Science.gov

Sample records for bacillus thuringiensis cry2ab2

  1. 40 CFR 174.519 - Bacillus thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Cry2Ab2 protein... thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry2Ab2 protein in or on corn or cotton are exempt from the requirement of...

  2. 40 CFR 174.519 - Bacillus thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry2Ab2 protein... thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry2Ab2 protein in or on corn or cotton are exempt from the requirement of...

  3. 40 CFR 174.519 - Bacillus thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry2Ab2 protein... thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry2Ab2 protein in or on corn or cotton are exempt from the requirement of...

  4. 40 CFR 174.519 - Bacillus thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry2Ab2 protein... thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry2Ab2 protein in or on corn or cotton are exempt from the requirement of...

  5. 40 CFR 174.519 - Bacillus thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry2Ab2 protein in corn and cotton; exemption from the requirement of a tolerance. 174.519 Section 174.519 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED...

  6. Toxicity and characterization of cotton expressing Bacillus thuringiensis Cry1Ac and Cry2Ab2 proteins for control of lepidopteran pests.

    PubMed

    Sivasupramaniam, S; Moar, W J; Ruschke, L G; Osborn, J A; Jiang, C; Sebaugh, J L; Brown, G R; Shappley, Z W; Oppenhuizen, M E; Mullins, J W; Greenplate, J T

    2008-04-01

    Cry1Ac protoxin (the active insecticidal toxin in both Bollgard and Bollgard II cotton [Gossypium hirsutum L.]), and Cry2Ab2 toxin (the second insecticidal toxin in Bollgard II cotton) were bioassayed against five of the primary lepidopteran pests of cotton by using diet incorporation. Cry1Ac was the most toxic to Heliothis virescens (F.) and Pectinophora gossypiella (Saunders), demonstrated good activity against Helicoverpa zea (Boddie), and had negligible toxicity against Spodoptera exigua (Hübner) and Spodoptera frugiperda (J. E. Smith). Cry2Ab2 was the most toxic to P. gossypiella and least toxic to S. frugiperda. Cry2Ab2 was more toxic to S. exigua and S. frugiperda than Cry1Ac. Of the three insect species most sensitive to both Bacillus thuringiensis (Bt) proteins (including H. zea), P. gossypiella was only three-fold less sensitive to Cry2Ab2 than Cry1Ac, whereas H. virescens was 40-fold less sensitive to Cry2Ab2 compared with CrylAc. Cotton plants expressing Cry1Ac only and both Cry1Ac and Cry2Ab2 proteins were characterized for toxicity against H. zea and S.frugiperda larvae in the laboratory and H. zea larvae in an environmental chamber. In no-choice assays on excised squares from plants of different ages, second instar H. zea larvae were controlled by Cry1Ac/Cry2Ab2 cotton with mortality levels of 90% and greater at 5 d compared with 30-80% mortality for Cry1Ac-only cotton, depending on plant age. Similarly, feeding on leaf discs from Cry1Ac/Cry2Ab2 cotton resulted in mortality of second instars of S.frugiperda ranging from 69 to 93%, whereas exposure to Cry1Ac-only cotton yielded 20-69% mortality, depending on plant age. When cotton blooms were infested in situ in an environmental chamber with neonate H. zea larvae previously fed on synthetic diet for 0, 24, or 48 h, 7-d flower abortion levels for Cry1Ac-only cotton were 15, 41, and 63%, respectively, whereas for Cry1Ac/Cry2Ab2 cotton, flower abortion levels were 0, 0, and 5%, respectively. Cry1Ac and

  7. F2 screen for resistance to Bacillus thuringiensis Cry2Ab2-maize in field populations of Spodoptera frugiperda (Lepidoptera: Noctuidae) from the southern United States.

    PubMed

    Niu, Ying; Qureshi, Jawwad A; Ni, Xinzhi; Head, Graham P; Price, Paula A; Meagher, Robert L; Kerns, David; Levy, Ronnie; Yang, Xiangbing; Huang, Fangneng

    2016-07-01

    The fall armyworm, Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae), is a target pest of transgenic maize and cotton expressing Bacillus thuringiensis (Bt) proteins in both North and South America. In 2013 and 2014, a total of 215 F2 two-parent families of S. frugiperda were established using single-pair mating of field individuals collected from seven locations in four states of the southern U.S.: Texas, Louisiana, Georgia, and Florida. The objective of the investigation was to detect resistance alleles in field populations to Cry2Ab2, a common Bt protein produced in transgenic maize and cotton. For each F2 family, 128 F2 neonates were screened on leaf tissue of Cry2Ab2 maize plants in the laboratory. A conservative estimate of the frequency of major Cry2Ab2 resistance alleles in S. frugiperda from the four states was 0.0023 with a 95% credibility interval of 0.0003-0.0064. In addition, six families were considered to likely possess minor resistance alleles at a frequency of 0.0082 with a 95% credibility interval of 0.0033-0.0152. One F2 family from Georgia (GA-15) was confirmed to possess a major resistance allele to the Cry2Ab2 protein. Larvae from this family survived well on whole maize plants expressing Cry2Ab2 protein and demonstrated a significant level (>15-fold) of resistance when fed with the same protein incorporated in a meridic diet. The detection of the major resistance allele along with the relatively abundant minor resistance alleles revealed in this study may have important implications for resistance management. PMID:27311896

  8. F2 screen for resistance to Bacillus thuringiensis Cry2Ab2-maize in field populations of Spodoptera frugiperda (Lepidoptera: Noctuidae) from the southern United States

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The fall armyworm, Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae), is a target of transgenic maize and cotton expressing Bacillus thuringiensis (Bt) proteins in both North and South America. In 2013 and 2014, a total of 215 F2 two-parent families of S. frugiperda were established usin...

  9. Identification, inheritance, and fitness costs of Cry2Ab2 resistance in a field-derived population of sugarcane borer, Diatraea saccharalis (F.) (Lepidoptera: Crambidae).

    PubMed

    Huang, Fangneng; Chen, Mao; Gowda, Anilkumar; Clark, Thomas L; McNulty, Brain C; Yang, Fei; Niu, Ying

    2015-09-01

    The sugarcane borer, Diatraea saccharalis (F.), is one of the major target pests of transgenic maize, Zea mays, expressing Bacillus thuringiensis (Bt) proteins in South America and mid-southern region of the U.S. The MON89034 maize expresses Cry2Ab2 and Cry1A.105 Bt proteins and it was developed to provide better control of key lepidopteran pests of maize including D. saccharalis. The objectives of this study were to select and characterize the resistance of D. saccharalis to Cry2Ab2 using a non-commercial Cry2Ab2 single gene Bt maize line. A Cry2Ab2-resistant strain (Cry2Ab2-RR) of D. saccharalis was established from 28 two-parent families collected from fields in northeast Louisiana, U.S. The Cry2Ab2-RR showed a high level of resistance to Cry2Ab2 in both diet-incorporated and whole maize plant bioassays. The Cry2Ab2 resistance in D. saccharalis was likely inherited as a single or a few tightly linked autosomal genes. The resistance was non-recessive and not associated with fitness costs. The results should provide valuable information in resistance monitoring, assessing resistance risk, and developing effective management strategies for the sustainable use of Bt maize technology for managing maize stalk borers. PMID:26205174

  10. Susceptibility of bollworm and tobacco budworm (Lepidoptera: Noctuidae) to Cry2Ab2 insecticidal protein.

    PubMed

    Ali, M I; Luttrell, R G

    2007-06-01

    Susceptibilities of 82 bollworm, Helicoverpa zea (Boddie), and 44 tobacco budworm, Heliothis virescens (F.) (Lepidoptera: Noctuidae), populations to Cry2Ab2 protein were measured in diet incorporated assays at the University of Arkansas from 2002 to 2005. Resulting data were used to calculate overall (pooled data) estimates of species susceptibility for future benchmarks of resistance. Variabilities among populations also were studied by comparing regressions for individual populations and calculating mean susceptibilities for different subgroups of the colonies studied. Individual lethal concentration (LC50) estimates for nine laboratory, seven laboratory-cross, and 28 field populations of H. virescens varied up to 48-fold when adjusted for the response of the most susceptible laboratory colony studied. Mean susceptibilities of all laboratory, laboratory-cross, or field colonies varied only two-fold. When grouped by host plants, populations collected on tobacco, Nicotiana tabacum (L.), seemed to be less susceptible than those collected on other host plants. Individual LC50 values for 82 laboratory, laboratory-cross and field populations of H. zea varied up to 37-fold. Mean LC50 values of all laboratory, laboratory-cross, or field populations varied only three-fold. Susceptibilities of populations from Bollgard cotton were up to four-fold less than those from Bacillus thuringiensis corn, Zea mays L. Field populations collected during late season were generally less susceptible than those collected early in the season. Across the two species, H. zea was less sensitive to Cry2Ab2 than H. virescens. Both species seem to be less sensitive to Cry2Ab2 than to CrylAc. PMID:17598557

  11. Susceptibility of Cry1Ab-resistant and -susceptible Sugarcane Borer (Lepidoptera: crambidae) to Four Bacillus thuringiensis Toxins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sugarcane borer, Diatraea saccharalis (F.), is a primary corn stalk borer pest targeted by transgenic corn expressing Bacillus thuringiensis (Bt) proteins in many areas of the mid-southern region of the United States. Recently, genes encoding for Cry1A.105 and Cry2Ab2 Bt proteins were transferred in...

  12. Bacillus thuringiensis

    PubMed Central

    Ibrahim, Mohamed A; Griko, Natalya; Junker, Matthew

    2010-01-01

    Bacillus thuringiensis (Bt) is a unique bacterium in that it shares a common place with a number of chemical compounds which are used commercially to control insects important to agriculture and public health. Although other bacteria, including B. popilliae and B. sphaericus, are used as microbial insecticides, their spectrum of insecticidal activity is quite limited compared to Bt. Importantly, Bt is safe for humans and is the most widely used environmentally compatible biopesticide worldwide. Furthermore, insecticidal Bt genes have been incorporated into several major crops, rendering them insect resistant, and thus providing a model for genetic engineering in agriculture. This review highlights what the authors consider the most relevant issues and topics pertaining to the genomics and proteomics of Bt. At least one of the authors (L.A.B.) has spent most of his professional life studying different aspects of this bacterium with the goal in mind of determining the mechanism(s) by which it kills insects. The other authors have a much shorter experience with Bt but their intellect and personal insight have greatly enriched our understanding of what makes Bt distinctive in the microbial world. Obviously, there is personal interest and bias reflected in this article notwithstanding oversight of a number of published studies. This review contains some material not published elsewhere although several ideas and concepts were developed from a broad base of scientific literature up to 2010. PMID:21327125

  13. Production and characterization of Bacillus thuringiensis Cry1Ac-resistant cotton bollworm Helicoverpa zea (Boddie).

    PubMed

    Anilkumar, Konasale J; Rodrigo-Simón, Ana; Ferré, Juan; Pusztai-Carey, Marianne; Sivasupramaniam, Sakuntala; Moar, William J

    2008-01-01

    Laboratory-selected Bacillus thuringiensis-resistant colonies are important tools for elucidating B. thuringiensis resistance mechanisms. However, cotton bollworm, Helicoverpa zea, a target pest of transgenic corn and cotton expressing B. thuringiensis Cry1Ac (Bt corn and cotton), has proven difficult to select for stable resistance. Two populations of H. zea (AR and MR), resistant to the B. thuringiensis protein found in all commercial Bt cotton varieties (Cry1Ac), were established by selection with Cry1Ac activated toxin (AR) or MVP II (MR). Cry1Ac toxin reflects the form ingested by H. zea when feeding on Bt cotton, whereas MVP II is a Cry1Ac formulation used for resistance selection and monitoring. The resistance ratio (RR) for AR exceeded 100-fold after 11 generations and has been maintained at this level for nine generations. This is the first report of stable Cry1Ac resistance in H. zea. MR crashed after 11 generations, reaching only an RR of 12. AR was only partially cross-resistant to MVP II, suggesting that MVP II does not have the same Cry1Ac selection pressure as Cry1Ac toxin against H. zea and that proteases may be involved with resistance. AR was highly cross-resistant to Cry1Ab toxin but only slightly cross-resistant to Cry1Ab expressing corn leaf powder. AR was not cross-resistant to Cry2Aa2, Cry2Ab2-expressing corn leaf powder, Vip3A, and cypermethrin. Toxin-binding assays showed no significant differences, indicating that resistance was not linked to a reduction in binding. These results aid in understanding why this pest has not evolved B. thuringiensis resistance, and highlight the need to choose carefully the form of B. thuringiensis protein used in experiments. PMID:18024681

  14. Bacillus thuringiensis (Bt)

    NASA Technical Reports Server (NTRS)

    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.

  15. Bacillus thuringiensis and Bacillus sphaericus biopesticides production.

    PubMed

    el-Bendary, Magda A

    2006-01-01

    The long residual action and toxicity of the chemical insecticides have brought about serious environmental problems such as the emergence and spread of insecticide resistance in many species of vectors, mammalian toxicity, and accumulation of pesticide residues in the food chain. All these problems have highlighted the need for alternative biological control agents. Entomo-pathogenic Bacillus thuringiensis (Bt) and Bacillus sphaericus (Bs) are two safe biological control agents. They have attracted considerable interest as possible replacements for the chemical insecticides. Although microbial insecticides based on Bt and Bs are available for use, their high cost makes large-scale application impracticable in developing countries. This review focuses on the economic production of these two microorganisms by submerged fermentation and solid state fermentation using agro-industrial by-products and other wastes. PMID:16598830

  16. Development and validation of real-time PCR screening methods for detection of cry1A.105 and cry2Ab2 genes in genetically modified organisms.

    PubMed

    Dinon, Andréia Z; Prins, Theo W; van Dijk, Jeroen P; Arisi, Ana Carolina M; Scholtens, Ingrid M J; Kok, Esther J

    2011-05-01

    Primers and probes were developed for the element-specific detection of cry1A.105 and cry2Ab2 genes, based on their DNA sequence as present in GM maize MON89034. Cry genes are present in many genetically modified (GM) plants and they are important targets for developing GMO element-specific detection methods. Element-specific methods can be of use to screen for the presence of GMOs in food and feed supply chains. Moreover, a combination of GMO elements may indicate the potential presence of unapproved GMOs (UGMs). Primer-probe combinations were evaluated in terms of specificity, efficiency and limit of detection. Except for specificity, the complete experiment was performed in 9 PCR runs, on 9 different days and by testing 8 DNA concentrations. The results showed a high specificity and efficiency for cry1A.105 and cry2Ab2 detection. The limit of detection was between 0.05 and 0.01 ng DNA per PCR reaction for both assays. These data confirm the applicability of these new primer-probe combinations for element detection that can contribute to the screening for GM and UGM crops in food and feed samples. PMID:21445662

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

    EPA Science Inventory

    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...

  18. [Bacillus thuringiensis: a biotechnology model].

    PubMed

    Sanchis, V; Lereclus, D

    1999-01-01

    This paper is on the different biotechnological approaches that have been used to improve Bacillus thuringiensis (Bt) for the control of agricultural insect pests and have contributed to the successful use of this biological control agent; it describes how a better knowledge of the high diversity of Bt strains and toxins genes together with the development of efficient host-vector systems has made it possible to overcome a number of the problems associated with Bt based insect control measures. First we present an overview of the biology of Bt and of the mode of action of its insecticidal toxins. We then describe some of the progress that has been made in furthering our knowledge of the genetics of Bt and of its insecticidal toxin genes and in the understanding of their regulation. The paper then deals with the use of recombinant DNA technology to develop new Bt strains for more effective pest control or to introduce the genes encoding partial-endotoxins directly into plants to produce insect-resistant trangenic plants. Several examples describing how biotechnology has been used to increase the production of insecticidal proteins in Bt or their persistence in the field by protecting them against UV degradation are presented and discussed. Finally, based on our knowledge of the mechanism of transposition of the Bt transposon Tn4430, we describe the construction of a new generation of recombinant strains of Bt, from which antibiotic resistance genes and other non-Bt DNA sequences were selectively eliminated, using a new generation of site-specific recombination vectors. In the future, continuing improvement of first generation products and research into new sources of resistance is essential to ensure the long-term control of insect pests. Chimeric toxins could also be produced so as to increase toxin activity or direct resistance towards a particular type of insect. The search for new insecticidal toxins, in Bt or other microorganisms, may also provide new weapons

  19. Narrow terahertz attenuation signatures in Bacillus thuringiensis.

    PubMed

    Zhang, Weidong; Brown, Elliott R; Viveros, Leamon; Burris, Kellie P; Stewart, C Neal

    2014-10-01

    Terahertz absorption signatures from culture-cultivated Bacillus thuringiensis were measured with a THz photomixing spectrometer operating from 400 to 1200 GHz. We observe two distinct signatures centered at ∼955 and 1015 GHz, and attribute them to the optically coupled particle vibrational resonance (surface phonon-polariton) of Bacillus spores. This demonstrates the potential of the THz attenuation signatures as "fingerprints" for label-free biomolecular detection. PMID:23821459

  20. Distribution of phenotypes among Bacillus thuringiensis strains

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An extensive collection of Bacillus thuringiensis isolates from around the world were phenotypically profiled using standard biochemical tests. Six phenotypic traits occurred in 20-86% of the isolates and were useful in distinguishing isolates: production of urease (U; 20.5% of isolates), hydrolysis...

