumer in corn fields and is therefore exposed to insecticidal proteins contained in the pollen of insect-resistant transgenic corn cultivars expressing Cry proteins derived from Bacillus thuringiensis (Bt). In the present study, the potential effect of Cry1Ab/2Aj- or Cry1Ac-containing transgenic Bt corn pollen on the fitness of P. japonica larvae was evaluated. The results show that the larval developmental time was significantly shorter when P. japonica larvae were fed pollen from Bt corn cultivars rather than control pollen but that pupation rate, eclosion rate, and adult fresh weight were not significantly affected. In the feeding experiments, the stability of the Cry proteins in the food sources was confirmed. When Bt corn pollen passed through the gut of P. japonica, 23% of Cry1Ab/2Aj was digested. The results demonstrate that consumption of Bt corn pollen containing Cry1Ab/2Aj or Cry1Ac has no detrimental effect on P. japonica larvae; the shortened developmental time of larvae that consumed these proteins was likely attributable to unknown differences in the nutritional composition between the Bt-transgenic and control corn pollen.

  6. Effects of Cry1Ab Transgenic Maize on Lifecycle and Biomarker Responses of the Earthworm, Eisenia Andrei

    Directory of Open Access Journals (Sweden)

    Mark Maboeta

    2012-12-01

    Full Text Available A 28-day study was conducted to determine the effects of the Bacillus thuringiensis Cry1Ab toxin on the earthworm Eisenia andrei. Previously, investigations have been limited to life-cycle level effects of this protein on earthworms, and mostly on E. fetida. In this study several endpoints were compared which included biomass changes, cocoon production, hatching success, a cellular metal-stress biomarker (Neutral Red Retention Time; NRRT and potential genotoxic effects in terms of Randomly Amplified Polymorphic DNA sequences (RAPDs. NRRT results indicated no differences between treatments (p > 0.36, and NRRT remained the same for both treatments at different times during the experiment (p = 0.18. Likewise, no significant differences were found for cocoon production (p = 0.32 or hatching success (p = 0.29. Conversely, biomass data indicated a significant difference between the control treatment and the Bt treatment from the second week onwards (p < 0.001, with the Bt treatment losing significantly more weight than the isoline treatment. Possible confounding factors were identified that might have affected the differences in weight loss between groups. From the RAPD profiles no conclusive data were obtained that could link observed genetic variation to exposure of E. andrei to Cry1Ab proteins produced by Bt maize.

  7. Bacillus thuringiensis Cry1A toxin-binding glycoconjugates present on the brush border membrane and in the peritrophic membrane of the Douglas-fir tussock moth are peritrophins

    Science.gov (United States)

    Algimantas P. Valaitis; John D. Podgwaite

    2013-01-01

    Bacillus thuringiensis (Bt) Cry1A toxin-binding sites in the Douglas fir tussock moth (DFTM) larval gut were localized using immunofluorescence microscopy. Cry1Aa, Cry1Ab and Cry1Ac all bound strongly to the DFTM peritrophic membrane (PM); weaker binding of the Cry1A toxins was observed along the apical brush border of the midgut epithelium....

  8. CRY 1AB trangenic cowpea obtained by nodal electroporation ...

    African Journals Online (AJOL)

    Electroporation-mediated genetic transformation was used to introduce Cry 1 Ab insecticidal gene into cowpea. Nodal buds were electroporated in planta with a plasmid carrying the Cry 1Ab and antibiotic resistance npt II genes driven by a 35S CaMV promoter. T1 seeds derived from electroporated branches were selected ...

  9. Inhibition of Helicoverpa zea (Lepidoptera: Noctuidae) Growth by Transgenic Corn Expressing Bt Toxins and Development of Resistance to Cry1Ab.

    Science.gov (United States)

    Reisig, Dominic D; Reay-Jones, Francis P F

    2015-08-01

    Transgenic corn, Zea mays L., that expresses the Bacillus thuringiensis (Bt) toxin Cry1Ab is only moderately toxic to Helicoverpa zea (Boddie) and has been planted commercially since 1996. Growth and development of H. zea was monitored to determine potential changes in susceptibility to this toxin over time. Small plots of corn hybrids expressing Cry1F, Cry1F × Cry1Ab, Cry1Ab × Cry3Bb1, Cry1A.105 × Cry2Ab2 × Cry3Bb1, Cry1A.105 × Cry2Ab2, and Vip3Aa20 × Cry1Ab × mCry3A were planted in both 2012 and 2013 inNorth and South Carolina with paired non-Bt hybrids from the same genetic background. H. zea larvae were sampled on three time periods from ears and the following factors were measured: kernel area injured (cm(2)) by H. zea larvae, larval number per ear, larval weight, larval length, and larval head width. Pupae were sampled on a single time period and the following factors recorded: number per ear, weight, time to eclosion, and the number that eclosed. There was no reduction in larval weight, number of insect entering the pupal stadium, pupal weight, time to eclosion, and number of pupae able to successfully eclose to adulthood in the hybrid expressing Cry1Ab compared with a non-Bt paired hybrid. As Cry1Ab affected these in 1996, H. zea may be developing resistance to Cry1Ab in corn, although these results are not comprehensive, given the limited sampling period, size, and geography. We also found that the negative impacts on larval growth and development were greater in corn hybrids with pyramided traits compared with single traits. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  10. Consumption of Bt maize pollen expressing Cry1Ab or Cry3Bb1 does not harm adult green Lacewings, Chrysoperla carnea (Neuroptera: Chrysopidae.

    Directory of Open Access Journals (Sweden)

    Yunhe Li

    Full Text Available Adults of the common green lacewing, Chrysoperla carnea (Stephens (Neuroptera: Chrysopidae, are prevalent pollen-consumers in maize fields. They are therefore exposed to insecticidal proteins expressed in the pollen of insect-resistant, genetically engineered maize varieties expressing Cry proteins derived from Bacillus thuringiensis (Bt. Laboratory experiments were conducted to evaluate the impact of Cry3Bb1 or Cry1Ab-expressing transgenic maize (MON 88017, Event Bt176 pollen on fitness parameters of adult C. carnea. Adults were fed pollen from Bt maize varieties or their corresponding near isolines together with sucrose solution for 28 days. Survival, pre-oviposition period, fecundity, fertility and dry weight were not different between Bt or non-Bt maize pollen treatments. In order to ensure that adults of C. carnea are not sensitive to the tested toxins independent from the plant background and to add certainty to the hazard assessment, adult C. carnea were fed with artificial diet containing purified Cry3Bb1 or Cry1Ab at about a 10 times higher concentration than in maize pollen. Artificial diet containing Galanthus nivalis agglutinin (GNA was included as a positive control. No differences were found in any life-table parameter between Cry protein containing diet treatments and control diet. However, the pre-oviposition period, daily and total fecundity and dry weight of C. carnea were significantly negatively affected by GNA-feeding. In both feeding assays, the stability and bioactivity of Cry proteins in the food sources as well as the uptake by C. carnea was confirmed. These results show that adults of C. carnea are not affected by Bt maize pollen and are not sensitive to Cry1Ab and Cry3Bb1 at concentrations exceeding the levels in pollen. Consequently, Bt maize pollen consumption will pose a negligible risk to adult C. carnea.

  11. Consumption of Bt maize pollen expressing Cry1Ab or Cry3Bb1 does not harm adult green Lacewings, Chrysoperla carnea (Neuroptera: Chrysopidae).

    Science.gov (United States)

    Li, Yunhe; Meissle, Michael; Romeis, Jörg

    2008-08-06

    Adults of the common green lacewing, Chrysoperla carnea (Stephens) (Neuroptera: Chrysopidae), are prevalent pollen-consumers in maize fields. They are therefore exposed to insecticidal proteins expressed in the pollen of insect-resistant, genetically engineered maize varieties expressing Cry proteins derived from Bacillus thuringiensis (Bt). Laboratory experiments were conducted to evaluate the impact of Cry3Bb1 or Cry1Ab-expressing transgenic maize (MON 88017, Event Bt176) pollen on fitness parameters of adult C. carnea. Adults were fed pollen from Bt maize varieties or their corresponding near isolines together with sucrose solution for 28 days. Survival, pre-oviposition period, fecundity, fertility and dry weight were not different between Bt or non-Bt maize pollen treatments. In order to ensure that adults of C. carnea are not sensitive to the tested toxins independent from the plant background and to add certainty to the hazard assessment, adult C. carnea were fed with artificial diet containing purified Cry3Bb1 or Cry1Ab at about a 10 times higher concentration than in maize pollen. Artificial diet containing Galanthus nivalis agglutinin (GNA) was included as a positive control. No differences were found in any life-table parameter between Cry protein containing diet treatments and control diet. However, the pre-oviposition period, daily and total fecundity and dry weight of C. carnea were significantly negatively affected by GNA-feeding. In both feeding assays, the stability and bioactivity of Cry proteins in the food sources as well as the uptake by C. carnea was confirmed. These results show that adults of C. carnea are not affected by Bt maize pollen and are not sensitive to Cry1Ab and Cry3Bb1 at concentrations exceeding the levels in pollen. Consequently, Bt maize pollen consumption will pose a negligible risk to adult C. carnea.

  12. Yield Losses in Transgenic Cry1Ab and Non-Bt Corn as Assessed Using a Crop-Life-Table Approach.

    Science.gov (United States)

    Silva, Gerson A; Picanço, Marcelo C; Ferreira, Lino R; Ferreira, Dalton O; Farias, Elizeu S; Souza, Thadeu C; Rodrigues-Silva, Nilson; Pereira, Eliseu José G

    2018-02-09

    In this study, we constructed crop life tables for Bacillus thuringiensis Berliner (Bt) Cry1Ab and non-Bt corn hybrids, in which yield-loss factors and abundance of predaceous arthropods were recorded during 2 yr at two locations. Corn kernel/grain was the yield component that had the heaviest losses and that determined the overall yield loss in the corn hybrids across years and locations. Yield losses in both corn hybrids were primarily caused by kernel-destroying insects. Helicoverpa zea (Boddie) and Spodoptera frugiperda (Smith) (Lepidoptera: Noctuidae) were the key loss factors at one location, while at the other, the key loss factor was the silk fly larvae, Euxesta spp. (Diptera: Ulidiidae). Although the realized yield of corn grains was not different (P > 0.05) between Cry1Ab and non-Bt corn hybrids, the Bt corn hybrid reduced (P corn fields. Various species of natural enemies were recorded, particularly the earwig Doru luteipes (Scudder) (Dermaptera: Forficulidae), which was the most abundant and frequent predaceous insect. These results indicate that integration of pest management practices should be pursued to effectively minimize losses by kernel-destroying insects during corn reproductive stages when growing non-Bt or certain low-dose Bt corn cultivars for fall armyworm and corn earworm, such as those producing Cry1Ab or other Cry toxins. © The Author(s) 2018. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  13. Characterization of cDNAs encoding serine proteases and their transcriptional responses to Cry1Ab protoxin in the gut of Ostrinia nubilalis larvae.

    Directory of Open Access Journals (Sweden)

    Jianxiu Yao

    Full Text Available Serine proteases, such as trypsin and chymotrypsin, are the primary digestive enzymes in lepidopteran larvae, and are also involved in Bacillus thuringiensis (Bt protoxin activation and protoxin/toxin degradation. We isolated and sequenced 34 cDNAs putatively encoding trypsins, chymotrypsins and their homologs from the European corn borer (Ostrinia nubilalis larval gut. Our analyses of the cDNA-deduced amino acid sequences indicated that 12 were putative trypsins, 12 were putative chymotrypsins, and the remaining 10 were trypsin and chymotrypsin homologs that lack one or more conserved residues of typical trypsins and chymotrypsins. Reverse transcription PCR analysis indicated that all genes were highly expressed in gut tissues, but one group of phylogenetically-related trypsin genes, OnTry-G2, was highly expressed in larval foregut and midgut, whereas another group, OnTry-G3, was highly expressed in the midgut and hindgut. Real-time quantitative PCR analysis indicated that several trypsin genes (OnTry5 and OnTry6 were significantly up-regulated in the gut of third-instar larvae after feeding on Cry1Ab protoxin from 2 to 24 h, whereas one trypsin (OnTry2 was down-regulated at all time points. Four chymotrypsin and chymotrypsin homolog genes (OnCTP2, OnCTP5, OnCTP12 and OnCTP13 were up-regulated at least 2-fold in the gut of the larvae after feeding on Cry1Ab protoxin for 24 h. Our data represent the first in-depth study of gut transcripts encoding expanded families of protease genes in O. nubilalis larvae and demonstrate differential expression of protease genes that may be related to Cry1Ab intoxication and/or resistance.

  14. Novel Vip3A Bacillus thuringiensis (Bt) maize approaches high-dose efficacy against Helicoverpa zea (Lepidoptera: Noctuidae) under field conditions: Implications for resistance management.

    Science.gov (United States)

    Burkness, Eric C; Dively, Galen; Patton, Terry; Morey, Amy C; Hutchison, William D

    2010-01-01

    Sweet corn, Zea mays L., transformed to express a novel vegetative insecticidal protein, Vip3A (event MIR162, Syngenta Seeds, Inc..), produced by the bacterium, Bacillus thuringiensis (Bt), was evaluated over four field seasons in Maryland and two field seasons in Minnesota for efficacy against the corn earworm, Helicoverpa zea (Boddie). Hybrids expressing the Vip3A protein and pyramided in hybrids also expressing the Cry1Ab Bt protein (event Bt11, ATTRIBUTE(®), Syngenta Seeds, Inc.) were compared to hybrids expressing only Cry1Ab or to genetically similar non-Bt hybrids each year. In addition to H. zea efficacy, results for Ostrinia nubilalis (Hübner) and Spodoptera frugiperda (J.E. Smith) are presented. Over all years and locations, the non-Bt hybrids, without insecticide protection, averaged between 43 and 100% ears infested with a range of 0.24 to 1.74 H. zea larvae per ear. By comparison, in the pyramided Vip3A x Cry1Ab hybrids, no larvae were found and only minimal kernel damage (likely due to other insect pests) was recorded. Hybrids expressing only Cry1Ab incurred a moderate level of H. zea feeding damage, with surviving larvae mostly limited to the first or second instar as a result of previously documented growth inhibition from Cry1Ab. These results suggest that the Vip3A protein, pyramided with Cry1Ab, appears to provide the first "high-dose" under field conditions and will be valuable for ongoing resistance management.

  15. Proteasas digestivas de tipo tripsina del taladro del maíz, "Sesamia nonagrioides" (Lepidoptera, Noctuidae), caracterización e interacción con la proteína insecticida Cry1Ab

    OpenAIRE

    Díaz Mendoza, María Mercedes

    2006-01-01

    Una de las plagas de insectos con mayor incidencia económica sobre el cultivo de maíz en el área Mediterránea es el taladro del maíz, Sesamia nonagrioides (Lepidoptera, Noctuidae). El maíz transgénico resistente a taladros que expresa un fragmento de la proteína insecticida Cry1Ab de la bacteria Bacillus Thuringiensis (Bt), comercializado en España desde 1998, ofrece una alternativa al uso de insecticidas químicos para el control de esta plaga. El mayor problema que puede presentar la tecnolo...

  16. Degradation and detection of transgenic Bacillus thuringiensis DNA and proteins in flour of three genetically modified rice events submitted to a set of thermal processes.

    Science.gov (United States)

    Wang, Xiaofu; Chen, Xiaoyun; Xu, Junfeng; Dai, Chen; Shen, Wenbiao

    2015-10-01

    This study aimed to investigate the degradation of three transgenic Bacillus thuringiensis (Bt) genes (Cry1Ab, Cry1Ac, and Cry1Ab/Ac) and the corresponding encoded Bt proteins in KMD1, KF6, and TT51-1 rice powder, respectively, following autoclaving, cooking, baking, or microwaving. Exogenous Bt genes were more stable than the endogenous sucrose phosphate synthase (SPS) gene, and short DNA fragments were detected more frequently than long DNA fragments in both the Bt and SPS genes. Autoclaving, cooking (boiling in water, 30 min), and baking (200 °C, 30 min) induced the most severe Bt protein degradation effects, and Cry1Ab protein was more stable than Cry1Ac and Cry1Ab/Ac protein, which was further confirmed by baking samples at 180 °C for different periods of time. Microwaving induced mild degradation of the Bt and SPS genes, and Bt proteins, whereas baking (180 °C, 15 min), cooking and autoclaving led to further degradation, and baking (200 °C, 30 min) induced the most severe degradation. The findings of the study indicated that degradation of the Bt genes and proteins somewhat correlated with the treatment intensity. Polymerase chain reaction, enzyme-linked immunosorbent assay, and lateral flow tests were used to detect the corresponding transgenic components. Strategies for detecting transgenic ingredients in highly processed foods are discussed. Copyright © 2015 Elsevier Ltd. All rights reserved.

  17. Estirpes de Bacillus thuringiensis efetivas contra insetos das ordens Lepidoptera, Coleoptera e Diptera Bacillus thuringiensis strains effective against insects of Lepidoptera, Coleoptera and Diptera orders

    Directory of Open Access Journals (Sweden)

    Lílian Botelho Praça

    2004-01-01

    Full Text Available O objetivo deste trabalho foi selecionar entre 300 estirpes de Bacillus thuringiensis as efetivas simultaneamente contra larvas de Spodoptera frugiperda J.E. Smith e Anticarsia gemmatalis Hübner (Lepidoptera: Noctuidae, Anthonomus grandis Boheman (Coleoptera: Curculionidae, Aedes aegypti Linnaeus e Culex quinquefasciatus Say (Diptera: Culicidae. Foram selecionadas duas estirpes de B. thuringiensis, denominadas S234 e S997, que apresentaram atividade contra as três ordens de insetos. As estirpes foram caracterizadas por métodos morfológicos, bioquímicos e moleculares. As mesmas apresentaram duas proteínas principais de 130 e 65 kDa, produtos de reação em cadeia da polimerase de tamanho esperado para a detecção dos genes cry1Aa, cry1Ab, cry1Ac, cry1B e cry2 e cristais bipiramidais, cubóides e esféricos.The aim of this work was to select among 300 strains of Bacillus thuringiensis those which are simultaneously effective against larvae of Spodoptera frugiperda J.E. Smith and Anticarsia gemmatalis Hübner (Lepidoptera: Noctuidae, Anthonomus grandis Boheman (Coleoptera: Curculionidae, Aedes aegypti Linnaeus and Culex quinquefasciatus Say (Diptera: Culicidae. Two strains of B. thuringiensis were selected, S234 and S997, which presented activity against those three insect orders. Both strains were characterized by morphological, biochemical and molecular methods. They have presented two main proteins with 130 and 65 kDa, polimerase chain reaction products with expected sizes for detection of the genes cry1Aa, cry1Ab, cry1Ac, cry1B and cry2 and bipiramidal, cubical and spherical crystals.

  18. A 90-day subchronic feeding study of genetically modified rice expressing Cry1Ab protein in Sprague-Dawley rats.

    Science.gov (United States)

    Song, Huan; He, Xiaoyun; Zou, Shiying; Zhang, Teng; Luo, Yunbo; Huang, Kunlun; Zhu, Zhen; Xu, Wentao

    2015-04-01

    Bacillus thuringiensis (Bt) transgenic rice line (mfb-MH86) expressing a synthetic cry1Ab gene can be protected against feeding damage from Lepidopteran insects, including Sesamia inferens, Chilo suppressalis, Tryporyza incertulas and Cnaphalocrocis medinalis. Rice flour from mfb-MH86 and its near-isogenic control MH86 was separately formulated into rodent diets at concentrations of 17.5, 35 and 70 % (w/w) for a 90-day feeding test with rats, and all of the diets were nutritionally balanced. In this study, the responses of rats fed diets containing mfb-MH86 were compared to those of rats fed flour from MH86. Overall health, body weight and food consumption were comparable between groups fed diets containing mfb-MH86 and MH86. Blood samples were collected prior to sacrifice and a few significant differences (p toxic effect was observed in the conditions of the experiment, based on the different parameters assessed. GM rice mfb-MH86 is as safe and nutritious as non-GM rice.

  19. Feeding behavior of neonate Ostrinia nubilalis (Lepidoptera: Crambidae) on Cry1Ab Bt corn: implications for resistance management.

    Science.gov (United States)

    Razze, J M; Mason, C E; Pizzolato, T D

    2011-06-01

    The European corn borer, Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae), is an economically important insect pest of corn, Zea mays L., in the United States and Canada. The development of genetically modified corn expressing genes derived from Bacillus thuringiensis (Bt) that encodes insecticidal crystalline (Cry) proteins has proven to be effective in controlling this insect. To assess the feeding behavior of neonate O. nubilalis on Bt corn, we examined differences in feeding behavior, based on presence of plant material in the gut, between Cry1Ab Bt corn and non-Bt near isoline corn for four intervals over a 48-h period. Feeding experiments revealed that there was significantly less feeding on Bt corn compared with non-Bt near isoline corn. The behavior of neonates on the plant corresponded with the differences in feeding on the two corn lines. The findings also showed that > 50% of the larvae initially left the plant before there was evidence in the gut of feeding regardless of whether the source was Bt or non-Bt corn. A higher quantity of plant material was found in the gut of larvae recovered from leaves of non-Bt compared with Bt corn. At the end of 48 h among the larvae that had left the plant, a greater proportion from Bt corn had plant material in the gut than did those from non-Bt corn.

  20. In vitro effect of Bacillus thuringiensis strains and Cry proteins in phytopathogenic fungi of paddy rice-field Efeito in vitro de cepas e proteínas Cry de Bacillus thuringiensis em fungos fitopatogênicos da cultura do arroz irrigado

    Directory of Open Access Journals (Sweden)

    Neiva Knaak

    2007-09-01

    Full Text Available Cry1Ab and Cry1Ac strains and proteins synthesized by Bacillus thuringiensis thuringiensis and B. thuringiensis kurstaki were assessed in the following phytopathogens: Rhizoctonia solani,Pyricularia grisea,Fusarium oxysporum and F. solani, which had their micelial growth decreased after incubation in the presence of the bacterial strains. As to Cry proteins, there were no inhibition halo development in the assessed concentrations.As cepas e proteínas Cry1Ab e Cry1Ac sintetizadas por Bacillus thuringiensis thuringiensis e B. thuringiensis kurstaki, foram avaliadas nos fitopatógenos: Rhizoctonia solani,Pyricularia grisea,Fusarium oxysporum e F. solani, os quais tiveram seu crescimento micelial reduzido após a incubação na presença das cepas bacterianas. Em relação às proteínas Cry, não houve formação de halo de inibição nas concentrações avaliadas.

  1. Histopathology and the lethal effect of Cry proteins and strains of Bacillus thuringiensis Berliner in Spodoptera frugiperda J.E. Smith Caterpillars (Lepidoptera, Noctuidae

    Directory of Open Access Journals (Sweden)

    N. Knaak

    Full Text Available Among the phytophagous insects which attack crops, the fall armyworm, Spodoptera frugiperda (J.E. Smith, 1797 (Lepidoptera, Noctuidae is particularly harmful in the initial growth phase of rice plants. As a potential means of controlling this pest, and considering that the entomopathogen Bacillus thuringiensis Berliner demonstrates toxicity due to synthesis of the Cry protein, the present study was undertaken to evaluate this toxic effect of B. thuringiensis thuringiensis 407 (pH 408 and B. thuringiensis kurstaki HD-73 on S. frugiperda. The following method was used. Both bacterial strains were evaluated in vitro in 1st instar S. frugiperda caterpillars, by means of histopathological assays. The Cry1Ab and Cry1Ac proteins, codified by the respective strains of B. thuringiensis, were evaluated in vivo by bioassays of 1st instar S. frugiperda caterpillars in order to determine the Mean Lethal Concentration (LC50. The results of the histopathological analysis of the midget of S. frugiperda caterpillars demonstrate that treatment with the B. thuringiensis thuringiensis strain was more efficient, because the degradations of the microvilosities started 9 hours after treatment application (HAT, while in the B. thuringiensis kurstaki the same effect was noticed only after 12 HAT. Toxicity data of the Cry1Ab and Cry1Ac proteins presented for the target-species LC50 levels of 9.29 and 1.79 μg.cm-2 respectively. The strains and proteins synthesised by B. thuringiensis thuringiensis and B. thuringiensis kurstaki are effective in controlling S. frugiperda, and may be used to produce new biopesticides or the genes may be utilised in the genetic transformation of Oryza sativa L.

  2. Antibiotics influence the toxicity of the delta endotoxins of Bacillus thuringiensis towards the cotton bollworm, Helicoverpa armigera

    Science.gov (United States)

    2014-01-01

    Background The cotton bollworm, Helicoverpa armigera is one of the most important crop pests worldwide. It has developed high levels of resistance to synthetic insecticides, and hence, Bacillus thuringiensis (Bt) formulations are used as a safer pesticide and the Bt genes have been deployed in transgenic crops for controlling this pest. There is an apprehension that H. armigera might develop resistance to transgenic crops in future. Therefore, we studied the role of gut microbes by eliminating them with antibiotics in H. armigera larvae on the toxicity of Bt toxins against this pest. Results Commercial formulation of Bt (Biolep®) and the pure Cry1Ab and Cry1Ac toxin proteins were evaluated at ED50, LC50, and LC90 dosages against the H. armigera larvae with and without antibiotics (which removed the gut microbes). Lowest H. armigera larval mortality due to Bt formulation and the Bt toxins Cry1Ab and Cry1Ac was recorded in insects reared on diets with 250 and 500 μg ml−1 diet of each of the four antibiotics (gentamicin, penicillin, rifampicin, and streptomycin), while the highest larval mortality was recorded in insects reared on diets without the antibiotics. Mortality of H. armigera larvae fed on diets with Bt formulation and the δ-endotoxins Cry1Ab and Cry1Ac was inversely proportional to the concentration of antibiotics in the artificial diet. Nearly 30% reduction in larval mortality was observed in H. armigera larvae from F1 to F3 generation when the larvae were reared on diets without antibiotics (with gut microbes) and fed on 0.15% Bt or 12 μg Cry1Ab or Cry1Ac ml−1 diet, indicating development of resistance to Bt in the presence of gut microflora. However, there were no differences in larval mortality due to Bt, Cry1Ab or Cry1Ac across generations in insects when they were reared on diets with 250 μg of each antibiotic ml−1 diet (without gut microflora). Conclusions The results suggested that antibiotics which eliminated gut microflora

  3. Bacillus thuringiensis (Bt)

    Science.gov (United States)

    2004-01-01

    Bacillus thuringiensis (Bt), a natural bacteria found all over the Earth, has a fairly novel way of getting rid of unwanted insects. Bt forms a protein substance (shown on the right) that is not harmful to humans, birds, fish or other vertebrates. When eaten by insect larvae the protein causes a fatal loss of appetite. For over 25 years agricultural chemical companies have relied heavily upon safe Bt pesticides. New space based research promises to give the insecticide a new dimension in effectiveness and applicability. Researchers from the Consortium for Materials Development in Space along with industrial affiliates such as Abott Labs and Pern State University flew Bt on a Space Shuttle mission in the fall of 1996. Researchers expect that the Shuttle's microgravity environment will reveal new information about the protein that will make it more effective against a wider variety of pests.

  4. Activity of Bacillus thuringiensis isolates against immature horn fly and stable fly (Diptera: Muscidae).

    Science.gov (United States)

    Lysyk, T J; Kalischuk-Tymensen, L D; Rochon, K; Selinger, L B

    2010-06-01

    We screened 85 isolates of Bacillus thuringiensis (Berliner), making up 57 different subspecies, and two isolates of Bacillus sphaericus (Meyer and Neide) for activity against immature horn flies, Haematobia irritans (L.), and stable flies, Stomoxys calcitrans (L.). The majority of B. thuringiensis and the B. sphaericus isolates had little or no activity against horn fly and stable fly. Approximately 87% of the isolates caused fly larvae and 64% caused stable fly, 95% of the isolates caused fly and stable fly immatures. These isolates were B. t. tolworthi 4L3, B. t. darmstadiensis 4M1, B. t. thompsoni 401, B. t. thuringiensis HD2, and B. t. kurstaki HD945. The LD50 values ranged from 2.2 to 7.9 x 10(6) spores per g manure for horn fly and from 6.3 to 35 x 10(6) spores per g media for stable fly. These were consistently more toxic compared with the B. t. israelensis isolates examined. All had DNA that hybridized with cry1Aa, cry1Ab, and cry1Ac toxin probes, three hybridized with a cry1B probe, and two hybridized with a cry2A probe. These may have potential for use in integrated management of pest flies.

  5. A mathematical model of exposure of non-target Lepidoptera to Bt-maize pollen expressing Cry1Ab within Europe

    Science.gov (United States)

    Perry, J. N.; Devos, Y.; Arpaia, S.; Bartsch, D.; Gathmann, A.; Hails, R. S.; Kiss, J.; Lheureux, K.; Manachini, B.; Mestdagh, S.; Neemann, G.; Ortego, F.; Schiemann, J.; Sweet, J. B.

    2010-01-01

    Genetically modified (GM) maize MON810 expresses a Cry1Ab insecticidal protein, derived from Bacillus thuringiensis (Bt), toxic to lepidopteran target pests such as Ostrinia nubilalis. An environmental risk to non-target Lepidoptera from this GM crop is exposure to harmful amounts of Bt-containing pollen deposited on host plants in or near MON810 fields. An 11-parameter mathematical model analysed exposure of larvae of three non-target species: the butterflies Inachis io (L.), Vanessa atalanta (L.) and moth Plutella xylostella (L.), in 11 representative maize cultivation regions in four European countries. A mortality–dose relationship was integrated with a dose–distance relationship to estimate mortality both within the maize MON810 crop and within the field margin at varying distances from the crop edge. Mortality estimates were adjusted to allow for physical effects; the lack of temporal coincidence between the susceptible larval stage concerned and the period over which maize MON810 pollen is shed; and seven further parameters concerned with maize agronomy and host-plant ecology. Sublethal effects were estimated and allowance made for aggregated pollen deposition. Estimated environmental impact was low: in all regions, the calculated mortality rate for worst-case scenarios was less than one individual in every 1572 for the butterflies and one in 392 for the moth. PMID:20053648

  6. 76 FR 14289 - Bacillus thuringiensis

    Science.gov (United States)

    2011-03-16

    ... a plant- incorporated protectant in accordance with the terms of Experimental Use Permit (EUP) No... chemistry data for eCry3.1Ab were required for a human health effects assessment. Even so, preliminary... EPA granting registrations or experimental use permits of Bacillus thuringiensis-based pesticides or...

  7. The end of a myth – Bt (Cry1Ab maize does not harm green lacewings

    Directory of Open Access Journals (Sweden)

    Joerg eRomeis

    2014-08-01

    Full Text Available A concern with Bt-transgenic insect-resistant plants is their potential to harm non-target organisms. Early studies reported that Cry1Ab-producing Bt maize and purified Cry1Ab harmed larvae of the green lacewing, Chrysoperla carnea. Although these effects could not be confirmed in subsequent studies, some authors still refer to them as evidence that Bt maize harms beneficial species. We provide a comprehensive review of the studies evaluating the effects of Bt (Cry1Ab maize on C. carnea. The evidence indicates that this important predator is not affected by Bt maize or by the produced Cry1Ab protein. We discuss how conceptual models can assist environmental risk assessments, and we emphasize the importance of robust and reproducible studies.

  8. The end of a myth-Bt (Cry1Ab) maize does not harm green lacewings.

    Science.gov (United States)

    Romeis, Jörg; Meissle, Michael; Naranjo, Steven E; Li, Yunhe; Bigler, Franz

    2014-01-01

    A concern with Bt-transgenic insect-resistant plants is their potential to harm non-target organisms. Early studies reported that Cry1Ab-producing Bt maize and purified Cry1Ab harmed larvae of the green lacewing, Chrysoperla carnea. Although these effects could not be confirmed in subsequent studies, some authors still refer to them as evidence that Bt maize harms beneficial species. We provide a comprehensive review of the studies evaluating the effects of Bt (Cry1Ab) maize on C. carnea. The evidence indicates that this important predator is not affected by Bt maize or by the produced Cry1Ab protein. We discuss how conceptual models can assist environmental risk assessments, and we emphasize the importance of robust and reproducible studies.

  9. The end of a myth—Bt (Cry1Ab) maize does not harm green lacewings

    Science.gov (United States)

    Romeis, Jörg; Meissle, Michael; Naranjo, Steven E.; Li, Yunhe; Bigler, Franz

    2014-01-01

    A concern with Bt-transgenic insect-resistant plants is their potential to harm non-target organisms. Early studies reported that Cry1Ab-producing Bt maize and purified Cry1Ab harmed larvae of the green lacewing, Chrysoperla carnea. Although these effects could not be confirmed in subsequent studies, some authors still refer to them as evidence that Bt maize harms beneficial species. We provide a comprehensive review of the studies evaluating the effects of Bt (Cry1Ab) maize on C. carnea. The evidence indicates that this important predator is not affected by Bt maize or by the produced Cry1Ab protein. We discuss how conceptual models can assist environmental risk assessments, and we emphasize the importance of robust and reproducible studies. PMID:25161661

  10. Application of Cry1Ab/Ac Bt strip for screening of resistance for ...

    African Journals Online (AJOL)

    ABU ZAYD

    2013-10-02

    Envirologix, 2008). ... gene, while the appearance of only the top (control) line indicates a negative response. Cry1Ab gene .... gene was found to be in Mendelian fashion in all the six cowpea crosses, the results indicate.

  11. Cry1Ab protein from Bt transgenic rice does not residue in rhizosphere soil

    International Nuclear Information System (INIS)

    Wang Haiyan; Ye Qingfu; Wang Wei; Wu Licheng; Wu Weixiang

    2006-01-01

    Expression of Cry1Ab protein in Bt transgenic rice (KMD) and its residue in the rhizosphere soil during the whole growth in field, as well as degradation of the protein from KMD straw in five soils under laboratory incubation were studied. The residue of Cry1Ab protein in KMD rhizosphere soil was undetectable (below the limit of 0.5 ng/g air-dried soil). The Cry1Ab protein contents in the shoot and root of KMD were 3.23-8.22 and 0.68-0.89 μg/g (fresh weight), respectively. The half-lives of the Cry1Ab protein in the soils amended with KMD straw (4%, w/w) ranged from 11.5 to 34.3 d. The residence time of the protein varied significantly in a Fluvio-marine yellow loamy soil amended with KMD straw at the rate of 3, 4 and 7%, with half-lives of 9.9, 13.8 and 18 d, respectively. In addition, an extraction method for Cry1Ab protein in soil was developed, with extraction efficiencies of 46.4-82.3%. - Cry1Ab protein was not detected in the rhizosphere soil of field-grown Bt transgenic rice

  12. Effects of Bacillus thuringiensis (Bt) corn on soil Folsomia fimetaria, Folsomia candida (Collembola), Hypoaspis aculeifer (Acarina) and Enchytraeus crypticus (Oligochaeta)

    DEFF Research Database (Denmark)

    Ke, X.; Krogh, P. H.

    The effects of the Cry1Ab toxin from Bacillus thuringiensis (corn variety Cascade Bt MON810 and DeKalb variety 618 Bt) were studied on survival and reproduction of the soil collembolan Folsomia fimetaria, Folsomia candida, the collembolan predator mite Hypoaspis aculeifer and enchytraeids....... There was a weak significant reduction by 30% on the reproduction of F. fimetaria fed Bt corn in Petri dishes for 21 days. Likewise there was a weak significant reduction by 40% of the reproduction of H. aculeifer by Bt corn in amounts corresponding to 20 g plant material kg-1 soil in the two species soil......-litter microcosm systems. There were no effects of Bt corn materials on the reproduction of F. fimetaria and E. crypticus in the single species soil-litter microcosms. No effects of Bt corn materials on mortality of all the 4 species were observed in all treatments. The tendency of effects of the Bt corn...

  13. Chitinolitic activity in proteic extracts of Bacillus thuringiensis toxic to boll weevil (Anthonomus grandis)

    Energy Technology Data Exchange (ETDEWEB)

    Silva, T.S; Rocha, T.L. [EMBRAPA Recursos Geneticos e Biotecnologia, DF (Brazil); Vasconcelos, E.A.R [Universidade de Brasilia (UnB), DF (Brazil); Grossi-de-Sa, M.F. [Universidade Catolica de Brasilia, DF (Brazil)

    2008-07-01

    Full text: Bacillus thuringiensis (Bt) is a spore forming bacteria, which produces Cry proteins toxic towards several insect orders. Bt S 811 strain produces at least three Cry toxins: Cry1Ab, Cry1Ia12, and Cry8, and shown toxicity to insects from Coleoptera order. In order to characterize the production of theses toxins, and check its activity against Boll weevil larvae, proteic extracts from Bt cells and supernatant proteins from the bacterial culture, were obtained at different stages of cell cycle; 8, 16, 24, and 32 hours after inoculation (HAI). Proteins from 32 HAI of the supernatant, and 8 HAI of the cellular fractions, shown highest activity towards the Boll weevil larvae. Western blotting assays using anti-Cry8 and anti-Cry1I were carried out to analyse these toxins in the Bt proteic extracts. The existence of a Cry8 was detected at 8 HAI in the cellular fraction, what allow associate this molecule with the toxicity of this fraction. However, toxicity observed at 32 HAI in the supernatant fraction, was not possible to be associated with Cry8 or Cry1Ia toxins, indicating that there are another protein(s) responsible for the toxicity. A protein homo log to Cry1Ab was identified by 'Peptide Mass Fingerprint' at 32 HAI of the supernatant fraction and a chitin binding protein was identified by 2DE/MS/MS in this same stage and chitinolitic activity was also observed by enzymatic assay. All our data suggest a possible synergism between Cry toxins and a chitinase in the activity of this strain towards Boll weevil.

  14. Chitinolitic activity in proteic extracts of Bacillus thuringiensis toxic to boll weevil (Anthonomus grandis)

    International Nuclear Information System (INIS)

    Silva, T.S; Rocha, T.L.; Vasconcelos, E.A.R; Grossi-de-Sa, M.F.

    2008-01-01

    Full text: Bacillus thuringiensis (Bt) is a spore forming bacteria, which produces Cry proteins toxic towards several insect orders. Bt S 811 strain produces at least three Cry toxins: Cry1Ab, Cry1Ia12, and Cry8, and shown toxicity to insects from Coleoptera order. In order to characterize the production of theses toxins, and check its activity against Boll weevil larvae, proteic extracts from Bt cells and supernatant proteins from the bacterial culture, were obtained at different stages of cell cycle; 8, 16, 24, and 32 hours after inoculation (HAI). Proteins from 32 HAI of the supernatant, and 8 HAI of the cellular fractions, shown highest activity towards the Boll weevil larvae. Western blotting assays using anti-Cry8 and anti-Cry1I were carried out to analyse these toxins in the Bt proteic extracts. The existence of a Cry8 was detected at 8 HAI in the cellular fraction, what allow associate this molecule with the toxicity of this fraction. However, toxicity observed at 32 HAI in the supernatant fraction, was not possible to be associated with Cry8 or Cry1Ia toxins, indicating that there are another protein(s) responsible for the toxicity. A protein homo log to Cry1Ab was identified by 'Peptide Mass Fingerprint' at 32 HAI of the supernatant fraction and a chitin binding protein was identified by 2DE/MS/MS in this same stage and chitinolitic activity was also observed by enzymatic assay. All our data suggest a possible synergism between Cry toxins and a chitinase in the activity of this strain towards Boll weevil

  15. Insecticidal Activity of Bacillus thuringiensis Strains Isolated from Soil and Water

    Directory of Open Access Journals (Sweden)

    Edyta Konecka

    2012-01-01

    Full Text Available We attempted to search novel Bacillus thuringiensis strains that produce crystals with potential utility in plant protection and with higher activity than strains already used in biopesticide production. Seven B. thuringiensis soil and water isolates were used in the research. We predicted the toxicity of their crystals by cry gene identification employing PCR method. The isolate MPU B63 with interesting, according to us, genes content was used in evaluating its crystal toxicity against Cydia pomonella caterpillars. The strain MPU B63 was cultured from water sample and had cry1Ab, cry1B, and cry15 genes. The LC50 crystals of MPU B63 were compared to LC50 of commercial bioinsecticide Foray determined against C. pomonella (codling moth. The activity of MPU B63 inclusions against codling moth larvae was approximately 24-fold higher than that of Foray. The results are a promising introduction for further study evaluating the potential usefulness of isolate MPU B63 crystals in plant protection.

  16. Insecticidal Activity of Bacillus thuringiensis Strains Isolated from Soil and Water

    Science.gov (United States)

    Konecka, Edyta; Baranek, Jakub; Hrycak, Anita; Kaznowski, Adam

    2012-01-01

    We attempted to search novel Bacillus thuringiensis strains that produce crystals with potential utility in plant protection and with higher activity than strains already used in biopesticide production. Seven B. thuringiensis soil and water isolates were used in the research. We predicted the toxicity of their crystals by cry gene identification employing PCR method. The isolate MPU B63 with interesting, according to us, genes content was used in evaluating its crystal toxicity against Cydia pomonella caterpillars. The strain MPU B63 was cultured from water sample and had cry1Ab, cry1B, and cry15 genes. The LC50 crystals of MPU B63 were compared to LC50 of commercial bioinsecticide Foray determined against C. pomonella (codling moth). The activity of MPU B63 inclusions against codling moth larvae was approximately 24-fold higher than that of Foray. The results are a promising introduction for further study evaluating the potential usefulness of isolate MPU B63 crystals in plant protection. PMID:22666145

  17. Lack of detrimental effects of Bacillus thuringiensis Cry toxins on the insect predator Chrysoperla carnea: a toxicological, histopathological, and biochemical analysis.

    Science.gov (United States)

    Rodrigo-Simón, Ana; de Maagd, Ruud A; Avilla, Carlos; Bakker, Petra L; Molthoff, Jos; González-Zamora, Jose E; Ferré, Juan

    2006-02-01

    The effect of Cry proteins of Bacillus thuringiensis on the green lacewing (Chrysoperla carnea) was studied by using a holistic approach which consisted of independent, complementary experimental strategies. Tritrophic experiments were performed, in which lacewing larvae were fed Helicoverpa armigera larvae reared on Cry1Ac, Cry1Ab, or Cry2Ab toxins. In complementary experiments, a predetermined amount of purified Cry1Ac was directly fed to lacewing larvae. In both experiments no effects on prey utilization or fitness parameters were found. Since binding to the midgut is an indispensable step for toxicity of Cry proteins to known target insects, we hypothesized that specific binding of the Cry1A proteins should be found if the proteins were toxic to the green lacewing. In control experiments, Cry1Ac was detected bound to the midgut epithelium of intoxicated H. armigera larvae, and cell damage was observed. However, no binding or histopathological effects of the toxin were found in tissue sections of lacewing larvae. Similarly, Cry1Ab or Cry1Ac bound in a specific manner to brush border membrane vesicles from Spodoptera exigua but not to similar fractions from green lacewing larvae. The in vivo and in vitro binding results strongly suggest that the lacewing larval midgut lacks specific receptors for Cry1Ab or Cry1Ac. These results agree with those obtained in bioassays, and we concluded that the Cry toxins tested, even at concentrations higher than those expected in real-life situations, do not have a detrimental effect on the green lacewing when they are ingested either directly or through the prey.

  18. Effect of Iranian Bt cotton on life table of Bemisia tabaci (Hemiptera: Alyrodidae and Cry 1Ab detection in the whitefly honeydew

    Directory of Open Access Journals (Sweden)

    Solmaz Azimi

    2016-09-01

    Full Text Available Transgenic cotton expressing the Cry 1Ab protein of Bacillus thuringiensis developing against Helocoverpa armigera may be affect on secondary pest such as Bemisia tabaci. In this study effects of Bt cotton on demographic parameters of B. tabaci were assessed and the data analyzed using the age specific, two-sex life table parameters. Results showed that getting to the adulthood stage, was faster on non-Bt cotton in comparison with Bt cotton. Also the fecundity was higher on non-Bt cotton than that on Bt cotton. Some of the population parameters (r, R0 and T of B. tabaci were affected by the Bt cotton significantly. The intrinsic rate of increase (r on Bt and non-Bt cotton was 0.07 day-1 and 0.1 day-1 , respectively. The net reproductive rate (R0 was 20.68 and 15.04 offspring/individual on Bt and non-Bt cotton, respectively. Mean generation time (T in non-Bt cotton was 27.22 and 34.62 days in Bt cotton. The results indicated that the life history of B. tabaci in the laboratory condition was influenced by host plant quality and Bt cotton was not a suitable host for B. tabaci. The western immunoblot method showed that the Cry protein detection in honeydew was positive which indicated that the Cry protein was ingested. Results revealed that the transgenic cotton could adversely affect the secondary pest and the natural enemies which feed on such pests as a host or their honeydew as a food source should be considered.

  19. Comparisons of transcriptional profiles of gut genes between cry1Ab-resistant and susceptible strains of Ostrinia nubilalis revealed genes possibly related to the adaptation of resistant larvae to transgenic cry1Ab corn

    Science.gov (United States)

    European corn borer (Ostrinia nubilalis) gut 2,895 unique genes expressions were quantified and compared between a laboratory-selected Cry1Ab-resistant (R) strain and a susceptible (S) strain after fed transgenic corn (MON810) leaves expressing Cry1Ab by microarray. A total of 398 gut genes were di...

  20. Efficient transformation of Bacillus thuringiensis requires nonmethylated plasmid DNA.

    OpenAIRE

    Macaluso, A; Mettus, A M

    1991-01-01

    The transformation efficiency of Bacillus thuringiensis depends upon the source of plasmid DNA. DNA isolated from B. thuringiensis, Bacillus megaterium, or a Dam- Dcm- Escherichia coli strain efficiently transformed several B. thuringiensis strains, B. thuringiensis strains were grouped according to which B. thuringiensis backgrounds were suitable sources of DNA for transformation of other B. thuringiensis strains, suggesting that B. thuringiensis strains differ in DNA modification and restri...

  1. The end of a myth – Bt(Cry1Ab) maize does not harm green lacewings

    Science.gov (United States)

    A concern with Bt-transgenic insect-resistant plants is their potential to harm non-target organisms. Early studies reported that Cry1Ab-producing Bt maize and purified Cry1Ab harmed larvae of the green lacewing, Chrysoperla carnea. Although these effects could not be confirmed in subsequent studies...

  2. Distribution and diversity of cry genes in native strains of Bacillus thuringiensis obtained from different ecosystems from Colombia.

    Science.gov (United States)

    Uribe, D; Martinez, W; Cerón, J

    2003-02-01

    Colombia is a tropical country located at the north of South America. It is considered to be one of the most important countries in terms of its biodiversity worldwide. One hundred and eight soil samples obtained from agricultural crops and wild ecosystems were evaluated in terms of the presence of Bacillus thuringiensis (Bt) native strains. One hundred and eight different Bt strains were isolated and characterized by the presence of crystal proteins by SDS-PAGE and a multiplex PCR with general and specific primers for cry1 and cry3, cry7, and cry8 gene detection. Most of the Bt strains (73%) reacted with the cry1 general primers; 27.8% of the Bt strains reacted with cry3, cry7, and cry8 general primers and 17.8% of strains did not react with any of these two sets of primers. Thirty different PCR profiles were found in the strains with cry1 genes when they were analyzed with specific primers (cry1A to cry1F). A high frequency of joint occurrence was observed for cry1Aa/cry1Ab, cry1Aa/cry1Ac, cry1Ab/cry1Ac, and cry1C/cry1D genes with a Pearson coefficient of 0.88, 0.74, 0.76, and 0.87, respectively. Other distinctive characteristics were found in the Colombian collection as the presence of 22.2% of native strains which presented, at the same time, lepidopteran and coleopteran active genes. Interesting relations were found as well between the cry gene distribution and the geographical areas sampled. Finally, some strains with moderate to high biopesticide activity against Spodoptera frugiperda (Lepidoptera) and Premnotrypes vorax (Coleoptera) insects were identified, this being important to explore future microbial strategies for the control of these crop pests in the region.

  3. Bacillus thuringiensis monogenic strains: screening and interactions with insecticides used against rice pests

    Science.gov (United States)

    Pinto, Laura M.N.; Dörr, Natália C.; Ribeiro, Ana Paula A.; de Salles, Silvia M.; de Oliveira, Jaime V.; Menezes, Valmir G.; Fiuza, Lidia M.

    2012-01-01

    The screening of Bacillus thuringiensis (Bt) Cry proteins with high potential to control insect pests has been the goal of numerous research groups. In this study, we evaluated six monogenic Bt strains (Bt dendrolimus HD-37, Bt kurstaki HD-1, Bt kurstaki HD-73, Bt thuringiensis 4412, Bt kurstaki NRD-12 and Bt entomocidus 60.5, which codify the cry1Aa, cry1Ab, cry1Ac, cry1Ba, cry1C, cry2A genes respectively) as potential insecticides for the most important insect pests of irrigated rice: Spodoptera frugiperda, Diatraea saccharalis, Oryzophagus oryzae, Oebalus poecilus and Tibraca limbativentris. We also analyzed their compatibility with chemical insecticides (thiamethoxam, labdacyhalothrin, malathion and fipronil), which are extensively used in rice crops. The bioassay results showed that Bt thuringiensis 4412 and Bt entomocidus 60.5 were the most toxic for the lepidopterans, with a 93% and 82% mortality rate for S. frugiperda and D. saccharalis, respectively. For O. oryzae, the Bt kurstaki NRD-12 (64%) and Bt dendrolimus HD-37 (62%) strains were the most toxic. The Bt dendrolimus HD-37 strain also caused high mortality (82%) to O. poecilus, however the strains assessed to T. limbativentris caused a maximum rate of 5%. The assays for the Bt strains interaction with insecticides revealed the compatibility of the six strains with the four insecticides tested. The results from this study showed the high potential of cry1Aa and cry1Ba genes for genetic engineering of rice plants or the strains to biopesticide formulations. PMID:24031872

  4. Assessment of the impact of Cry1Ab expression on insects dwelling on the maize plants

    Czech Academy of Sciences Publication Activity Database

    Habuštová, Oxana; Doležal, Petr; Hussein, H. M.; Spitzer, Lukáš; Turanli, F.; Růžička, Vlastimil; Sehnal, František

    2007-01-01

    Roč. 37, supplement 1 (2007), s. 50-51 ISSN 1738-2297. [International Congress of Insect Biotechnology and Industry. 19.08.2007-24.08.2007, Daegu] R&D Projects: GA AV ČR KJB6007304 Institutional research plan: CEZ:AV0Z50070508 Keywords : GM crops * Cry1Ab endotoxin * European corn borer Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection

  5. Discrimination of Transgenic Rice containing the Cry1Ab Protein using Terahertz Spectroscopy and Chemometrics

    Science.gov (United States)

    Xu, Wendao; Xie, Lijuan; Ye, Zunzhong; Gao, Weilu; Yao, Yang; Chen, Min; Qin, Jianyuan; Ying, Yibin

    2015-01-01

    Spectroscopic techniques combined with chemometrics methods have proven to be effective tools for the discrimination of objects with similar properties. In this work, terahertz time-domain spectroscopy (THz-TDS) combined with discriminate analysis (DA) and principal component analysis (PCA) with derivative pretreatments was performed to differentiate transgenic rice (Hua Hui 1, containing the Cry1Ab protein) from its parent (Ming Hui 63). Both rice samples and the Cry1Ab protein were ground and pressed into pellets for terahertz (THz) measurements. The resulting time-domain spectra were transformed into frequency-domain spectra, and then, the transmittances of the rice and Cry1Ab protein were calculated. By applying the first derivative of the THz spectra in conjunction with the DA model, the discrimination of transgenic from non-transgenic rice was possible with accuracies up to 89.4% and 85.0% for the calibration set and validation set, respectively. The results indicated that THz spectroscopic techniques and chemometrics methods could be new feasible ways to differentiate transgenic rice. PMID:26154950

  6. Siderophores of Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis.

    Science.gov (United States)

    Wilson, Melissa K; Abergel, Rebecca J; Raymond, Kenneth N; Arceneaux, Jean E L; Byers, B Rowe

    2006-09-15

    Three Bacillus anthracis Sterne strains (USAMRIID, 7702, and 34F2) and Bacillus cereus ATCC 14579 excrete two catecholate siderophores, petrobactin (which contains 3,4-dihydroxybenzoyl moieties) and bacillibactin (which contains 2,3-dihydroxybenzoyl moieties). However, the insecticidal organism Bacillus thuringiensis ATCC 33679 makes only bacillibactin. Analyses of siderophore production by previously isolated [Cendrowski et al., Mol. Microbiol. 52 (2004) 407-417] B. anthracis mutant strains revealed that the B. anthracis bacACEBF operon codes for bacillibactin production and the asbAB gene region is required for petrobactin assembly. The two catecholate moieties also were synthesized by separate routes. PCR amplification identified both asbA and asbB genes in the petrobactin producing strains whereas B. thuringiensis ATCC 33679 retained only asbA. Petrobactin synthesis is not limited to the cluster of B. anthracis strains within the B. cereus sensu lato group (in which B. cereus, B. anthracis, and B. thuringiensis are classified), although petrobactin might be prevalent in strains with pathogenic potential for vertebrates.

  7. Phage-Mediated Competitive Chemiluminescent Immunoassay for Detecting Cry1Ab Toxin by Using an Anti-Idiotypic Camel Nanobody.

    Science.gov (United States)

    Qiu, Yulou; Li, Pan; Dong, Sa; Zhang, Xiaoshuai; Yang, Qianru; Wang, Yulong; Ge, Jing; Hammock, Bruce D; Zhang, Cunzheng; Liu, Xianjin

    2018-01-31

    Cry toxins have been widely used in genetically modified organisms for pest control, raising public concern regarding their effects on the natural environment and food safety. In this work, a phage-mediated competitive chemiluminescent immunoassay (c-CLIA) was developed for determination of Cry1Ab toxin using anti-idiotypic camel nanobodies. By extracting RNA from camels' peripheral blood lymphocytes, a naive phage-displayed nanobody library was established. Using anti-Cry1Ab toxin monoclonal antibodies (mAbs) against the library for anti-idiotypic antibody screening, four anti-idiotypic nanobodies were selected and confirmed to be specific for anti-Cry1Ab mAb binding. Thereafter, a c-CLIA was developed for detection of Cry1Ab toxin based on anti-idiotypic camel nanobodies and employed for sample testing. The results revealed a half-inhibition concentration of developed assay to be 42.68 ± 2.54 ng/mL, in the linear range of 10.49-307.1 ng/mL. The established method is highly specific for Cry1Ab recognition, with negligible cross-reactivity for other Cry toxins. For spiked cereal samples, the recoveries of Cry1Ab toxin ranged from 77.4% to 127%, with coefficient of variation of less than 9%. This study demonstrated that the competitive format based on phage-displayed anti-idiotypic nanobodies can provide an alternative strategy for Cry toxin detection.

  8. Testing potential effects of Maize expressing the Bacillus thuringiensisCry1 Ab endotoxin (Bt Maize) on mycorrhizal fungal communities via DNA- and RNA-based pyrosequencing and molecular fingerprinting.

    NARCIS (Netherlands)

    Verbruggen, E.; Kuramae, E.E.; Hillekens, R.; de Hollander, M.; Kiers, E.T.; Roling, W.F.M.; Kowalchuk, G.A.; van der Heijden, M.G.A.

    2012-01-01

    The cultivation of genetically modified (GM) crops has increased significantly over the last decades. However, concerns have been raised that some GM traits may negatively affect beneficial soil biota, such as arbuscular mycorrhizal fungi (AMF), potentially leading to alterations in soil

  9. BOOK REVIEW – BACILLUS THURINGIENSIS: A CORNERSTONE OF MODERN AGRICULTURE BACILLUS THURINGIENSIS

    Science.gov (United States)

    Are you interested in the technical issues surrounding the use of Bacillus thuringiensis pesticidal traits as sprays and as plant incorporated protectants (transgenic crops)? Should the dimensions of human health, ecology, entomology, risk assessment, resistance management, and d...

  10. Does Cry1Ab protein affect learning performances of the honey bee Apis mellifera L. (Hymenoptera, Apidae)?

    Science.gov (United States)

    Ramirez-Romero, R; Desneux, N; Decourtye, A; Chaffiol, A; Pham-Delègue, M H

    2008-06-01

    Genetically modified Bt crops are increasingly used worldwide but side effects and especially sublethal effects on beneficial insects remain poorly studied. Honey bees are beneficial insects for natural and cultivated ecosystems through pollination. The goal of the present study was to assess potential effects of two concentrations of Cry1Ab protein (3 and 5000 ppb) on young adult honey bees. Following a complementary bioassay, our experiments evaluated effects of the Cry1Ab on three major life traits of young adult honey bees: (a) survival of honey bees during sub-chronic exposure to Cry1Ab, (b) feeding behaviour, and (c) learning performance at the time that honey bees become foragers. The latter effect was tested using the proboscis extension reflex (PER) procedure. The same effects were also tested using a chemical pesticide, imidacloprid, as positive reference. The tested concentrations of Cry1Ab protein did not cause lethal effects on honey bees. However, honey bee feeding behaviour was affected when exposed to the highest concentration of Cry1Ab protein, with honey bees taking longer to imbibe the contaminated syrup. Moreover, honey bees exposed to 5000 ppb of Cry1Ab had disturbed learning performances. Honey bees continued to respond to a conditioned odour even in the absence of a food reward. Our results show that transgenic crops expressing Cry1Ab protein at 5000 ppb may affect food consumption or learning processes and thereby may impact honey bee foraging efficiency. The implications of these results are discussed in terms of risks of transgenic Bt crops for honey bees.

  11. Preliminary investigations reveal that Bacillus thuringiensis δ ...

    African Journals Online (AJOL)

    The imminent introduction of transgenic crops into Kenya requires a rigorous assessment of the potential risks involved. This study focused on the possible effect of Bacillus thuringiensisδ-endotoxin [CryIA(c)] on arbuscular mycorrhizal fungi (AMF) associated with sorghum. In green house experiments, sorghum seedlings ...

  12. Isolation and characterization of native Bacillus thuringiensis ...

    African Journals Online (AJOL)

    ... (<30% mortality) or non insecticidal activity. However, results of motility, hemolytic activity, antibiotic-susceptibility patterns, and crystal shape, seem to suggest that many of our Bt isolates may exhibit parasporins activity. Key words: Bacillus thuringiensis, parasporal crystal, isolation, biochemical type, insecticidal, cry gene, ...

  13. Purification And Characterization Of Marine Bacillus Thuringiensis ...

    African Journals Online (AJOL)

    Urease was purified to homogeneity from Bacillus thuringiensis N2 using different purification steps namely, 55% acetone precipitation, DEAE-Sephadex A50 anion exchange column and Sephadex G120-200 gel filtration chromatography. The enzyme was purified 95.27 fold and showed a final specific activity of 10.48 ...

  14. Aminopeptidase N isoforms from the midgut of Bombyx mori and Plutella xylostella -- their classification and the factors that determine their binding specificity to Bacillus thuringiensis Cry1A toxin.

    Science.gov (United States)

    Nakanishi, Kazuko; Yaoi, Katsuro; Nagino, Yasushi; Hara, Hirotaka; Kitami, Madoka; Atsumi, Shogo; Miura, Nami; Sato, Ryoichi

    2002-05-22

    Novel aminopeptidase N (APN) isoform cDNAs, BmAPN3 and PxAPN3, from the midguts of Bombyx mori and Plutella xylostella, respectively, were cloned, and a total of eight APN isoforms cloned from B. mori and P. xylostella were classified into four classes. Bacillus thuringiensis Cry1Aa and Cry1Ab toxins were found to bind to specific APN isoforms from the midguts of B. mori and P. xylostella, and binding occurred with fragments that corresponded to the BmAPN1 Cry1Aa toxin-binding region of each APN isoform. The results suggest that APN isoforms have a common toxin-binding region, and that the apparent specificity of Cry1Aa toxin binding to each intact APN isoform seen in SDS-PAGE is determined by factors such as expression level in conjunction with differences in binding affinity.

  15. Fate of Cry1Ab protein in agricultural systems under slurry management of cows fed genetically modified maize (Zea mays L.) MON810: a quantitative assessment.

    Science.gov (United States)

    Gruber, Helga; Paul, Vijay; Guertler, Patrick; Spiekers, Hubert; Tichopad, Ales; Meyer, Heinrich H D; Muller, Martin

    2011-07-13

    The objective of the study was to track the fate of recombinant Cry1Ab protein in a liquid manure field trial when feeding GM maize MON810 to dairy cows. A validated ELISA was applied for quantification of Cry1Ab in the agricultural chain from GM maize plants, feed, liquid manure and soil to crops grown on manured fields. Starting with 23.7 μg of Cry1Ab g(-1) dry weight GM maize material, a rapid decline of Cry1Ab levels was observed as 2.6% and 0.9% of Cry1Ab from the GM plant were detected in feed and liquid manure, respectively. Half of this residual Cry1Ab persisted during slurry storage for 25 weeks. After application to experimental fields, final degradation of Cry1Ab to below detectable levels in soil was reported. Cry1Ab exhibited a higher rate of degradation compared to total protein in the agricultural processes. Immunoblotting revealed a degradation of the 65 kDa Cry1Ab into immunoreactive fragments of lower size in all analyzed materials.

  16. DNA degradation in genetically modified rice with Cry1Ab by food processing methods: implications for the quantification of genetically modified organisms.

    Science.gov (United States)

    Xing, Fuguo; Zhang, Wei; Selvaraj, Jonathan Nimal; Liu, Yang

    2015-05-01

    Food processing methods contribute to DNA degradation, thereby affecting genetically modified organism detection and quantification. This study evaluated the effect of food processing methods on the relative transgenic content of genetically modified rice with Cry1Ab. In steamed rice and rice noodles, the levels of Cry1Ab were ⩾ 100% and <83%, respectively. Frying and baking in rice crackers contributed to a reduction in Pubi and Cry1Ab, while microwaving caused a decrease in Pubi and an increase in Cry1Ab. The processing methods of sweet rice wine had the most severe degradation effects on Pubi and Cry1Ab. In steamed rice and rice noodles, Cry1Ab was the most stable, followed by SPS and Pubi. However, in rice crackers and sweet rice wine, SPS was the most stable, followed by Cry1Ab and Pubi. Therefore, Cry1Ab is a better representative of transgenic components than is Pubi because the levels of Cry1Ab were less affected compared to Pubi. Copyright © 2014 Elsevier Ltd. All rights reserved.

  17. Histopathological effects and determination of the putative receptor of Bacillus thuringiensis Cry1Da toxin in Spodoptera littoralis midgut.

    Science.gov (United States)

    BenFarhat-Touzri, Dalel; Saadaoui, Marwa; Abdelkefi-Mesrati, Lobna; Saadaoui, Imen; Azzouz, Hichem; Tounsi, Slim

    2013-02-01

    Bacillus thuringiensis subsp. aizawai strain HD133, known by its effectiveness against Spodoptera species, produces many insecticidal proteins including Cry1Ab, Cry1Ca and Cry1Da. In the present study, the insecticidal activity of Cry1Da against Spodoptera littoralis was investigated. It showed toxicity with an LC(50) of 224.4 ng/cm(2) with 95% confidence limits of (178.61-270.19) and an LC(90) of 467.77 ng/cm(2) with 95% confidence limits of (392.89-542.65). The midgut histopathology of Cry1Da fed larvae showed vesicle formation in the apical region, vacuolization and destruction of epithelial cells. Biotinylated-activated Cry1Da toxin bound protein of about 65 kDa on blots of S. littoralis brush border membrane preparations. This putative receptor differs in molecular size from those recognized by Cry1C and Vip3A which are active against this polyphagous insect. This difference in midgut receptors strongly supports the use of Cry1Da as insecticidal agent, particularly in case of Cry and/or Vip-resistance management. Copyright © 2012 Elsevier Inc. All rights reserved.

  18. Ostrinia nubilalis parasitism and the field abundance of non-target insects in transgenic Bacillus thuringiensis corn (Zea mays).

    Science.gov (United States)

    Bourguet, Denis; Chaufaux, Josette; Micoud, Annie; Delos, Marc; Naibo, Bernard; Bombarde, Fany; Marque, Gilles; Eychenne, Nathalie; Pagliari, Carine

    2002-10-01

    In this study, we evaluated in field trials the effects on non-target species, of transgenic corn producing the Cry1Ab toxin of Bacillus thuringiensis (Bt). In 1998, we collected Ostrinia nubilalis (Hübner) larvae from transgenic Bt corn (Novartis Hybrid 176) and non-Bt corn at four geographical sites. We found a significant variation in parasitism by the tachinids Lydella thompsoni (Herting) and Pseudoperichaeta nigrolineata (Walker) among sites, and more parasitism in non-Bt than in Bt fields. The Bt effect did not vary significantly among fields. In 1999, we performed a field experiment at two sites, comparing the temporal abundance of non-target arthropods in Bt corn (Monsanto Hybrid MON810) and non-Bt corn. The non-target insects studied included the aphids Metopolophium dirhodum (Walker), Rhopalosiphum padi (L.) and Sitobion avenae (F.), the bug Orius insidiosus (Say), the syrphid Syrphus corollae (Meigen), the ladybird Coccinella septempunctata (L.), the lacewing Chrysoperla carnea (Stephens), thrips and hymenopteran parasitoids. For all species but one, the number of individuals varied greatly over the season but did not differ between the types of corn. The only exception was thrips which, at one site, was significantly more abundant in Bt corn than in non-Bt corn. However this difference did not remain significant when we took the multiple tests into account. Implications for pest resistance management, population dynamics and risk assessment are discussed.

  19. Binding specificity of Bacillus thuringiensis Cry1Aa for purified, native Bombyx mori aminopeptidase N and cadherin-like receptors

    Directory of Open Access Journals (Sweden)

    Jenkins Jeremy L

    2001-10-01

    Full Text Available Abstract Background To better understand the molecular interactions of Bt toxins with non-target insects, we have examined the real-time binding specificity and affinity of Cry1 toxins to native silkworm (Bombyx mori midgut receptors. Previous studies on B. mori receptors utilized brush border membrane vesicles or purifed receptors in blot-type assays. Results The Bombyx mori (silkworm aminopeptidase N (APN and cadherin-like receptors for Bacillus thuringiensis insecticidal Cry1Aa toxin were purified and their real-time binding affinities for Cry toxins were examined by surface plasmon resonance. Cry1Ab and Cry1Ac toxins did not bind to the immobilized native receptors, correlating with their low toxicities. Cry1Aa displayed moderate affinity for B. mori APN (75 nM, and unusually tight binding to the cadherin-like receptor (2.6 nM, which results from slow dissociation rates. The binding of a hybrid toxin (Aa/Aa/Ac was identical to Cry1Aa. Conclusions These results indicate domain II of Cry1Aa is essential for binding to native B. mori receptors and for toxicity. Moreover, the high-affinity binding of Cry1Aa to native cadherin-like receptor emphasizes the importance of this receptor class for Bt toxin research.