  1. The Complete Genome Sequence of Bacillus thuringiensis AlHakam

    SciTech Connect

    Challacombe, Jean F.; Altherr, Michael R.; Xie, Gary; 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.; Han, Cliff S.; Hill, Karen K.; Hitchcock, Penny; Jackson, Paul J.; Keim, Paul; Kewalramani, Avinash Ramesh; Longmire, Jon; Lucas, Susan; Malfatti, Stephanie; Martinez, Diego; McMurry, Kim; Meincke, Linda J.; Misra, Monica; Moseman, Bernice L.; Mundt, Mark; Munk,A. Christine; Okinaka, Richard T.; Parson-Quintana, B.; Reilly, LeePhilip; 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.; Gilna, Paul; Brettin, Thomas S.

    2007-04-01

    Bacillus thuringiensis is an insect pathogen that is widelyused as a biopesticide (3). Here we report the finished, annotated genomesequence of B. thuringiensis Al Hakam, which was collected in Iraq by theUnited Nations Special Commission (2).

  2. A pangenomic study of Bacillus thuringiensis.

    PubMed

    Fang, Yongjun; Li, Zhaolong; Liu, Jiucheng; Shu, Changlong; Wang, Xumin; Zhang, Xiaowei; Yu, Xiaoguang; Zhao, Duojun; Liu, Guiming; Hu, Songnian; Zhang, Jie; Al-Mssallem, Ibrahim; Yu, Jun

    2011-12-20

    Bacillus thuringiensis (B. thuringiensis) is a soil-dwelling Gram-positive bacterium and its plasmid-encoded toxins (Cry) are commonly used as biological alternatives to pesticides. In a pangenomic study, we sequenced seven B. thuringiensis isolates in both high coverage and base-quality using the next-generation sequencing platform. The B. thuringiensis pangenome was extrapolated to have 4196 core genes and an asymptotic value of 558 unique genes when a new genome is added. Compared to the pangenomes of its closely related species of the same genus, B. thuringiensis pangenome shows an open characteristic, similar to B. cereus but not to B. anthracis; the latter has a closed pangenome. We also found extensive divergence among the seven B. thuringiensis genome assemblies, which harbor ample repeats and single nucleotide polymorphisms (SNPs). The identities among orthologous genes are greater than 84.5% and the hotspots for the genome variations were discovered in genomic regions of 2.3-2.8Mb and 5.0-5.6Mb. We concluded that high-coverage sequence assemblies from multiple strains, before all the gaps are closed, are very useful for pangenomic studies. PMID:22196399

  3. Complete Genome Sequence of Bacillus thuringiensis Bacteriophage BMBtp2.

    PubMed

    Dong, Zhaoxia; Peng, Donghai; Wang, Yueying; Zhu, Lei; Ruan, Lifang; Sun, Ming

    2013-01-01

    Bacillus thuringiensis is an insect pathogen which has been widely used for biocontrol. During B. thuringiensis fermentation, lysogenic bacteriophages cause severe losses of yield. Here, we announce the complete genome sequence of a bacteriophage, BMBtp2, which is induced from B. thuringiensis strain YBT-1765, which may be helpful to clarify the mechanism involved in bacteriophage contamination. PMID:23405296

  4. Bacillus thuringiensis and Its Pesticidal Crystal Proteins

    PubMed Central

    Schnepf, E.; Crickmore, N.; Van Rie, J.; Lereclus, D.; Baum, J.; Feitelson, J.; Zeigler, D. R.; Dean, D. H.

    1998-01-01

    During the past decade the pesticidal bacterium Bacillus thuringiensis has been the subject of intensive research. These efforts have yielded considerable data about the complex relationships between the structure, mechanism of action, and genetics of the organism’s pesticidal crystal proteins, and a coherent picture of these relationships is beginning to emerge. Other studies have focused on the ecological role of the B. thuringiensis crystal proteins, their performance in agricultural and other natural settings, and the evolution of resistance mechanisms in target pests. Armed with this knowledge base and with the tools of modern biotechnology, researchers are now reporting promising results in engineering more-useful toxins and formulations, in creating transgenic plants that express pesticidal activity, and in constructing integrated management strategies to insure that these products are utilized with maximum efficiency and benefit. PMID:9729609

  5. 75 FR 35805 - Pesticide Product Registrations; Conditional Approvals

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-06-23

    ... products (EPA File Symbols 524-LTL and 524-LTA) containing the active ingredients, Bacillus thuringiensis... with the proposed use of the Bacillus thuringiensis Cry1A.105 and Cry2Ab2 proteins and the genetic... use of the Bacillus thuringiensis Cry1A.105 and Cry2Ab2 proteins and the genetic material...

  6. Distribution and diversity of Dipteran-specific cry and cyt genes in native Bacillus thuringiensis strains obtained from different ecosystems of Iran.

    PubMed

    Jouzani, Gholamreza Salehi; Abad, Ali Pourjan; Seifinejad, Ali; Marzban, Rasoul; Kariman, Khalil; Maleki, Bahram

    2008-02-01

    One hundred and twenty-eight Bacillus thuringiensis isolates from fields of different ecological regions of Iran were collected to study the distribution and diversity of Dipteran-specific cry and cyt genes. The percentage of samples with Bt showed significant differences between different regions and also between different fields. The most Bt frequency was observed in the soil samples collected from Caspianic zone (7%) and soils of cotton (17%). Characterization of isolates was based on morphological characteristics of crystals, plasmid profiles and protein band patterns as well as PCR analysis using general and specific primers for 22 different cry and cyt genes encoding proteins active against mosquitoes. Thirty-eight different cry gene profiles were detected in this collection. Several of them were found to be different from all previously published profiles and none of the previous researches reported these numbers of profiles. Strains containing cry2-type genes were the most abundant and represent 57.1% of the isolates. Strains harboring cry24 and cry10 genes were also highly abundant (38.7 and 32.8%, respectively). cry11, cry4, cry17, cry19, cry21, cry29, cyt1, and cry9 genes were less abundant, found in 25.7, 14.3, 11.4, 1.4, 4.3, 1.4, and 10% of the strains, respectively. Among the cry2 gene containing isolates, 37.5% strains harbored cry2Aa, 55% cry2Ab, 2.5% cry2Ac, and 5% other or novel cry2-type genes. Among the cry4 gene containing isolates, 0% strains harbored cry4A, 60% cry4B, and 40% cry4C, cry4D or novel cry4 type genes. Finally, based on crystal morphology, protein patterns and PCR, 21 strains were selected as potentially high Dipteran-active for bioassays. Also our results showed that some of the isolates may harbor minimum a putative novel cry gene. PMID:17999100

  7. Fluorescent Amplified Fragment Length Polymorphism Analysis of Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis Isolates

    PubMed Central

    Hill, Karen K.; Ticknor, Lawrence O.; Okinaka, Richard T.; Asay, Michelle; Blair, Heather; Bliss, Katherine A.; Laker, Mariam; Pardington, Paige E.; Richardson, Amber P.; Tonks, Melinda; Beecher, Douglas J.; Kemp, John D.; Kolstø, Anne-Brit; Wong, Amy C. Lee; Keim, Paul; Jackson, Paul J.

    2004-01-01

    DNA from over 300 Bacillus thuringiensis, Bacillus cereus, and Bacillus anthracis isolates was analyzed by fluorescent amplified fragment length polymorphism (AFLP). B. thuringiensis and B. cereus isolates were from diverse sources and locations, including soil, clinical isolates and food products causing diarrheal and emetic outbreaks, and type strains from the American Type Culture Collection, and over 200 B. thuringiensis isolates representing 36 serovars or subspecies were from the U.S. Department of Agriculture collection. Twenty-four diverse B. anthracis isolates were also included. Phylogenetic analysis of AFLP data revealed extensive diversity within B. thuringiensis and B. cereus compared to the monomorphic nature of B. anthracis. All of the B. anthracis strains were more closely related to each other than to any other Bacillus isolate, while B. cereus and B. thuringiensis strains populated the entire tree. Ten distinct branches were defined, with many branches containing both B. cereus and B. thuringiensis isolates. A single branch contained all the B. anthracis isolates plus an unusual B. thuringiensis isolate that is pathogenic in mice. In contrast, B. thuringiensis subsp. kurstaki (ATCC 33679) and other isolates used to prepare insecticides mapped distal to the B. anthracis isolates. The interspersion of B. cereus and B. thuringiensis isolates within the phylogenetic tree suggests that phenotypic traits used to distinguish between these two species do not reflect the genomic content of the different isolates and that horizontal gene transfer plays an important role in establishing the phenotype of each of these microbes. B. thuringiensis isolates of a particular subspecies tended to cluster together. PMID:14766590

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

    EPA Science Inventory

    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...

  9. CHARACTERIZATION OF THE PARASPORAL INCLUSION OF BACILLUS THURINGIENSIS VAR. KYUSHUENSIS

    EPA Science Inventory

    Bacillus thuringiensis var. kyushuensis synthesizes an irregularly shaped parasporal inclusion during sporulation. lectron microscopy revealed that the inclusions are composed of a relatively homogeneous appearing center surrounded by a thick, electron dense coating. urified incl...

  10. Bacillus thuringiensis Conjugation in Simulated Microgravity

    NASA Astrophysics Data System (ADS)

    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.

  11. Sludge based Bacillus thuringiensis biopesticides: viscosity impacts.

    PubMed

    Brar, S K; Verma, M; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2005-08-01

    Viscosity studies were performed on raw, pre-treated (sterilised and thermal alkaline hydrolysed or both types of treatment) and Bacillus thuringiensis (Bt) fermented sludges at different solids concentration (10-40 g/L) for production of biopesticides. Correlations were established among rheological parameter (viscosity), solids (total and dissolved) concentration and entomotoxicity (Tx) of Bt fermented sludges. Exponential and power laws were preferentially followed by hydrolysed fermented compared to raw fermented sludge. Soluble chemical oxygen demand variation corroborated with increase in dissolved solids concentration on pre-treatments, contributing to changes in viscosity. Moreover, Tx was higher for hydrolysed fermented sludge in comparison to raw fermented sludge owing to increased availability of nutrients and lower viscosity that improved oxygen transfer. The shake flask results were reproducible in fermenter. This study will have major impact on selecting fermentation, harvesting and formulation techniques of Bt fermented sludges for biopesticide production. PMID:15979118

  12. Complete Genome Sequence of Bacillus thuringiensis Bacteriophage Smudge.

    PubMed

    Cornell, Jessica L; Breslin, Eileen; Schuhmacher, Zachary; Himelright, Madison; Berluti, Cassandra; Boyd, Charles; Carson, Rachel; Del Gallo, Elle; Giessler, Caris; Gilliam, Benjamin; Heatherly, Catherine; Nevin, Julius; Nguyen, Bryan; Nguyen, Justin; Parada, Jocelyn; Sutterfield, Blake; Tukruni, Muruj; Temple, Louise

    2016-01-01

    Smudge, a bacteriophage enriched from soil using Bacillus thuringiensis DSM-350 as the host, had its complete genome sequenced. Smudge is a myovirus with a genome consisting of 292 genes and was identified as belonging to the C1 cluster of Bacillus phages. PMID:27540049

  13. Complete Genome Sequence of Bacillus thuringiensis Bacteriophage Smudge

    PubMed Central

    Cornell, Jessica L.; Breslin, Eileen; Schuhmacher, Zachary; Himelright, Madison; Berluti, Cassandra; Boyd, Charles; Carson, Rachel; Del Gallo, Elle; Giessler, Caris; Gilliam, Benjamin; Heatherly, Catherine; Nevin, Julius; Nguyen, Bryan; Nguyen, Justin; Parada, Jocelyn; Sutterfield, Blake; Tukruni, Muruj

    2016-01-01

    Smudge, a bacteriophage enriched from soil using Bacillus thuringiensis DSM-350 as the host, had its complete genome sequenced. Smudge is a myovirus with a genome consisting of 292 genes and was identified as belonging to the C1 cluster of Bacillus phages. PMID:27540049

  14. Pathogenomic Sequence Analysis of Bacillus cereus and Bacillus thuringiensis Isolates Closely Related to Bacillus anthracis†

    PubMed Central

    Han, Cliff S.; Xie, Gary; Challacombe, Jean F.; Altherr, Michael R.; Bhotika, Smriti S.; 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.; 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

    2006-01-01

    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. PMID:16621833

  15. Complete genome sequence of Bacillus thuringiensis strain HD521.

    PubMed

    Li, Qiao; Xu, Li Z; Zou, Ting; Ai, Peng; Huang, Gang H; Li, Ping; Zheng, Ai P

    2015-01-01

    Bacillus thuringiensis is the most widely used biological pesticide in the world. It belongs to the Bacillus cereus sensu lato group, which contains six species. Among these six species, B. thuringiensis, B. anthracis, and B. cereus have a low genetic diversity. B. thuringiensis strain HD521 shows maroon colony which is different from most of the B. thuringiensis strains. Strain HD521 also displays an ability to inhibit plant sheath blight disease pathogen (Rhizoctonia solani AG1 IB) growth and can form bipyramidal parasporal crystals consisting of three cry7 genes. These crystals have an insecticidal activity against Henosepilachna vigintioctomaculata larva (Coleoptera). Here we report the complete genome sequence of strain HD521, which has one chromosome and six circular plasmids. PMID:26380647

  16. Mechanism of Insect Resistance to the Microbial Insecticide Bacillus thuringiensis

    NASA Astrophysics Data System (ADS)

    van Rie, J.; McGaughey, W. H.; Johnson, D. E.; Barnett, B. D.; van Mellaert, H.

    1990-01-01

    Receptor binding studies show that resistance of a laboratory-selected Plodia interpunctella strain to a Bacillus thuringiensis insecticidal crystal protein (ICP) is correlated with a 50-fold reduction in affinity of the membrane receptor for this protein. The strain is sensitive to a second type of ICP that apparently recognizes a different receptor. Understanding the mechanism of resistance will provide strategies to prevent or delay resistance and hence prolong the usefulness of B. thuringiensis ICPs as environmentally safe insecticides.

  17. Bacillus thuringiensis: a genomics and proteomics perspective.

    PubMed

    Ibrahim, Mohamed A; Griko, Natalya; Junker, Matthew; Bulla, Lee A

    2010-01-01

    Bacillus thuringiensis (Bt) is a unique bacterium in that it shares a common place with a number of chemical compounds which are used commercially to control insects important to agriculture and public health. Although other bacteria, including B. popilliae and B. sphaericus, are used as microbial insecticides, their spectrum of insecticidal activity is quite limited compared to Bt. Importantly, Bt is safe for humans and is the most widely used environmentally compatible biopesticide worldwide. Furthermore, insecticidal Bt genes have been incorporated into several major crops, rendering them insect resistant, and thus providing a model for genetic engineering in agriculture.This review highlights what the authors consider the most relevant issues and topics pertaining to the genomics and proteomics of Bt. At least one of the authors (L.A.B.) has spent most of his professional life studying different aspects of this bacterium with the goal in mind of determining the mechanism(s) by which it kills insects. The other authors have a much shorter experience with Bt but their intellect and personal insight have greatly enriched our understanding of what makes Bt distinctive in the microbial world. Obviously, there is personal interest and bias reflected in this article notwithstanding oversight of a number of published studies. This review contains some material not published elsewhere although several ideas and concepts were developed from a broad base of scientific literature up to 2010. PMID:21327125

  18. Bacteriocins synthesized by Bacillus thuringiensis: generalities and potential applications

    PubMed Central

    Salazar-Marroquín, Elma Laura; Galán-Wong, Luis J.; Moreno-Medina, Víctor Ricardo; Reyes-López, Miguel Ángel; Pereyra-Alférez, Benito

    2016-01-01

    The members of the Bacillus thuringiensis group, commonly known as Bt, produce a huge number of metabolites, which show biocidal and antagonistic activity. B. thuringiensis is widely known for synthesizing Cry, Vip and Cyt proteins, active against insects and other parasporins with biocidal activity against certain types of cancerous cells. Nevertheless, B. thuringiensis also synthesizes compounds with antimicrobial activity, especially bacteriocins. Some B. thuringiensis bacteriocins resemble lantibiotics and other small linear peptides (class IIa) from the lactic acid bacteria bacteriocins classification system. Although many bacteriocins produced by Bt have been reported, there is no proper classification for them. In this work, we have grouped these based on molecular weight and functionality. Bacteriocins are small peptides synthesized by bacteria, presenting inhibitory activity against Gram-positive and Gram-negative bacteria and to a lesser extent against fungi. These molecules represent a good study model in the search for microbial control alternatives. Lactic acid bacteria produces a huge number of these types of molecules with great potential. Nonetheless, members of the Bacillus, cereus group, especially B. thuringiensis, emerge as an attractive alternative for obtaining bacteriocins showing novel activities. This review describes the potential applications of B. thuringiensis bacteriocins in the control of foodborne pathogens, environment and medical area. PMID:27340340

  19. Regulation of cry gene expression in Bacillus thuringiensis.

    PubMed

    Deng, Chao; Peng, Qi; Song, Fuping; Lereclus, Didier

    2014-01-01

    Bacillus thuringiensis differs from the closely related Bacillus cereus group species by its ability to produce crystalline inclusions. The production of these crystals mainly results from the expression of the cry genes, from the stability of their transcripts and from the synthesis, accumulation and crystallization of large amounts of insecticidal Cry proteins. This process normally coincides with sporulation and is regulated by various factors operating at the transcriptional, post-transcriptional, metabolic and post-translational levels. PMID:25055802

  20. Occurrence and significance of Bacillus thuringiensis on wine grapes.

    PubMed

    Bae, Sungsook; Fleet, Graham H; Heard, Gillian M

    2004-08-01

    Wine grapes harvested at different stages during cultivation from several vineyards in New South Wales, Australia, harboured Bacillus thuringiensis at viable populations of 10(2)-10(6) cfu/g. Commercial preparations of B. thuringiensis had been sprayed onto the grapes as a biological insecticide. B. thuringiensis (10(1)-10(3) cfu/ml) was isolated from grape juice and fermenting grape juice in a commercial winery. Although B. thuringiensis remained viable when inoculated at 10(3)-10(4) cfu/ml into grape juice and wine (pH 3.0-6.0), it did not grow. Using in vitro agar culture assays, B. thuringiensis inhibited several grape-associated yeasts and bacteria as well as various species of fungi associated with grape spoilage and ochratoxin A production. B. thuringiensis did not inhibit Saccharomyces cerevisiae in agar culture or during alcoholic fermentation of grape juice. B. thuringiensis inhibited the malolactic bacterium, Oenococcus oeni, in agar culture but not during mixed cultures in a liquid medium. PMID:15246241

  1. Parallel Evolution of Bacillus thuringiensis Toxin Resistance in Lepidoptera

    PubMed Central

    Baxter, Simon W.; Badenes-Pérez, Francisco R.; Morrison, Anna; Vogel, Heiko; Crickmore, Neil; Kain, Wendy; Wang, Ping; Heckel, David G.; Jiggins, Chris D.

    2011-01-01

    Despite the prominent and worldwide use of Bacillus thuringiensis (Bt) insecticidal toxins in agriculture, knowledge of the mechanism by which they kill pests remains incomplete. Here we report genetic mapping of a membrane transporter (ABCC2) to a locus controlling Bt Cry1Ac toxin resistance in two lepidopterans, implying that this protein plays a critical role in Bt function. PMID:21840855

  2. Interactions of transgenic Bacillus thuringiensis insecticidal crops with spiders (Araneae)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genetically modified crops expressing insecticidal proteins from Bacillus thuringiensis (Bt) have dramatically increased in acreage since their introduction in the mid-1990’s. Although the insecticidal mechanisms of Bt target specific pests, concerns persist regarding direct and indirect effects on...

  3. Impact of Entomopathogens on Pest Resistance to Bacillus thuringiensis.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Adaptation by pest insects to the bacterium Bacillus thuringiensis (Bt) can alter their susceptibility to other pathogens. As the number of acres planted in crops engineered to produce Bt toxin increases, many key agricultural pests undergo strong selection to evolve resistance to Bt. In conjuncti...

  4. TRANSGENIC PLANTS EXPRESSING BACILLUS THURINGIENSIS DELTA-ENDOTOXINS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Commercial varieties of transgenic Bacillus thuringiensis (Bt) plants have been developed in many countries to control target pests. Initially, the expression of native Bt genes in plants was low due to mRNA instability, improper splicing, and post-translation modifications. Subsequently, modificati...

  5. Bollgard II cotton: compositional analysis and feeding studies of cottonseed from insect-protected cotton (Gossypium hirsutum L.) producing the Cry1Ac and Cry2Ab2 proteins.

    PubMed

    Hamilton, Kathryn A; Pyla, Paul D; Breeze, Matthew; Olson, Tammy; Li, Menghe; Robinson, Edwin; Gallagher, Sean P; Sorbet, Roy; Chen, Yin

    2004-11-17

    Bollgard II cotton event 15985 producing the Cry1Ac and Cry2Ab2 proteins has been developed by genetic modification to broaden the spectrum of insects to which the plant is tolerant and to provide an insect resistance management tool to impede the onset of resistance. The purpose of this study was to evaluate the composition and nutrition of Bollgard II cotton, relative to the use for food and animal feed, compared to that of conventional cotton varieties. Compositional analyses were conducted to measure proximate, fiber, amino acid, fatty acid, gossypol, and mineral contents of cottonseed from a total of 14 U.S. field sites over two years. Compositional analysis results showed that the cottonseed and cottonseed oil from Bollgard II cotton were comparable in their composition to those of the conventional control cotton line and other commercial varieties. The composition data are supported by nutritional safety studies conducted with dairy cows, catfish, and quail. Results from these studies showed that Bollgard II performed similarly to the conventional control cotton varieties. These data demonstrate that Bollgard II cotton is compositionally and nutritionally equivalent to conventional cotton varieties. These data support the conclusion that Bollgard II cotton is as safe and nutritious as conventional cotton for food and feed use. PMID:15537305

  6. The Complete Genome Sequence of Bacillus thuringiensis Al Hakam▿

    PubMed Central

    Challacombe, Jean F.; Altherr, Michael R.; Xie, Gary; 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.; Han, Cliff S.; Hill, Karen K.; Hitchcock, Penny; Jackson, Paul J.; Keim, Paul; Kewalramani, Avinash Ramesh; Longmire, Jon; Lucas, Susan; Malfatti, Stephanie; Martinez, Diego; 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.; Saunders, Elizabeth; Tapia, Roxanne; Tesmer, Judith G.; Thayer, Nina; Thompson, Linda S.; Tice, Hope; Ticknor, Lawrence O.; Wills, Patti L.; Gilna, Paul; Brettin, Thomas S.

    2007-01-01

    Bacillus thuringiensis is an insect pathogen that is widely used as a biopesticide (E. Schnepf, N. Crickmore, J. Van Rie, D. Lereclus, J. Baum, J. Feitelson, D. R. Zeigler, and D. H. Dean, Microbiol. Mol. Biol. Rev. 62:775-806, 1998). Here we report the finished, annotated genome sequence of B. thuringiensis Al Hakam, which was collected in Iraq by the United Nations Special Commission (L. Radnedge, P. Agron, K. Hill, P. Jackson, L. Ticknor, P. Keim, and G. Andersen, Appl. Environ. Microbiol. 69:2755-2764, 2003). PMID:17337577

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

    PubMed Central

    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

  8. Phages preying on Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis: past, present and future.

    PubMed

    Gillis, Annika; Mahillon, Jacques

    2014-07-01

    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

  9. Genetic Differentiation between Sympatric Populations of Bacillus cereus and Bacillus thuringiensis

    PubMed Central

    Vilas-Boas, Gislayne; Sanchis, Vincent; Lereclus, Didier; Lemos, Manoel Victor F.; Bourguet, Denis

    2002-01-01

    Little is known about genetic exchanges in natural populations of bacteria of the spore-forming Bacillus cereus group, because no population genetics studies have been performed with local sympatric populations. We isolated strains of Bacillus thuringiensis and B. cereus from small samples of soil collected at the same time from two separate geographical sites, one within the forest and the other at the edge of the forest. A total of 100 B. cereus and 98 B. thuringiensis strains were isolated and characterized by electrophoresis to determine allelic composition at nine enzymatic loci. We observed genetic differentiation between populations of B. cereus and B. thuringiensis. Populations of a given Bacillus species—B. thuringiensis or B. cereus—were genetically more similar to each other than to populations of the other Bacillus species. Hemolytic activity provided further evidence of this genetic divergence, which remained evident even if putative clones were removed from the data set. Our results suggest that the rate of gene flow was higher between strains of the same species, but that exchanges between B. cereus and B. thuringiensis were nonetheless possible. Linkage disequilibrium analysis revealed sufficient recombination for B. cereus populations to be considered panmictic units. In B. thuringiensis, the balance between clonal proliferation and recombination seemed to depend on location. Overall, our data indicate that it is not important for risk assessment purposes to determine whether B. cereus and B. thuringiensis belong to a single or two species. Assessment of the biosafety of pest control based on B. thuringiensis requires evaluation of the extent of genetic exchange between strains in realistic natural conditions. PMID:11872495

  10. Genetic differentiation between sympatric populations of Bacillus cereus and Bacillus thuringiensis.

    PubMed

    Vilas-Boas, Gislayne; Sanchis, Vincent; Lereclus, Didier; Lemos, Manoel Victor F; Bourguet, Denis

    2002-03-01

    Little is known about genetic exchanges in natural populations of bacteria of the spore-forming Bacillus cereus group, because no population genetics studies have been performed with local sympatric populations. We isolated strains of Bacillus thuringiensis and B. cereus from small samples of soil collected at the same time from two separate geographical sites, one within the forest and the other at the edge of the forest. A total of 100 B. cereus and 98 B. thuringiensis strains were isolated and characterized by electrophoresis to determine allelic composition at nine enzymatic loci. We observed genetic differentiation between populations of B. cereus and B. thuringiensis. Populations of a given Bacillus species--B. thuringiensis or B. cereus--were genetically more similar to each other than to populations of the other Bacillus species. Hemolytic activity provided further evidence of this genetic divergence, which remained evident even if putative clones were removed from the data set. Our results suggest that the rate of gene flow was higher between strains of the same species, but that exchanges between B. cereus and B. thuringiensis were nonetheless possible. Linkage disequilibrium analysis revealed sufficient recombination for B. cereus populations to be considered panmictic units. In B. thuringiensis, the balance between clonal proliferation and recombination seemed to depend on location. Overall, our data indicate that it is not important for risk assessment purposes to determine whether B. cereus and B. thuringiensis belong to a single or two species. Assessment of the biosafety of pest control based on B. thuringiensis requires evaluation of the extent of genetic exchange between strains in realistic natural conditions. PMID:11872495

  11. Occurrence of Bacillus thuringiensis in fresh waters of Japan.

    PubMed

    Ichimatsu, T; Mizuki, E; Nishimura, K; Akao, T; Saitoh, H; Higuchi, K; Ohba, M

    2000-04-01

    Bacillus thuringiensis was recovered at a relatively high frequency from both running and still fresh waters in natural environments of Kyushu, Japan. Of 107 water samples examined, 53 (49.5%) contained this organism. The frequency of B. thuringiensis colonies was 4.4% among 4414 colonies of the Bacillus cereus/B. thuringiensis group. The density of this bacterium in fresh waters averaged 0.45 cfu/ml. Serologically, B. thuringiensis isolates were assigned to 26 H serotypes. Of these, H14/36 (H serovar israelensis/malaysiensis) was the predominant, followed by the serotypes H3abc (kurstaki), H27 (mexicanensis), H3ad (sumiyoshiensis), and H35 (seoulensis). Of 195 isolates, 52 (26.7%) exhibited larvicidal activity against aquatic Diptera; 21 killed Culex pipiens molestus (Culicidae) only, and 31 were active on both the culicine mosquito and the moth-fly, Clogmia albipunctata (Psychodidae). The Diptera-toxic isolates produced spherical or irregularly pointed parasporal inclusions. PMID:10688688

  12. Photoprotection of Bacillus thuringiensis kurstaki from ultraviolet irradiation

    SciTech Connect

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

    1991-05-01

    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.

  13. SinR Controls Enterotoxin Expression in Bacillus thuringiensis Biofilms

    PubMed Central

    Økstad, Ole-Andreas; Verplaetse, Emilie; Gilois, Nathalie; Bennaceur, Imène; Perchat, Stéphane; Gominet, Myriam; Aymerich, Stéphane; Kolstø, Anne-Brit; Lereclus, Didier; Gohar, Michel

    2014-01-01

    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. PMID:24498128

  14. Genome Sequence of the Endophytic Bacterium Bacillus thuringiensis Strain KB1, a Potential Biocontrol Agent against Phytopathogens

    PubMed Central

    Jo, Sung Hee; Hong, Chi Eun

    2016-01-01

    Bacillus thuringiensis is the most widely known microbial pesticide used in agricultural applications. Herein, we report a draft genome sequence of the endophytic bacterium Bacillus thuringiensis strain KB1, which exhibits antagonism against phytopathogens. PMID:27103716

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

    PubMed

    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. PMID:16881328

  16. 40 CFR 174.509 - Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 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...

  17. 40 CFR 174.504 - Bacillus thuringiensis Cry1F protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Cry1F protein...-INCORPORATED PROTECTANTS Tolerances and Tolerance Exemptions § 174.504 Bacillus thuringiensis Cry1F protein; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1F protein in the...

  18. 40 CFR 174.509 - Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 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...

  19. 40 CFR 174.509 - Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 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...

  20. 40 CFR 174.509 - Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 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...

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

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

    PubMed

    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; Ercolini, Danilo; Pennacchio, Francesco

    2016-08-23

    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

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

    PubMed

    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. PMID:25283838

  4. Bacillus thuringiensis resistance in Plutella - too many trees?

    PubMed

    Crickmore, Neil

    2016-06-01

    Plutella xylostella was the first insect for which resistance to Bacillus thuringiensis was reported in the field, yet despite many studies on the nature of this resistance phenotype its genetic and molecular basis remains elusive. Many different factors have been proposed as contributing to resistance, although in many cases it has not been possible to establish a causal link. Indeed, there are so many studies published that it has become very difficult to 'see the wood for the trees'. This article will attempt to clarify our current understanding of Bt resistance in P. xylostella and consider the criteria that are used when validating a particular model. PMID:27436736

  5. Microbial control and biotechnology research on Bacillus thuringiensis in China.

    PubMed

    Huang, Da-Fang; Zhang, Jie; Song, Fu-Ping; Lang, Zhi-Hong

    2007-07-01

    The current status of production and application of biopesticides for pest control in China is briefly reviewed, with a focus on research advances in microbial control with Bacillus thuringiensis (Bt). These have led to improvements in Bt production, exploitation of Bt gene resources, and development of engineered Bt insecticides and transgenic Bt crops that have expanded host ranges and increased efficacy against target pests. Both conventional and biotechnology approaches need to be employed to achieve further progress in discovery, production technology, formulation processing, development of quality standards and recommended use patterns. PMID:17481651

  6. Insecticidal proteins from Bacillus thuringiensis protect corn from corn rootworms.

    PubMed

    Moellenbeck, D J; Peters, M L; Bing, J W; Rouse, J R; Higgins, L S; Sims, L; Nevshemal, T; Marshall, L; Ellis, R T; Bystrak, P G; Lang, B A; Stewart, J L; Kouba, K; Sondag, V; Gustafson, V; Nour, K; Xu, D; Swenson, J; Zhang, J; Czapla, T; Schwab, G; Jayne, S; Stockhoff, B A; Narva, K; Schnepf, H E; Stelman, S J; Poutre, C; Koziel, M; Duck, N

    2001-07-01

    Field tests of corn co-expressing two new delta-endotoxins from Bacillus thuringiensis (Bt) have demonstrated protection from root damage by western corn rootworm (Diabrotica virgifera virgifera LeConte). The level of protection exceeds that provided by chemical insecticides. In the bacterium, these proteins form crystals during the sporulation phase of the growth cycle, are encoded by a single operon, and have molecular masses of 14 kDa and 44 kDa. Corn rootworm larvae fed on corn roots expressing the proteins showed histopathological symptoms in the midgut epithelium. PMID:11433280