  20. Bacillus thuringiensis Vip3Aa Toxin Resistance in Heliothis virescens (Lepidoptera: Noctuidae).

    Science.gov (United States)

    Pickett, Brian R; Gulzar, Asim; Ferré, Juan; Wright, Denis J

    2017-05-01

    Laboratory selection with Vip3Aa of a field-derived population of Heliothis virescens produced >2,040-fold resistance in 12 generations of selection. The Vip3Aa-selected (Vip-Sel)-resistant population showed little cross-resistance to Cry1Ab and no cross-resistance to Cry1Ac. Resistance was unstable after 15 generations without exposure to the toxin. F 1 reciprocal crosses between Vip3Aa-unselected (Vip-Unsel) and Vip-Sel insects indicated a strong paternal influence on the inheritance of resistance. Resistance ranged from almost completely recessive (mean degree of dominance [ h ] = 0.04 if the resistant parent was female) to incompletely dominant (mean h = 0.53 if the resistant parent was male). Results from bioassays on the offspring from backcrosses of the F 1 progeny with Vip-Sel insects indicated that resistance was due to more than one locus. The results described in this article provide useful information for the insecticide resistance management strategies designed to overcome the evolution of resistance to Vip3Aa in insect pests. IMPORTANCE Heliothis virescens is an important pest that has the ability to feed on many plant species. The extensive use of Bacillus thuringiensis (Bt) crops or spray has already led to the evolution of insect resistance in the field for some species of Lepidoptera and Coleoptera. The development of resistance in insect pests is the main threat to Bt crops. The effective resistance management strategies are very important to prolong the life of Bt plants. Lab selection is the key step to test the assumption and predictions of management strategies prior to field evaluation. Resistant insects offer useful information to determine the inheritance of resistance and the frequency of resistance alleles and to study the mechanism of resistance to insecticides. Copyright © 2017 American Society for Microbiology.

  1. Transgenic Bacillus thuringiensis (Bt rice is safer to aquatic ecosystems than its non-transgenic counterpart.

    Directory of Open Access Journals (Sweden)

    Guangsheng Li

    Full Text Available Rice lines genetically modified with the crystal toxin genes from Bacillus thuringiensis (Bt have experienced rapid development, with biosafety certificates for two Bt rice lines issued in 2009. There has still been no commercial release of these lines yet due to public concerns about human health and environmental risks. Some studies confirmed that Bt rice was as safe as conventional rice to non-target organisms when pesticides were not applied, however, pesticides are still required in Bt rice to control non-lepidopteran pests. In this study, we assessed the environmental effects of two Bt rice lines expressing either the cry1Ab/1Ac or cry2A genes, respectively, by using zooplanktons as indicator species under normal field management practices using pesticides when required. In the whole rice growing season, non-Bt rice was sprayed 5 times while Bt rice was sprayed 2 times, which ensured both rice achieved a normal yield. Field investigations showed that rice type (Bt and non-Bt significantly influenced zooplankton abundance and diversity, which were up to 95% and 80% lower in non-Bt rice fields than Bt rice fields. Laboratory rearing showed that water from non-Bt rice fields was significantly less suitable for the survival and reproduction of Daphnia magna and Paramecium caudatum in comparison with water from Bt rice fields. Higher pesticide residues were detected in the water from non-Bt than Bt rice fields, accounting for the bad performance of zooplankton in non-Bt field water. Our results demonstrate that Bt rice is safer to aquatic ecosystems than non-Bt rice, and its commercialization will be beneficial for biodiversity restoration in rice-based ecosystems.

  2. Isolation of bacillus thuringiensis from different samples from Mansehra District

    International Nuclear Information System (INIS)

    Younis, F.; Lodhi, A.F.; Raza, G.

    2009-01-01

    The insecticidal activity of Bacillus thuringiensis has made it very interesting for the control of a variety of agricultural pests and human disease vectors. The present study is an attempt to explore the potential and diversity. of Bacillus thuringiensis. from the local environment for the control of cotton spotted bollworm (Earias sp.), a major pest of cotton. Two hundred and ninety eight samples of soil, grain dust, wild animal dung, birds dropping, decaying leaves and dead insects were collected from different ecological environments of Mansehra District yielding 438 Bacillus thuringiensis isolates that produce parasporal crystalline inclusions. In this study the soil samples were found to be the richest source for Bacillus thuringiensis. (author)

  3. Impacts of Bacillus thuringiensis var. israelensis and Bacillus ...

    African Journals Online (AJOL)

    The study assessed the impact of bio-larvicides- Bacillus thuringiensis var. israelensis (Bti) and B. sphaericus (Bs) on anopheline mosquito larval densities in four selected areas of Lusaka urban district. Larval densities were determined using a standard WHO protocol at each study area prior to and after larviciding.

  4. Occurrence of Bacillus thuringiensis in faeces of herbivorous farm ...

    African Journals Online (AJOL)

    Bacillus thuringiensis (Berliner), the insect pathogen has been isolated from a variety of habitat. It is understood that the habitat of B. thuringiensis has always been associated with their biological activity. In the present study, B. thuringiensis was isolated from faeces of cows and goats. The phenotypic characterization ...

  5. Comparisons of Transcriptional Profiles of Gut Genes between Cry1Ab-Resistant and Susceptible Strains of Ostrinia nubilalis Revealed Genes Possibly Related to the Adaptation of Resistant Larvae to Transgenic Cry1Ab Corn

    Directory of Open Access Journals (Sweden)

    Jianxiu Yao

    2017-01-01

    Full Text Available A microarray developed on the basis of 2895 unique transcripts from larval gut was used to compare gut gene expression profiles between a laboratory-selected Cry1Ab-resistant (R strain and its isoline susceptible (S strain of the European corn borer (Ostrinia nubilalis after the larvae were fed the leaves of transgenic corn (MON810 expressing Cry1Ab or its non-transgenic isoline for 6 h. We revealed 398 gut genes differentially expressed (i.e., either up- or down-regulated genes with expression ratio ≥2.0 in S-strain, but only 264 gut genes differentially expressed in R-strain after being fed transgenic corn leaves. Although the percentages of down-regulated genes among the total number of differentially expressed genes (50% in S-strain and 45% in R-strain were similar between the R- and S-strains, the expression ratios of down-regulated genes were much higher in S-strain than in R-strain. We revealed that 17 and 9 significantly up- or down-regulated gut genes from S and R-strain, respectively, including serine proteases and aminopeptidases. These genes may be associated with Cry1Ab toxicity by degradation, binding, and cellular defense. Overall, our study suggests enhanced adaptation of Cry1Ab-resistant larvae on transgenic Cry1Ab corn as revealed by lower number and lower ratios of differentially expressed genes in R-strain than in S-strain of O. nubilalis.

  6. A 52-week safety study in cynomolgus macaques for genetically modified rice expressing Cry1Ab/1Ac protein.

    Science.gov (United States)

    Mao, Jie; Sun, Xing; Cheng, Jian-Hua; Shi, Yong-Jie; Wang, Xin-Zheng; Qin, Jun-Jie; Sang, Zhi-Hong; He, Kun; Xia, Qing

    2016-09-01

    A 52-week feeding study in cynomolgus macaques was carried out to evaluate the safety of Bt rice Huahui 1 (HH1), a transgenic rice line expressing Cry1Ab/1Ac protein. Monkeys were fed a diet with 20% or 60% HH1 rice, 20% or 60% parental rice (Minghui 63, MH63), normal diet, normal diet spiked with purified recombinant Cry1Ab/1Ac fusion protein or bovine serum albumin (BSA) respectively. During the feeding trail, clinical observations were conducted daily, and multiple parameters, including body weight, body temperature, electrocardiogram, hematology, blood biochemistry, serum metabolome and gut microbiome were examined at regular intervals. Upon sacrifice, the organs were weighted, and the macroscopic, microscopic and electron microscopic examinations were performed. The results show no adverse or toxic effects of Bt rice HH1 or Cry1Ab/1Ac fusion protein on monkeys. Therefore, the present 52-week primate feeding study suggests that the transgenic rice containing Cry 1Ab/1Ac is equivalent to its parental rice line MH63. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Isolation and characterization of Bacillus thuringiensis from soils in ...

    African Journals Online (AJOL)

    Of 110 Bacillus thuringiensis isolates analyzed for the presence of crystal protein genes, 7 tested positive for cry 4, cry 11, and cyt toxin genes. Sequencing of these genes in positive strains demonstrated 99–100 % homology to known mosquitocidal cry and cyt genes in Bacillus thuringiensis subsp. israelensis. The present ...

  8. by lipase from Bacillus thuringiensis and Lysinibacillus sphaericus

    African Journals Online (AJOL)

    This study reported production of lipase by Bacillus thuringiensis and Lysinibacillus sphaericus. Bacteria isolates were screened on Bushnell-Hass Mineral Salt medium containing 1% PMS for oil degradation. Two potent isolates were identified using 16S rRNA as Bacillus thuringiensis and Lysinibacillus sphaericus.

  9. An ultrasensitive label-free electrochemiluminescent immunosensor for measuring Cry1Ab level and genetically modified crops content.

    Science.gov (United States)

    Gao, Hongfei; Wen, Luke; Wu, Yuhua; Fu, Zhifeng; Wu, Gang

    2017-11-15

    The development of genetically modified (GM) insect-resistant crops has aroused great public concern about the risks on the eco-environment resulting from a release of toxic Cry proteins (such as Cry1Ab) to the soil. Therefore, it is of crucial importance to measure the Cry proteins level and the GM crops content. Here, we have tested for the first time a method that uses novel carbon nanospheres (CNPs) label-free electrochemiluminescent (ECL) immunosensor for the ultrasensitive quantification of Cry1Ab and GM crops. In this work, novel CNPs were prepared from printer toner with a very facile approach, and linked with anti-Cry1Ab antibodies to modify a golden working electrode. The immunoreaction between Cry1Ab and its antibody formed an immunocomplex on the bioreceptor region of the sensor, which inhibited electron transfer between the electrode surface and the ECL substance, leading to a decrease of ECL response. Under the optimal conditions, the fabricated label-free ECL immunosensor determined Cry1Ab down to 3.0pgmL -1 within a linear range of 0.010-1.0ngmL -1 , showing significant improvement of sensitivity than that of most previous reports. Meanwhile, the proposed method was successfully applied for GM rice BT63 and GM maize MON810 detections down to 0.010% and 0.020%, respectively. Due to its outstanding advantages such as high sensitivity, ideal selectivity, simple fabrication, rapid detection, and low cost, the developed method can be considered as a powerful and pioneering tool for GM crops detection. Its use can also be extended to other toxin protein sensing in foods. Copyright © 2017. Published by Elsevier B.V.

  10. Association of bioassays and molecular characterization to select new Bacillus thuringiensis isolates effective against Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae)

    International Nuclear Information System (INIS)

    Fatoretto, Julio C.; Sena, Janete A.D.; Lemos, Manoel V.F.; Junior Boica, Arlindo L. , Jaboticabal, SP . Faculdade de Ciencias Agrarias e Veterinarias. Dept. de Fitossanidade; Brazil)

    2007-01-01

    The fall armyworm, Spodoptera frugiperda (J. E. Smith), is one of the main corn pests and Bacillus thuringiensis is important in its control because of its entomopathogenic property. The objective of this study was the molecular characterization of B. thuringiensis isolates for cry1 locus presence and the assessment of the efficiency of these isolates in controlling S. frugiperda caterpillars. Gral-cry1 was used in the PCR analyses to confirm the presence of the cry1 locus in 15 isolates. A 3 x 108 spore/ml suspension bathed the diet used to feed 30 caterpillars per isolate, with three replications. The cry1 locus type genes of the different isolates were identified for five gene subclasses; linear regression analyses were carried out to ascertain possible associations between the presence of an individual cry1 locus gene and high levels of toxicity. All the DNAs amplified with Gral-cry1 presented an amplification product with the expected size. Regarding the levels of insecticide efficiency against the cob worm, 41 isolates presented 100% mortality and 16 presented an index between 70% and 90%. The cry1Ab gene was present in 80 isolates, cryb in 69 isolates, cry1Ac in all the isolates and cryv and cry1E in 93 and 27 isolates, respectively. The values regarding the individual effect of each gene on caterpillar mortality were significant at 1% probability for the cry1Ac and cry1E genes. (author)

  11. Isolation and characterization of a new Bacillus thuringiensis strain Lip harboring a new cry1Aa gene highly toxic to Ephestia kuehniella (Lepidoptera: Pyralidae) larvae.

    Science.gov (United States)

    El Khoury, Micheline; Azzouz, Hichem; Chavanieu, Alain; Abdelmalak, Nouha; Chopineau, Joël; Awad, Mireille Kallassy

    2014-06-01

    The aim of this study was to characterize new Bacillus thuringiensis strains that have a potent insecticidal activity against Ephestia kuehniella larvae. Strains harboring cry1A genes were tested for their toxicity, and the Lip strain showed a higher insecticidal activity compared to that of the reference strain HD1 (LC50 of Lip and HD1 were 33.27 and 128.61 μg toxin/g semolina, respectively). B. thuringiensis Lip harbors and expresses cry1Aa, cry1Ab, cry1Ac, cry1Ad and cry2A. DNA sequencing revealed several polymorphisms in Lip Cry1Aa and Cry1Ac compared to the corresponding proteins of HD1. The activation process using Ephestia kuehniella midgut juice showed that Lip Cry1A proteins were more stable in the presence of larval proteases. Moreover, LipCry1A proteins exhibited higher insecticidal activity against these larvae. These results indicate that Lip is an interesting strain that could be used as an alternative to the worldwide used strain HD1.

  12. Transferrin Impacts Bacillus thuringiensis Biofilm Levels

    Directory of Open Access Journals (Sweden)

    Bianca Garner

    2016-01-01

    Full Text Available The present study examined the impact of transferrin on Bacillus thuringiensis biofilms. Three commercial strains, an environmental strain (33679, the type strain (10792, and an isolate from a diseased insect (700872, were cultured in iron restricted minimal medium. All strains produced biofilm when grown in vinyl plates at 30°C. B. thuringiensis 33679 had a biofilm biomass more than twice the concentration exhibited by the other strains. The addition of transferrin resulted in slightly increased growth yields for 2 of the 3 strains tested, including 33679. In contrast, the addition of 50 μg/mL of transferrin resulted in an 80% decrease in biofilm levels for strain 33679. When the growth temperature was increased to 37°C, the addition of 50 μg/mL of transferrin increased culture turbidity for only strain 33679. Biofilm levels were again decreased in strain 33679 at 37°C. Growth of B. thuringiensis cultures in polystyrene resulted in a decrease in overall growth yields at 30°C, with biofilm levels significantly decreased for 33679 in the presence of transferrin. These findings demonstrate that transferrin impacts biofilm formation in select strains of B. thuringiensis. Identification of these differences in biofilm regulation may be beneficial in elucidating potential virulence mechanisms among the differing strains.

  13. BACILLUS THURINGIENSIS ELASTASES WITH INSECTICIDE ACTIVITY

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    E. V. Matseliukh

    2015-10-01

    Full Text Available The purpose of the research was a screening of proteases with elastase activity among Bacillus thuringiensis strains, their isolation, partially purification, study of physicochemical properties and insecticide activity in relation to the larvae of the Colorado beetle. The objects of the investigation were 18 strains of B. thuringiensis, isolated from different sources: sea water, dry biological product "Bitoksibatsillin" and also from natural populations of Colorado beetles of the Crimea, Kherson, Odesa, Mykolaiv and Zaporizhiia regions of Ukraine. Purification of enzymes with elastase activity isolated from above mentioned strains was performed by gel-chromatography and insecticide activity was studied on the 3–4 larvae instar of Colorado beetle. The ability of a number of B. thuringiensis strains to synthesize the proteases with elastase activity has been established. The most active were enzymes obtained from strains IMV B-7465, IMV B-7324 isolated from sea water, and strains 9, 902, Bt-H and 0-239 isolated from Colorado beetles. The study of the physicochemical properties of the partially purified proteases of these strains showed that they belonged to enzymes of the serine type. Peptidases of a number of B. thuringiensis strains (IMV B-7324, IMV B-7465, 902, 0-239, 9 are metal-dependent enzymes. Optimal conditions of action of all tested enzymes are the neutral and alkaline рН values and the temperatures of 30–40 °С. The studies of influence of the complex enzyme preparations and partially purified ones of B. thuringiensis strains on the larvae instar of Colorado beetles indicated that enzymes with elastase activity could be responsible for insecticide action of the tested strains.

  14. Expressed sequence tags from larval gut of the European corn borer (Ostrinia nubilalis: Exploring candidate genes potentially involved in Bacillus thuringiensis toxicity and resistance

    Directory of Open Access Journals (Sweden)

    Crespo Andre LB

    2009-06-01

    Full Text Available Abstract Background Lepidoptera represents more than 160,000 insect species which include some of the most devastating pests of crops, forests, and stored products. However, the genomic information on lepidopteran insects is very limited. Only a few studies have focused on developing expressed sequence tag (EST libraries from the guts of lepidopteran larvae. Knowledge of the genes that are expressed in the insect gut are crucial for understanding basic physiology of food digestion, their interactions with Bacillus thuringiensis (Bt toxins, and for discovering new targets for novel toxins for use in pest management. This study analyzed the ESTs generated from the larval gut of the European corn borer (ECB, Ostrinia nubilalis, one of the most destructive pests of corn in North America and the western world. Our goals were to establish an ECB larval gut-specific EST database as a genomic resource for future research and to explore candidate genes potentially involved in insect-Bt interactions and Bt resistance in ECB. Results We constructed two cDNA libraries from the guts of the fifth-instar larvae of ECB and sequenced a total of 15,000 ESTs from these libraries. A total of 12,519 ESTs (83.4% appeared to be high quality with an average length of 656 bp. These ESTs represented 2,895 unique sequences, including 1,738 singletons and 1,157 contigs. Among the unique sequences, 62.7% encoded putative proteins that shared significant sequence similarities (E-value ≤ 10-3with the sequences available in GenBank. Our EST analysis revealed 52 candidate genes that potentially have roles in Bt toxicity and resistance. These genes encode 18 trypsin-like proteases, 18 chymotrypsin-like proteases, 13 aminopeptidases, 2 alkaline phosphatases and 1 cadherin-like protein. Comparisons of expression profiles of 41 selected candidate genes between Cry1Ab-susceptible and resistant strains of ECB by RT-PCR showed apparently decreased expressions in 2 trypsin-like and 2

  15. Association of bioassays and molecular characterization to select new Bacillus thuringiensis isolates effective against Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae); Associacao de bioensaios e caracterizacao molecular para selecao de novos isolados de Bacillus thuringiensis efetivos contra Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae)

    Energy Technology Data Exchange (ETDEWEB)

    Fatoretto, Julio C.; Sena, Janete A.D.; Lemos, Manoel V.F. [UNESP, Jaboticabal, SP (Brazil). Faculdade de Ciencias Agrarias e Veterinarias. Dept. de Biologia Aplicada a Agropecuaria; Barreto, Marliton R. [Universidade Federal do Mato Grosso (UFMT), Cuiaba, MT (Brazil). Inst. Universitario do Norte Matogrossense (IUNMAT)]. E-mail: mrbarreto@pop.com.br; Junior Boica, Arlindo L. (UNESP), Jaboticabal, SP (Brazil). Faculdade de Ciencias Agrarias e Veterinarias. Dept. de Fitossanidade)

    2007-09-15

    The fall armyworm, Spodoptera frugiperda (J. E. Smith), is one of the main corn pests and Bacillus thuringiensis is important in its control because of its entomopathogenic property. The objective of this study was the molecular characterization of B. thuringiensis isolates for cry1 locus presence and the assessment of the efficiency of these isolates in controlling S. frugiperda caterpillars. Gral-cry1 was used in the PCR analyses to confirm the presence of the cry1 locus in 15 isolates. A 3 x 108 spore/ml suspension bathed the diet used to feed 30 caterpillars per isolate, with three replications. The cry1 locus type genes of the different isolates were identified for five gene subclasses; linear regression analyses were carried out to ascertain possible associations between the presence of an individual cry1 locus gene and high levels of toxicity. All the DNAs amplified with Gral-cry1 presented an amplification product with the expected size. Regarding the levels of insecticide efficiency against the cob worm, 41 isolates presented 100% mortality and 16 presented an index between 70% and 90%. The cry1Ab gene was present in 80 isolates, cryb in 69 isolates, cry1Ac in all the isolates and cryv and cry1E in 93 and 27 isolates, respectively. The values regarding the individual effect of each gene on caterpillar mortality were significant at 1% probability for the cry1Ac and cry1E genes. (author)

  16. A Fast and Sensitive Quantitative Lateral Flow Immunoassay for Cry1Ab Based on a Novel Signal Amplification Conjugate

    OpenAIRE

    Chen, Chunxiang; Wu, Jian

    2012-01-01

    A novel lateral flow immunoassay (LFIA) signal amplification strategy for the detection of Cry1Ab based on amplification via a polylysine (PL) chain and biotin-streptavidin system (BSAS) is described. In this system, multiple fluorescence dyes (FL) were directly coated on the surface of PL and conjugated with antibody via the BSAS for construction of novel signal amplification (FLPL-BSAS-mAb1) conjugates, in which FL, PL and BSAS were employed to improve the sensitivity of LFIA. Compared with...

  17. A fast and sensitive quantitative lateral flow immunoassay for Cry1Ab based on a novel signal amplification conjugate.

    Science.gov (United States)

    Chen, Chunxiang; Wu, Jian

    2012-01-01

    A novel lateral flow immunoassay (LFIA) signal amplification strategy for the detection of Cry1Ab based on amplification via a polylysine (PL) chain and biotin-streptavidin system (BSAS) is described. In this system, multiple fluorescence dyes (FL) were directly coated on the surface of PL and conjugated with antibody via the BSAS for construction of novel signal amplification (FLPL-BSAS-mAb1) conjugates, in which FL, PL and BSAS were employed to improve the sensitivity of LFIA. Compared with conventional LFIA, the sensitivity of FLPL-BSAS-mAb1-based LFIA was increased by approximately 100-fold. Quantified linearity was achieved in the value range of 0-1,000 pg/mL. The limit of detection (LOD) was reached 10 pg/mL after optimization of reaction conditions. To our knowledge, this represents one of the most sensitive LFIA for Cry1Ab yet reported. Furthermore, the detection time for this method was about 10 min. Therefore, it should be an attractive alternative compared to conventional immunoassays in routine control for Cry1Ab.

  18. Isolation and Characterization of Bacillus thuringiensis strains from ...

    African Journals Online (AJOL)

    Twelve isolates of Bacillus thuringiensis were obtained from soil samples collected from farmers' fields in Kakamega and Machakos districts. Soil samples from Machakos yielded more B. thuringiensis isolates t han those from Kakamega. Three isolates K13-1, 12F-K and K10-2 were obtained from Kakamega soil samples.

  19. Characterization of Bacillus thuringiensis strains from Jordan and ...

    African Journals Online (AJOL)

    Eight serotypes with Bacillus thuringiensis israelensis being the most common. Out of the twenty-six isolated strains, five strains (serotype: kenyae, kurstaki, kurstaki HD1 and thuringiensis) that produced bipyramid crystal proteins were toxic to the lepidoptera larvae of Ephestia kuehniella Zeller. The SDS-PAGE protein ...

  20. Ecology and diversity of Bacillus thuringiensis in soil environment ...

    African Journals Online (AJOL)

    Bacillus thuringiensis populations ranged between 4.23 x 105, 6.52 x 105 cfu/g soil and consist of 11 types of isolates with 3 polymorphic, 7 spherical and 1 bipyramidal type of crystals. Polymorphic crystal containing isolates were further characterized. B. thuringiensis isolates were circular, white, flat and undulate or entire.

  1. BOOK REVIEW: BACILLUS THURINGIENSIS: A CORNERSTONE OF MODERN AGRICULTURE

    Science.gov (United States)

    Are you interested in the technical issues surrounding the use of Bacillus thuringiensis pesticidal traits as sprays and as plant incorporated protectants (transgenic crops)? Should the dimensions of human health, ecology, entomology, risk assessment, resistance management, and d...

  2. Cytotoxic effects of delfin insecticide ( Bacillus thuringiensis ) on cell ...

    African Journals Online (AJOL)

    Cytotoxic effects of delfin insecticide ( Bacillus thuringiensis ) on cell behaviour, phagocytosis, contractile vacuole activity and macronucleus in a protozoan ciliate Paramecium caudatum. ... macronucleus, fragmentation, vacuolization and complete diffusion of macronucleus were observed and were dose dependent.

  3. Ecological aspects of Bacillus thuringiensis in an Oxisol Ecologia do Bacillus thuringiensis num Latossolo

    Directory of Open Access Journals (Sweden)

    Lessandra Heck Paes Leme Ferreira

    2003-02-01

    Full Text Available Bacillus thuringiensis is a Gram positive, sporangial bacterium, known for its insecticidal habilities. Survival and conjugation ability of B. thuringiensis strains were investigated; vegetative cells were evaluated in non-sterile soil. Vegetative cells decreased rapidly in number, and after 48 hours the population was predominantly spores. No plasmid transfer was observed in non-sterile soil, probably because the cells died and the remaining cells sporulated quickly. Soil is not a favorable environment for B. thuringiensis multiplication and conjugation. The fate of purified B. thuringiensis toxin was analyzed by extractable toxin quantification using ELISA. The extractable toxin probably declined due to binding on surface-active particles in the soil.O comportamento de células vegetativas do Bacillus thuringiensis foi estudado em solo não esterilizado. Após o inóculo grande parte das células morrem e o restante esporula em 24 horas. Não foi observada conjugação provavelmente porque poucas células sobrevivem no solo e rapidamente esporulam, mostrando que este não é o ambiente propício para a multiplicação e conjugação desta bactéria. A toxina purificada, portanto livre de células, diminui rapidamente sua quantidade em solo não esterilizado. Provavelmente a ligação da toxina na fração argilosa do solo é a principal responsável por este fenômeno.

  4. Bacillus thuringiensis Conjugation in Simulated Microgravity

    Science.gov (United States)

    Beuls, Elise; van Houdt, Rob; Leys, Natalie; Dijkstra, Camelia; Larkin, Oliver; Mahillon, Jacques

    2009-10-01

    Spaceflight experiments have suggested a possible effect of microgravity on the plasmid transfer among strains of the Gram-positive Bacillus thuringiensis, as opposed to no effect recorded for Gram-negative conjugation. To investigate these potential effects in a more affordable experimental setup, three ground-based microgravity simulators were tested: the Rotating Wall Vessel (RWV), the Random Positioning Machine (RPM), and a superconducting magnet. The bacterial conjugative system consisted in biparental matings between two B. thuringiensis strains, where the transfer frequencies of the conjugative plasmid pAW63 and its ability to mobilize the nonconjugative plasmid pUB110 were assessed. Specifically, potential plasmid transfers in a 0-g position (simulated microgravity) were compared to those obtained under 1-g (normal gravity) condition in each device. Statistical analyses revealed no significant difference in the conjugative and mobilizable transfer frequencies between the three different simulated microgravitational conditions and our standard laboratory condition. These important ground-based observations emphasize the fact that, though no stimulation of plasmid transfer was observed, no inhibition was observed either. In the case of Gram-positive bacteria, this ability to exchange plasmids in weightlessness, as occurs under Earth's conditions, should be seen as particularly relevant in the scope of spread of antibiotic resistances and bacterial virulence.

  5. Modification of hormonal balance in larvae of the corn borer Sesamia nonagrioides (Lepidoptera: Noctuidae) due to sublethal Bacillus thuringiensis protein ingestion.

    Science.gov (United States)

    Pérez-Hedo, Meritxell; Albajes, Ramon; Eizaguirre, Matilde

    2011-06-01

    Bacillus thuringiensis (Bt) corn, Zea mays L., is highly efficient against the corn borer Sesamia nonagrioides (Lefèbvre) (Lepidoptera: Noctuidae) when the larvae feed only on the transgenic plants. However, when they feed on Bt leaves during only part of their development, thus ingesting sublethal amounts of Bt toxins, some larvae survive. A previous study reported a prolonged development and precocious diapause induction in larvae fed on a diet with sublethal amounts of Cry1Ab protein. To determine whether these effects were accompanied by a modification of the hormonal balance, S. nonagrioides larvae were fed on sublethal amounts of Bt protein provided in Bt leaves or in the diet. The larvae that survived had higher levels of juvenile hormone (JH), whereas their level of ecdysteroids did not increase sufficiently to allow pupation, leading to a longer larval development and more larval molts. This response may be considered a defense mechanism that allows some larvae to survive toxin ingestion; it is similar the response to insecticidal toxins or viruses observed in other larvae. Changes in the hormone levels in diapausing larvae were undetectable, probably because these changes were masked by the higher level of JH in the hemolymph of diapausing larvae and because of lack of ecdysteroid titer increase, a phenomenon that is usually observed a few days before pupation in nondiapausing larvae. These results should be taken into account in the establishment of non-Bt refuges to prevent development of Bt-resistance in S. non-agrioides populations.

  6. A high-throughput, in-vitro assay for Bacillus thuringiensis insecticidal proteins.

    Science.gov (United States)

    Izumi Willcoxon, Michi; Dennis, Jaclyn R; Lau, Sabina I; Xie, Weiping; You, You; Leng, Song; Fong, Ryan C; Yamamoto, Takashi

    2016-01-10

    A high-throughput, in-vitro assay for Bacillus thuringiensis (Bt) insecticidal proteins designated as Cry was developed and evaluated for screening a large number of Cry protein variants produced by DNA shuffling. This automation-amenable assay exploits an insect cell line expressing a single receptor of Bt Cry proteins. The Cry toxin used to develop this assay is a variant of the Cry1Ab protein called IP1-88, which was produced previously by DNA shuffling. Cell mortality caused by the activated Bt Cry toxin was determined by chemical cell viability assay in 96/384-well microtiter plates utilizing CellTiter 96(®) obtained from Promega. A widely-accepted mode-of-action theory of certain Bt Cry proteins suggests that the activated toxin binds to one or more receptors and forms a pore through the insect gut epithelial cell apical membrane. A number of insect proteins such as cadherin-like protein (Cad), aminopeptidase-N (APN), alkaline phosphatase (ALP) and ABC transporter (ABCC) have been identified as the receptors of Bt Cry toxins. In this study, Bt Cry toxin receptors Ostrinia nubilalis (European corn borer) cadherin-like protein (On-Cad) and aminopeptidase-N 1 and 3 (On-APN1, On-APN3) and Spodoptera frugiperda (fall armyworm) cadherin-like protein (Sf-Cad) were cloned in an insect cell line, Sf21, and a mammalian cell line, Expi293F. It was observed by ligand blotting and immunofluorescence microscopy that trypsin-activated IP1-88 bound to On-Cad and On-APN1, but not Sf-Cad or On-APN3. In contrast, IP1-88 bound only to APN1 in BBMV (Brush Border Membrane Vesicles) prepared from the third and fourth-instar O. nubilalis larval midgut. The sensitivity of the recombinant cells to the toxin was then tested. IP1-88 showed no toxicity to non-recombinant Sf21 and Expi293F. Toxicity was observed only when the On-Cad gene was cloned and expressed. Sf-Cad and On-APN1 were not able to make those cells sensitive to the toxin. Since the expression of On-Cad alone was

  7. The use of two biological formulations of Bacillus Thuringiensis and ...

    African Journals Online (AJOL)

    Two biological formulations of the microbial agents Bacillus thuringiensis and Bacillus sphaericus, known by their trade names Vectobac 12 AS and VectoLex CG (Corn Cob) granules, respectively, were obtained from Valent Biosciences Company (formerly Abbott Laboratories) of North Chicago, USA, and applied to control ...

  8. Antagonistic activity of selected strains of Bacillus thuringiensis ...

    African Journals Online (AJOL)

    The aim of this work was to determine, in vitro, the antagonistic effectiveness of 60 strains of Bacillus thuringiensis against damping-off and root and stem rot caused by Rhizoctonia solani. The strains were obtained from the International Collection of Entomopathogenic Bacillus at the FCB-UANL. During the in vitro dual ...

  9. SR450 and Superhawk XP Applications of Bacillus Thuringiensis Israelensis Against Culex Quinquefasciatus

    Science.gov (United States)

    2014-09-01

    SR450 AND SUPERHAWK XP APPLICATIONS OF BACILLUS THURINGIENSIS ISRAELENSIS AGAINST CULEX QUINQUEFASCIATUS1 JAMES C. DUNFORD,2 CRAIG A. STOOPS,3 ALDEN...thermal fogger applications of VectobacH WDG Bacillus thuringiensis israelensis (Bti) against Culex quinquefasciatus. Bacillus thuringiensis ...3. DATES COVERED 00-00-2014 to 00-00-2014 4. TITLE AND SUBTITLE SR450 and Superhawk XP Applications of Bacillus Thuringiensis Israelensis

  10. Inheritance Patterns, Dominance and Cross-Resistance of Cry1Ab- and Cry1Ac-Selected Ostrinia furnacalis (Guenée

    Directory of Open Access Journals (Sweden)

    Tiantao Zhang

    2014-09-01

    Full Text Available Two colonies of Asian corn borer, Ostrinia furnacalis (Guenée, artificially selected from a Bt-susceptible colony (ACB-BtS for resistance to Cry1Ab (ACB-AbR and Cry1Ac (ACB-AcR toxins, were used to analyze inheritance patterns of resistance to Cry1 toxins. ACB-AbR and ACB-AcR evolved significant levels of resistance, with resistance ratios (RR of 39-fold and 78.8-fold to Cry1Ab and Cry1Ac, respectively. The susceptibility of ACB-AbR larvae to Cry1Ac and Cry1F toxins, which had not previously been exposed, were significantly reduced, being >113-fold and 48-fold, respectively. Similarly, susceptibility of ACB-AcR larvae to Cry1Ab and Cry1F were also significantly reduced (RR > nine-fold, RR > 18-fold, respectively, indicating cross-resistance among Cry1Ab, Cry1Ac, and Cry1F toxins. However, ACB-AbR and ACB-AcR larvae were equally susceptible to Cry1Ie as were ACB-BtS larvae, indicating no cross-resistance between Cry1Ie and Cry1Ab or Cry1Ac toxins; this may provide considerable benefits in preventing or delaying the evolution of resistance in ACB to Cry1Ab and Cry1Ac toxins. Backcrossing studies indicated that resistance to Cry1Ab toxin was polygenic in ACB-AbR, but monogenic in ACB-AcR, whilst resistance to Cry1Ac toxin was primarily monogenic in both ACB-AbR and ACB-AcR, but polygenic as resistance increased.

  11. Initial Study of Production of Bacillus thuringiensis israelensis Using Locally Obtained Substrates

    OpenAIRE

    Soesanto, Soesanto

    2015-01-01

    Bacillus thuringiensis israelensis diketahui sebagai entomopatogen nyamuk ades aegypti yang merupakan vektor penyakit demam berdarah.Telah dilakukan percobaan produksi Bacillus thuringiensis israelensis dengan menggunakan medium yang mengandung bahan lokal terasi udang, melase dan tepung kedelai. Hasil uji toksisitas (bioassay) terhadap Final Whole Culture (FWC) dengan bahan lokal yang murah menunjukkan potensi sebagai agensia bioinsektisida.Key Words: entomopathogen - Bacillus thuringiensis ...

  12. 40 CFR 180.1011 - Viable spores of the microorganism Bacillus thuringiensis Berliner; exemption from the...

    Science.gov (United States)

    2010-07-01

    ... Bacillus thuringiensis Berliner; exemption from the requirement of a tolerance. 180.1011 Section 180.1011... microorganism Bacillus thuringiensis Berliner; exemption from the requirement of a tolerance. (a) For the... authentic strain of Bacillus thuringiensis Berliner conforming to the morphological and biochemical...

  13. Genetic Basis of Cry1F-Resistance in a Laboratory Selected Asian Corn Borer Strain and Its Cross-Resistance to Other Bacillus thuringiensis Toxins.

    Directory of Open Access Journals (Sweden)

    Yueqin Wang

    Full Text Available The Asian corn borer (ACB, Ostrinia furnacalis (Guenée (Lepidoptera: Crambidae, is the most destructive insect pest of corn in China. Susceptibility to the Cry1F toxin derived from Bacillus thuringiensis has been demonstrated for ACB, suggesting the potential for Cry1F inclusion as part of an insect pest management program. Insects can develop resistance to Cry toxins, which threatens the development and use of Bt formulations and Bt crops in the field. To determine possible resistance mechanisms to Cry1F, a Cry1F-resistant colony of ACB (ACB-FR that exhibited more than 1700-fold resistance was established through selection experiments after 49 generations of selection under laboratory conditions. The ACB-FR strain showed moderate cross-resistance to Cry1Ab and Cry1Ac of 22.8- and 26.9-fold, respectively, marginally cross-resistance to Cry1Ah (3.7-fold, and no cross-resistance to Cry1Ie (0.6-fold. The bioassay responses of progeny from reciprocal F1 crosses to different Cry1 toxin concentrations indicated that the resistance trait to Cry1Ab, Cry1Ac and Cry1F has autosomal inheritance with no maternal effect or sex linked. The effective dominance (h of F1 offspring was calculated at different concentrations of Cry1F, showing that h decreased as concentration of Cry1F increased. Finally, the analysis of actual and expected mortality of the progeny from a backcross (F1 × resistant strain indicated that the inheritance of the resistance to Cry1F in ACB-FR was due to more than one locus. The present study provides an understanding of the genetic basis of Cry1F resistance in ACB-FR and also shows that pyramiding Cry1F with Cry1Ah or Cry1Ie could be used as a strategy to delay the development of ACB resistance to Bt proteins.

  14. Pathogenomic sequence analysis of Bacillus cereus and Bacillus thuringiensis isolates closely related to Bacillus anthracis.

    Science.gov (United States)

    Han, Cliff S; Xie, Gary; Challacombe, Jean F; Altherr, Michael R; Bhotika, Smriti S; Brown, Nancy; Bruce, David; Campbell, Connie S; Campbell, Mary L; Chen, Jin; Chertkov, Olga; Cleland, Cathy; Dimitrijevic, Mira; Doggett, Norman A; Fawcett, John J; Glavina, Tijana; Goodwin, Lynne A; Green, Lance D; Hill, Karen K; Hitchcock, Penny; Jackson, Paul J; Keim, Paul; Kewalramani, Avinash Ramesh; Longmire, Jon; Lucas, Susan; Malfatti, Stephanie; McMurry, Kim; Meincke, Linda J; Misra, Monica; Moseman, Bernice L; Mundt, Mark; Munk, A Christine; Okinaka, Richard T; Parson-Quintana, B; Reilly, Lee Philip; Richardson, Paul; Robinson, Donna L; Rubin, Eddy; Saunders, Elizabeth; Tapia, Roxanne; Tesmer, Judith G; Thayer, Nina; Thompson, Linda S; Tice, Hope; Ticknor, Lawrence O; Wills, Patti L; Brettin, Thomas S; Gilna, Paul

    2006-05-01

    Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis are closely related gram-positive, spore-forming bacteria of the B. cereus sensu lato group. While independently derived strains of B. anthracis reveal conspicuous sequence homogeneity, environmental isolates of B. cereus and B. thuringiensis exhibit extensive genetic diversity. Here we report the sequencing and comparative analysis of the genomes of two members of the B. cereus group, B. thuringiensis 97-27 subsp. konkukian serotype H34, isolated from a necrotic human wound, and B. cereus E33L, which was isolated from a swab of a zebra carcass in Namibia. These two strains, when analyzed by amplified fragment length polymorphism within a collection of over 300 of B. cereus, B. thuringiensis, and B. anthracis isolates, appear closely related to B. anthracis. The B. cereus E33L isolate appears to be the nearest relative to B. anthracis identified thus far. Whole-genome sequencing of B. thuringiensis 97-27and B. cereus E33L was undertaken to identify shared and unique genes among these isolates in comparison to the genomes of pathogenic strains B. anthracis Ames and B. cereus G9241 and nonpathogenic strains B. cereus ATCC 10987 and B. cereus ATCC 14579. Comparison of these genomes revealed differences in terms of virulence, metabolic competence, structural components, and regulatory mechanisms.

  15. Cry1Ab treatment has no effects on viability of cultured porcine intestinal cells, but triggers Hsp70 expression.

    Directory of Open Access Journals (Sweden)

    Angelika Bondzio

    Full Text Available In vitro testing can contribute to reduce the risk that the use of genetically modified (GM crops and their proteins show unintended toxic effects. Here we introduce a porcine intestinal cell culture (IPEC-J2 as appropriate in vitro model and tested the possible toxic potential of Cry1Ab protein, commonly expressed in GM-maize. For comprehensive risk assessment we used WST-1 conversion and ATP content as metabolic markers for proliferation, lactate dehydrogenase release as indicator for cells with compromised membrane and transepithelial electrical resistance as parameter indicating membrane barrier function. The results were compared to the effects of valinomycin, a potassium ionophore, known to induce cytotoxic effects in most mammalian cell types. Whereas no toxicity was observed after Cry1Ab treatment, valinomycin induced a decrease in IPEC-J2 viability. This was confirmed by dynamic monitoring of cellular responses. Additionally, two dimensional differential in-gel electrophoresis was performed. Only three proteins were differentially expressed. The functions of these proteins were associated with responses to stress. The up-regulation of heat shock protein Hsp70 was verified by Western blotting as well as by enzyme-linked immunosorbent assay and may be related to a protective function. These findings suggest that the combination of in vitro testing and proteomic analysis may serve as a promising tool for mechanism based safety assessment.

  16. Variation in microRNA levels between Cry1Ab-resistant and susceptible strains of the European corn borer, Ostrinia nubilalis (Hübner)

    Science.gov (United States)

    Transgenic maize hybrids that express insecticidal Bacillus thuringiensis (Bt) crystalline (Cry) protein toxins effectively protect against feeding damage inflicted by the insect pest the European corn borer, Ostrinia nubilalis. Field monitoring and laboratory selections have detected varying level...

  17. Ultra-violet-resistant mutants of Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Jones, D.R.; Karunakaran, V.; Hacking, A.J.

    1991-01-01

    One of the main disadvantages of using Bacillus thuringiensis as an insecticide is that the spore and crystal preparations applied to foliage are readily washed away by rain and are inactivated by sunlight. Spores from some strains of B. thuringiensis have been shown to be highly sensitive to u.v. light. This study has demonstrated how mutants with increased resistance to u.v., isolated by successive rounds of u.v. irradiation, and additionally with increased specific pathogenicity can be isolated. These techniques should be applied to strains that are frequently used in the industrial production of B.thuringiensis toxin. (author)

  18. Plasmid-associated sensitivity of Bacillus thuringiensis to UV light

    International Nuclear Information System (INIS)

    Benoit, T.G.; Wilson, G.R.; Bull, D.L.; Aronson, A.I.

    1990-01-01

    Spores and vegetative cells of Bacillus thuringiensis were more sensitive to UV light than were spores or cells of plasmid-cured B. thuringiensis strains or of the closely related Bacillus cereus. Introduction of B. thuringiensis plasmids into B. cereus by cell mating increased the UV sensitivity of the cells and spores. Protoxins encoded by one or more B. thuringiensis plasmids were not involved in spore sensitivity, since a B. thuringiensis strain conditional for protoxin accumulation was equally sensitive at the permissive and nonpermissive temperatures. In addition, introduction of either a cloned protoxin gene, the cloning vector, or another plasmid not containing a protoxin gene into a plasmid-cured strain of B. thuringiensis all increased the UV sensitivity of the spores. Although the variety of small, acid-soluble proteins was the same in the spores of all strains examined, the quantity of dipicolinic acid was about twice as high in the plasmid-containing strains, and this may account for the differences in UV sensitivity of the spores. The cells of some strains harboring only B. thuringiensis plasmids were much more sensitive than cells of any of the other strains, and the differences were much greater than observed with spores

  19. Effects of 60-Week Feeding Diet Containing Bt Rice Expressing the Cry1Ab Protein on the Offspring of Inbred Wuzhishan Pigs Fed the Same Diet.

    Science.gov (United States)

    Liu, Qiang; Yang, Weigang; Li, Mingjie; Wu, Yi; Wang, Yingzheng; Wu, Shuaishuai; Gao, Hui; Han, Ying; Yang, Feng; Feng, Shutang; Zeng, Shenming

    2017-11-29

    We evaluated the chronic effects of Bt rice carrying the Cry1Ab protein (1.64 mg/kg) on offspring of highly inbred WZSP, fed with Bt rice for 360 days, in a 60-week feeding study. The WZSP offspring (n = 27) were assigned to two groups (Minghui86 group, female n = 6, male n = 5; Bt group, female n = 11, male n = 5). The average obtained Cry1Ab protein dosage for female and male pigs was 1.003 and 1.234 mg/kg body weight after 10 weeks of feeding, respectively. The experimental feed in the study was nutritionally matched in both groups. The average daily gain and feed conversion ratio of the females in week 3 and males from weeks 1 to 10 were different between the Bt and Minghui86 groups (P rice carrying Cry1Ab protein exerts no unintended adverse effects on WZSP offspring.

  20. A system for the directed evolution of the insecticidal protein from Bacillus thuringiensis.

    Science.gov (United States)

    Ishikawa, Hiroshi; Hoshino, Yasushi; Motoki, Yutaka; Kawahara, Takuma; Kitajima, Mika; Kitami, Madoka; Watanabe, Ayako; Bravo, Alejandra; Soberon, Mario; Honda, Atsuko; Yaoi, Katsuro; Sato, Ryoichi

    2007-06-01

    Theoretically, the activity of AB-type toxin molecules such as the insecticidal toxin (Cry toxin) from B. thuringiensis, which have one active site and two binding site, is improved in parallel with the binding affinity to its receptor. In this experiment, we tried to devise a method for the directed evolution of Cry toxins to increase the binding affinity to the insect receptor. Using a commercial T7 phage-display system, we expressed Cry1Aa toxin on the phage surface as fusions with the capsid protein 10B. These recombinant phages bound to a cadherin-like protein that is one of the Cry1Aa toxin receptors in the model target insect Bombyx mori. The apparent affinity of Cry1Aa-expressing phage for the receptor was higher than that of Cry1Ab-expressing phage. Phages expressing Cry1Aa were isolated from a mixed suspension of phages expressing Cry1Ab and concentrated by up to 130,000-fold. Finally, random mutations were made in amino acid residues 369-375 in domain 2 of Cry1Aa toxin, the mutant toxins were expressed on phages, and the resulting phage library was screened with cadherin-like protein-coated beads. As a result, phages expressing abnormal or low-affinity mutant toxins were excluded, and phages with high-affinity mutant toxins were selected. These results indicate that a method combining T7 phage display with selection using cadherin-like protein-coated magnetic beads can be used to increase the activity of easily obtained, low-activity Cry toxins from bacteria.

  1. Occurrence of natural Bacillus thuringiensis contaminants and residues of Bacillus thuringiensis-based insecticides on fresh fruits and vegetables

    DEFF Research Database (Denmark)

    Frederiksen, Kristine; Rosenquist, Hanne; Jørgensen, Kirsten

    2006-01-01

    A total of 128 Bacillus cereus-like strains isolated from fresh fruits and vegetables for sale in retail shops in Denmark were characterized. Of these strains, 39% (50/128) were classified as Bacillus thuringiensis on the basis of their content of cry genes determined by PCR or crystal proteins v...... isolated from fruits and vegetables. The same was seen for a third enterotoxin, CytK. In conclusion, the present study strongly indicates that residues of B. thuringiensis-based insecticides can be found on fresh fruits and vegetables and that these are potentially enterotoxigenic....

  2. Degradation of endogenous and exogenous genes of genetically modified rice with Cry1Ab during food processing.

    Science.gov (United States)

    Zhang, Wei; Xing, Fuguo; Selvaraj, Jonathan Nimal; Liu, Yang

    2014-05-01

    In order to assess the degradation of endogenous and exogenous genes during food processing, genetically modified rice with Cry1Ab was used as raw material to produce 4 processed foods: steamed rice, rice noodles, rice crackers, and sweet rice wine. The results showed various processing procedures caused different degrees of degradation of both endogenous and exogenous genes. During the processing of steamed rice and rice noodles, the procedures were so mild that only genes larger than 1500 bp were degraded, and no degradation of NOS terminator and Hpt gene was detected. For rice crackers, frying was the most severe procedure, followed by microwaving, baking, boiling, 1st drying, and 2nd drying. For sweet rice wine, fermentation had more impact on degradation of genes than the other processing procedures. All procedures in this study did not lead to degradation of genes to below 200 bp, except for NOS terminator. In the case of stability of the genes studied during processing of rice crackers and sweet rice wine, SPS gene was the most, followed by the Cry1Ab gene, Hpt gene, Pubi promoter, and NOS terminator. In our study, we gained some information about the degradation of endogenous and exogenous genes during 4 foods processing, compared the different stabilities between endogenous and exogenous genes, and analyzed different effects of procedure on degradation of genes. In addition, the fragments of endogenous and exogenous genes about 200 bp could be detected in final products, except NOS terminator. As a result, we provided some base information about risk assessment of genetically modified (GM) food and appropriate length of fragment to detect GM component in processed foods. © 2014 Institute of Food Technologists®

  3. The colonization of Bacillus thuringiensis strains in bryophytes

    Czech Academy of Sciences Publication Activity Database

    Lin, Q.; Zhu, P.; Carballar-Lejarazú, R.; Gelbič, Ivan; Guan, X.; Xu, L.; Zhang, L.

    2017-01-01

    Roč. 41, č. 1 (2017), s. 41-48 ISSN 1300-0152 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * GFP * plant colonization Subject RIV: EE - Microbiology, Virology OBOR OECD: Microbiology Impact factor: 1.038, year: 2016 http://journals.tubitak.gov.tr/biology/issues/biy-17-41-1/biy-41-1-5-1510-16.pdf

  4. Profile of cry from native Bacillus thuringiensis isolates and ...

    African Journals Online (AJOL)

    oyaide

    2013-05-29

    May 29, 2013 ... The characterization of 255 Bacillus thuringiensis isolates of Coorg, Sharavatti and BR hills, containing genes known to be active against coleopteran and lepidopteran insect species was done through PCR amplification using the specific and degenerate primers. The isolates were also tested for their.

  5. Effects of Ingesting Bacillus Thuringiensis (Berliner) Spores on ...

    African Journals Online (AJOL)

    Bacillus thuringiensis Berliner was isolated from dead Sesamia calamistis Hampson (Lepidoptera: Noctuidae) larvae collected from maize farms in Cape Coast, Ghana. Spores produced from the vegetative cells were incorporated into an artificial diet and fed to 2nd instar S. calamistis larvae. The duration of larval and pupal ...

  6. Laboratory evaluation of toxicity of Bacillus thuringiensis , neem oil ...

    African Journals Online (AJOL)

    Diamondback moth (DBM), Plutella xylostella (L.), remains a major pest of brassica crops worldwide. Chemical control of this pest remains difficult due to the rapid development of resistance to insecticides and to their effect on natural enemies. The objective of this study was to assess the toxicity of Bacillus thuringiensis (Bt), ...

  7. Bacillus thuringiensis and its application in agriculture | Ali | African ...

    African Journals Online (AJOL)

    Presently, a number of approaches to pest control via genetic engineering have been developed and genetically engineered crops expressing insecticidal characteristics are under cultivation for the last 15 years. Use of Bacillus thuringiensis genes encoding o̅ endotoxins with insecticidal characteristics is the major ...

  8. Evaluation of potency of native Bacillus thuringiensis against maize ...

    African Journals Online (AJOL)

    Bacillus thuringiensis is the most used biological control agent to date. Among major constraints to maize production, safety and hence food sufficiency in Kenya is infestation, damage and contamination by insect pests. Maize grains are adversely damaged by. Prostephanus truncatus which occasionally paves way for the ...

  9. Effects of Ingesting Bacillus Thuringiensis (Berliner) Spores on ...

    African Journals Online (AJOL)

    Effects of Ingesting Bacillus Thuringiensis (Berliner) Spores on Developmental Stages and Fecundity of Surviving Sesamia Calamistis (Hampson) (Lepidoptera: ... The PDF file you selected should load here if your Web browser has a PDF reader plug-in installed (for example, a recent version of Adobe Acrobat Reader).

  10. Screening of Local Bacillus thuringiensis Isolates for Toxicity to ...

    African Journals Online (AJOL)

    Stem borers are a major source of pre-harvest maize crop losses in Kenya and many Sub- Saharan African countries. This menace needs to be addressed if food security is to be realized in this region. Seven local isolates of Bacillus thuringiensis (Bt) strains were isolated from soils collected from Kakamega and Machakos ...

  11. The Impact of Bacillus Thuringiensis Israelensis (Bti) on Adult and ...

    African Journals Online (AJOL)

    In the year 2007, the Ministry of Health (MoH) initiated a larviciding program using Bacillus thuringiensis israelensis (Bti) to mitigate the effects of black fly bites. This study was aimed at assessing the impact of Bti on adult and larvae black fly populations. Baseline data was collected prior to Bti application and after ...

  12. Profile of cry from native Bacillus thuringiensis isolates and ...

    African Journals Online (AJOL)

    The characterization of 255 Bacillus thuringiensis isolates of Coorg, Sharavatti and BR hills, containing genes known to be active against coleopteran and lepidopteran insect species was done through PCR amplification using the specific and degenerate primers. The isolates were also tested for their insecticidal activity ...

  13. Detection of Bacillus thuringiensis genes in transgenic maize by the ...

    African Journals Online (AJOL)

    We optimized the PCR method to detect genetically engineered Bacillus thuringiensis (Bt) maize in open quarantine fields in Kenya. Many factors affect the extraction of the DNA from plants, such as the amount of tissue available, the condition of the plant material, the numbers of steps involved in the extraction procedure, ...

  14. 40 CFR 180.1107 - Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas...

    Science.gov (United States)

    2010-07-01

    ... thuringiensis variety kurstaki encapsulated into killed Pseudomonas fluorescens; exemption from the requirement... killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas fluorescens is exempt from the...

  15. Effects of ensiling of Bacillus thuringiensis (Bt) maize (MON810) on ...

    African Journals Online (AJOL)

    ajl yemi

    2011-11-30

    Nov 30, 2011 ... Cry1Ab protein, neutral detergent fiber, acid detergent fiber, acid detergent lignin and crude protein. Silage from two of the Bt maize varieties had ... Mechanical disruption of cell walls and membranes during ... study was to determine the effects of ensiling Bt maize on concentration of the Cry1Ab protein and ...

  16. Microarray detection and qPCR screening of potential biomarkers of Folsomia candida (Collembola: Isotomidae) exposed to Bt proteins (Cry1Ab and Cry1Ac)

    DEFF Research Database (Denmark)

    Yuan, Yiyang; Krogh, Paul Henning; Bai, Xue

    2014-01-01

    The impact of Bt proteins on non-target arthropods is less understood than their effects on target organisms where the mechanism of toxic action is known. Here, we report the effects of two Bt proteins, Cry1Ab and Cry1Ac, on gene expression in the non-target collembolan, Folsomia candida....... A customized microarray was used to study gene expression in F. candida specimens that were exposed to Cry1Ab and Cry1Ac. All selected transcripts were subsequently confirmed by qPCR. Eleven transcripts were finally verified, and three of them were annotated. The responses of all eleven transcripts were...... tested in specimens for both Cry1Ab and Cry1Ac at a series of concentrations. These transcripts were separated into two and three groups for Cry1Ab and Cry1Ac, respectively, depend on their expression levels. However, those eleven transcripts did not respond to the Bt proteins in Bt-rice residues....

  17. Comparative genomics of extrachromosomal elements in Bacillus thuringiensis subsp. israelensis.

    Science.gov (United States)

    Bolotin, Alexandre; Gillis, Annika; Sanchis, Vincent; Nielsen-LeRoux, Christina; Mahillon, Jacques; Lereclus, Didier; Sorokin, Alexei

    2017-05-01

    Bacillus thuringiensis subsp. israelensis is one of the most important microorganisms used against mosquitoes. It was intensively studied following its discovery and became a model bacterium of the B. thuringiensis species. Those studies focused on toxin genes, aggregation-associated conjugation, linear genome phages, etc. Recent announcements of genomic sequences of different strains have not been explicitly related to the biological properties studied. We report data on plasmid content analysis of four strains using ultra-high-throughput sequencing. The strains were commercial product isolates, with their putative ancestor and type B. thuringiensis subsp. israelensis strain sequenced earlier. The assembled contigs corresponding to published and novel data were assigned to plasmids described earlier in B. thuringiensis subsp. israelensis and other B. thuringiensis strains. A new 360 kb plasmid was identified, encoding multiple transporters, also found in most of the earlier sequenced strains. Our genomic data show the presence of two toxin-coding plasmids of 128 and 100 kb instead of the reported 225 kb plasmid, a co-integrate of the former two. In two of the sequenced strains, only a 100 kb plasmid was present. Some heterogeneity exists in the small plasmid content and structure between strains. These data support the perception of active plasmid exchange among B. thuringiensis subsp. israelensis strains in nature. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  18. A Comprehensive Assessment of the Effects of Transgenic Cry1Ac/Cry1Ab Rice Huahui 1 on Adult Micraspis discolor (Fabricius (Coleoptera: Coccinellidae.

    Directory of Open Access Journals (Sweden)

    Xia Zhou

    Full Text Available Micraspis discolor (Fabricius (Coleoptera: Coccinellidae is a widely distributed coleoptera predator in southern Asia in rice ecosystem, and adult M. discolor feed on both rice pollen and soft-bodied arthropods. Bitrophic bioassay and tritrophic bioassay were conducted to evaluate the potential impact of Cry1Ac/Cry1Ab-expressing rice Huahui 1 and its non-transgenic counterpart Minghui 63 on fitness parameters of adult M. discolor. The results showed that the survival, and fecundity of this beetle' adults were not different when they fed on Bt rice or non-Bt rice pollen or Nilaparvata lugens (Stål reared on Bt rice or non-Bt rice. Toxicity assessment to ensure M. discolor adults were not sensitive to Cry1Ab or Cry1Ac protein independent from the pollen background, M. discolor adults were fed with an artificial diet containing Cry1Ac, Cry1Ab or both protein approximately 10 times higher concentration than in Huahui 1 rice pollen. No difference was detected for any of the life-table parameters tested between Cry protein-containing and pure diet. Artificial diet containing E-64 (N-(trans-Epoxysuccinyl-L-leucine 4-guanidinobutylamide was included as a positive control. In contrast, the pre-oviposition and fecundity of M. discolor were significantly adversely affected by feeding on E-64-containing diet. In both bioassays, the uptakes of Cry protein by adult M. discolor were tested by ELISA measurements. These results indicated that adults of M. discolor are not affected by Cry1Ab- or Cry1Ac-expressing rice pollen and are not sensitive to Cry protein at concentrations exceeding the levels in rice pollen in Huahui1. This suggests that M. discolor adults would not be harmed by Cry1Ac/Cry1Ab rice if Bt rice Huahui 1 were commercialized.

  19. A Comprehensive Assessment of the Effects of Transgenic Cry1Ac/Cry1Ab Rice Huahui 1 on Adult Micraspis discolor (Fabricius) (Coleoptera: Coccinellidae).

    Science.gov (United States)

    Zhou, Xia; Guo, Yunling; Kong, Hua; Zuo, Jiao; Huang, Qixing; Jia, Ruizong; Guo, Anping; Xu, Lin

    2016-01-01

    Micraspis discolor (Fabricius) (Coleoptera: Coccinellidae) is a widely distributed coleoptera predator in southern Asia in rice ecosystem, and adult M. discolor feed on both rice pollen and soft-bodied arthropods. Bitrophic bioassay and tritrophic bioassay were conducted to evaluate the potential impact of Cry1Ac/Cry1Ab-expressing rice Huahui 1 and its non-transgenic counterpart Minghui 63 on fitness parameters of adult M. discolor. The results showed that the survival, and fecundity of this beetle' adults were not different when they fed on Bt rice or non-Bt rice pollen or Nilaparvata lugens (Stål) reared on Bt rice or non-Bt rice. Toxicity assessment to ensure M. discolor adults were not sensitive to Cry1Ab or Cry1Ac protein independent from the pollen background, M. discolor adults were fed with an artificial diet containing Cry1Ac, Cry1Ab or both protein approximately 10 times higher concentration than in Huahui 1 rice pollen. No difference was detected for any of the life-table parameters tested between Cry protein-containing and pure diet. Artificial diet containing E-64 (N-(trans-Epoxysuccinyl)-L-leucine 4-guanidinobutylamide) was included as a positive control. In contrast, the pre-oviposition and fecundity of M. discolor were significantly adversely affected by feeding on E-64-containing diet. In both bioassays, the uptakes of Cry protein by adult M. discolor were tested by ELISA measurements. These results indicated that adults of M. discolor are not affected by Cry1Ab- or Cry1Ac-expressing rice pollen and are not sensitive to Cry protein at concentrations exceeding the levels in rice pollen in Huahui1. This suggests that M. discolor adults would not be harmed by Cry1Ac/Cry1Ab rice if Bt rice Huahui 1 were commercialized.

  20. A Comprehensive Assessment of the Effects of Transgenic Cry1Ac/Cry1Ab Rice Huahui 1 on Adult Micraspis discolor (Fabricius) (Coleoptera: Coccinellidae)

    Science.gov (United States)

    Zhou, Xia; Guo, Yunling; Kong, Hua; Zuo, Jiao; Huang, Qixing; Jia, Ruizong; Guo, Anping; Xu, Lin

    2016-01-01

    Micraspis discolor (Fabricius) (Coleoptera: Coccinellidae) is a widely distributed coleoptera predator in southern Asia in rice ecosystem, and adult M. discolor feed on both rice pollen and soft-bodied arthropods. Bitrophic bioassay and tritrophic bioassay were conducted to evaluate the potential impact of Cry1Ac/Cry1Ab-expressing rice Huahui 1 and its non-transgenic counterpart Minghui 63 on fitness parameters of adult M. discolor. The results showed that the survival, and fecundity of this beetle’ adults were not different when they fed on Bt rice or non-Bt rice pollen or Nilaparvata lugens (Stål) reared on Bt rice or non-Bt rice. Toxicity assessment to ensure M. discolor adults were not sensitive to Cry1Ab or Cry1Ac protein independent from the pollen background, M. discolor adults were fed with an artificial diet containing Cry1Ac, Cry1Ab or both protein approximately 10 times higher concentration than in Huahui 1 rice pollen. No difference was detected for any of the life-table parameters tested between Cry protein-containing and pure diet. Artificial diet containing E-64 (N-(trans-Epoxysuccinyl)-L-leucine 4-guanidinobutylamide) was included as a positive control. In contrast, the pre-oviposition and fecundity of M. discolor were significantly adversely affected by feeding on E-64-containing diet. In both bioassays, the uptakes of Cry protein by adult M. discolor were tested by ELISA measurements. These results indicated that adults of M. discolor are not affected by Cry1Ab- or Cry1Ac-expressing rice pollen and are not sensitive to Cry protein at concentrations exceeding the levels in rice pollen in Huahui1. This suggests that M. discolor adults would not be harmed by Cry1Ac/Cry1Ab rice if Bt rice Huahui 1 were commercialized. PMID:26914608

  1. Bacillus thuringiensis and its application in agriculture

    African Journals Online (AJOL)

    USER

    2010-04-05

    Apr 5, 2010 ... Presently, a number of approaches to pest control via genetic engineering have been developed and genetically engineered crops ... of this bacterium, which are effective against Lepidop-. *Corresponding author. E-mail: .... provide a useful tool for management of resistance to B. thuringiensis toxins ...

  2. Phages Preying on Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis: Past, Present and Future

    Science.gov (United States)

    Gillis, Annika; Mahillon, Jacques

    2014-01-01

    Many bacteriophages (phages) have been widely studied due to their major role in virulence evolution of bacterial pathogens. However, less attention has been paid to phages preying on bacteria from the Bacillus cereus group and their contribution to the bacterial genetic pool has been disregarded. Therefore, this review brings together the main information for the B. cereus group phages, from their discovery to their modern biotechnological applications. A special focus is given to phages infecting Bacillus anthracis, B. cereus and Bacillus thuringiensis. These phages belong to the Myoviridae, Siphoviridae, Podoviridae and Tectiviridae families. For the sake of clarity, several phage categories have been made according to significant characteristics such as lifestyles and lysogenic states. The main categories comprise the transducing phages, phages with a chromosomal or plasmidial prophage state, γ-like phages and jumbo-phages. The current genomic characterization of some of these phages is also addressed throughout this work and some promising applications are discussed here. PMID:25010767

  3. Phages Preying on Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis: Past, Present and Future

    Directory of Open Access Journals (Sweden)

    Annika Gillis

    2014-07-01

    Full Text Available Many bacteriophages (phages have been widely studied due to their major role in virulence evolution of bacterial pathogens. However, less attention has been paid to phages preying on bacteria from the Bacillus cereus group and their contribution to the bacterial genetic pool has been disregarded. Therefore, this review brings together the main information for the B. cereus group phages, from their discovery to their modern biotechnological applications. A special focus is given to phages infecting Bacillus anthracis, B. cereus and Bacillus thuringiensis. These phages belong to the Myoviridae, Siphoviridae, Podoviridae and Tectiviridae families. For the sake of clarity, several phage categories have been made according to significant characteristics such as lifestyles and lysogenic states. The main categories comprise the transducing phages, phages with a chromosomal or plasmidial prophage state, γ-like phages and jumbo-phages. The current genomic characterization of some of these phages is also addressed throughout this work and some promising applications are discussed here.

  4. Bacillus thuringiensis HD-1 Cry- : development of a safe, non-insecticidal simulant for Bacillus anthracis.

    Science.gov (United States)

    Bishop, A H; Robinson, C V

    2014-09-01

    A representative simulant for spores of Bacillus anthracis is needed for field testing. Bacillus thuringiensis is gaining recognition as a suitable organism. A strain that does not form the insecticidal, parasporal crystals that are characteristic of this species is a more accurate physical representative of B. anthracis spores. We developed noninsecticidal derivatives of two isolates of B. thuringiensis HD-1. Two plasmid-cured derivatives of B. thuringiensis HD-1, unable to make crystal toxins ('Cry(-) '), were isolated. These isolates and the existing Cry(-) strain, B. thuringiensis Al Hakam, were probed with PCR assays against the known insecticidal genes cry, vip and cyt. Their genomic DNA was sequenced to demonstrate a lack of insecticidal genes. This was confirmed by bioassays against a number of invertebrate species. Real-time PCR assays were developed to identify the B. thuringiensis HD-1 Cry(-) derivatives and an effective differential and selective medium was assessed. All three Cry(-) isolates are devoid of known insecticidal determinants. The B. thuringiensis HD-1 Cry(-) derivatives can easily be recovered from soil and identified by PCR with some selectivity. The B. thuringiensis HD-1 Cry(-) derivatives represent accurate, nongenetically manipulated simulants for B. anthracis with excellent human and environmental safety records. © 2014 Crown Copyright. Journal of Applied Microbiology © 2014 Society for Applied Microbiology This article is published with the permission of the Controller of HMSO and the Queen's Printer for Scotland.