  7. Existence of lysogenic bacteriophages in Bacillus thuringiensis type strains.

    PubMed

    Roh, Jong Yul; Park, Jong Bin; Liu, Qin; Kim, Song Eun; Tao, Xueying; Choi, Tae Woong; Choi, Jae Young; Kim, Woo Jin; Jin, Byung Rae; Je, Yeon Ho

    2013-07-01

    We screened the existence of bacteriophages in 67 Bacillus thuringiensis type strains by phage DNA extraction and PCR using phage terminase small subunit (TerS)-specific primers to the supernatants and the precipitated pellets of Bt cultures, and by transmission electron microscopy. The various bacteriophages were observed from the supernatants of 22 type strains. Ten type strains showed the extracted phage DNAs and the amplified fragment by TerS PCR but 12 type strains showed only the phage DNAs. Their morphological characteristic suggests that they belong to Family Siphoviridae which had a long tail and symmetrical head. PMID:23632013

  8. Isolation and molecular characterisation of alkaline protease producing Bacillus thuringiensis.

    PubMed

    Agasthya, Annapurna S; Sharma, Naresh; Mohan, Anand; Mahal, Prabhpreet

    2013-05-01

    Proteases are of particular interest because of their action on insoluble keratin substrates and generally on a broad range of protein substrates. Proteases are one of the most important groups of industrial enzymes used in detergent, protein, brewing, meat, photographic, leather, dairy, pharmaceutical and food industry. In the present study, the organism isolated from the protein rich soil sample was identified by biochemical and molecular characterisation as Bacillus thuringiensis and further optimum conditions for alkaline protease synthesis were determined. The growth conditions for B. thuringiensis was optimised by inoculating into yeast extract casein medium at different pH and incubating at different temperatures. The maximum protease production occurred at pH 8 and at 37 °C. B. thuringiensis showed proteolytic activity at various culture conditions. Optimum conditions for the protease activity were found to be 47 °C and pH 8. In the later stage, the blood removing action of crude and partially purified protease was found to be effective within 25 min in the presence of commercial detergents indicating the possible use of this enzyme in detergent industry. Enzyme also showed good activity against hair substrate keratin and can be used for dehairing. PMID:22826099

  9. Modified Bacillus thuringiensis Toxins and a Hybrid B. thuringiensis Strain Counter Greenhouse-Selected Resistance in Trichoplusia ni▿

    PubMed Central

    Franklin, Michelle T.; Nieman, Christal L.; Janmaat, Alida F.; Soberón, Mario; Bravo, Alejandra; Tabashnik, Bruce E.; Myers, Judith H.

    2009-01-01

    Resistance of greenhouse-selected strains of the cabbage looper, Trichoplusia ni, to Bacillus thuringiensis subsp. kurstaki was countered by a hybrid strain of B. thuringiensis and genetically modified toxins Cry1AbMod and Cry1AcMod, which lack helix α-1. Resistance to Cry1AbMod and Cry1AcMod was >100-fold less than resistance to native toxins Cry1Ab and Cry1Ac. PMID:19592525

  10. 40 CFR 174.530 - Bacillus thuringiensis Cry2Ae protein in cotton; temporary exemption from the requirement of a...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry2Ae protein... thuringiensis Cry2Ae protein in cotton; temporary exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry2Ae protein in or on the food commodities of cotton, cotton; cotton,...

  11. Two novel strains of Bacillus thuringiensis toxic to coleopterans.

    PubMed Central

    Rupar, M J; Donovan, W P; Groat, R G; Slaney, A C; Mattison, J W; Johnson, T B; Charles, J F; Dumanoir, V C; de Barjac, H

    1991-01-01

    Two novel strains of Bacillus thuringiensis were isolated from native habitats by the use of genes coding for proteins toxic to coleopterans (cryIII genes) as hybridization probes. Strain EG2838 (isolated by the use of the cryIIIA probe) contained a cryIIIA-hybridizing plasmid of approximately 100 MDa and synthesized crystal proteins of approximately 200 (doublet), 74, 70, 32, and 28 kDa. Strain EG4961 (isolated by the use of a cryIIIA-related probe) contained a cryIIIA-hybridizing plasmid of approximately 95 MDa and synthesized crystal proteins of 74, 70, and 30 kDa. Structural relationships among the crystal proteins of strains EG2838 and EG4961 were detected; antibodies to the CryIIIA protein toxic to coleopterans reacted with the 74- and 70-kDa proteins of EG2838 and EG4961, antibodies to the 32-kDa plus 28-kDa proteins of EG2838 reacted with the 30-kDa protein of EG4961, and antibodies to the 200-kDa proteins of EG2838 reacted with the 28-kDa protein of EG2838. Experiments with B. thuringiensis flagella antibody reagents demonstrated that EG2838 belongs to H serotype 9 (reference strain B. thuringiensis subsp. tolworthi) and that EG4961 belongs to H serotype 18 (reference strain B. thuringiensis subsp. kumamotoensis). A mixture of spores plus crystal proteins of either EG2838 or EG4961 was toxic to the larvae of Colorado potato beetle (Leptinotarsa decemlineata), and significantly, the EG4961 mixture was also toxic to the larvae of southern corn rootworm (Diabrotica undecimpunctata howardi). DNA restriction blot analysis suggested that strains EG2838 and EG4961 each contained a unique gene coding for a protein toxic to coleopterans. Images PMID:1781691

  12. Two novel strains of Bacillus thuringiensis toxic to coleopterans.

    PubMed

    Rupar, M J; Donovan, W P; Groat, R G; Slaney, A C; Mattison, J W; Johnson, T B; Charles, J F; Dumanoir, V C; de Barjac, H

    1991-11-01

    Two novel strains of Bacillus thuringiensis were isolated from native habitats by the use of genes coding for proteins toxic to coleopterans (cryIII genes) as hybridization probes. Strain EG2838 (isolated by the use of the cryIIIA probe) contained a cryIIIA-hybridizing plasmid of approximately 100 MDa and synthesized crystal proteins of approximately 200 (doublet), 74, 70, 32, and 28 kDa. Strain EG4961 (isolated by the use of a cryIIIA-related probe) contained a cryIIIA-hybridizing plasmid of approximately 95 MDa and synthesized crystal proteins of 74, 70, and 30 kDa. Structural relationships among the crystal proteins of strains EG2838 and EG4961 were detected; antibodies to the CryIIIA protein toxic to coleopterans reacted with the 74- and 70-kDa proteins of EG2838 and EG4961, antibodies to the 32-kDa plus 28-kDa proteins of EG2838 reacted with the 30-kDa protein of EG4961, and antibodies to the 200-kDa proteins of EG2838 reacted with the 28-kDa protein of EG2838. Experiments with B. thuringiensis flagella antibody reagents demonstrated that EG2838 belongs to H serotype 9 (reference strain B. thuringiensis subsp. tolworthi) and that EG4961 belongs to H serotype 18 (reference strain B. thuringiensis subsp. kumamotoensis). A mixture of spores plus crystal proteins of either EG2838 or EG4961 was toxic to the larvae of Colorado potato beetle (Leptinotarsa decemlineata), and significantly, the EG4961 mixture was also toxic to the larvae of southern corn rootworm (Diabrotica undecimpunctata howardi). DNA restriction blot analysis suggested that strains EG2838 and EG4961 each contained a unique gene coding for a protein toxic to coleopterans. PMID:1781691

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

    PubMed Central

    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

  14. Architecture and High-Resolution Structure of Bacillus thuringiensis and Bacillus cereus Spore Coat Surfaces

    SciTech Connect

    Plomp, M; Leighton, T; Wheeler, K; Malkin, A

    2005-02-18

    We have utilized atomic force microscopy (AFM) to visualize the native surface topology and ultrastructure of Bacillus thuringiensis and Bacillus cereus spores in water and in air. AFM was able to resolve the nanostructure of the exosporium and three distinctive classes of appendages. Removal of the exosporium exposed either a hexagonal honeycomb layer (B. thuringiensis) or a rodlet outer spore coat layer (B. cereus). Removal of the rodlet structure from B. cereus spores revealed an underlying honeycomb layer similar to that observed with B. thuringiensis spores. The periodicity of the rodlet structure on the outer spore coat of B. cereus was {approx}8 nm, and the length of the rodlets was limited to the cross-patched domain structure of this layer to {approx}200 nm. The lattice constant of the honeycomb structures was {approx}9 nm for both B. cereus and B. thuringiensis spores. Both honeycomb structures were composed of multiple, disoriented domains with distinct boundaries. Our results demonstrate that variations in storage and preparation procedures result in architectural changes in individual spore surfaces, which establish AFM as a useful tool for evaluation of preparation and processing ''fingerprints'' of bacterial spores. These results establish that high-resolution AFM has the capacity to reveal species-specific assembly and nanometer scale structure of spore surfaces. These species-specific spore surface structural variations are correlated with sequence divergences in a spore core structural protein SspE.

  15. Susceptibility, mechanisms of response and resistance to Bacillus thuringiensis toxins in Spodoptera spp.

    PubMed

    Herrero, Salvador; Bel, Yolanda; Hernández-Martínez, Patricia; Ferré, Juan

    2016-06-01

    Bioinsecticides based on Bacillus thuringiensis have long been used as an alternative to synthetic insecticides to control insect pests. In this review, we focus on insects of the genus Spodoptera, including relevant polyphagous species that are primary and secondary pests of many crops, and how B. thuringiensis toxins can be used for Spodoptera spp. pest management. We summarize the main findings related to susceptibility, midgut binding specificity, mechanisms of response and resistance of this insect genus to B. thuringiensis toxins. PMID:27436737

  16. Survival of diverse bacillus thuringiensis strains in gypsy moth (Lepidotera: Lymantriidae) is correlated with urease production

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacillus thuringiensis is an entomopathogenic bacterium that can kill a variety of pest insects, but seldom causes epizootics because it replicates poorly in insects. By attempting to repeatedly pass lepidopteran-active B. thuringiensis strains through gypsy moth larvae, we found that only those str...

  17. Activity of Bacillus thuringiensis against Pryeria sinica(Lepidoptera: Zygaenidae), an invasive pest of Euonymus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pryeria sinica Moore (Lepidoptera: Zygaenidae), an invasive pest of Euonymus, is susceptible in the second instar to the Bacillus thuringiensis Berliner product Thuricide®, and to several strains isolated from other B. thuringiensis products. Third instars are also susceptible, while susceptibility...

  18. CHARACTERIZATION OF THE MAMMALIAN TOXICITY OF THE CRYSTAL POLYPEPTIDES OF BACILLUS THURINGIENSIS SUBSPECIES ISRAELENSIS

    EPA Science Inventory

    Solubilized crystal polypeptide preparations of Bacillus thuringiensis subsp. israelertsis (BTI) were fractionated by immunoaffinity chromatography using a bound monoclonal antibody formed against the 28K crystal polypeptide. The 28K polypeptide was confirmed to be hemolytic and ...

  19. 40 CFR 180.1011 - Viable spores of the microorganism Bacillus thuringiensis Berliner; exemption from the...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... production is a Bacillus thuringiensis strain which does not produce β-exotoxin under standard manufacturing... it is applied either to growing crops, or when it is applied after harvest in accordance with...

  20. Pathogenicity of Bacillus thuringiensis variety kurstaki to Ixodes scapularis (Acari: Ixodidae)

    USGS Publications Warehouse

    Zhioua, E.; Heyer, K.; Browning, M.; Ginsberg, H.S.; LeBrun, R.A.

    1999-01-01

    Pathogenicity of the entomopathogenic bacterium Bacillus thuringiensis var. kurstaki de Barjac & Lemille was tested against the black-legged tick, Ixodes scapularis Say. Engorged larvae dipped in a solution of 108 spores per ml showed 96% mortality, 3 wk post-infection. The LC50 value for engorged larvae (concentration required to kill 50% of ticks) was 107 spores/ml. Bacillus thuringiensis shows considerable potential as a microbial control agent for the management of Ixodes scapularis.

  1. A strategy for shuffling numerous Bacillus thuringiensis crystal protein domains.

    PubMed

    Knight, Jacqueline S; Broadwell, Andrew H; Grant, Warwick N; Shoemaker, Charles B

    2004-12-01

    Bacillus thuringiensis that produce Cry1Ba are toxic to Lucilia cuprina Wiedemann blow fly maggots in vivo, and when applied in quantity to sheep fleece, provide up to 6 wk protection against flystrike in the field. These strains also are toxic to Epiphyas postvittana (Walker) light brown apple moth caterpillars. B. thuringiensis expressing Cry1Db are toxic only to E. postvittana. When Cry1Ba and Cry1Db proteins are expressed within Escherichia coli, the recombinant bacteria have the same toxicity profile as the wild-type B. thuringiensis strain. In an effort to develop a Cry protein with improved blow fly toxicity, three different internal regions of Cry1Ba coding DNA, encoding all or part of domains I, II and III respectively were systematically exchanged with the corresponding region from a pool of other Cry protein coding DNAs. The chimeric products were then expressed in recombinant E. coli, and the resulting bacteria assayed for toxicity on L. cuprina and E. postvittana. Clones having insecticide bioactivity were characterized to identify the source of the replacement Cry domain. Despite successfully expressing a large number and variety of chimeric proteins within E. coli, many with measurable insecticidal activity, none of the chimeras had greater potency against L. cuprina than the wild-type Cry1Ba. Chimeric replacements involving domains I and II were rarely active, whereas a much higher proportion of domain III chimeras had some bioactivity. We conclude that shuffling of Cry coding regions through joining at the major conserved sequence motifs is an effective means for the production of a diverse number of chimeric Cry proteins but that such toxins with enhanced bioactive properties will be rare or nonexistent. PMID:15666731

  2. Production of Polyhydroxyalkanoate Co-polymer by Bacillus thuringiensis.

    PubMed

    Singh, Mamtesh; Kumar, Prasun; Patel, Sanjay K S; Kalia, Vipin C

    2013-03-01

    Integrative processes for the production of bioenergy and biopolymers are gaining importance in recent years as alternatives to fossil fuels and synthetic plastics. In the present study, Bacillus thuringiensis strain EGU45 has been used to generate hydrogen (H2), polyhydroxybutyrate (PHB) and new co-polymers (NP). Under batch culture conditions with 250 ml synthetic media, B. thuringiensis EGU45 produced up to 0.58 mol H2/mol of glucose. Effluent from the H2 production stage was incubated under shaking conditions leading to the production of PHB up to 95 mg/l along with NP of levulinic acid up to 190 mg/l. A twofold to fourfold enhancement in PHB and up to 1.5 fold increase in NP yields was observed on synthetic medium (mixture of M-9+GM-2 medium in 1:1 ratio) containing at 1-2 % glucose concentration. The novelty of this work lies in developing modified physiological conditions, which induce bacterial culture to produce NP. PMID:24426082

  3. Glucose induced fractal colony pattern of Bacillus thuringiensis.

    PubMed

    Roy, Manas K; Banerjee, Paromita; Sengupta, Tapas K; Dattagupta, Sushanta

    2010-08-01

    Growing colonies of bacteria on the surface of thin agar plates exhibit fractal patterns as a result of nonlinear response to environmental conditions, such as nutrients, solidity of the agar medium and temperature. Here, we examine the effect of glucose on pattern formation by growing colonies of Bacillus thuringiensis isolate KPWP1. We also present the theoretical modeling of the colony growth of KPWP1 and the associated spatio-temporal patterns. Our experimental results are in excellent agreement with simulations based on a reaction-diffusion model that describes diffusion-limited aggregation and branching, in which individual cells move actively in the periphery, but become immotile in the inner regions of the growing colony. We obtain the Hausdorff fractal dimension of the colony patterns: D(H.Expt)=1.1969 and D(H, R.D.=)1.1965, for experiment and reaction-diffusion model, respectively. Results of our experiments and modeling clearly show how glucose at higher concentration can prove to be inhibitory for motility of growing colonies of B. thuringiensis cells on semisolid support and be responsible for changes in the growth pattern. PMID:20553734

  4. Mode of action of mosquitocidal Bacillus thuringiensis toxins.

    PubMed

    Soberón, Mario; Fernández, Luisa E; Pérez, Claudia; Gill, Sarjeet S; Bravo, Alejandra