  5. Bioaccumulation of Cry1Ab protein from an herbivore reduces anti-oxidant enzyme activities in two spider species.

    Directory of Open Access Journals (Sweden)

    Ji Zhou

    Full Text Available Cry proteins are expressed in rice lines for lepidopteran pest control. These proteins can be transferred from transgenic rice plants to non-target arthropods, including planthoppers and then to a predatory spider. Movement of Cry proteins through food webs may reduce fitness of non-target arthropods, although recent publications indicated no serious changes in non-target populations. Nonetheless, Cry protein intoxication influences gene expression in Cry-sensitive insects. We posed the hypothesis that Cry protein intoxication influences enzyme activities in spiders acting in tri-trophic food webs. Here we report on the outcomes of experiments designed to test our hypothesis with two spider species. We demonstrated that the movement of CryAb protein from Drosophila culture medium into fruit flies maintained on the CryAb containing medium and from the flies to the spiders Ummeliata insecticeps and Pardosa pseudoannulata. We also show that the activities of three key metabolic enzymes, acetylcholine esterase (AchE, glutathione peroxidase (GSH-Px, and superoxide dismutase (SOD were significantly influenced in the spiders after feeding on Cry1Ab-containing fruit flies. We infer from these data that Cry proteins originating in transgenic crops impacts non-target arthropods at the physiological and biochemical levels, which may be one mechanism of Cry protein-related reductions in fitness of non-target beneficial predators.

  6. Photoprotection of Bacillus thuringiensis kurstaki from ultraviolet irradiation

    International Nuclear Information System (INIS)

    Cohen, E.; Rozen, H.; Joseph, T.; Braun, S.; Margulies, L.

    1991-01-01

    Irradiation of Bacillus thuringiensis var. kurstaki HD1 at 300-350 nm for up to 12 hr using a photochemical reactor results in a rapid loss of its toxicity to larvae of Heliothis armigera. Photoprotection of the toxic component was obtained by adsorption of cationic chromophores such as acriflavin (AF), methyl green, and rhodamine B to B. thuringiensis. AF gave the best photoprotection and a level of 0.42 mmol/g dye absorbed per gram of B. thuringiensis was highly toxic even after 12 hr of ultraviolet (uv) irradiation as compared to the control (77.5 and 5% of insect mortality, respectively). Ultraviolet and Fourier-transform infrared spectroscopic studies indicate molecular interactions between B. thuringiensis and AF. The nature of these interactions and energy or charge transfer as possible mechanisms of photoprotection are discussed. It is speculated that tryptophan residues are essential for the toxic effect of B. thuringiensis. It is suggested that photoprotection is attained as energy is transferred from the excited tryptophan moieties to the chromophore molecules

  7. SinR controls enterotoxin expression in Bacillus thuringiensis biofilms.

    Directory of Open Access Journals (Sweden)

    Annette Fagerlund

    Full Text Available The entomopathogen Bacillus thuringiensis produces dense biofilms under various conditions. Here, we report that the transition phase regulators Spo0A, AbrB and SinR control biofilm formation and swimming motility in B. thuringiensis, just as they control biofilm formation and swarming motility in the closely related saprophyte species B. subtilis. However, microarray analysis indicated that in B. thuringiensis, in contrast to B. subtilis, SinR does not control an eps operon involved in exopolysaccharides production, but regulates genes involved in the biosynthesis of the lipopeptide kurstakin. This lipopeptide is required for biofilm formation and was previously shown to be important for survival in the host cadaver (necrotrophism. Microarray analysis also revealed that the SinR regulon contains genes coding for the Hbl enterotoxin. Transcriptional fusion assays, Western blots and hemolysis assays confirmed that SinR controls Hbl expression, together with PlcR, the main virulence regulator in B. thuringiensis. We show that Hbl is expressed in a sustained way in a small subpopulation of the biofilm, whereas almost all the planktonic population transiently expresses Hbl. The gene coding for SinI, an antagonist of SinR, is expressed in the same biofilm subpopulation as hbl, suggesting that hbl transcription heterogeneity is SinI-dependent. B. thuringiensis and B. cereus are enteric bacteria which possibly form biofilms lining the host intestinal epithelium. Toxins produced in biofilms could therefore be delivered directly to the target tissue.

  8. [Bioconversion of sewage sludge to biopesticide by Bacillus thuringiensis].

    Science.gov (United States)

    Chang, Ming; Zhou, Shun-gui; Lu, Na; Ni, Jin-ren

    2006-07-01

    Feasibility of bioconversion of sewage sludge to biopesticide by Bacillus thuringiensis was studied using sewage sludge as a raw material. The fermentation was also compared with conventional medium. Results showed that without any pretreatment, the nutrients contained in sewage sludge were almost sufficient for Bacillus thuringiensis growth, even with a rapid multiplicational rate. Higher viable cells and viable spores values were obtained earlier at 24 h, with 9.48 x 10(8) CFU x mL(-1) and 8.51 x 10(8) CFU x mL(-1) respectively, which was 12 hours earlier and nearly 20 percent higher than conventional medium. SEM of 36 h samples gave a clear phenomenon that the metabolizability in sludge was much faster with spores and crystals spreading around. The crystals in sludge seemed rather bigger and more regular. Also a better crystal protein yield of 2.80 mg x mL(-1) was observed in sludge medium compared to conventional medium at the end of fermentation. Sludge fermentation for Bacillus thuringiensis reduces the producing cost, and gives better fermentation capabilities. It's expected to be a new method for sludge disposal.

  9. Down-regulation of aminopeptidase N and ABC transporter subfamily G transcripts in Cry1Ab and Cry1Ac resistant Asian corn borer, Ostrina furnacalis (Lepidoptera: Crambidae)

    Science.gov (United States)

    Bacillus thuringiensis (Bt) crystalline protein (Cry) toxins cause mortality by a mechanism involving pore formation or signal transduction following toxin binding to receptors along the midgut lumen of susceptible insects, but this mechanism and mutations therein that lead to resistance remain poor...

  10. 40 CFR 174.517 - Bacillus thuringiensis Cry9C protein in corn; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry9C protein... PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.517 Bacillus thuringiensis... Bacillus thuringiensis Cry9C protein in corn is exempted from the requirement of a tolerance for residues...

  11. 40 CFR 174.509 - Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry3A protein...-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.509 Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry3A protein are exempted...

  12. 40 CFR 174.520 - Bacillus thuringiensis Cry1F protein in corn; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1F protein... PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.520 Bacillus thuringiensis Cry1F protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis...

  13. 40 CFR 174.502 - Bacillus thuringiensis Cry1A.105 protein; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1A.105...-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.502 Bacillus thuringiensis Cry1A.105 protein; exemption from the requirement of a tolerance. (a) Residues of Bacillus thuringiensis Cry1A.105 protein in...

  14. 40 CFR 174.530 - Bacillus thuringiensis Cry2Ae protein in cotton; temporary exemption from the requirement of a...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry2Ae protein... Bacillus thuringiensis Cry2Ae protein in or on the food commodities of cotton, cotton; cotton, undelinted... byproducts are exempt temporarily from the requirement of a tolerance when Bacillus thuringiensis Cry2Ae...

  15. First application of a microsphere-based immunoassay to the detection of genetically modified organisms (GMOs): quantification of Cry1Ab protein in genetically modified maize.

    Science.gov (United States)

    Fantozzi, Anna; Ermolli, Monica; Marini, Massimiliano; Scotti, Domenico; Balla, Branko; Querci, Maddalena; Langrell, Stephen R H; Van den Eede, Guy

    2007-02-21

    An innovative covalent microsphere immunoassay, based on the usage of fluorescent beads coupled to a specific antibody, was developed for the quantification of the endotoxin Cry1Ab present in MON810 and Bt11 genetically modified (GM) maize lines. In particular, a specific protocol was developed to assess the presence of Cry1Ab in a very broad range of GM maize concentrations, from 0.1 to 100% [weight of genetically modified organism (GMO)/weight]. Test linearity was achieved in the range of values from 0.1 to 3%, whereas fluorescence signal increased following a nonlinear model, reaching a plateau at 25%. The limits of detection and quantification were equal to 0.018 and 0.054%, respectively. The present study describes the first application of quantitative high-throughput immunoassays in GMO analysis.

  16. 40 CFR 180.1154 - CryIA(c) and CryIC derived delta-endotoxins of Bacillus thuringiensis var. kurstaki encapsulated...

    Science.gov (United States)

    2010-07-01

    ...-endotoxins of Bacillus thuringiensis var. kurstaki encapsulated in killed Pseudomonas fluorescens, and the... Bacillus thuringiensis var. kurstaki encapsulated in killed Pseudomonas fluorescens, and the expression... thuringiensis var. kurstaki encapsulated in killed Pseudomonas fluorescens and the expression plasmid and...

  17. SR450 and Superhawk XP applications of Bacillus thuringiensis israelensis de Barjac against Culex quinquefasciatus Say

    Science.gov (United States)

    Sprayer comparisons and larval morality assays were conducted following SR450 backpack mist blower and Superhawk XP thermal fogger applications of Vectobac® WDG Bacillus thuringiensis israelensis (Bti) de Barjac against Culex quinquefasciatus Say. Bacillus thuringiensis israelensis was applied at m...

  18. A Fast and Sensitive Quantitative Lateral Flow Immunoassay for Cry1Ab Based on a Novel Signal Amplification Conjugate

    Directory of Open Access Journals (Sweden)

    Jian Wu

    2012-08-01

    Full Text Available A novel lateral flow immunoassay (LFIA signal amplification strategy for the detection of Cry1Ab based on amplification via a polylysine (PL chain and biotin-streptavidin system (BSAS is described. In this system, multiple fluorescence dyes (FL were directly coated on the surface of PL and conjugated with antibody via the BSAS for construction of novel signal amplification (FLPL-BSAS-mAb1 conjugates, in which FL, PL and BSAS were employed to improve the sensitivity of LFIA. Compared with conventional LFIA, the sensitivity of FLPL-BSAS-mAb1-based LFIA was increased by approximately 100-fold. Quantified linearity was achieved in the value range of 0–1,000 pg/mL. The limit of detection (LOD was reached 10 pg/mL after optimization of reaction conditions. To our knowledge, this represents one of the most sensitive LFIA for Cry1Ab yet reported. Furthermore, the detection time for this method was about 10 min. Therefore, it should be an attractive alternative compared to conventional immunoassays in routine control for Cry1Ab.

  19. Midgut microbiota and host immunocompetence underlie Bacillus thuringiensis killing mechanism

    Science.gov (United States)

    Caccia, Silvia; Di Lelio, Ilaria; La Storia, Antonietta; Marinelli, Adriana; Varricchio, Paola; Franzetti, Eleonora; Banyuls, Núria; Tettamanti, Gianluca; Casartelli, Morena; Giordana, Barbara; Ferré, Juan; Gigliotti, Silvia; Pennacchio, Francesco

    2016-01-01

    Bacillus thuringiensis is a widely used bacterial entomopathogen producing insecticidal toxins, some of which are expressed in insect-resistant transgenic crops. Surprisingly, the killing mechanism of B. thuringiensis remains controversial. In particular, the importance of the septicemia induced by the host midgut microbiota is still debated as a result of the lack of experimental evidence obtained without drastic manipulation of the midgut and its content. Here this key issue is addressed by RNAi-mediated silencing of an immune gene in a lepidopteran host Spodoptera littoralis, leaving the midgut microbiota unaltered. The resulting cellular immunosuppression was characterized by a reduced nodulation response, which was associated with a significant enhancement of host larvae mortality triggered by B. thuringiensis and a Cry toxin. This was determined by an uncontrolled proliferation of midgut bacteria, after entering the body cavity through toxin-induced epithelial lesions. Consequently, the hemolymphatic microbiota dramatically changed upon treatment with Cry1Ca toxin, showing a remarkable predominance of Serratia and Clostridium species, which switched from asymptomatic gut symbionts to hemocoelic pathogens. These experimental results demonstrate the important contribution of host enteric flora in B. thuringiensis-killing activity and provide a sound foundation for developing new insect control strategies aimed at enhancing the impact of biocontrol agents by reducing the immunocompetence of the host. PMID:27506800

  20. Bacillus thuringiensis membrane-damaging toxins acting on mammalian cells.

    Science.gov (United States)

    Celandroni, Francesco; Salvetti, Sara; Senesi, Sonia; Ghelardi, Emilia

    2014-12-01

    Bacillus thuringiensis is widely used as a biopesticide in forestry and agriculture, being able to produce potent species-specific insecticidal toxins and considered nonpathogenic to other animals. More recently, however, repeated observations are documenting the association of this microorganism with various infectious diseases in humans, such as food-poisoning-associated diarrheas, periodontitis, bacteremia, as well as ocular, burn, and wound infections. Similar to B. cereus, B. thuringiensis produces an array of virulence factors acting against mammalian cells, such as phosphatidylcholine- and phosphatidylinositol-specific phospholipase C (PC-PLC and PI-PLC), hemolysins, in particular hemolysin BL (HBL), and various enterotoxins. The contribution of some of these toxins to B. thuringiensis pathogenicity has been studied in animal models of infection, following intravitreous, intranasal, or intratracheal inoculation. These studies lead to the speculation that the activities of PC-PLC, PI-PLC, and HBL are responsible for most of the pathogenic properties of B. thuringiensis in nongastrointestinal infections in mammals. This review summarizes data regarding the biological activity, the genetic basis, and the structural features of these membrane-damaging toxins. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  1. Methodology for fast evaluation of Bacillus thuringiensis crystal protein content

    Directory of Open Access Journals (Sweden)

    Alves Lúcia M. Carareto

    2000-01-01

    Full Text Available The development of the production and use of Bacillus thuringiensis in Brazil at a commercial scale faces certain difficulties, among them the establishment of efficient methodologies for the quantitation of toxic products to be commercialized. Presently, the amount of toxin is given in percentage by analyzing the samples total protein content. Such methodology however, does not measure the actual amount of active protein present in the product, since most strains express different endotoxin genes and might even produce b-toxin. Since the various types of toxins exhibit different antigenic characteristics, this work has as objective the utilization of fast immunological techniques to quantify the level of crystal protein. Crystal protein produced by a subspecies of Bacillus thuringiensis var. israelensis was purified by ultracentrifugation and utilized to immunize rabbits and to produce hiperimmune sera. Such sera were latter used to evaluate the level of proteins on commercial bioinsecticide and on laboratory cultures of B. thuringiensis through the immunodot technique. The results were obtained by comparison of data obtained from reactions with known concentrations of crystal protein permitting to evaluate the level of such protein on various materials.

  2. Multilocus sequence analysis of Bacillus thuringiensis serovars navarrensis, bolivia and vazensis and Bacillus weihenstephanensis reveals a common phylogeny.

    Science.gov (United States)

    Soufiane, Brahim; Baizet, Mathilde; Côté, Jean-Charles

    2013-01-01

    The Bacillus cereus group sensu lato includes six closely-related bacterial species: Bacillus cereus, Bacillus anthracis, Bacillus thuringiensis, Bacillus mycoides, Bacillus pseudomycoides and Bacillus weihenstephanensis. B. thuringiensis is distinguished from the other species mainly by the appearance of an inclusion body upon sporulation. B. weihenstephanensis is distinguished based on its psychrotolerance and the presence of specific signature sequences in the 16S rRNA gene and cspA genes. A total of seven housekeeping genes (glpF, gmK, ilvD, pta, purH, pycA and tpi) from different B. thuringiensis serovars and B. weihenstephanensis strains were amplified and their nucleotide sequences determined. A maximum likelihood phylogenetic tree was inferred from comparisons of the concatenated sequences. B. thuringiensis serovars navarrensis, bolivia and vazensis clustered not with the other B. thuringiensis serovars but rather with the B. weihenstephanensis strains, indicative of a common phylogeny. In addition, specific signature sequences and single nucleotide polymorphisms common to B. thuringiensis serovars navarrensis, bolivia and vazensis and the B. weihenstephanensis strains, and absent in the other B. thuringiensis serovars, were identified.

  3. Construction of Bacillus thuringiensis Simulant Strains Suitable for Environmental Release.

    Science.gov (United States)

    Park, Sangjin; Kim, Changhwan; Lee, Daesang; Song, Dong Hyun; Cheon, Ki Cheol; Lee, Hong Suk; Kim, Seong Joo; Kim, Jee Cheon; Lee, Sang Yup

    2017-05-01

    For a surrogate bacterium to be used in outdoor studies, it is important to consider environmental and human safety and ease of detection. Recently, Bacillus thuringiensis , a popular bioinsecticide bacterium, has been gaining attention as a surrogate bacterium for use in biodefense. In this study, we constructed simulant strains of B. thuringiensis with enhanced characteristics for environmental studies. Through transposon mutagenesis, pigment genes were inserted into the chromosome, producing yellow-colored colonies for easy detection. To prevent persistence of spores in the environment, a genetic circuit was designed to produce a spore without sporulation capability. Two loxP sites were inserted, one on each side of the spo0A gene, which encodes a sporulation master regulator, and a sporulation-dependent Cre expression cassette was inserted into the chromosome. This genetic circuit successfully deleted spo0A during sporulation, producing spores that lacked the spo0A gene. In addition, two major α/β-type small acid-soluble spore protein (SASP) genes, predicted by synteny analysis, were deleted. The spores of the mutant strain showed increased UV-C sensitivity and quickly lost viability when tested in a solar simulator. When the spores of the mutant strain were administered to the lungs of BALB/c mice, cells were quickly removed from the body, suggesting enhanced in vivo safety. All strains constructed in this study contain no antibiotic resistance markers and all heterologous genes were inserted into the chromosome, which are useful features for simulants to be released into the environment. IMPORTANCE B. thuringiensis has recently been receiving increasing attention as a good spore simulant in biodefense research. However, few studies were done to properly address many important features of B. thuringiensis as a simulant in environmental studies. Since spores can persist in the environment for years after release, environmental contamination is a big problem

  4. Chitinase production by Bacillus thuringiensis and Bacillus licheniformis: their potential in antifungal biocontrol.

    Science.gov (United States)

    Gomaa, Eman Zakaria

    2012-02-01

    Thirty bacterial strains were isolated from the rhizosphere of plants collected from Egypt and screened for production of chitinase enzymes. Bacillus thuringiensis NM101-19 and Bacillus licheniformis NM120-17 had the highest chitinolytic activities amongst those investigated. The production of chitinase by B. thuringiensis and B. licheniformis was optimized using colloidal chitin medium amended with 1.5% colloidal chitin, with casein as a nitrogen source, at 30°C after five days of incubation. An enhancement of chitinase production by the two species was observed by addition of sugar substances and dried fungal mats to the colloidal chitin media. The optimal conditions for chitinase activity by B. thuringiensis and B. licheniformis were at 40°C, pH 7.0 and pH 8.0, respectively. Na(+), Mg(2+), Cu(2+), and Ca(2+) caused enhancement of enzyme activities whereas they were markedly inhibited by Zn(2+), Hg(2+), and Ag(+). In vitro, B. thuringiensis and B. licheniformis chitinases had potential for cell wall lysis of many phytopathogenic fungi tested. The addition of B. thuringiensis chitinase was more effective than that of B. licheniformis in increasing the germination of soybean seeds infected with various phytopathogenic fungi.

  5. [Isolation of Bacillus thuringiensis IMV B-7324 fibrinolytic peptidase].

    Science.gov (United States)

    Nidialkova, N A; Matseliukh, O V; Varbanets', L D

    2012-01-01

    Fibrinolytic peptidase of Bacillus thuringiensis IMV B-7324 was isolated by ammonium sulfate fractionation, gel-filtration and ion exchange chromatography on TSK-gels--Toyopearl HW-55 and DEAE 650 (M). Fibrinolytic activity of the purified enzyme was 87.9 U/mg of protein that was 19.9 times higher compared with the supernatant cultural liquid, the yield on its activity reached 31%. The gel-filtration on Sepharose 6B and by SDS-PAGE electrophoresis demonstrated the homogeneity of the purified fibrinolytic peptidase, which molecular weight was approximately 24 kDa.

  6. Bacillus thuringiensis subsp. israelensis and Its Dipteran-Specific Toxins

    Science.gov (United States)

    Ben-Dov, Eitan

    2014-01-01

    Bacillus thuringiensis subsp. israelensis (Bti) is the first Bacillus thuringiensis to be found and used as an effective biological control agent against larvae of many mosquito and black fly species around the world. Its larvicidal activity resides in four major (of 134, 128, 72 and 27 kDa) and at least two minor (of 78 and 29 kDa) polypeptides encoded respectively by cry4Aa, cry4Ba, cry11Aa, cyt1Aa, cry10Aa and cyt2Ba, all mapped on the 128 kb plasmid known as pBtoxis. These six δ-endotoxins form a complex parasporal crystalline body with remarkably high, specific and different toxicities to Aedes, Culex and Anopheles larvae. Cry toxins are composed of three domains (perforating domain I and receptor binding II and III) and create cation-selective channels, whereas Cyts are composed of one domain that acts as well as a detergent-like membrane perforator. Despite the low toxicities of Cyt1Aa and Cyt2Ba alone against exposed larvae, they are highly synergistic with the Cry toxins and hence their combinations prevent emergence of resistance in the targets. The lack of significant levels of resistance in field mosquito populations treated for decades with Bti-bioinsecticide suggests that this bacterium will be an effective biocontrol agent for years to come. PMID:24686769

  7. Bacillus thuringiensis subsp. israelensis and Its Dipteran-Specific Toxins

    Directory of Open Access Journals (Sweden)

    Eitan Ben-Dov

    2014-03-01

    Full Text Available Bacillus thuringiensis subsp. israelensis (Bti is the first Bacillus thuringiensis to be found and used as an effective biological control agent against larvae of many mosquito and black fly species around the world. Its larvicidal activity resides in four major (of 134, 128, 72 and 27 kDa and at least two minor (of 78 and 29 kDa polypeptides encoded respectively by cry4Aa, cry4Ba, cry11Aa, cyt1Aa, cry10Aa and cyt2Ba, all mapped on the 128 kb plasmid known as pBtoxis. These six δ-endotoxins form a complex parasporal crystalline body with remarkably high, specific and different toxicities to Aedes, Culex and Anopheles larvae. Cry toxins are composed of three domains (perforating domain I and receptor binding II and III and create cation-selective channels, whereas Cyts are composed of one domain that acts as well as a detergent-like membrane perforator. Despite the low toxicities of Cyt1Aa and Cyt2Ba alone against exposed larvae, they are highly synergistic with the Cry toxins and hence their combinations prevent emergence of resistance in the targets. The lack of significant levels of resistance in field mosquito populations treated for decades with Bti-bioinsecticide suggests that this bacterium will be an effective biocontrol agent for years to come.

  8. Cry1Ab protein quantification in leaves, stems and grains, and effectiveness to control Spodoptera frugiperda and Helicoverpa zea on two hybrids of genetically modified corn

    Directory of Open Access Journals (Sweden)

    Geraldo Balieiro Neto

    2013-01-01

    Full Text Available A study was carried out to evaluate the infestation and associated damages to the presence of the Spodoptera frugiperda and Helicoverpa zea caterpillars, in two genetically modified (GM corn, Dekalb DKB390 and Agroceres AG8088, expressing the cry1Ab protein. For this objective, an split-splot design with two factors (hybrid x gene was carried out. Negative controls were made with the same corn hybrids without the gene cry1Ab (NoGM. The concentration of the protein Cry1Ab was determined by the ELISA (enzyme linked immuno sorbent assay technique in previously dehydrated stems, leaves and grains of GM corns. Caterpillars sampling of S. frugiperda and associated damage survey were accomplished at 15, 22, 29, 36 and 42 days after the sowing, according to a damage scale with 5 levels (0- pest absence to 5- dead plant. Countings of H. zea caterpillars and associated damage were assessed at 57, 71, 78 and 85 days after the sowing, according to a damage scale with 4 levels (0-pest absence curse to 4-gallery in the corn cob minor than 3cm. Sampled caterpillars were divided in two groups, smaller or equal to 15mm and bigger than 15mm. No insecticide application was accomplished in the GM blocks while NoGM blocks were sprayed with deltametrina (2,8%, 42 days after the sowing. The infestation level and associated damage due to S. frugiperda presence was significantly smaller (p < 0,05 in the GM corns in comparison to NoGM corns. Nevertheless, the number and associated damage of S. frugiperda caterpillars, smaller than 15 mm, were superior in the GM DKB390 corn when compared to the GM AG8088 corn. Differences were not observed in the S. frugiperda infestation and associated damage between GM corns and between NoGM corns. On average, the concentration of Cry1Ab protein was significantly superior in leaves and stems in comparison to the grain and, usually, superior in the GM AG8088 corn comparatively to GM DKB390 corn. No differences were found on level damages

  9. Molluscicidal activity of Bacillus thuringiensis strains against Biomphalaria alexandrina snails

    Directory of Open Access Journals (Sweden)

    Amany M. Abd El-Ghany

    2017-12-01

    Full Text Available Schistosomiasis is a parasitic disease transmitted to man and different warm blooded animals by means of snails. Great effort has been made to control the transmission of the disease by many strategies. Consequently, the utilization of particular molluscicides is viewed as a standout amongst the best measures for molluscs control. Recently, microbial pathogen used as non-traditional molluscicides which have attracted significant research attention due to the increasing, worldwide development of resistance to chemical molluscicides in molluscs populations. The present work aimed to study the molluscicidal impacts of eleven isolates of Bacillus thuringiensis which were isolated from soils of six Egyptian governorates toward Biomphalaria alexandrina snails. Results showed that, B. thuringiensis provides an effective biological control agent against B. alexandrina snails. Out of the tested isolates, four isolates; Qalyubia, Asyut 1, Qena and North Sinai 2 isolates show high-level molluscicidal activity. The obtained results indicated that LC50 and LC90 values were ranged between 133.27–457.74 mg/mL and 270.32–781.05 mg/mL, respectively. The most noteworthy molluscicidal impact was displayed by Qalyubia isolate which isolated from Qalyubia governorate with mortality rate extended from 20% to100% at five treatment concentrations of 100–500 mg/mL. The LC50 and LC90 values for Qalyubia isolate were 133.27 mg/mL and 270.32 mg/mL, respectively. Keywords: Biomphalaria alexandrina, Bacillus thuringiensis, Molluscicidal activity, Biological control

  10. Bitrophic and Tritrophic Effects of Transgenic cry1Ab/cry2Aj Maize on the Beneficial, Nontarget Harmonia axyridis (Coleoptera: Coccinellidae).

    Science.gov (United States)

    Chang, Xue; Lu, Zengbin; Shen, Zhicheng; Peng, Yufa; Ye, Gongyin

    2017-10-01

    Harmonia axyridis (Pallas) is a common and abundant predator in China and may be exposed to Cry toxins that are produced in Bt crops either by feeding on plant parts or by feeding on target or nontarget herbivorous insects. A new Bt maize line, expressing the Cry1Ab/Cry2Aj fused protein, has been developed and should be rigorously assessed for the ecological risks on the natural enemy. Laboratory experiments were carried out to study the effects of this Bt maize on nontarget predator H. axyridis via bitrophic interaction of adult H. axyridis feeding on Bt maize pollen and tritrophic interaction of H. axyridis consuming the lepidopteran prey. Spodoptera exigua (Hübner) neonate larvae were used to transfer Bt protein because they could survive after ingesting transgenic cry1Ab/cry2Aj maize kernels in the previous study. ELISA bioassays confirmed that the Bt protein could be transferred, but diluted through Bt maize-prey-predator. Life history parameters such as survival, development, weight, fecundity, and egg hatching rate were not significantly different when H. axyridis consumed prey that had been reared on Bt maize compared with prey reared on a nontransformed parental control. Furthermore, feeding directly on Bt maize pollen also had no detrimental effects on fitness, survival, and weight of female and male adults. In conclusion, our results indicate that transgenic cry1Ab/cry2Aj maize poses no ecological risks on the nontarget predator H. axyridis. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  11. 40 CFR 180.1108 - Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas...

    Science.gov (United States)

    2010-07-01

    ... thuringiensis variety San Diego encapsulated into killed Pseudomonas fluorescens; exemption from the requirement... into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas fluorescens is...

  12. Persistence of Bacillus thuringiensis subsp. kurstaki in Urban Environments following Spraying▿†‡

    Science.gov (United States)

    Van Cuyk, Sheila; Deshpande, Alina; Hollander, Attelia; Duval, Nathan; Ticknor, Lawrence; Layshock, Julie; Gallegos-Graves, LaVerne; Omberg, Kristin M.

    2011-01-01

    Bacillus thuringiensis subsp. kurstaki is applied extensively in North America to control the gypsy moth, Lymantria dispar. Since B. thuringiensis subsp. kurstaki shares many physical and biological properties with Bacillus anthracis, it is a reasonable surrogate for biodefense studies. A key question in biodefense is how long a biothreat agent will persist in the environment. There is some information in the literature on the persistence of Bacillus anthracis in laboratories and historical testing areas and for Bacillus thuringiensis in agricultural settings, but there is no information on the persistence of Bacillus spp. in the type of environment that would be encountered in a city or on a military installation. Since it is not feasible to release B. anthracis in a developed area, the controlled release of B. thuringiensis subsp. kurstaki for pest control was used to gain insight into the potential persistence of Bacillus spp. in outdoor urban environments. Persistence was evaluated in two locations: Fairfax County, VA, and Seattle, WA. Environmental samples were collected from multiple matrices and evaluated for the presence of viable B. thuringiensis subsp. kurstaki at times ranging from less than 1 day to 4 years after spraying. Real-time PCR and culture were used for analysis. B. thuringiensis subsp. kurstaki was found to persist in urban environments for at least 4 years. It was most frequently detected in soils and less frequently detected in wipes, grass, foliage, and water. The collective results indicate that certain species of Bacillus may persist for years following their dispersal in urban environments. PMID:21926205

  13. Occurrence and significance of Bacillus cereus and Bacillus thuringiensis in ready-to-eat food

    DEFF Research Database (Denmark)

    Rosenquist, Hanne; Ørum-Smidt, Lasse; Andersen, Sigrid R

    2005-01-01

    Among 48,901 samples of ready-to-eat food products at the Danish retail market, 0.5% had counts of Bacillus cereus-like bacteria above 10(4) cfu g(-1). The high counts were most frequently found in starchy, cooked products, but also in fresh cucumbers and tomatoes. Forty randomly selected strains...... had at least one gene or component involved in human diarrhoeal disease, while emetic toxin was related to only one B. cereus strain. A new observation was that 31 out of the 40 randomly selected B. cereus-like strains could be classified as Bacillus thuringiensis due to crystal production and...

  14. Constitutive Activation of the Midgut Response to Bacillus thuringiensis in Bt-Resistant Spodoptera exigua

    NARCIS (Netherlands)

    Hernandez-Martinez, P.; Navarro-Cerrillo, G.; Caccia, S.; Maagd, de R.A.; Moar, W.J.; Ferre, J.; Escriche, B.; Herrero, S.

    2010-01-01

    Bacillus thuringiensis is the most effective microbial control agent for controlling numerous species from different insect orders. The main threat for the long term use of B. thuringiensis in pest control is the ability of insects to develop resistance. Thus, the identification of insect genes

  15. 40 CFR 174.504 - Bacillus thuringiensis Cry1F protein in cotton; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1F protein... PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.504 Bacillus thuringiensis Cry1F protein in cotton; exemption from the requirement of a tolerance. Residues of Bacillus...

  16. A 90 day safety assessment of genetically modified rice expressing Cry1Ab/1Ac protein using an aquatic animal model.

    Science.gov (United States)

    Zhu, Hao-Jun; Chen, Yi; Li, Yun-He; Wang, Jia-Mei; Ding, Jia-Tong; Chen, Xiu-Ping; Peng, Yu-Fa

    2015-04-15

    In fields of transgenic Bt rice, frogs are exposed to Bt proteins through consumption of both target and nontarget insects. In the present study, we assessed the risk posed by transgenic rice expressing a Cry1Ab/1Ac fusion protein (Huahui 1, HH1) on the development of Xenopus laevis. For 90 days, froglets were fed a diet with 30% HH1 rice, 30% parental rice (Minghui 63, MH63), or no rice as a control. Body weight and length were measured every 15 days. After sacrificing the froglets, we performed a range of biological, clinical, and pathological assessments. No significant differences were found in body weight (on day 90: 27.7 ± 2.17, 27.4 ± 2.40, and 27.9 ± 1.67 g for HH1, MH63, and control, respectively), body length (on day 90: 60.2 ± 1.55, 59.3 ± 2.33, and 59.7 ± 1.64 mm for HH1, MH63, and control, respectively), animal behavior, organ weight, liver and kidney function, or the microstructure of some tissues between the froglets fed on the HH1-containing diet and those fed on the MH63-containing or control diets. This indicates that frog development was not adversely affected by dietary intake of Cry1Ab/1Ac protein.

  17. Germination and persistence of Bacillus anthracis and Bacillus thuringiensis in soil microcosms.

    Science.gov (United States)

    Bishop, A H

    2014-11-01

    Decontaminating large, outdoor spaces of Bacillus anthracis spores presents significant problems, particularly in soil. Proof was sought that the addition of germinant chemicals could cause spores of B. anthracis and Bacillus thuringiensis, a commonly used simulant of the threat agent, to convert to the less resistant vegetative form in a microcosm. Nonsterile plant/soil microcosms were inoculated with spores of B. thuringiensis and two nonpathogenic strains of B. anthracis. A combination of L-alanine (100 mmol l(-1)) and inosine (10 mmol l(-1)) resulted in a 6 log decrease in spore numbers in both strains of B. anthracis over 2 weeks at 22°C; a 3 log decrease in B. anthracis Sterne spore numbers was observed after incubation for 2 weeks at 10°C. Negligible germination nor a decrease in viable count occurred in either strain when the concentration of L-alanine was decreased to 5 mmol l(-1). Germinated spores of B. thuringiensis were able to persist in vegetative form in the microcosms, whereas those of B. anthracis rapidly disappeared. The pleiotropic regulator PlcR, which B. anthracis lacks, does not contribute to the persistence of B. thuringiensis in vegetative form in soil. The principle of adding germinants to soil to trigger the conversion of spores to vegetative form has been demonstrated. Bacillus anthracis failed to persist in vegetative form or resporulate in the microcosms after it had been induced to germinate. The large scale, outdoor decontamination of B. anthracis spores may be facilitated by the application of simple, defined combinations of germinants. © 2014 Crown Copyright. Journal of Applied Microbiology © 2014 Society for Applied Microbiology This article is Published with the permission of the Controller of HMSO and Queen's Printer for Scotland.

  18. Chitinolytic activities in Bacillus thuringiensis and their synergistic effects on larvicidal activity.

    Science.gov (United States)

    Liu, M; Cai, Q X; Liu, H Z; Zhang, B H; Yan, J P; Yuan, Z M

    2002-01-01

    To investigate the distribution of chitinase in Bacillus thuringiensis strains, and the enhancing effects of the chitinase-producing B. thuringiensis strains on insecticidal toxicity of active B. thuringiensis strain against Spodoptera exigua larvae. The chitinolytic activities of B.thuringiensis strains representing the 70 serotypes were investigated by the whitish opaque halo and the colorimetric method. Thirty-eight strains produced different levels of chitinase at pH 7.0, and so did 17 strains at pH 10.0. The strain T04A001 exhibited the highest production, reaching a specific activity of 355 U ml(-1) in liquid medium. SDS-PAGE and Western blotting showed that the chitinase produced by some B. thuringiensis strains had a molecular weight of about 61 kDa. The bioassay results indicated that the chitinase-producing B. thuringiensis strains could enhance the insecticidal activity of B. thuringiensis strain DL5789 against S. exigua larvae, with an enhancing ratio of 2.35-fold. This study demonstrated that chitinase was widely produced in B. thuringiensis strains and some of the strains could enhance the toxicity of active B. thuringiensis strain. This is the first investigation devoted exclusively to analyse the distribution of chitinase in B. thuringiensis. It infers that the chitinase produced by B. thuringiensis might play a role in the activity of the biopesticide.

  19. Improvement of Bacillus thuringiensis bacteriocin production through culture conditions optimization.

    Science.gov (United States)

    Kamoun, Fakher; Zouari, Nabil; Saadaoui, Imen; Jaoua, Samir

    2009-01-01

    BUPM4 is a Bacillus thuringiensis subsp. kurstaki strain, isolated from Tunisian soil, producing an original bacteriocin named Bacthuricin F4. The optimization of the latter production conditions was carried out under several physicochemical conditions. It was found that the highest bacteriocin activity was reached at low aeration while bacteriocin synthesis yields were strongly reduced at higher ones. A balance between growth and bacteriocin synthesis, both highly dependent on aeration, was taken into account for the overproduction of bacteriocin. Both glucose and glycerol were shown to be necessary for Bacthuricin F4 maximal synthesis. In addition, the optimal carbon/nitrogen ratio for bacteriocin production is 9. In such optimal conditions, more than 4-fold greater bacteriocin production was obtained than when using TSB medium.

  20. Evaluation of a new formulation of Bacillus thuringiensis israelensis

    Directory of Open Access Journals (Sweden)

    J. Lopes

    Full Text Available The aim of this study was to determine the potency (ITU and efficacy of a liquid formulation of Bacillus thuringiensis israelensis developed by the State University of Londrina named BioUel, against early fourth instar larvae of Aedes aegypti and Culex quinquefasciatus. The ITU/mg of BioUel was 960, the LC50 was of 0.271 (± 0.39 ppm, and the LC95 was 0.634 (± 0.099 ppm, in larvae of C. quinquefasciatus. In A. aegypti larvae, LC50 was 0.332 (± 0.042 ppm and LC95 was 0.694 (± 0.073 ppm. The ITU level of BioUel and its control results were similar to most commercial products tested. Stability was of approximately 90 days, which allows for local production.

  1. Presence survival spores of Bacillus thuringiensis varieties in grain warehouse

    Directory of Open Access Journals (Sweden)

    Sánchez-Yáñez Juan Manuel

    2016-08-01

    Full Text Available Genus Bacillus thuringiensis (Bt synthesized spores and crystals toxic to pest-insects in agriculture. Bt is comospolitan then possible to isolate some subspecies or varieties from warehouse. The aims of study were: i to isolate Bt varieties from grain at werehouse ii to evaluate Bt toxicity on Spodoptera frugiperda and Shit-ophilus zeamaisese iii to analyze Bt spores persistence in Zea mays grains at werehouse compared to same Bt on grains exposed to sun radiation. Results showed that at werehouse were recovered more than one variety of Bt spores. According to each isolate Bt1 o Bt2 were toxic to S. frugiperda or S. zeamaisese. One those Bt belong to var morrisoni. At werehouse these spores on Z. mays grains surviving more time, while the same spores exposed to boicide sun radiation they died.

  2. Nanoscale imaging of Bacillus thuringiensis flagella using atomic force microscopy

    Science.gov (United States)

    Gillis, Annika; Dupres, Vincent; Delestrait, Guillaume; Mahillon, Jacques; Dufrêne, Yves F.

    2012-02-01

    Because bacterial flagella play essential roles in various processes (motility, adhesion, host interactions, secretion), studying their expression in relation to function is an important challenge. Here, we use atomic force microscopy (AFM) to gain insight into the nanoscale surface properties of two wild-type and four mutant strains of Bacillus thuringiensis exhibiting various levels of flagellation. We show that, unlike AFM in liquid, AFM in air is a simple and reliable approach to observe the morphological details of the bacteria, and to quantify the density and dimensions of their flagella. We found that the amount of flagella expressed by the six strains, as observed at the nanoscale, correlates with their microscopic swarming motility. These observations provide novel information on flagella expression in Gram-positive bacteria and demonstrate the power of AFM in genetic studies for the fast assessment of the phenotypic characteristics of bacterial strains altered in cell surface appendages.Because bacterial flagella play essential roles in various processes (motility, adhesion, host interactions, secretion), studying their expression in relation to function is an important challenge. Here, we use atomic force microscopy (AFM) to gain insight into the nanoscale surface properties of two wild-type and four mutant strains of Bacillus thuringiensis exhibiting various levels of flagellation. We show that, unlike AFM in liquid, AFM in air is a simple and reliable approach to observe the morphological details of the bacteria, and to quantify the density and dimensions of their flagella. We found that the amount of flagella expressed by the six strains, as observed at the nanoscale, correlates with their microscopic swarming motility. These observations provide novel information on flagella expression in Gram-positive bacteria and demonstrate the power of AFM in genetic studies for the fast assessment of the phenotypic characteristics of bacterial strains altered in

  3. Binding site alteration is responsible for field-isolated resistance to Bacillus thuringiensis Cry2A insecticidal proteins in two Helicoverpa species.

    Directory of Open Access Journals (Sweden)

    Silvia Caccia

    Full Text Available BACKGROUND: Evolution of resistance by target pests is the main threat to the long-term efficacy of crops expressing Bacillus thuringiensis (Bt insecticidal proteins. Cry2 proteins play a pivotal role in current Bt spray formulations and transgenic crops and they complement Cry1A proteins because of their different mode of action. Their presence is critical in the control of those lepidopteran species, such as Helicoverpa spp., which are not highly susceptible to Cry1A proteins. In Australia, a transgenic variety of cotton expressing Cry1Ac and Cry2Ab (Bollgard II comprises at least 80% of the total cotton area. Prior to the widespread adoption of Bollgard II, the frequency of alleles conferring resistance to Cry2Ab in field populations of Helicoverpa armigera and Helicoverpa punctigera was significantly higher than anticipated. Colonies established from survivors of F(2 screens against Cry2Ab are highly resistant to this toxin, but susceptible to Cry1Ac. METHODOLOGY/PRINCIPAL FINDINGS: Bioassays performed with surface-treated artificial diet on neonates of H. armigera and H. punctigera showed that Cry2Ab resistant insects were cross-resistant to Cry2Ae while susceptible to Cry1Ab. Binding analyses with (125I-labeled Cry2Ab were performed with brush border membrane vesicles from midguts of Cry2Ab susceptible and resistant insects. The results of the binding analyses correlated with bioassay data and demonstrated that resistant insects exhibited greatly reduced binding of Cry2Ab toxin to midgut receptors, whereas no change in (125I-labeled-Cry1Ac binding was detected. As previously demonstrated for H. armigera, Cry2Ab binding sites in H. punctigera were shown to be shared by Cry2Ae, which explains why an alteration of the shared binding site would lead to cross-resistance between the two Cry2A toxins. CONCLUSION/SIGNIFICANCE: This is the first time that a mechanism of resistance to the Cry2 class of insecticidal proteins has been reported

  4. Pengaruh Bacillus Thuringiensis terhadap penggerek batang jagung Ostrinia Furnacalis (Lep. Pyralidae

    Directory of Open Access Journals (Sweden)

    Harnoto Harnoto

    2017-02-01

    Full Text Available The effect of Bacillus thuringiensis to the mortality of corn stemborer Ostrinia furnacalis (Lep. Pyralidae. The study was conducted at the laboratory of Bogor Research Institute for Agricultural Biotechnology and Genetic Resources during 2005. The objective of this study was to evaluate the effect of the dosages of Bacillus. thuringiensis var. aizawai and var. Kurstaki on the mortalities of larvae of corn stemborer Ostrinia furnacalis. Completely randomize design was used with seven treatments. Each treatment was repeated four time with ten larvae per replication. The treatments were three formulation dosages of B. thuringiensis var. aizawai, i.e. 0,5; 1,0; and 2,0 g/l, thee formulation dosages of B. thuringiensis var. Kurstaki, i.e. 0,5; 1,0; and 2,0 g/l, and untreated control. Second instar larvae of O. furnacalis was used in this study. B. thuringiensis was contaminated to the surface of artificial diet with a small paint brush. The result showed that B. thuringiensis var. kurstaki at the dose rate of 1,0 g/l was toxic to the test insect while B. thuringiensis var. aizawai at the dose rate of 2,0 g/l was toxic to the test insect B. thuringiensis var. kurstaki was more toxic than  B. thuringiensis var. aizawai to the corn stemborer.

  5. Occurrence of Toxigenic Bacillus cereus and Bacillus thuringiensis in Doenjang, a Korean Fermented Soybean Paste.

    Science.gov (United States)

    Park, Kyung Min; Kim, Hyun Jung; Jeong, Moon Cheol; Koo, Minseon

    2016-04-01

    This study determined the prevalence and toxin profile of Bacillus cereus and Bacillus thuringiensis in doenjang, a fermented soybean food, made using both traditional and commercial methods. The 51 doenjang samples tested were broadly contaminated with B. cereus; in contrast, only one sample was positive for B. thuringiensis. All B. cereus isolates from doenjang were positive for diarrheal toxin genes. The frequencies of nheABC and hblACD in traditional samples were 22.7 and 0%, respectively, whereas 5.1 and 5.1% of B. cereus isolates from commercial samples possessed nheABC and hblACD, respectively. The detection rate of ces gene was 10.8%. The predominant toxin profile among isolates from enterotoxigenic B. cereus in doenjang was profile 4 (entFM-bceT-cytK). The major enterotoxin genes in emetic B. cereus were cytK, entFM, and nheA genes. The B. thuringiensis isolate was of the diarrheagenic type. These results provide a better understanding of the epidemiology of the enterotoxigenic and emetic B. cereus groups in Korean fermented soybean products.

  6. Evaluation of Bt (Bacillus thuringiensis) rice varieties against stem borer (Chilo suppressalis).

    Science.gov (United States)

    Kiani, Ghaffar; Nematzadeh, Ghorban Ali; Ghareyazie, Behzad; Sattari, Majid

    2008-02-15

    Three transgenic rice varieties namely Khazar, Neda and Nemat, all containing a cry1Ab gene, were evaluated through PCR analysis and field examinations for their resistance at natural infestation of insect pests during 2007. The results showed that all transgenic varieties produced 1.2 kb PCR product derived from application of cry1Ab gene. In field conditions, transgenic varieties exhibited high levels of resistance against natural infestation of stem borer and the damaged plants based on dead heart or white heat for them were less than 1%. Moreover, in stem-cut bioassay 100% of released larvae died within four days after infestation. These results demonstrate that expression of cry1Ab gene in the genome of transgenic varieties provided season-long protection from the natural infestation of lepidopteran insects.

  7. Lack of Detection of Bt Sugarcane Cry1Ab and NptII DNA and Proteins in Sugarcane Processing Products Including Raw Sugar

    Directory of Open Access Journals (Sweden)

    Adriana Cheavegatti-Gianotto

    2018-03-01

    Full Text Available Brazil is the largest sugarcane producer and the main sugar exporter in the world. The industrial processes applied by Brazilian mills are very efficient in producing highly purified sugar and ethanol. Literature presents evidence of lack of DNA/protein in these products, regardless of the nature of sugarcane used as raw material. Recently CTNBio, the Brazilian biosafety authority, has approved the first biotechnology-derived sugarcane variety for cultivation, event CTC175-A, which expresses the Cry1Ab protein to control the sugarcane borer (Diatraea saccharalis. The event also expresses neomycin-phosphotransferase type II (NptII protein used as selectable marker during the transformation process. Because of the high purity of sugar and ethanol produced from genetically modified sugarcane, these end-products should potentially be classified as “pure substances, chemically defined,” by Brazilian Biosafety Law No. 11.105. If this classification is to be adopted, these substances are not considered as “GMO derivatives” and fall out of the scope of Law No. 11.105. In order to assess sugar composition and quality, we evaluate Cry1Ab and NptII expression in several sugarcane tissues and in several fractions from laboratory-scale processing of event CTC175-A for the presence of these heterologous proteins as well as for the presence of traces of recombinant DNA. The results of these studies show that CTC175-A presents high expression of Cry1Ab in leaves and barely detectable expression of heterologous proteins in stalks. We also evaluated the presence of ribulose-1,5-bisphosphate carboxylase/oxygenase protein and DNA in the fractions of the industrial processing of conventional Brazilian sugarcane cultivars. Results from both laboratory and industrial processing were concordant, demonstrating that DNA and protein are not detected in the clarified juice and downstream processed fractions, including ethanol and raw sugar, indicating that protein

  8. Screening of Bacillus thuringiensis strains effective against mosquitoes Prospecção de estirpes de Bacillus thuringiensis efetivas contra mosquitos

    Directory of Open Access Journals (Sweden)

    Rose Gomes Monnerat

    2005-02-01

    Full Text Available The objective of this work was to evaluate 210 Bacillus thuringiensis strains against Aedes aegypti and Culex quinquefasciatus larvae to select the most effective. These strains were isolated from different regions of Brazil and are stored in a Bacillus spp. collection at Embrapa Recursos Genéticos e Biotecnologia, Brasília, Brazil. The selected strains were characterized by morphological (microscopy, biochemical (SDS-PAGE 10% and molecular (PCR methods. Six B. thuringiensis strains were identified as mosquito-toxic after the selective bioassays. None of the strains produced the expected PCR products for detection of cry4, cry11 and cyt1A genes. These results indicate that the activity of mosquitocidal Brazilian strains are not related with Cry4, Cry11 or Cyt proteins, so they could be used as an alternative bioinsecticide against mosquitoes.Neste trabalho foram realizados testes de patogenicidade com 210 estirpes de Bacillus thuringiensis contra larvas de Aedes aegypti e Culex quinquefasciatus, a fim de se determinar as mais eficazes. Estas estirpes foram isoladas de diversas regiões do Brasil e estão armazenadas na coleção de Bacillus spp. da Embrapa Recursos Genéticos e Biotecnologia. As estirpes selecionadas foram caracterizadas por métodos morfológicos (microscopia, bioquímicos (SDS-PAGE 10% e moleculares (Reação em Cadeia da Polimerase. Foram selecionadas seis estirpes entomopatogênicas de Bacillus thuringiensis. Nenhuma das estirpes de Bacillus thuringiensis apresentou produtos de PCR esperados para a detecção dos genes cry4, cry11 e cyt1A. A patogenicidade das estirpes não está associada à presença das toxinas Cry4, Cry11 ou Cyt, assim, essas estirpes poderão ser utilizadas para a formatação de um bioinseticida alternativo contra mosquitos.

  9. Kelimpahan Arthropoda Karnivora di Pertanaman Padi Ratun di Sawah Lebak yang Diaplikasikan Bioinsektisida Bacillus Thuringiensis

    OpenAIRE

    Sunariah, Fila; Herlinda, Siti; Windusari, Yuanita

    2016-01-01

    Kelimpahan Arthropoda karnivora di pertanaman padi ratun di sawah lebak yang diaplikasikan bioinsektisida Bacillus thuringiensis telah dilakukan di sawah lebak Pemulutan, Ogan Ilir Sumatera Selatan selama Agustus-Oktober 2013. Penelitian bertujuan untuk mengetahui kelimpahan Arthropoda karnivora padi ratun yang diberi aplikasi Bacillus thurngiensis dan tanpa diberi aplikasi bioinsektisida. Pengambilan sampel spesies Arthropoda dilakukan dengan menggunakan jaring serangga pada tajuk tanaman pa...

  10. Phytophagous mites on genetically modified maize with Bacillus thuringiensis genes

    Directory of Open Access Journals (Sweden)

    Tamara Esteves Ferreira

    2017-08-01

    Full Text Available ABSTRACT: The study of non-target organisms such as mites, can provide information about the possible effects of transgenic maize cultivars on the arthropod community. This study aimed to evaluate the abundance of phytophagous mites in the area of Bt maize and evaluate the instantaneous population growth rate, and food preference of phytophagous mites on Bt maize. We registered the species occurrence and the number of adult mite individuals on four maize cultivars, one non-Bt 30F35, and three containing the proteins Cry1Ab (30F35 Yg, Cry1F (30F35 Hx and Vip3a (Impacto Viptera. Cry proteins disrupt the midgut epithelium of insect pests. The food preference and instantaneous population growth rate (ri were evaluated using the two-spotted spider mite Tetranychus urticae as model. The species Catarhinus tricholaenae and Aceria zeala were recorded. The Bt maize cultivars did not significantly affect the average number of C. tricholaenae and A. zeala mites compared to conventional cultivar. The population growth rates of T. urticae were similar for the different maize cultivars. T. urticae showed no preference between the leaf of Bt or conventional cultivars. Bt maize did not affect the abundance of species in phytophagous mite.

  11. How Quorum Sensing Connects Sporulation to Necrotrophism in Bacillus thuringiensis.

    Directory of Open Access Journals (Sweden)

    Stéphane Perchat

    2016-08-01

    Full Text Available Bacteria use quorum sensing to coordinate adaptation properties, cell fate or commitment to sporulation. The infectious cycle of Bacillus thuringiensis in the insect host is a powerful model to investigate the role of quorum sensing in natural conditions. It is tuned by communication systems regulators belonging to the RNPP family and directly regulated by re-internalized signaling peptides. One such RNPP regulator, NprR, acts in the presence of its cognate signaling peptide NprX as a transcription factor, regulating a set of genes involved in the survival of these bacteria in the insect cadaver. Here, we demonstrate that, in the absence of NprX and independently of its transcriptional activator function, NprR negatively controls sporulation. NprR inhibits expression of Spo0A-regulated genes by preventing the KinA-dependent phosphorylation of the phosphotransferase Spo0F, thus delaying initiation of the sporulation process. This NprR function displays striking similarities with the Rap proteins, which also belong to the RNPP family, but are devoid of DNA-binding domain and indirectly control gene expression via protein-protein interactions in Bacilli. Conservation of the Rap residues directly interacting with Spo0F further suggests a common inhibition of the sporulation phosphorelay. The crystal structure of apo NprR confirms that NprR displays a highly flexible Rap-like structure. We propose a molecular regulatory mechanism in which key residues of the bifunctional regulator NprR are directly and alternatively involved in its two functions. NprX binding switches NprR from a dimeric inhibitor of sporulation to a tetrameric transcriptional activator involved in the necrotrophic lifestyle of B. thuringiensis. NprR thus tightly coordinates sporulation and necrotrophism, ensuring survival and dissemination of the bacteria during host infection.

  12. 40 CFR 174.518 - Bacillus thuringiensis Cry3Bb1 protein in corn; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry3Bb1 protein... PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.518 Bacillus thuringiensis Cry3Bb1 protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis...

  13. Field and laboratory studies on the impact of two Bt rice lines expressing a fusion protein Cry1Ab/1Ac on aquatic organisms.

    Science.gov (United States)

    Wang, Yongmo; Huang, Jiacheng; Hu, Huawei; Li, Jianhong; Liu, Biao; Zhang, Guoan

    2013-06-01

    Genetically modified (GM) rice expressing Bt toxins is at the edge of commercial release in China. However, little information is available concerning the impact of Bt rice on aquatic organisms which are abundant in paddy field. A two-year study was conducted to assess the effects of two GM rice lines expressing a fusion protein Cry1Ab/1Ac (Bt rice) on three groups of zooplankton, rotifers, cladocerans and copepods in field conditions. Multi-factor ANOVA revealed that the population densities of rotifers, cladocerans and copepods in paddy field varied significantly between years and rice developmental stages, but did not differ significantly between Bt and non-Bt rice treatments. In all the field investigations, only one significant difference was found on copepods in the tillering stage of 2009, but the difference was not related to the presence of the Cry toxin. Under open-air conditions, we simulated the farming practice of straw mulch, using Bt rice straw as a food source for the water flea Daphnia hyalina. After one and two months of culture, the density of D. hyalina did not differ between Bt rice treatments and non-Bt rice treatments. A laboratory experiment found that purified Bt toxins Cry1Ab and Cry1Ac had no toxic effect on D. hyalina even in the treatment in which the Bt toxin concentration was as high as 2500ng/ml. Those above results indicate that the two Bt rice lines have no negative effect on the three groups of zooplankton. However, further studies are needed to compare the effects of Bt rice and non-Bt rice on the paddy zooplankton community in the context of integrated pest management which includes the use of pesticides. Copyright © 2013 Elsevier Inc. All rights reserved.

  14. Detection by real-time PCR and pyrosequencing of the cry1Ab and cry1Ac genes introduced in genetically modified (GM) constructs.

    Science.gov (United States)

    Debode, Frederic; Janssen, Eric; Bragard, Claude; Berben, Gilbert

    2017-08-01

    The presence of genetically modified organisms (GMOs) in food and feed is mainly detected by the use of targets focusing on promoters and terminators. As some genes are frequently used in genetically modified (GM) construction, they also constitute excellent screening elements and their use is increasing. In this paper we propose a new target for the detection of cry1Ab and cry1Ac genes by real-time polymerase chain reaction (PCR) and pyrosequencing. The specificity, sensitivity and robustness of the real-time PCR method were tested following the recommendations of international guidelines and the method met the expected performance criteria. This paper also shows how the robustness testing was assessed. This new cry1Ab/Ac method can provide a positive signal with a larger number of GM events than do the other existing methods using double dye-probes. The method permits the analysis of results with less ambiguity than the SYBRGreen method recommended by the European Reference Laboratory (EURL) GM Food and Feed (GMFF). A pyrosequencing method was also developed to gain additional information thanks to the sequence of the amplicon. This method of sequencing-by-synthesis can determine the sequence between the primers used for PCR. Pyrosequencing showed that the sequences internal to the primers present differences following the GM events considered and three different sequences were observed. The sensitivity of the pyrosequencing was tested on reference flours with a low percentage GM content and different copy numbers. Improvements in the pyrosequencing protocol provided correct sequences with 50 copies of the target. Below this copy number, the quality of the sequence was more random.

  15. Environmental Distribution and Diversity of Insecticidal Proteins of Bacillus thuringiensis Berliner

    Directory of Open Access Journals (Sweden)

    Xavier, R.

    2007-01-01

    Full Text Available Bacillus thuringiensis Berliner based biopesticides have been successfully used world over for the control of agricultural pests and vectors of human diseases. Currently there are more than 200 B. thuringiensis strains with differing insecticidal activities are available as biocontrol agents and for developing transgenic plants. However, two major disadvantages are the development of insect resistance and high target specificity (narrow host range. Globally there is a continuous search for new B. thuringiensis strains with novel insecticidal activities. The present study aims to study the environmental distribution of B. thuringiensis and their toxic potential against insect pests. Soil and grain samples were collected from different environments and were processed by a modified acetate selection method. Initially B. thuringiensis isolates were screened on the basis of colony morphology and phase contrast microscopy for the presence of parasporal crystal inclusions. The population dynamics showed that B. thuringiensis is abundant in sericulture environment compared to other niches. Relative abundance of B. thuringiensis strains in sericulture environment shows the persistent association of B. thuringiensis with Bombyx mori (silk worm as insect pathogen. The protein profiles of the selected strains were studied by SDS-PAGE. The protein profiles of majority of B. thuringiensis isolates from grain storage facilities predominantly showing the 130 kDa and 68 kDa proteins, which is characteristics of lepidopteran active B. thuringiensis. However, one isolate BTRX-4 has 80-85 kDa protein, which is novel in that, this strain also exhibits antilepidopteran activity, which is normally presented by B. thuringiensis strains having 130 kDa and 68 kDa proteins. The protein profile of B. thuringiensis isolates from sericulture environment shows two different protein profiles. B. thuringiensis isolates BTRX-16 to BTRX-22 predominantly show 130 kDa protein

  16. Germination and conjugation of Bacillus thuringiensis subsp. israelensis in the intestine of gnotobiotic rats

    DEFF Research Database (Denmark)

    Wilcks, Andrea; Ørum-Smidt, Lasse; Bahl, Martin Iain

    2008-01-01

    Aims: To study the ability of Bacillus thuringiensis subsp. israelensis spores to germinate and subsequently transfer a conjugative plasmid in the intestinal tract of gnotobiotic rats. Methods and Results: Germination was studied by feeding germ-free rats with spores of a B. thuringiensis strain...... harbouring a plasmid encoding green fluorescent protein (GFP), which enabled quantification of germinated bacteria by flow cytometry. To study in vivo conjugation, germ-free rats were first associated with a B. thuringiensis recipient strain and after 1 week an isogenic donor strain harbouring...... the conjugative plasmid pXO16 was introduced. Both strains were given as spores and transfer of pXO16 was observed from the donor to the recipient strain. Conclusions: Bacillus thuringiensis is able to have a full life cycle in the intestine of gnotobiotic rats including germination of spores, several cycles...

  17. Side effects of Bacillus thuringiensis var. kurstaki on the hymenopterous parasitic wasp Trichogramma chilonis.

    Science.gov (United States)

    Amichot, Marcel; Curty, Christine; Benguettat-Magliano, Olivia; Gallet, Armel; Wajnberg, Eric

    2016-02-01

    Most of the detrimental effects of using conventional insecticides to control crop pests are now well identified and are nowadays major arguments for replacing such compounds by the use of biological control agents. In this respect, the bacterium Bacillus thuringiensis var. kurstaki and Trichogramma (Hymenoptera: Trichogrammatidae) parasitic wasp species are both effective against lepidopterous pests and can actually be used concomitantly. In this work, we studied the potential side effects of B. thuringiensis var. kurstaki on Trichogramma chilonis females. We first evidenced an acute toxicity of B. thuringiensis on T. chilonis. Then, after ingestion of B. thuringiensis at sublethal doses, we focused on life history traits of T. chilonis such as longevity, reproductive success and the time spent on host eggs patches. The reproductive success of T. chilonis was not modified by B. thuringiensis while a significant effect was observed on longevity and the time spent on host eggs patches. The physiological and ecological meanings of the results obtained are discussed.

  18. Persistence of Bacillus thuringiensis bioinsecticides in the gut of human-flora-associated rats

    DEFF Research Database (Denmark)

    Wilcks, Andrea; Hansen, Bjarne Munk; Hendriksen, Niels Bohse

    2006-01-01

    The capability of two bioinsecticide strains of Bacillus thuringiensis (ssp. israelensis and ssp. kurstaki) to germinate and persist in vivo in the gastrointestinal tract of human-flora-associated rats was studied. Rats were dosed either with vegetative cells or spores of the bacteria for 4...... consecutive days. In animals fed spores, B. thuringiensis cells were detected in faecal and intestinal samples of all animals, whereas vegetative cells only poorly survived the gastric passage. Heat-treatment of intestinal samples, which kills vegetative cells, revealed that B. thuringiensis spores were...... capable of germination in the gastrointestinal tract. In one animal fed spores of B. thuringiensis ssp. kurstaki, these bacteria were detected at high density (10(3)-10(4) CFU g(-1) faecal and intestinal samples) even 2 weeks after the last dosage. In the same animal, passage of B. thuringiensis ssp...

  19. Isolation and identification of some Bacillus thuringiensis strains with insecticidal activity against Ceratitis capitata

    International Nuclear Information System (INIS)

    Majdoub, Nihed

    2010-01-01

    The aims of the present work is to study the effect of toxins (delta-endotoxins), extracted from different strains of Bacillus thuringiensis on Ceratitis capitata, a devastating of citrus and fruit trees. Strains of B. thuringiensis were isolated from the mud of Sebket Sejoumi. Among 70 isolates tested, 15 showed a significant identicalness activity in which 5 isolates led to mortality rates ≥ 90 pour cent . These mortality rates are caused by endotoxins of B. thuringiensis. Analysis of proteins profiles of different isolates of B. thuringiensis revealed variability between them. The preliminary results of this study encourage us towards the characterization of the insecticidal activity produced by B. thuringiensis strains for large scale application.

  20. Translocation and insecticidal activity of Bacillus thuringiensis living inside of plants

    OpenAIRE

    Monnerat, Rose Gomes; Soares, Carlos Marcelo; Capdeville, Guy; Jones, Gareth; Martins, Érica Soares; Praça, Lilian; Cordeiro, Bruno Arrivabene; Braz, Shélida Vasconcelos; Dos Santos, Roseane Cavalcante; Berry, Colin

    2009-01-01

    Summary The major biological pesticide for the control of insect infestations of crops, Bacillus thuringiensis was found to be present naturally within cotton plants from fields that had never been treated with commercial formulations of this bacterium. The ability of B. thuringiensis to colonize plants as an endophyte was further established by the introduction of a strain marked by production of green fluorescent protein (GFP). After inoculation of this preparation close to the roots of cot...

  1. Extended genetic analysis of Brazilian isolates of Bacillus cereus and Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Viviane Zahner

    2013-02-01

    Full Text Available Multiple locus sequence typing (MLST was undertaken to extend the genetic characterization of 29 isolates of Bacillus cereus and Bacillus thuringiensis previously characterized in terms of presence/absence of sequences encoding virulence factors and via variable number tandem repeat (VNTR. Additional analysis involved polymerase chain reaction for the presence of sequences (be, cytK, inA, pag, lef, cya and cap, encoding putative virulence factors, not investigated in the earlier study. MLST analysis ascribed novel and unique sequence types to each of the isolates. A phylogenetic tree was constructed from a single sequence of 2,838 bp of concatenated loci sequences. The strains were not monophyletic by analysis of any specific housekeeping gene or virulence characteristic. No clear association in relation to source of isolation or to genotypic profile based on the presence or absence of putative virulence genes could be identified. Comparison of VNTR profiling with MLST data suggested a correlation between these two methods of genetic analysis. In common with the majority of previous studies, MLST was unable to provide clarification of the basis for pathogenicity among members of the B. cereus complex. Nevertheless, our application of MLST served to reinforce the notion that B. cereus and B. thuringiensis should be considered as the same species.

  2. Synergistic activity of Bacillus thuringiensis toxins against Simulium spp. larvae.

    Science.gov (United States)

    Monnerat, Rose; Pereira, Eleny; Teles, Beatriz; Martins, Erica; Praça, Lilian; Queiroz, Paulo; Soberon, Mario; Bravo, Alejandra; Ramos, Felipe; Soares, Carlos Marcelo

    2014-09-01

    Species of Simulium spread diseases in humans and animals such as onchocerciasis and mansonelosis, causing health problems and economic loses. One alternative for controlling these insects is the use of Bacillus thuringiensis serovar israelensis (Bti). This bacterium produces different dipteran-active Cry and Cyt toxins and has been widely used in blackfly biological control programs worldwide. Studies on other insect targets have revealed the role of individual Cry and Cyt proteins in toxicity and demonstrated a synergistic effect among them. However, the insecticidal activity and interactions of these proteins against Simulium larvae have not been reported. In this study we demonstrate that Cry4Ba is the most effective toxin followed by Cry4Aa and Cry11Aa. Cry10Aa and Cyt1Aa were not toxic when administered alone but both were able to synergise the activity of Cry4B and Cry11Aa toxins. Cyt1Aa is also able to synergise with Cry4Aa. The mixture of all toxin-producing strains showed the greatest level of synergism, but still lower than the Bti parental strain. Copyright © 2014 Elsevier Inc. All rights reserved.