    2007-04-01

    Cry toxins from Bacillus thuringiensis (Bt) are used for insect control. Their primary action is to lyse midgut epithelial cells. In lepidopteran insects, Cry1A monomeric toxins interact with a first receptor and this interaction triggers toxin oligomerization. The oligomeric structure interacts then with a second GPI-anchored receptor that induces insertion into membrane microdomains and larvae death. In the case of mosquitocidal Bt strains, two different toxins participate, Cry and Cyt. These toxins have a synergistic effect and Cyt1Aa overcomes Cry toxin-resistance. We will summarize recent findings on the identification of Cry receptors in mosquitoes and the mechanism of synergism: Cyt1Aa synergizes or suppresses resistance to Cry toxins by functioning as a Cry membrane-bound receptor. PMID:17145072

  5. Bacillus thuringiensis: a specific gamma-cyclodextrin producer strain.

    PubMed

    Goo, Bon Geun; Hwang, You Jin; Park, Jae Kweon

    2014-03-11

    An anaerobic microbial isolate Bacillus species, designated B. thuringiensis GU-2, was isolated from soil as a specific γ-cyclodextrin (CD) producer strain in alkaline medium under anaerobic conditions. The optimum pH and temperature for bacterial growth and γ-CD production were estimated to be pH 8.5 and 37°C in the presence of 1.0% starch substrate, respectively. A high purity yield >95% of γ-CD from the total CD yield in the reaction mixture was obtained from starch that was supposed to be converted by gamma-cyclodextrin glycotransferase, tentatively named as γ-CGTase. The maximum γ-CGTase activity was estimated at 2.45U/mL under optimized condition. This is the first report demonstrating the generation of a specific γ-cyclodextrin (CD) producer strain by the action of a γ-CGTase under anaerobic conditions. PMID:24456970

  6. Novel fermentation media for production of Bacillus thuringiensis subsp. israelensis.

    PubMed

    Poopathi, Subbiah; Kumar, K Anup

    2003-08-01

    The production of Bacillus thuringiensis subsp. israelensis (deBarjac) (Bti) as a biopesticide is not cost-effective using existing fermentation technology. In this study, we explored the use of several less expensive alternative culture media (potato, common sugar, and Bengal gram) for the growth and production of Bti. Growth was obtained in all tested media and was comparable to that obtained in conventional medium (Luria-Bertani). Toxicity assays showed that the toxin produced from the novel growth media were effective in killing larvae of Culex quinquefasciatus, Anopheles stephensi, and Aedes aegypti and toxicity was comparable to that produced from Luria-Bertani medium. These observations suggest that potato can be used as a cheap source of culture medium for the production of Bti toxin in mosquito control programs. PMID:14503573

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

    NASA Astrophysics Data System (ADS)

    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

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

    PubMed

    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. PMID:27052865

  9. 40 CFR 180.1107 - Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Delta endotoxin of Bacillus... From Tolerances § 180.1107 Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta endotoxin...

  10. 40 CFR 180.1107 - Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Delta endotoxin of Bacillus... From Tolerances § 180.1107 Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta endotoxin...

  11. 40 CFR 180.1107 - Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Delta endotoxin of Bacillus... From Tolerances § 180.1107 Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta endotoxin...

  12. 40 CFR 180.1107 - Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Delta endotoxin of Bacillus... From Tolerances § 180.1107 Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta endotoxin...

  13. Recombinant Strain of Bacillus thuringiensis Producing Cyt1A, Cry11B, and the Bacillus sphaericus Binary Toxin

    PubMed Central

    Park, Hyun-Woo; Bideshi, Dennis K.; Federici, Brian A.

    2003-01-01

    A novel recombinant Bacillus thuringiensis subsp. israelensis strain that produces the B. sphaericus binary toxin, Cyt1Aa, and Cry11Ba is described. The toxicity of this strain (50% lethal concentration [LC50] = 1.7 ng/ml) against fourth-instar Culex quinquefasciatus was higher than that of B. thuringiensis subsp. israelensis IPS-82 (LC50 = 7.9 ng/ml) or B. sphaericus 2362 (LC50 = 12.6 ng/ml). PMID:12571069

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

    PubMed

    Perchat, Stéphane; Talagas, Antoine; Poncet, Sandrine; Lazar, Noureddine; Li de la Sierra-Gallay, Inès; Gohar, Michel; Lereclus, Didier; Nessler, Sylvie

    2016-08-01

    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. PMID:27483473

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

    PubMed Central

    Poncet, Sandrine; Lazar, Noureddine; Li de la Sierra-Gallay, Inès; Gohar, Michel; Lereclus, Didier; Nessler, Sylvie

    2016-01-01

    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. PMID:27483473

  16. Spider mite infestations reduce Bacillus thuringiensis toxin concentration in corn leaves and predators avoid spider mites that have fed on Bacillus thuringiensis corn

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenic crops containing pyramid-stacked genes for Bacillus thuringiensis derived toxins for controlling coleopteran and lepidopteran pests are increasingly common. As part of environmental risk assessments, these crops are evaluated for toxicity against non-target organisms, and for their poten...

  17. Comparative genomics analysis of the companion mechanisms of Bacillus thuringiensis Bc601 and Bacillus endophyticus Hbe603 in bacterial consortium

    PubMed Central

    Jia, Nan; Ding, Ming-Zhu; Gao, Feng; Yuan, Ying-Jin

    2016-01-01

    Bacillus thuringiensis and Bacillus endophyticus both act as the companion bacteria, which cooperate with Ketogulonigenium vulgare in vitamin C two-step fermentation. Two Bacillus species have different morphologies, swarming motility and 2-keto-L-gulonic acid productivities when they co-culture with K. vulgare. Here, we report the complete genome sequencing of B. thuringiensis Bc601 and eight plasmids of B. endophyticus Hbe603, and carry out the comparative genomics analysis. Consequently, B. thuringiensis Bc601, with greater ability of response to the external environment, has been found more two-component system, sporulation coat and peptidoglycan biosynthesis related proteins than B. endophyticus Hbe603, and B. endophyticus Hbe603, with greater ability of nutrients biosynthesis, has been found more alpha-galactosidase, propanoate, glutathione and inositol phosphate metabolism, and amino acid degradation related proteins than B. thuringiensis Bc601. Different ability of swarming motility, response to the external environment and nutrients biosynthesis may reflect different companion mechanisms of two Bacillus species. Comparative genomic analysis of B. endophyticus and B. thuringiensis enables us to further understand the cooperative mechanism with K. vulgare, and facilitate the optimization of bacterial consortium. PMID:27353048

  18. Comparative genomics analysis of the companion mechanisms of Bacillus thuringiensis Bc601 and Bacillus endophyticus Hbe603 in bacterial consortium.

    PubMed

    Jia, Nan; Ding, Ming-Zhu; Gao, Feng; Yuan, Ying-Jin

    2016-01-01

    Bacillus thuringiensis and Bacillus endophyticus both act as the companion bacteria, which cooperate with Ketogulonigenium vulgare in vitamin C two-step fermentation. Two Bacillus species have different morphologies, swarming motility and 2-keto-L-gulonic acid productivities when they co-culture with K. vulgare. Here, we report the complete genome sequencing of B. thuringiensis Bc601 and eight plasmids of B. endophyticus Hbe603, and carry out the comparative genomics analysis. Consequently, B. thuringiensis Bc601, with greater ability of response to the external environment, has been found more two-component system, sporulation coat and peptidoglycan biosynthesis related proteins than B. endophyticus Hbe603, and B. endophyticus Hbe603, with greater ability of nutrients biosynthesis, has been found more alpha-galactosidase, propanoate, glutathione and inositol phosphate metabolism, and amino acid degradation related proteins than B. thuringiensis Bc601. Different ability of swarming motility, response to the external environment and nutrients biosynthesis may reflect different companion mechanisms of two Bacillus species. Comparative genomic analysis of B. endophyticus and B. thuringiensis enables us to further understand the cooperative mechanism with K. vulgare, and facilitate the optimization of bacterial consortium. PMID:27353048

  19. ssp genes and spore osmotolerance in Bacillus thuringiensis israelensis and Bacillus sphaericus.

    PubMed

    Cucchi, A; Sanchez de Rivas, C

    1995-10-01

    It was shown previously that spores and vegetative cells of Bacillus sphaericus (Bf) and Bacillus thuringiensis israelensis (Bti) are very sensitive to osmotic variations. Since spore osmotolerance has been associated with their SASP (small acid soluble spore proteins) content coded by ssp genes, hybridization assays were performed with sspE and sspA genes from B. subtilis as probes and showed that Bti and Bf strains could lack an sspE-like gene. The B. subtilis sspE gene was then introduced into Bti 4Q2 strain; spores were obtained and showed a 65 to 650 times higher level of osmotolerance to NaCl, without affecting other important properties: hypoosmotic resistance in vegetative cells, spore UV resistance, and larvicidal activity against diptera larvae. PMID:7549769

  20. 40 CFR 174.517 - Bacillus thuringiensis Cry9C protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry9C protein... Cry9C protein in corn; exemption from the requirement of a tolerance. The plant-incorporated protectant Bacillus thuringiensis Cry9C protein in corn is exempted from the requirement of a tolerance for...

  1. 40 CFR 174.517 - Bacillus thuringiensis Cry9C protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry9C protein... Cry9C protein in corn; exemption from the requirement of a tolerance. The plant-incorporated protectant Bacillus thuringiensis Cry9C protein in corn is exempted from the requirement of a tolerance for...

  2. 40 CFR 174.517 - Bacillus thuringiensis Cry9C protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry9C protein... Cry9C protein in corn; exemption from the requirement of a tolerance. The plant-incorporated protectant Bacillus thuringiensis Cry9C protein in corn is exempted from the requirement of a tolerance for...

  3. 40 CFR 174.517 - Bacillus thuringiensis Cry9C protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Cry9C protein... Cry9C protein in corn; exemption from the requirement of a tolerance. The plant-incorporated protectant Bacillus thuringiensis Cry9C protein in corn is exempted from the requirement of a tolerance for...

  4. 40 CFR 174.517 - Bacillus thuringiensis Cry9C protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry9C protein... Cry9C protein in corn; exemption from the requirement of a tolerance. The plant-incorporated protectant Bacillus thuringiensis Cry9C protein in corn is exempted from the requirement of a tolerance for...

  5. 40 CFR 174.504 - Bacillus thuringiensis Cry1F protein in cotton; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry1F protein... Cry1F protein in cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1F protein in cotton are exempt from the requirement of a tolerance when used as a...

  6. 40 CFR 174.530 - Bacillus thuringiensis Cry2Ae protein in cotton; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry2Ae protein... Cry2Ae protein in cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry2Ae protein in or on the food and feed commodities of cotton; cotton, undelinted seed;...

  7. 40 CFR 174.530 - Bacillus thuringiensis Cry2Ae protein in cotton; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Cry2Ae protein... Cry2Ae protein in cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry2Ae protein in or on the food and feed commodities of cotton; cotton, undelinted seed;...

  8. 40 CFR 174.502 - Bacillus thuringiensis Cry1A.105 protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... protein; exemption from the requirement of a tolerance. 174.502 Section 174.502 Protection of Environment...-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...

  9. 40 CFR 174.529 - Bacillus thuringiensis modified Cry1Ab protein as identified under OECD Unique Identifier SYN...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... protein as identified under OECD Unique Identifier SYN-IR67B-1 in cotton; exemption from the requirement... 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...

  10. 40 CFR 174.520 - Bacillus thuringiensis Cry1F protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry1F protein... Cry1F protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1F protein in corn are exempt from the requirement of a tolerance when used as...

  11. 40 CFR 174.530 - Bacillus thuringiensis Cry2Ae protein in cotton; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry2Ae protein... Cry2Ae protein in cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry2Ae protein in or on the food and feed commodities of cotton; cotton, undelinted seed;...

  12. 40 CFR 174.502 - Bacillus thuringiensis Cry1A.105 protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... protein; exemption from the requirement of a tolerance. 174.502 Section 174.502 Protection of Environment...-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...

  13. 40 CFR 174.520 - Bacillus thuringiensis Cry1F protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry1F protein... Cry1F protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1F protein in corn are exempt from the requirement of a tolerance when used as...

  14. 40 CFR 174.529 - Bacillus thuringiensis modified Cry1Ab protein as identified under OECD Unique Identifier SYN...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... protein as identified under OECD Unique Identifier SYN-IR67B-1 in cotton; exemption from the requirement... 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...

  15. 40 CFR 174.502 - Bacillus thuringiensis Cry1A.105 protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... protein; exemption from the requirement of a tolerance. 174.502 Section 174.502 Protection of Environment...-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...

  16. 40 CFR 174.504 - Bacillus thuringiensis Cry1F protein in cotton; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry1F protein... Cry1F protein in cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1F protein in cotton are exempt from the requirement of a tolerance when used as a...

  17. 40 CFR 174.529 - Bacillus thuringiensis modified Cry1Ab protein as identified under OECD Unique Identifier SYN...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... protein as identified under OECD Unique Identifier SYN-IR67B-1 in cotton; exemption from the requirement... 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...

  18. 40 CFR 174.502 - Bacillus thuringiensis Cry1A.105 protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... protein; exemption from the requirement of a tolerance. 174.502 Section 174.502 Protection of Environment...-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...

  19. 40 CFR 174.504 - Bacillus thuringiensis Cry1F protein in cotton; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry1F protein... Cry1F protein in cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1F protein in cotton are exempt from the requirement of a tolerance when used as a...

  20. 40 CFR 174.520 - Bacillus thuringiensis Cry1F protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry1F protein... Cry1F protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1F protein in corn are exempt from the requirement of a tolerance when used as...

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

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... protein as identified under OECD Unique Identifier SYN-IR67B-1 in cotton; exemption from the requirement... 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...

  2. Expression of Bacillus thuringiensis delta-endotoxin genes during vegetative growth.

    PubMed Central

    Mettus, A M; Macaluso, A

    1990-01-01

    Bacillus thuringiensis delta-endotoxin (crystal protein) genes are normally expressed only during sporulation. It is possible to produce crystal protein during vegetative growth by placing B. thuringiensis crystal protein genes downstream of a strong vegetative promoter. By removing a possible transcriptional terminator of the tetracycline resistance gene of pBC16 and inserting a multiple cloning site, delta-endotoxin genes can be cloned downstream from the tetracycline resistance gene promoter. This construct allows for readthrough transcription from the strong vegetative promoter. Crystal protein is then produced during vegetative growth as well as during sporulation in both B. thuringiensis and Bacillus megaterium. This construct also allows for production of delta-endotoxin in B. thuringiensis strains that do not normally produce delta-endotoxin because of a defect in sporulation. Images PMID:2160219

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

    PubMed Central

    Zahner, Viviane; Silva, Ana Carolina Telles de Carvalho e; de Moraes, Gabriela Pinhel; McIntosh, Douglas; de Filippis, Ivano

    2013-01-01

    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. PMID:23440117

  4. IS231A from Bacillus thuringiensis is functional in Escherichia coli: transposition and insertion specificity.

    PubMed Central

    Hallet, B; Rezsöhazy, R; Delcour, J

    1991-01-01

    A kanamycin resistance gene was introduced within the insertion sequence IS231A from Bacillus thuringiensis, and transposition of the element was demonstrated in Escherichia coli. DNA sequencing at the target sites showed that IS231A transposition results in direct repeats of variable lengths (10, 11, and 12 bp). These target sequences resemble the terminal inverted repeats of the transposon Tn4430, which are the preferred natural insertion sites of IS231 in B. thuringiensis. Images PMID:1648561

  5. The impact of secondary pests on Bacillus thuringiensis (Bt) crops.

    PubMed

    Catarino, Rui; Ceddia, Graziano; Areal, Francisco J; Park, Julian

    2015-06-01

    The intensification of agriculture and the development of synthetic insecticides enabled worldwide grain production to more than double in the last third of the 20th century. However, the heavy dependence and, in some cases, overuse of insecticides has been responsible for negative environmental and ecological impacts across the globe, such as a reduction in biodiversity, insect resistance to insecticides, negative effects on nontarget species (e.g. natural enemies) and the development of secondary pests. The use of recombinant DNA technology to develop genetically engineered insect-resistant crops could mitigate many of the negative side effects of insecticides. One such genetic alteration enables crops to express toxic crystalline (Cry) proteins from the soil bacteria Bacillus thuringiensis (Bt). Despite the widespread adoption of Bt crops, there are still a range of unanswered questions concerning longer term agro-ecosystem interactions. For instance, insect species that are not susceptible to the expressed toxin can develop into secondary pests and cause significant damage to the crop. Here, we review the main causes surrounding secondary pest dynamics in Bt crops and the impact of such outbreaks. Regardless of the causes, if nonsusceptible secondary pest populations exceed economic thresholds, insecticide spraying could become the immediate solution at farmers' disposal, and the sustainable use of this genetic modification technology may be in jeopardy. Based on the literature, recommendations for future research are outlined that will help to improve the knowledge of the possible long-term ecological trophic interactions of employing this technology. PMID:25832330

  6. Raman spectroscopy of Bacillus thuringiensis physiology and inactivation

    NASA Astrophysics Data System (ADS)

    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. Role of receptors in Bacillus thuringiensis crystal toxin activity.