  3. Bacillus thuringiensis: fermentation process and risk assessment: a short review

    Directory of Open Access Journals (Sweden)

    Deise M. F Capalbo

    1995-02-01

    Full Text Available Several factors make the local production of Bacillus thuringiensis (Bt highly appropriate for pest control in developing nations. Bt can be cheaply produced on a wide variety of low cost, organic substrates. Local production results in considerable savings in hard currency which otherwise would be spent on importation of chemical and biological insecticides. The use of Bt in Brazil has been limited in comparison with chemical insecticides. Although Bt is imported, some Brazilian researchers have been working on its development and production. Fermentation processes (submerged and semi-solid were applied, using by-products from agro-industries. As the semi-solid fermentation process demonstrated to be interesting for Bt endotoxins production, it could be adopted for small scale local production. Although promising results had been achieved, national products have not been registered due to the absence of a specific legislation for biological products. Effective actions are being developed in order to solve this gap. Regardless of the biocontrol agents being considered atoxic and harmless to the environment, information related to direct and indirect effects of microbials are still insufficient in many cases. The risk analysis of the use of microbial control agents is of upmost importance nowadays, and is also discussed.

  4. Bacillus thuringiensis Toxins: An Overview of Their Biocidal Activity

    Science.gov (United States)

    Palma, Leopoldo; Muñoz, Delia; Berry, Colin; Murillo, Jesús; Caballero, Primitivo

    2014-01-01

    Bacillus thuringiensis (Bt) is a Gram positive, spore-forming bacterium that synthesizes parasporal crystalline inclusions containing Cry and Cyt proteins, some of which are toxic against a wide range of insect orders, nematodes and human-cancer cells. These toxins have been successfully used as bioinsecticides against caterpillars, beetles, and flies, including mosquitoes and blackflies. Bt also synthesizes insecticidal proteins during the vegetative growth phase, which are subsequently secreted into the growth medium. These proteins are commonly known as vegetative insecticidal proteins (Vips) and hold insecticidal activity against lepidopteran, coleopteran and some homopteran pests. A less well characterized secretory protein with no amino acid similarity to Vip proteins has shown insecticidal activity against coleopteran pests and is termed Sip (secreted insecticidal protein). Bin-like and ETX_MTX2-family proteins (Pfam PF03318), which share amino acid similarities with mosquitocidal binary (Bin) and Mtx2 toxins, respectively, from Lysinibacillus sphaericus, are also produced by some Bt strains. In addition, vast numbers of Bt isolates naturally present in the soil and the phylloplane also synthesize crystal proteins whose biological activity is still unknown. In this review, we provide an updated overview of the known active Bt toxins to date and discuss their activities. PMID:25514092

  5. Adhesion of Spores of Bacillus thuringiensis on a Planar Surface

    Energy Technology Data Exchange (ETDEWEB)

    Chung, Eunhyea [Georgia Institute of Technology; Kweon, Hyojin [Georgia Institute of Technology; Yiacoumi, Sotira [Georgia Institute of Technology; Lee, Ida [University of Tennessee, Knoxville (UTK); Joy, David Charles [ORNL; Palumbo, Anthony Vito [ORNL; Tsouris, Costas [ORNL

    2010-01-01

    Adhesion of spores of Bacillus thuringiensis (Bt) and spherical silica particles on surfaces was experimentally and theoretically investigated in this study. Topography analysis via atomic force microscopy (AFM) and electron microscopy indicates that Bt spores are rod shaped, {approx}1.3 {mu}m in length and {approx}0.8 {mu}m in diameter. The adhesion force of Bt spores and silica particles on gold-coated glass was measured at various relative humidity (RH) levels by AFM. It was expected that the adhesion force would vary with RH because the individual force components contributing to the adhesion force depend on RH. The adhesion force between a particle and a planar surface in atmospheric environments was modeled as the contribution of three major force components: capillary, van der Waals, and electrostatic interaction forces. Adhesion force measurements for Bt spore (silica particle) and the gold surface system were comparable with calculations. Modeling results show that there is a critical RH value, which depends on the hydrophobicity of the materials involved, below which the water meniscus does not form and the contribution of the capillary force is zero. As RH increases, the van der Waals force decreases while the capillary force increases to a maximum value.

  6. Raman spectroscopy of Bacillus thuringiensis physiology and inactivation

    Science.gov (United States)

    Morrow, J. B.; Almeida, J.; Cole, K. D.; Reipa, V.

    2012-12-01

    The ability to detect spore contamination and inactivation is relevant to developing and determining decontamination strategy success for food and water safety. This study was conducted to develop a systematic comparison of nondestructive vibrational spectroscopy techniques (Surface-Enhanced Raman Spectroscopy, SERS, and normal Raman) to determine indicators of Bacillus thuringiensis physiology (spore, vegetative, outgrown, germinated and inactivated spore forms). SERS was found to provide better resolution of commonly utilized signatures of spore physiology (dipicolinic acid at 1006 cm-1 and 1387 cm-1) compared to normal Raman and native fluorescence indigenous to vegetative and outgrown cell samples was quenched in SERS experiment. New features including carotenoid pigments (Raman features at 1142 cm-1, 1512 cm-1) were identified for spore cell forms. Pronounced changes in the low frequency region (300 cm-1 to 500 cm-1) in spore spectra occurred upon germination and inactivation (with both free chlorine and by autoclaving) which is relevant to guiding decontamination and detection strategies using Raman techniques.

  7. Bacillus thuringiensis Toxins: An Overview of Their Biocidal Activity

    Directory of Open Access Journals (Sweden)

    Leopoldo Palma

    2014-12-01

    Full Text Available Bacillus thuringiensis (Bt is a Gram positive, spore-forming bacterium that synthesizes parasporal crystalline inclusions containing Cry and Cyt proteins, some of which are toxic against a wide range of insect orders, nematodes and human-cancer cells. These toxins have been successfully used as bioinsecticides against caterpillars, beetles, and flies, including mosquitoes and blackflies. Bt also synthesizes insecticidal proteins during the vegetative growth phase, which are subsequently secreted into the growth medium. These proteins are commonly known as vegetative insecticidal proteins (Vips and hold insecticidal activity against lepidopteran, coleopteran and some homopteran pests. A less well characterized secretory protein with no amino acid similarity to Vip proteins has shown insecticidal activity against coleopteran pests and is termed Sip (secreted insecticidal protein. Bin-like and ETX_MTX2-family proteins (Pfam PF03318, which share amino acid similarities with mosquitocidal binary (Bin and Mtx2 toxins, respectively, from Lysinibacillus sphaericus, are also produced by some Bt strains. In addition, vast numbers of Bt isolates naturally present in the soil and the phylloplane also synthesize crystal proteins whose biological activity is still unknown. In this review, we provide an updated overview of the known active Bt toxins to date and discuss their activities.

  8. Biopesticide production from Bacillus thuringiensis: an environmentally friendly alternative.

    Science.gov (United States)

    Rosas-García, Ninfa M

    2009-01-01

    Since its discovery as a microbial insecticide, Bacillus thuringiensis has been widely used to control insect pests important in agriculture, forestry, and medicine. The wide variety of formulations based on spore-crystal complexes intended for ingestion by target insects, are the result of many years of research. The development of a great variety of matrices for support of the spore-crystal complex enables many improvements, such as an increase in toxic activity, higher palatability to insects, or longer shelf lives. These matrices use many chemical, vegetable or animal compounds to foster contact between crystals and insect midguts, without harming humans or the environment. Biotechnology companies are tasked with the production of these kinds of bioinsecticides. These companies must not only provide formulations tailored to specific crops and the insect pests, but they must also search for and produce bioinsecticides based on new strains of high potency, whether wild or genetically improved. It is expected that new products will appear on the market soon, providing an increased activity spectrum and applicability to many other pest-impacted crops. These products may help develop a more organic agriculture. This review article discusses recent patents related to bioinsecticides.

  9. The complete genome sequence of Bacillus thuringiensis serovar Hailuosis YWC2-8.

    Science.gov (United States)

    Zhu, Jun; Zhang, Qinbin; Cao, Ye; Li, Qiao; Zhu, Zizhong; Wang, Linxia; Li, Ping

    2016-02-10

    Bacillus thuringiensis, a typical aerobic, Gram-positive, spore-forming bacterium, is an important microbial insecticide widely used in the control of agricultural pests. B. thuringiensis serovar Hailuosis YWC2-8 with high insecticidal activity against Diptera and Lepidoptera insects has three insecticidal crystal protein genes, such as cry4Cb2, cry30Ea2, and cry56Aa1. In this study, the complete genome sequence of B. thuringiensis YWC2-8 was analyzed, which contains one circular gapless chromosome and six circular plasmids. Copyright © 2015 Elsevier B.V. All rights reserved.

  10. A Bacillus thuringiensis S-Layer Protein Involved in Toxicity against Epilachna varivestis (Coleoptera: Coccinellidae)

    Science.gov (United States)

    Peña, Guadalupe; Miranda-Rios, Juan; de la Riva, Gustavo; Pardo-López, Liliana; Soberón, Mario; Bravo, Alejandra

    2006-01-01

    The use of Bacillus thuringiensis as a biopesticide is a viable alternative for insect control since the insecticidal Cry proteins produced by these bacteria are highly specific; harmless to humans, vertebrates, and plants; and completely biodegradable. In addition to Cry proteins, B. thuringiensis produces a number of extracellular compounds, including S-layer proteins (SLP), that contribute to virulence. The S layer is an ordered structure representing a proteinaceous paracrystalline array which completely covers the surfaces of many pathogenic bacteria. In this work, we report the identification of an S-layer protein by the screening of B. thuringiensis strains for activity against the coleopteran pest Epilachna varivestis (Mexican bean beetle; Coleoptera: Coccinellidae). We screened two B. thuringiensis strain collections containing unidentified Cry proteins and also strains isolated from dead insects. Some of the B. thuringiensis strains assayed against E. varivestis showed moderate toxicity. However, a B. thuringiensis strain (GP1) that was isolated from a dead insect showed a remarkably high insecticidal activity. The parasporal crystal produced by the GP1 strain was purified and shown to have insecticidal activity against E. varivestis but not against the lepidopteran Manduca sexta or Spodoptera frugiperda or against the dipteran Aedes aegypti. The gene encoding this protein was cloned and sequenced. It corresponded to an S-layer protein highly similar to previously described SLP in Bacillus anthracis (EA1) and Bacillus licheniformis (OlpA). The phylogenetic relationships among SLP from different bacteria showed that these proteins from Bacillus cereus, Bacillus sphaericus, B. anthracis, B. licheniformis, and B. thuringiensis are arranged in the same main group, suggesting similar origins. This is the first report that demonstrates that an S-layer protein is directly involved in toxicity to a coleopteran pest. PMID:16391064

  11. Prevalence and Toxin Characteristics of Bacillus thuringiensis Isolated from Organic Vegetables.

    Science.gov (United States)

    Kim, Jung-Beom; Choi, Ok-Kyung; Kwon, Sun-Mok; Cho, Seung-Hak; Park, Byung-Jae; Jin, Na Young; Yu, Yong Man; Oh, Deog-Hwan

    2017-08-28

    The prevalence and toxin characteristics of Bacillus thuringiensis isolated from 39 organic vegetables were investigated. B. thuringiensis was detected in 30 out of the 39 organic vegetables (76.9%) with a mean value of 2.60 log CFU/g. Twenty-five out of the 30 B. thuringiensis isolates (83.3%) showed insecticidal toxicity against Spodoptera exigua . The hblCDA, nheABC , and entFM genes were found to be the major toxin genes, but the ces gene was not detected in any of the tested B. thuringiensis isolates. The hemolysin BL enterotoxin was detected in all 30 B. thuringiensis isolates (100%). The non-hemolytic enterotoxin complex was found in 27 out of 30 B. thuringiensis isolates (90.0%). The B. thuringiensis tested in this study had similar toxin gene characteristics to B. cereus , which possessed more than one toxin gene. B. thuringiensis could have the potential risk of foodborne illness based on the toxin genes and toxin-producing ability.

  12. Effect of Bacillus thuringiensis on microbial functional groups in sorghum rhizosphere Efeito do Bacillus thuringiensis sobre grupos funcionais de microrganismos na rizosfera de sorgo

    Directory of Open Access Journals (Sweden)

    Carlos Brasil

    2006-05-01

    Full Text Available The objective of this work was to assess the effect of two strains of Bacillus thuringiensis var. kurstaki on sorghum rhizosphere microorganisms. The strains were HD1, that produces the bioinsecticidal protein, and 407, that is a mutant non-producer. The strains do not influence microbial population, but reduce plant growth and improve mycorrhizal colonization and free living fixing N2 community.O objetivo deste trabalho foi avaliar o efeito de duas cepas de Bacillus thuringiensis var. kurstaki sobre microrganismos na rizosfera do sorgo. As cepas foram a HD1, produtora do cristal bioinseticida, e a 407, uma mutante não-produtora. As duas cepas não influenciam a comunidade microbiana, mas reduzem o crescimento da planta. A colonização micorrízica e a população de fixadores de N2 de vida livre aumentaram.

  13. Effect of Cry1Ab protein on rhizobacterial communities of Bt-maize over a four-year cultivation period.

    Directory of Open Access Journals (Sweden)

    Jorge Barriuso

    Full Text Available BACKGROUND: Bt-maize is a transgenic variety of maize expressing the Cry toxin from Bacillus turingiensis. The potential accumulation of the relative effect of the transgenic modification and the cry toxin on the rhizobacterial communities of Bt-maize has been monitored over a period of four years. METHODOLOGY/PRINCIPAL FINDINGS: The accumulative effects of the cultivation of this transgenic plant have been monitored by means of high throughput DNA pyrosequencing of the bacterial DNA coding for the 16S rRNA hypervariable V6 region from rhizobacterial communities. The obtained sequences were subjected to taxonomic, phylogenetic and taxonomic-independent diversity studies. The results obtained were consistent, indicating that variations detected in the rhizobacterial community structure were possibly due to climatic factors rather than to the presence of the Bt-gene. No variations were observed in the diversity estimates between non-Bt and Bt-maize. CONCLUSIONS/SIGNIFICANCE: The cultivation of Bt-maize during the four-year period did not change the maize rhizobacterial communities when compared to those of the non-Bt maize. This is the first study to be conducted with Bt-maize during such a long cultivation period and the first evaluation of rhizobacterial communities to be performed in this transgenic plant using Next Generation Sequencing.

  14. Advances in developing Bacillus thuringiensis-based insecticde formulations Avances en el desarrollo de formulaciones insecticidas a base de Bacillus thuringiensis

    OpenAIRE

    Rosas-García Ninfa María

    2008-01-01

    Developing Bacillus thuringiensis-based formulations is an old technology which has been revived during recent decades. The spore-crystal complex (being the main ingredient in these preparations) has been the main objective of this research, involving the search for new or improved strains. The type of materials used included a wide variety of completely biodegradable ingredients which could have been leaves, stems or fruit which when dried and ground could serve as feeding stimulants, as wel...

  15. Contributions of gut bacteria to Bacillus thuringiensis-induced mortality vary across a range of Lepidoptera

    Science.gov (United States)

    Broderick, Nichole A; Robinson, Courtney J; McMahon, Matthew D; Holt, Jonathan; Handelsman, Jo; Raffa, Kenneth F

    2009-01-01

    Background Gut microbiota contribute to the health of their hosts, and alterations in the composition of this microbiota can lead to disease. Previously, we demonstrated that indigenous gut bacteria were required for the insecticidal toxin of Bacillus thuringiensis to kill the gypsy moth, Lymantria dispar. B. thuringiensis and its associated insecticidal toxins are commonly used for the control of lepidopteran pests. A variety of factors associated with the insect host, B. thuringiensis strain, and environment affect the wide range of susceptibilities among Lepidoptera, but the interaction of gut bacteria with these factors is not understood. To assess the contribution of gut bacteria to B. thuringiensis susceptibility across a range of Lepidoptera we examined larval mortality of six species in the presence and absence of their indigenous gut bacteria. We then assessed the effect of feeding an enteric bacterium isolated from L. dispar on larval mortality following ingestion of B. thuringiensis toxin. Results Oral administration of antibiotics reduced larval mortality due to B. thuringiensis in five of six species tested. These included Vanessa cardui (L.), Manduca sexta (L.), Pieris rapae (L.) and Heliothis virescens (F.) treated with a formulation composed of B. thuringiensis cells and toxins (DiPel), and Lymantria dispar (L.) treated with a cell-free formulation of B. thuringiensis toxin (MVPII). Antibiotics eliminated populations of gut bacteria below detectable levels in each of the insects, with the exception of H. virescens, which did not have detectable gut bacteria prior to treatment. Oral administration of the Gram-negative Enterobacter sp. NAB3, an indigenous gut resident of L. dispar, restored larval mortality in all four of the species in which antibiotics both reduced susceptibility to B. thuringiensis and eliminated gut bacteria, but not in H. virescens. In contrast, ingestion of B. thuringiensis toxin (MVPII) following antibiotic treatment

  16. [Septicaemia of chironomid larvae (Diptera: Chironomidae) promoted by Bacillus cereus and B. thuringiensis].

    Science.gov (United States)

    Khodyrev, V P

    2012-01-01

    Natural factors regulating the population of chironomids were studied. The bacteria Bacillus cereus were isolated from chironomids sampled from Kuyalnitskii Firth after epizooty of Chironomus sp., and bacteria Bacillus thuringiensis spp. israelensis (Bti) were isolated from dead larva of Chironomus plumosus sampled in the Sea of Azov (3-m depth). Bti were characterized by high insecticide activity on midges Anopheles messeae Fall., Aedes cireneus Mg., and Culex pipiens pipiens f. pipiens L.

  17. Isolation and partial characterization of a mutant of Bacillus thuringiensis producing melanin Isolamento e caracterização parcial de um mutante de Bacillus thuringiensis produtor de melanina

    Directory of Open Access Journals (Sweden)

    Gislayne T. Vilas-Bôas

    2005-09-01

    Full Text Available A mutant (407-P of Bacillus thuringiensis subsp. thuringiensis strain 407 producing a melanin was obtained after treatment with the mutagenic agent ethyl-methane-sulfonate. Several microbiological and biochemical properties of the two strains were analyzed and the results were similar. The mutant 407-P was also incorporated into non-sterilized soil samples, recovered, easily identified, and quantified, what enables its use in ecology of B. thuringiensis.Um mutante (407-P da linhagem Bacillus thuringiensis subsp. thuringiensis 407 produtor de melanina foi obtido após tratamento com o agente mutagênico etil-metano-sulfonato. Diversas propriedades microbiológicas e bioquímicas das duas linhagens foram analisadas e os resultados foram similares. O mutante 407-P foi incorporado em amostras de solo não esterilizado, recuperado, facilmente identificado e quantificado, possibilitando seu uso em estudos de ecologia de B. thuringiensis.

  18. Differentiation between Bacillus thuringiensis strains by gyrB PCR-Sau3AI fingerprinting.

    Science.gov (United States)

    Awad, Mireille Kallassy; Saadaoui, Imène; Rouis, Souad; Tounsi, Slim; Jaoua, Samir

    2007-02-01

    gyrB DNA fragments of seven Bacillus thuringiensis local collection family representatives were amplified by PCR and sequenced. Several differences in their corresponding sequences were evidenced. Both in silico and in vitro restriction maps of gyrB sequences and fragments respectively confirmed that EcoRI and Sau3AI could be used to differentiate between B. thuringiensis strains. However, the phylogeny analysis showed that only the gyrB PCR-Sau3AI allows a strains classification that correlates very well with that obtained on the basis of the sequences analysis. Thus, these finds show that gyrB PCR- Sau3AI digestion could be considered as an efficient, rapid, and easy method to make a distinction, not only between strains belonging to the Bacillus cereus group, but also between those belonging to B. thuringiensis.

  19. A method for in Vivo radiolabeling Bacillus thuringiensis native δ-endotoxin crystals

    Science.gov (United States)

    C. Noah Koller; Leah S. Bauer; Robert M. Hollingworth

    1995-01-01

    The entomocidal CryIIIA δ-endotoxin protein of Bacillus thuringiensis var. tenebrionis is distinctive in chemistry and host range. In contrast to other δ-endotoxins, the CryIIIA parasporal crystals are toxic within the acidic midgut environment of several coleopteran species, particularly those in the family...

  20. Effect of chemical additives on Bacillus thuringiensis (Bacillales: Bacillaceae) against Plutella xylostella (Lepidoptera: Pyralidae)

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Qiu, S.; Huang, T.; Huang, Z.; Xu, L.; Wu, C.; Gelbič, Ivan; Guan, X.

    2013-01-01

    Roč. 106, č. 3 (2013), s. 1075-1080 ISSN 0022-0493 Institutional research plan: CEZ:AV0Z50070508 Keywords : additives * Bacillus thuringiensis * biocontrol Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection Impact factor: 1.605, year: 2013 http://www.bioone.org/doi/pdf/10.1603/EC12288

  1. Resistance: a threat to the insecticidal crystal proteins of Bacillus thuringiensis

    Science.gov (United States)

    Leah S. Bauer

    1995-01-01

    Insecticidal crystal proteins (also known as d-endotoxins) synthesized by the bacterium Bacillus thuringiensis Berliner (Bt) are the active ingredient of various environmentally friendly insecticides that are 1) highly compatible with natural enemies and other nontarget organisms due to narrow host specificity, 2) harmless to vertebrates, 3) biodegradable in the...

  2. A new formulation of Bacillus thuringiensis: UV protection and sustained release mosquito larvae studies

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Zhang, X.; Zhang, Y.; Wu, S.; Gelbič, Ivan; Xu, L.; Guan, X.

    2016-01-01

    Roč. 6, DEC 22 (2016), č. článku 39425. ISSN 2045-2322 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * pest control * UV protection Subject RIV: EE - Microbiology, Virology Impact factor: 4.259, year: 2016 http://www.nature.com/articles/srep39425

  3. Potato expressing beetle-specific Bacillus thuringiensis Cry3Aa toxin reduces performance of a moth

    Czech Academy of Sciences Publication Activity Database

    Hussein, H. M.; Habuštová, Oxana; Turanli, Ferit; Sehnal, František

    2006-01-01

    Roč. 32, č. 1 (2006), s. 1-13 ISSN 0098-0331 R&D Projects: GA ČR(CZ) GA522/02/1507 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis * Spodoptera littoralis * Leptinotarsa decemlineata Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection Impact factor: 1.896, year: 2006

  4. Characterization of Baculovirus Insecticides Expressing Tailored Bacillus thuringiensis CryIA(b) Crystal Proteins

    NARCIS (Netherlands)

    Martens, John W M; Knoester, Marga; Weijts, Franci; Groffen, Sander J A; Hu, Zhihong; Bosch, Dirk; Vlak, Just M.

    1995-01-01

    Full-length, truncated, and mature forms of the CryIA(b) insecticidal crystal protein gene of Bacillus thuringiensis were engineered into the p10 locus of Autographa californica nuclear polyhedrosis virus (AcNPV). A signal sequence of Heliothis virescens juvenile hormone esterase was introduced at

  5. Laboratory Assessment of the Effects of Bacillus thuringiensis on Native Lepidoptera

    Science.gov (United States)

    John W. Peacock; Dale F. Schweitzer; Jane L. Carter; Normand R. Dubois

    1998-01-01

    The effect of 2 formulations of Bacillus thuringiensis subsp. kurstaki (Foray 48B and Dipel 8AF) was evaluated on 42 species of native Lepidoptera in laboratory bioassays using instars that are present in the field at the time of gypsy moth suppression applications. Mortality was significant for 27 of the 42 species evaluated...

  6. Laboratory and field studies on the effects of Bacillus thuringiensis on non-target lepidoptera

    Science.gov (United States)

    John Peacock; Stephen Talley; Taylor Williams; Richard. Reardon

    1992-01-01

    Bacillus thuringiensis (B.t.) is one of the insecticides considered effective for suppression of gypsy moth infestations, and it is considered to one of the most selective in terms of its effects on other insects. Although B.t. is touted to be "environmentally safe", there is a paucity of field data to support this claim, particularly as...

  7. Use of spent mushroom substrate for production of Bacillus thuringiensis by solid-state fermentation

    Czech Academy of Sciences Publication Activity Database

    Wu, S.; Lan, Y.; Huang, D.; Peng, Y.; Huang, Z.; Xu, L.; Gelbič, Ivan; Carballar-Lejarazu, R.; Guan, X.; Zhang, L.; Zou, S.

    2014-01-01

    Roč. 107, č. 1 (2014), s. 137-143 ISSN 0022-0493 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * spent mushroom substrate * solid-state fermentation Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection Impact factor: 1.506, year: 2014 http://www.bioone.org/doi/pdf/10.1603/EC13276

  8. Biological activity of Bacillus thuringiensis (Bacillales: Bacillaceae) chitinase against Caenorhabditis elegans (Rhabditida: Rhabditidae)

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Yu, J.; Xie, Y.; Lin, H.; Huang, Z.; Xu, L.; Gelbič, Ivan; Guan, X.

    2014-01-01

    Roč. 107, č. 2 (2014), s. 551-558 ISSN 0022-0493 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * Caenorhabditis elegans * chitinase Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection Impact factor: 1.506, year: 2014 http://www.bioone.org/doi/ pdf /10.1603/EC13201

  9. The occurrence of Photorhabdus-like toxin complexes in Bacillus thuringiensis

    Science.gov (United States)

    Recently, genomic sequencing of a Bacillus thuringiensis (Bt) isolate from our collection revealed the presence of an apparent operon encoding an insecticidal toxin complex (Tca) similar to that first described from the entomopathogen Photorhabdus luminescens. To determine whether these genes are w...

  10. Characterization of cry1Cb3 and cry1Fb7 from Bacillus thuringiensis subsp. galleriae

    Czech Academy of Sciences Publication Activity Database

    Huang, T.; Xiao, Y.; Pan, J.; Zhang, L.; Gelbič, Ivan; Guan, X.

    2015-01-01

    Roč. 10, č. 1 (2015), s. 521-528 ISSN 2391-5412 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis subsp. galleriae * PCR-RFLP * cloning Subject RIV: EB - Genetics ; Molecular Biology http://www.degruyter.com/view/j/biol.2015.10.issue-1/biol-2015-0054/biol-2015-0054.xml

  11. Solubilization, Activation, and Insecticidal Activity of Bacillus thuringiensis Serovar thompsoni HD542 Crystal Proteins

    NARCIS (Netherlands)

    Naimov, S.; Boncheva, R.; Karlova, R.B.; Dukiandjiev, S.; Minkov, I.; Maagd, de R.A.

    2008-01-01

    Cry15Aa protein, produced by Bacillus thuringiensis serovar thompsoni HD542 in a crystal together with a 40 kDa accompanying protein is one of a small group of non-typical, less well-studied members of the Cry family of insecticidal proteins, and may provide an alternative for the more commonly used

  12. Investigation of Cytocidal Activity of Bacillus Thuringiensis Parasporal Toxin on CCRF-CEM Cell Line

    Directory of Open Access Journals (Sweden)

    Elham Moazamian

    2013-03-01

    Full Text Available Background & Objective: Parasporin is a parasporal protein of Bacillus thuringiensis and exhibits special cytocidal activity against human cancer cells. Similar to other insecticidal Bacillus thuringiensis crystal toxins, parasporin shows target specificity and damages the cellular membrane. In this study, different strains of Bacillus thuringiensis isolated from various regions of Iran and their cytocidal activity against CCRF-CEM cell line and human erythrocyte were investigated.   Materials & Methods: Fifty soil samples were collected from different Iranian provinces, and characterization was performed based on protein crystal morphology by phase-contrast microscope and variations of Cry protein toxin using SDS-PAGE. After parasporin was processed with proteinase K, the active form was produced and protein activity on the cell line was evaluated. Results: Parasporal inclusion proteins showed different cytotoxicity against acute lymphoblastic leukemia cells (ALL, but not against normal lymphocyte. Isolated parasporin demonstrated no hemolytic activity against human erythrocyte. It appears that these proteins have the ability to differentiate between normal lymphocytes and leukemia cells and have specific receptors on specific cancer cell lines. Conclusion: Our results provide evidence that the parasporin-producing organism is a common member in Bacillus thuringiensis populations occurring in the natural environments of Iran.

  13. IMPACT OF BT ( BACILLUS THURINGIENSIS ) CROPS ON BAT ACTIVITY IN SOUTH TEXAS AGROECOSYSTEMS

    Science.gov (United States)

    The widespread adoption of transgenic insecticidal crops raises concerns that nontarget species may be harmed and food webs disrupted. The goal of this research is to determine how transgenic Bt (Bacillus thuringiensis) crops impact the activity of Brazilian freetailed bats (Tada...

  14. Regulation by gut bacertia of immune response, Bacillus thuringiensis susceptibility and hemolin expression in Plodia interpunctella

    Science.gov (United States)

    Plodia interpunctella (Hübner) is an important stored grain insect pest worldwide, and the first lepidopteran with reported resistance to Bacillus thuringiensis (Bt) toxins. Since gut bacteria may affect Bt insecticidal activity, we determined whether P. interpunctella lacking gut enterobacteria had...

  15. Recent field experiences with Bacillus thuringiensis in Canada and research needs

    Science.gov (United States)

    Oswald N. Morris

    1985-01-01

    The CANUSA working group on the use of B.t. against the spruce budworm has prepared a document entitled "Guidelines for the operational use of Bacillus thuringiensis (B.t.) against the spruce budworm" following six years of extensive cooperative field trials in Canada and the U.S.A. (Morris et al 1984). The document summarized below (Table...

  16. Bacillus thuringiensis toxins trigger receptor shedding from gypsy moth midgut cells

    Science.gov (United States)

    Algimantas P. Valaitis

    2007-01-01

    The mechanism of action of the Cry1 insecticidal proteins produced by Bacillus thuringiensis (Bt) begins with the processing of these proteins in the larval gut. After proteolytic activation, the Bt toxins bind to specific midgut receptors and insert into the membrane of the gut epithelial cells, causing insect death.

  17. Response of the Cottonwood Leaf Beetle (Coleoptera: Chrysomelidae) to Bacillus thuringiensis var. san diego

    Science.gov (United States)

    Leah S. Bauer

    1990-01-01

    A standardized laboratory bioassay was used to quantify the lethal and sub-lethal responses of larval and adult cottonwood leaf beetles, Chrysomela scripta F., to Bacillus thuringiensis var. san diego, formulated as M-One standard powder (Mycogen Corporation, San Diego). The median lethal concentration (LC

  18. Effects of Bacillus thuringiensis CRY1A(c) δ-endotoxin on growth ...

    African Journals Online (AJOL)

    STORAGESEVER

    2010-01-04

    Jan 4, 2010 ... The recent introduction of Bt maize and Bt cotton transgenic crops into Africa has raised concerns on their potential short and long-term ecological effects on the environment. The effects of Bacillus thuringiensis (Bt) Cry1A(c) δ-endotoxin on the growth, nodulation and productivity of two leguminous.

  19. Effects of Bacillus thuringiensis CRY1A(c) d-endotoxin on growth ...

    African Journals Online (AJOL)

    The recent introduction of Bt maize and Bt cotton transgenic crops into Africa has raised concerns on their potential short and long-term ecological effects on the environment. The effects of Bacillus thuringiensis (Bt) Cry1A(c) d-endotoxin on the growth, nodulation and productivity of two leguminous plants grown in clay soil ...

  20. Chemical modulators of the innate immune response alter gypsy moth larval susceptibility to Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Broderick Nichole A

    2010-04-01

    Full Text Available Abstract Background The gut comprises an essential barrier that protects both invertebrate and vertebrate animals from invasion by microorganisms. Disruption of the balanced relationship between indigenous gut microbiota and their host can result in gut bacteria eliciting host responses similar to those caused by invasive pathogens. For example, ingestion of Bacillus thuringiensis by larvae of some species of susceptible Lepidoptera can result in normally benign enteric bacteria exerting pathogenic effects. Results We explored the potential role of the insect immune response in mortality caused by B. thuringiensis in conjunction with gut bacteria. Two lines of evidence support such a role. First, ingestion of B. thuringiensis by gypsy moth larvae led to the depletion of their hemocytes. Second, pharmacological agents that are known to modulate innate immune responses of invertebrates and vertebrates altered larval mortality induced by B. thuringiensis. Specifically, Gram-negative peptidoglycan pre-treated with lysozyme accelerated B. thuringiensis-induced killing of larvae previously made less susceptible due to treatment with antibiotics. Conversely, several inhibitors of the innate immune response (eicosanoid inhibitors and antioxidants increased the host's survival time following ingestion of B. thuringiensis. Conclusions This study demonstrates that B. thuringiensis infection provokes changes in the cellular immune response of gypsy moth larvae. The effects of chemicals known to modulate the innate immune response of many invertebrates and vertebrates, including Lepidoptera, also indicate a role of this response in B. thuringiensis killing. Interactions among B. thuringiensis toxin, enteric bacteria, and aspects of the gypsy moth immune response may provide a novel model to decipher mechanisms of sepsis associated with bacteria of gut origin.

  1. Binding analysis of Bacillus thuringiensis Cry1 proteins in the sugarcane borer, Diatraea saccharalis (Lepidoptera: Crambidae).

    Science.gov (United States)

    Davolos, Camila C; Hernández-Martinez, Patricia; Crialesi-Legori, Paula C B; Desidério, Janete A; Ferré, Juan; Escriche, Baltasar; Lemos, Manoel Victor F

    2015-05-01

    Sugarcane borer (Diatraea saccharalis, F.) is an important corn pest in South America and United States. The aim of the present study was to analyze the susceptibility and binding interactions of three Cry1A proteins and Cry1Fa in a Brazilian D. saccharalis population. The results showed that Cry1Ab was the most active, followed by Cry1Ac, Cry1Fa and Cry1Aa. All Cry1-biotinylated proteins tested bound specifically to the D. saccharalis brush border membrane vesicles (BBMV). Heterologous competition assays showed shared binding sites for all Cry1A proteins and another one shared by Cry1Fa and Cry1Ab. Thus, pyramiding Cry1Aa/Cry1Ac and Cry1F proteins would be a recommended strategy for managing this pest. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. New Bacillus thuringiensis toxin combinations for biological control of lepidopteran larvae.

    Science.gov (United States)

    Elleuch, Jihen; Zghal, Raida Zribi; Jemaà, Mohamed; Azzouz, Hichem; Tounsi, Slim; Jaoua, Samir

    2014-04-01

    Cyt1Aa from Bacillus thuringiensis israelensis is known by its synergistical activity with B. thuringiensis and Bacillus sphaericus toxins. It is able to improve dipteran specific toxins activity and can prevent or overcome larval resistance to those proteins. The objective of the current study was to investigate the possible improvement of larvicidal activity of B. thuringiensis kurstaki expressing heterogeneous proteins Cyt1A and P20. cyt1A98 and p20 genes encoding the cytolytic protein (Cyt1A98) and the accessory protein (P20), respectively, were introduced individually and in combination into B. thuringiensis kurstaki strain BNS3. Immunoblot analysis evidenced the expression of these genes in the recombinant strains and hinted that P20 acts as molecular chaperone protecting Cyt1A98 from proteolytic attack in BNS3. The toxicities of recombinant strains were studied and revealed that BNS3pHTp20 exhibited higher activity than that of the negative control (BNS3pHTBlue) toward Ephestia kuehniella, but not toward Spodoptera littoralis. When expressed in combination with P20, Cyt1A98 enhanced BNS3 activity against E. kuehniella and S. littoralis. Thus, Cyt1Aa protein could enhance lepidopteran Cry insecticidal activity and would prevent larval resistance to the most commercialized B. thuringiensis kurstaki toxins. Copyright © 2014 Elsevier B.V. All rights reserved.

  3. Genomic and transcriptomic insights into the efficient entomopathogenicity of Bacillus thuringiensis.

    Science.gov (United States)

    Zhu, Lei; Peng, Donghai; Wang, Yueying; Ye, Weixing; Zheng, Jinshui; Zhao, Changming; Han, Dongmei; Geng, Ce; Ruan, Lifang; He, Jin; Yu, Ziniu; Sun, Ming

    2015-09-28

    Bacillus thuringiensis has been globally used as a microbial pesticide for over 70 years. However, information regarding its various adaptions and virulence factors and their roles in the entomopathogenic process remains limited. In this work, we present the complete genomes of two industrially patented Bacillus thuringiensis strains (HD-1 and YBT-1520). A comparative genomic analysis showed a larger and more complicated genome constitution that included novel insecticidal toxicity-related genes (ITRGs). All of the putative ITRGs were summarized according to the steps of infection. A comparative genomic analysis showed that highly toxic strains contained significantly more ITRGs, thereby providing additional strategies for infection, immune evasion, and cadaver utilization. Furthermore, a comparative transcriptomic analysis suggested that a high expression of these ITRGs was a key factor in efficient entomopathogenicity. We identified an active extra urease synthesis system in the highly toxic strains that may aid B. thuringiensis survival in insects (similar to previous results with well-known pathogens). Taken together, these results explain the efficient entomopathogenicity of B. thuringiensis. It provides novel insights into the strategies used by B. thuringiensis to resist and overcome host immune defenses and helps identify novel toxicity factors.

  4. Sensitivity of the bacterium Bacillus Thuringiensis as an insect disease agent to gamma-rays

    International Nuclear Information System (INIS)

    Merdam, A.I.; Abdu, R.M.

    1977-01-01

    The effect of gamma radiation on the viability of the entomopathogenic spore-forming bacterium, Bacillus thuringiensis, was tested. The different gamma doses varied much in their effect on such bacterium. All irradiated Bacillus suspensions with doses below 85 krad showed different degrees of inhibitory activity. However, bacterial suspensions irradiated at a dose of 90 krad. proved to promote spore germination. Changes in the physiological, and morphological characters of the irradiated Bacillus at these levels were detected. The new observed characters were induced at a particular dose level of 90 krad. These new characters are assumed to be due to genetic changes induced at this particular gamma dose

  5. Analysis of Bacillus thuringiensis Population Dynamics and Its Interaction With Pseudomonas fluorescens in Soil

    Science.gov (United States)

    Rojas-Ruiz, Norma Elena; Sansinenea-Royano, Estibaliz; Cedillo-Ramirez, Maria Lilia; Marsch-Moreno, Rodolfo; Sanchez-Alonso, Patricia; Vazquez-Cruz, Candelario

    2015-01-01

    Background: Bacillus thuringiensis is the most successful biological control agent, however, studies so far have shown that B. thuringiensis is very sensitive to environmental factors such as soil moisture and pH. Ultraviolet light from the sun had been considered as the main limiting factor for its persistence in soil and it has recently been shown that the antagonism exerted by other native soil organisms, such as Pseudomonas fluorescens, is a determining factor in the persistence of this bacterium under in vitro culture conditions. Objectives: The aim of the present investigation was to analyze the population dynamics of B. thuringiensis and its interaction with P. fluorescens using microbiological and molecular methods in soil, under different conditions, and to determinate the effect of nutrients and moisture on its interaction. Materials and Methods: The monitoring was performed by microbiological methods, such as viable count of bacteria, and molecular methods such as Polymerase Chain Reaction (PCR) and hybridization, using the direct extraction of DNA from populations of inoculated soil. Results: The analysis of the interaction between B. thuringiensis and P. fluorescens in soil indicated that the disappearance of B. thuringiensis IPS82 is not dependent on the moisture but the composition of nutrients that may be affecting the secretion of toxic compounds in the environment of P. fluorescens. The results showed that the recovered cells were mostly spores and not vegetative cells in all proved treatments. The molecular methods were effective for monitoring bacterial population inoculated in soil. Conclusions: Bacillus thuringiensis is very sensitive to the interaction of P. fluorescens, however is capable to survive in soil due to its capacity of sporulate. Some of the cells in the form of spores germinated and folded slightly and remained in a constant cycle of sporulation and germination. This confirms that B. thuringiensis IPS82 can germinate, grow and

  6. RAP-PCR fingerprinting reveals time-dependent expression of development-related genes following differentiation process of Bacillus thuringiensis

    Czech Academy of Sciences Publication Activity Database

    Huang, T.; Yu, X.; Gelbič, Ivan; Guan, X.

    2015-01-01

    Roč. 61, č. 9 (2015), s. 683-690 ISSN 0008-4166 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * development * RNA arbitrarily primed PCR Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.335, year: 2015

  7. Resistance to Bacillus thuringiensis linked with a cadherin transmembrane mutation affecting cellular trafficking in pink bollworm from China

    Science.gov (United States)

    Transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) are cultivated extensively worldwide. However, their benefits are being eroded by increasingly rapid evolution of resistance in pests. In some previously analyzed strains of three major lepidopteran pests, resistance t...

  8. Interactions between Bacillus thuringiensis and parasitoids of late-instar larvae of the spruce budworm (Lepidoptera: Tortricidae)

    NARCIS (Netherlands)

    Schoenmaker, A.; Cusson, M.; Frankenhuyzen, van K.

    2001-01-01

    We investigated interactions between Bacillus thuringiensis Berliner var. kurstaki and parasitoids that attack late instars of the eastern spruce budworm, Choristoneura fumiferana (Clemens). In a petri-dish arena, females of Tranosema rostrale rostrale (Brishke) (Hymenoptera: Ichneumonidae) were

  9. Microimaging of Bacillus thuringiensis Toxin-binding proteins in gypsy moth larval gut using confocal fluorescence microscopy

    Science.gov (United States)

    Daniel J. Krofcheck; Algimantas P. Valaitis

    2010-01-01

    After ingestion by susceptible insect larvae, Bacillus thuringiensis (Bt) insecticidal proteins bind to the brush border membranes of gut epithelial cells and disrupt the integrity of the plasma membrane by forming...

  10. Comparison of sampling methods to recover germinated Bacillus anthracis and Bacillus thuringiensis endospores from surface coupons.

    Science.gov (United States)

    Mott, T M; Shoe, J L; Hunter, M; Woodson, A M; Fritts, K A; Klimko, C P; Quirk, A V; Welkos, S L; Cote, C K

    2017-05-01

    In an attempt to devise decontamination methods that are both effective and minimally detrimental to the environment, we evaluated germination induction as an enhancement to strategies for Bacillus anthracis spore decontamination. To determine an optimal method for the recovery of germinating spores from different matrices, it was critical to ensure that the sampling procedures did not negatively impact the viability of the germinating spores possibly confounding the results and downstream analyses of field trial data. Therefore, the two main objectives of this study were the following: (i) development of an effective processing protocol capable of recovering the maximum number of viable germinating or germinated spores from different surface materials; and (ii) using a model system of spore contamination, employ this protocol to evaluate the potential applicability of germination induction to wide-area decontamination of B. anthracis spores. We examined parameters affecting the sampling efficiencies of B. anthracis and the surrogate species Bacillus thuringiensis on nonporous and porous materials. The most efficient extraction from all matrices was observed using PBS with 0·01% Tween 80 extraction buffer. The addition of a sonication and/or extended vortex treatment did not yield significant increases in spore or germinated spore recovery. Our data demonstrate that previous germination-induction experiments performed in suspension can be reproduced when Bacillus spores are deposited onto reference surfaces materials. Our proof of concept experiment illustrated that a germination pretreatment step significantly improves conventional secondary decontamination strategies and remediation plans. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.

  11. Decreased toxicity of Bacillus thuringiensis subsp. israelensis to mosquito larvae after contact with leaf litter.

    Science.gov (United States)

    Tetreau, Guillaume; Stalinski, Renaud; Kersusan, Dylann; Veyrenc, Sylvie; David, Jean-Philippe; Reynaud, Stéphane; Després, Laurence

    2012-08-01

    Bacillus thuringiensis subsp. israelensis is a bacterium producing crystals containing Cry and Cyt proteins, which are toxic for mosquito larvae. Nothing is known about the interaction between crystal toxins and decaying leaf litter, which is a major component of several mosquito breeding sites and represents an important food source. In the present work, we investigated the behavior of B. thuringiensis subsp. israelensis toxic crystals sprayed on leaf litter. In the presence of leaf litter, a 60% decrease in the amount of Cyt toxin detectable by immunology (enzyme-linked immunosorbent assays [ELISAs]) was observed, while the respective proportions of Cry toxins were not affected. The toxicity of Cry toxins toward Aedes aegypti larvae was not affected by leaf litter, while the synergistic effect of Cyt toxins on all B. thuringiensis subsp. israelensis Cry toxins was decreased by about 20% when mixed with leaf litter. The toxicity of two commercial B. thuringiensis subsp. israelensis strains (VectoBac WG and VectoBac 12AS) and a laboratory-produced B. thuringiensis subsp. israelensis strain decreased by about 70% when mixed with leaf litter. Taken together, these results suggest that Cyt toxins interact with leaf litter, resulting in a decreased toxicity of B. thuringiensis subsp. israelensis in litter-rich environments and thereby dramatically reducing the efficiency of mosquitocidal treatments.

  12. Decreased Toxicity of Bacillus thuringiensis subsp. israelensis to Mosquito Larvae after Contact with Leaf Litter

    Science.gov (United States)

    Stalinski, Renaud; Kersusan, Dylann; Veyrenc, Sylvie; David, Jean-Philippe; Reynaud, Stéphane; Després, Laurence

    2012-01-01

    Bacillus thuringiensis subsp. israelensis is a bacterium producing crystals containing Cry and Cyt proteins, which are toxic for mosquito larvae. Nothing is known about the interaction between crystal toxins and decaying leaf litter, which is a major component of several mosquito breeding sites and represents an important food source. In the present work, we investigated the behavior of B. thuringiensis subsp. israelensis toxic crystals sprayed on leaf litter. In the presence of leaf litter, a 60% decrease in the amount of Cyt toxin detectable by immunology (enzyme-linked immunosorbent assays [ELISAs]) was observed, while the respective proportions of Cry toxins were not affected. The toxicity of Cry toxins toward Aedes aegypti larvae was not affected by leaf litter, while the synergistic effect of Cyt toxins on all B. thuringiensis subsp. israelensis Cry toxins was decreased by about 20% when mixed with leaf litter. The toxicity of two commercial B. thuringiensis subsp. israelensis strains (VectoBac WG and VectoBac 12AS) and a laboratory-produced B. thuringiensis subsp. israelensis strain decreased by about 70% when mixed with leaf litter. Taken together, these results suggest that Cyt toxins interact with leaf litter, resulting in a decreased toxicity of B. thuringiensis subsp. israelensis in litter-rich environments and thereby dramatically reducing the efficiency of mosquitocidal treatments. PMID:22610426

  13. 40 CFR 174.505 - Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis modified Cry3A... of Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn are exempt from the requirement of... REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.505 Bacillus...

  14. 40 CFR 174.519 - Bacillus thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry2Ab2 protein... Bacillus thuringiensis Cry2Ab2 protein in or on corn or cotton are exempt from the requirement of a... REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.519 Bacillus...

  15. 40 CFR 174.501 - Bacillus thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Vip3Aa protein... Bacillus thuringiensis Vip3Aa proteins in or on corn or cotton are exempt from the requirement of a... REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.501 Bacillus...

  16. 40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry34Ab1 and... Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of a... REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.506 Bacillus...

  17. 40 CFR 174.510 - Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1Ac protein... PLANT-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.510 Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus...

  18. Immunotoxicological Evaluation of Genetically Modified Rice Expressing Cry1Ab/Ac Protein (TT51-1) by a 6-Month Feeding Study on Cynomolgus Monkeys

    Science.gov (United States)

    Tan, Xiaoyan; Zhou, Xiaobing; Tang, Yao; Lv, Jianjun; Zhang, Lin; Sun, Li; Yang, Yanwei; Miao, Yufa; Jiang, Hua; Chen, Gaofeng; Huang, Zhiying; Wang, Xue

    2016-01-01

    The present study was performed to evaluate the food safety of TT51-1, a new type of genetically modified rice that expresses the Cry1Ab/Ac protein (Bt toxin) and is highly resistant to most lepidopteran pests. Sixteen male and 16 female cynomolgus monkeys were randomly divided into four groups: conventional rice (non-genetically modified rice, non-GM rice), positive control, 17.5% genetically modified rice (GM rice) and 70% GM rice. Monkeys in the non-GM rice, positive control, and GM rice groups were fed on diets containing 70% non-GM rice, 17.5% GM rice or 70% GM rice, respectively, for 182 days, whereas animals in the positive group were intravenously injected with cyclophosphamide every other day for a total of four injections before the last treatment. Six months of treatment did not yield abnormal observations. Specifically, the following parameters did not significantly differ between the non-GM rice group and GM rice groups: body weight, food consumption, electrocardiogram, hematology, immuno-phenotyping of lymphocytes in the peripheral blood, mitogen-induced peripheral blood lymphocyte proliferation, splenocyte proliferation, KLH-T cell-dependent antibody response, organ weights and ratios, and histological appearance (p>0.05). Animals from the GM rice group differed from animals in the non-GM rice group (p<0.05) in several parameters: specifically, their body temperatures and serum alanine aminotransferase (ALT) levels were higher, whereas their levels of serum K+, Cl- and cytokines (IL-2, IL-4 and IL-5) were lower. Because dose- or time-dependent changes were not observed in this study and animals appeared histologically normal, the aforementioned differences were not considered to be adverse or related to the treatment with GM rice. In conclusion, a 6-month feeding study of TT51-1 did not show adverse immunotoxicological effects on cynomolgus monkeys. PMID:27684490

  19. 40 CFR 174.532 - Bacillus thuringiensis eCry3.1Ab protein in corn; temporary exemption from the requirement of a...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis eCry3.1Ab... Bacillus thuringiensis eCry3.1Ab protein in corn, in or on the food and feed commodities of corn; corn... Bacillus thuringiensis eCry3.1Ab protein in corn is used as a plant-incorporated protectant in accordance...

  20. Evaluation de l'efficacité de la Bacillus thuringiensis contre les larves et imagos de Andrector ruficornis sur les plants de Solanum tuberosum au Cameroun

    OpenAIRE

    Ouzounov, IS.; Omokolo, ND.; Ambang, Z.

    2002-01-01

    Evaluation of the Efficiency of Bacillus thuringiensis on Larvae and Adults of Andrector ruficornis on Solanum tuberosum plants in Cameroon. The efficiency of Bacillus thuringiensis as an insecticide on the larvae and adults of A. ruficornis (a leaf eating pest) was investigated on Solanum tuberosum plants grown in the field. The results show that, one hour following the treatment of A. ruficornis with a 1.4 g/l suspension of Bacillus thuringiensis, larvae and adults are paralysed on the plan...

  1. Frequency and distribution of Bacillus Thuringiensis from Ethiopian ...

    African Journals Online (AJOL)

    Two hundred and thirteen B. thuringiensis isolates were tested for larvicidal activity against An. arabiensis (Diptera) and Plutella xylostella (Lepidoptera) larvae. Of the tested isolates, 44 (21%) killed 50-100% of An. arabiensis larvae within 48 hours. Isolates that killed 100% larvae within 24 hours were all from tepid to cool ...

  2. Structural Insights into Bacillus thuringiensis Cry, Cyt and Parasporin Toxins

    Science.gov (United States)

    Xu, Chengchen; Wang, Bi-Cheng; Yu, Ziniu; Sun, Ming

    2014-01-01

    Since the first X-ray structure of Cry3Aa was revealed in 1991, numerous structures of B. thuringiensis toxins have been determined and published. In recent years, functional studies on the mode of action and resistance mechanism have been proposed, which notably promoted the developments of biological insecticides and insect-resistant transgenic crops. With the exploration of known pore-forming toxins (PFTs) structures, similarities between PFTs and B. thuringiensis toxins have provided great insights into receptor binding interactions and conformational changes from water-soluble to membrane pore-forming state of B. thuringiensis toxins. This review mainly focuses on the latest discoveries of the toxin working mechanism, with the emphasis on structural related progress. Based on the structural features, B. thuringiensis Cry, Cyt and parasporin toxins could be divided into three categories: three-domain type α-PFTs, Cyt toxin type β-PFTs and aerolysin type β-PFTs. Structures from each group are elucidated and discussed in relation to the latest data, respectively. PMID:25229189

  3. Structural Insights into Bacillus thuringiensis Cry, Cyt and Parasporin Toxins

    Directory of Open Access Journals (Sweden)

    Chengchen Xu

    2014-09-01

    Full Text Available Since the first X-ray structure of Cry3Aa was revealed in 1991, numerous structures of B. thuringiensis toxins have been determined and published. In recent years, functional studies on the mode of action and resistance mechanism have been proposed, which notably promoted the developments of biological insecticides and insect-resistant transgenic crops. With the exploration of known pore-forming toxins (PFTs structures, similarities between PFTs and B. thuringiensis toxins have provided great insights into receptor binding interactions and conformational changes from water-soluble to membrane pore-forming state of B. thuringiensis toxins. This review mainly focuses on the latest discoveries of the toxin working mechanism, with the emphasis on structural related progress. Based on the structural features, B. thuringiensis Cry, Cyt and parasporin toxins could be divided into three categories: three-domain type α-PFTs, Cyt toxin type β-PFTs and aerolysin type β-PFTs. Structures from each group are elucidated and discussed in relation to the latest data, respectively.

  4. Toxicity studies for indigenous Bacillus thuringiensis isolates from Malang City, East Java on Aedes aegypti larvae.

    Science.gov (United States)

    Gama, Zulfaidah Penata; Nakagoshi, Nobukazu; Suharjono; Setyowati, Faridah

    2013-02-01

    To investigate the toxicity of indigenous Bacillus thuringiensis (B. thuringiensis)isolates from Malang City for controlling Aedes aegypti (Ae. aegypti) larvae. Soil samples were taken from Purwantoro and Sawojajar sub-districts. Bacterial isolation was performed using B. thuringiensis selective media. Phenotypic characteristics of the isolates were obtained with the simple matching method. The growth and prevalence of spores were determined by the Total Plate Count method, and toxicity tests were also performed on the third instar larval stage of Ae. aegypti. The percentage of larval mortality was analysed using probit regression. The LC50 was analysed by ANOVA, and the Tukey HSD interval was 95%. Among the 33 selected bacterial isolates, six were obtained (PWR4-31, PWR4-32, SWJ4-2b, SWJ4-4b, SWJ-4k and SWJ5-1) that had a similar phenotype to reference B. thuringiensis. Based on the dendrogram, all of the bacterial isolates were 71% similar. Three isolates that had a higher prevalence of reference B. thuringiensis were PWR4-32, SWJ4-4b and SW5-1, of which the spore prevalence was 52.44%, 23.59%, 34.46%, respectively. These three indigenous isolates from Malang City successfully killed Ae. aegypti larvae. The PWR4-32 isolates were the most effective at killing the larvae. Six indigenous B. thuringiensis isolates among the 33 bacterial isolates found in the Sawojajar and Purwantoro sub-districts were toxic to the third instar larvae of Ae. aegypti. The PWR4-32 isolates were identical to the reference B. thuringiensis and had 88% phenotype similarity. The PWR4-32 isolates had the highest spore prevalence (52.44%), and the early stationary phase occurred at 36 h. The PWR4-32 isolates were the most effective at killing Ae. aegypti larvae (LC50-72 h=2.3×10(8) cells/mL).

  5. Complete genome sequence and bioinformatics analyses of Bacillus thuringiensis strain BM-BT15426.

    Science.gov (United States)

    Liu, Junyan; Li, Lin; Peters, Brian M; Li, Bing; Chen, Dingqiang; Xu, Zhenbo; Shirtliff, Mark E

    2017-07-01

    This study aimed to investigate the genetic characteristics of Bacillus thuringiensis strain BM-BT15426. B. thuringiensis strain was identified by sequencing the PCR product (amplifying 16S rRNA gene) using ABI Prism 377 DNA Sequencer. The genome was sequenced using PacBio RS II sequencers and assembled de novo using HGAP. Also, further genome annotation was performed. The genome of B. thuringiensis strain BM-BT15426 has a length of 5,246,329 bp and contains 5409 predicted genes with an average G + C content of 35.40%. Three genes were involved in the "Infectious diseases: Amoebiasis" pathway. A total of 21 virulence factors and 9 antibiotic resistant genes were identified. The major pathogenic factors of B. thuringiensis strain BM-BT15426 were identified through complete genome sequencing and bioinformatics analyses which contributes to further study on pathogenic mechanism and phenotype of B. thuringiensis. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Translocation of Bacillus thuringiensis in Phaseolus vulgaris tissues and vertical transmission in Arabidopsis thaliana.

    Science.gov (United States)

    García-Suárez, R; Verduzco-Rosas, L A; Del Rincón-Castro, M C; Délano-Frier, J P; Ibarra, J E

    2017-04-01

    To demonstrate the ability of Bacillus thuringiensis to penetrate as spore-crystal complex to the internal tissues of bean plants, and keep its insecticidal activity. To test the vertical transmission of the spore-crystal complex in Arabidopsis thaliana. The experimental strain was transformed with the pMUTIN-gfp plasmid which labelled the spores of B. thuringiensis HD-73 with the GFP protein. Once the rhizosphere of the bean plants was inoculated with the labelled strain, the bacterium was recovered from leaves, stems, and petioles. Furthermore, toxicity of treated plants was significantly higher than control plants when bio-assayed on cabbage looper larvae. The labelled strain was recovered from the dead insects. When the rhizosphere of A. thaliana plants was inoculated with the labelled strain, mature seeds from these plants were surface-sterilized and grown under in vitro conditions. The labelled strain was recovered from the seedlings. We showed that B. thuringiensis subsp. kurstaki (HD-73) in the rhizosphere can translocate to upper tissues of bean plants, and keep its insecticidal activity. Transmission of the labelled B. thuringiensis strain passed to the next generation of A. thaliana. The role of B. thuringiensis as a potential facultative endophyte bacterium and the possible biotechnological repercussions are discussed. © 2017 The Society for Applied Microbiology.

  7. SR450 And Superhawk XP Applications Of Bacillus thuringiensis israelensis Against Culex quinquefasciatus.

    Science.gov (United States)

    Dunford, James C; Stoops, Craig A; Estep, Alden S; Britch, Seth C; Richardson, Alec G; Walker, Todd W; Farooq, Muhammad; Hoel, David F; Platt, Raymond R; Smith, Vincent L; Wirtz, Robert A; Kerce, Jerry D

    2014-09-01

    Sprayer comparisons and larval morality assays were conducted following SR450 backpack mist blower and Superhawk XP thermal fogger applications of Vectobac® WDG Bacillus thuringiensis israelensis (Bti) against Culex quinquefasciatus. Bacillus thuringiensis israelensis was applied at maximum label rate in a 232.26-m(2) field plot located in north-central Florida with containers placed at 2 heights (ground level and 1.52 m above ground) on stakes positioned 3.04, 6.09, 9.14, 12.19, and 15.24 m from the spray line. Results indicated that there was no significant (P > 0.05) difference in 24- and 48-h larval mortality between the 2 sprayers or between the 2 heights. There was significant difference (P 70% larval mortality 3.04-9.14 m from the spray line, and control mosquito larvae.

  8. Laser He-Ne effect on bacillus thuringiensis var. kurstaki strain LBT-24

    International Nuclear Information System (INIS)

    Dias, Adriana; Barreara, Lenay; Otero, Isabel; Rabelo, Yanet; Rodriguez, Dania

    1999-01-01

    Bacillus thuringiensis toxin is one of the world widely used entomopathogen. It presents an strong insecticide activity on Lepidoptera, Coleoptera and Diptera. It was studied the effect of Laser He-Ne on Bacillus thuringiensis var. kurstaki strain LBT-24. Growing curves were made and were calculated the duplication time and the specific growing speed of each one. The curves were statistically compared. It was also analysed the phage induction with and without Laser red light influence. Also, it was observed the presence of the d-endotoxin crystal with this treatment. The red Laser He-Ne enhanced the growth of this micro-organism under laboratory conditions and didn't have any effect over the other characteristics analysed

  9. Enhancement of virulence of bacillus thuringiensis and serratia marcescens by chemicals

    International Nuclear Information System (INIS)

    Khan, K. A.

    2006-01-01

    Studies were conducted on the enhancement of pathogenicity of Bacillus thuringiensis by 1% boric acid against various species of termites. The increase in virulence of Serratia marcescens by 1% potassium chloride or 1% Sodium citrate against the workers of M. championi has also been established. The increase in virulence is confirmed by the enhancement ratio, which are ranging from about 1.5 to 1.8 for Bacillus thuringiensis and 1.3 to 1.6 for Serratia marcescens. It was also noted that 1% boric acid alone was found toxic to various species of termites. However, Potassium chloride and Sodium citrate in a concentration of 1% were non-toxic to the workers of M. championi. (author)

  10. Efektifitas Bacillus Thuringiensis Yang Diisolasi Dari Sampel Tanah Di YOGYAKARTA Terhadap Larva Culex Quinquefasciatus Invitro

    OpenAIRE

    Suryani, Lilis

    2008-01-01

    Filariasis disease is a public health problem, especially in rural areas. One kinds of mosquitoes that transmited it is Culex quinquefasciatus. Since 1972 chemical insecticides has been usedfor vector control in Indonesia. However, in 1987 it was reported that there are many areas has been resistant against chemical insecticides. Bacillus thuringiensis is a gram positif bacteria, rod, aerobic and spore shape. There are many strain of this bacteria produces a toxic protein to insect. This rese...

  11. Genetical and radiobiological characteristics of phage Tg13 of Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Takubova, R.M.; Azizbekyan, R.R.

    1979-01-01

    The radiation-genetical aspects of interrelations between phages and cells of the spore-forming bacteria Bacillus thurin-giensis were studied. The phage Tg13 liberates C-mutants, forming transparent negative colonies, both spontaneously and under the effect of UV irradiation. UV-radiation increases reliably the level of C-mutants in the population. The phenotype of the observed mutants is, evidently, caused by the specific features of interaction in the system: preudolysogenic culture -phage Tg13

  12. The potential of the novel mosquitocidal Bacillus thuringiensis strain LLP29 for use in practice

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Wu, S.; Peng, Y.; Li, M.; Sun, L.; Huang, E.; Guan, X.; Gelbič, Ivan

    2011-01-01

    Roč. 36, č. 2 (2011), s. 458-460 ISSN 1081-1710 R&D Projects: GA MŠk 2B08003 Grant - others:National Natural Science Foundation of China (CN) 31071745; Science Foundation of the Ministry of Education of China (CN) 20093515110010; Science Foundation of the Ministry of Education of China (CN) 20093515120010 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis Subject RIV: ED - Physiology Impact factor: 0.885, year: 2011

  13. Distinct clpP Genes Control Specific Adaptive Responses in Bacillus thuringiensis

    OpenAIRE

    Fedhila, Sinda; Msadek, Tarek; Nel, Patricia; Lereclus, Didier

    2002-01-01

    ClpP and ClpC are subunits of the Clp ATP-dependent protease, which is ubiquitous among prokaryotic and eukaryotic organisms. The role of these proteins in stress tolerance, stationary-phase adaptive responses, and virulence in many bacterial species has been demonstrated. Based on the amino acid sequences of the Bacillus subtilis clpC and clpP genes, we identified one clpC gene and two clpP genes (designated clpP1 and clpP2) in Bacillus thuringiensis. Predicted proteins ClpP1 and ClpP2 have ...

  14. Synergistic effect of certain insecticides combined with Bacillus thuringiensis on mosquito larvae

    Directory of Open Access Journals (Sweden)

    C.P. Narkhede

    2017-04-01

    Full Text Available For effective vector control it is essential to formulate new preparations having multiple action against the vector pest. Developing combined formulation of biopesticide and chemical pesticide is one of the novel concept to fight against the vectors with new weapons; however, compatibility of biopesticide i.e. Bacillus thuringiensis (Bt and chemical pesticide is a real hurdle. In this investigation, local isolate Bacillus thuringiensis SV2 (BtSV2 was tested for its compatibility with various available mosquito larvicides. Temephos was most compatible with BtSV2 than with other tested pesticides. These two compatible agents were tested for larvicidal potential. Our study revealed that the synergistic effect of both agents reduces LC50 value by 30.68 and 22.36% against the Ae. aegypti and An. stephensi, respectively. The larvicidal potential increased when compared to individual pesticides. It was also observed a biochemical change in larvae after the TBT (Temephos + Bacillus thuringiensis combination treatment; it involves decreased level of alpha esterase, acetylcholine esterase and protein while level of beta esterase and acid phosphatase was unchanged and alkaline phosphatase activity was increased. Increased potential of combined formulation may be due to altered physiological condition.

  15. Experimental design and Bayesian networks for enhancement of delta-endotoxin production by Bacillus thuringiensis.

    Science.gov (United States)

    Ennouri, Karim; Ayed, Rayda Ben; Hassen, Hanen Ben; Mazzarello, Maura; Ottaviani, Ennio

    2015-12-01

    Bacillus thuringiensis (Bt) is a Gram-positive bacterium. The entomopathogenic activity of Bt is related to the existence of the crystal consisting of protoxins, also called delta-endotoxins. In order to optimize and explain the production of delta-endotoxins of Bacillus thuringiensis kurstaki, we studied seven medium components: soybean meal, starch, KH₂PO₄, K₂HPO₄, FeSO₄, MnSO₄, and MgSO₄and their relationships with the concentration of delta-endotoxins using an experimental design (Plackett-Burman design) and Bayesian networks modelling. The effects of the ingredients of the culture medium on delta-endotoxins production were estimated. The developed model showed that different medium components are important for the Bacillus thuringiensis fermentation. The most important factors influenced the production of delta-endotoxins are FeSO₄, K2HPO₄, starch and soybean meal. Indeed, it was found that soybean meal, K₂HPO₄, KH₂PO₄and starch also showed positive effect on the delta-endotoxins production. However, FeSO4 and MnSO4 expressed opposite effect. The developed model, based on Bayesian techniques, can automatically learn emerging models in data to serve in the prediction of delta-endotoxins concentrations. The constructed model in the present study implies that experimental design (Plackett-Burman design) joined with Bayesian networks method could be used for identification of effect variables on delta-endotoxins variation.