    PubMed

    Pigott, Craig R; Ellar, David J

    2007-06-01

    Bacillus thuringiensis produces crystalline protein inclusions with insecticidal or nematocidal properties. These crystal (Cry) proteins determine a particular strain's toxicity profile. Transgenic crops expressing one or more recombinant Cry toxins have become agriculturally important. Individual Cry toxins are usually toxic to only a few species within an order, and receptors on midgut epithelial cells have been shown to be critical determinants of Cry specificity. The best characterized of these receptors have been identified for lepidopterans, and two major receptor classes have emerged: the aminopeptidase N (APN) receptors and the cadherin-like receptors. Currently, 38 different APNs have been reported for 12 different lepidopterans. Each APN belongs to one of five groups that have unique structural features and Cry-binding properties. While 17 different APNs have been reported to bind to Cry toxins, only 2 have been shown to mediate toxin susceptibly in vivo. In contrast, several cadherin-like proteins bind to Cry toxins and confer toxin susceptibility in vitro, and disruption of the cadherin gene has been associated with toxin resistance. Nonetheless, only a small subset of the lepidopteran-specific Cry toxins has been shown to interact with cadherin-like proteins. This review analyzes the interactions between Cry toxins and their receptors, focusing on the identification and validation of receptors, the molecular basis for receptor recognition, the role of the receptor in resistant insects, and proposed models to explain the sequence of events at the cell surface by which receptor binding leads to cell death. PMID:17554045

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

    PubMed Central

    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

  9. [Effect of pesticides on Bacillus thuringiensis strains under controlled conditions].

    PubMed

    Salerno, C; Dias, S; Sagardoy, M

    1999-01-01

    Little is known about native populations of Bacillus thuringiensis (Bt) isolated from soils of Argentina. We undertook this study to determine the resistance to different pesticides of two commercial and fourteen native strains of Bt under in vitro conditions. An agar plate bioassay test conducted with ten pesticides and sixteen strains of Bt showed that Basagran, Scepter, Fungoxan and Decis were not toxic for the bioinsecticide bacteria at recommended application rates (RAR). In contrast, low concentrations (3.2% RAR) of Agil, Select and Isomero showed a deleterious effect on the bacteria investigated. Simultaneously, four of the pesticides were able to produce phenotypical changes on the Bt colonies grown on nutrient agar. Moreover, in a greenhouse experiment, seven pesticides applied at 1.6%, 12.5% and 100% RAR on soybean leaves were not as inhibitory as under in vitro conditions for two Bt strains (HD-1 and A61). However, survival of these strains in the phyllosphere of soybean differed significantly between untreated leaves and leaves treated with pesticides after 20 days of study (P < 0.05). Finally, and from an ecological point of view, these findings suggest that the addition of some pesticides to soybean leaves in lower concentrations than those recommended could be favourable for the persistence of Bt in this environment. PMID:10425660

  10. Evolution of Bacillus thuringiensis Cry toxins insecticidal activity

    PubMed Central

    Bravo, Alejandra; Gómez, Isabel; Porta, Helena; García-Gómez, Blanca Ines; Rodriguez-Almazan, Claudia; Pardo, Liliana; Soberón, Mario

    2013-01-01

    Insecticidal Cry proteins produced by Bacillus thuringiensis are use worldwide in transgenic crops for efficient pest control. Among the family of Cry toxins, the three domain Cry family is the better characterized regarding their natural evolution leading to a large number of Cry proteins with similar structure, mode of action but different insect specificity. Also, this group is the better characterized regarding the study of their mode of action and the molecular basis of insect specificity. In this review we discuss how Cry toxins have evolved insect specificity in nature and analyse several cases of improvement of Cry toxin action by genetic engineering, some of these examples are currently used in transgenic crops. We believe that the success in the improvement of insecticidal activity by genetic evolution of Cry toxins will depend on the knowledge of the rate-limiting steps of Cry toxicity in different insect pests, the mapping of the specificity binding regions in the Cry toxins, as well as the improvement of mutagenesis strategies and selection procedures. PMID:22463726

  11. Continuous evolution of Bacillus thuringiensis toxins overcomes insect resistance.

    PubMed

    Badran, Ahmed H; Guzov, Victor M; Huai, Qing; Kemp, Melissa M; Vishwanath, Prashanth; Kain, Wendy; Nance, Autumn M; Evdokimov, Artem; Moshiri, Farhad; Turner, Keith H; Wang, Ping; Malvar, Thomas; Liu, David R

    2016-05-01

    The Bacillus thuringiensis δ-endotoxins (Bt toxins) are widely used insecticidal proteins in engineered crops that provide agricultural, economic, and environmental benefits. The development of insect resistance to Bt toxins endangers their long-term effectiveness. Here we have developed a phage-assisted continuous evolution selection that rapidly evolves high-affinity protein-protein interactions, and applied this system to evolve variants of the Bt toxin Cry1Ac that bind a cadherin-like receptor from the insect pest Trichoplusia ni (TnCAD) that is not natively bound by wild-type Cry1Ac. The resulting evolved Cry1Ac variants bind TnCAD with high affinity (dissociation constant Kd = 11-41 nM), kill TnCAD-expressing insect cells that are not susceptible to wild-type Cry1Ac, and kill Cry1Ac-resistant T. ni insects up to 335-fold more potently than wild-type Cry1Ac. Our findings establish that the evolution of Bt toxins with novel insect cell receptor affinity can overcome insect Bt toxin resistance and confer lethality approaching that of the wild-type Bt toxin against non-resistant insects. PMID:27120167

  12. Bacillus thuringiensis: A story of a successful bioinsecticide

    PubMed Central

    Bravo, Alejandra; Likitvivatanavong, Supaporn; Gill, Sarjeet S.; Soberón, Mario

    2013-01-01

    Bacillus thuringiensis (Bt) bacteria are insect pathogens that rely on insecticidal pore forming proteins known as Cry and Cyt toxins to kill their insect larval hosts. At least four different non-structurally related families of proteins form the Cry toxin group of toxins. The expression of certain Cry toxins in transgenic crops has contributed to an efficient control of insect pests resulting in a significant reduction in chemical insecticide use. The mode of action of the three domain Cry toxin family involves sequential interaction of these toxins with several insect midgut proteins facilitating the formation of a pre-pore oligomer structure and subsequent membrane insertion that leads to the killing of midgut insect cells by osmotic shock. In this manuscript we review recent progress in understanding the mode of action of this family of proteins in lepidopteran, dipteran and coleopteran insects. Interestingly, similar Cry-binding proteins have been identified in the three insect orders, as cadherin, aminopeptidase-N and alkaline phosphatase suggesting a conserved mode of action. Also, recent data on insect responses to Cry toxin attack is discussed. Finally, we review the different Bt based products, including transgenic crops, that are currently used in agriculture. PMID:21376122

  13. Development of sludge based stable aqueous Bacillus thuringiensis formulations.

    PubMed

    Brar, S K; Verma, M; Tyagi, R D; Valéro, J R; Surampalli, R Y; Banerji, S K

    2004-01-01

    This study focuses on development of aqueous flowable (suspension) formulations for Bacillus thuringiensis (Bt) based biopesticides from wastewater sludge. Different inerts like sorbitol, sodium monophosphate, sodium metabisulphite, sorbic acid, propionic acid, Tween-80, Triton X-100 and glycerol were tested for formulations. Five different formulations for non-hydrolyzed (NH) secondary sludges were tried and the best combination selected on the basis of various physical parameters like viscosity, particle size, suspendibility, entomotoxicity, and microbiological purity tests. F5 formulations (for secondary sludge) comprising sorbitol, sodium monophosphate and sodium metabisulphite gave better physical and biological characteristics with a small effect on entomotoxicity and spore concentration after 120 days at pH 6, 6.5 and temperatures 40 and 50 degrees C and viscosity change at 40 and 50 degrees C. The formulations were more stable at pH 4.0 to 5.0 and temperatures 4 to 30 degrees C whereas at pH 6.0 and 6.5 and temperatures 40 and 50 degrees C, there was degeneration of the product. Lower proteolytic activity and physical factors like ionic strength and surface group changes at pH 6 and 6.5 were responsible for the instability of the formulation. PMID:15581017

  14. Starch industry wastewater-based stable Bacillus thuringiensis liquid formulations.

    PubMed

    Brar, Satinder K; Verma, M; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2005-12-01

    Liquid formulations were developed from Bacillus thuringiensis (Bt)-fermented broths of starch industry wastewater (SIW) and of soya medium. Stability studies were carried out for 1 yr. Storage stability was tested by studying various physical and chemical (e.g., viscosity, particle size, corrosion, and suspendibility) and biological (e.g., microbial contamination, viable spores, and entomotoxicity) parameters at different pH levels and temperatures. Three suspending agents, sorbitol, sodium monophosphate, and sodium metabisulfite, were added to fermented broth in different concentrations. Sorbitol and sodium monophosphate in the ratio 3:1 was the best suspending agent combination for both formulations. Starch industry wastewater fermentation yielded cell and viable spore counts 10- and 4-fold greater than those from soya medium, respectively, and a 1.7-fold increase in entomotoxicity. However, both formulations started deteriorating at pH 6 and 6.5 and 40 and 50 degrees C. There were no signs of corrosion and microbial contamination in both types of formulations. PMID:16539110

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

    SciTech Connect

    Chung, Eunhyea; Kweon, Hyojin; Yiacoumi, Sotira; Lee, Ida; Joy, David Charles; Palumbo, Anthony Vito; Tsouris, Costas

    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.

  16. Investigation of lead(II) uptake by Bacillus thuringiensis 016.

    PubMed

    Chen, Zhi; Pan, Xiaohong; Chen, Hui; Lin, Zhang; Guan, Xiong

    2015-11-01

    In this work, we investigated the lead(II) biosorption mechanism of Bacillus thuringiensis (Bt) 016 through batch and microscopic experiments. We found that the maximum lead(II) biosorption capacity of Bt 016 was 164.77 mg/g (dry weight). The pH value could affect the biosorption of lead(II) in a large extent. Fourier transform infrared analyses and selective passivation experiments suggested that the carboxyl, amide and phosphate functional groups of Bt 016 played an important role in lead(II) biosorption. Scanning electron microscopy observation showed that noticeable lead(II) precipitates were accumulated on bacterial surfaces. Further transmission electron microscopy thin section analysis coupled with energy dispersive X-ray spectroscopy as well as selected area electron diffraction indicated that lead(II) immobilized on the bacteria could be transformated into random-shaped crystalline lead-containing minerals eventually. This work provided a new insight into lead(II) uptake of Bt, highlighting the potential of Bt in the restoration of lead(II) contaminated repositories. PMID:26271773

  17. Monarch larvae sensitivity to Bacillus thuringiensis- purified proteins and pollen

    PubMed Central

    Hellmich, Richard L.; Siegfried, Blair D.; Sears, Mark K.; Stanley-Horn, Diane E.; Daniels, Michael J.; Mattila, Heather R.; Spencer, Terrence; Bidne, Keith G.; Lewis, Leslie C.

    2001-01-01

    Laboratory tests were conducted to establish the relative toxicity of Bacillus thuringiensis (Bt) toxins and pollen from Bt corn to monarch larvae. Toxins tested included Cry1Ab, Cry1Ac, Cry9C, and Cry1F. Three methods were used: (i) purified toxins incorporated into artificial diet, (ii) pollen collected from Bt corn hybrids applied directly to milkweed leaf discs, and (iii) Bt pollen contaminated with corn tassel material applied directly to milkweed leaf discs. Bioassays of purified Bt toxins indicate that Cry9C and Cry1F proteins are relatively nontoxic to monarch first instars, whereas first instars are sensitive to Cry1Ab and Cry1Ac proteins. Older instars were 12 to 23 times less susceptible to Cry1Ab toxin compared with first instars. Pollen bioassays suggest that pollen contaminants, an artifact of pollen processing, can dramatically influence larval survival and weight gains and produce spurious results. The only transgenic corn pollen that consistently affected monarch larvae was from Cry1Ab event 176 hybrids, currently <2% corn planted and for which re-registration has not been applied. Results from the other types of Bt corn suggest that pollen from the Cry1Ab (events Bt11 and Mon810) and Cry1F, and experimental Cry9C hybrids, will have no acute effects on monarch butterfly larvae in field settings. PMID:11559841

  18. Complete genome sequence of Bacillus thuringiensis subsp. chinensis strain CT-43.

    PubMed

    He, Jin; Wang, Jieping; Yin, Wen; Shao, Xiaohu; Zheng, Huajun; Li, Mingshun; Zhao, Youwen; Sun, Ming; Wang, Shengyue; Yu, Ziniu

    2011-07-01

    Bacillus thuringiensis has been widely used as an agricultural biopesticide for a long time. As a producing strain, B. thuringiensis subsp. chinensis strain CT-43 is highly toxic to lepidopterous and dipterous insects. It can form various parasporal crystals consisting of Cry1Aa3, Cry1Ba1, Cry1Ia14, Cry2Aa9, and Cry2Ab1. During fermentation, it simultaneously generates vegetative insecticidal protein Vip3Aa10 and the insecticidal nucleotide analogue thuringiensin. Here, we report the finished, annotated genome sequence of B. thuringiensis strain CT-43. PMID:21551307

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

    PubMed Central

    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

  20. Interaction between the predator Podisus nigrispinus (Hemiptera: Pentatomidae) and the entomopathogenic bacteria Bacillus thuringiensis.

    PubMed

    Carvalho, V F P; Vacari, A M; Pomari, A F; De Bortoli, C P; Ramalho, D G; De Bortoli, S A

    2012-12-01

    Plutella xylostella (L.) is susceptible to both the entomopathogen Bacillus thuringiensis and the predator, Brazilian spined soldier bug [Podisus nigrispinus (Dallas)]. The objective of this study was to measure the interaction between the bacterium B. thuringiensis and the predator P. nigrispinus. We also studied the behavior of P. nigrispinus in relation to its choice between B. thuringiensis-infected and healthy P. xylostellais larvae. In the first treatment, P. nigrispinus nymphs were fed daily with B. thuringiensis-infected P. xylostella larvae and distilled water. In the second treatment, nymphs were fed daily with healthy larvae and a suspension of B. thuringiensis as a source of water. The control nymphs were fed daily with healthy larvae and water. Adult P. nigrispinus were separated by sex, couples were formed, and they were fed daily with P. xylostella larvae derived from the treatments. We followed the development of P. nigrispinus and measured its biological characteristics. On the basis of these data, parameters were determined for the construction of life tables. A choice test was used to compare infected and healthy larvae. The HD1 strain of B. thuringiensis does not affect the biological characteristics of P. nigrispinus when fed infected larvae and water or healthy larvae and B. thuringiensis suspension. Our study shows that integrated management of P. xylostella, a pest of the Brassicaceae, is feasible by using the HD1 strain of B. thuringiensis and the predator P. nigrispinus, because the predator shows no preference for infected or healthy P. xylostella larvae. PMID:23321092

  1. Field evaluation of Bacillus thuringiensis H-14 against Aedes mosquitoes.

    PubMed

    Lee, Y W; Zairi, J

    2006-06-01

    Studies were carried out on the residual efficacy of Bacillus thuringiensis H-14 (water dispersible granule, VectoBac ABG 6511) as direct application in the control of Aedes larvae in the field. Field Aedes sp populations in the earthen and glass jars were predetermined before initiation of the trial. On confirmation of the presence of Aedes species in the designated area, Sungai Nibong Kecil, Penang Island, Malaysia, Bti was introduced in the 55L earthen and 3L glass jars). Two test designs were carried out. The first design had treated water replenished daily with 6L of seasoned water and the second design is without the replenishment of water but evaporated water was replenished. Bti was effective in the field for at least 35 days with more than 80% reduction in the Aedes larvae in the treated containers. For earthen jars with daily replenishment of water, 100% reduction was recorded for the first 3 days, while more than 80% reduction was recorded up to day 40. At day 60, Bti still provided an efficacy of 54.32 +/- 4.61 (%) of reduction. Whilst for earthen jars without daily replenishment of water, 100% reduction was recorded for the first 5 days, while more than 80% of reduction was recorded up to day 40. For the glass jars studied, similar efficacy was observed. In jars with daily replenishment of water a better larval control was observed. Percentage of reduction from day 50 to 60 for replenishment of water was between 50 to 70% compared to without replenishment of water with less than 40%. PMID:17041550