  16. Novel Cell Wall Hydrolase CwlC from Bacillus thuringiensis Is Essential for Mother Cell Lysis.

    Science.gov (United States)

    Chen, Xiaomin; Gao, Tantan; Peng, Qi; Zhang, Jie; Chai, Yunrong; Song, Fuping

    2018-04-01

    In this study, a sporulation-specific gene (tentatively named cwlC ) involved in mother cell lysis in Bacillus thuringiensis was characterized. The encoded CwlC protein consists of an N-terminal N -acetylmuramoyl-l-alanine amidase (Mur N Ac-LAA) domain and a C-terminal amidase02 domain. The recombinant histidine-tagged CwlC proteins purified from Escherichia coli were able to directly bind to and digest the B. thuringiensis cell wall. The CwlC point mutations at the two conserved glutamic acid residues (Glu-24 and Glu-140) shown to be critical for the catalytic activity in homologous amidases resulted in a complete loss of cell wall lytic activity, suggesting that CwlC is an N -acetylmuramoyl-l-alanine amidase. Results of transcriptional analyses indicated that cwlC is transcribed as a monocistronic unit and that its expression is dependent on sporulation sigma factor K (σ K ). Deletion of cwlC completely blocked mother cell lysis during sporulation without impacting the sporulation frequency, Cry1Ac protein production, and insecticidal activity. Taken together, our data suggest that CwlC is an essential cell wall hydrolase for B. thuringiensis mother cell lysis during sporulation. Engineered B. thuringiensis strains targeting cwlC , which allows the crystal inclusion to remain encapsulated in the mother cell at the end of sporulation, may have the potential to become more effective biological control agents in agricultural applications since the crystal inclusion remains encapsulated in the mother cell at the end of sporulation. IMPORTANCE Mother cell lysis has been well studied in Bacillus subtilis , which involves three distinct yet functionally complementary cell wall hydrolases. In this study, a novel cell wall hydrolase, CwlC, was investigated and found to be essential for mother cell lysis in Bacillus thuringiensis CwlC of B. thuringiensis only shows 9 and 21% sequence identity with known B. subtilis mother cell hydrolases CwlB and CwlC, respectively

  17. A novel Bacillus thuringiensis strain LLB6, isolated from bryophytes, and its new cry2Ac-type gene.

    Science.gov (United States)

    Zhang, L L; Lin, J; Luo, L; Guan, C Y; Zhang, Q L; Guan, Y; Zhang, Y; Ji, J T; Huang, Z P; Guan, X

    2007-03-01

    To isolate and characterize the novel Bacillus thuringiensis strains from bryophytes collected from Wuyi Mountain, Fujian Province of China, and identify new B. thuringiensis strains and toxins active against mosquitoes. Twelve novel B. thuringiensis strains were isolated from 76 bryophyte samples. According to the results of this preliminary screening, LLB6 was the most toxic to Aedes albopictus. Then phase-contrast as well as scanning electron microscopy, bioassays, cloning, sequencing and expression were performed to characterize the novel isolate LLB6 and its new gene cry2Ac5. Bacillus thuringiensis occurred naturally on bryophytes. LLB6 isolated from Physcomitrium japonicum was toxic to A. albopictus. A new cry2Ac5 gene of LLB6 was detected, cloned and expressed successfully. Bioassays on A. albopictus showed that the expressed Cry2Ac5 was also toxic to the third instar larvae. This is the first report of B. thuringiensis strains isolated from bryophytes. It represents a specific source of new B. thuringiensis strains and is of great importance for the knowledge of the ecology of B. thuringiensis. Novel LLB6 harboring the new gene cry2Ac5 and its expressed Cry2Ac5 protein revealed activity against A. albopictus and became a new member of B. thuringiensis toxins.

  18. Spore prevalence and toxigenicity of Bacillus cereus and Bacillus thuringiensis isolates from U.S. retail spices.

    Science.gov (United States)

    Hariram, Upasana; Labbé, Ronald

    2015-03-01

    Recent incidents of foodborne illness associated with spices as the vehicle of transmission prompted this examination of U.S. retail spices with regard to Bacillus cereus. This study focused on the levels of aerobic-mesophilic spore-forming bacteria and B cereus spores associated with 247 retail spices purchased from five states in the United States. Samples contained a wide range of aerobic-mesophilic bacterial spore counts ( 10(7) CFU/g). Using a novel chromogenic agar, B. cereus and B. thuringiensis spores were isolated from 77 (31%) and 11 (4%) samples, respectively. Levels of B. cereus were thuringiensis isolates possessed at least one type of enterotoxin gene: HBL (hemolysin BL) or nonhemolytic enterotoxin (NHE). None of the 88 isolates obtained in this study possessed the emetic toxin gene (ces). Using commercially available immunological toxin detection kits, the toxigenicity of the isolates was confirmed. The NHE enterotoxin was expressed in 98% of B. cereus and 91% of B. thuringiensis isolates that possessed the responsible gene. HBL enterotoxin was detected in 87% of B. cereus and 100% of B. thuringiensis PCR-positive isolates. Fifty-two percent of B. cereus and 54% of B. thuringiensis isolates produced both enterotoxins. Ninety-seven percent of B. cereus isolates grew at 12°C, although only two isolates grew well at 9°C. The ability of these spice isolates to form spores, produce diarrheal toxins, and grow at moderately abusive temperatures makes retail spices an important potential vehicle for foodborne illness caused by B. cereus strains, in particular those that produce diarrheal toxins.

  19. Dark fermentative bioconversion of glycerol to hydrogen by Bacillus thuringiensis.

    Science.gov (United States)

    Kumar, Prasun; Sharma, Rishi; Ray, Subhasree; Mehariya, Sanjeet; Patel, Sanjay K S; Lee, Jung-Kul; Kalia, Vipin C

    2015-04-01

    Biodiesel manufacturing units discharge effluents rich in glycerol. The need is to convert crude glycerol (CG) into useful products such as hydrogen (H2). Under batch culture, Bacillusthuringiensis EGU45 adapted on pure glycerol (PG, 2% v/v) resulted in an H2 yield of 0.646 mol/mol glycerol consumed on minimal media (250 mL) supplemented with 1% ammonium nitrate at 37°C over 4 days. Here, H2 constituted 67% of the total biogas. Under continuous culture, at 2 days of hydraulic retention time, B. thuringiensis immobilized on ligno-cellulosic materials (banana leaves - BL, 10% v/v) resulted in a H2 yield of 0.386 mol/mol PG consumed. On CG, the maximal H2 yield of 0.393 mol/mol feed consumed was recorded. In brief, B. thuringiensis could transform CG, on limited resources - minimal medium with sodium nitrate, by immobilizing them on cheap and easily available biowaste, which makes it a suitable candidate for H2 production on a large scale. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Translocation and insecticidal activity of Bacillus thuringiensis living inside of plants.

    Science.gov (United States)

    Monnerat, Rose Gomes; Soares, Carlos Marcelo; Capdeville, Guy; Jones, Gareth; Martins, Erica Soares; Praça, Lilian; Cordeiro, Bruno Arrivabene; Braz, Shélida Vasconcelos; dos Santos, Roseane Cavalcante; Berry, Colin

    2009-07-01

    The major biological pesticide for the control of insect infestations of crops, Bacillus thuringiensis was found to be present naturally within cotton plants from fields that had never been treated with commercial formulations of this bacterium. The ability of B. thuringiensis to colonize plants as an endophyte was further established by the introduction of a strain marked by production of green fluorescent protein (GFP). After inoculation of this preparation close to the roots of cotton and cabbage seedlings, GFP-marked bacteria could be re-isolated from all parts of the plant, having entered the roots and migrated through the xylem. Leaves taken from the treated plants were able to cause toxicity when fed to the Lepidoptera Spodoptera frugiperda (cotton) and Plutella xylostella (cabbage). These results open up new horizons for understanding the natural ecology and evolution of B. thuringiensis and use of B. thuringiensis in insect control. Journal compilation © 2009 Society for Applied Microbiology and Blackwell Publishing Ltd. No claim to original Brazilian government works.

  1. Effect of Bacillus thuringiensis parasporal toxin on stimulating of IL-2 and IL-5 cytokines production

    Directory of Open Access Journals (Sweden)

    Marzieh Soleimany

    2018-03-01

    Full Text Available Introduction:Bacillus thuringiensis, is a Gram-positive spore-forming bacterium that produces crystalline parasporal protein (Cry during sporulation. Some of these Cry toxins do not show cytotoxicity against insects but they are capable to kill some human and animal cancer cells. The aim of this study was to verify whether cytocidal parasporal of B thuringiensis strains have immunostimulatory activity on human peripheral blood mononuclear cells (PBMNC and to evaluate the ability of IL-2 and IL-5 production. Materials and methods: B. thuringiensis toxin with cytocidal activity was isolated and treated with proteinase K. PBMNC was cultured and treated with activated crystal proteins. We evaluated the ability of different cytokines production with Flow Cytometry. Results: In this study, immune stimulatory toxins Cry1 were distinguished. This toxin can stimulate production of cytokines IL-2 and stop production of IL-5. Discussion and conclusion: According to anti-cancer effect of B. thuringiensis toxins and also immune stimulatory effect, with more research these toxins can be introduced as immunotherapy drug in cancer treatment.

  2. Assessment of microbial larvicide spraying with Bacillus thuringiensis israelensis, for the prevention of malaria.

    Science.gov (United States)

    Kinde-Gazard, D; Baglo, T

    2012-03-01

    The aim of this study was to assess the contribution of microbial larvicide spraying, Bacillus thuringiensis israelensis, as prevention strategy against malaria. An experimental study consisted in spraying B. thuringiensis israelensis in a district during 1 year has been conducted. Another district (control) was not sprayed. Eight hundred and two children were evaluated, thick drop and swab examination was performed for those presenting with fever. The larval density was calculated in their habitats as well as larvicide remanence. Capture of mosquitoes with human bait allowed determining human exposure to bites at night, and identifying anopheles after dissection. The incidence of pediatric malaria was 13.8% in the sprayed district and 31.4% in the control district. The parasitic load ranged from 2000 to 42,000 parasites/μL in the sprayed district and 2000 to 576,000 parasites/μL in the control district. Plasmodium falciparum was the most frequent (97.8%) plasmodial species. In the control district, at least 20 larvae by liter of water were counted; anopheles larvae were found in 11 larval habitats out of 15 (73.33%). The human exposure to anopheles bites at night was 14.25 in the sprayed district and 33.13 in the control district. The remanence of B. thuringiensis israelensis was estimated at 9 days in the sprayed district. The larvicide B. thuringiensis israelensis may be used in vector control strategy for the prevention of malaria. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  3. Effect of inherited sterility and bacillus thuringiensis on mortality and reproduction of phthorimaea opercullela zeller (lepidoptera: gelechidae)

    International Nuclear Information System (INIS)

    Makee, H.; Tlas, M. D.; Amer, S.; Abdulla, J.

    2008-01-01

    The effect of a commercial formulation of Bacillus thuringiensis (Dipel 2X) upon F 1 progeny of irradiated and unirradiated phthorimaea operculella male parents was investigated. F 1 progeny of irradiated parents was more susceptible to B. thuringiensis than that of unirradiated parents. A combination of irradiation and B. thuringiensis led to higher mortality in F 1 progeny of P. operculella. The LC 50 was 0.406 g/100ml for F 1 progeny of unirradiated parents, but 0.199 g/100ml for those of irradiated parents. There was a great reduction in the pupal weight, fecundity and egg hatchability of F 1 progeny of irradiated patents compared to those unirradiated parents. Such reduction was increased by applying higher concentration of B. thuringiensis. A combination between inherited sterility technique and B. thuringiensis application could give a good controlling result against P. operculella. (author)

  4. Screening and identification of a Bacillus thuringiensis strain S1/4 with large and efficient insecticidal activities.

    OpenAIRE

    Jaoua, Samir; Sellami, Sameh; Zghal, Taheni; Cherif, Maroua; Zalila-Kolsi, Imen; Jaoua, Samir; Jamoussi, Kaïs

    2013-01-01

    The bacterium Bacillus thuringiensis was recognized for its entomopathogenic activities related to Cry and Cyt proteins forming the δ-endotoxins and some extracellular activities like the vegetative insecticidal proteins (VIP) and Cry1I. These activities may act specifically against diverse organisms and some of them typically characterize each strain. Here, we screened a set of 212 B. thuringiensis strains to search the higher insecticidal activities. These strains had bipyramidal and cubic ...

  5. Evaluation of Bacillus thuringiensis Pathogenicity for a Strain of the Tick, Rhipicephalus microplus, Resistant to Chemical Pesticides

    OpenAIRE

    Fern?ndez-Ruvalcaba, Manuel; Pe?a-Chora, Guadalupe; Romo-Mart?nez, Armando; Hern?ndez-Vel?zquez, V?ctor; de Parra, Alejandra Bravo; De La Rosa, Diego P?rez

    2010-01-01

    The pathogenicity of four native strains of Bacillus thuringiensis against Rhipicephalus (Boophilus) microplus (Canestrine) (Acari: Ixodidae) was evaluated. A R. microplus strain that is resistant to organophosphates, pyrethroids, and amidines, was used in this study. Adult R. microplus females were bioassayed using the immersion test of Drummond against 60 B. thuringiensis strains. Four strains, GP123, GP138, GP130, and GP140, were found to be toxic. For the immersion test, the total protein...

  6. Expression of chitinase-encoding genes in Bacillus thuringiensis and toxicity of engineered B. thuringiensis subsp. aizawai toward Lymantria dispar larvae.

    Science.gov (United States)

    Lertcanawanichakul, Monthon; Wiwat, Chanpen; Bhumiratana, Amaret; Dean, Donald Harry

    2004-03-01

    Chitinase genes from Aeromonas hydrophila and Bacillus circulans No.4.1 were cloned into the plasmid pHY300PLK and designated as pHYA2 and pHYB43, respectively. Both plasmids were introduced into various strains of B. thuringiensis by electroporation. Plasmid pHYB43 was generally structurally stable, but showed lower segregrational stability than pHYA2 in B. thuringiensis subsp. aizawai when grown under nonselective conditions. The production of chitinase from B. thuringiensis subsp. aizawai harboring pHYB43 or pHYA2 could be detected after native polyacrylamide gel electrophoresis by using 4-methylumbelliferyl beta-D- N,N'- diacetylchitobioside as the substrate. Moreover, B. thuringiensis subsp. aizawai harboring pHYB43 gave 15 times higher chitinase activity than when harboring pHYA2, as determined by means of a colorimetric method using glycol chitin as the substrate. In addition, B. thuringiensis subsp. aizawai harboring pHYB43 was more toxic to gypsy moth larvae ( Lymantria dispar) than parental B. thuringiensis subsp. aizawai or its clone harboring pHYA2.

  7. Expression of Bacillus thuringiensis serovar. israelensis toxins in Asticcacaulis excentricus to control dipteran larvae of vectors of diseases

    Directory of Open Access Journals (Sweden)

    Óscar Enrique Guevara

    2004-01-01

    Full Text Available Bacillus thuringiensis cry genes encode for a diverse group of crystal-forming proteins that exhibit insecticidal activity towards dipteran, lepidopteran and coleopteran larvae. The effectiveness of insecticides based on mosquito larvicidal B. thuringiensis strains can be enhanced by using aquatic prosthecated bacteria as alternative hosts, since they do not sink, cytoplasmic located toxins are protected f rom UV radiation and, most importantly, mosquito larvae feed on them. An Asticcacaulis excentricus reference strain was transformed with the cry1 1Aa gene from Bacillus thuringiensis serovar. israelensis. Western blot and electrophoresis were used to test recombinant protein expression; Western blot revealed a 72 kDa protein corresponding to B. thuringiensis serovar. israelensis Cry1 1 Aa. These aquatic bacte­rias toxicity achieved 50% mortality at 23 ng/mL concentration in f irst instar Culex quinquefasciatus larvae. Other bioassays indicated that recombinant A. excentricus is toxic against Aedes aegyptiand Anopheles albimanus first instar larvae. Buoyancy tests demonstrated the advantage of A. excentricus over B. thuringiensis. Key words: Asticcacaulis excentricus, Bacillus thuringiensis, prosthecated bacteria, dengue, malaria.

  8. Are nematodes a missing link in the confounded ecology of the entomopathogen Bacillus thuringiensis?

    Science.gov (United States)

    Ruan, Lifang; Crickmore, Neil; Peng, Donghai; Sun, Ming

    2015-06-01

    Bacillus thuringiensis, which is well known as an entomopathogen, has been accepted by the public as a safe bioinsecticide. The natural ecology of this bacterium has never been particularly clear, with views ranging from it being an obligate pathogen to an opportunist pathogen that can otherwise exist as a soil saprophyte or a plant endophyte. This confusion has recently led to it being considered as an environmental pathogen that has evolved to occupy a diverse set of environmental niches in which it can thrive without needing a host. A significant driving force behind this classification is the fact that B. thuringiensis is found in high numbers in environments that are not occupied by the insect hosts to which it is pathogenic. It is our opinion that the ubiquitous presence of this bacterium in the environment is the result of a variety of vectoring systems, particularly those that include nematodes. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Larvicidal activity of Bacillus thuringiensis var. israelensis Cry11Aa toxin against Haemonchus contortus.

    Science.gov (United States)

    DE Lara, Ana Paula DE Souza Stori; Lorenzon, Lucas Bigolin; Vianna, Ana Muñoz; Santos, Francisco Denis Souza; Pinto, Luciano Silva; Aires Berne, Maria Elisabeth; Leite, Fábio Pereira Leivas

    2016-10-01

    Effective control of gastrointestinal parasites is necessary in sheep production. The development of anthelmintics resistance is causing the available chemically based anthelmintics to become less effective. Biological control strategies present an alternative to this problem. In the current study, we tested the larvicidal effects of Bacillus thuringiensis var. israelensis Cry11Aa toxin against Haemonchus contortus larvae. Bacterial suspensions [2 × 108 colony-forming units (CFU) g-1 of the feces] of B. thuringiensis var. israelensis and recombinant Escherichia coli expressing Cry11Aa toxin were added to naturally H. contortus egg-contaminated feces. The larvae were quantified, and significant reductions of 62 and 81% (P var. israelensis and recombinant E. coli expressing Cry11Aa toxin were then orally administered to lambs naturally infected with H. contortus. Twelve hours after administration, feces were collected and submitted to coprocultures. Significant larvae reductions (P var. israelensis is a promising new class of biological anthelmintics for treating sheep against H. contortus.

  10. Diagnostic properties of three conventional selective plating media for selection of Bacillus cereus, B. thuringiensis and B. weihenstephanensis

    DEFF Research Database (Denmark)

    Hendriksen, Niels Bohse; Hansen, Bjarne Munk

    2011-01-01

    The aim of this study was to assess the diagnostic properties of the two selective plating media and a chromogenic medium for identification of Bacillus cereus. The 324 isolates were B. cereus (37%), Bacillus weihenstephanensis (45%) or Bacillus thuringiensis (18%), as identified by a new...... combination of techniques. All isolates were growing on mannitol–egg yolk–polymyxin agar (MYP), and they did not form acid from mannitol. However, a significant lower number of B. thuringiensis isolates did not show lecithinase activity. All isolates were also growing on polymyxin–egg yolk...... recommended selective plating media MYP and PEMBA for detection of B. cereus group bacteria both have their limitations for identification of some B. cereus, B. weihenstephanensis or B. thuringiensis. However, MYP is preferable compared to PEMBA. The chromogenic medium has its own advantages and limitations...

  11. Antagonistic activity of selected strains of Bacillus thuringiensis ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-05-02

    May 2, 2008 ... funguicidas. Memorias Primera Convención Mundial del Chile. León,. Guanajuato, México. Resumen, pp. 144-150. Podile AR, Laxmi VDV (1998). Seed bacterization with Bacillus subtilis. AF1 increases phenylalanine ammnonia lyase and reduces the incidence of fusarial wilt in pigeonpea. J. Phytophatol.

  12. Transcriptional profile of tomato roots exhibiting Bacillus thuringiensis-induced resistance to Ralstonia solanacearum.

    Science.gov (United States)

    Takahashi, Hideki; Nakaho, Kazuhiro; Ishihara, Takeaki; Ando, Sugihiro; Wada, Takumi; Kanayama, Yoshinori; Asano, Shinichiro; Yoshida, Shigenobu; Tsushima, Seiya; Hyakumachi, Mitsuro

    2014-01-01

    Activation of SA-dependent signaling pathway and suppression of JA-dependent signaling pathway seem to play key roles inB. thuringiensis-induced resistance toR. solanacearumin tomato plants. Bacillus thuringiensis, a well-known and effective bio-insecticide, has attracted considerable attention as a potential biological control agent for the suppression of plant diseases. Treatment of tomato roots with a filter-sterilized cell-free filtrate (CF) of B. thuringiensis systemically suppresses bacterial wilt caused by Ralstonia solanacearum through systemic activation of the plant defense system. Comparative analysis of the expression of the Pathogenesis-Related 1(P6) gene, a marker for induced resistance to pathogens, in various tissues of tomato plants treated with CF on their roots suggested that the B. thuringiensis-induced defense system was activated in the leaf, stem, and main root tissues, but not in the lateral root tissue. At the same time, the growth of R. solanacearum was significantly suppressed in the CF-treated main roots but not in the CF-treated lateral roots. This distinct activation of the defense reaction and suppression of R. solanacearum were reflected by the differences in the transcriptional profiles of the main and lateral tissues in response to the CF. In CF-treated main roots, but not CF-treated lateral roots, the expression of several salicylic acid (SA)-responsive defense-related genes was specifically induced, whereas jasmonic acid (JA)-related gene expression was either down-regulated or not induced in response to the CF. On the other hand, genes encoding ethylene (ET)-related proteins were induced equally in both the main and lateral root tissues. Taken together, the co-activation of SA-dependent signaling pathway with ET-dependent signaling pathway and suppression of JA-dependent signaling pathway may play key roles in B. thuringiensis-induced resistance to R. solanacearum in tomato.

  13. Identification of Distinct Bacillus thuringiensis 4A4 Nematicidal Factors Using the Model Nematodes Pristionchus pacificus and Caenorhabditis elegans

    Science.gov (United States)

    Iatsenko, Igor; Nikolov, Angel; Sommer, Ralf J.

    2014-01-01

    Bacillus thuringiensis has been extensively used for the biological control of insect pests. Nematicidal B. thuringiensis strains have also been identified; however, virulence factors of such strains are poorly investigated. Here, we describe virulence factors of the nematicidal B. thuringiensis 4A4 strain, using the model nematodes Pristionchus pacificus and Caenorhabditis elegans. We show that B. thuringiensis 4A4 kills both nematodes via intestinal damage. Whole genome sequencing of B. thuringiensis 4A4 identified Cry21Ha, Cry1Ba, Vip1/Vip2 and β-exotoxin as potential nematicidal factors. Only Cry21Ha showed toxicity to C. elegans, while neither Cry nor Vip toxins were active against P. pacificus, when expressed in E. coli. Purified crystals also failed to intoxicate P. pacificus, while autoclaved spore-crystal mixture of B. thuringiensis 4A4 retained toxicity, suggesting that primary β-exotoxin is responsible for P. pacificus killing. In support of this, we found that a β-exotoxin-deficient variant of B. thuringiensis 4A4, generated by plasmid curing lost virulence to the nematodes. Thus, using two model nematodes we revealed virulence factors of the nematicidal strain B. thuringiensis 4A4 and showed the multifactorial nature of its virulence. PMID:25025708

  14. Functional analysis of the sporulation-specific diadenylate cyclase CdaS in Bacillus thuringiensis

    Science.gov (United States)

    Zheng, Cao; Ma, Yang; Wang, Xun; Xie, Yuqun; Ali, Maria K.; He, Jin

    2015-01-01

    Cyclic di-AMP (c-di-AMP) is a recently discovered bacterial secondary messenger molecule, which is associated with various physiological functions. In the genus Bacillus, the intracellular level and turnover of c-di-AMP are mainly regulated by three diadenylate cyclases (DACs), including DisA, CdaA and CdaS, and two c-di-AMP-specific phosphodiesterases (GdpP and PgpH). In this study, we demonstrated that CdaS protein from B. thuringiensis is a hexameric DAC protein that can convert ATP or ADP to c-di-AMP in vitro and the N-terminal YojJ domain is essential for the DAC activity. Based on the markerless gene knock-out method, we demonstrated that the transcription of cdaS was initiated by the sporulation-specific sigma factor σH and the deletion of cdaS significantly delayed sporulation and parasporal crystal formation. These findings contrast with similar experiments conducted using B. subtilis, wherein transcription of its cdaS was initiated by the sigma factor σG. Deletion of all the three DAC genes from a single strain was unsuccessful, suggesting that c-di-AMP is an indispensable molecule in B. thuringiensis. Phylogenetic analysis indicated increased diversity of CdaS in the B. cereus and B. subtilis Bacillus subgroups. In summary, this study identifies important aspects in the regulation of c-di-AMP in the genus Bacillus. PMID:26441857

  15. Characteristics of a broad lytic spectrum endolysin from phage BtCS33 of Bacillus thuringiensis.

    Science.gov (United States)

    Yuan, Yihui; Peng, Qin; Gao, Meiying

    2012-12-19

    Endolysins produced by bacteriophages lyse bacteria, and are thus considered a novel type of antimicrobial agent. Several endolysins from Bacillus phages or prophages have previously been characterized and used to target Bacillus strains that cause disease in animals and humans. B. thuringiensis phage BtCS33 is a Siphoviridae family phage and its genome has been sequenced and analyzed. In the BtCS33 genome, orf18 was found to encode an endolysin protein (PlyBt33). Bioinformatic analyses showed that endolysin PlyBt33 was composed of two functional domains, the N-terminal catalytic domain and the C-terminal cell wall binding domain. In this study, the entire endolysin PlyBt33, and both the N- and C-termini,were expressed in Escherichia coli and then purified. The lytic activities of PlyBt33 and its N-terminus were tested on bacteria. Both regions exhibited lytic activity, although PlyBt33 showed a higher lytic activity than the N-terminus. PlyBt33 exhibited activity against all Bacillus strains tested from five different species, but was not active against Gram-negative bacteria. Optimal conditions for PlyBt33 reactivity were pH 9.0 and 50 °C. PlyBt33 showed high thermostability, with 40% of initial activity remaining following 1 h of treatment at 60 °C. The C-terminus of PlyBt33 bound to B. thuringiensis strain HD-73 and Bacillus subtilis strain 168. This cell wall binding domain might be novel, as its amino acid sequence showed little similarity to previously reported endolysins. PlyBt33 showed potential as a novel antimicrobial agent at a relatively high temperature and had a broad lytic spectrum within the Bacillus genus. The C-terminus of PlyBt33 might be a novel kind of cell wall binding domain.

  16. Characteristics of a broad lytic spectrum endolysin from phage BtCS33 of Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Yuan Yihui

    2012-12-01

    Full Text Available Abstract Background Endolysins produced by bacteriophages lyse bacteria, and are thus considered a novel type of antimicrobial agent. Several endolysins from Bacillus phages or prophages have previously been characterized and used to target Bacillus strains that cause disease in animals and humans. B. thuringiensis phage BtCS33 is a Siphoviridae family phage and its genome has been sequenced and analyzed. In the BtCS33 genome, orf18 was found to encode an endolysin protein (PlyBt33. Results Bioinformatic analyses showed that endolysin PlyBt33 was composed of two functional domains, the N-terminal catalytic domain and the C-terminal cell wall binding domain. In this study, the entire endolysin PlyBt33, and both the N- and C-termini,were expressed in Escherichia coli and then purified. The lytic activities of PlyBt33 and its N-terminus were tested on bacteria. Both regions exhibited lytic activity, although PlyBt33 showed a higher lytic activity than the N-terminus. PlyBt33 exhibited activity against all Bacillus strains tested from five different species, but was not active against Gram-negative bacteria. Optimal conditions for PlyBt33 reactivity were pH 9.0 and 50°C. PlyBt33 showed high thermostability, with 40% of initial activity remaining following 1 h of treatment at 60°C. The C-terminus of PlyBt33 bound to B. thuringiensis strain HD-73 and Bacillus subtilis strain 168. This cell wall binding domain might be novel, as its amino acid sequence showed little similarity to previously reported endolysins. Conclusions PlyBt33 showed potential as a novel antimicrobial agent at a relatively high temperature and had a broad lytic spectrum within the Bacillus genus. The C-terminus of PlyBt33 might be a novel kind of cell wall binding domain.

  17. A novel mosquitocidal Bacillus thuringiensis strain LLP29 isolated from the phylloplane of Magnolia denudata

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Huang, E.; Lin, J.; Gelbič, Ivan; Zhang, Q.; Guan, Y.; Huang, T.; Guan, X.

    2010-01-01

    Roč. 165, č. 2 (2010), s. 133-141 ISSN 0944-5013 R&D Projects: GA MŠk 2B08003 Grant - others:United Fujian Provincial Health and Education Project for Tackling Key Research(CN) WKJ2008-2-44; Talented Youth Project of Fujian Province(CN) 2008F3012; Educational Department of Fujian Province(CN) JA08080; Fujian Agriculture and Forestry University(CN) 08A01 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis * cyt1 * mosquito Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection Impact factor: 1.958, year: 2010

  18. Comparative Genomics of Bacillus thuringiensis Reveals a Path to Specialized Exploitation of Multiple Invertebrate Hosts.

    Science.gov (United States)

    Zheng, Jinshui; Gao, Qiuling; Liu, Linlin; Liu, Hualin; Wang, Yueying; Peng, Donghai; Ruan, Lifang; Raymond, Ben; Sun, Ming

    2017-08-08

    Understanding the genetic basis of host shifts is a key genomic question for pathogen and parasite biology. The Bacillus cereus group, which encompasses Bacillus thuringiensis and Bacillus anthracis , contains pathogens that can infect insects, nematodes, and vertebrates. Since the target range of the essential virulence factors (Cry toxins) and many isolates is well known, this group presents a powerful system for investigating how pathogens can diversify and adapt to phylogenetically distant hosts. Specialization to exploit insects occurs at the level of the major clade and is associated with substantial changes in the core genome, and host switching between insect orders has occurred repeatedly within subclades. The transfer of plasmids with linked cry genes may account for much of the adaptation to particular insect orders, and network analysis implies that host specialization has produced strong associations between key toxin genes with similar targets. Analysis of the distribution of plasmid minireplicons shows that plasmids with orf156 and orf157 , which carry genes encoding toxins against Lepidoptera or Diptera, were contained only by B. thuringiensis in the specialized insect clade (clade 2), indicating that tight genome/plasmid associations have been important in adaptation to invertebrate hosts. Moreover, the accumulation of multiple virulence factors on transposable elements suggests that cotransfer of diverse virulence factors is advantageous in terms of expanding the insecticidal spectrum, overcoming insect resistance, or through gains in pathogenicity via synergistic interactions between toxins. IMPORTANCE Population genomics have provided many new insights into the formation, evolution, and dynamics of bacterial pathogens of humans and other higher animals, but these pathogens usually have very narrow host ranges. As a pathogen of insects and nematodes, Bacillus thuringiensis , which produces toxins showing toxicity to many orders of insects and

  19. [Experiments with Bacillus thuringiensis protoplasts. I. Isolation of protoplasts and their reversion to bacillary form].

    Science.gov (United States)

    Tsenin, A N; Nesterenko, A V; Rybchin, V N; Potokin, I L; Pisarevskiĭ, Iu S

    1983-04-01

    A method for protoplastization of crystal- and spore-forming Bacillus thuringiensis bacterian and consequent cell wall regeneration on a solid hypertonic medium is presented. Up to 50% of the protoplasts prepared were viable and formed colonies under special conditions; at the same time, less than 0,01% of the cells treated with lysozyme were resistant to the osmotic shock; bacterial autolytic system takes part in protoplasts formation. Electron microscopic studies of protoplasts and cells confirm the fact of cell wall removal and support the proposed mechanism of protoplast formation.

  20. Avances en el desarrollo de formulaciones insecticidas a base de bacillus thuringiensis

    OpenAIRE

    Rosas-García, Ninfa María

    2008-01-01

    El desarrollo de las formulaciones insecticidas elaboradas a base de la bacteria Bacillus thuringiensis es una tecnología centenaria que ha recibido un fuerte impacto en décadas recientes. La mezcla de esporas y cristales, que es el principio activo de estas preparaciones, ha sido objeto de estudio constante y en ello se destaca la búsqueda de cepas cada vez más potentes o mejoradas. Así mismo, los materiales utilizados incluyen una amplia variedad de ingredientes completamente biodegradables...

  1. Isolation of strains of Bacillus thuringiensis insecticidal biological activity against Ceratitis capitata

    International Nuclear Information System (INIS)

    Hmaied, Ezzedine; Ben Mbarek, Wael

    2010-01-01

    The present work is to study the effect of toxins (δ-endotoxins) extracted from strains of Bacillus thuringiensis isolated from the mud on the fly Sabkhat Dejoumi Ceratitis capitata, a pest of citrus and fruit trees. Among 51 isolated tested, 15 showed a very significant insecticidal activity, characterized by mortality rates exceeding 80 pour cent. These mortality rates are caused by endotoxins of Bt revealed variability between them. The preliminary results of this study encourage us towards the characterization of the insecticidal activity produced by strains of Bt for large scale application.

  2. Two new Brazilian isolates of Bacillus thuringiensis toxic to Anticarsia gemmatalis (Lepidoptera: Noctuidae

    Directory of Open Access Journals (Sweden)

    LM. Fiuza

    Full Text Available Bacillus thuringiensis is a bacterium used for biopesticides production and pest-resistant plants due to the synthesis of protein crystals by cry genes, which are effective in controlling several insect orders such as Lepidoptera. This work aimed at the evaluation and characterisation of two new B. thuringiensis isolates active against A. gemmatalis (Hübner 1818 larvae, which is the soybean major pest. The results showed that Bt117-4 isolate amplified fragments corresponding to cry2 and cry9 genes, and synthesised protein fragments equivalent to 130, 90 and 45 kDa. The Bt3146-4 isolate amplified DNA fragments corresponding to cry9 gene and synthesised protein fragments of 70, 58 and 38 kDa. Transmission electron microscopy revealed the presence of protein crystals in both isolates. CL50 with Cry purified proteins from Bt117-4 and Bt3146-4, corresponded to 0.195 and 0.191 µg larvae-1, respectively. The two B. thuringiensis isolates selected in this study were effective to control velvetbean caterpillar at laboratory conditions. Field tests should be carried on to develop new biopesticides formulation as well for cry genes resource for Anticarsia gemmatalis resistant transgenic plants.

  3. Phosphorus Availability in Wheat, in Volcanic Soils Inoculated with Phosphate-Solubilizing Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Jorge Delfim

    2018-01-01

    Full Text Available The use of phosphate solubilizing bacteria (PSB is an ecological strategy that allows for increasing the availability of phosphorus (P in soil. The objective of this study was to evaluate P availability in wheat, in soils derived from volcanic ash (Andisol and Ultisol, and inoculated with phosphate-solubilizing Bacillus thuringiensis, the experiment was conducted in pots under greenhouse conditions using a completely randomized design. Wheat plants were inoculated and re-inoculated at 20 and 46 days after sowing (DAS, respectively, with B. thuringiensis; and, soil and plant sampling was performed after 46, 66, and 87 days based on the Zadoks growth scale (Z. The inoculation resulted in an 11% increase in P of the rhizosphere at Z46 (Ultisol, P also increased 34% and 67% in aerial tissues at Z46 (Andisol and Ultisol, respectively, while an increase of 75% was observed in root tissues at Z87 (Ultisol. Similarly, the inoculation resulted in increases in acid phosphatase activity (Andisol, soil microbial biomass (Andisol and Ultisol, and root biomass in plants (Ultisol, without achieving increase of the aerial biomass of the plants. The phosphate solubilizing B. thuringiensis strain showed some positive, but also negative effects in soils and plants, depending on the soil.

  4. Efficacy of genetically modified Bt toxins alone and in combinations against pink bollworm resistant to Cry1Ac and Cry2Ab

    Science.gov (United States)

    Evolution of resistance in pests threatens the long-term success of transgenic crops that produce insecticidal proteins from Bacillus thuringiensis (Bt). Previous work showed that genetically modified Bt toxins Cry1AbMod and Cry1AcMod effectively countered resistance to native Bt toxins Cry1Ab and ...

  5. Biological Activity of Bacillus thuringiensis (Bacillales: Bacillaceae) in Anastrepha fraterculus (Diptera: Tephritidae).

    Science.gov (United States)

    Martins, Liliane Nachtigall; Lara, Ana Paula de Souza Stori de; Ferreira, Márcio Soares; Nunes, Adrise Medeiros; Bernardi, Daniel; Leite, Fábio Pereira Leivas; Garcia, Flávio Roberto Mello

    2018-01-18

    Anastrepha fraterculus (Wiedemann) (Diptera: Tephritidae) is considered to be one of the major pest insects in fruit orchards worldwide. Bacillus thuringiensis Berliner (Bacillales: Bacillaceae) strains are widely used as biological control agents and show high biological activity against different insect species. The objective of this study was to evaluate the biological activity of different strains of B. thuringiensis against A. fraterculus larvae and adults. Bioassays were performed using suspensions of bacterial spores/crystals of B. thuringiensis var. israelensis (Bti), kurstaki (Btk), and oswaldocruzi (Bto) strains at three concentrations [2 × 107, 2 × 108, and 2 × 109 colony-forming units per ml (CFU ml-1)]. At a concentration of 2 × 109 CFU ml-1, a significant larval effect (mortality 60%) was observed when compared with the control treatment. Larvae that ingested spore/crystal suspensions of Bti, Btk, or Bto bacterial strains exhibited significant larval and pupal deformations, leading to a significant decrease (~50%) in the completion of the insects' biological cycle (egg to adult). The B. thuringiensis strains (Bti, Btk, or Bto) at a concentration of 2 × 109 CFU ml-1 in combination with one food attractant (BioAnastrepha 3% or CeraTrap 1.5%) in formulations of toxic baits provided high mortality (mortality > 85%) of A. fraterculus adults 7 d after treatment. However, the Btk strain in combination with CeraTrap 1.5% caused mortality of 40%. On the basis of these results, the native bacterial strains Bti, Btk, and Bto were considered to be promising candidates as biological control agents against A. fraterculus. © The Author(s) 2018. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  6. Cloning of the Bacillus thuringiensis serovar sotto chitinase (Schi gene and characterization of its protein

    Directory of Open Access Journals (Sweden)

    Wan-Fang Zhong

    2005-12-01

    Full Text Available Chitinase plays a positive role in the pathogenicity of Bacillus thuringiensis to insect pests. We used touchdown PCR to clone the chitinase (Schi gene from Bacillus thuringiensis serovar sotto (Bt sotto chromosomal DNA. Our DNA sequencing analysis revealed that the Bt sotto Schi gene consists of an open reading frame (ORF of 2067 nucleotides with codes for the chitinase precursor. We also found that the putative promoter consensus sequences (the -35 and -10 regions of the Bt soto Schi gene are identical to those of the chiA71 gene from Bt Pakistani, the chiA74 gene from Bt kenyae and the ichi gene from Bt israelensis. The Schi chitinase precursor is 688 amino acids long with an estimated molecular mass of 75.75 kDa and a theoretical isoelectric point of 5.74, and contains four domains, which are, in sequence, a signal peptide, an N-terminal catalytic domain, a fibronectin type III like domain and a C-terminal chitin-binding domain. Sequence comparison and the evolutionary relationship of the Bt sotto Schi chitinase to other chitinase and chitinase-like proteins are also discussed.

  7. Adaptive Strategies of Bacillus thuringiensis Isolated from Acid Mine Drainage Site in Sabah, Malaysia.

    Science.gov (United States)

    Yik, Low Yi; Chin, Grace Joy Wei Lie; Budiman, Cahyo; Joseph, Collin Glenn; Musta, Baba; Rodrigues, Kenneth Francis

    2018-06-01

    The adaptive process in bacteria is driven by specific genetic elements which regulate phenotypic characteristics such as tolerance to high metal ion concentrations and the secretion of protective biofilms. Extreme environments such as those associated with heavy metal pollution and extremes of acidity offer opportunities to study the adaptive mechanisms of microorganisms. This study focused on the genome analysis of Bacillus thuringiensis (Bt MCMY1), a gram positive rod shaped bacterium isolated from an acid mine drainage site in Sabah, Malaysia by using a combination of Single Molecule Real Time DNA Sequencing, Scanning Electron Microscopy (SEM) and Fourier Transform Infrared Spectroscopy (FTIR). The genome size of Bt MCMY1 was determined to be 5,458,152 bases which was encoded on a single chromosome. Analysis of the genome revealed genes associated with resistance to Copper, Mercury, Arsenic, Cobalt, Zinc, Cadmium and Aluminum. Evidence from SEM and FTIR indicated that the bacterial colonies form distinct films which bear the signature of polyhydroxyalkanoates (PHA) and this finding was supported by the genome data indicating the presence of a genetic pathway associated with the biosynthesis of PHAs. This is the first report of a Bacillus sp. isolated from an acid mine drainage site in Sabah, Malaysia and the genome sequence will provide insights into the manner in which B. thuringiensis adapts to acid mine drainage.

  8. Bacillus thuringiensis strain NG, a Novel Isolated Strain for production of Various Polyhydroxyalkanoates

    Directory of Open Access Journals (Sweden)

    Nazila Gholamveisi

    2017-12-01

    Full Text Available Introduction: Microbial biopolymers such as polyhydroxyalkanoates (PHA are proper alternatives for petroleum-derived plastics. These biopolymers have many advantages over conventional plastics such as biodegradability, environmental friendly and infinite as a renewable resource. Therefore, our study was aimed to isolate a bacterial strain capable of producing polyhydroxybutyrate (PHB; a highly applicable type of PHA. Materials and methods: To this aim, a total of 6 PHA-producing bacteria were isolated from waste water exit site of a brewery factory. The 6 isolates were studied by Sudan black-staining technique and the most stained isolate was selected for further studies. Next, the selected isolate was identified based on morphological, biochemical and phylogenetic analyses. Finally, the ability of strain in producing PHB as well as other PHAs was analyzed via GC-MS technique. Results: Strain NG had the highest yield of PHB, according to Sudan black-staining technique and it was selected for further studies. The strain NG was identified as a new strain of Bacillus thuringiensis. According to GC-MS results, this strain was able to produce PHB as well as 4 other PHAs including hexadecanoic acid methyl ester, octadecanoic acid methyl ester, tetradecanoic acid methyl ester, 8-octadecenoic acid methyl ester. Discussion and conclusion: It was the first report on producing various PHAs at the same time by a strain of Bacillus thuringiensis.

  9. Characterization of a Chitin-Binding Protein from Bacillus thuringiensis HD-1.

    Directory of Open Access Journals (Sweden)

    Naresh Arora

    Full Text Available Strains of Bacillus thuringiensis produce insecticidal proteins. These strains have been isolated from diverse ecological niches, such as soil, phylloplane, insect cadavers and grain dust. To effectively propagate, these strains produce a range of molecules that facilitate its multiplication in a competing environment. In this report, we have examined synthesis of a chitin-binding protein and evaluated its effect on fungi encountered in environment and its interaction with insecticidal proteins synthesized by B. thuringiensis. The gene encoding chitin-binding protein has been cloned and expressed. The purified protein has been demonstrated to interact with Cry insecticidal protein, Cry1Ac by Circular Dichrosim spectroscopy (CD and in vitro pull down assays. The chitin-binding protein potentiates insecticidal activity of bacillar insecticidal protein, Cry1Ac. Further, chitin-binding protein was fungistatic against several soil fungi. The chitin binding protein is expressed in spore mother cell and deposited along with insecticidal protein, Cry1Ac. It interacts with Cry1Ac to potentiate its insecticidal activity and facilitate propagation of Bacillus strain in environment by inhibiting growth of certain fungi.

  10. Optimization of process parameters for enhanced biodegradation of acid red 119 by Bacillus thuringiensis SRDD

    Directory of Open Access Journals (Sweden)

    Riddhi H. Dave

    2012-02-01

    Full Text Available Developed Bacillus thuringiensis SRDD showed degradation of C.I. Acid red 119 and growth under the extremecondition of temperature 70°C, pH 3-8, heavy metals concentration of 0.8 mM, NaCl up to 900 mM and 1000 ppm dye. Cottonseed, caster cake and corn cake powders were found to be better and cheaper nutrient supplements for the Bacillus thuringiensisSRDD for biodegradation as compared to molasses. After development of the culture and the process, more than99% degradation was achieved in less than 2 hrs of contact time even on 18th cycles of addition of 100 ppm AR-119 dye. Thedeveloped process showed AR-119 biodegradation rate as high as 220 mg L-1 h-1, which is found to be 130 times more ascompared to the reported data. U.V., FTIR, TLC and HPLC analysis data confirmed biodegradation ability of the Bacillusthuringiensis for AR-119.

  11. Characterization of a Chitin-Binding Protein from Bacillus thuringiensis HD-1.

    Science.gov (United States)

    Arora, Naresh; Sachdev, Bindiya; Gupta, Rani; Vimala, Y; Bhatnagar, Raj K

    2013-01-01

    Strains of Bacillus thuringiensis produce insecticidal proteins. These strains have been isolated from diverse ecological niches, such as soil, phylloplane, insect cadavers and grain dust. To effectively propagate, these strains produce a range of molecules that facilitate its multiplication in a competing environment. In this report, we have examined synthesis of a chitin-binding protein and evaluated its effect on fungi encountered in environment and its interaction with insecticidal proteins synthesized by B. thuringiensis. The gene encoding chitin-binding protein has been cloned and expressed. The purified protein has been demonstrated to interact with Cry insecticidal protein, Cry1Ac by Circular Dichrosim spectroscopy (CD) and in vitro pull down assays. The chitin-binding protein potentiates insecticidal activity of bacillar insecticidal protein, Cry1Ac. Further, chitin-binding protein was fungistatic against several soil fungi. The chitin binding protein is expressed in spore mother cell and deposited along with insecticidal protein, Cry1Ac. It interacts with Cry1Ac to potentiate its insecticidal activity and facilitate propagation of Bacillus strain in environment by inhibiting growth of certain fungi.

  12. Characterization of a Chitin-Binding Protein from Bacillus thuringiensis HD-1

    Science.gov (United States)

    Gupta, Rani; Vimala, Y.; Bhatnagar, Raj K.

    2013-01-01

    Strains of Bacillus thuringiensis produce insecticidal proteins. These strains have been isolated from diverse ecological niches, such as soil, phylloplane, insect cadavers and grain dust. To effectively propagate, these strains produce a range of molecules that facilitate its multiplication in a competing environment. In this report, we have examined synthesis of a chitin-binding protein and evaluated its effect on fungi encountered in environment and its interaction with insecticidal proteins synthesized by B. thuringiensis. The gene encoding chitin-binding protein has been cloned and expressed. The purified protein has been demonstrated to interact with Cry insecticidal protein, Cry1Ac by Circular Dichrosim spectroscopy (CD) and in vitro pull down assays. The chitin-binding protein potentiates insecticidal activity of bacillar insecticidal protein, Cry1Ac. Further, chitin-binding protein was fungistatic against several soil fungi. The chitin binding protein is expressed in spore mother cell and deposited along with insecticidal protein, Cry1Ac. It interacts with Cry1Ac to potentiate its insecticidal activity and facilitate propagation of Bacillus strain in environment by inhibiting growth of certain fungi. PMID:23824872

  13. Bacillus thuringiensis Cry1Ca-resistant Spodoptera exigua lacks expression of one of four Aminopeptidase N genes

    NARCIS (Netherlands)

    Herrero, S.; Gechev, T.; Bakker, P.L.; Moar, W.; Maagd, de R.A.

    2005-01-01

    BACKGROUND: Insecticidal toxins from Bacillus thuringiensis bind to receptors on midgut epithelial cells of susceptible insect larvae. Aminopeptidases N (APNs) from several insect species have been shown to be putative receptors for these toxins. Here we report the cloning and expression analysis of

  14. A hybrid Bacillus thuringiensis delta-endotoxin gene gives resistance against a coleopteran and a lepidopteran pest in transgenic potato

    NARCIS (Netherlands)

    Naimov, S.; Dukiandjiev, S.; Maagd, de R.A.

    2003-01-01

    Expression of Bacillus thuringiensis delta-endotoxins has proven to be a successful strategy for obtaining insect resistance in transgenic plants. Drawbacks of expression of a single resistance gene are the limited target spectrum and the potential for rapid adaptation of the pest. Hybrid toxins

  15. Characterization of the pH-Mediated solubility of Bacillus thuringiensis var. san diego native δ-endotoxin crystals

    Science.gov (United States)

    C. N. Koller; L. S. Bauer; R. M. Hollingworth

    1992-01-01

    Native crystals of Bacillus thuringiensis var. san diego, a coleopteran-specific δ-endotoxin, were metabolically labeled with [35S]methionine. Specific activity was 82,000 CPM/μg (2.44 Ci/mmol). Using a universal buffer formulated with the same ionic strength at every pH, we determined that...

  16. Bacillus thuringiensis Cry3Aa protoxin intoxication of Tenebrio molitor induces widespread changes in the expression of serine peptidase transcripts

    Science.gov (United States)

    The yellow mealworm, Tenebrio molitor, is a pest of stored grain products and is sensitive to the coleopteran-specific Cry3Aa toxin from Bacillus thuringiensis (Bt). Larvae digest protein initially with cysteine peptidases in the anterior midgut and further with serine peptidases in middle and poste...

  17. Survival of Bacillus thuringiensis strains in gypsy moth (Lymantria dispar) larvae is correlated with production of urease

    Science.gov (United States)

    Phyllis A.W. Martin; Robert R. Jr. Farrar; Michael B. Blackburn

    2011-01-01

    We tested 50 lepidopteran-toxic Bacillus thuringiensis Berliner (Bt) strains with diverse phenotypes for the ability to survive repeated passages through larvae of the gypsy moth, Lymantria dispar (L.), without intervening growth on artificial media. These experiments have revealed a remarkable correlation...

  18. Isolation of transcripts from Diabrotica virgifera virgifera LeConte responsive to the Bacillus thuringiensis toxin Cry3Bb1

    Science.gov (United States)

    Crystal proteins derived from Bacillus thuringiensis (Bt) have been widely used as a method of insect pest management for several decades. In recent years, a transgenic corn expressing the Cry3Bb1 toxin has been successfully used for protection against corn rootworm larvae (Genus...

  19. Expression of Bacillus thuringiensis cytolytic toxin (Cyt2Ca1) in citrus roots to control Diaprepes abbreviatus larvae

    Science.gov (United States)

    Diaprepes abbreviatus (L.) is an important pest of citrus in the USA. Currently, no effective management strategies of Diaprepes abbreviatus exist in citriculture. To protect citrus against Diaprepes abbreviatus a transgenic citrus rootstock expressing Bacillus thuringiensis Cyt2Ca1, an insect toxin...

  20. Bacillus thuringiensis delta-endotoxin Cry1 hybrid proteins with increased activity against the Colorado potato beetle

    NARCIS (Netherlands)

    Naimov, S.; Weemen-Hendriks, M.; Dukiandjiev, S.; Maagd, de R.A.

    2001-01-01

    Cry1 delta-endotoxins of Bacillus thuringiensis are generally active against lepidopteran insects, but Cry1Ba and Cry1Ia have additional, though low, levels of activity against coleopterans such as the Colorado potato beetle. Here we report the construction of Cry1Ba/Cry1Ia hybrid toxins which have

  1. The introduction of integrated pest management in the Ethiopian horticultural sector : Bacillus thuringiensis strains and its toxicity

    NARCIS (Netherlands)

    Belder, den E.; Elderson, J.

    2012-01-01

    1 Introduction As hazards of conventional broad acting pesticides are documented, researchers, poli cymakers and growers look for pesticides that are toxic only to the target pest, have no impact on other such as beneficial species, and have fewer environmental effects. Bacillus thuringiensis (Bt)

  2. Recovery of Bacillus thuringiensis and related spore-forming bacteria from soil after application for gypsy moth control

    Science.gov (United States)

    Phyllis A.W. Martin; Elizabeth A. Mongeon; Michael B. Blackburn; Dawn E. Gundersen-Rindal

    2011-01-01

    Bacillus thuringiensis Berliner (Bt) has been applied for gypsy moth (Lymantria dispar L.) control in forests in the northeastern U.S. for many years. The subspecies of Bt that is used (urstaki) is not common in U.S. soil. We attempted to recover Bt from...

  3. Characterization of eight Bacillus thuringiensis isolates originated from fecal samples of Fuzhou Zoo and Fuzhou Panda Center

    Czech Academy of Sciences Publication Activity Database

    Wu, Ch.; Wu, L.; Zhang, L.; Gelbič, Ivan; Xu, L.; Guan, X.

    2014-01-01

    Roč. 17, č. 3 (2014), s. 395-397 ISSN 1226-8615 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * faeces * microscophy Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection Impact factor: 0.946, year: 2014 http://www.sciencedirect.com/science/article/pii/S122686151400034X

  4. Isolation and characterization of a novel native Bacillus thuringiensis strain BRC-HZM2 capable of degrading chlorpyrifos

    Czech Academy of Sciences Publication Activity Database

    Wu, S.; Peng, Y.; Huang, Z.; Huang, Z.; Xu, L.; Gelbič, Ivan; Guan, X.; Zhang, L.; Zou, S.

    2015-01-01

    Roč. 55, č. 3 (2015), s. 389-397 ISSN 0233-111X Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * biopesticide * biodegradation Subject RIV: EE - Microbiology, Virology Impact factor: 1.585, year: 2015 http://onlinelibrary.wiley.com/doi/10.1002/jobm.201300501/epdf

  5. Localization of Bacillus thuringiensis Cry1A toxin-binding molecules in gypsy moth larval gut sections using fluorescence microscopy

    Science.gov (United States)

    Algimantas P. Valaitis

    2011-01-01

    The microbial insecticide Bacillus thuringiensis (Bt) produces Cry toxins, proteins that bind to the brush border membranes of gut epithelial cells of insects that ingest it, disrupting the integrity of the membranes, and leading to cell lysis and insect death. In gypsy moth, Lymantria dispar, two toxin-binding molecules for the...

  6. 77 FR 6471 - Bacillus thuringiensis Cry2Ae Protein in Cotton; Exemption from the Requirement of a Tolerance

    Science.gov (United States)

    2012-02-08

    ... requirement of a tolerance for residues of Bacillus thuringiensis Cry2Ae protein in or on the food and feed... transformation using plasmid pTEM12. This PIP provides event GHB119 cotton protection against feeding damage by... for event GHB119 is BCS-GH005-8. The cry2Ae gene was isolated from Bt subspecies dakota and its...

  7. Nontarget impact of Bacillus thuringiensis kurstaki in central Appalachian mixed broadleaf-pine forests: long-term evaluation of arthropods

    Science.gov (United States)

    John S. Strazanac; George E. Seidel; Vicki Kondo; Cynthia J. Fritzler; Linda Butler

    2007-01-01

    Current measures for gypsy moth (Lymantria dispar L.) control emphasize the use of pheromones, growth regulators, and biopesticides. One of the biopesticides, Bacillus thuringiensis kurstaki (Btk), will continue to be necessary for immediate control of gypsy moth and other forest lepidopteran outbreaks. Although...

  8. Effect of proteolytic and detoxification enzyme inhibitors on Bacillus thuringiensis var. israelensis tolerance in the mosquito Aedes aegypti

    Czech Academy of Sciences Publication Activity Database

    Hu, X.; Guo, Y.; Wu, S.; Liu, Z.; Fu, T.; Shao, E.; Carballar-Lejarazú, R.; Zhao, G.; Huang, Z.; Gelbič, Ivan; Guan, X.; Zou, S.; Xu, L.; Zhang, L.

    2017-01-01

    Roč. 27, č. 2 (2017), s. 169-179 ISSN 0958-3157 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * Bti * Aedes aegypti Subject RIV: ED - Physiology OBOR OECD: Biology (theoretical, mathematical, thermal, cryobiology, biological rhythm), Evolutionary biology Impact factor: 0.919, year: 2016

  9. Beetle-specific Bacillus thuringiensis Cry3Aa toxin reduces larval growth and curbs reproduction in Spodoptera littoralis (Boisd.)

    Czech Academy of Sciences Publication Activity Database

    Hussein, Hany; Habuštová, Oxana; Sehnal, František

    2005-01-01

    Roč. 61, - (2005), s. 1186-1192 ISSN 1526-498X R&D Projects: GA ČR(CZ) GA522/02/1507 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis * Spodoptera littoralis * Bt applications Subject RIV: ED - Physiology Impact factor: 1.175, year: 2005

  10. Natural phytosanitary products effects on Bacillus Thuringiensis SUBSP. Kurstaki (BerlinerEfeito de produtos fitossanitários naturais sobre Bacillus Thuringiensis subesp. Kurstaki (Berliner

    Directory of Open Access Journals (Sweden)

    Everton Ricardi Lozano da Silva

    2012-12-01

    Full Text Available This work aimed to evaluate the effect of natural phytossanitary products (NPP on spores and crystal toxicity of Bacillus thuringiensis subsp. kurstaki – HD1 (Btk. For this commercial products (Agromos, Biogermex, Bovemax, Bordeaux mixture, Ecolife®, Dalneen, Matan Plus, Pyronin and Stüble-Aid® were used at three different concentrations. The effect of NPP on spores was assessed by comparing a suspension of Btk + NPP with sterile distilled water (SDW and another suspension with nutrient broth (NB, inoculated on nutrient agar (NA in Petri dishes to quantify the number of CFU/mL, 18 h after inoculation and incubation. The effect of NPP on crystals was evaluated with a suspension of Btk+SDW+NPP added to the artificial diet supplied for Anticarsia gemmatalis Hub. (Lepidoptera: Noctuidae quantifying the number of dead larvae at 12, 24, 48 and 72 h. Matan Plus was the only natural product that did not present effect on spores. All other products, regardless of concentration, decreased significantly CFU/mL Regarding crystals, Bordeaux mixture was the only one that reduced significantly Btk insecticidal activity at three concentrations. Este trabalho objetivou avaliar o efeito dos produtos fitossanitários naturais (PFN sobre esporos e sobre a toxicidade dos cristais de Bacillus thuringiensis subespécie kurstaki – HD1 (Btk. Para tal foram usados os produtos comerciais (Agromos, Biogermex, Bovemax, Calda Bordalesa, Ecolife®, Dalneen, Matan Plus, Pironin e Stüble –Aid® em três diferentes concentrações. O efeito dos PFN sobre esporos foi avaliado comparando-se suspensões de Btk + PFN com água destilada esterelizada (ADE e suspensões com caldo nutriente (CB, inoculadas em agar nutriente (AN, em placas de Petri quantificando-se o número de unidades formadoras de colônias (UFC / mL, 18 h após a inoculação e incubação. O efeito dos PFN sobre cristais foi avaliado com suspensões de Btk + ADE + PFN adicionados à dieta artificial

  11. The impact of inducing germination of Bacillus anthracis and Bacillus thuringiensis spores on potential secondary decontamination strategies.

    Science.gov (United States)

    Omotade, T O; Bernhards, R C; Klimko, C P; Matthews, M E; Hill, A J; Hunter, M S; Webster, W M; Bozue, J A; Welkos, S L; Cote, C K

    2014-12-01

    Decontamination and remediation of a site contaminated by the accidental or intentional release of fully virulent Bacillus anthracis spores are difficult, costly and potentially damaging to the environment. Development of novel decontamination strategies that have minimal environmental impacts remains a high priority. Although ungerminated spores are amongst the most resilient organisms known, once exposed to germinants, the germinating spores, in some cases, become susceptible to antimicrobial environments. We evaluated the concept that once germinated, B. anthracis spores would be less hazardous and significantly easier to remediate than ungerminated dormant spores. Through in vitro germination and sensitivity assays, we demonstrated that upon germination, B. anthracis Ames spores and Bacillus thuringiensis Al Hakam spores (serving as a surrogate for B. anthracis) become susceptible to environmental stressors. The majority of these germinated B. anthracis and B. thuringiensis spores were nonviable after exposure to a defined minimal germination-inducing solution for prolonged periods of time. Additionally, we examined the impact of potential secondary disinfectant strategies including bleach, hydrogen peroxide, formaldehyde and artificial UV-A, UV-B and UV-C radiation, employed after a 60-min germination-induction step. Each secondary disinfectant employs a unique mechanism of killing; as a result, germination-induction strategies are better suited for some secondary disinfectants than others. These results provide evidence that the deployment of an optimal combination strategy of germination-induction/secondary disinfection may be a promising aspect of wide-area decontamination following a B. anthracis contamination event. By inducing spores to germinate, our data confirm that the resulting cells exhibit sensitivities that can be leveraged when paired with certain decontamination measures. This increased susceptibility could be exploited to devise more

  12. Influence of multi-year Bacillus thuringiensis subsp. israelensis on the abundance of B. cereus group populations in Swedish riparian wetland soils

    DEFF Research Database (Denmark)

    Hendriksen, Niels Bohse; Schneider, Salome; Tajrin, Tania

    Bacillus thuringiensis subsp. israelensis (Bti) is a soil-born bacterium affiliated to the B. cereus group (Bcg, a group including the pathogens B. cereus, B. thuringiensis, and B. anthracis) and used in biocontrol products against nematoceran larvae. However, knowledge is limited on how long...

  13. Nano-Mechanical Properties of Heat Inactivated Bacillus anthracis and Bacillus thuringiensis Spores

    Science.gov (United States)

    2008-03-01

    spores were grown in plastic petri dishes on Criterion Dehydrated Culture Media , which contained per liter of formula 15 grams agar , 5 grams gelatin...became reality in 2001 when terrorists sent spores in a powdered form in letters to two senators and several news media offices, killing five people...is the causative agent of the disease anthrax. B. thuringiensis is often used in pesticides and bioengineering pest resistant crops because of its

  14. Proteolysis, histopathological effects, and immunohistopathological localization of delta-endotoxins of Bacillus thuringiensis subsp. kurstaki in the midgut of lepidopteran olive tree pathogenic insect Prays oleae.

    Science.gov (United States)

    Rouis, S; Chakroun, M; Saadaoui, I; Jaoua, S

    2007-02-01

    Considering the fact that Prays oleae is one of the most pathogenic insects to the olive tree in the Mediterranean basin, particularly in Tunisia, the mode of action of Cry insecticidal toxins of Bacillus thuringiensis kurstaki in Prays oleae midgut was investigated. The proteolysis of Bacillus thuringiensis delta-endotoxins in the midgut was a key step in determining their potency against Prays oleae. The latter's proteases activated the delta-endotoxins early, yielding stable toxins. The in vitro and in vivo binding of these toxins to Prays oleae larvae midgut was studied immunohistochemically, evidencing a midgut columnar cell vacuolization, microvilli damage, and then a pass of epithelium cell content into the larvae midgut. Moreover, Bacillus thuringiensis toxins were shown to bind to the apical microvilli of the midgut epithelial cells. The in vitro study of the interaction of Prays oleae midgut proteins with biotinylated Bacillus thuringiensis toxins allowed the prediction of four suitable receptor proteins in Prays oleae.

  15. Lack of detrimental effects of Bacillus thuringiensis Cry toxins on the insect predator Chrysoperla carnea: a toxicological, histopathological, and biochemical analysis

    NARCIS (Netherlands)

    Rodrigo-Simón, A.; Maagd, de R.A.; Avilla, C.; Bakker, P.L.; Molthoff, J.W.; González-Zamora, J.; Ferré, J.

    2006-01-01

    The effect of Cry proteins of Bacillus thuringiensis on the green lacewing (Chrysoperla carnea) was studied by using a holistic approach which consisted of independent, complementary experimental strategies. Tritrophic experiments were performed, in which lacewing larvae were fed Helicoverpa

  16. Susceptibility of the endangered Karner blue butterfly (Lepidoptera: Lycaenidae) to Bacillus thuringiensis var. kurstaki used for gypsy moth suppression in Michigan

    Science.gov (United States)

    Catherine Papp Herms; Deborah G. McCullough; Leah S. Bauer; Robert A. Haack; Norman R. Dubois

    1997-01-01

    We investigated the phenological and physiological susceptibility of the endangered Karner blue butterfly (Lycaeides melissa samuelis) to Bacillus thuringiensis var. kurstaki (Bt), a product widely used for gypsy moth (Lymantria dispar) suppression in Michigan and other infested states. We...

  17. Produccion por tecnologia de fermentacion de bacillus thuringiensis utilizando medios alternativos

    Directory of Open Access Journals (Sweden)

    Yaneth Amparo Muñoz-Peñalosa

    2000-01-01

    Full Text Available In the production by fermentation technology Bacillus thuringiensis of five alternative methods they were studied. The results of cell growth, working-level 100ml in static culture and temperature of 28 ° C, mostraronque the optimal substrate corresponded to the environment in which molasses and rice powder was added (alternative Medium No. 1] The cell development using 100 ml of substrate was studied with reciprocating shaking 110 rpm. in this test was determined filter curve creciemiento medio.The inoculum, allowed tiempode set the process in 6 hours. For the development of fermentations, are counted with the experimental equipment, glass bioreactor in two liters of capacity and aeration devices, mechanical stirring, sampling and output gases.The fermentation in the production of Bacillus thuringiensis is the type discontinuous submerged aerobic process and growth into account .Teniendo bibliographic information and preliminary results of the study, fixed working parameters were determined for production by fermentation of Bacillus thuringiensis, being alternative means No. 1, volume 1 liter temperature 28 ° C and cell concentration of the inoculum. To determine the optimum parameters of fermantacion was used a factorial design of experiments of the type 22, (two variables at two levels, with aeration (3.2-0.5 VVM and agitation (110-210 rpm .The fermentations performed 7, 4 design and 3 the average level of the variables. For monitoring fermentation sample was taken every 12 hours and cell concentration (Chamber of Neuvauer and pH was analyzed. The results of cell concentration measurement for fermentations at 60 hours shows that optimum working conditions and limitations correspond to the values ​​of the variable, 3.2 VVM aeration and agitation 210 Variable rpm.Significant was the aeration of pH in the fermentation media change neutral to acid and ended as a staple. A fermentations I were efectuo controlde microbiological quality, Gram

  18. Heat and desiccation are the predominant factors affecting inactivation of Bacillus licheniformis and Bacillus thuringiensis spores during simulated composting.

    Science.gov (United States)

    Stanford, K; Harvey, A; Barbieri, R; Xu, S; Reuter, T; Amoako, K K; Selinger, L B; McAllister, T A

    2016-01-01

    The suitability of composting for disposal of livestock mortalities due to Bacillus anthracis was assessed by measuring viability of surrogate spores from two strains each of Bacillus licheniformis and Bacillus thuringiensis after a heating cycle modelled on a cattle composting study. Sporulation was attempted from 10 to 37°C, but poor yields at lower temperatures resulted in 25, 30 and 37°C being selected to generate sufficient spores (8 log10  CFU ml(-1) ) for experiments. Spores were inoculated into 3 g autoclaved dried-ground compost rehydrated with 6 ml water or silica beads in a factorial design for each strain, sporulation temperature, matrix and sampling day (0, 25, 50, 100, 150). Maximum incubation temperature was 62°C, but spores were maintained at ≥55°C for 78 of 150 days. Although significant differences existed among Bacillus strains and sporulation temperatures, numbers of viable spores after 150 days averaged 1·3 log10  CFU g(-1) , a 5·2 log10 reduction from day 0. Spore inactivation was likely due to heat and desiccation as matrices were autoclaved prior to incubation, negating impacts of microflora. Results support composting for disposal of anthrax mortalities, provided long-term thermophillic heating is achieved. Due to limited sporulation at 10°C, livestock mortalities from anthrax at this or lower ambient temperatures would likely be of lower risk for disease transmission. © 2015 The Society for Applied Microbiology.

  19. Bacillus thuringiensis subsp. sichuansis strain MC28 produces a novel crystal protein with activity against Culex quinquefasciatus larvae.