  2. Spatio-Temporal Evolution of Sporulation in Bacillus thuringiensis Biofilm.

    PubMed

    El-Khoury, Nay; Majed, Racha; Perchat, Stéphane; Kallassy, Mireille; Lereclus, Didier; Gohar, Michel

    2016-01-01

    Bacillus thuringiensis can produce a floating biofilm which includes two parts: a ring and a pellicle. The ring is a thick structure which sticks to the culture container, while the pellicle extends over the whole liquid surface and joins the ring. We have followed over time, from 16 to 96 h, sporulation in the two biofilm parts. Sporulation was followed in situ in 48-wells polystyrene microtiterplates with a fluorescence binocular stereomicroscope and a spoIID-yfp transcriptional fusion. Sporulation took place much earlier in the ring than in the pellicle. In 20 h-aged biofilms, spoIID was expressed only in the ring, which could be seen as a green fluorescent circle surrounding the non-fluorescent pellicle. However, after 48 h of culture, the pellicle started to express spoIID in specific area corresponding to protrusions, and after 96 h both the ring and the whole pellicle expressed spoIID. Spore counts and microscopy observations of the ring and the pellicle harvested separately confirmed these results and revealed that sporulation occured 24 h-later in the pellicle comparatively to the ring, although both structures contained nearly 100% spores after 96 h of culture. We hypothesize that two mechanisms, due to microenvironments in the biofilm, can explain this difference. First, the ring experiences a decreased concentration of nutrients earlier than the pellicle, because of a lower exchange area with the culture medium. An second, the ring is exposed to partial dryness. Both reasons could speed up sporulation in this biofilm structure. Our results also suggest that spores in the biofilm display a phenotypic heterogeneity. These observations might be of particular significance for the food industry, since the biofilm part sticking to container walls - the ring - is likely to contain spores and will therefore resist both to washing and to cleaning procedures, and will be able to restart a new biofilm when food production has resumed. PMID:27536298

  3. Spatio-Temporal Evolution of Sporulation in Bacillus thuringiensis Biofilm

    PubMed Central

    El-Khoury, Nay; Majed, Racha; Perchat, Stéphane; Kallassy, Mireille; Lereclus, Didier; Gohar, Michel

    2016-01-01

    Bacillus thuringiensis can produce a floating biofilm which includes two parts: a ring and a pellicle. The ring is a thick structure which sticks to the culture container, while the pellicle extends over the whole liquid surface and joins the ring. We have followed over time, from 16 to 96 h, sporulation in the two biofilm parts. Sporulation was followed in situ in 48-wells polystyrene microtiterplates with a fluorescence binocular stereomicroscope and a spoIID-yfp transcriptional fusion. Sporulation took place much earlier in the ring than in the pellicle. In 20 h-aged biofilms, spoIID was expressed only in the ring, which could be seen as a green fluorescent circle surrounding the non-fluorescent pellicle. However, after 48 h of culture, the pellicle started to express spoIID in specific area corresponding to protrusions, and after 96 h both the ring and the whole pellicle expressed spoIID. Spore counts and microscopy observations of the ring and the pellicle harvested separately confirmed these results and revealed that sporulation occured 24 h-later in the pellicle comparatively to the ring, although both structures contained nearly 100% spores after 96 h of culture. We hypothesize that two mechanisms, due to microenvironments in the biofilm, can explain this difference. First, the ring experiences a decreased concentration of nutrients earlier than the pellicle, because of a lower exchange area with the culture medium. An second, the ring is exposed to partial dryness. Both reasons could speed up sporulation in this biofilm structure. Our results also suggest that spores in the biofilm display a phenotypic heterogeneity. These observations might be of particular significance for the food industry, since the biofilm part sticking to container walls – the ring – is likely to contain spores and will therefore resist both to washing and to cleaning procedures, and will be able to restart a new biofilm when food production has resumed. PMID:27536298

  4. Transfer of Bacillus thuringiensis plasmids coding for delta-endotoxin among strains of B. thuringiensis and B. cereus.

    PubMed Central

    González, J M; Brown, B J; Carlton, B C

    1982-01-01

    The recently discovered high-frequency transfer of plasmids between strains of Bacillus thuringiensis was used to study the genetic relationship between plasmids and production of the insecticidal delta-endotoxin crystal. Three strains of B. thuringiensis transmitted the Cry+ (crystal-producing) phenotype to Cry- (acrystalliferous) B. thuringiensis recipients. Agarose gel electrophoresis showed that one specific plasmid from each donor strain was always present in Cry+ "transcipients." The size of the transmissible crystal-coding plasmid varied with the donor strain, being 75 MDal (megadaltons) in size in HD-2, 50 MDal in HD-73, and 44 MDal in HD-263. Immunological analysis showed the Cry+ transcipients to be hybrid strains, having flagella of the recipient serotype and crystals of the donor serotype. These results demonstrate that the structural genes for the delta-endotoxin are plasmid borne. Crystal-coding plasmids also transferred into two strains of the related species Bacillus cereus and yielded transcipients that produced crystals of the same antigenicity as the donor strain. Images PMID:6294667

  5. Urea-Mercaptoethanol-Soluble Protein from Spores of Bacillus thuringiensis and Other Species

    PubMed Central

    Somerville, H. J.; Delafield, F. P.; Rittenberg, S. C.

    1970-01-01

    Treatment with urea-mercaptoethanol of purified spores of Bacillus thuringiensis, other Bacillus species, and Clostridium roseum solubilizes a protein fraction between 5 and 12% of the dry weight of the spores. This fraction behaves identically to the crystal protein of B. thuringiensis on acrylamide-gel electrophoresis. The protein from all of the Bacillus species shows partial homology with crystal protein, using the Ouchterlony immunodiffusion technique. A further fraction, similar in amount, can be removed from spores of B. thuringiensis by the addition of sodium lauryl sulfate to the urea-mercaptoethanol. Spores of B. thuringiensis extracted in these ways show no difference when compared to untreated spores with respect to viability or resistance to heat and ultraviolet-irradiation. The extracted spores do show differences in their germination requirements and their susceptibility to phase-darkening by lysozyme. It is concluded that an urea-mercaptoethanol-soluble protein or class of protein is a widespread component of bacterial spores, possibly located in the spore coat, and that this protein may be related to the crystal protein of B. thuringiensis. Images PMID:4984077

  6. 40 CFR 174.501 - Bacillus thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Vip3Aa protein... thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Vip3Aa proteins in or on corn or cotton are exempt from the requirement of...

  7. 40 CFR 174.501 - Bacillus thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Vip3Aa protein... thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Vip3Aa proteins in or on corn or cotton are exempt from the requirement of...

  8. 40 CFR 174.501 - Bacillus thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Vip3Aa protein... thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Vip3Aa proteins in or on corn or cotton are exempt from the requirement of...

  9. 40 CFR 174.501 - Bacillus thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Vip3Aa protein... thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Vip3Aa proteins in or on corn or cotton are exempt from the requirement of...

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

    PubMed Central

    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 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. PMID:21255282

  11. A preliminary study of the bioactivity of vegetative proteins extracted from Malaysian Bacillus thuringiensis isolates.

    PubMed

    Ramasamy, B; Nadarajah, V D; Soong, Z K; Lee, H L; Mohammad, S M

    2008-04-01

    Vegetative proteins from Malaysian strains of Bacillus thuringiensis israelensis strains (Bt 11, Bt 12, Bt 15, Bt 16, Bt 17, Bt 21 and Bt 22) and Bacillus sphaericus H-25 strains (Bs 1 and Bs 2) were screened for haemolytic, cytotoxic and larvicidal activity. SDS-PAGE profiles of the Bacillus thuringiensis strains studied consistently showed major bands of 33-37 kDa and 47 kDa. Bt 16 also showed two bands of 66 kDa and 45 kDa similar to the previously reported binary vegetative protein, Vip1Ac (66 kDa) and Vip 2Ac (45 kDa). Both the Bacillus sphaericus strains showed a 35 kDa band that was similiar to a previously reported vegetative protein, the Mtx2 protein. Bs 2 also contains a 37 kDa band, similar to another vegetative protein, the Mtx 3 protein. With the exception of Bt 17 and Bt 21, vegetative proteins from all Bacillus thuringiensis and Bacillus sphaericus strains were highly haemolytic to human erythrocytes, causing more than 75% haemolysis at the highest concentration of 200 microg/ml. High haemolytic activity was associated with high cytotoxic activity with most of the haemolytic strains being indiscriminately cytotoxic to both CEM-SS (human T lymphoblastoid) and HeLa (human uterus cervical cancer) cell lines. Interestingly, the less haemolytic vegetative proteins from Bt 17 and Bt 21 demonstrated cytotoxic activity comparable to that of the highly haemolytic vegetative proteins. Bt 21 displayed toxicity towards both cell lines while Bt 17 was more toxic towards CEM-SS cells. Bioassay against Aedes aegypti and Culex quinquefasciatus larvae revealed that vegetative proteins from the Bacillus thuringiensis strains had activity against both species of larvae but vegetative proteins from Bacillus sphaericus were weakly larvicidal towards Cx. quinquefasciatus only. PMID:18600206

  12. Analysis of opportunities and challenges in patenting of Bacillus thuringiensis insecticidal crystal protein genes.

    PubMed

    Swamy, H M Mahadeva; Asokan, R; Rajasekaran, P E; Mahmood, Riaz; Nagesha, S N; Arora, D K

    2012-04-01

    Bacillus thuringiensis (Bt) is the most widely used microbial control agent. The broad spectrum of susceptible hosts, production on artificial media and ease of application has caused the widespread use of this bacterium against several pests in agriculture, forest and vectors of human diseases. B.thuringiensis toxins are highly species specific which provide economic, environmental benefits, potential for future control and spread of the technology worldwide. This makes the B. thuringiensis crystal proteins an interesting tool for the implementation in integrated pest management programs. It has gained importance over the last 100 years for its biocontrol properties which is used in this review as a case study and analysis of the patents granted on B. thuringiensis was carried out. This study categorizes a number of patents related to B.thuringiensis insecticidal crystal proteins, application of B.thuringiensis insecticidal crystal proteins and the development of patentable technologies. The analyses were done using various criteria like patenting trends over the years, assignees playing a major role, comparison of the technology used in different patents and the patenting activity across the insect orders. Patent documents related to bacterium B.thuringiensis contain a trove of technical and commercial information and thus, patent analysis is considered as a useful tool for R management and techno economical development. Patent analysis also helps identifying and evaluating new and alternate technologies, keeping abreast with latest technologies for business interests, finding solutions to technical problems and ideas for new innovative trends. PMID:22239684

  13. Bacillus thuringiensis as a surrogate for Bacillus anthracis in aerosol research.

    PubMed

    Tufts, Jenia A M; Calfee, M Worth; Lee, Sang Don; Ryan, Shawn P

    2014-05-01

    Characterization of candidate surrogate spores prior to experimental use is critical to confirm that the surrogate characteristics are as closely similar as possible to those of the pathogenic agent of interest. This review compares the physical properties inherent to spores of Bacillus anthracis (Ba) and Bacillus thuringiensis (Bt) that impact their movement in air and interaction with surfaces, including size, shape, density, surface morphology, structure and hydrophobicity. Also evaluated is the impact of irradiation on the physical properties of both Bacillus species. Many physical features of Bt and Ba have been found to be similar and, while Bt is considered typically non-pathogenic, it is in the B. cereus group, as is Ba. When cultured and sporulated under similar conditions, both microorganisms share a similar cylindrical pellet shape, an aerodynamic diameter of approximately 1 μm (in the respirable size range), have an exosporium with a hairy nap, and have higher relative hydrophobicities than other Bacillus species. While spore size, morphology, and other physical properties can vary among strains of the same species, the variations can be due to growth/sporulation conditions and may, therefore, be controlled. Growth and sporulation conditions are likely among the most important factors that influence the representativeness of one species, or preparation, to another. All Bt spores may, therefore, not be representative of all Ba spores. Irradiated spores do not appear to be a good surrogate to predict the behavior of non-irradiated spores due to structural damage caused by the irradiation. While the use of Bt as a surrogate for Ba in aerosol testing appears to be well supported, this review does not attempt to narrow selection between Bt strains. Comparative studies should be performed to test the hypothesis that viable Ba and Bt spores will behave similarly when suspended in the air (as an aerosol) and to compare the known microscale characteristics

  14. Phylogenetic distribution of phenotypic traits in bacillus thuringiensis analyzed by multilocus sequence typing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Strains from a collection of 3,639 diverse Bacillus thuringiensis isolates were classified based on phenotypic profiles resulting from six biochemical tests, including production of amylase (T), lecithinase (L), urease (U), acid from sucrose (S) and salicin (A), and the hydrolysis of esculin (E). St...

  15. Toxicity of "Bacillus thuringiensis var. Kurstaki" to the Painted Lady Butterfly, Vanessa cardui.

    ERIC Educational Resources Information Center

    Stalter, Richard; Nadal, Gerard; Kincaid, Dwight

    2000-01-01

    Reports the effects of Bacillus thuringiensis var. Kurstaki (BT), which is highly toxic, to a non-target lepidopteran, the Painted Lady butterfly. Indicates that BT kills some Painted Lady butterfly larvae at the lowest dilution tested after 48 hours. (ASK)

  16. Bacillus thuringiensis Cry3Aa toxin increases the susceptibility of Crioceris quatuordecimpunctata to Beauveria bassiana infection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The spotted asparagus beetle, Crioceris quatuordecimpunctata (Coleoptera: Chrysomelidae), is one of the most devastating pests of asparagus in China and elsewhere. In this study, we investigated the interaction of Bacillus thuringiensis (Bt) Cry3Aa toxin and the entomopathogenic fungus Beauveria bas...

  17. Complete Genome Sequence of Bacillus thuringiensis Serovar Tolworthi Strain Pasteur Institute Standard

    PubMed Central

    Kanda, Kohzo; Nakashima, Kaede

    2015-01-01

    The genome sequence of Bacillus thuringiensis serovar tolworthi strain Pasteur Institute Standard was determined. The genome consists of a 5.9-Mb chromosome and eight plasmids, one of which is linear. The second largest plasmid (293 kb) carries the genes encoding insecticidal proteins. PMID:26139717

  18. Transcriptome of the gypsy moth (Lymantria dispar) larval midgut in response to infection by Bacillus thuringiensis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transcriptomic profiles of the lepidopteran insect pest Lymantria dispar (gypsy moth) were characterized in the larval midgut in response to infection by the biopesticide Bacillus thuringiensis kurstaki. RNA-Seq approaches were used to define a set of 49,613 assembled transcript sequences, of which...

  19. A Novel Tenebrio molitor Cadherin is a Functional Receptor for Bacillus thuringiensis Toxin Cry3Aa

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cry toxins produced by the bacterium Bacillus thuringiensis (Bt) are effective biological insecticides. Cadherin-like proteins have been reported as functional Cry1A toxin receptors in Lepidoptera. We present the first report demonstrating a functional interaction between the coleopteran-specific ...

  20. COMPARATIVE ANALYSIS OF PROTEINASE ACTIVITIES OF BACILLUS THURINGIENSIS-RESISTANT AND -SUSCEPTIBLE OSTRINIA NUBILALIS (LEPIDOPTERA: CRAMBIDAE)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Proteinase activities were compared in soluble and membrane fractions of gut tissues of Bacillus thuringiensis-resistant and -susceptible Ostrinia nubilalis larvae. The soluble trypsin-like proteinase activity of the resistant strain was reduced 56%, significantly lower than that of the susceptibl...

  1. 40 CFR 180.1107 - Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Delta endotoxin of Bacillus thuringiensis variety kurstaki encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. 180.1107 Section 180.1107 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS TOLERANCES...

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

    EPA Science Inventory

    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...

  3. INGESTION AND ADSORPTION OF 'BACILLUS THURINGIENSIS' SUBSP. 'ISRAELENSIS' BY 'GAMMARUS LACUSTRIS' IN THE LABORATORY

    EPA Science Inventory

    Several groups of Gammarus lacustris adults were exposed to solutions containing 0.5 and 5.0 mg of Bacillus thuringiensis subsp. israelensis per liter for 1- or 24-hour periods by using traditional static bioassay exposure procedures. The experiments verified that traditional exp...

  4. Mineralization of the Bacillus thuringiensis Cry1Ac endotoxin in soil

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Despite a number of studies describing the fate of Bacillus thuringiensis insecticidal endotoxins in soil have been conducted in the past decade, conflicting information on persistence of this class of insecticidal toxins exists. In the present experiment, 14C from glucose was incorporated into the ...