    Science.gov (United States)

    Guan, Peng; Dai, Xiaojuan; Zhu, Jun; Li, Qiao; Li, Shuangcheng; Wang, Shiquan; Li, Ping; Zheng, Aiping

    2014-04-01

    The Bacillus thuringiensis subsp. sichuansis MC28 strain produces spherical parasporal crystals during sporulation and exhibits remarkable insecticidal activity against dipteran and lepidopteran pests. We characterized a novel cry gene (cry69Aa1), which was found in the pMC95 plasmid of the MC28 strain. The cry69Aa1 gene was inserted into a shuttle vector (pSTK) and expressed in an acrystalliferous mutant B. thuringiensis HD73⁻. In this transformant, a large number of spherical parasporal crystals, which were toxic to Culex quinquefasciatus (Diptera), were formed.

  20. Study of the Bacillus thuringiensis Vip3Aa16 histopathological effects and determination of its putative binding proteins in the midgut of Spodoptera littoralis

    OpenAIRE

    Abdelkefi-Mesrati, Lobna; Boukedi, Hanen; Dammak-Karray, Mariam; Sellami-Boudawara, Tahya; Jaoua, Samir; Tounsi, Slim

    2011-01-01

    The bacterium Bacillus thuringiensis produces, at the vegetative stage of its growth, Vip3A proteins with activity against a broad spectrum of lepidopteran insects. The Egyptian cotton leaf worm (Spodoptera littoralis) is an important agricultural pest that is susceptible to the Vip3Aa16 protein of Bacillus thuringiensis kurstaki strain BUPM95. The midgut histopathology of Vip3Aa fed larvae showed vacuolization of the cytoplasm, brush border membrane destruction, vesicle formation in the apic...

  1. c-di-GMP Regulates Various Phenotypes and Insecticidal Activity of Gram-Positive Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Yang Fu

    2018-02-01

    Full Text Available C-di-GMP has been well investigated to play significant roles in the physiology of many Gram-negative bacteria. However, its effect on Gram-positive bacteria is less known. In order to more understand the c-di-GMP functions in Gram-positive bacteria, we have carried out a detailed study on the c-di-GMP-metabolizing enzymes and their physiological functions in Bacillus thuringiensis, a Gram-positive entomopathogenic bacterium that has been applied as an insecticide successfully. We performed a systematic study on the ten putative c-di-GMP-synthesizing enzyme diguanylate cyclases (DGCs and c-di-GMP-degrading enzyme phosphodiesterases (PDEs in B. thuringiensis BMB171, and artificially elevated the intracellular c-di-GMP level in BMB171 by deleting one or more pde genes. We found increasing level of intracellular c-di-GMP exhibits similar activities as those in Gram-negative bacteria, including altered activities in cell motility, biofilm formation, and cell-cell aggregation. Unexpectedly, we additionally found a novel function exhibited by the increasing level of c-di-GMP to promote the insecticidal activity of this bacterium against Helicoverpa armigera. Through whole-genome transcriptome profile analyses, we found that 4.3% of the B. thuringiensis genes were differentially transcribed when c-di-GMP level was increased, and 77.3% of such gene products are involved in some regulatory pathways not reported in other bacteria to date. In summary, our study represents the first comprehensive report on the c-di-GMP-metabolizing enzymes, their effects on phenotypes, and the transcriptome mediated by c-di-GMP in an important Gram-positive bacterium.

  2. c-di-GMP Regulates Various Phenotypes and Insecticidal Activity of Gram-Positive Bacillus thuringiensis

    Science.gov (United States)

    Fu, Yang; Yu, Zhaoqing; Liu, Shu; Chen, Bo; Zhu, Li; Li, Zhou; Chou, Shan-Ho; He, Jin

    2018-01-01

    C-di-GMP has been well investigated to play significant roles in the physiology of many Gram-negative bacteria. However, its effect on Gram-positive bacteria is less known. In order to more understand the c-di-GMP functions in Gram-positive bacteria, we have carried out a detailed study on the c-di-GMP-metabolizing enzymes and their physiological functions in Bacillus thuringiensis, a Gram-positive entomopathogenic bacterium that has been applied as an insecticide successfully. We performed a systematic study on the ten putative c-di-GMP-synthesizing enzyme diguanylate cyclases (DGCs) and c-di-GMP-degrading enzyme phosphodiesterases (PDEs) in B. thuringiensis BMB171, and artificially elevated the intracellular c-di-GMP level in BMB171 by deleting one or more pde genes. We found increasing level of intracellular c-di-GMP exhibits similar activities as those in Gram-negative bacteria, including altered activities in cell motility, biofilm formation, and cell-cell aggregation. Unexpectedly, we additionally found a novel function exhibited by the increasing level of c-di-GMP to promote the insecticidal activity of this bacterium against Helicoverpa armigera. Through whole-genome transcriptome profile analyses, we found that 4.3% of the B. thuringiensis genes were differentially transcribed when c-di-GMP level was increased, and 77.3% of such gene products are involved in some regulatory pathways not reported in other bacteria to date. In summary, our study represents the first comprehensive report on the c-di-GMP-metabolizing enzymes, their effects on phenotypes, and the transcriptome mediated by c-di-GMP in an important Gram-positive bacterium. PMID:29487570

  3. c-di-GMP Regulates Various Phenotypes and Insecticidal Activity of Gram-PositiveBacillus thuringiensis.

    Science.gov (United States)

    Fu, Yang; Yu, Zhaoqing; Liu, Shu; Chen, Bo; Zhu, Li; Li, Zhou; Chou, Shan-Ho; He, Jin

    2018-01-01

    C-di-GMP has been well investigated to play significant roles in the physiology of many Gram-negative bacteria. However, its effect on Gram-positive bacteria is less known. In order to more understand the c-di-GMP functions in Gram-positive bacteria, we have carried out a detailed study on the c-di-GMP-metabolizing enzymes and their physiological functions in Bacillus thuringiensis , a Gram-positive entomopathogenic bacterium that has been applied as an insecticide successfully. We performed a systematic study on the ten putative c-di-GMP-synthesizing enzyme diguanylate cyclases (DGCs) and c-di-GMP-degrading enzyme phosphodiesterases (PDEs) in B. thuringiensis BMB171, and artificially elevated the intracellular c-di-GMP level in BMB171 by deleting one or more pde genes. We found increasing level of intracellular c-di-GMP exhibits similar activities as those in Gram-negative bacteria, including altered activities in cell motility, biofilm formation, and cell-cell aggregation. Unexpectedly, we additionally found a novel function exhibited by the increasing level of c-di-GMP to promote the insecticidal activity of this bacterium against Helicoverpa armigera . Through whole-genome transcriptome profile analyses, we found that 4.3% of the B. thuringiensis genes were differentially transcribed when c-di-GMP level was increased, and 77.3% of such gene products are involved in some regulatory pathways not reported in other bacteria to date. In summary, our study represents the first comprehensive report on the c-di-GMP-metabolizing enzymes, their effects on phenotypes, and the transcriptome mediated by c-di-GMP in an important Gram-positive bacterium.

  4. Isolation and characterization of a new Bacillus thuringiensis strain with a promising toxicity against Lepidopteran pests.

    Science.gov (United States)

    Boukedi, Hanen; Sellami, Sameh; Ktari, Sonia; Belguith-Ben Hassan, Najeh; Sellami-Boudawara, Tahya; Tounsi, Slim; Abdelkefi-Mesrati, Lobna

    2016-01-01

    Insecticides derived from Bacillus thuringiensis are gaining worldwide importance as environmentally desirable alternatives to chemicals for the control of pests in public health and agriculture. Isolation and characterization of new strains with higher and broader spectrum of activity is an ever growing field. In the present work, a novel Tunisian B. thuringiensis isolate named BLB459 was characterized and electrophoresis assay showed that among a collection of 200 B. thuringiensis strains, the plasmid profile of BLB459 was distinctive. SmaI-PFGE typing confirmed the uniqueness of the DNA pattern of this strain, compared with BUPM95 and HD1 reference strains. PCR and sequencing assays revealed that BLB459 harbored three cry genes (cry30, cry40 and cry54) corresponding to the obtained molecular sizes in the protein pattern. Interestingly, PCR-RFLP assay demonstrated the originality of the BLB459 cry30-type gene compared to the other published cry30 genes. Insecticidal bioassays showed that BLB459 spore-crystal suspension was highly toxic to both Ephestia kuehniella and Spodoptera littoralis with LC50 values of about 64 (53-75) and 80 (69-91) μg of toxin cm(-2), respectively, comparing with that of the commercial strain HD1 used as reference. Important histopathological effects of BLB459 δ-endotoxins on the two tested larvae midguts were detected, traduced by the vacuolization of the apical cells, the damage of microvilli, and the disruption of epithelial cells. These results proved that BLB459 strain could be of a great interest for lepidopteran biocontrol. Copyright © 2016 Elsevier GmbH. All rights reserved.

  5. Decolorization of dyehouse effluent and biodegradation of Congo red by Bacillus thuringiensis RUN1.

    Science.gov (United States)

    Olukanni, Olumide David; Osuntoki, Akinniyi A; Awotula, Ayodeji Olushola; Kalyani, Dayanand C; Gbenle, George Olabode; Govindwar, Sanjay P

    2013-06-28

    A dye-decolorizing bacterium was isolated from a soil sample and identified as Bacillus thuringiensis using 16S rRNA sequencing. The bacterium was able to decolorize three different textile dyes, namely, Reactive blue 13, Reactive red 58, and Reactive yellow 42, and a real dyehouse effluent up to 80-95% within 6 h. Some non-textile industrially important dyes were also decolorized to different extents. Fourier transform infrared spectroscopy and gas chromatography-mass spectrometer analysis of the ethyl acetate extract of Congo red dye and its metabolites showed that the bacterium could degrade it by the asymmetric cleavage of the azo bonds to yield sodium (4- amino-3-diazenylnaphthalene-1-sulfonate) and phenylbenzene. Sodium (4-amino-3-diazenylnaphthalene-1-sulfonate) was further oxidized by the ortho-cleavage pathway to yield 2- (1-amino-2-diazenyl-2-formylvinyl) benzoic acid. There was induction of the activities of laccase and azoreductase during the decolorization of Congo red, which suggests their probable role in the biodegradation. B. thuringiensis was found to be versatile and could be used for industrial effluent biodegradation.

  6. High instability of a nematicidal Cry toxin plasmid in Bacillus thuringiensis.

    Science.gov (United States)

    Sheppard, Anna E; Nakad, Rania; Saebelfeld, Manja; Masche, Anna C; Dierking, Katja; Schulenburg, Hinrich

    2016-01-01

    In bacterial pathogens, virulence factors are often carried on plasmids and other mobile genetic elements, and as such, plasmid evolution is central in understanding pathogenicity. Bacillus thuringiensis is an invertebrate pathogen that uses plasmid-encoded crystal (Cry) toxins to establish infections inside the host. Our study aimed to quantify stability of two Cry toxin-encoding plasmids, BTI_23p and BTI_16p, under standard laboratory culturing conditions. These two plasmids are part of the genome of the B. thuringiensis strain MYBT18679, which is of particular interest because of its high pathogenicity towards nematodes. One of the plasmids, BTI_23p, was found to be highly unstable, with substantial loss occurring within a single growth cycle. Nevertheless, longer term experimental evolution in the absence of a host revealed maintenance of the plasmid at low levels in the bacterial populations. BTI_23p encodes two nematicidal Cry toxins, Cry21Aa2 and Cry14Aa1. Consistent with previous findings, loss of the plasmid abolished pathogenicity towards the nematode Caenorhabditis elegans, which could be rescued by addition of Cry21Aa2-expressing Escherichia coli. These results implicate BTI_23p as a plasmid that is required for successful infection, yet unstable when present at high frequency in the population, consistent with the role of Cry toxins as public goods. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Cytotoxicity Analysis of Three Bacillus thuringiensis Subsp. israelensis δ-Endotoxins towards Insect and Mammalian Cells

    Science.gov (United States)

    Teixeira Corrêa, Roberto Franco; Ardisson-Araújo, Daniel Mendes Pereira; Monnerat, Rose Gomes; Ribeiro, Bergmann Morais

    2012-01-01

    Three members of the δ-endotoxin group of toxins expressed by Bacillus thuringiensis subsp. israelensis, Cyt2Ba, Cry4Aa and Cry11A, were individually expressed in recombinant acrystalliferous B. thuringiensis strains for in vitro evaluation of their toxic activities against insect and mammalian cell lines. Both Cry4Aa and Cry11A toxins, activated with either trypsin or Spodoptera frugiperda gastric juice (GJ), resulted in different cleavage patterns for the activated toxins as seen by SDS-PAGE. The GJ-processed proteins were not cytotoxic to insect cell cultures. On the other hand, the combination of the trypsin-activated Cry4Aa and Cry11A toxins yielded the highest levels of cytotoxicity to all insect cells tested. The combination of activated Cyt2Ba and Cry11A also showed higher toxic activity than that of toxins activated individually. When activated Cry4Aa, Cry11A and Cyt2Ba were used simultaneously in the same assay a decrease in toxic activity was observed in all insect cells tested. No toxic effect was observed for the trypsin-activated Cry toxins in mammalian cells, but activated Cyt2Ba was toxic to human breast cancer cells (MCF-7) when tested at 20 µg/mL. PMID:23029407

  8. Quantification of Bacillus thuringiensis Vip3Aa16 Entomopathogenic Toxin Using Its Hemolytic Activity.

    Science.gov (United States)

    Boukedi, Hanen; Ben Khedher, Saoussen; Ghribi, Dhouha; Dammak, Mariam; Tounsi, Slim; Abdelkefi-Mesrati, Lobna

    2017-05-01

    Vegetative insecticidal proteins produced by some Bacillus thuringiensis strains are specifically toxic to different agricultural pests such as the polyphagous Spodoptera and several other Lepidopteran insects, but one of the major problems found in the use of these biopesticides was the lack of an easy and credible method of quantification of such secreted toxins. Heterologous expression of B. thuringiensis Vip3Aa16 toxin was performed in Escherichia coli then the protein was purified by chromatography. Using blood agar as well as blood agar overlay (zymogram assay), we reported, for the first time, the capacity of Vip3Aa16 to induce hemolysis. The hemolytic activity of this protein was shown to be relatively stable after treatment at 40 °C and at a range of pH between 6.5 and 9. Moreover, a linear relationship was shown between hemolysis levels and Vip3Aa16 concentrations. The model established in the present study could quantify Vip3A toxin as a function of hemolytic activity and the assay proposed showed to be a simple and low-cost method to readily assess Vip3A toxins in liquid cultures and facilitate the use of this kind of bioinsecticides in pest management programs.

  9. Two distinct pathways lead Bacillus thuringiensis to commit to sporulation in biofilm.

    Science.gov (United States)

    Verplaetse, Emilie; Slamti, Leyla; Gohar, Michel; Lereclus, Didier

    2017-05-01

    The spore-forming bacterium Bacillus thuringiensis is an efficient biofilm producer, responsible for persistent contamination of industrial food processing systems. B. thuringiensis biofilms are highly heterogeneous bacterial structures in which three distinct cell types controlled by quorum sensing regulators were identified: PlcR-controlled virulent cells, NprR-dependent necrotrophic cells and cells committed to sporulation, a differentiation process controlled by Rap phosphatases and Spo0A-P. Interestingly, a cell lineage study revealed that, in LB medium or in insect larvae, only necrotrophic cells became spores. Here we analyzed cellular differentiation undertaken by cells growing in biofilm in a medium optimized for sporulation. No virulent cells were identified; surprisingly, two distinct routes could lead to differentiation as a spore in this growth condition: the NprR-dependent route, followed by the majority of cells, and the newly identified NprR-independent route, which is followed by 20% of sporulating cells. Copyright © 2016. Published by Elsevier Masson SAS.

  10. Susceptibility of Grapholita molesta (Busck, 1916) to formulations of Bacillus thuringiensis, individual toxins and their mixtures.

    Science.gov (United States)

    Ricietto, Ana Paula Scaramal; Gomis-Cebolla, Joaquín; Vilas-Bôas, Gislayne Trindade; Ferré, Juan

    2016-11-01

    The Oriental fruit moth, Grapholita molesta (Lepidoptera: Tortricidae), is a major pest of fruit trees worldwide, such as peach and apple. Bacillus thuringiensis has been shown to be an efficient alternative to synthetic insecticides in the control of many agricultural pests. The objective of this study was to evaluate the effectiveness of B. thuringiensis individual toxins and their mixtures for the control of G. molesta. Bioassays were performed with Cry1Aa, Cry1Ac, Cry1Ca, Vip3Aa, Vip3Af and Vip3Ca, as well as with the commercial products DiPel® and XenTari®. The most active proteins were Vip3Aa and Cry1Aa, with LC 50 values of 1.8 and 7.5ng/cm 2 , respectively. Vip3Ca was nontoxic to this insect species. Among the commercial products, DiPel® was slightly, but significantly, more toxic than XenTari®, with LC 50 values of 13 and 33ng commercial product/cm 2 , respectively. Since Vip3A and Cry1 proteins are expressed together in some insect-resistant crops, we evaluated possible synergistic or antagonistic interactions among them. The results showed moderate to high antagonism in the combinations of Vip3Aa with Cry1Aa and Cry1Ca. Copyright © 2016. Published by Elsevier Inc.

  11. Effects of Bacillus thuringiensis strains virulent to Varroa destructor on larvae and adults of Apis mellifera.

    Science.gov (United States)

    Alquisira-Ramírez, Eva Vianey; Peña-Chora, Guadalupe; Hernández-Velázquez, Víctor Manuel; Alvear-García, Andrés; Arenas-Sosa, Iván; Suarez-Rodríguez, Ramón

    2017-08-01

    The sublethal effects of two strains of Bacillus thuringiensis, which were virulent in vitro to Varroa destructor, were measured on Apis mellifera. The effects of five concentrations of total protein (1, 5, 25, 50 and 100μg/mL) from the EA3 and EA26.1 strains on larval and adult honey bees were evaluated for two and seven days under laboratory conditions. Based on the concentrations evaluated, total protein from the two strains did not affect the development of larvae, the syrup consumption, locomotor activity or proboscis extension response of adults. These same parameters were also tested for the effects of three concentrations (1, 10 and 15μg/kg) of cypermethrin as a positive control. Although no significant differences were observed after two days of treatment with cypermethrin, a dose-response relationship in syrup consumption and locomotor activity was observed. A significant reduction in the proboscis extension response of the bees treated with cypermethrin was also observed. Therefore, in contrast to cypermethrin, our results indicate that the EA3 and EA26.1 strains of B. thuringiensis can be used in beehives to control V. destructor and reduce the negative effects of this mite on colonies without adverse effects on the larvae and adults of A. mellifera. Additionally, the overuse of synthetic miticides, which produce both lethal and sublethal effects on bees, can be reduced. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Cytotoxicity analysis of three Bacillus thuringiensis subsp. israelensis δ-endotoxins towards insect and mammalian cells.

    Directory of Open Access Journals (Sweden)

    Roberto Franco Teixeira Corrêa

    Full Text Available Three members of the δ-endotoxin group of toxins expressed by Bacillus thuringiensis subsp. israelensis, Cyt2Ba, Cry4Aa and Cry11A, were individually expressed in recombinant acrystalliferous B. thuringiensis strains for in vitro evaluation of their toxic activities against insect and mammalian cell lines. Both Cry4Aa and Cry11A toxins, activated with either trypsin or Spodoptera frugiperda gastric juice (GJ, resulted in different cleavage patterns for the activated toxins as seen by SDS-PAGE. The GJ-processed proteins were not cytotoxic to insect cell cultures. On the other hand, the combination of the trypsin-activated Cry4Aa and Cry11A toxins yielded the highest levels of cytotoxicity to all insect cells tested. The combination of activated Cyt2Ba and Cry11A also showed higher toxic activity than that of toxins activated individually. When activated Cry4Aa, Cry11A and Cyt2Ba were used simultaneously in the same assay a decrease in toxic activity was observed in all insect cells tested. No toxic effect was observed for the trypsin-activated Cry toxins in mammalian cells, but activated Cyt2Ba was toxic to human breast cancer cells (MCF-7 when tested at 20 µg/mL.

  13. Analysis of abrB Expression during the Infectious Cycle of Bacillus thuringiensis Reveals Population Heterogeneity

    Directory of Open Access Journals (Sweden)

    Samia Ben Rejeb

    2017-12-01

    Full Text Available Using the model host/pathogen pair Galleria mellonella/Bacillus thuringiensis, we have shown that these bacteria could kill their insect host, survive in its cadaver and form spores by sequentially activating virulence, necrotrophism and sporulation genes. However, the population isolated from the cadavers was heterogeneous, including non-sporulating cells in an unknown physiological state. To characterize these bacteria, we used a transcriptional fusion between the promoter of a gene expressed during early exponential growth (abrB and a reporter gene encoding a destabilized version of GFP, in combination with a fluorescent reporter of the necrotrophic state. The composition of the bacterial population during infection was then analyzed by flow cytometry. We showed that the PabrB promoter was activated in the population that had turned on the necrotrophic reporter, suggesting a re-entry into vegetative growth. Strikingly, the cells that did not go through the necrotrophic state did not activate the PabrB promoter and appear as a dormant subpopulation. We propose a new model describing the B. thuringiensis cell types during infection.

  14. Fate of insecticidal Bacillus thuringiensis Cry protein in soil: differences between purified toxin and biopesticide formulation.

    Science.gov (United States)

    Hung, Truong Phuc; Truong, Le Van; Binh, Ngo Dinh; Frutos, Roger; Quiquampoix, Hervé; Staunton, Siobhán

    2016-12-01

    Bacillus thuringiensis produces insecticidal proteins known as Cry, and its efficiency and absence of side effects make it the most widely used biopesticide. There is little information on the role of soils in the fate of Cry proteins from commercial biopesticide formulations, unlike toxins from genetically modified crops, which have been intensively studied in recent years. The persistence of Cry in soil was followed under field and laboratory conditions. Sunlight accelerated loss of detectable Cry under laboratory conditions, but little effect of shade was observed under field conditions. The half-life of biopesticide proteins in soil under natural conditions was about 1 week. Strong temperature effects were observed, but they differed for biopesticide and purified protein, indicating different limiting steps. For the biopesticide, the observed decline in detectable protein was due to biological factors, possibly including the germination of B. thuringiensis spores, and was favoured by higher temperature. In contrast, for purified proteins, the decline in detectable protein was slower at low temperature, probably because the conformational changes of the soil-adsorbed protein, which cause fixation and hence reduced extraction efficiency, are temperature dependent. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  15. Isolation, characterization and toxicity of native Bacillus thuringiensis isolates from different hosts and habitats in Iran

    Directory of Open Access Journals (Sweden)

    Ghassemi-Kahrizeh Akbar

    2017-09-01

    Full Text Available Bacillus thuringiensis is a Gram-positive, aerobic, facultative anaerobic and endospore-forming bacterium. Different strains of this species have the ability to produce parasporal crystalline inclusions which are toxic to larvae of different insect orders and other invertebrates and cause rapid death of the host. To determine the importance of this species in microbial control, we collected native strains and studied their virulence on the diamondback moth, Plutella xylostella. More than 148 samples were collected from Alborz, Guilan and Mazandaran Provinces. Experimental samples, including soil samples from forests, fruit gardens, agricultural fields, diseased and dead larvae, were transferred to a laboratory in sterile plastic containers. For evaluating B. thuringiensis isolates virulence, a cabbage leaf dip method with 106 cell · ml−1 concentration of various Bt isolates was applied to diamondback moths. Larval mortality was recorded 72 h after treatment. Based on bioassay results, all isolates were classified into three high, medium and low virulence groups. Protein level characterization based on the SDS-PAGE gel analysis showed that two isolates from a high virulence group have proteins of high molecular masses of 121 and 109 kDa. Results revealed that there is a positive correlation between protein masses and virulence of isolates. In addition, this research introduced nine strains that are highly toxic to P. xylostella and would be valuable as insecticidal agents for controlling lepidopteran pests.

  16. Probiotic Enterococcus mundtii Isolate Protects the Model Insect Tribolium castaneum against Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Thorben Grau

    2017-07-01

    Full Text Available Enterococcus mundtii strains isolated from the larval feces of the Mediterranean flour moth Ephestia kuehniella show antimicrobial activity against a broad spectrum of Gram-positive and Gram-negative bacteria. The in vitro probiotic characterization of one isolate revealed a high auto-aggregation score, a hydrophilic cell surface, tolerance for low pH, no hemolytic activity, and susceptibility to all tested antibiotics. We used the red flour beetle Tribolium castaneum, an established model organism, for the in vivo characterization of one probiotic E. mundtii isolate from E. kuehniella larvae. Tribolium castaneum larvae were fed orally with the probiotic isolate or the corresponding supernatant and then infected with either the entomopathogen Bacillus thuringiensis or Pseudomonas entomophila. Larvae exposed to the isolate or the supernatant showed increased survival following infection with B. thuringiensis but not P. entomophila. Heat treatment or treatment with proteinase K reduced the probiotic effect of the supernatant. However, the increased resistance attracts a fitness penalty manifested as a shorter lifespan and reduced fertility. T. castaneum has, pending on further research, the potential as an alternative model for the pre-screening of probiotics.

  17. Novel toxicity of Bacillus thuringiensis strains against the melon fruit fly, Bactrocera cucurbitae (Diptera: Tephritidae).

    Science.gov (United States)

    Shishir, Md Asaduzzaman; Akter, Asma; Bodiuzzaman, Md; Hossain, M Aftab; Alam, Md Musfiqul; Khan, Shakil Ahmed; Khan, Shakila Nargis; Hoq, M Mozammel

    2015-01-01

    Bactrocera cucurbitae (melon fruit fly) is one of the most detrimental vegetable-damaging pests in Bangladesh. The toxicity of Bacillus thuringiensis (Bt) has been reported against a few genera of Bactrocera in addition to numerous other insect species. Bt strains, harbouring cry1A-type genes were, therefore, assayed in vivo against the 3(rd) instar larvae of B. cucurbitae in this study. The biotype-based prevalence of cry1 and cry1A genes was calculated to be 30.8% and 11.16%, respectively, of the test strains (n=224) while their prevalence was greatest in biotype kurstaki. Though three indigenous Bt strains from biotype kurstaki with close genetic relationship exhibited higher toxicity, maximum mortalities were recorded for Btk HD-73 (96%) and the indigenous Bt JSc1 (93%). LC50 and LC99 values were determined to be 6.81 and 8.32 for Bt JSc1, 7.30 and 7.92 for Bt SSc2, and 6.99 and 7.67 for Btk HD-73, respectively. The cause of toxicity and its variation among the strains was found to be correlated with the synergistic toxic effects of cry1, cry2, cry3 and cry9 gene products, i.e. relevant Cry proteins. The novel toxicity of the B. thuringiensis strains against B. cucurbitae revealed in the present study thus will help in developing efficient and eco-friendly control measures such as Bt biopesticides and transgenic Bt cucurbits.

  18. Structure and distribution of the Bacillus thuringiensis Cry4Ba toxin in lipid membranes

    Energy Technology Data Exchange (ETDEWEB)

    Puntheeranurak, Theeraporn [Institute for Biophysics, Johannes Kepler University of Linz, Altenbergerstr. 69, A-4040 Linz (Austria); Laboratory of Molecular Biophysics, Institute of Molecular Biology and Genetics, Mahidol University, Salaya Campus, Nakornpathom 73170 (Thailand); Stroh, Cordula [Institute for Biophysics, Johannes Kepler University of Linz, Altenbergerstr. 69, A-4040 Linz (Austria); Zhu Rong [Institute for Biophysics, Johannes Kepler University of Linz, Altenbergerstr. 69, A-4040 Linz (Austria); Angsuthanasombat, Chanan [Laboratory of Molecular Biophysics, Institute of Molecular Biology and Genetics, Mahidol University, Salaya Campus, Nakornpathom 73170 (Thailand); Hinterdorfer, Peter [Institute for Biophysics, Johannes Kepler University of Linz, Altenbergerstr. 69, A-4040 Linz (Austria)]. E-mail: peter.hinterdorfer@jku.at

    2005-11-15

    Bacillus thuringiensis Cry {delta}-endotoxins cause death of susceptible insect larvae by forming lytic pores in the midgut epithelial cell membranes. The 65 kDa trypsin activated Cry4Ba toxin was previously shown to be capable of permeabilizing liposomes and forming ionic channels in receptor-free planar lipid bilayers. Here, magnetic ACmode (MACmode) atomic force microscopy (AFM) was used to characterize the lateral distribution and the native molecular structure of the Cry4Ba toxin in the membrane. Liposome fusion and the Langmuir-Blodgett technique were employed for supported lipid bilayer preparations. The toxin preferentially inserted in a self-assembled structure, rather than as a single monomeric molecule. In addition, the spontaneous insertion into receptor-free lipid bilayers lead to formation of characteristic pore-like structures with four-fold symmetry, suggesting that tetramers are the preferred oligomerization state of this toxin.

  19. Bacillus thuringiensis Cry1Aa toxin-binding region of Bombyx mori aminopeptidase N.

    Science.gov (United States)

    Yaoi, K; Nakanishi, K; Kadotani, T; Imamura, M; Koizumi, N; Iwahana, H; Sato, R

    1999-12-17

    The Bacillus thuringiensis Cry1Aa toxin-binding region of Bombyx mori aminopeptidase N (APN) was analyzed, to better understand the molecular mechanism of susceptibility to the toxin and the development of resistance in insects. APN was digested with lysylendopeptidase and the ability of the resulting fragments to bind to Cry1Aa and 1Ac toxins was examined. The binding abilities of the two toxins to these fragments were different. The Cry1Aa toxin bound to the fragment containing 40-Asp to 313-Lys, suggesting that the Cry1Aa toxin-binding site is located in the region between 40-Asp and 313-Lys, while Cry1Ac toxin bound exclusively to mature APN. Next, recombinant APN of various lengths was expressed in Escherichia coli cells and its ability to bind to Cry1Aa toxin was examined. The results localized the Cry1Aa toxin binding to the region between 135-Ile and 198-Pro.

  20. Effects and mechanisms of Bacillus thuringiensis crystal toxins for mosquito larvae.

    Science.gov (United States)

    Zhang, Qi; Hua, Gang; Adang, Michael J

    2017-10-01

    Bacillus thuringiensis is a Gram-positive aerobic bacterium that produces insecticidal crystalline inclusions during sporulation phases of the mother cell. The virulence factor, known as parasporal crystals, is composed of Cry and Cyt toxins. Most Cry toxins display a common 3-domain topology. Cry toxins exert intoxication through toxin activation, receptor binding and pore formation in a suitable larval gut environment. The mosquitocidal toxins of Bt subsp. israelensis (Bti) were found to be highly active against mosquito larvae and are widely used for vector control. Bt subsp. jegathesan is another strain which possesses high potency against broad range of mosquito larvae. The present review summarizes characterized receptors for Cry toxins in mosquito larvae, and will also discuss the diversity and effects of 3-D mosquitocidal Cry toxin and the ongoing research for Cry toxin mechanisms generated from investigations of lepidopteran and dipteran larvae. © 2016 Institute of Zoology, Chinese Academy of Sciences.

  1. Structure and distribution of the Bacillus thuringiensis Cry4Ba toxin in lipid membranes

    International Nuclear Information System (INIS)

    Puntheeranurak, Theeraporn; Stroh, Cordula; Zhu Rong; Angsuthanasombat, Chanan; Hinterdorfer, Peter

    2005-01-01

    Bacillus thuringiensis Cry δ-endotoxins cause death of susceptible insect larvae by forming lytic pores in the midgut epithelial cell membranes. The 65 kDa trypsin activated Cry4Ba toxin was previously shown to be capable of permeabilizing liposomes and forming ionic channels in receptor-free planar lipid bilayers. Here, magnetic ACmode (MACmode) atomic force microscopy (AFM) was used to characterize the lateral distribution and the native molecular structure of the Cry4Ba toxin in the membrane. Liposome fusion and the Langmuir-Blodgett technique were employed for supported lipid bilayer preparations. The toxin preferentially inserted in a self-assembled structure, rather than as a single monomeric molecule. In addition, the spontaneous insertion into receptor-free lipid bilayers lead to formation of characteristic pore-like structures with four-fold symmetry, suggesting that tetramers are the preferred oligomerization state of this toxin

  2. Bacillus thuringiensis in caterpillars and associated materials collected from protected tropical forests in northwestern Costa Rica.

    Science.gov (United States)

    Rodríguez-Sánchez, César; Sittenfeld, Ana; Janzen, Daniel H; Espinoza, Ana M

    2006-06-01

    Bacillus thuringiensis (Bt) synthesizes crystalline inclusions that are toxic to caterpillars (Lepidoptera) and other orders of invertebrates. Materials associated with 37 caterpillars from 16 species, collected while feeding on 15 different species of host plants in dry, cloud and rain forests located in the Area de Conservación Guanacaste in northwestern Costa Rica, were examined for the presence of Bt. From a total of 101 derived samples, 25 Bt isolates were cultured: 56% from host plant leaves, 8% from caterpillar guts and 36% from caterpillar fecal pellets. Bt was isolated from at least one sample in 38% of the systems constituted by the food plant, gut and fecal pellets corresponding to a single caterpillar. Four different morphologies of crystalline inclusions were observed, with bipyramidal and irregular crystal morphologies being the most prevalent.

  3. How the insect pathogen bacteria Bacillus thuringiensis and Xenorhabdus/Photorhabdus occupy their hosts.

    Science.gov (United States)

    Nielsen-LeRoux, Christina; Gaudriault, Sophie; Ramarao, Nalini; Lereclus, Didier; Givaudan, Alain

    2012-06-01

    Insects are the largest group of animals on earth. Like mammals, virus, fungi, bacteria and parasites infect them. Several tissue barriers and defense mechanisms are common for vertebrates and invertebrates. Therefore some insects, notably the fly Drosophila and the caterpillar Galleria mellonella, have been used as models to study host-pathogen interactions for several insect and mammal pathogens. They are excellent tools to identify pathogen determinants and host tissue cell responses. We focus here on the comparison of effectors used by two different groups of bacterial insect pathogens to accomplish the infection process in their lepidopteran larval host: Bacillus thuringiensis and the nematode-associated bacteria, Photorhabdus and Xenorhabdus. The comparison reveals similarities in function and expression profiles for some genes, which suggest that such factors are conserved during evolution in order to attack the tissue encountered during the infection process. Copyright © 2012 Elsevier Ltd. All rights reserved.

  4. Overcome of Carbon Catabolite Repression of Bioinsecticides Production by Sporeless Bacillus thuringiensis through Adequate Fermentation Technology

    Directory of Open Access Journals (Sweden)

    Saoussen Ben Khedher

    2014-01-01

    Full Text Available The overcoming of catabolite repression, in bioinsecticides production by sporeless Bacillus thuringiensis strain S22 was investigated into fully controlled 3 L fermenter, using glucose based medium. When applying adequate oxygen profile throughout the fermentation period (75% oxygen saturation, it was possible to partially overcome the catabolite repression, normally occurring at high initial glucose concentrations (30 and 40 g/L glucose. Moreover, toxin production yield by sporeless strain S22 was markedly improved by the adoption of the fed-batch intermittent cultures technology. With 22.5 g/L glucose used into culture medium, toxin production was improved by about 36% when applying fed-batch culture compared to one batch. Consequently, the proposed fed-batch strategy was efficient for the overcome of the carbon catabolite repression. So, it was possible to overproduce insecticidal crystal proteins into highly concentrated medium.

  5. Sub-chronic lung inflammation after airway exposures to Bacillus thuringiensis biopesticides in mice

    DEFF Research Database (Denmark)

    Barfod, Kenneth K; Poulsen, Steen Seier; Hammer, Maria

    2010-01-01

    The aim of the present study was to assess possible health effects of airway exposures to Bacillus thuringiensis (Bt) based biopesticides in mice. Endpoints were lung inflammation evaluated by presence of inflammatory cells in bronchoalveolar lavage fluid (BALF), clearance of bacteria from the lung...... lumen and histological alterations of the lungs. Hazard identifications of the biopesticides were carried out using intratracheal (i.t.) instillation, followed by an inhalation study. The two commercial biopesticides used were based on the Bt. subspecies kurstaki and israelensis, respectively. Groups...... of BALB/c mice were i.t instilled with one bolus (3.5 × 105 or 3.4 × 106 colony forming units (CFU) per mouse) of either biopesticide. Control mice were instilled with sterile water. BALFs were collected and the inflammatory cells were counted and differentiated. The BALFs were also subjected to CFU...

  6. Microbiological ecology and association of Bacillus thuringiensis in chicken feces originating from feed

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Peng, Y.; Wu, S.; Sun, L.; Huang, E.; Huang, T.; Xu, L.; Wu, Ch.; Gelbič, Ivan; Guan, X.

    2012-01-01

    Roč. 65, č. 6 (2012), s. 784-791 ISSN 0343-8651 R&D Projects: GA MŠk 2B08003 Grant - others:National Natural Science Foundation of China(CN) 31071745; Ministry of Education of China(CN) 20093515110010; Ministry of Education of China(CN) 20093515120010; Transformation Fund for Agricultural Science and Technology Achievements(CN) 2010GB2C400212; Education Department Foundation(CN) JA12092 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis Subject RIV: EH - Ecology, Behaviour Impact factor: 1.520, year: 2012 http://link.springer.com/content/pdf/10.1007%2Fs00284-012-0231-3

  7. Cost-effective production of Bacillus thuringiensis by solid-state fermentation.

    Science.gov (United States)

    Devi, P S Vimala; Ravinder, T; Jaidev, C

    2005-02-01

    Production of Bacillus thuringiensis (Bt) was standardized on wheat bran based media in 250 ml Erlenmeyer flasks. Scale-up of Bt production on the best medium in plastic tubs with aeration at 8 h intervals starting 16 h after incubation yielded a significant increase in spore count and toxin content of the product. Maximum lysis of Bt cells was obtained by 60 h of incubation at 30 degrees C. This protocol was suitable for production of Bt strains and local isolates. The Bt produced proved highly effective at 0.1% concentration against larvae of castor semilooper, Achaea janata L, resulting in complete mortality by three days in laboratory bioassays. In field trials, the population of castor semilooper larvae on the castor bean crop was reduced significantly by three days after application. The cost for material production of 1 kg of Bt was approximately US dollars 0.70.

  8. Efficiency of Intergeneric Recombinants Between Bacillus Thuringiensis and Bacillus Subtilis for Increasing Mortality Rate in Cotten Leaf Worm

    Science.gov (United States)

    AlOtaibi, Saad Aied

    2012-12-01

    In this study , two strains of Bacillus belonging to two serotypes and four of their transconjugants were screened with respect to their toxicity against lepidopterous cotton pest. . Bacterial transconjugants isolated from conjugation between both strains were evaluated for their transconjugant efficiency caused mortality in Spodoptera littoralis larvae . Two groups of bioinsecticides ; crystals , crystals and spores have been isolated from Bacillusstrains and their transconjugants . Insecticidal crystal protein ( ICP ) was specific for lepidopteran insects because of the toxin sufficient both for insect specificity and toxicity . The toxicities of these two groups against larvae of Spodoptera littoralis was expressed as transconjugant efficiency , which related to the mean number of larvae died expressed as mortality percentage . The results showed transconjugant efficiency in reducing the mean number of Spodoptera littoralis larvae feeding on leaves of Ricinus communis sprayed with bioinsecticides of Bt transconjugants. Most values of positive transconjugant efficiency related to increasing mortality percentage are due to toxicological effects appeared in response to the treatments with crystals + endospores than that of crystals alone .This indicated that crystals + endospores was more effective for increasing mortality percentage than that resulted by crystals . Higher positive transconjugant efficiency in relation to the mid parents and better parent was appeared at 168 h of treatment . The results indicated that recombinant Bacillus thuringiensis are important control agents for lepidopteran pests , as well as , susceptibility decreased with larval development . The results also suggested a potential for the deployment of these recominant entomopathogens in the management of Spodoptera. littoralis larvae .

  9. Mosquito biolarvicide production by sequential fermentation with dual strains of Bacillus thuringiensis subsp. israelensis and Bacillus sphaericus using sewage sludge.

    Science.gov (United States)

    Zhuang, Li; Zhou, Shungui; Wang, Yueqiang; Chang, Min

    2011-01-01

    This study demonstrated the bioconversion of sewage sludge into a composite biolarvicide for mosquito control based on sequential fermentation with dual strains of Bacillus thuringiensis subsp. israelensis (Bti) and Bacillus sphaericus (Bs). Results showed that sewage sludge was a suitable fermentation substrate for supporting growth, sporulation and mosquitocidal proteins synthesis by Bti and Bs. Through sequential fermentation with dual strains, a 10-L bench-scale fermentor was capable of producing Bti and Bs at a cell concentration of 2.1×10(9) and 6.8×10(8) CFU/mL, respectively. Such sequential fermentation can save half of raw materials and energy consumption comparing with the sludge fermentation with single strain. The toxic activity and persistence of the composite biolarvicide against mosquito larvae in the polluted waters were enhanced by the increased toxin complexity and synergistic interactions. This study, for the first time, validates the technical feasibility of using sewage sludge to produce a cost-effective composite biolarvicide based on Bti and Bs. Copyright © 2010 Elsevier Ltd. All rights reserved.

  10. Purification and characterization of a new Bacillus thuringiensis bacteriocin active against Listeria monocytogenes, Bacillus cereus and Agrobacterium tumefaciens.

    Science.gov (United States)

    Kamoun, Fakher; Fguira, Ines Ben; Hassen, Najeh Belguith Ben; Mejdoub, Hafedh; Lereclus, Didier; Jaoua, Samir

    2011-09-01

    This study reports on the identification, characterization and purification of a new bacteriocin, named Bacthuricin F103, from a Bacillus thuringiensis strain BUPM103. Bacthuricin F103 production began in the early exponential phase and reached a maximum in the middle of the same phase. Two chromatographic methods based on high performance liquid chromatography and fast protein liquid chromatography systems were used to purify Bacthuricin F103. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis revealed that this bacteriocin had a molecular weight of approximately 11 kDa. It also showed a wide range of thermostability of up to 80 °C for 60 min and a broad spectrum of antimicrobial activity over a pH range of 3.0-10.0. This bacteriocin was noted, and for the first time, to exhibit potent antimicrobial activity against Agrobacterium subsp. strains, the major causal agents of crown gall disease in tomato and vineyard crops, and against several challenging organisms in food, such as Listeria monocytogenes and Bacillus cereus. Complete killing with immediate impact on cells was observed within a short period of time. The sequence obtained for Bacthuricin F103 by direct N-terminal sequencing shared considerable homology with hemolysin. Bacthuricin F103 was noted to act through the depletion of intracellular ions, which suggest that the cell membrane was a possible target to Bacthuricin F103.

  11. Growth of hydroxyapatite on the cellular membrane of the bacterium Bacillus thuringiensis for the preparation of hybrid biomaterials

    International Nuclear Information System (INIS)

    Cervantes, Eric Reyes; Torres, Maykel González; Muñoz, Susana Vargas; Rosas, Efraín Rubio

    2016-01-01

    This study aimed to grow hydroxyapatite (HAp) crystals on the cellular wall of the Gram-positive bacterium Bacillus thuringiensis using a bio-mimetic method. Several strains were phenotypically and genotypically characterized using multilocus sequence typing (MLST) gene markers to differentiate the strains and confirm the identity of the isolated species to guarantee that the selected species was not harmful to human health or the environment. Three of the analyzed strains were selected because they exhibited the best nucleation and growth of HAp on the bacterial surface. This innovative method to grow HAp crystals on a cellular membrane helps to elucidate the mechanisms by which osseous tissue is formed in nature. The optimum concentration for the simulated physiological fluid (SPF) was 1.5 ×. The hybrid materials were characterized by optical microscopy, atomic force microscopy (AFM), scanning electron microscopy (SEM), X-ray powder diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR). - Highlights: • HAp crystals are grown on the cellular wall of a GP bacteria Bacillus thuringiensis. • The growing was carried out by using a bio-mimetic method. • Hybrid materials were characterized with morphological and spectroscopic techniques. • The reported method allows understanding the mechanisms to produce osseous tissue. • The membrane of Bacillus thuringiensis can grow more HAp than Bacillus halodurans.

  12. Production of the bioinsecticide Bacillus thuringiensis subsp. israelensis with deltamethrin increases toxicity towards mosquito larvae.

    Science.gov (United States)

    Tetreau, G; Patil, C D; Chandor-Proust, A; Salunke, B K; Patil, S V; Després, L

    2013-08-01

    Bacillus thuringiensis subsp. israelensis is a bioinsecticide used for larval mosquito control and it represents a safe alternative to chemical insecticides. Despite its environmental safety, it is less efficient and persistent than chemical insecticides. To bypass these limitations, we propose to combine the advantages of chemical and biological insecticides by producing Bti in a medium supplemented with a chemical insecticide (DDT, deltamethrin, permethrin, propoxur or temephos). Among the investigated insecticides, the addition of deltamethrin in the medium induced a higher toxicity (over 6.72-fold) of the composite deltamethrin-Bti towards mosquito larvae as compared to Bti alone. This was mainly due to the insertion of deltamethrin into the membranes of Bti spores, as evidenced by a quantification of membrane-extracted deltamethrin by HPLC. This composite larvicide is a promising tool to decrease the quantity of chemicals dispersed in the environment, to increase the efficacy of Bti and to facilitate its widespread use as a transition between chemical and biological insecticides. Further experiments are required to characterize the mechanisms that underline the incorporation of deltamethrin into Bti to optimize the production and the toxicity of this composite larvicide. This study is the first report of an increased efficacy of the mosquitocidal bioinsecticide Bacillus thuringiensis subsp. israelensis (Bti) when produced with a chemical insecticide. The results clearly demonstrate that deltamethrin is able to synergize the insecticidal activity of Bti through inclusion into spore membranes, reducing off-target and nonspecific toxicity occurring when the chemical is used alone as sprays. This new composite chemical-biological insecticide can become an invaluable tool as an intermediate between single chemical usage and the widespread use of Bti, notably in developing countries with limited financial resources for intensive mosquito control campaigns. © 2013

  13. RESIDUAL TOXICITY OF BACILLUS THURINGIENSIS H-14 (VCRC B17 IN SOME TYPES OF BREEDING PLACES OF AEDES AEGYPTI

    Directory of Open Access Journals (Sweden)

    Salamun Salamun

    2012-09-01

    Full Text Available Bacillus thuringiensis H-14, adalah agensia mikrobial yang sangat spesifik terhadap serangga sasaran, aman terhadap golongan mamalia, dan tidak mencemari lingkungan, sehingga dapat dikembangkan sebagai agensia untuk pengendalian vektor, khususnya vektor demam berdarah dengue di Indonesia. Toksisitas residual B. thuringiensis H-14 (VCRC B17 terhadap larva instar III Aedes aegypti pada beberapa tipe tempat penampung air telah dievaluasi di dalam laboratorium. Hasil evaluasi menunjukkan bahwa angka kematian larva uji lebih dari 80% oleh pengaruh B. thuringiensis H-14 (VCRC B17 pada konsentrasi antara 1 sampai 25 mg/l di dalam tipe tempat penampung air dari semen, tanah liat, dan plastik masing-masing adalah 16 sampai 60 hari, 18 sampai 36 hari, dan 12 sampai 42 hari.

  14. Selection of optimum conditions of medium acidity and aeration for submerget cultivation of Bacillus thuringiensis and Beauveria bassiana

    Directory of Open Access Journals (Sweden)

    O. A. Dregval

    2010-06-01

    Full Text Available The paper deals with the influence of medium pH and aeration rate on growth and sporulation of Bacillus thuringiensis and Вeauveria bassiana, which are main constituents of the complex microbial insecticide. It was established optimal medium pH for B. thuringiensis – 6.0 and for В. bassiana – 6.0–7.0. The maximum productivity of the studied microorganisms was observed in the same range of aeration – 7– 14 mmol O2/l/h. The selected conditions of cultivation are necessary for the production of complex biological insecticide based on the association of B. thuringiensis and B. bassiana.

  15. Isolation of Bacillus thuringiensis from the state of Amazonas, in Brazil, and screening against Aedes aegypti (Diptera, Culicidae

    Directory of Open Access Journals (Sweden)

    Joelma Soares-da-Silva

    2015-03-01

    Full Text Available We investigated the use of Bacillus thuringiensis isolated in the state of Amazonas, in Brazil, for the biological control of the dengue vector Aedes aegypti. From 25 soil samples collected in nine municipalities, 484 bacterial colonies were obtained, 57 (11.78% of which were identified as B. thuringiensis. Six isolates, IBt-03, IBt-06, IBt-07, IBt-28, IBt-30, and BtAM-27 showed insecticidal activity, and only BtAM-27 presents the five genes investigated cry4Aa, cry4Ba, cry10Aa, cry11Aa, and cry11Ba. The IBt-07 and IBt- 28, with lower LC50 values, showed equal toxicity compared to the standards. The isolates of B. thuringiensis from Amazonas constitute potential new means of biological control for A. aegypti, because of their larvicidal activity and the possibility that they may also contain new combinations of toxins.

  16. Evaluation of Bacillus thuringiensis Pathogenicity for a Strain of the Tick, Rhipicephalus microplus, Resistant to Chemical Pesticides

    Science.gov (United States)

    Fernández-Ruvalcaba, Manuel; Peña-Chora, Guadalupe; Romo-Martínez, Armando; Hernández-Velázquez, Víctor; de Parra, Alejandra Bravo; De La Rosa, Diego Pérez

    2010-01-01

    The pathogenicity of four native strains of Bacillus thuringiensis against Rhipicephalus (Boophilus) microplus (Canestrine) (Acari: Ixodidae) was evaluated. A R. microplus strain that is resistant to organophosphates, pyrethroids, and amidines, was used in this study. Adult R. microplus females were bioassayed using the immersion test of Drummond against 60 B. thuringiensis strains. Four strains, GP123, GP138, GP130, and GP140, were found to be toxic. For the immersion test, the total protein concentration for each bacterial strain was 1.25 mg/ml. Mortality, oviposition, and egg hatch were recorded. All of the bacterial strains had significant effects compared to the controls, but no significant differences were seen between the 4 strains. It is evident that these B. thuringiensis strains have a considerable detrimental effect on the R. microplus strain that is resistant to pesticides. PMID:21062139

  17. Isolation and characterization of radioresistant mutants in Bacillus subtilis and Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Kalinin, V.L.; Petrov, V.N.; Petrova, T.M.

    1981-01-01

    Vegetative cells of Bac. thuringiensis var. galleriae (the wild-type strain 351) are much more sensitive to lethal effects of UV light and 60 Co-γ-rays than those of Bac. subtilis (the wild-type strain 168). This difference is less pronounced for spores of these strains. By means of repeated γ-irradiation-regrowth cycles radioresistant mutants Bac. thuringiensis Gamsup(r) 14 and Bac. subtilis Gamsup(r) 9 were selected. The vegetative cells of these mutants are correspondingly 19 times and 3.9 times more resistant to lethal effects of γ-radiation than the cells of the parental strains. The resistance of the Gamsup(r) mutant cells to lethal effects of UV light and H 2 O 2 is also increased. The spores of the Gamsup(r) 14 mutant are 1.5-1.7 times more resistant to γ-radiation and UV light than the wild-type spores. The radioresistant mutants and the parental strains do not vary in their capacity for host-cell reactivation of UV- or γ-irradiated phages Tg13 and 105

  18. The chromosome map of Bacillus thuringiensis subsp. canadensis HD224 is highly similar to that of the Bacillus cereus type strain ATCC 14579.

    Science.gov (United States)

    Carlson, C R; Johansen, T; Kolstø, A B

    1996-08-01

    A physical map of the Bacillus thuringiensis subsp. canadensis HD224 chromosome based on AscI, NotI, and SfiI restriction sites has been established. The chromosome map of 4.3 Mb was similar to a revised map of the chromosome of the B. cereus type strain ATCC 14579, except that the B. thuringiensis subsp. canadensis HD224 chromosome lacked a NotI site and had two additional AscI sites. The positions of 27 probes were identical in the common macromap. A probe for the insecticidal toxin gene, cryIA, hybridized only to the B. thuringiensis subsp. canadensis HD224 chromosome. The BssHII ribotype patterns were almost identical confirming the similarity between the two strains.

  19. New Role for DCR-1/Dicer in Caenorhabditis elegans Innate Immunity against the Highly Virulent Bacterium Bacillus thuringiensis DB27

    Science.gov (United States)

    Iatsenko, Igor; Sinha, Amit; Rödelsperger, Christian

    2013-01-01

    Bacillus thuringiensis produces toxins that target invertebrates, including Caenorhabditis elegans. Virulence of Bacillus strains is often highly specific, such that B. thuringiensis strain DB27 is highly pathogenic to C. elegans but shows no virulence for another model nematode, Pristionchus pacificus. To uncover the underlying mechanisms of the differential responses of the two nematodes to B. thuringiensis DB27 and to reveal the C. elegans defense mechanisms against this pathogen, we conducted a genetic screen for C. elegans mutants resistant to B. thuringiensis DB27. Here, we describe a B. thuringiensis DB27-resistant C. elegans mutant that is identical to nasp-1, which encodes the C. elegans homolog of the nuclear-autoantigenic-sperm protein. Gene expression analysis indicated a substantial overlap between the genes downregulated in the nasp-1 mutant and targets of C. elegans dcr-1/Dicer, suggesting that dcr-1 is repressed in nasp-1 mutants, which was confirmed by quantitative PCR. Consistent with this, the nasp-1 mutant exhibits RNA interference (RNAi) deficiency and reduced longevity similar to those of a dcr-1 mutant. Building on these surprising findings, we further explored a potential role for dcr-1 in C. elegans innate immunity. We show that dcr-1 mutant alleles deficient in microRNA (miRNA) processing, but not those deficient only in RNAi, are resistant to B. thuringiensis DB27. Furthermore, dcr-1 overexpression rescues the nasp-1 mutant's resistance, suggesting that repression of dcr-1 determines the nasp-1 mutant's resistance. Additionally, we identified the collagen-encoding gene col-92 as one of the downstream effectors of nasp-1 that play an important role in resistance to DB27. Taken together, these results uncover a previously unknown role for DCR-1/Dicer in C. elegans antibacterial immunity that is largely associated with miRNA processing. PMID:23918784

  20. Bacillus cereus and Bacillus thuringiensis spores in Korean rice: prevalence and toxin production as affected by production area and degree of milling.

    Science.gov (United States)

    Kim, Booyoung; Bang, Jihyun; Kim, Hoikyung; Kim, Yoonsook; Kim, Byeong-Sam; Beuchat, Larry R; Ryu, Jee-Hoon

    2014-09-01

    We determined the prevalence of and toxin production by Bacillus cereus and Bacillus thuringiensis in Korean rice as affected by production area and degree of milling. Rough rice was collected from 64 farms in 22 agricultural areas and polished to produce brown and white rice. In total, rice samples were broadly contaminated with B. cereus spores, with no effect of production area. The prevalence and counts of B. cereus spores declined as milling progressed. Frequencies of hemolysin BL (HBL) production by isolates were significantly (P ≤ 0.01) reduced as milling progressed. This pattern corresponded with the presence of genes encoding the diarrheal enterotoxins. The frequency of B. cereus isolates positive for hblC, hblD, or nheB genes decreased as milling progressed. Because most B. cereus isolates from rice samples contained six enterotoxin genes, we concluded that B. cereus in rice produced in Korea is predominantly of the diarrheagenic type. The prevalence of B. thuringiensis in rice was significantly lower than that of B. cereus and not correlated with production area. All B. thuringiensis isolates were of the diarrheagenic type. This study provides information useful for predicting safety risks associated with B. cereus and B. thuringiensis in rough and processed Korean rice. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. Evidence of two mechanisms involved in Bacillus thuringiensis israelensis decreased toxicity against mosquito larvae: Genome dynamic and toxins stability.

    Science.gov (United States)

    Elleuch, Jihen; Zribi Zghal, Raida; Lacoix, Marie Noël; Chandre, Fabrice; Tounsi, Slim; Jaoua, Samir

    2015-07-01

    Biopesticides based on Bacillus thuringiensis israelensis are the most used and most successful around the world. This bacterium is characterized by a dynamic genome able to win or lose genetic materials which leads to a decrease in its effectiveness. The detection of such phenomena is of great importance to monitor the stability of B. thuringiensis strains in industrial production processes of biopesticides. New local B. thuringiensis israelensis isolates were investigated. They present variable levels of delta-endotoxins production and insecticidal activities against Aedes aegypti larvae. Searching on the origin of this variability, molecular and biochemical analyses were performed. The obtained results describe two main reasons of the decrease of B. thuringiensis israelensis insecticidal activity. The first reason was the deletion of cry4Aa and cry10Aa genes from the 128-kb pBtoxis plasmid as evidenced in three strains (BLB124, BLB199 and BLB506) among five. The second was the early degradation of Cry toxins by proteases in larvae midgut mainly due to some amino acids substitutions evidenced in Cry4Ba and Cry11Aa δ-endotoxins detected in BLB356. Before biological treatment based on B. thuringiensis israelensis, the studies of microflore in each ecosystem have a great importance to succeed pest management programs. Copyright © 2015 Elsevier GmbH. All rights reserved.

  2. Decolorization of textile dye RB19 using volcanic rock matrix immobilized Bacillus thuringiensis cells with surface displayed laccase.

    Science.gov (United States)

    Wan, Juan; Sun, Xiaowen; Liu, Cheng; Tang, Mengjun; Li, Lin; Ni, Hong

    2017-06-01

    A triplicate volcanic rock matrix-Bacillus thuringiensis-laccase WlacD (VRMs-Bt-WlacD) dye decolorization system was developed. WlacD was displayed on the B. thuringiensis MB174 cell surface to prepare a whole-cell laccase biocatalyst by using two repeat N-terminal domains of autolysin Mbg (Mbgn) 2 as the anchoring motif. Immunofluorescence microscopic assays confirmed that the fusion protein (Mbgn) 2 -WlacD was anchored on the surface of the recombinant B. thuringiensis MB174. After optimization by a single factor test, L 9 (3 4 )-orthogonal test, Plackett-Burman test, steepest ascent method, and Box-Behnken response surface methodology, the whole-cell specific laccase activity of B. thuringiensis MB174 was improved to 555.2 U L -1 , which was 2.25 times than that of the primary culture condition. Optimized B. thuringiensis MB174 cells were further adsorbed by VRMs to prepare VRMs-Bt-WlacD, an immobilized whole-cell laccase biocatalyst. Decolorization capacity of as-prepared VRMs-Bt-WlacD toward an initial concentration of 500 mg L -1 of an textile dye reactive blue 19 (RB19) aqueous solution reached 72.36% at a solid-to-liquid ratio of 10 g-100 mL. Repeated decolorization-activation operations showed the high decolorization capacity of VRMs-Bt-WlacD and have the potential for large-scale or continuous operations.

  3. The Pathogenomic Sequence Analysis of B. cereus and B.thuringiensis Isolates Closely Related to Bacillus anthracis

    Energy Technology Data Exchange (ETDEWEB)

    Han, Cliff S.; Xie, Gary; Challacombe, Jean F.; Altherr, MichaelR.; Smriti, B.; Bruce, David; Campbell, Connie S.; Campbell, Mary L.; Chen, Jin; Chertkov, Olga; Cleland, Cathy; Dimitrijevic-Bussod, M.; Doggett, Norman A.; Fawcett, John J.; Glavina, Tijana; Goodwin, Lynne A.; Hill, Karen K.; Hitchcock, Penny; Jackson, Paul J.; Keim, Paul; Kewalramani, Avinash Ramesh; Longmire, Jon; Lucas, Susan; Malfatti,Stephanie; McMurry, Kim; Meincke, Linda J.; Misra, Monica; Moseman,Bernice L.; Mundt, Mark; Munk, A. Christine; Okinaka, Richard T.; Parson-Quintana, B.; Reilly, Lee P.; Richardson, Paul; Robinson, DonnaL.; Rubin, Eddy; Saunders, Elizabeth; Tapia, Roxanne; Tesmer, Judith G.; Thayer, Nina; Thompson, Linda S.; Tice, Hope; Ticknor, Lawrence O.; Wills, Patti L.; Gilna, Payl; Brettin, Thomas S.

    2005-08-18

    The sequencing and analysis of two close relatives of Bacillus anthracis are reported. AFLP analysis of over 300 isolates of B.cereus, B. thuringiensis and B. anthracis identified two isolates as being very closely related to B. anthracis. One, a B. cereus, BcE33L, was isolated from a zebra carcass in Nambia; the second, a B. thuringiensis, 97-27, was isolated from a necrotic human wound. The B. cereus appears to be the closest anthracis relative sequenced to date. A core genome of over 3,900 genes was compiled for the Bacillus cereus group, including Banthracis. Comparative analysis of these two genomes with other members of the B. cereus group provides insight into the evolutionary relationships among these organisms. Evidence is presented that differential regulation modulates virulence, rather than simple acquisition of virulence factors. These genome sequences provide insight into the molecular mechanisms contributing to the host range and virulence of this group of organisms.

  4. A constitutively expressed 36 kDa exochitinase from Bacillus thuringiensis HD-1.

    Science.gov (United States)

    Arora, Naresh; Ahmad, Tarannum; Rajagopal, R; Bhatnagar, Raj K

    2003-08-01

    A 36 kDa chitinase was purified by ion exchange and gel filtration chromatography from the culture supernatant of Bacillus thuringiensis HD-1. The chitinase production was independent of the presence of chitin in the growth medium and was produced even in the presence of glucose. The purified chitinase was active at acidic pH, had an optimal activity at pH 6.5, and showed maximum activity at 65 degrees C. Of the various substrates, the enzyme catalyzed the hydrolysis of the disaccharide 4-MU(GlnAc)(2) most efficiently and was therefore classified as an exochitinase. The sequence of the tryptic peptides showed extensive homology with Bacillus cereus 36 kDa exochitinase. The 1083 bp open reading frame encoding 36 kDa chitinase was amplified with primers based on the gene sequence of B. cereus 36 kDa exochitinase. The deduced amino-acid sequence showed that the protein contained an N-terminal signal peptide and consisted of a single catalytic domain. The two conserved signature sequences characteristic of family 18 chitinases were mapped at positions 105-109 and 138-145 of Chi36. The recombinant chitinase was expressed in a catalytically active form in Escherichia coli in the vector pQE-32. The expressed 36 kDa chitinase potentiated the insecticidal effect of the vegetative insecticidal protein (Vip) when used against neonate larvae of Spodoptera litura.

  5. Modular columns to study depth-dependence behavior of mosquito larvae and toxicity of Bacillus thuringiensis subsp. israelensis.

    Science.gov (United States)

    Meir, Michal; Ben-Dov, Eitan; Zaritsky, Arieh

    2011-03-01

    Modular transparent column system was designed to study depth-dependence behavior of mosquito larvae. The system was used in preliminary experiments to evaluate the effect of water depth on the larvicidal activity of Bacillus thuringiensis subsp. israelensis de Barjac against bottom feeder larvae of Aedes aegypti (Linn.) (Diptera: Culicidae), and suggestions for increasing the efficiency of the device are discussed. Copyright © 2010 Elsevier B.V. All rights reserved.

  6. Mannose Phosphate Isomerase Isoenzymes in Plutella xylostella Support Common Genetic Bases of Resistance to Bacillus thuringiensis Toxins in Lepidopteran Species

    OpenAIRE

    Herrero, Salvador; Ferré, Juan; Escriche, Baltasar

    2001-01-01

    A strong correlation between two mannose phosphate isomerase (MPI) isoenzymes and resistance to Cry1A toxins from Bacillus thuringiensis has been found in a Plutella xylostella population. MPI linkage to Cry1A resistance had previously been reported for a Heliothis virescens population. The fact that the two populations share similar biochemical, genetic, and cross-resistance profiles of resistance suggests the occurrence of homologous resistance loci in both species.

  7. Extraction and separation of water soluble proteins from Bacillus thuringiensis-transgenic and non-transgenic maize species by CZE

    Czech Academy of Sciences Publication Activity Database

    Sázelová, Petra; Kašička, Václav; Ibanez, E.; Cifuentes, A.

    2009-01-01

    Roč. 32, č. 21 (2009), s. 3801-3808 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GA203/08/1428 Grant - others:GA ČR(CZ) GA203/09/0675 Program:GA Institutional research plan: CEZ:AV0Z40550506 Keywords : Bacillus thuringiensis -transgenic maize * CZE-UV profiling * Maize proteins Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.551, year: 2009

  8. Effect of ultraviolet and gamma rays on the activity of delta-endotoxin protein crystals of Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Burges, H.D.; Hillyer, S.; Chanter, D.O.

    1975-01-01

    Sensitive bioassays with larvae of Pieris brassicae revealed no reduction of insecticidal activity as a result of severe gamma or ultraviolet irradiation of crystals of Bacillus thuringiensis (serotype V). The measured response was the inhibition of larval feeding by the crystals over exposure periods short enough for the presence of live spores not to influence feeding. The results were analyzed using a logistic model. (U.S.)

  9. Fly ash based Bacillus thuringiensis var. israelensis formulation for use against Culex quinquefasciatus, the vector of filariasis in natural ecosystems

    OpenAIRE

    S Tamilselvan; P Jambulingam; V Manoharan; R Shanmugasundaram; G Vivekanandan; A M Manonmani

    2015-01-01

    Background & objectives: Fly ash is produced in huge quantities by the various thermal power stations in India. This thermal waste has been employed as a carrier material in the preparation of a biopesticidal water dispersible powder (WDP) formulation for use against mosquitoes. In the present investigation, this newly developed fly ash based WDP formulation was evaluated in natural breeding habitats of mosquito. Methods: Fly ash based WDP formulation of Bacillus thuringiensis var. israele...

  10. Low translocation of Bacillus thuringiensis israelensis to inner organs in mice after pulmonary exposure to commercial biopesticide

    DEFF Research Database (Denmark)

    Barfod, Kenneth Klingenberg; Ørum-Smidt, Lasse; Krogfelt, Karen A.

    2010-01-01

    Translocation of viable cells from a Bacillus thuringiensis israelensis-based biopesticide to inner organs in a mouse model was studied. Mice were exposed to the originally formulated product through the lungs and gastrointestinal tract by intratracheal instillation. Colony forming units (CFU) were...... grown from lungs, caecum, spleen and liver on Bacillus cereus-specific agar (BCSA) after 24 h and finally determined to be biopesticide strain B. t. israelensis by large plasmid profile. No CFU were found in spleen or liver of the control mice or in any aerosol background or material. We have shown...... biopesticides in the future....

  11. Application of statistical experimental design for optimisation of bioinsecticides production by sporeless Bacillus thuringiensis strain on cheap medium

    Directory of Open Access Journals (Sweden)

    Saoussen Ben Khedher

    2013-09-01

    Full Text Available In order to overproduce bioinsecticides production by a sporeless Bacillus thuringiensis strain, an optimal composition of a cheap medium was defined using a response surface methodology. In a first step, a Plackett-Burman design used to evaluate the effects of eight medium components on delta-endotoxin production showed that starch, soya bean and sodium chloride exhibited significant effects on bioinsecticides production. In a second step, these parameters were selected for further optimisation by central composite design. The obtained results revealed that the optimum culture medium for delta-endotoxin production consists of 30 g L-1 starch, 30 g L-1 soya bean and 9g L-1 sodium chloride. When compared to the basal production medium, an improvement in delta-endotoxin production up to 50% was noted. Moreover, relative toxin yield of sporeless Bacillus thuringiensis S22 was improved markedly by using optimised cheap medium (148.5 mg delta-endotoxins per g starch when compared to the yield obtained in the basal medium (94.46 mg delta-endotoxins per g starch. Therefore, the use of optimised culture cheap medium appeared to be a good alternative for a low cost production of sporeless Bacillus thuringiensis bioinsecticides at industrial scale which is of great importance in practical point of view.

  12. Application of statistical experimental design for optimisation of bioinsecticides production by sporeless Bacillus thuringiensis strain on cheap medium.

    Science.gov (United States)

    Ben Khedher, Saoussen; Jaoua, Samir; Zouari, Nabil

    2013-01-01

    In order to overproduce bioinsecticides production by a sporeless Bacillus thuringiensis strain, an optimal composition of a cheap medium was defined using a response surface methodology. In a first step, a Plackett-Burman design used to evaluate the effects of eight medium components on delta-endotoxin production showed that starch, soya bean and sodium chloride exhibited significant effects on bioinsecticides production. In a second step, these parameters were selected for further optimisation by central composite design. The obtained results revealed that the optimum culture medium for delta-endotoxin production consists of 30 g L(-1) starch, 30 g L(-1) soya bean and 9 g L(-1) sodium chloride. When compared to the basal production medium, an improvement in delta-endotoxin production up to 50% was noted. Moreover, relative toxin yield of sporeless Bacillus thuringiensis S22 was improved markedly by using optimised cheap medium (148.5 mg delta-endotoxins per g starch) when compared to the yield obtained in the basal medium (94.46 mg delta-endotoxins per g starch). Therefore, the use of optimised culture cheap medium appeared to be a good alternative for a low cost production of sporeless Bacillus thuringiensis bioinsecticides at industrial scale which is of great importance in practical point of view.

  13. FORMULASI PADAT RHIZOBAKTERIA INDIGENUS BACILLUS THURINGIENSIS TS2 DAN WAKTU PENYIMPANAN UNTUK MENGENDALIKAN PENYAKIT PUSTUL BAKTERI XANTHOMONAS AXONOPODIS PV. GLYCINES

    Directory of Open Access Journals (Sweden)

    Yulmira Yanti

    2017-05-01

    Full Text Available Solid formulations of indigenous rhizobacteria Bacillus thuringiensis TS2 and storage time to control bacterial pustule disease Xanthomonas axonopodis pv. glycines. Bacterial pustule disease caused by Xanthomonas axonopodis pv. Glycines is a major constraint in soybean cultivation. Indigenous rhizobacteria Bacillus thuringiensis TS2 from soybean rhizosphere acquired from previous research is the best isolate which can control soybean bacterial pustule disease and increase growth rate of soybean. To increased its stability and interaction with soybean plants, Bacillus thuringiensis TS2 was urged to test furthermore especially its formulation with based formula tapioca powder, peat and bulk. The most effective storage time also need to test. Result showed that all rhizobacterial formula had ability to decrease incidence of bacterial pustule disease compared to control. Moreover, all the three formula could increase plant growth, total of leaves, total of branch and yields. Flowering time was also advanced by 1-8 days compared to control. Decreasing of disease rate and increasing of plant growth rate variated between different formulations.

  14. Leaf morphology and ultrastructure responses to elevated O3 in transgenic Bt (cry1Ab/cry1Ac) rice and conventional rice under fully open-air field conditions.

    Science.gov (United States)

    Li, Chunyan; Liu, Biao; Li, Chunhua; Zeng, Qing; Hao, Mingzhuo; Han, Zhengmin; Zhu, Jianguo; Li, Xiaogang; Shen, Wenjing

    2013-01-01

    Elevated tropospheric ozone severely affects not only yield but also the morphology, structure and physiological functions of plants. Because of concerns regarding the potential environmental risk of transgenic crops, it is important to monitor changes in transgenic insect-resistant rice under the projected high tropospheric ozone before its commercial release. Using a free-air concentration enrichment (FACE) system, we investigated the changes in leaf morphology and leaf ultrastructure of two rice varieties grown in plastic pots, transgenic Bt Shanyou 63 (Bt-SY63, carrying a fusion gene of cry1Ab and cry1Ac) and its non-transgenic counterpart (SY63), in elevated O3 (E-O3) versus ambient O3 (A-O3) after 64-DAS (Days after seeding), 85-DAS and 102-DAS. Our results indicated that E-O3 had no significant effects on leaf length, leaf width, leaf area, stomatal length and stomatal density for both Bt-SY63 and SY63. E-O3 increased the leaf thickness of Bt-SY63, but decreased that of SY63. O3 stress caused early swelling of the thylakoids of chloroplasts, a significant increase in the proportion of total plastoglobule area in the entire cell area (PCAP) and a significant decrease in the proportion of total starch grain area in the entire cell area (SCAP), suggesting that E-O3 accelerated the leaf senescence of the two rice genotypes. Compared with SY63, E-O3 caused early swelling of the thylakoids of chloroplasts and more substantial breakdown of chloroplasts in Bt-SY63. Our results suggest that the incorporation of cry1Ab/Ac into SY63 could induce unintentional changes in some parts of plant morphology and that O3 stress results in greater leaf damage to Bt-SY63 than to SY63, with the former coupled with higher O3 sensitivity in CCAP (the proportions of total chloroplast area in the entire cell area), PCAP and SCAP. This study provides valuable baseline information for the prospective commercial release of transgenic crops under the projected future climate.

  15. Leaf morphology and ultrastructure responses to elevated O3 in transgenic Bt (cry1Ab/cry1Ac rice and conventional rice under fully open-air field conditions.

    Directory of Open Access Journals (Sweden)

    Chunyan Li

    Full Text Available BACKGROUND: Elevated tropospheric ozone severely affects not only yield but also the morphology, structure and physiological functions of plants. Because of concerns regarding the potential environmental risk of transgenic crops, it is important to monitor changes in transgenic insect-resistant rice under the projected high tropospheric ozone before its commercial release. METHODOLOGY/PRINCIPAL FINDINGS: Using a free-air concentration enrichment (FACE system, we investigated the changes in leaf morphology and leaf ultrastructure of two rice varieties grown in plastic pots, transgenic Bt Shanyou 63 (Bt-SY63, carrying a fusion gene of cry1Ab and cry1Ac and its non-transgenic counterpart (SY63, in elevated O3 (E-O3 versus ambient O3 (A-O3 after 64-DAS (Days after seeding, 85-DAS and 102-DAS. Our results indicated that E-O3 had no significant effects on leaf length, leaf width, leaf area, stomatal length and stomatal density for both Bt-SY63 and SY63. E-O3 increased the leaf thickness of Bt-SY63, but decreased that of SY63. O3 stress caused early swelling of the thylakoids of chloroplasts, a significant increase in the proportion of total plastoglobule area in the entire cell area (PCAP and a significant decrease in the proportion of total starch grain area in the entire cell area (SCAP, suggesting that E-O3 accelerated the leaf senescence of the two rice genotypes. Compared with SY63, E-O3 caused early swelling of the thylakoids of chloroplasts and more substantial breakdown of chloroplasts in Bt-SY63. CONCLUSIONS/SIGNIFICANCE: Our results suggest that the incorporation of cry1Ab/Ac into SY63 could induce unintentional changes in some parts of plant morphology and that O3 stress results in greater leaf damage to Bt-SY63 than to SY63, with the former coupled with higher O3 sensitivity in CCAP (the proportions of total chloroplast area in the entire cell area, PCAP and SCAP. This study provides valuable baseline information for the prospective

  16. Micro-Etched Platforms for Thermal Inactivation of Bacillus Anthracis and Bacillus Thuringiensis Spores

    Science.gov (United States)

    2008-03-01

    gentlemen are approachable and extremely well versed in their fields. Dr. Burggraf’s passion for learning coupled with the patience necessary to allow...wide spread employment on civilian targets has increased. Indeed, the well known biological agent Anthrax, Bacillus anthracis (B.a.), was recently...reached at 0.5 seconds with the heat from the fireball lasting a total of 4 seconds ( Orson , J.A. 2003; 104). From this data, the short time-temperature

  17. Structural studies of δ-endotoxin Cry 1 C from Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Guimaraes, B.G.; Garratt, R.C.; Oliva, G.; Lemos, M.V.F.; Arantes, O.M.N.

    1996-01-01

    Full text. The δ-endotoxins are a family of crystal protein by a soil bacterium, Bacillus thuringiensis. The study of these proteins has been of great interest due to their highly specific activity against insects of the orders Lepidoptera, Diptera and Coleoptera. Thus, the δa-endotoxins have been used for more than two decades as biological insecticides to control agricultural pests and, more recently, insects vectors of some diseases. The knowledge of their three-dimensional structures is very important to understand their mechanism of action and their high specificity. To date, the structure of only three proteins of the δ-endotoxins family have been reported: Cry3A, a coleopteran-specific toxin (beetle toxin) 1 , Cry1Aa, a lepidopteran-specific toxin (butterfly toxin) 2 and CytB, a dipteran-specific toxin (mosquito toxin) 3 Our work is aimed at the determination of the crystallographic structure by X-ray diffraction of δ-endotoxin Cry1C, also toxic to insects of the Lepidoptera order but towards families other than those affected by Cry1Aa. A comparison between these structures may lead to important conclusions about the reasons for the specificity and would allow the planning of mutants with more efficient activity. The cry1C gene was cloned into an adequate vector and expressed in an acrystalliferous B. thuringiensis strain. After cell culture and sporulation the microcrystals of Cry1C were separated by ultra-centrifugation in sacharose. The protoxin inclusion bodies were activated by commercial trpsin and the protease-resistant core was purified by anion-exchange chromatography. Crystallization experiments are being conducted in order to obtain single crystals suitable for diffraction measurements. We intend to use the Protein Crystallograph Station of the LNLS to collect data as soon as it is available and we have suitable crystals. (author)

  18. Identification and Characterization of Three Previously Undescribed Crystal Proteins from Bacillus thuringiensis subsp. jegathesan

    Science.gov (United States)

    Sun, Yunjun; Zhao, Qiang; Ding, Xuezhi; Hu, Quanfang; Federici, Brian A.

    2013-01-01

    The total protoxin complement in the parasporal body of mosquitocidal strain, Bacillus thuringiensis subsp. jegathesan 367, was determined by use of a polyacrylamide gel block coupled to mass spectrometry. A total of eight protoxins were identified from this strain, including five reported protoxins (Cry11Ba, Cry19Aa, Cry24Aa, Cry25Aa, and Cyt2Bb), as well as three previously undescribed (Cry30Ca, Cry60Aa, and Cry60Ba) in this isolate. It was interesting that the encoding genes of three new protoxins existed as cry30Ca-gap-orf2 and cry60Ba-gap-cry60Aa. The cry30Ca and a downstream orf2 gene were oriented in the same direction and separated by 114 bp, and cry60Ba was located 156 bp upstream from and in the same orientation to cry60Aa. The three new protoxin genes were cloned from B. thuringiensis subsp. jegathesan and expressed in an acrystalliferous strain under the control of cyt1A gene promoters and the STAB-SD stabilizer sequence. Recombinant strain containing only cry30Ca did not produce visible inclusion under microscope observation, while that containing both cry30Ca and orf2 could produce large inclusions. Cry60Aa and Cry60Ba synthesized either alone or together in the acrystalliferous host could yield large inclusions. In bioassays using the fourth-instar larvae of Culex quinquefasciatus, Cry60Aa and Cry60Ba alone or together had estimated 50% lethal concentrations of 2.9 to 7.9 μg/ml; however, Cry30Ca with or without ORF2 was not toxic to this mosquito. PMID:23524673

  19. Potato flour mediated solid-state fermentation for the enhanced production of Bacillus thuringiensis-toxin.

    Science.gov (United States)

    Smitha, Robinson Babysarojam; Jisha, Veloorvalappil Narayanan; Pradeep, Selvanesan; Josh, Moolakkariyil Sarath; Benjamin, Sailas

    2013-11-01

    In this study, we explored the efficacy of raw potato flour (PF) as supplement to the conventional LB medium (LB control, designated as M1) for enhancing the concomitant production of endospores and δ-endotoxin from Bacillus thuringiensis subsp. kurstaki by solid-state fermentation (SSF). Of different concentrations and combinations of media tested, 10% (w/v) PF supplemented LB medium (M2) was found as the best source for the maximum yield of toxin. After 12 h submerged fermentation (SmF) at 37°C and 125 rpm, M2 was made into a wet-solid matter for SSF by removing the supernatant (1000 ×g, 10 min); the resultant pellet subsequently incubated statically (37°C) for the production of B. thuringiensis subsp. kurstaki toxin (Btk-toxin). In comparison to M1, yield of δ-endotoxin purified by sucrose density gradient centrifugation method from M2 was about 6-fold higher (53% recovery). This maximum yield from M2 was obtained at 48 h (as against 72 h from M1), thus the gestation period of M2 was reduced by 24 h with higher yield. In addition to the quantitative data, qualitative photomicrographs taken by image analyzer, scanning electron and fluorescent microscopes and digital camera showed physical evidences for the upper hand of SSF over conventional SmF for the enhanced production of Btk-toxin. SDS-PAGE image of the purified δ-endotoxin showed three major fractions with apparent MWs 66, 45 and 30 kDa. Briefly, if low-cost agricultural products like PF is used as supplement to LB, by SSF strategy, production of Btk-toxin could be enhanced to 6-fold in short gestation time without losing its entomotoxicity efficiency. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  20. Gamma Radiation to Increase Efficiency of Bacillus thuringiensis Thai Strain for Insect Pets Control

    International Nuclear Information System (INIS)

    Chanpaisaeng, Jariya; Keawsompong, Suttipun; Piadang, Nattaya; Tephan, Prakai; Keawchingduang, Wannapa

    2006-09-01

    Bacillus thuringiensis (Bt) isolates JCPT16 and JCPT68 were gamma-irradiated at 2, 4, 6 and 8 kGy. The efficiency of these Bt isolates on S. litura control was also undertaken. It was found that the 4 kGy irradiated JCPT16 isolate had lowest LC 50 of 6.6x10 3 spore/ml while the non-irradiated JCPT 16 isolate had LC 50 of 6.2x10 3 spore/ml. Whereas the irradiated JCPT68 isolate at 8 kGy was noticed to have the lowest LC 50 of 2.7 x 10 3 spores/ml, the non-irradiated JCPT68 had LC 50 of 1.8x10 3 spores/ml. The efficiency test of B. thuringiensis isolate on S. exigua showed that the 2 kGy irradiated JCPT16 isolate had the lowest LC 50 of 2.52x10 4 spores/ml while the non-irradiated JCPT16 isolate had LC 50 of 6.04x10 3 spores/ml. The irradiated JCPT68 isolate at 4 kGy had the lowest LC 50 of 5.41x10 4 spores/ml, the non irradiated JCPT68 had LC 50 of 1.51x10 4 spores/ml. According to LC 50 values, there were no significant differences of efficiency on S. litura and S. exigua control among Bt isolates irradiated at various concentrations. The isolate JCPT16, JCPT35, JCPT50 and JCPT68 irradiated at dose of 10 kGy showed higher UV tolerance. After expose by UV ray, most of irradiated isolates still displayed high efficiency of controlling S. litura, S. exigua and Plutella xylostell.

  1. Long lasting persistence of Bacillus thuringiensis Subsp. israelensis (Bti in mosquito natural habitats.

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    Mathieu Tilquin

    Full Text Available BACKGROUND: The detrimental effects of chemical insecticides on the environment and human health have lead to the call for biological alternatives. Today, one of the most promising solutions is the use of spray formulations based on Bacillus thuringiensis subsp. israelensis (Bti in insect control programs. As a result, the amounts of Bti spread in the environment are expected to increase worldwide, whilst the common belief that commercial Bti is easily cleared from the ecosystem has not yet been clearly established. METHODOLOGY/MAIN FINDINGS: In this study, we aimed to determine the nature and origin of the high toxicity toward mosquito larvae found in decaying leaf litter collected in several natural mosquito breeding sites in the Rhône-Alpes region. From the toxic fraction of the leaf litter, we isolated B. cereus-like bacteria that were further characterized as B. thuringiensis subsp. israelensis using PCR amplification of specific toxin genes. Immunological analysis of these Bti strains showed that they belong to the H14 group. We finally used amplified length polymorphism (AFLP markers to show that the strains isolated from the leaf litter were closely related to those present in the commercial insecticide used for field application, and differed from natural worldwide genotypes. CONCLUSIONS/SIGNIFICANCE: Our results raise the issue of the persistence, potential proliferation and environmental accumulation of human-spread Bti in natural mosquito habitats. Such Bti environmental persistence may lengthen the exposure time of insects to this bio-insecticide, thereby increasing the risk of resistance acquisition in target insects, and of a negative impact on non-target insects.

  2. Molecular characterization and genetic diversity of insecticidal crystal protein genes in native Bacillus thuringiensis isolates.

    Science.gov (United States)

    Mahadeva Swamy, H M; Asokan, R; Mahmood, Riaz; Nagesha, S N

    2013-04-01

    The Western Ghats of Karnataka natural ecosystem are among the most diverse and is one of the eight hottest hotspots of biological diversity in the world, that runs along the western part of India through four states including Karnataka. Bacillus thuringiensis (Bt) strains were isolated from soils of Western Ghats of Karnataka and characterized by molecular and analytical methods as a result of which 28 new Bt-like isolates were identified. Bt strains were isolated from soil samples using sodium acetate selection method. The morphology of crystals was studied using light and phase contrast microscopy. Isolates were further characterized for insecticidal cry gene by PCR, composition of toxins in bacterial crystals by SDS-PAGE cloning, sequencing and evaluation of toxicity was done. As a result 28 new Bt-like isolates were identified. Majority of the isolates showed the presence of a 55 kDa protein bands on SDS-PAGE while the rest showed 130, 73, 34, and 25 kDa bands. PCR analysis revealed predominance of Coleopteran-active cry genes in these isolates. The variations in the nucleotide sequences, crystal morphology, and mass of crystal protein(s) purified from the Bt isolates revealed genetic and molecular diversity. Three strains containing Coleopteran-active cry genes showed higher activity against larvae Myllocerus undecimpustulatus undatus Marshall (Coleoptera: Curculionidae) than B. thuringiensis subsp. Morrisoni. Results indicated that Bt isolates could be utilized for bioinsecticide production, aiming to reduce the use of chemical insecticide which could be useful to use in integrated pest management to control agriculturally important pests for sustainable crop production.

  3. Biochemical, immunological and toxicological characteristics of the crystal proteins of Bacillus thuringiensis subsp. medellin

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    Sergio Orduz

    1996-04-01

    Full Text Available Characterization of the insecticidal and hemolytic activity of solubilized crystal proteins of Bacillus thuringiensis (Bt subsp. medellin (Btmed was performed and compared to solubilized crystal proteins of isolates 1884 of B. thuringiensis subsp. israelensis (Bti and isolate PG-14 of B. thuringiensis subsp. morrisoni (Btm. In general, at acid pH values solubilization of the Bt crystalline parasporal inclusions (CPI was lower than at alkaline pH. The larvicidal activity demonstrated by the CPI of Btmed indicated that optimal solubilization of CPI takes place at a pH value of 11.3, in Bti at pH values from 5.03 to 11.3 and in Btm at pH values from 9.05 to 11.3. Hemolytic activity against sheep red blood cells was mainly found following extraction at pH 11.3 in all Bt strains tested. Polyacrylamide gel electrophoresis under denaturing conditions revealed that optimal solubilization of the CPI in all Bt strains takes place at the alkaline pH values from 9.05 to 11.3. An enriched preparation of Btmed crystals was obtained, solubilized and crystal proteins were separated on a size exclusion column (Sephacryl S-200. Three main protein peaks were observed on the chromatogram. The first peak had two main proteins that migrate between 90 to 100 kDa. These proteins are apparently not common to other Bt strains isolated to date. The second and third peaks obtained from the size exclusion column yielded polypeptides of 68 and 28-30 kDa, respectively. Each peak independently, showed toxicity against 1st instar Culex quinquefasciatus larvae. Interestingly, combinations of the fractions corresponding to the 68 and 30 kDa protein showed an increased toxicity. These results suggest that the 94 kDa protein is an important component of the Btmed toxins with the highest potency to kill mosquito larvae. When crystal proteins of Bti were probed with antisera raised independently against the three main protein fractions of Btmed, the only crystal protein that showed

  4. Characterization, N-terminal sequencing and classification of Tolworthcin 524: A bacteriocin produced by Bacillus thuringiensis subsp. tolworthi.

    Science.gov (United States)

    Pacheco-Cano, Rubén D; de la Fuente-Salcido, Norma M; Salcedo-Hernández, Rubén; León-Galván, M Fabiola; Bideshi, Dennis K; Hernández-Guzmán, Gustavo; Barboza-Corona, J Eleazar

    2014-12-01

    Bacteriocins synthesized by entomopathogenic Bacillus thuringiensis are gaining attention owing to their inhibitory effects against a wide variety of pathogenic bacteria. In the present study, we purified and characterized Tolworthcin 524, a bacteriocin synthesized by B. thuringiensis subsp. tolworthi, and compared it with other bacteriocins synthesized by B. thuringiensis. Tolworthcin 524 was separated and purified from the secretome of B. thuringiensis by fast protein liquid chromatography with a gel filtration column to obtain yields of 17% and a specific activity of ∼3600U/mgprotein. The purified product showed two peptides of ∼9 and 6kDa with antimicrobial activity in a gel-screening assay. The purified product was analyzed by two-dimensional electrophoresis and the resolved peptides of ∼9 and 6kDa with isoelectric points of ∼8 were sequenced. Partial sequences (METPVVQPR and DWTCWSCLVCAACS) were obtained suggesting that the ∼9 and 6kDa correspond to the prebacteriocin and mature Tolworthcin 524, respectively. Sequences showed high identity with Thurincin H and Thuricin 17 and had a conserved motif with other bacteriocins of B. thuringiensis. Based on sequence data, Tolworthcin 524 was classified in subclass II.2 (Thuricin-like peptides) of the Bacillus bacteriocin classification scheme. The larger peptide did not harbor a sequence suggestive of a signal peptide neither did it contain the double-glycine (GG) motif characteristic of the secretion leader recognized by the ABC transport system. Implications of these properties in Tolworthcin 524 secretion are discussed. Copyright © 2014 Elsevier GmbH. All rights reserved.

  5. Detection of β-exotoxin synthesis in Bacillus thuringiensis using an easy bioassay with the nematode Caenorhabditis elegans.

    Science.gov (United States)

    Sánchez-Soto, A I; Saavedra-González, G I; Ibarra, J E; Salcedo-Hernández, R; Barboza-Corona, J E; Del Rincón-Castro, M C

    2015-12-01

    The insecticidal activity of Bacillus thuringiensis is owing to the action of Cry and Cyt proteins. In addition to the synthesis of insecticidal proteins, some strains are able to synthesize β-exotoxin, which is highly toxic to humans. In this regard, it is very important to have a simple method to detect β-exotoxin to avoid the commercial production of this type of strains. In this work, we developed a simple and fast method, using the nematode Caenorhabditis elegans to detect indirectly the synthesis of β-exotoxin by B. thuringiensis strain. Using this assay, we detected that ~60% of Mexican native strains (i.e. LBIT-471, 491, 492, 497, 507, 511, 515, 536 and 537) were toxic to the nematode (44-97% mortalities) and their β-exotoxin (βEx(+) ) production, including a positive control (NRD-12), was confirmed by HPLC. In addition, the negative controls (βEx(-) ) LBIT-436 (HD-1) and LBIT-438 and also the native strains LBIT-499, 500, 521, 522, 533 and 542, did not show a detrimental effect against nematodes larvae, neither the synthesis of β-exotoxin as determined by HPLC. Finally, we did not find a correlation between B. thuringiensis strains with similar plasmid patterns and the β-exotoxin production. In this work, we implemented a qualitative and fast bioassay using the nematode Caenorhabditis elegans to detect the production of β-exotoxin in different strains of Bacillus thuringiensis. We show that this assay is useful to detect β-exotoxin in B. thuringiensis with high reliability, helping to discriminate strains that could not be used as bioinsecticides because of their putative risk to humans. Data show that qualitative bioassay with nematodes is a potential alternative to fly larvae bioassays, and correlated with the determination of β-exotoxin by HPLC. © 2015 The Society for Applied Microbiology.

  6. Bacillus thuringiensis in caterpillars and associated materials collected from protected tropical forests in northwestern Costa Rica

    Directory of Open Access Journals (Sweden)

    César Rodríguez-Sánchez

    2006-06-01

    Full Text Available Bacillus thuringiensis (Bt synthesizes crystalline inclusions that are toxic to caterpillars (Lepidoptera and other orders of invertebrates. Materials associated with 37 caterpillars from 16 species, collected while feeding on 15 different species of host plants in dry, cloud and rain forests located in the Área de Conservación Guanacaste in northwestern Costa Rica, were examined for the presence of Bt. From a total of 101 derived samples, 25 Bt isolates were cultured: 56% from host plant leaves, 8% from caterpillar guts and 36% from caterpillar fecal pellets. Bt was isolated from at least one sample in 38% of the systems constituted by the food plant, gut and fecal pellets corresponding to a single caterpillar. Four different morphologies of crystalline inclusions were observed, with bipyramidal and irregular crystal morphologies being the most prevalent. Rev. Biol. Trop. 54(2: 265-271. Epub 2006 Jun 01.Bacillus thuringiensis (Bt sintetiza inclusiones cristalinas que resultan tóxicas para algunas larvas de lepidópteros y otros órdenes de invertebrados. Su presencia fue examinada en materiales asociados a 37 orugas de mariposas de 16 especies, las cuales fueron colectadas mientras se alimentaban en 15 especies diferentes de plantas hospederas en bosques secos, nubosos y húmedos localizados dentro del Área de Conservación Guanacaste (ACG en el noroeste de Costa Rica. A partir de un total de 101 muestras se obtuvo 25 aislamientos de Bt: 56% a partir de material foliar de las plantas hospederas, 8% a partir del contenido intestinal de las larvas y 36% a partir de sus excrementos. Esta bacteria fue cultivada a partir de al menos uno de los 3 diferentes tipos de muestra asociados a una oruga particular (planta hospedera, intestino, excremento en 38% de los casos posibles. En la colección de aislamientos obtenida se observaron cuatro morfologías de inclusiones cristalinas, siendo aquellas bipiramidales e irregulares las más prevalentes.

  7. Susceptibility of The Asian Corn Borer, Ostrinia furnacalis, to Bacillus thuringiensis Toxin CRY1AC

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    Aye Kyawt Kyawt Ei

    2008-07-01

    Full Text Available The larval susceptibility of the Asian corn borer, Ostrinia furnacalis (Guenee (Lepidoptera: Crambidae, to a Bacillus thuringiensis protein (Cry1Ac was evaluated using insect feeding bioassays. The founding population of O. furnacalis was originally collected from the experimental station of UGM at Kalitirto and had been reared in the laboratory for three generations using an artificial diet “InsectaLf”. The tested instars were exposed on diets treated with a series of concentrations of Cry1Ac for one week. The LC50 values on the seventh day after treatment for 1st, 2nd, 3rd and 4th instars were 7.79, 21.12, 113.66, and 123.17 ppm, respectively, showing that the higher the instars the lesser the susceptibility to Cry1Ac. When the neonates were exposed to sublethal concentrations of Cry1Ac (0.0583, 0.116, and 0.5830 ppm, growth and development of the surviving larvae were inhibited. The fecundity and viability of females produced from treated larvae decreased with increasing the concentrations. These findings indicate that Cry1Ac is toxic to larva of O. furnacalis and has chronic effects to larvae surviving from Cry1Ac ingestion.   Kepekaan larva penggerek batang jagung Asia, Ostrinia furnacalis (Guenee (Lepidoptera: Crambidae, terhadap protein Bacillus thuringiensis Cry1Ac diuji dengan metode celup pakan. Larva berasal dari pertanaman jagung di KP-4, UGM di Kalitirto dan telah dikembangbiakkan di laboratorium menggunakan pakan buatan (InsectaLF selama tiga generasi sebelum digunakan untuk pengujian. Larva O. furnacalis yang diuji dipaparkan pada pakan buatan yang telah dicelupkan pada seri konsentrasi Cry1Ac. Nilai LC50 pada hari ketujuh setelah perlakukan untuk instar 1, 2, 3, dan 4 berturut-turut adalah 0,79; 21,12; 113,66; dan 123,17 ppm. Hal ini menunjukkan bahwa instar yang semakin tinggi tingkat kepekaannya terhadap Cry1Ac semakin menurun. Larva yang baru menetas dan diberi pakan yang telah dicelupkan pada konsentrasi sublethal Cry1Ac

  8. Characterization of Tunisian Bacillus thuringiensis strains with abundance of kurstaki subspecies harbouring insecticidal activities against the lepidopteran insect Ephestia kuehniella.

    Science.gov (United States)

    Saadaoui, Imen; Al-Thani, Roda; Al-Saadi, Fatma; Belguith-Ben Hassan, Najeh; Abdelkefi-Mesrati, Lobna; Schultz, Patrick; Rouis, Souad; Jaoua, Samir

    2010-12-01

    The study of 257 crystal-producing Bacillus thuringiensis isolates from bioinsecticide free soil samples collected from different sites in Tunisia, was performed by PCR amplification, using six primer pairs specific for cry1, cry2, cry3, cry4, and vip3A genes, by the investigation of strain plasmid pattern, crystal morphology and delta-endotoxin content and by the assessment of insecticidal activities against the lepidopteran insect Ephestia kuehniella. Based on plasmid pattern study, 11 representative strains of the different classes were subjected to morphological and molecular analyses. The comparison of the PFGE fingerprints confirmed the heterogeneity of these strains. B. thuringiensis kurstaki strains, harbouring at the same time the genes cry1A, cry2, cry1Ia, and vip3A, were the most abundant (65.4%). 33.34% of the new isolates showed particular delta-endotoxin profiles but no PCR products with the used primer sets. B. thuringiensis israelensis was shown to be also very rare among the Tunisian B. thuringiensis isolates diversity. These findings could have considerable impacts for the set up of new pest control biological agents.

  9. Larvicidal efficacy of Catharanthus roseus Linn. (Family: Apocynaceae) leaf extract and bacterial insecticide Bacillus thuringiensis against Anopheles stephensi Liston.

    Science.gov (United States)

    Panneerselvam, Chellasamy; Murugan, Kadarkarai; Kovendan, Kalimuthu; Kumar, Palanisamy Mahesh; Ponarulselvam, Sekar; Amerasan, Duraisamy; Subramaniam, Jayapal; Hwang, Jiang-Shiou

    2013-11-01

    To explore the larvicidal activity of Catharanthus roseus (C. roseus) leaf extract and Bacillus thuringiensis (B. thuringiensis) against the malarial vector Anopheles stephensi (An. stephensi), when being used alone or together. The larvicidal activity was assayed at various concentrations under the laboratory and field conditions. The LC50 and LC90 values of the C. roseus leaf extract were determined by probit analysis. The plant extract showed larvicidal effects after 24 h of exposure; however, the highest larval mortality was found in the petroleum ether extract of C. roseus against the first to fourth instars larvae with LC50=3.34, 4.48, 5.90 and 8.17 g/L, respectively; B. thuringiensis against the first to fourth instars larvae with LC50=1.72, 1.93, 2.17 and 2.42 g/L, respectively; and the combined treatment with LC50=2.18, 2.41, 2.76 and 3.22 g/L, respectively. No mortality was observed in the control. The petroleum ether extract of C. roseus extract and B. thuringiensis have potential to be used as ideal eco-friendly agents for the control of An. stephensi in vector control programs. The combined treatment with this plant crude extract and bacterial toxin has better larvicidal efficacy against An. stephensi. Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  10. [Bacillus thuringiensis: general aspects. An approach to its use in the biological control of lepidopteran insects behaving as agricultural pests].

    Science.gov (United States)

    Sauka, Diego H; Benintende, Graciela B

    2008-01-01

    Bacillus thuringiensis is the most widely applied biological pesticide used to control insects that affect agriculture and forestry and which transmit human and animal pathogens. During the past decades B. thuringiensis has been the subject of intensive research. These efforts have yielded considerable data about the relationships between the structure, mechanism of action, and genetics of their pesticidal crystal proteins. As a result, a coherent picture of these relationships has emerged. Other studies have focused on the ecological role of the B. thuringiensis crystal proteins and their performance in agricultural and other natural settings. With this knowledge as background and the help of biotechnological tools, researchers are now reporting promising results in the development of more useful toxins, recombinant bacteria, new formulations and transgenic plants that express pesticidal activity, in order to assure that these products are utilized with the best efficiency and benefit. This article is an attempt to integrate all these recent developments in the study of B. thuringiensis into a context of biological control of lepidopteran insect pest of agricultural importance.

  11. Avaliação de produtos à base de Bacillus thuringiensis no controle da traça-das-crucíferas Evaluation of insecticides based on Bacillus thuringiensis in the control of the diamondback moth

    Directory of Open Access Journals (Sweden)

    Patrícia T Medeiros

    2006-06-01

    Full Text Available Avaliou-se em dois experimentos a suscetibilidade da traça-das-crucíferas a inseticidas à base de Bacillus thuringiensis em repolho cv. Itiban. O delineamento do primeiro experimento (de julho a setembro/03, em área de plantio comercial em Brazlândia (DF, foi de blocos casualizados, com seis tratamentos e dez repetições; os bioinseticidas utilizados foram B. thuringiensis kurstaki (S1450CO, B. thuringiensis aizawai comercial (Bta e três produtos formulados com as estirpes S1450BB, S811BB, S845BB de B. thuringiensis pertencentes ao Banco de Bacillus spp. da Embrapa Recursos Genéticos e Biotecnologia. No segundo experimento, realizado no campo experimental da Embrapa (DF, de outubro/03 a janeiro/04, o delineamento foi de blocos casualizados, com seis tratamentos e quatro repetições; os inseticidas utilizados foram os mesmos do primeiro experimento, com a adição de Spinosad, e retirado o tratamento S811BB. Os produtos foram aplicados quando foi atingido o nível de dano de seis furos nas quatro folhas centrais do repolho. O Bta comercial foi o produto mais eficaz no primeiro experimento, tendo sido aplicado cinco vezes e diferiu estaticamente dos demais produtos. Os formulados S845BB e S1450BB não apresentaram diferenças quando comparados ao produto comercial S1450 e foram aplicados seis vezes. O produto S811BB também foi aplicado seis vezes, mas sua eficácia foi inferior aos demais produtos e não diferiu da testemunha. Já no segundo experimento, o S1450 comercial foi aplicado cinco vezes e os demais produtos à base de Bt, seis vezes. Todos os produtos utilizados não diferiram entre si, diferindo apenas no número de aplicações.Two experiments were performed to evaluate the susceptibility of the diamondback moth to insecticides based on Bacillus thuringiensis in cabbage cv. Itiban. The first experiment was carried out from July to September 2003, in a production area in Brazlândia (DF, Brazil. Randomized blocks with six

  12. Elucidation of the mechanisms of CryIIIA overproduction in a mutagenized strain of Bacillus thuringiensis var. tenebrionis

    International Nuclear Information System (INIS)

    Adams, L.F.; Mathewes, S.; O'Hara, P.; Peterson, A.; Gürtler, H.

    1994-01-01

    NB176 is a Bacillus thuringiensis mutant derived by λ-irradiation of NB125 Bacillus thuringiensis var. tenebrionis (Krieg). It exhibits two interesting phenotypes: (i) oligosporogeny and (ii) twofold to threefold overproduction of the CryIIIA protein. Southern profiles of the NB176 strain showed an additional copy(s) of the cryIIIA gene located on a 4 kb HindIII fragment, in addition to the expected cryIIIA gene on a 3 kb HindIII fragment. Each cryIIIA gene-bearing HindIII fragment was cloned from NB176. The restriction map of the 3 kb HindIII fragment was identical to that published by Donovan and coworkers. Sequencing of the 4 kb HindIII fragment showed no alterations in the promoter region of the cryIIIA gene but did show replacement of the region immediately following the cryIIIA open reading frame with a sequence encoding a transposase with 50% amino acid homology to that of Tn 1000. These findings suggest that the overproduction phenotype of NB176 results from extra copies of the cryIIIA gene produced from a transposition event(s) induced or stabilized by γ-irradiation. Integration of additional copies of the cryIIIA gene into the native 90MDa plasmid of the wild-type B. thuringiensis var. tenebrionis strain resulted in strains that made enormous crystals, many possessing greatly enhanced insecticidal activity

  13. The Synergist Effect of The Henna Plant, Lawsonia alba on Bacillus thuringiensis var. kurstaki Against Third Larval Instar of Pistachio Leaf Borer, Ocneria terebinthina Strg. (Lep.: Lymanteriidae)

    OpenAIRE

    Zahra Sheibani

    2017-01-01

    Ocneria terebinthina Strg. (Lep.: Lymanteriidae) is a leaf borer pest in pistachio orchard. The toxicity of Bacillus thuringiensis var. kurstaki alone and in combination with henna powder was investigated on third larval instar of O. terebinthina under laboratory conditions. Bioassay was carried out using spray technique on pistachio offshoot. Probit analysis of concentration-mortality data was conducted to estimate the LC50 value. The LC50 value of B. thuringiensis on third instar of larvae ...

  14. Cellular responses in Bacillus thuringiensis CS33 during bacteriophage BtCS33 infection.

    Science.gov (United States)

    Wu, Dandan; Yuan, Yihui; Liu, Pengming; Wu, Yan; Gao, Meiying

    2014-04-14

    Bacillus thuringiensis (Bt) has been widely used for 50years as a biopesticide for controlling insect pests. However, bacteriophage infection can cause failures in 50%-80% of the batches during Bt fermentation, resulting in severe losses. In the present work, the physiological and biochemical impacts of Bt strain CS33 have been studied during bacteriophage infection. This study adopted a gel-based proteomics approach to probe the sequential changed proteins in phage-infected Bt cells. To phage, it depressed the host energy metabolism by suppressing the respiration chain, the TCA cycle, and the utilization of PHB on one hand; on the other hand, it hijacked the host translational machine for its own macromolecular synthesis. To host, superinfection exclusion might be triggered by the changes of S-layer protein and flagella related proteins, which were located on the cell surface and might play as the candidates for the phage recognition. More importantly, the growth rate, cell mass, and ICPs yield were significantly decreased. The low yield of ICPs was mainly due to the suppressed utilization of PHB granules. Further functional study on these altered proteins may lead to a better understanding of the pathogenic mechanisms and the identification of new targets for phage control. B. thuringiensis (Bt) has been widely used for 50years as a safe biopesticide for controlling agricultural and sanitary insect pests. However, bacteriophage infection can cause severe losses during B. thuringiensis fermentation. The processes and consequences of interactions between bacteriophage and Bt were still poorly understood, and the molecular mechanisms involved were more unknown. This study adopted a gel-based proteomics approach to probe the physiological and biochemical impacts of Bt strain CS33 after phage-infection. The interactions between phage BtCS33 and its host Bt strain CS33 occurred mainly on four aspects. First, phage synthesized its nucleic acids through metabolic

  15. Hot, humid air decontamination of a C-130 aircraft contaminated with spores of two acrystalliferous Bacillus thuringiensis strains, surrogates for Bacillus anthracis.

    Science.gov (United States)

    Buhr, T L; Young, A A; Bensman, M; Minter, Z A; Kennihan, N L; Johnson, C A; Bohmke, M D; Borgers-Klonkowski, E; Osborn, E B; Avila, S D; Theys, A M G; Jackson, P J

    2016-04-01

    To develop test methods and evaluate survival of Bacillus thuringiensis kurstaki cry(-) HD-1 and B. thuringiensis Al Hakam spores after exposure to hot, humid air inside of a C-130 aircraft. Bacillus thuringiensis spores were either pre-inoculated on 1 × 2 or 2 × 2 cm substrates or aerosolized inside the cargo hold of a C-130 and allowed to dry. Dirty, complex surfaces (10 × 10 cm) swabbed after spore dispersal showed a deposition of 8-10 log10 m(-2) through the entire cargo hold. After hot, humid air decontamination at 75-80°C, 70-90% relative humidity for 7 days, 87 of 98 test swabs covering 0·98 m(2) , showed complete spore inactivation. There was a total of 1·67 log10 live CFU detected in 11 of the test swabs. Spore inactivation in the 98 test swabs was measured at 7·06 log10 m(-2) . Laboratory test methods for hot, humid air decontamination were scaled for a large-scale aircraft field test. The C-130 field test demonstrated that hot, humid air can be successfully used to decontaminate an aircraft. Transition of a new technology from research and development to acquisition at a Technology Readiness Level 7 is unprecedented. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

  16. Transgenic organisms expressing genes from Bacillus thuringiensis to combat insect pests.

    Science.gov (United States)

    Zaritsky, Arieh; Ben-Dov, Eitan; Borovsky, Dov; Boussiba, Sammy; Einav, Monica; Gindin, Galina; Horowitz, A Rami; Kolot, Mikhail; Melnikov, Olga; Mendel, Zvi; Yagil, Ezra

    2010-01-01

    Various subspecies (ssp.) of Bacillus thuringiensis (Bt) are considered the best agents known so far to control insects, being highly specific and safe, easily mass produced and with long shelf life.1 The para-crystalline body that is produced during sporulation in the exosporium includes polypeptides named δ-endotoxins, each killing a specific set of insects. The different entomopathogenic toxins of various Bt ssp. can be manipulated genetically in an educated way to construct more efficient transgenic bacteria or plants that express combinations of toxin genes to control pests.2 Joint research projects in our respective laboratories during the last decade demonstrate what can be done by implementing certain ideas using molecular biology with Bt ssp. israelensis (Bti) as a model system. Here, we describe our progress achieved with Gram-negative bacterial species, including cyanobacteria, and some preliminary experiments to form transgenic plants, mainly to control mosquitoes (Diptera), but also a particular Lepidopteran and Coleopteran pest species. In addition, a system is described by which environment-damaging genes can be removed from the recombinants thus alleviating procedures for obtaining permits to release them in nature. © 2010 Landes Bioscience

  17. Bacillus thuringiensis isolates entomopathogenic for Culex quinquefasciatus (Diptera: Culicidae and Anticarsia gemmatalis (Lepidoptera: Noctuidae

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    V. Gobatto

    Full Text Available Samples of the Bacillus thuringiensis (Bt were collected from soil and insects. Eight isolates were selected from rural soil, 15 from urban soil and 11 from insects. These were evaluated for entomopathogenicity against larvae of Anticarsia gemmatalis and Culex quinquefasciatus. The pathogenicity tests showed that a higher percentage of isolates were active against A. gemmatalis (60% compared to C. quinquefasciatus (31%. Probit analysis (LC50 indicated that against A. gemmatalis four of the isolates presented values similar to the reference strain against A. gemmatalis, while against C. quinquefasciatus one isolate showed an LC50 similar to the reference strain (IPS-82. SDS-PAGE characterisation of two isolates showed a 27 kDa protein fraction related to the Bt subspecies israelensis cytolytic toxin (cyt gene. One 130 kDa protein, possibly related to the Bt crystal inclusions (cry1 gene, was identified in the other two isolates, which were more toxic for lepidoptera; another isolate presented a protein of 100 kDa. Some new local Bt isolates had similar LC50 probit values to the reference strains.

  18. Toxicity and affecting factors of Bacillus thuringiensis var. israelensis on Chironomus kiiensis larvae.

    Science.gov (United States)

    Cao, Chuan-Wang; Sun, Li-Li; Wen, Rong-Rong; Li, Xiao-Peng; Wu, Hong-Qu; Wang, Zhi-Ying

    2012-01-01

    Abstract Bacillus thuringiensis var. israelensis (Bti) is a suitable agent for controlling Chironomus kiiensis, a major pest polluting water. In this study, laboratory bioassays were used to study toxicity and affecting factors of Bti on C. kiiensis larvae. Tests were conducted using three commercial Bti formulations (oil miscible suspension, 1,200 ITU/mL; wettable power, 1,200 ITU/mg; technical material, 5,000 ITU/mg) of Bti. The toxicity of Bti formulations to third and fourth instar C. kiiensis larvae was in decreasing order of technical material, oil miscible suspension, and wettable powder, based on the 12 and 24 hour LC50 values. Increasing larval densities (from 10 to 30 per bioassay cup) increased the LC50 values for fourth instar C. kiiensis larvae. The LC50 values for fourth instar larvae reared in sand substrate were higher than those from soil substrate, and autoclaved substrates significantly increased the LC50 values. The technical material of Bti at 12 and 24 hours responded similarly to changes in temperature between 30° C and 15° C, but the LC50 values at a range of tested temperatures showed distinct differences in time points.

  19. SUSCETIBILIDADE DE Helicoverpa armigera Hübner A FORMULADOS À BASE DE Bacillus thuringiensis BERLINER

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    Victor Luiz de Souza Lima

    2017-01-01

    Full Text Available O inseto Helicoverpa armigera recentemente registrado no Brasil é uma das maiores pragas da agricultura mundial. Pode atacar mais de 200 espécies de plantas e possui populações resistentes a diversos inseticidas. A utilização de microrganismos com potencial patogênico contra insetos é uma alternativa aos inseticidas. Essa pesquisa foi realizada com o objetivo de determinar a suscetibilidade de lagartas de H. armigera à produtos formulados à base de Bacillus thuringiensis (Bt Para os experimentos, foram utilizados os produtos comerciais Dipel® e Agree®, os quais tiveram sua concentração ajustada para 108 conídios viáveis ml-1. Essa concentração foi aplicada sobre dieta artificial, a qual foi colocada em uma placa de Petri que continha 10 lagartas de primeiro instar. Foram realizadas cinco repetições para cada produto. As avaliações foram feitas a cada 24h durante sete dias. Os produtos comerciais Dipel® e Agree® causaram, respectivamente, 100% e 94% de mortalidade das lagartas de H. armigera. Esse resultado mostra o potencial de produtos à base de Bt sobre H. armigera.

  20. Quantitative spectral light scattering polarimetry for monitoring fractal growth pattern of Bacillus thuringiensis bacterial colonies

    Science.gov (United States)

    Banerjee, Paromita; Soni, Jalpa; Ghosh, Nirmalya; Sengupta, Tapas K.

    2013-02-01

    It is of considerable current interest to develop various methods which help to understand and quantify the cellular association in growing bacterial colonies and is also important in terms of detection and identification of a bacterial species. A novel approach is used here to probe the morphological structural changes occurring during the growth of the bacterial colony of Bacillus thuringiensis under different environmental conditions (in normal nutrient agar, in presence of glucose - acting as additional nutrient and additional 3mM arsenate as additional toxic material). This approach combines the quantitative Mueller matrix polarimetry to extract intrinsic polarization properties and inverse analysis of the polarization preserving part of the light scattering spectra to determine the fractal parameter H (Hurst exponent) using Born approximation. Interesting differences are observed in the intrinsic polarization parameters and also in the Hurst exponent, which is a measurement of the fractality of a pattern formed by bacteria while growing as a colony. These findings are further confirmed with optical microscopic studies of the same sample and the results indicate a very strong and distinct dependence on the environmental conditions during growth, which can be exploited to quantify different bacterial species and their growth patterns.

  1. Toxicity of radiation-resistant strains of Bacillus thuringiensis (Berl.) to larval Plutella xylostella (L.)

    International Nuclear Information System (INIS)

    Jangi, M.S.; Ibrahim, Hasan

    1983-01-01

    A total of 24 isolates of Bacillus thuringiensis (Berliner), resistant to a γ-radiation dose of 100 krad, were screened for their toxicity to larval silkworms, Bombyxmori(L.), and 15 of them were subsequently tested for their toxicity to larval diamond-back moth, Plutella xylostella(L.). The LC 50 's of these isolates to B. mori ranged from 1.6 X 10 5 to 6.0 X 10 3 spores/mL or from 5.9 to 0.3 μg cellular protein/mL. The irradiation treatment produced isolates which were significantly more toxic to P. xylostella (LC 50 4 spores/mL or 3.7 μg cellular protein/mL) and/ or less toxic to B. mori (LC 50 > 2.3 X 10 4 spores/mL or 1.0 μg cellular protein/mL) than the parent commercial strain

  2. Molecular Approaches to Improve the Insecticidal Activity of Bacillus thuringiensis Cry Toxins

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    Wagner A. Lucena

    2014-08-01

    Full Text Available Bacillus thuringiensis (Bt is a gram-positive spore-forming soil bacterium that is distributed worldwide. Originally recognized as a pathogen of the silkworm, several strains were found on epizootic events in insect pests. In the 1960s, Bt began to be successfully used to control insect pests in agriculture, particularly because of its specificity, which reflects directly on their lack of cytotoxicity to human health, non-target organisms and the environment. Since the introduction of transgenic plants expressing Bt genes in the mid-1980s, numerous methodologies have been used to search for and improve toxins derived from native Bt strains. These improvements directly influence the increase in productivity and the decreased use of chemical insecticides on Bt-crops. Recently, DNA shuffling and in silico evaluations are emerging as promising tools for the development and exploration of mutant Bt toxins with enhanced activity against target insect pests. In this report, we describe natural and in vitro evolution of Cry toxins, as well as their relevance in the mechanism of action for insect control. Moreover, the use of DNA shuffling to improve two Bt toxins will be discussed together with in silico analyses of the generated mutations to evaluate their potential effect on protein structure and cytotoxicity.

  3. Persistence of detectable insecticidal proteins from Bacillus thuringiensis (Cry) and toxicity after adsorption on contrasting soils.

    Science.gov (United States)

    Hung, T P; Truong, L V; Binh, N D; Frutos, R; Quiquampoix, H; Staunton, S

    2016-01-01

    Insecticidal Cry, or Bt, proteins are produced by the soil-endemic bacterium, Bacillus thuringiensis and some genetically modified crops. Their environmental fate depends on interactions with soil. Little is known about the toxicity of adsorbed proteins and the change in toxicity over time. We incubated Cry1Ac and Cry2A in contrasting soils subjected to different treatments to inhibit microbial activity. The toxin was chemically extracted and immunoassayed. Manduca sexta was the target insect for biotests. Extractable toxin decreased during incubation for up to four weeks. Toxicity of Cry1Ac was maintained in the adsorbed state, but lost after 2 weeks incubation at 25 °C. The decline in extractable protein and toxicity were much slower at 4 °C with no significant effect of soil sterilization. The major driving force for decline may be time-dependent fixation of adsorbed protein, leading to a decrease in the extraction yield in vitro, paralleled by decreasing solubilisation in the larval gut. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Extraction of flocculants from a strain of Bacillus thuringiensis and analysis of their properties

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    Jingrong Wu

    2017-07-01

    Full Text Available In a preliminary screening study, our laboratory isolated from the biofloc in aquaculture waters a strain of Bacillus thuringiensis, which produced highly efficient bio-flocculants. In the present study, we extracted the crude flocculants from this strain and analyzed their properties. Distribution analysis indicated that the flocculants were mainly distributed in the supernatant of the fermentation liquid. The flocculants were extracted using an ethanol extraction method, and the chemical compositions and morphology of the crude flocculants were analyzed using the Molish reaction, Fehling reaction, ninhydrin reaction, biuret reaction, phenol-sulfuric acid assay, Coomassie brilliant blue staining, ultraviolet scanning, infrared scanning and scanning electron microscopy. The carbohydrate composition of the polysaccharides in the flocculants was analyzed with thin layer chromatography. The results indicated that flocculants were solid substances with an ivory white color and their texture was loose and soft. Visualization under scanning electron microscopy revealed that their ultra-morphology consisted of small, long and fiber-like shapes. Chemical and physical analyses indicated that polysaccharides accounted for 34.5% of the components in the crude flocculants. The monosaccharides present in crude flocculants included mainly glucose, galactose and mannitose.

  5. Increase in midgut microbiota load induces an apparent immune priming and increases tolerance to Bacillus thuringiensis.

    Science.gov (United States)

    Hernández-Martínez, Patricia; Naseri, Bahram; Navarro-Cerrillo, Gloria; Escriche, Baltasar; Ferré, Juan; Herrero, Salvador

    2010-10-01

    The insect immune system is comprised of both humoral and cellular components that are mobilized in response to parasitic or pathogenic infections. Activation of the immune response implies a considerable expenditure of energy and that is why insects rely on inducible pathways that are activated after coming into contact with the pathogenic agent. Known as immune priming, insects can prolong the activation of the immune response and transmit their immune status to the next generation. Starting from a laboratory colony of the lepidopteran Spodoptera exigua and using the lytic zone assay as a measure of the immune status, we selected for a sub-colony with high levels of immune activity in the absence of external challenging with bacteria. Immune-activated insect showed characteristics that are typical reported for immune primed insects, such as increased tolerance to pathogens (Bacillus thuringiensis in our case), fitness-cost associated to the immune status, and maternal transmission of the immune status. However, additional analysis revealed that the selection for the immune-activated insects was based on the selection of insects carrying a higher bacterial load in the midgut. Our results suggest that activation of the immune system in S. exigua may not only occur as consequence of the immune priming but also from an increase in midgut microbiota load. © 2010 Society for Applied Microbiology and Blackwell Publishing Ltd.

  6. Proteomic Analysis of Bacillus thuringiensis Strain 4.0718 at Different Growth Phases

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    Xiaohui Li

    2012-01-01

    Full Text Available The growth process of Bacillus thuringiensis Bt4.0718 strain was studied using proteomic technologies. The proteins of Bt whole cells at three phases—middle vegetative, early sporulation, and late sporulation—were extracted with lysis buffer, followed with separation by 2-DE and identified by MALDI-TOF/TOF MS. Bioactive factors such as insecticidal crystal proteins (ICPs including Cry1Ac(3, Cry2Aa, and BTRX28, immune inhibitor (InhA, and InhA precursor were identified. InhA started to express at the middle vegetative phase, suggesting its contribution to the survival of Bt in the host body. At the early sporulation phase, ICPs started their expression. CotJC, OppA, ORF1, and SpoIVA related to the formation of crystals and spores were identified, the expression characteristics of which ensured the stable formation of crystals and spores. This study provides an important foundation for further exploration of the stable expression of ICPs, the smooth formation of crystals, and the construction of recombinant strains.

  7. Bacillus thuringiensis: a successful insecticide with new environmental features and tidings.

    Science.gov (United States)

    Jouzani, Gholamreza Salehi; Valijanian, Elena; Sharafi, Reza

    2017-04-01

    Bacillus thuringiensis (Bt) is known as the most successful microbial insecticide against different orders of insect pests in agriculture and medicine. Moreover, Bt toxin genes also have been efficiently used to enhance resistance to insect pests in genetically modified crops. In light of the scientific advantages of new molecular biology technologies, recently, some other new potentials of Bt have been explored. These new environmental features include the toxicity against nematodes, mites, and ticks, antagonistic effects against plant and animal pathogenic bacteria and fungi, plant growth-promoting activities (PGPR), bioremediation of different heavy metals and other pollutants, biosynthesis of metal nanoparticles, production of polyhydroxyalkanoate biopolymer, and anticancer activities (due to parasporins). This review comprehensively describes recent advances in the Bt whole-genome studies, the last updated known Bt toxins and their functions, and application of cry genes in plant genetic engineering. Moreover, the review thoroughly describes the new features of Bt which make it a suitable cell factory that might be used for production of different novel valuable bioproducts.

  8. Isolation and characterization of Bacillus thuringiensis strains active against Elasmopalpus lignosellus (Zeller, 1848 (Lepidoptera, Pyralidae

    Directory of Open Access Journals (Sweden)

    Janaina Zorzetti

    2017-08-01

    Full Text Available Elasmopalpus lignosellus (Zeller, 1848 (Lepidoptera, Pyralidae is an insect pest of 60 economically important crops, including sugarcane, wheat, soybean, rice, beans, sorghum, peanuts, and cotton. The aim of this work was to select and characterize Bacillus thuringiensis isolates with insecticidal activity against E. Lignosellus that could be used as an alternative method of control. Selective bioassays were done to evaluate the toxicity of 47 isolates against first instar larvae of E. lignosellus. For the most toxic bacterial strains, the lethal concentration (LC50 was estimated and morphological, biochemical and molecular methods were used to characterize the isolates. Among the 47 isolates tested, 12 caused mortality above 85% and showed LC50 values from 0.038E+8 to 0.855E+8 spores mL-1. Isolates BR83, BR145, BR09, BR78, S1534, and S1302 had the lowest LC50 values and did not differ from the standard HD-1 strain; the exception was BR83.The protein profiles produced bands with molecular masses of 60-130 kDa. The genes cry1, cry2, cry3, and cry11 were identified in the molecular characterization. The morphological analysis identified three different crystal inclusions: bipyramidal, spherical and cuboidal. Among the tested isolates, 12 isolates have potential for biotechnological control of E. Lignosellus by development of new biopesticides or genetically modified plants.

  9. Safety of Bacillus thuringiensis Cry1C protein for Daphnia magna based on different functional traits.

    Science.gov (United States)

    Chen, Yi; Yang, Yan; Zhu, Haojun; Romeis, Jörg; Li, Yunhe; Peng, Yufa; Chen, Xiuping

    2018-01-01

    Cry1C is a Bacillus thuringiensis (Bt) insecticidal protein and it can be produced by transgenic rice lines developed in China. Cladocera species are common aquatic arthropods that may be exposed to insecticidal proteins produced in Bt-transgenic plants through ingestion of pollen or crop residues in water. As the cladoceran Daphnia magna plays an important role in the aquatic food chain, it is important to assess the possible effects of Bt crops to this species. To evaluate the safety of the Cry1C protein for D. magna, individuals were exposed to different concentrations of purified Cry1C protein in M4 medium for 21 days. Potassium dichromate (K 2 Cr 2 O 7 ), a known toxicant to D. magna, was added to M4 medium as a positive control treatment, and pure M4 medium was used as a negative control. Our results show that developmental, reproductive, and biochemical parameters of D. magna were not significantly different between Cry1C and negative control treatments but were significantly inhibited by the positive control. We thus conclude that D. magna is insensitive to Cry1C. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Characterization of Bacillus thuringiensis soil isolates from Cuba, with insecticidal activity against mosquitoes.

    Science.gov (United States)

    González, Aileen; Díaz, Raúl; Díaz, Manuel; Borrero, Yainais; Bruzón, Rosa Y; Carreras, Bertha; Gato, René

    2011-09-01

    Chemical insecticides may be toxic and cause environmental degradation. Consequently, biological control for insects represents an alternative with low ecological impact. In this work, three soil isolates (A21, A51 and C17) from different regions of the Cuban archipelago were identified, characterized and evaluated against Aedes aegypti and Culex quinquefasciatus. The new isolates were compared with reference IPS82 strain and two strains isolated from biolarvicides Bactivec and Bactoculicida, respectively. The differentiation was done by morphological, biochemical, bioassays activity and molecular methods (SDS-PAGE, plasmid profile and random amplified polymorphic analysis). All isolates were identified as Bacillus thuringiensis. The A21, A51 and C17 isolates showed higher larvicide activity than Bactivec's isolated reference strain, against both A. aegypti and C. quinquefasciatus. A21 isolate had a protein profile similar to IPS82 and Bactivec strain. A51 and C17 isolates produced a characteristic proteins pattern. A21 and A51 isolates had plasmid patterns similar to IPS82 standard strain, while C17 isolate had different both plasmid profile and protein bands. All the studied isolates showed a diverse RAPD patterns and were different from the strains previously used in biological control in Cuba.

  11. Vegetative insecticidal protein of Bacillus thuringiensis BLB459 and its efficiency against Lepidoptera.

    Science.gov (United States)

    Boukedi, Hanen; Ben Khedher, Saoussen; Hadhri, Rania; Jaoua, Samir; Tounsi, Slim; Abdelkefi-Mesrati, Lobna

    2017-04-01

    Bacillus thuringiensis strain BLB459 supernatant showed a promising activity against Lepidopteran pests with extremely damages in the larvae midgut. Investigations of the genes that encode secreted toxin demonstrated that this strain harbored a vip3-type gene named vip3(459). Based on its original nucleotide and amino acid sequences, this gene was cloned into pET-14b vector and overexpressed in Escherichia coli. The expressed protein was purified and tested against different insects and interestingly the novel toxin demonstrated a remarkable activity against the stored products pest Ephestia kuehniella and the polyphagous insects Spodoptera littoralis and Agrotis segetum. As demonstrated, the acute activity of Vip3(459) protein against A. segetum can be due to its original amino acids sequence and the putative receptors of this toxin in the larvae midgut. These results demonstrated that this Vip3 toxin showed a wide spectrum of activity against Lepidoptera and support its use as a biological control agent. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. EFEKTIVITAS Bacillus thuringiensis H-14 STRAIN LOKAL DALAM BUAH KELAPA TERHADAP LARVA Anopheles sp dan Culex sp di KAMPUNG LAUT KABUPATEN CILACAP

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    Blondine Ch. P

    2013-07-01

    Full Text Available Abstrak Bacillus thuringiensis serotipe H-14 strain lokal adalah bakteri patogen bersifat target spesifiknya larva nyamuk, aman bagi mamalia dan lingkungan. Penelitian bertujuan menentukan efektivitas B. thuringiensis H-14 strain lokal yang dikembangbiakkan dalam buah kelapa untuk pengendalian larva Anopheles sp dan Culex sp. Rancangan eksperimental semu, terdiri dari kelompok perlakuan dan kontrol. Bacillus thuringiensis H-14 strain lokal dikembangbiakan dalam10 buah kelapa umur 6–8 bulan, dengan berat kira-kira 1 kg, telah berisi air kelapa sekitar 400-500 ml/buah kelapa yang diperoleh dari Desa Klaces, Kampung Laut, Kabupaten Cilacap. Diinkubasi selama 14 hari pada temperatur kamar dan ditebarkan di 6 kolam yang menjadi habitat perkembangbiakan larva nyamuk dengan luas berkisar 3–100 m2.Hasil yang diperoleh menunjukkan efektivitas B. thuringiensis H-14 strain lokal terhadap larva Anopheles sp dan Culex sp selama 1 hari sesudah penebaran kematian larva berturut-turut sebesar 80–100% dan 79,31–100%. Sedangkan pada hari ke-14 sebesar 69,30–76,71% dan 67,69–86,04%. Buah kelapa dapat digunakan sebagai media lokal alternatif untuk pengembangbiakan B. thuringiensis H-14 strain lokal Kata kunci: B. thuringiensis H-14,  strain  lokal, buah kelapa, pengendalian larva Abstract Bacillus thuringiensis serotype H-14 local strain is pathogenic bacteria which specific  target to mosquito larvae. It is safe for mammals and enviroment. The aims of this study was to determine the effectivity of B. thuringiensis H-14 local strain which culturing in thecoconut wates against Anopheles sp and Culex sp mosquito larvae. This research is quasi experiment which consist of treated  and control groups. Bacillus thuringiensis H-14 local strain was cultured in 10 coconuts with 6–8 months age with weight around 1 kg that contained were approximately 400-500 ml/coconut were taken from Klaces village, Kampung Laut. After that the coconuts incubated for 14

  13. Avaliação da eficiência de formulações de Bacillus thuringiensis para o controle de traça-das-crucíferas em repolho no Distrito Federal Efficiency of Bacillus thuringiensis formulations in controlling Diamondback Moth in cabbage in the Federal District

    Directory of Open Access Journals (Sweden)

    Marina Castelo Branco

    1999-11-01

    Full Text Available A traça-das-crucíferas (Plutella xylostella é a praga mais importante do repolho no Distrito Federal. Seu controle é feito basicamente com inseticidas. Novos produtos são constantemente avaliados para o controle da praga e neste trabalho duas novas formulações de Bacillus thuringiensis [Bacillus thuringiensis var. aizawai (350 e 500 g/hae B. thuringiensis var. kurstaki x B. thuringiensis var. aizawai (350 e 500 ml/ha] foram avaliadas no período de maio a outubro de 1995. Os tratamentos B. thuringiensis var. kurstaki (500 ml/ha, deltametrina (240 ml/ha e uma testemunha sem pulverização foram também incluídos no experimento. O delineamento foi blocos ao acaso, com sete tratamentos e quatro repetições. Os resultados mostraram que B. thuringiensis var. aizawai nas duas dosagens avaliadas e B. thuringiensis var. kurstaki x B. thuringiensis var. aizawai (500 ml/ha foram os produtos mais eficientes. Ao final do experimento larvas e pupas de traça-das-crucíferas foram coletadas no campo e a primeira geração foi submetida a um teste de laboratório onde discos de folhas de repolho foram tratados com as dosagens dos inseticidas a base de B. thuringiensis utilizadas no campo. Larvas de segundo estádio foram colocadas sobre os discos tratados e a mortalidade de larvas avaliada após 72 h. Todos os tratamentos causaram mais de 97% de mortalidade de larvas. O resultado do teste de laboratório sugere que a menor eficiência de B. thuringiensis var. kurstaki x B. thuringiensis var. aizawai (350 ml/ha e B. thuringiensis var. kurstaki (500 ml/ha no teste de campo quando comparada aos demais Bacillus, pode ser devido à mais rápida degradação destes produtos no ambiente.The Diamondback Moth (Plutella xylostella is the most important cabbage pest in the Federal District. New insecticides are frequently tested for its control and in this study two new Bacillus thuringiensis formulations [B. thuringiensis var. aizawai (350 and 500 g/ha and B

  14. Toxicity, activation process, and histopathological effect of Bacillus thuringiensis vegetative insecticidal protein Vip3Aa16 on Tuta absoluta.

    Science.gov (United States)

    Sellami, Sameh; Cherif, Maroua; Abdelkefi-Mesrati, Lobna; Tounsi, Slim; Jamoussi, Kaïs

    2015-02-01

    Tuta absoluta is a destructive moth of Solanaceae plants and especially tomatoes. Here, we considered the entomopathogenic activity of the Bacillus thuringiensis Vip3Aa16 protein heterologously produced by Escherichia coli against T. absoluta. Purified Vip3Aa16 showed lower lethal concentration 50 % against third instar larvae (Toxin/tomato leaf) (335 ± 17 ng/cm(2)) compared to that of B. thuringiensis kurstaki HD1 δ-endotoxins (955 ± 4 ng/cm(2)) (P larvae gut soluble proteases, yielding derivative proteins essentially of about 62 and 33 kDa. The gut-soluble proteases could contain trypsin-like enzymes implicated in Vip3Aa16 activation since the proteolysis was inhibited using specific protease inhibitors. Additionally, we showed that the histopathological effect of Vip3Aa16 on T. absoluta larva midguts consisted on a microvillus damage and an epithelial cell rupture.

  15. Biological control of the spruce coneworm Dioryctria abietella: Spraying with Bacillus thuringiensis reduced damage in a seed orchard

    Energy Technology Data Exchange (ETDEWEB)

    Weslien, Jan [Forestry Research Inst. of Sweden, Uppsala (Sweden)

    1999-08-01

    Spraying of cones with Bacillus thuringiensis var. Kurstaki x Aizawa in a Norway spruce (Picea abies (L.) Karst.) seed orchard reduced damage by Dioryctria abietella Den. et Schiff. (Lepidoptera, Pyralidae). The incidence of D. abietella attacks was about 80% among unsprayed cones, but less than 15% showed attacks among cones that had been sprayed three times at 9-day intervals with a 0.2% suspension (weight/weight) of the B. thuringiensis preparation in water (Turex 50 WP, 25 000 IU/mg). Attacked, sprayed cones had fewer D. abietella larvae per cone than attacked, unsprayed cones. Spraying did not reduce the damage by Cydia strobilella (L.) (Lepidoptera, Tortricidae) or by Strobilomyia anthracina Czerny (Diptera, Anthomyiidae) 13 refs, 3 tabs

  16. Identification of new isolates of Bacillus thuringiensis using rep-PCR products and delta-endotoxin electron microscopy

    Directory of Open Access Journals (Sweden)

    A.S.G. Lima

    2002-01-01

    Full Text Available PCR has been used to analyze the distribution of REP (Repetitive Extragenic Palindromic and ERIC (Enterobacterial Repetitive Intergenic Consensus sequences (rep-PCR found within the genome of the bacterium Bacillus thuringiensis, with the purpose to analyze the genetic similarities among 56 subspecies samples and 95 field isolates. The PCR products were analyzed by EB-AGE (ethidium bromide-agarose electrophoresis and then submitted to banding comparisons, based on the Phyllip software algorithm. When the banding similarities were considered for comparison purposes among all the strains, the phylogenic tree patterns varied according to the rep-PCR primers considered, but, from a broader point of view, the ERIC sequences produced better results, which, together with electron microscopy analysis of the released parasporal bodies and colony morphology characteristics, allowed to detect two possible new subspecies of B. thuringiensis.

  17. Identification of new isolates of Bacillus thuringiensis using rep-PCR products and d-endotoxin electron microscopy

    Directory of Open Access Journals (Sweden)

    Lima A.S.G.

    2002-01-01

    Full Text Available PCR has been used to analyze the distribution of REP (Repetitive Extragenic Palindromic and ERIC (Enterobacterial Repetitive Intergenic Consensus sequences (rep-PCR found within the genome of the bacterium Bacillus thuringiensis, with the purpose to analyze the genetic similarities among 56 subspecies samples and 95 field isolates. The PCR products were analyzed by EB-AGE (ethidium bromide-agarose electrophoresis and then submitted to banding comparisons, based on the Phyllip software algorithm. When the banding similarities were considered for comparison purposes among all the strains, the phylogenic tree patterns varied according to the rep-PCR primers considered, but, from a broader point of view, the ERIC sequences produced better results, which, together with electron microscopy analysis of the released parasporal bodies and colony morphology characteristics, allowed to detect two possible new subspecies of B. thuringiensis.

  18. Complete genome sequence of Bacillus thuringiensis YC-10, a novel active strain against plant-parasitic nematodes.

    Science.gov (United States)

    Cheng, Feixue; Wang, Jian; Song, Zhiqiang; Cheng, Ju'e; Zhang, Deyong; Liu, Yong

    2015-09-20

    Bacillus thuringiensis is an important microbial biopesticide for controlling agricultural pests by the production of toxic parasporal crystals proteins.Here,we report the finished annotated genome sequence of B. thuringiensis YC-10,which is highly toxic to nematodes.The complete genome sequence consists of a circular chromosome and nine circular plasmids,which the biggest plasmid harbors six parasporal crystals proteins genes consisting of cry1Aa, cry1Ac, cry1Ia, cry2Aa, cry2Ab and cryB1. The crystals proteins of Cry1Ia and Cry1Aa have high nematicidal activity against Meloidogyne incognita. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Effects of temperature on mortality of larval stable fly (Diptera: Muscidae) caused by five isolates of Bacillus thuringiensis.

    Science.gov (United States)

    Lysyk, T J; Selinger, L B

    2012-04-01

    We examined the effects of temperature on mortality of larval stable fly [Stomoxys calcitrans (L.)] caused by Bacillus thuringiensis tolworthi 4L3, B. t. darmastedensis 4M1, B. t. thompsoni 401, B. t. thuringiensis HD2, and B. t. kurstaki HD945. At moderate doses, mortality caused by all isolates ranged from 87 to 99% at 15 degrees C and declined to 29-63% as temperature increased to 30 degrees C. A similar pattern was seen when a higher dose was used, except that the reduction in mortality at warmer temperatures was not as great as was seen with the moderate doses. Insecticidal activity of each isolate against first-instar larvae was reduced by only 5-15% after 5 d in the medium. Mortality of second- and third-instar larvae ranged from 2 to 21%, suggesting the isolates were less effective against larger larvae.

  20. Persistence of Toxic Activity of Fermentation Extracts from Bacillus thuringiensis var. israelensis after More Than Three Decades of Storage

    Directory of Open Access Journals (Sweden)

    Luis Jesús Galán-Wong

    2017-01-01

    Full Text Available This study was carried out to determine the persistence of toxicity of fermentation extracts of Bacillus thuringiensis var. israelensis after more than three decades of storage. For this purpose, a population of Aedes aegypti was established. The mortality rate of 20 spore-crystal extracts purified using the acetone-lactose coprecipitation method was measured and evaluated by bioassays according to a modified WHO protocol. The extracts with the highest mortality rate were determined in triplicate by their LD50 and LD98. All extracts showed toxicity at the highest tested dose (1000 ppm and some, such as strains 3260 and 3501, still killed larvae at doses as low as 0.01 ppm. These data are surprising because no study on the activity of B. thuringiensis toxic proteins after such a long storage time has been reported.

  1. Toxicity of Tolyltriazole to Bacillus Microorganisms.

    Science.gov (United States)

    2000-03-01

    Bacillus coagulans Microbacterium lacticum Jupiter Bacillus thuringiensis Bacillus thuringiensis Bacillus cereus Bacillus Bacillus thuringiensis...TOXICITY OF TOLYLTRIAZOLE TO BACILLUS MICROORGANISMS THESIS Christopher J. Leonard, First Lieutenant, USAF AFIT/GEE/ENV/OOM-12 Approved for...AND DATES COVERED Master’s Thesis 4. TITLE AND SUBTITLE TOXICITY OF TOLYLTRIAZOLE TO BACILLUS MICROORGANISMS 6. AUTHOR(S) Christopher J

  2. Molecular characterization of a DNA fragment harboring the replicon of pBMB165 from Bacillus thuringiensis subsp. tenebrionis

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    Yu Ziniu

    2006-10-01

    Full Text Available Abstract Background Bacillus thuringiensis belongs to the Bacillus cereus sensu lato group of Gram-positive and spore-forming bacteria. Most isolates of B. thuringiensis can bear many endogenous plasmids, and the number and size of these plasmids can vary widely among strains or subspecies. As far as we know, the replicon of the plasmid pBMB165 is the first instance of a plasmid replicon being isolated from subsp. tenebrionis and characterized. Results A 20 kb DNA fragment containing a plasmid replicon was isolated from B. thuringiensis subsp. tenebrionis YBT-1765 and characterized. By Southern blot analysis, this replicon region was determined to be located on pBMB165, the largest detected plasmid (about 82 kb of strain YBT-1765. Deletion analysis revealed that a replication initiation protein (Rep165, an origin of replication (ori165 and an iteron region were required for replication. In addition, two overlapping ORFs (orf6 and orf10 were found to be involved in stability control of plasmid. Sequence comparison showed that the replicon of pBMB165 was homologous to the pAMβ1 family replicons, indicating that the pBMB165 replicon belongs to this family. The presence of five transposable elements or remnants thereof in close proximity to and within the replicon control region led us to speculate that genetic exchange and recombination are potentially responsible for the divergence among the replicons of this plasmid family. Conclusion The replication and stability features of the pBMB165 from B. thuringiensis subsp. tenebrionis YBT-1765 were identified. Of particular interest is the homology and divergence shared between the pBMB165 replicon and other pAMβ1 family replicons.

  3. Re-aerosolization of Bacillus thuringiensis spores from concrete and turf.

    Science.gov (United States)

    Bishop, A H; O'Sullivan, C M; Lane, A; Butler Ellis, M C; Sellors, W J

    2017-05-01

    Spores of Bacillus anthracis deposited on surfaces can become airborne again as a result of air currents and mechanical forces. As such, they are a potential source of infection by inhalation. Spores of Bacillus thuringiensis were used to quantify this phenomenon in a simulation of outdoor conditions. Concrete and turf surfaces were inoculated by aerosol to produce high spore densities (greater than 1 × 10 9  CFU per m 2 ) which were then subjected to the passage of air at 10 ms -1 with and without simulated walking. Re-aerosolized spores were sampled by wetted wall cyclone air samplers. The mean total re-aerosolization rate from concrete (m -2  min -1 ) was 1·16 × 10 -3 for wind alone and 3·2 × 10 -3 for wind and simulated walking while for turf the respective values were 2·7 × 10 -4 and 6·7 × 10 -4 . Following the malicious and/or accidental release of an aerosol of Bacillus anthracis spores, the immediate risk of human inhalation would decrease as the spores were deposited on surfaces or diluted by wind flow. There is, however, a concern that the deposited spores could become re-aerosolized and so present an ongoing hazard. Using an accurate simulant for B. anthracis spores a method is reported here that allowed the enumeration of re-aerosolized spores from concrete and turf by wind flow and footfall. Under the conditions used, the rates of re-aerosolization were low. These findings will need to be verified under real outdoor conditions before the true significance in terms of secondary exposure to pathogenic spores can be assessed. © 2017 Crown copyright. Letter of Applied Microbiology © 2017 The Society for Applied Microbiology. This article is published with the permission of the Controller of HMSO and the Queen's Printer for Scotland.

  4. A strong promoter of a non-cry gene directs expression of the cry1Ac gene in Bacillus thuringiensis.

    Science.gov (United States)

    Zhang, Xin; Gao, Tantan; Peng, Qi; Song, Lai; Zhang, Jie; Chai, Yunrong; Sun, Dongmei; Song, Fuping

    2018-04-01

    Bacillus thuringiensis bacteria show insecticidal activities that rely upon the production of insecticidal crystal proteins, which are encoded by cry or cyt genes and can target a variety of insect pests. It has been shown that cry1Ac is the only cry gene in B. thuringiensis subsp. kurstaki HD73 (B. thuringiensis HD73) and its expression is controlled by both σ E and σ K . Here, we report a novel σ E -dependent strong promoter of a non-cry gene (HD73_5014), which can direct strong cry1Ac gene expression in B. thuringiensis HD73. We constructed an E. coli-B. thuringiensis shuttle vector (pHT315-P 5014 -1Ac) for cry1Ac gene expression, using the HD73_5014 gene promoter. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blot analysis showed that expression of the cry1Ac gene directed by the HD73_5014 gene promoter was at the same level as that directed by the previously known strongest cry promoter, P cry8E . However, this strain did not form typical bipyramidal crystals in mother cells, as observed by transmission electron microscopy and atomic force microscope. The strain with Cry1Ac protein expression under the control of the HD73_5014 gene promoter (P 5014 -cry1Ac) showed insecticidal activity against Plutella xylostella similar to that under the control of the orf1cry8E gene promoter (P cry8E -cry1Ac). Collectively, these results suggest that the HD73_5014 gene promoter, as a non-cry gene promoter, would be an efficient transcriptional element for cry gene expression. These data also show the possibility for improving Cry production by searching for transcriptional elements in not only cry genes, but also non-cry genes.

  5. Larvicidal Activities of Indigenous Bacillus thuringiensis Isolates and Nematode Symbiotic Bacterial Toxins against the Mosquito Vector, Culex pipiens (Diptera: Culicidae

    Directory of Open Access Journals (Sweden)

    Ashraf M Ahmed

    2017-06-01

    Full Text Available Background: The incidence of mosquito-borne diseases and the resistance of mosquitoes to conventional pesticides have recently caused a panic to the authorities in the endemic countries. This study was conducted to identify native larvicidal biopesticides against Culex pipiens for utilization in the battle against mosquito-borne diseases.Methods: Larvicidal activities of new indigenous Bacillus thuringiensis isolates and crude toxin complexes (TCs of two nematode bacterial-symbionts, Photorhabdus luminescens akhurstii (HRM1 and Ph. luminescens akhurstii (HS1 that tested against Cx. pipiens. B. thuringiensis isolates were recovered from different environmental samples in Saudi Arabia, and the entomopathogenic nematodes, Heterorhabditis indica (HRM1 and He. sp (HS1 were iso­lated from Egypt. Larvicidal activities (LC50 and LC95 of the potentially active B. thuringiensis strains or TCs were then evaluated at 24 and 48h post-treatment.Results: Three B. thuringiensis isolates were almost as active as the reference B. thuringiensis israelensis (Bti-H14, and seven isolates were 1.6–5.4 times more toxic than Bti-H14. On the other hand, the TCs of the bacterial sym­bionts, HRM1 and HS1, showed promising larvicidal activities. HS1 showed LC50 of 2.54 folds that of HRM1 at 24h post-treatment. Moreover, histopathological examinations of the HS1-treated larvae showed deformations in midgut epithelial cells at 24h post-treatment.Conclusion: Synergistic activity and molecular characterization of these potentially active biocontrol agents are currently being investigated. These results may lead to the identification of eco-friend mosquito larvicidal product(s that could contribute to the battle against mosquito-borne diseases.

  6. Distribution of Bacillus thuringiensis subsp. israelensis in Soil of a Swiss Wetland Reserve after 22 Years of Mosquito Control▿†

    Science.gov (United States)

    Guidi, Valeria; Patocchi, Nicola; Lüthy, Peter; Tonolla, Mauro

    2011-01-01

    Recurrent treatments with Bacillus thuringiensis subsp. israelensis are required to control the floodwater mosquito Aedes vexans that breeds in large numbers in the wetlands of the Bolle di Magadino Reserve in Canton Ticino, Switzerland. Interventions have been carried out since 1988. In the present study, the spatial distribution of resting B. thuringiensis subsp. israelensis spores in the soil was measured. The B. thuringiensis subsp. israelensis concentration was determined in soil samples collected along six transects covering different elevations within the periodically flooded zones. A total of 258 samples were processed and analyzed by quantitative PCR that targeted an identical fragment of 159 bp for the B. thuringiensis subsp. israelensis cry4Aa and cry4Ba genes. B. thuringiensis subsp. israelensis spores were found to persist in soils of the wetland reserve at concentrations of up to 6.8 log per gram of soil. Continuous accumulation due to regular treatments could be excluded, as the decrease in spores amounted to 95.8% (95% confidence interval, 93.9 to 97.7%). The distribution of spores was correlated to the number of B. thuringiensis subsp. israelensis treatments, the elevation of the sampling point, and the duration of the flooding periods. The number of B. thuringiensis subsp. israelensis treatments was the major factor influencing the distribution of spores in the different topographic zones (P thuringiensis subsp. israelensis spores are rather immobile after their introduction into the environment. PMID:21498758

  7. The Synergist Effect of The Henna Plant, Lawsonia alba on Bacillus thuringiensis var. kurstaki Against Third Larval Instar of Pistachio Leaf Borer, Ocneria terebinthina Strg. (Lep.: Lymanteriidae

    Directory of Open Access Journals (Sweden)

    Zahra Sheibani

    2017-12-01

    Full Text Available Ocneria terebinthina Strg. (Lep.: Lymanteriidae is a leaf borer pest in pistachio orchard. The toxicity of Bacillus thuringiensis var. kurstaki alone and in combination with henna powder was investigated on third larval instar of O. terebinthina under laboratory conditions. Bioassay was carried out using spray technique on pistachio offshoot. Probit analysis of concentration-mortality data was conducted to estimate the LC50 value. The LC50 value of B. thuringiensis on third instar of larvae was estimated as 2817.30 ppm. The synergist effect of henna powder on efficacy of B. thuringiensis was also evaluated. Three concentrations of henna powder were combined with minimum lethal dose of B. thuringiensis against third larval instar. The results revealed that the henna powder had synergist effect on B. thuringiensis. The combination of B. thuringiensis and henna powder (6000 ppm caused 80.68% mortality, whereas B. thuringiensis alone caused 15.91% mortality. Our results suggested that henna powder can increase the efficacy of B. thuringiensis in controlling O. terebinthina in pest integrated management.

  8. Molecular characterization of indigenous Bacillus thuringiensis strains isolated from Kashmir valley.

    Science.gov (United States)

    Reyaz, A L; Gunapriya, L; Indra Arulselvi, P

    2017-06-01

    Bacillus thuringiensis (Bt) being an eco-friendly bioinsecticide is effectively used in pest management strategies and, therefore, isolation and identification of new strains effective against a broad range of target pests is important. In the present study, new indigenous B. thuringiensis strains were isolated and investigated so that these could be used as an alternative and/or support the current commercial strains/cry proteins in use. For this, 159 samples including soil, leaf and spider webs were collected from ten districts of Kashmir valley (India). Of 1447 bacterial strains screened, 68 Bt strains were identified with 4 types of crystalline inclusions. Crystal morphology ranking among the Bt strains was spherical (69.11%) > spore attached (8.82%) > rod (5.88%) = bipyramidal (5.88%) > spherical plus rod (4.41%) > spherical plus bipyramidal (2.94%) = cuboidal (2.94%). SDS-PAGE investigation of the spore-crystal mixture demonstrated Bt strains contained proteins of various molecular weights ranging from 150 to 28 kDa. Insecticidal activity of the 68 indigenous Bt strains against Spodoptera litura neonates showed that Bt strain SWK1 strain had the highest mortality. Lepidopteron active genes (cry1, cry2Ab, cry2Ab) were present in six Bt strains. Further, analysis of a full-length cry2A gene (~1.9 kb) by PCR-RFLP in strain SWK1 revealed that it was a new cry2A gene in Bt strain SWK1 and was named as cry2Al1 (GenBank Accession No. KJ149819.1) using the Bt toxin nomenclature ( http://www.btnomenclature.info ). Insect bioassays with neonate larvae of S. litura and H. armigera showed that the purified Cry2Al1 is toxic to S. litura with LC 50 2.448 µg/ml and H. armigera with LC 50 3.374 µg/ml, respectively. However, it did not produce any mortality in third instar larvae of Aedes aegypti, Culex quinquefasciatus and Anopheles stephensi larvae/pupae insects (100 µg/ml) at 28 ± 2 °C and 75 to 85% relative humidity under a photoperiod of 14L:10D.

  9. Response surface modeling for hot, humid air decontamination of materials contaminated with Bacillus anthracis ∆Sterne and Bacillus thuringiensis Al Hakam spores

    Science.gov (United States)

    2014-01-01

    Response surface methodology using a face-centered cube design was used to describe and predict spore inactivation of Bacillus anthracis ∆Sterne and Bacillus thuringiensis Al Hakam spores after exposure of six spore-contaminated materials to hot, humid air. For each strain/material pair, an attempt was made to fit a first or second order model. All three independent predictor variables (temperature, relative humidity, and time) were significant in the models except that time was not significant for B. thuringiensis Al Hakam on nylon. Modeling was unsuccessful for wiring insulation and wet spores because there was complete spore inactivation in the majority of the experimental space. In cases where a predictive equation could be fit, response surface plots with time set to four days were generated. The survival of highly purified Bacillus spores can be predicted for most materials tested when given the settings for temperature, relative humidity, and time. These predictions were cross-checked with spore inactivation measurements. PMID:24949256

  10. Test methods and response surface models for hot, humid air decontamination of materials contaminated with dirty spores of Bacillus anthracis ∆Sterne and Bacillus thuringiensis Al Hakam.

    Science.gov (United States)

    Buhr, T L; Young, A A; Barnette, H K; Minter, Z A; Kennihan, N L; Johnson, C A; Bohmke, M D; DePaola, M; Cora-Laó, M; Page, M A

    2015-11-01

    To develop test methods and evaluate survival of Bacillus anthracis ∆Sterne or Bacillus thuringiensis Al Hakam on materials contaminated with dirty spore preparations after exposure to hot, humid air using response surface modelling. Spores (>7 log10 ) were mixed with humic acid + spent sporulation medium (organic debris) or kaolin (dirt debris). Spore samples were then dried on five different test materials (wiring insulation, aircraft performance coating, anti-skid, polypropylene, and nylon). Inoculated materials were tested with 19 test combinations of temperature (55, 65, 75°C), relative humidity (70, 80, 90%) and time (1, 2, 3 days). The slowest spore inactivation kinetics was on nylon webbing and/or after addition of organic debris. Hot, humid air effectively decontaminates materials contaminated with dirty Bacillus spore preparations; debris and material interactions create complex decontamination kinetic patterns; and B. thuringiensis Al Hakam is a realistic surrogate for B. anthracis. Response surface models of hot, humid air decontamination were developed which may be used to select decontamination parameters for contamination scenarios including aircraft. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.

  11. Uso de productos derivados de Bacillus thuringiensis como alternativa de control en nematodos de importancia veterinaria. Revisión

    Directory of Open Access Journals (Sweden)

    Alejandro Vázquez-Pineda

    2012-01-01

    Full Text Available La bacteria entomopatógena Bacillus thuringiensis produce cristales proteicos con actividad citotóxica en contra de insectos y nematodos. La toxicidad de B. thuringiensis en plagas agrícolas es ampliamente conocida, pero poco se conoce acerca de su actividad en contra de nematodos parásitos. Recientemente, la actividad nematicida de las proteínas derivadas de B. thuringiensis se demostró en parásitos de mamíferos como Haemonchus, Teladorsagia, Nippostrongylus, y Ancylostoma, y en nematodos de plantas, Globodera y Meloidogyne. Entre el grupo de B. thuringiensis con efecto nematicida, las proteínas de la cepa IB-16 han mostrado actividad letal de 50 a 100 % en contra de diferentes estadios del principal género de rumiantes, Haemonchus contortus. Asimismo, los géneros de nematodos de vida libre, Panagrellus redivivus y Caenorhabditis elegans han sido blanco de estudios de la acción nematicida de B. thuringiensis. Por ejemplo, el efecto tóxico de la proteína Cry5B de B. thuringiensis se observó en las células intestinales de C. elegans, además esta acción parece involucrar receptores celulares específicos, similares a los que se han notificado en contra de plagas agrícolas. Asimismo, la unión de la proteína Cry5B ocurre en receptores específicos, como moléculas de carbohidratos, las cuales están presentes en la membrana de las células de intestino de los nematodos, ocasionando daño y muerte. A través de este tipo de estudios, los derivados de B. thuringiensis podrían considerarse una alternativa de control en nematodos que afectan a los animales domésticos, como rumiantes, así como en contra de otros nematodos patógenos de mamíferos e incluso de plantas agrícolas.

  12. PENGARUH KETINGGIAN HABITAT KELAPA (Cocos nucifera TERHADAP PENGEMBANGBIAKAN Bacillus thuringiensis H-14 DAN TOKSISITASNYA TERHADAP JENTIK (Anopheles aconitus

    Directory of Open Access Journals (Sweden)

    Lulus Susanti

    2013-05-01

    Full Text Available Abstrak Penggunaan Bacillus thuringiensis H-14 sebagai biolarvasida sudah banyak diketahui di masyarakat. Balai Besar Penelitian dan Pengembangan Vektor dan Reservoir Penyakit (B2P2VRP telah menguji penggunaan media buah kelapa sebagai media pengembangbiakan B.thuringiensis H-14. Pada penelitian ini dilakukan pengembangbiakkan B.thuringiensis H-14 galur lokal dengan menggunakan media air kelapa yang diambil dari berbagai wilayah dengan memperhatikan ketinggian wilayahnya. Lokasi pengambilan sampel air kelapa adalah daerah dengan ketinggian < 20 m dpl (pantai Parangtritis, 21 – 250 m dpl (Kabupaten Purworejo, 251 – 500 m dpl (Kabupaten Semarang dan 501 – 750 m dpl (Kota Salatiga. Penelitian ini dilakukan dengan mengambil air buah kelapa hijau umur 4 – 6 bulan atau berat mencapai > 600 gram dari masing-masing wilayah penelitian. Kemudian air kelapa dari masing-masing wilayah penelitian diambil secara random untuk dilakukan pengujian kandungan nutrisinya, sedangkan sisanya di sterilisasi untuk dijadikan media pengembangbiakan B.thuringiensis H-14. Tujuan dari penelitian ini untuk mengetahui ada atau tidaknya perbedaan hasil pengembangbiakan di media air kelapa yang didapatkan dari lokasi dengan ketinggian habitat yang berbeda-beda, serta efek toksisitasnya terhadap jentik Anopheles aconitus. Hasil uji analisa air kelapa dari pantai Parangtritis adalah kadar karbohidrat 1,82%, dengan lemak 0,02%, protein 0,04% dan gula reduksi sebesar 1,67%. Air kelapa dari kabupaten Purworejo kandungan karbohidrat 1,92%, lemak 0,01%. protein 0,06% dan gula reduksi 1,87%. Air kelapa dari Kabupaten Semarang kandungan karbohidrat 1,68%, lemak 0,01%, protein 0,12% dan gula reduksi 1,52%. Sedangkan kandungan karbohidrat dari air kelapa kota Salatiga adalah 3,12% merupakan kandungan yang tertinggi dibandingkan dari daerah lain, kandungan lemak 0,01%, protein 0,11% dan gula reduksi 2,97%, merupakan kandungan tertinggi dibandingkan dengan hasil dari daerah lain

  13. Production of Chymotrypsin-Resistant Bacillus thuringiensis Cry2Aa1 δ-Endotoxin by Protein Engineering

    OpenAIRE

    Audtho, Mongkon; Valaitis, Algimantas P.; Alzate, Oscar; Dean, Donald H.

    1999-01-01

    Cleavage of the Cry2Aa1 protoxin (molecular mass, 63 kDa) from Bacillus thuringiensis by midgut juice of gypsy moth (Lymantria dispar) larvae resulted in two major protein fragments: a 58-kDa fragment which was highly toxic to the insect and a 49-kDa fragment which was not toxic. In the midgut juice, the protoxin was processed into a 58-kDa toxin within 1 min, but after digestion for 1 h, the 58-kDa fragment was further cleaved within domain I, resulting in the protease-resistant 49-kDa fragm...

  14. Chimeric vip3Aa16TC Gene Encoding the Toxic Core of the Vegetative Insecticidal Protein Enhanced Bacillus thuringiensis Entomopathogenicity

    OpenAIRE

    Sameh Sellami; Maroua Cherif; Samir Jaoua; Kaïs Jamoussi

    2015-01-01

    Vip3 insecticidal protein is produced by Bacillus thuringiensis during the vegetative stage. Its proteolysis by the midgut juice of susceptible larvae formed four major products of approximately 66, 45, 33 and 22 kDa. In this study, we cloned the vip3Aa16TC DNA encoding the “Vip3Aa16 toxic core (TC)” of 33 kDa corresponding to the Vip3Aa16 region from amino acid 200 to 456. The vip3Aa16TC chimeric gene carried by the pHT-vip3Aa16TC plasmid was under the control of the sporulation ...

  15. Enhancement of intrinsic antitumor activity in spore-endotoxin mixtures of Bacillus thuringiensis by exposure to ultraviolet radiation

    International Nuclear Information System (INIS)

    Zamola, B.; Karminski-Zamola, G.; Fuks, Z.; Kubovic, M.; Wrishcer, M.

    1985-01-01

    Irradiation of spore-endotoxin mixtures from Bacillus thuringiensis cultures at 254 nm (60 μW cm -2 ) enhances their intrinsic antitumor potency as well as that of either component. The extent of enhancement depends on the length of exposure (optimum: 35 min) and may thus be due to photochemical changes of the endotoxin protein or/and to photoproduction of additional compounds with antitumor activity. Antitumor effects, expressed as survival rates of C57BL/6 mice inoculated with Lewis' mouse lung carcinoma and subjected to treatments 24 h later, depended on the number of doses of preparations administered (mixture, separated components). (author)

  16. Estudio de las bases de la resistencia a las proteínas insecticidas de bacillus thuringiensis en ostrinia nubilalis

    OpenAIRE

    Crava, Maria C.

    2013-01-01

    Ostrinia nubilalis (Hübner) es una de las plagas más devastadoras de los cultivos de maíz de Europa y Norte América, y a nivel económico la obtención de un control eficaz de esta plaga es un logro fundamental. En 1996, se permitió la comercialización de las plantas transgénicas que llevan insertado en el genoma un gen procedente de Bacillus thuringiensis (Berliner) (Bt) y que codifica para una proteína insecticida de las que esta bacteria produce en forma de cristales paraesporales (proteínas...

  17. Operational Evaluation Of Vectomax® WSP (Bacillus thuringiensis Subsp. israelensis+Bacillus sphaericus) Against Larval Culex pipiens in Septic Tanks (1).

    Science.gov (United States)

    Cetin, Huseyin; Oz, Emre; Yanikoglu, Atila; Cilek, James E

    2015-06-01

    The residual effectiveness of VectoMax® WSP (a water-soluble pouch formulation containing a combination of Bacillus thuringiensis subsp. israelensis strain AM65-52 and B. sphaericus strain ABTS 1743) when applied to septic tanks against 3rd- and 4th-stage larvae of Culex pipiens L. was evaluated in this study. This formulation was evaluated at operational application rates of 1 pouch (10 g) and 2 pouches (20 g) per septic tank. Both application rates resulted in >96% control of larvae for 24 days. Operationally, VectoMax WSP has proven to be a useful tool for the nonchemical control of Culex species in septic tank environments.

  18. pH-Controlled Bacillus thuringiensis Cry1Ac Protoxin Loading and Release from Polyelectrolyte Microcapsules

    Science.gov (United States)

    Yang, Wenhui; He, Kanglai; Zhang, Jie; Guo, Shuyuan

    2012-01-01

    Crystal proteins synthesized by Bacillus thuringiensis (Bt) have been used as biopesticides because of their toxicity to the insect larval hosts. To protect the proteins from environmental stress to extend their activity, we have developed a new microcapsule formulation. Poly (acrylic acid) (PAH) and poly (styrene sulfonate) (PSS) were fabricated through layer-by-layer self-assembly based on a CaCO3 core. Cry1Ac protoxins were loaded into microcapsules through layer-by-layer self-assembly at low pH, and the encapsulated product was stored in water at 4°C. Scanning electron microscopy (SEM) was used to observe the morphology of the capsules. To confirm the successful encapsulation, the loading results were observed with a confocal laser scattering microscope (CLSM), using fluorescein-labeled Cry1Ac protoxin (FITC-Cry1Ac). The protoxins were released from the capsule under the alkaline condition corresponding to the midgut of certain insects, a condition which seldom exists elsewhere in the environment. The following bioassay experiment demonstrated that the microcapsules with Cry1Ac protoxins displayed approximately equivalent insecticidal activity to the Asian corn borer compared with free Cry1Ac protoxins, and empty capsules proved to have no effect on insects. Further result also indicated that the formulation could keep stable under the condition of heat and desiccation. These results suggest that this formulation provides a promising methodology that protects protoxins from the environment and releases them specifically in the target insects’ midgut, which has shown potential as biopesticide in the field. PMID:23024810

  19. [Experimental observation of toxic effect of Bacillus thuringiensis var. israelensis against Aedes, Culex and Anopheles larvae].

    Science.gov (United States)

    Li, Ju-Lin; Zhu, Guo-Ding; Zhou, Hua-Yun; Tang, Jian-Xia; Cao, Jun

    2014-02-01

    To evaluate the toxic effect of Bacillus thuringiensis var. israelensis (Bti) wettable powder against Aedes, Culex and Anopheles larvae. The biological assay was applied to test the lethal concentration of 50% (LC50) of Bti wettable powder against Aedes, Culex and Anopheles larvae. The LC50(s) of Bti wettable powder against Aedes albopictus, Culex pipiens pallens and Anopheles sinensis larvae were 0.104, 0.160 microg/ml and 0.324 microg/ml, respectively; its biological potencies against them were 0.125, 0.192 IU/ml and 0.389 IU/ml, respectively. The LC50(s) of continuous contact of Bti wettable powder with An. sinensis stage III larvae for 1, 2 d and 3 d were 0.324, 0.092 microg/ml and 0.032 microg/ml, respectively, and its biological potencies were 0.389, 0.110 IU/ml and 0.038 IU/ml, respectively. The LC50(s) of the bacteria against An. sinensis stage I , II, III, IV were 0.024, 0.137, 0.324 microg/ml and 0.450 microg/ml, respectively, and the biological potencies were 0.029, 0.164, 0.389 IU/ml and 0.540 IU/ml, respectively. Bti wettable powder has a good toxicity to Aedes, Culex and Anopheles larvae, especially for the latter two. It is better to apply the bacteria at the early stage of mosquito larvae.

  20. Persistence of detectable insecticidal proteins from Bacillus thuringiensis (Cry) and toxicity after adsorption on contrasting soils

    International Nuclear Information System (INIS)

    Hung, T.P.; Truong, L.V.; Binh, N.D.; Frutos, R.; Quiquampoix, H.; Staunton, S.

    2016-01-01

    Insecticidal Cry, or Bt, proteins are produced by the soil-endemic bacterium, Bacillus thuringiensis and some genetically modified crops. Their environmental fate depends on interactions with soil. Little is known about the toxicity of adsorbed proteins and the change in toxicity over time. We incubated Cry1Ac and Cry2A in contrasting soils subjected to different treatments to inhibit microbial activity. The toxin was chemically extracted and immunoassayed. Manduca sexta was the target insect for biotests. Extractable toxin decreased during incubation for up to four weeks. Toxicity of Cry1Ac was maintained in the adsorbed state, but lost after 2 weeks incubation at 25 °C. The decline in extractable protein and toxicity were much slower at 4 °C with no significant effect of soil sterilization. The major driving force for decline may be time-dependent fixation of adsorbed protein, leading to a decrease in the extraction yield in vitro, paralleled by decreasing solubilisation in the larval gut. - Graphical abstract: Biotest, presenting Cry-contaminated feed to Manduca sexta larvae in individual Perspex boxes. Display Omitted - Highlights: • Toxicity of Cry protein is initially conserved after adsorption on soil. • Toxicity and extractability decline with time, more rapidly at 25 °C than 4 °C. • Similar dynamics of Cry1AC and Cry2A on soil with varying texture and organic C. • Sterilization of soil does not change Cry dynamics or temperature effect in soil. • Cry decline is determined by progressive fixation on soil not microbial breakdown. - Toxicity was initially maintained after adsorption on soil and both extractable Cry and toxicity declined rapidly, more slowly at low temperature, due to different fixation dynamics. Toxicity of Cry protein is initially conserved after adsorption on soil.

  1. pH regulates pore formation of a protease activated Vip3Aa from Bacillus thuringiensis.

    Science.gov (United States)

    Kunthic, Thittaya; Watanabe, Hirokazu; Kawano, Ryuji; Tanaka, Yoshikazu; Promdonkoy, Boonhiang; Yao, Min; Boonserm, Panadda

    2017-11-01

    Vip3Aa insecticidal protein is produced from Bacillus thuringiensis and exerts a broad spectrum of toxicity against lepidopteran insect species. Although Vip3Aa has been effectively used as part of integrated pest management strategies, the mechanism of the toxin remains unclear. Here, we investigated the effect of pH in a range from 5.0 to 10.0 on the pore-forming activity of the trypsin activated Vip3Aa (actVip3Aa) by in vitro pore-forming assays. Based on calcein release assay, actVip3Aa could permeabilize the artificial neutral liposomes under all the pH tested, except pH10.0. The maximum membrane permeability of actVip3Aa was detected at pH8.0 and the permeability decreased and abolished when exposing to acidic and alkaline conditions, respectively. The planar lipid bilayer experiment revealed that actVip3Aa formed ion channels at pH5.0-8.0 but no current signals were detected at pH10.0, consistent with the observation from calcein release assay. The toxin formed ion channels with a diameter of 1.4nm at pH8.0 and pore size was gradually decreased when reducing the pH. This study provided a view of the molecular mechanism of Vip3Aa by which the pore-forming activity is regulated by pH. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. An ABC transporter mutation is correlated with insect resistance to Bacillus thuringiensis Cry1Ac toxin.

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    Linda J Gahan

    2010-12-01

    Full Text Available Transgenic crops producing insecticidal toxins from Bacillus thuringiensis (Bt are commercially successful in reducing pest damage, yet knowledge of resistance mechanisms that threaten their sustainability is incomplete. Insect resistance to the pore-forming Cry1Ac toxin is correlated with the loss of high-affinity, irreversible binding to the mid-gut membrane, but the genetic factors responsible for this change have been elusive. Mutations in a 12-cadherin-domain protein confer some Cry1Ac resistance but do not block this toxin binding in in vitro assays. We sought to identify mutations in other genes that might be responsible for the loss of binding. We employed a map-based cloning approach using a series of backcrosses with 1,060 progeny to identify a resistance gene in the cotton pest Heliothis virescens that segregated independently from the cadherin mutation. We found an inactivating mutation of the ABC transporter ABCC2 that is genetically linked to Cry1Ac resistance and is correlated with loss of Cry1Ac binding to membrane vesicles. ABC proteins are integral membrane proteins with many functions, including export of toxic molecules from the cell, but have not been implicated in the mode of action of Bt toxins before. The reduction in toxin binding due to the inactivating mutation suggests that ABCC2 is involved in membrane integration of the toxin pore. Our findings suggest that ABC proteins may play a key role in the mode of action of Bt toxins and that ABC protein mutations can confer high levels of resistance that could threaten the continued utilization of Bt-expressing crops. However, such mutations may impose a physiological cost on resistant insects, by reducing export of other toxins such as plant secondary compounds from the cell. This weakness could be exploited to manage this mechanism of Bt resistance in the field.

  3. An ABC transporter mutation is correlated with insect resistance to Bacillus thuringiensis Cry1Ac toxin.

    Science.gov (United States)

    Gahan, Linda J; Pauchet, Yannick; Vogel, Heiko; Heckel, David G

    2010-12-16

    Transgenic crops producing insecticidal toxins from Bacillus thuringiensis (Bt) are commercially successful in reducing pest damage, yet knowledge of resistance mechanisms that threaten their sustainability is incomplete. Insect resistance to the pore-forming Cry1Ac toxin is correlated with the loss of high-affinity, irreversible binding to the mid-gut membrane, but the genetic factors responsible for this change have been elusive. Mutations in a 12-cadherin-domain protein confer some Cry1Ac resistance but do not block this toxin binding in in vitro assays. We sought to identify mutations in other genes that might be responsible for the loss of binding. We employed a map-based cloning approach using a series of backcrosses with 1,060 progeny to identify a resistance gene in the cotton pest Heliothis virescens that segregated independently from the cadherin mutation. We found an inactivating mutation of the ABC transporter ABCC2 that is genetically linked to Cry1Ac resistance and is correlated with loss of Cry1Ac binding to membrane vesicles. ABC proteins are integral membrane proteins with many functions, including export of toxic molecules from the cell, but have not been implicated in the mode of action of Bt toxins before. The reduction in toxin binding due to the inactivating mutation suggests that ABCC2 is involved in membrane integration of the toxin pore. Our findings suggest that ABC proteins may play a key role in the mode of action of Bt toxins and that ABC protein mutations can confer high levels of resistance that could threaten the continued utilization of Bt-expressing crops. However, such mutations may impose a physiological cost on resistant insects, by reducing export of other toxins such as plant secondary compounds from the cell. This weakness could be exploited to manage this mechanism of Bt resistance in the field.

  4. Bacillus thuringiensis-derived Cry5B has potent anthelmintic activity against Ascaris suum.

    Science.gov (United States)

    Urban, Joseph F; Hu, Yan; Miller, Melanie M; Scheib, Ulrike; Yiu, Ying Y; Aroian, Raffi V

    2013-01-01

    Ascaris suum and Ascaris lumbricoides are two closely related geo-helminth parasites that ubiquitously infect pigs and humans, respectively. Ascaris suum infection in pigs is considered a good model for A. lumbricoides infection in humans because of a similar biology and tissue migration to the intestines. Ascaris lumbricoides infections in children are associated with malnutrition, growth and cognitive stunting, immune defects, and, in extreme cases, life-threatening blockage of the digestive tract and aberrant migration into the bile duct and peritoneum. Similar effects can be seen with A. suum infections in pigs related to poor feed efficiency and performance. New strategies to control Ascaris infections are needed largely due to reduced treatment efficacies of current anthelmintics in the field, the threat of resistance development, and the general lack of new drug development for intestinal soil-transmitted helminths for humans and animals. Here we demonstrate for the first time that A. suum expresses the receptors for Bacillus thuringiensis crystal protein and novel anthelmintic Cry5B, which has been previously shown to intoxicate hookworms and which belongs to a class of proteins considered non-toxic to vertebrates. Cry5B is able to intoxicate A. suum larvae and adults and triggers the activation of the p38 mitogen-activated protein kinase pathway similar to that observed with other nematodes. Most importantly, two moderate doses of 20 mg/kg body weight (143 nM/kg) of Cry5B resulted in a near complete cure of intestinal A. suum infections in pigs. Taken together, these results demonstrate the excellent potential of Cry5B to treat Ascaris infections in pigs and in humans and for Cry5B to work effectively in the human gastrointestinal tract.

  5. Heterologous expression, purification and biochemical characterization of endochitinase ChiA74 from Bacillus thuringiensis.

    Science.gov (United States)

    Casados-Vázquez, Luz Edith; Avila-Cabrera, Salvador; Bideshi, Dennis K; Barboza-Corona, J Eleazar

    2015-05-01

    ChiA74 is a secreted endochitinase produced by Bacillus thuringiensis. Previously we have partially characterized the physical parameters that affect enzymatic activity of ChiA74 in crude preparations of bacterial secretomes. In the present study, we cloned the chiA74 open reading frame (ORF) lacking the 5' sequence coding for its secretion signal peptide (chiA74Δsp) into a cold shock expression vector (pColdI) for production of the enzyme in Escherichia coli BL21-Rosetta 2. As a result, the N-terminal end of ChiA74Δsp ORF was fused to an artificial sequence of 28 amino acid, including a 6× histidine tag for purification of recombinant 6×His tagged-ChiA74Δsp (rChiA74, ∼74kDa). Along with a protein of ∼74kDa, we co-purified its ∼55kDa processed form which was confirmed by Western blot analysis. Optimal endochitinase activity of purified rChiA74 occurred at pH 7 and 40°C. Most divalent cations (e.g. Ba(+2), Ca(+2), Mn(+2), Mg(+2), Zn(+2) and Cu(+2)) at concentration of 10mM reduced chitinase activity by ∼30%, and Hg(+2) (10mM) drastically inhibited ChiA74 activity by ∼75-100%. The Vmax, Km and kcat for rChiA74 were 0.11±0.01nmol/min, 2.15μM±0.45 and 3.81s(-1), respectively, using 4-MU-GlcNAc3 as substrate. Using purified rChiA74 and colloidal chitin as substrate, chitin-derived oligosaccharides with degree of polymerization of 2 and 1 were detected. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Bacillus thuringiensis: mechanism of action, resistance, and new applications: a review.

    Science.gov (United States)

    Melo, André Luiz de Almeida; Soccol, Vanete Thomaz; Soccol, Carlos Ricardo

    2016-01-01

    Since the first report by Ishiwata in 1902 of a Bombyx mori infection, followed by the description by Berliner, Bacillus thuringiensis (Bt) has become the main microorganism used in biological control. The application of Bt to combat invertebrates of human interest gained momentum with the growing demand for food free of chemical pesticides and with the implementation of agriculture methods that were less damaging to the environment. However, the mechanisms of action of these products have not been fully elucidated. There are two proposed models: the first is that Bt causes an osmotic imbalance in response to the formation of pores in a cell membrane, and the second is that it causes an opening of ion channels that activate the process of cell death. There are various ways in which Bt resistance can develop: changes in the receptors that do not recognize the Cry toxin, the synthesis of membrane transporters that eliminate the peptides from the cytosol and the development of regulatory mechanisms that disrupt the production of toxin receptors. Besides the potential for formulation of biopesticides and the use in developing genetically modified cultivars, recent studies with Bt have discussed promising applications in other branches of science. Chitinase, an enzyme that degrades chitin, increases the efficiency of Bt insecticides, and there has been of increasing interest in the industry, given that its substrate is extremely abundant in nature. Another promising field is the potential for Bt proteins to act against cancer cells. Parasporins, toxins of Bt that do not have an entomopathogenic effect, have a cytotoxic effect on the cells changed by some cancers. This demonstrates the potential of the microorganism and new opportunities opening for future applications.

  7. Density Dependence and Growth Rate: Evolutionary Effects on Resistance Development to Bt (Bacillus thuringiensis).

    Science.gov (United States)

    Martinez, Jeannette C; Caprio, Michael A; Friedenberg, Nicholas A

    2018-02-09

    It has long been recognized that pest population dynamics can affect the durability of a pesticide, but dose remains the primary component of insect resistance management (IRM). For transgenic pesticidal traits such as Bt (Bacillus thuringiensis Berliner (Bacillales: Bacillaceae)), dose (measured as the mortality of susceptibles caused by a toxin) is a relatively fixed characteristic and often falls below the standard definition of high dose. Hence, it is important to understand how pest population dynamics modify durability and what targets they present for IRM. We used a deterministic model of a generic arthropod pest to examine how timing and strength of density dependence interacted with population growth rate and Bt mortality to affect time to resistance. As in previous studies, durability typically reached a minimum at intermediate doses. However, high population growth rates could eliminate benefits of high dose. The timing of density dependence had a more subtle effect. If density dependence operated simultaneously with Bt mortality, durability was insensitive to its strengths. However, if density dependence was driven by postselection densities, decreasing its strength could increase durability. The strength of density dependence could affect durability of both single traits and pyramids, but its influence depended on the timing of density dependence and size of the refuge. Our findings suggest the utility of a broader definition of high dose, one that incorporates population-dynamic context. That maximum growth rates and timing and strength of interactions causing density dependent mortality can all affect durability, also highlights the need for ecologically integrated approaches to IRM research. © The Author(s) 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  8. Lipopeptide biosurfactant from Bacillus thuringiensis pak2310: A potential antagonist against Fusarium oxysporum.

    Science.gov (United States)

    Deepak, R; Jayapradha, R

    2015-03-01

    The aims of the study were to evaluate the effects of a biosurfactant obtained from a novel Bacillus thuringiensis on Fusarium oxysporum to determine the morphological changes in the structure of the fungi and its biofilm in the presence of the biosurfactant and to evaluate the toxicity of the biosurfactant on HEp-2 human epithelial cell lines. The strain was screened and isolated from petroleum contaminated soil based on the E24 emulsification index. The biosurfactant was produced on glycerol, extracted using chloroform:methanol system and purified using HPLC. The purified fraction showing both surface activity (emulsification and oil-spread activity) and anti-fusarial activity (agar well diffusion method) was studied using FT-IR and MALDI-TOF MS, respectively. The minimum inhibitory concentration (MIC) and the biofilm inhibitory concentration (BIC) were determined using dilution method. The effect of biosurfactant on the morphology of Fusarium oxysporum was monitored using light microscopy and confocal laser scanning microscopy (for biofilm). The purified surfactant showed the presence of functional groups like that of surfactin in the FT-IR spectra and MALDI-TOF MS estimated the molecular weight as 700Da. The MIC and BIC were estimated to be 0.05 and 0.5mg/mL, respectively. The molecule was also non-toxic to HEp-2 cell lines at 10× MIC. A non-toxic and effective anti-Fusarium biosurfactant, that is both safe for human use and to the environment, has been characterized. The growth and metabolite production using glycerol (major byproduct of biodiesel and soap industries) also adds up to the efficiency and ecofriendly nature of this biosurfactant. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  9. Pathogenicity of Bacillus thuringiensis isolated from two species of Acromyrmex (Hymenoptera, Formicidae

    Directory of Open Access Journals (Sweden)

    L. M. N. Pinto

    Full Text Available The control of Acromyrmex leaf-cutting ants is necessary due to the severe damage they cause to diverse crops. A possibility was to control them using the bacterium Bacillus thuringiensis (Bt that characteristically produces insecticidal crystal proteins (ICPs. The ICPs have been effective in controlling lepidopterans, dipterans, and coleopterans, but their action against hymenopterans is unknown. This paper describes an attempt to isolate Bt from ants of two Acromyrmex species, to evaluate its pathogenicity towards these ants, and to test isolates by PCR. Bacterial isolates of Bt obtained from A. crassispinus and A. lundi have been assayed against A. lundi in the laboratory. The bioassays were carried out in BOD at 25°C, with a 12-hour photoperiod, until the seventh day after treatment. The Bt isolates obtained were submitted to total DNA extraction and tested by PCR with primers specific to cry genes. The results showed Bt presence in 40% of the assessed samples. The data from the in vivo assays showed a mortality rate higher than 50% in the target population, with the Bt HA48 isolate causing 100% of corrected mortality. The PCR results of Bt isolates showed a magnification of DNA fragments relative to cry1 genes in 22% of the isolates, and cry9 in 67%. Cry2, cry3, cry7, and cry8 genes were not detected in the tested samples, and 22% had no magnified DNA fragments corresponding to the assessed cry genes. The results are promising not only regarding allele identification in new isolates, but also fort the assays aimed at determining the Bt HA48 LC50's, which can eventually be applied in controlling of Acromyrmex leaf-cutting ants.

  10. Bacillus thuringiensis Cyt2Aa2 toxin disrupts cell membranes by forming large protein aggregates.

    Science.gov (United States)

    Tharad, Sudarat; Toca-Herrera, José L; Promdonkoy, Boonhiang; Krittanai, Chartchai

    2016-10-01

    Bacillus thuringiensis (Bt) Cyt2Aa2 showed toxicity against Dipteran insect larvae and in vitro lysis activity on several cells. It has potential applications in the biological control of insect larvae. Although pore-forming and/or detergent-like mechanisms were proposed, the mechanism underlying cytolytic activity remains unclear. Analysis of the haemolytic activity of Cyt2Aa2 with osmotic stabilizers revealed partial toxin inhibition, suggesting a distinctive mechanism from the putative pore formation model. Membrane permeability was studied using fluorescent dye entrapped in large unilamellar vesicles (LUVs) at various protein/lipid molar ratios. Binding of Cyt2Aa2 monomer to the lipid membrane did not disturb membrane integrity until the critical protein/lipid molar ratio was reached, when Cyt2Aa2 complexes and cytolytic activity were detected. The complexes are large aggregates that appeared as a ladder when separated by agarose gel electrophoresis. Interaction of Cyt2Aa2 with Aedes albopictus cells was investigated by confocal microscopy and total internal reflection fluorescent microscopy (TIRF). The results showed that Cyt2Aa2 binds on the cell membrane at an early stage without cell membrane disruption. Protein aggregation on the cell membrane was detected later which coincided with cell swelling. Cyt2Aa2 aggregations on supported lipid bilayers (SLBs) were visualized by AFM. The AFM topographic images revealed Cyt2Aa2 aggregates on the lipid bilayer at low protein concentration and subsequently disrupts the lipid bilayer by forming a lesion as the protein concentration increased. These results supported the mechanism whereby Cyt2Aa2 binds and aggregates on the lipid membrane leading to the formation of non-specific hole and disruption of the cell membrane. © 2016 The Author(s).

  11. Production of wetland Chironomidae (Diptera) and the effects of using Bacillus thuringiensis israelensis for mosquito control.

    Science.gov (United States)

    Lundström, J O; Schäfer, M L; Petersson, E; Persson Vinnersten, T Z; Landin, J; Brodin, Y

    2010-02-01

    Massive mosquito nuisance problems, caused by the flood-water mosquito Aedes sticticus, occur after floods in the flood plains of the River Dalälven, central Sweden. Since 2002, the biological mosquito larvicide Bacillus thuringiensis var. israelensis (Bti) has been used to control these mosquitoes. Here, we report results from the first six years of monitoring Chironomidae, the most susceptible non-target organisms, in three wetlands with Bti-treatment against mosquitoes and in three wetlands without treatment. Emergence traps were used for continuous insect sampling from May to September each year, 2002-2007, and were emptied once a week. A total of 21,394 chironomids of 135 species were collected, and the subfamily Orthocladiinae dominated the fauna followed by Chironominae and Tanypodinae. The annual chironomid production in the wetlands was low, with an average of 1917 individuals per m(2), and 42 g ash-free dry weight per m(2). We found no reduced production of chironomids at neither family, nor subfamily level, in Bti-treated as compared to untreated wetlands. This is the first long-term follow-up study of the possible effects of Bti-based mosquito larval control on chironomid species production. In the short-term view, one species had higher production in treated areas. In the long-term view, four species had higher and one species had lower production in treated areas. We conclude that the Bti-based control of flood-water mosquitoes does not cause any major direct negative effects on chironomid production, and therefore does not seem to induce any risk for indirect negative effects on birds, bats or any other predators feeding on chironomids.

  12. Improvement and efficient display of Bacillus thuringiensis toxins on M13 phages and ribosomes.

    Science.gov (United States)

    Pacheco, Sabino; Cantón, Emiliano; Zuñiga-Navarrete, Fernando; Pecorari, Frédéric; Bravo, Alejandra; Soberón, Mario

    2015-12-01

    Bacillus thuringiensis (Bt) produces insecticidal proteins that have been used worldwide in the control of insect-pests in crops and vectors of human diseases. However, different insect species are poorly controlled by the available Bt toxins or have evolved resistance to these toxins. Evolution of Bt toxicity could provide novel toxins to control insect pests. To this aim, efficient display systems to select toxins with increased binding to insect membranes or midgut proteins involved in toxicity are likely to be helpful. Here we describe two display systems, phage display and ribosome display, that allow the efficient display of two non-structurally related Bt toxins, Cry1Ac and Cyt1Aa. Improved display of Cry1Ac and Cyt1Aa on M13 phages was achieved by changing the commonly used peptide leader sequence of the coat pIII-fusion protein, that relies on the Sec translocation pathway, for a peptide leader sequence that relies on the signal recognition particle pathway (SRP) and by using a modified M13 helper phage (Phaberge) that has an amber mutation in its pIII genomic sequence and preferentially assembles using the pIII-fusion protein. Also, both Cry1Ac and Cyt1Aa were efficiently displayed on ribosomes, which could allow the construction of large libraries of variants. Furthermore, Cry1Ac or Cyt1Aa displayed on M13 phages or ribosomes were specifically selected from a mixture of both toxins depending on which antigen was immobilized for binding selection. These improved systems may allow the selection of Cry toxin variants with improved insecticidal activities that could counter insect resistances.

  13. Field-evolved resistance by western corn rootworm to multiple Bacillus thuringiensis toxins in transgenic maize.

    Science.gov (United States)

    Gassmann, Aaron J; Petzold-Maxwell, Jennifer L; Clifton, Eric H; Dunbar, Mike W; Hoffmann, Amanda M; Ingber, David A; Keweshan, Ryan S

    2014-04-08

    The widespread planting of crops genetically engineered to produce insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt) places intense selective pressure on pest populations to evolve resistance. Western corn rootworm is a key pest of maize, and in continuous maize fields it is often managed through planting of Bt maize. During 2009 and 2010, fields were identified in Iowa in which western corn rootworm imposed severe injury to maize producing Bt toxin Cry3Bb1. Subsequent bioassays revealed Cry3Bb1 resistance in these populations. Here, we report that, during 2011, injury to Bt maize in the field expanded to include mCry3A maize in addition to Cry3Bb1 maize and that laboratory analysis of western corn rootworm from these fields found resistance to Cry3Bb1 and mCry3A and cross-resistance between these toxins. Resistance to Bt maize has persisted in Iowa, with both the number of Bt fields identified with severe root injury and the ability western corn rootworm populations to survive on Cry3Bb1 maize increasing between 2009 and 2011. Additionally, Bt maize targeting western corn rootworm does not produce a high dose of Bt toxin, and the magnitude of resistance associated with feeding injury was less than that seen in a high-dose Bt crop. These first cases of resistance by western corn rootworm highlight the vulnerability of Bt maize to further evolution of resistance from this pest and, more broadly, point to the potential of insects to develop resistance rapidly when Bt crops do not achieve a high dose of Bt toxin.

  14. pH-controlled Bacillus thuringiensis Cry1Ac protoxin loading and release from polyelectrolyte microcapsules.

    Directory of Open Access Journals (Sweden)

    Wenhui Yang

    Full Text Available Crystal proteins synthesized by Bacillus thuringiensis (Bt have been used as biopesticides because of their toxicity to the insect larval hosts. To protect the proteins from environmental stress to extend their activity, we have developed a new microcapsule formulation. Poly (acrylic acid (PAH and poly (styrene sulfonate (PSS were fabricated through layer-by-layer self-assembly based on a CaCO(3 core. Cry1Ac protoxins were loaded into microcapsules through layer-by-layer self-assembly at low pH, and the encapsulated product was stored in water at 4°C. Scanning electron microscopy (SEM was used to observe the morphology of the capsules. To confirm the successful encapsulation, the loading results were observed with a confocal laser scattering microscope (CLSM, using fluorescein-labeled Cry1Ac protoxin (FITC-Cry1Ac. The protoxins were released from the capsule under the alkaline condition corresponding to the midgut of certain insects, a condition which seldom exists elsewhere in the environment. The following bioassay experiment demonstrated that the microcapsules with Cry1Ac protoxins displayed approximately equivalent insecticidal activity to the Asian corn borer compared with free Cry1Ac protoxins, and empty capsules proved to have no effect on insects. Further result also indicated that the formulation could keep stable under the condition of heat and desiccation. These results suggest that this formulation provides a promising methodology that protects protoxins from the environment and releases them specifically in the target insects' midgut, which has shown potential as biopesticide in the field.

  15. Susceptibility of Aedes aegypti larvae to temephos and Bacillus thuringiensis var israelensis in integrated control Susceptibilidade de larvas de Aedes aegypti ao tratamento integrado com temephos e Bacillus thuringiensis var israelensis

    Directory of Open Access Journals (Sweden)

    Carlos Fernando S. de Andrande

    1991-06-01

    Full Text Available The susceptibility of field collected Aedes aegypti larvae was evaluated in terms of median lethal time (LT50 and final mortality, when treated with temephos, Bacillus thuringiensis var israelensis as well as mixtures of these two agents. Third instar larvae were shown to be more susceptible than early and late fourth instar ones to the entomopathogen. Survival of some individuals when exposed to temephos suggest possible resistance. Temporal synergism in early fourth instar larvae was detected when they were exposed to mixtures of Bti-temephos. The possibility of this integrated treatment is commented on.A susceptibilidade de larvas de Aedes aegypti coletadas no campo foi avaliada em termos do tempo letal mediano (TL50 e da mortalidade final, quando tratadas com temephos, Bacillus thuringiensis var israelensis ou misturas desses dois agentes. As larvas de terceiro estádio mostraram-se mais suceptíveis ao patógeno do que aquelas no início ou no fim do quarto estádio. A sobrevivência de alguns indivíduos aos tratamentos com temephos permite sugerir a possibilidade de resistência. Foi detectada a existência de sinergismo temporal, quando larvas no início do quarto estádio foram tratadas com as misturas do Bti com o temephos. A possibilidade do tratamento integrado é comentada.

  16. EFIKASI DAN EFEK RESIDU LARVISIDA Bacillus thuringiensis H-14 FORMULASI GRANULER TERHADAP LARVA Culex quinguefasciatus DI LABORATORIUM

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    Astri Maharani I.P.

    2012-11-01

    Full Text Available Bacillus thuringiensis was one of the insect pathogen which would be developed into biolarvicide. The research was conducted to study the efficacy and the residual effect of B.thuringiensis serotype H-14 larvicide granular formula against Culex quinquefasciatus larvae in the laboratory. This research used an amount of 1 gr/50 lt, 0,5 gr/50 lt, 0,25 gr/50 lt, 0,125 gr/50 lt, 0,0625 gr/50 lt, 0,031 gr/50 lt, 0,015 gr/50 lt, 0,0078 gr/50 lt, 0,0039 gr/50 lt as doses of granular formula. Dosages were given with a repetition of three times. The result showed that B.thuringiensis H-14 larvicide granular formula which dozes of 1 gr/50 lt, 0,5 gr/50 lt, 0,25 gr/50 lt,0,125 gr/50 lt, 0,0625 gr/50 lt were effective to kill 100% of Cx. quinquefasciatus larvae after 24 hours exposure. Residual effect of B. thmingiensis H-14 larvicide granular formula could kill more than 70% larvae until as longer 4 weeks. Applying the anova analysis, there was asignificant different number of death larvae at different doses and at different times. Keywords: Effectiveness, residual effect, biolarvacide

  17. Plasmid patterns of efficient and inefficient strains of Bacillus thuringiensis against Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae).

    Science.gov (United States)

    Fagundes, R B S; Picoli, E A T; Lana, U G P; Valicente, Fernando H

    2011-10-01

    Bacillus thuringiensis harbors genes encoding Cry proteins found in chromosomes or plasmids of different sizes (4-150 Mb). Although the smaller plasmids are more abundant in B. thuringiensis, their specific function is unknown. As for the megaplasmids, their main recognized function is to harbor cry genes, although the sequencing of some of these plasmids indicates the occurrence of other important genes. This work used a new protocol for practical and rapid extraction of plasmid DNA in order to characterize the plasmid patterns of Brazilian strains belonging to Embrapa Milho e Sorgo research center B. thuringiensis bank. We tried to further assess the relationship of plasmid patterns with strains belonging to the same serovars and strains causing 100% and no mortality to Spodoptera frugiperda (J.E. Smith) larvae. It was possible to characterize 59 strains based on the migration of bands in agarose gel. Strains belonging to the same serovars showed different plasmid sizes (from 1,636 bp to 23,200 bp), with the exception of two strains belonging to serovar galleriae. The strain T09 Bt tolworthi showed a plasmid migration pattern identical to strains belonging to serovar galleriae. Plasmid patterns differed for 46 strains, confirming that this is a useful tool to discriminate specific strains. However, it was not possible to associate the plasmid pattern or the occurrence of particular plasmids with the pathogenicity of a given species towards S. frugiperda larvae.

  18. Small RNA-mediated Cry toxin silencing allows Bacillus thuringiensis to evade Caenorhabditis elegans avoidance behavioral defenses

    Science.gov (United States)

    Peng, Donghai; Luo, Xiaoxia; Zhang, Ni; Guo, Suxia; Zheng, Jinshui; Chen, Ling

    2018-01-01

    Abstract Pathogen avoidance behavior protects animal hosts against microbial pathogens. Pathogens have evolved specific strategies during coevolution in response to such host defenses. However, these strategies for combatting host avoidance behavioral defenses remain poorly understood. Here, we used Caenorhabditis elegans and its bacterial pathogen Bacillus thuringiensis as a model and determined that small RNA (sRNA)-mediated Cry toxin silencing allowed pathogens to evade host avoidance behavioral defenses. The B. thuringiensis strain YBT-1518, which encodes three nematicidal cry genes, is highly toxic to C. elegans. However, the expression of the most potent toxin, Cry5Ba, was silenced in this strain when YBT-1518 was outside the host. Cry5Ba silencing was due to the sRNA BtsR1, which bound to the RBS site of the cry5Ba transcript via direct base pairing and inhibited Cry5Ba expression. Upon ingestion by C. elegans, Cry5Ba was expressed in vivo by strain YBT-1518. Cry5Ba silencing may allow B. thuringiensis to avoid nematode behavioral defenses and then express toxins once ingested to kill the host and gain a survival advantage. Our work describes a novel model of sRNA-mediated regulation to aid pathogens in combating host avoidance behavioral defenses. PMID:29069426

  19. Crystalline protein profiling and cry gene detection in Bacillus thuringiensis strains isolated during epizootics in Cydia pomonella L.

    Directory of Open Access Journals (Sweden)

    Konecka Edyta

    2014-12-01

    Full Text Available The composition of Bacillus thuringiensis crystalline inclusions was characterized in 18 strains: 12 isolates were obtained from the intestinal tract of Cydia pomonella larvae during epizootics, 2 isolates were cultured from Leucoma salicis larvae taken from their natural populations, and 4 reference strains. The number and molecular mass of B. thuringiensis crystalline proteins (Cry and Cyt was estimated by the sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE. The crystals contained 1-8 proteins with molecular masses of 36-155 kDa. The toxin profiles differed both quantatively and qualitatively. The B. thuringiensis MPU B9 isolate had the highest number and diversity of Cry toxins. The analysis of crystal composition by SDS-PAGE was insufficient to detect groups and subgroups of Cry proteins. We identified 20 groups and 3 subgroups of Cry and Cyt crystalline toxins. Only one epizootic strain harboured cry25. In single reference strains, the cry1H, cry10 and cry25 genes were found. We did not find any correlation between the occurrence of cry genes and electrophoretic protein profiles of crystalline toxins.

  20. Low persistence of Bacillus thuringiensis serovar israelensis spores in four mosquito biotopes of a salt marsh in southern France.

    Science.gov (United States)

    Hajaij, Myriam; Carron, Alexandre; Deleuze, Julien; Gaven, Bruno; Setier-Rio, Marie-Laure; Vigo, Gerard; Thiéry, Isabelle; Nielsen-LeRoux, Christina; Lagneau, Christophe

    2005-11-01

    We studied the persistence of Bacillus thuringiensis serovar israelensis (Bti) in a typical breeding site of the mosquito Ochlerotatus caspius in a particularly sensitive salt marsh ecosystem following two Bti-based larvicidal applications (Vectobac 12AS, 1.95 L/ha). The treated area was composed of four larval biotopes that differed in terms of the most representative plant species (Sarcocornia fruticosa, Bolboschoenus maritimus, Phragmites australis, and Juncus maritimus) and the physical and chemical characteristics of the soil. We sampled water, soil, and plants at various times before and after the applications (from spring to autumn, 2001) and quantified the spores of B. thuringiensis (Bt) and Bacillus species. The B. cereus group accounted for between 0% and 20% of all Bacillus spp. before application depending on the larval biotope. No Bti were found before application. The variation in the quantity of bacilli during the mosquito breeding season depended more on the larval biotope than on the season or the larvicidal application. More bacilli were found in soil (10(4)-10(6) spores/g) than on plant samples (10(2)-10(4) spores/g). The abundance in water (10(5) to 10(7) spores/L) appeared to be correlated to the water level of the breeding site. The number of Bti spores increased just after application, after declining; no spores were detected in soil or water 3 months after application. However, low numbers of Bti spores were present on foliage from three of the four studied plant strata. In conclusion, the larvicidal application has very little impact on Bacillus spp. flora after one breeding season (two applications).