  5. Screening Bacillus thuringiensis strains for toxicity against Manduca sexta and Plutella xylostella

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Screening Bacillus thuringiensis (Bt) isolates or strains for toxicity has traditionally been performed with one bacterial isolate at time versus a specific insect. By testing of Bt strains in groups, we identified 28 of 147 Bt isolates as toxic to either diamondback moth, Plutella xylostella (L.),...

  6. Fulminant phlegmonitis of the esophagus, stomach, and duodenum due to Bacillus thuringiensis.

    PubMed

    Matsumoto, Hisatake; Ogura, Hiroshi; Seki, Masafumi; Ohnishi, Mitsuo; Shimazu, Takeshi

    2015-03-28

    We report a case of phlegmonitis of the esophagus, stomach, and duodenum in patient in an immunocompromised state. Culture of gastric juice and blood yielded Bacillus thuringiensis. This case showed that even low-virulence bacilli can cause lethal gastrointestinal phlegmonous gastritis in conditions of immunodeficiency. PMID:25834344

  7. Molecular markers to determine ecological fate of Bacillus thuringiensis subsp. kurstaki

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacillus thuringiensis (“Bt”) is a ubiquitous soil bacterium with entomopathogenic properties. One strain, Bt subsp. kurstaki (“Btk”), is highly toxic to lepidopteran larvae and used in many commercial products for biological pest control. We designed a set of DNA markers that successfully identifi...

  8. Recovery of Bacillus thuringiensis and insect toxic related strains from forest soil

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We attempted to recover Bacillus thuringiensis (Bt) from soil that had been sprayed two years prior with Bt for gypsy moth control. By amplifying the bacteria found in the soil on bacterial agar and feeding this diverse microbial population to tobacco hornworm larvae, 15 spore-forming bacteria from ...

  9. 40 CFR 180.1011 - Viable spores of the microorganism Bacillus thuringiensis Berliner; exemption from the...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Viable spores of the microorganism Bacillus thuringiensis Berliner; exemption from the requirement of a tolerance. 180.1011 Section 180.1011 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS TOLERANCES AND EXEMPTIONS FOR PESTICIDE CHEMICAL...

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  11. Binding of Bacillus thuringiensis toxin CrylAc to multiple sites of cadherin in pink bollworm

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Toxins from Bacillus thuringiensis (Bt) are widely used for pest control. In particular, Bt toxin Cry lAc produced by transgenic cotton kills some key lepidopteran pests. We found that CrylAc binds to recombinant peptides corresponding to extracellular regions of a cadherin protein (BtR) in a major ...

  12. ANALYSIS OF MIDGUT PROTEINASES FROM BACILLUS THURINGIENSIS-SUSCEPTIBLE AND -RESISTANT HELIOTHIS VIRESCENS (LEPIDOPTERA: NOCTUIDAE)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Insects with altered proteinases can avoid intoxication by Bacillus thuringiensis (Bt) toxins. Therefore, proteinase activities from gut extracts of Bt-susceptible (YDK) and -resistant (YHD2-B, CXC and KCBhyb) H. virescens strains were compared. The overall pH of gut extracts from YDK and CXC were...

  13. VERTEBRATE TOXICOLOGY OF THE SOLUBILIZED PROTEINS OF BACILLUS THURINGIENSIS SUBSP. ISRAELENSIS

    EPA Science Inventory

    This review summarizes the studies done with the mammalian toxic Bacillus thuringiensis subsp. israelensis (Bti) 28 kDa cytA protein. The data is relevant to hazard identification studies with bacterial pesticides. The data shows that cytA produces lethal physiological changes in...

  14. EFFECT OF REMOVAL OF THE CYTOLYTIC FACTOR OF 'BACILLUS THURINGIENSIS' SUBSP. 'ISRAELENSIS' ON MOSQUITO TOXICITY

    EPA Science Inventory

    Solubilized crystal protein of Bacillus thuringiensis subsp. israelensis was fractionated by affinity chromotography using a monoclonal antibody directed against the crystal's 28 kDa peptide. The 28 kDa peptide ws found to be relatively nontoxic to mosquito larvae although it doe...

  15. INTERACTIONS BETWEEN BACILLUS THURINGIENSIS SUBSP. ISRAELENSIS AND FATHEAD MINNOWS, PIMEPHALES PROMELAS RAFINESQUE, UNDER LABORATORY CONDITIONS

    EPA Science Inventory

    Interactions between Bacillus thuringiensis subsp. israelensis and fathead minnows, Pimephales promelas, were studied in laboratory exposures to two commercial formulations, Vectobac-G and Mosquito Attack. ortality among fatheads exposed to 2.0 x 10 6 to 6.5 x 10 6 CFU/ml with bo...

  16. INSECT RESISTANCE TO BACILLUS THURINGIENSIS: ALTERATIONS IN THE INDIANMEAL MOTH LARVAL GUT PROTEOME

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Insect resistance to the Cry toxins of Bacillus thuringiensis (Bt) has been examined previously using a number of traditional biochemical and molecular techniques. In this study, we utilized a proteomic approach involving two-dimensional differential gel electrophoresis, mass spectrometry and funct...

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  18. BACILLUS THURINGIENSIS VAR. KURSTAKI AFFECTS A BENEFICIAL INSECT, THE CINNABAR MOTH (LEPIDOPTERA: ARCTIIDAE)

    EPA Science Inventory

    The microbial insecticide bacillus thuringiensis Berliner var. kurstaki is used to control forest pests in regions where tansy ragwort, Senecio jacobaea L. occurs. iological control of this noxious weed may be compromised if the cinnabar moth, Tyria jacobaeae (L), is susceptible ...

  19. Insecticidal toxins from Bacillus thuringiensis subsp. kenyae: gene cloning and characterization and comparison with B. thuringiensis subsp. kurstaki CryIA(c) toxins.

    PubMed Central

    Von Tersch, M A; Robbins, H L; Jany, C S; Johnson, T B

    1991-01-01

    Genes encoding insecticidal crystal proteins were cloned from three strains of Bacillus thuringiensis subsp. kenyae and two strains of B. thuringiensis subsp. kurstaki. Characterization of the B. thuringiensis subsp. kenyae toxin genes showed that they are most closely related to cryIA(c) from B. thuringiensis subsp. kurstaki. The cloned genes were introduced into Bacillus host strains, and the spectra of insecticidal activities of each Cry protein were determined for six pest lepidopteran insects. CryIA(c) proteins from B. thuringiensis subsp. kenyae are as active as CryIA(c) proteins from B. thuringiensis subsp. kurstaki against Trichoplusia ni, Lymantria dispar, Heliothis zea, and H. virescens but are significantly less active against Plutella xylostella and, in some cases, Ostrinia nubilalis. The sequence of a cryIA(c) gene from B. thuringiensis subsp. kenyae was determined (GenBank M35524) and compared with that of cryIA(c) from B. thuringiensis subsp. kurstaki. The two genes are more than 99% identical and show seven amino acid differences among the predicted sequences of 1,177 amino acids. Images PMID:2014985

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

    PubMed Central

    2010-01-01

    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. PMID:20423490

  1. [Biology of two lysogenic phages from Bacillus thuringiensis MZ1].

    PubMed

    Liao, Wei; Sun, Fan; Song, Shao-yun; Shi, Wei; Pang, Yi

    2007-02-01

    A Bacillus thuringiensis (Bt) fermentative strain MZ1 (subsp. kurstaki) , from a company in Meixian County of Guangdong Province, produce toxins during sporulation and are extensively used in the field to control pest insects (Lepidoptera) in China. But some unknown or random factors that inhibited or stopped B. t growth in the fermentation can be regarded as reflecting the exist of lysogenic phage. Therefore, strain MZI and its lysogenic phages were studied in this paper. With indicator strain ZK1, two kind of phage plaques, one with about 3mm diameter and the other with about 1mm diameter, can be observed after strain MZ1 cultured in plates or flasks was induced by mitomycin C. Then, two lysogenic phages, namely MZTP01 and MZTP02, were isolated and characterized in biology. They belonged to family Siphoviridae, which had icosahedral heads (MZTP01 :82nm x 85nm; MZTP02: 75nm x 55nm) and long tails (MZTP01: 220nm x 18nm; MZTP02: 183nm x 12nm) without flexibility. Host range examination showed that six and seven (including indicator strain ZKl) out of 113 B. t strains saved in our laboratory were sensitive to MZTP01 and MZTP02, respectively. MZTP01 was more stable than MZTP02 against pH value, ultraviolet and heat treatment, but contrary against organic solvents. For MZTP01 and MZTP02, K values in the neutralization reactions were 45 and 326, respectively. Both phages had no relationship in their antigenicity. Burst size of phage MZTP02 was 175, two times more than that of MZTP01. Latent time of MZTP02 was 40min, one times shorter than that of MZTP01. It was suggested that both phages DNA be linear dsDNA through the typical absorption curves, reaction with diphenylamine, DNase sensitivity and acridine orange staining. And this was in good accord with the previous findings that all tailed phages being dsDNA moleculars. The genomic DNA and their restriction maps showed that both molecular weights should be between 9.4 - 23kb. Both phage genomic DNAs were digested by

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

    PubMed Central

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

    2015-01-01

    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. PMID:26411888

  3. Response of Heliothis virescens (Lepidoptera: Noctuidae) strains to Bacillus thuringiensis Cry1Ac incorporated into different insect artificial diets

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Susceptibility to the Cry1Ac toxin from Bacillus thuringiensis in Heliothis virescens is usually measured by performing bioassays under laboratory conditions. Currently there is great interest and research devoted to this insect because it is one of the main targets of B. thuringiensis-expressing tr...

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

    PubMed Central

    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

  5. Effects of four entomopathogenic nematode species on fitness costs of pink bollworm resistance to Bacillus thuringiensis toxin Cry1Ac

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Evolution of resistance by pests can reduce the efficacy of transgenic crops that produce insecticidal toxins from the bacterium Bacillus thuringiensis (Bt). However, fitness costs can slow the evolution of resistance. We tested whether four species of entomopathogenic nematodes (Steinernematidae ...

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

    PubMed Central

    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

  7. A novel metalloproteinase virulence factor is involved in Bacillus thuringiensis pathogenesis in nematodes and insects.

    PubMed

    Peng, Donghai; Lin, Jian; Huang, Qiong; Zheng, Wen; Liu, Guoqiang; Zheng, Jinshui; Zhu, Lei; Sun, Ming

    2016-03-01

    The Gram-positive soil bacterium Bacillus thuringiensis has been developed as the leading microbial insecticide for years. The pathogenesis of B. thuringiensis requires common extracellular factors that depend on the PlcR regulon, which regulates a large number of virulence factors; however, the precise role of many of these proteins is not known. In this study, we describe the complete lifecycle of a nematicidal B. thuringiensis strain in the free living nematode Caenorhabditis elegans using in vitro and in vivo molecular techniques to follow host and bacterial effectors during the infection process. We then focus on the metalloproteinase ColB, a collagenase, which was found highly important for destruction of the intestine thereby facilitates the adaptation and colonization of B. thuringiensis in C. elegans. In vivo green fluorescent protein (GFP) reporter-gene studies showed that ColB expression is highly induced and regulated by the global activator PlcR. Finally, we demonstrated that ColB also takes part in B. thuringiensis virulence in an insect model following injection and oral infection. Indeed, addition of purified ColB accelerates the action of Cry toxin proteins in insects, too. These results give novel insights into host adaptation for B. thuringiensis and other B. cereus group bacteria and highlight the role of collagenase metalloproteases to synergize infection process. PMID:26995589

  8. Activation of MAP kinase pathways in Galleria mellonella infected with Bacillus thuringiensis.

    PubMed

    Wojda, Iwona; Koperwas, Konrad; Jakubowicz, Teresa

    2014-01-01

    We followed changes in the level of phospho-MAP kinases in the greater wax moth Galleria mellonella after infection with Bacillus thuringiensis. We observed an enhanced level of phosphorylated p38 and JNK in fat bodies of the infected larvae. In hemocytes, injection of B. thuringiensis caused the highest increase in phospho-JNK, however, all pathways were activated after aseptic injection. We report that Galleria mellonella larvae exposed to heat shock before infection showed an enhanced level of phosphorylated JNK in fat body. This finding is relevant in the light of our previous reports, which submit evidence that pre-shocked animals are more resistant to infection. PMID:24455757

  9. Ubiquity of parasporin-1 producers in Bacillus thuringiensis natural populations of Japan

    NASA Astrophysics Data System (ADS)

    Uemori, Akiko; Maeda, Minoru; Yasutake, Koichi; Ohgushi, Akira; Kagoshima, Kumiko; Mizuki, Eiichi; Ohba, Michio

    2007-01-01

    Parasporin, a Bacillus thuringiensis parasporal protein, is unique in having a strong cytocidal activity preferential for human cancer cells. In this study, we characterized parasporin activities associated with three novel geographical isolates of B. thuringiensis. Parasporal inclusion proteins of the three isolates were highly toxic to human uterus cervix cancer cells (HeLa), but not to non-cancer uterine smooth muscle cells (UtSMC). Inclusions of the isolates lacked insect toxicity and hemolytic activity against sheep erythrocytes. Ouchterlony immunodiffusion tests revealed that the proteins of the three isolates are immunologically closely related to parasporin-1 (Cry31A), but dissimilar to the three other existing parasporin groups. Our results provide evidence that the parasporin-1-producing organism is a common member in B. thuringiensis populations occurring in natural environments of Japan.

  10. Microcalorimetric investigation on the growth model and the protein yield of Bacillus thuringiensis.

    PubMed

    Xiaoyan, Lin; Yi, Liu; Peng, Liu; Songsheng, Qu; Ziniu, Yu

    2004-06-30

    A novel microcalorimetric technique based on the bacterial heat output was applied to evaluate the special growth model, the protein expression and the generation time of Bacillus thuringiensis for the first time. The thermogenic curves of the aerobic metabolism of B. thuringiensis strains YBT-833, YBT-1520 and YBT-833-2-1 were determined by using an LKB-2277 BioActivity Monitor. The analysis of the thermogenic curves indicated both the mutant strain and the wild-type strains followed the same linear growth model during sporulation. The metabolism heat output revealed heat output was correlated to the yield of the insecticidal crystal proteins (ICPs) very well, the more protein product, and the less heat output. Based on the data acquired, we proposed that this method could be a useful tool in monitoring the fermentation of B. thuringiensis. PMID:15165757

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

    PubMed Central

    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

  12. Ecological consequences of ingestion of Bacillus cereus on Bacillus thuringiensis infections and on the gut flora of a lepidopteran host.

    PubMed

    Raymond, Ben; Lijek, Rebeccah S; Griffiths, Robert I; Bonsall, Michael B

    2008-09-01

    The Bacillus cereus group comprises a diverse array of non-pathogenic bacteria as well as pathogens such as Bacillus thuringiensis. Their spores are found together in soil and leaves and are therefore likely to commonly interact within hosts. Mixed infections of pathogenic B. thuringiensis and non-pathogenic strains have been little studied, despite their potential impact on biological control and the evolutionary ecology of virulence. Antibiotic secreting strains of B. cereus have been shown to be able to synergize B. thuringiensis (Bt) infections. We explored the ecology of these mixed infections more broadly in the diamondback moth (DBM). We tested whether antibiotic-expressing B. cereus can synergize Bt infections initiated with spores, investigated whether ingestion of antibiotic-expressing B. cereus had any consequences for the larval gut flora and whether synergistic interactions with B. cereus increase Bt reproduction. Ingestion of high-antibiotic secreting B. cereus synergized infections of B. thuringiensis in diamondback moth larvae, but at a lower level than previously reported. Coinfection also increased slightly the number of Bt spores found in cadavers. Culture independent analysis of gut homogenates indicated that ingestion of an antibiotic-expressing strain of B. cereus reduced the abundance of the gut flora and led to gut communities being dominated bacteria with DGGE profiles very similar to pure B. cereus cultures. Ingestion of B. cereus, regardless of genotype, reduced densities of an enteric isolate of Enterobacter sp. These findings support the hypothesis that antibiotic secretion in the gut synergizes B. thuringiensis infections by reducing the abundance of the commensal gut flora and facilitating invasion by bacteria in the B. cereus group. PMID:18533180

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

    PubMed

    Zhang, L; Qiu, S; Huang, T; Huang, Z; Xu, L; Wu, C; Gelbic, I; Guan, X

    2013-06-01

    To examine the effect of chemical additives on Bacillus thuringiensis (Berliner) against Plutella xylostella (L.), inorganic salts, nitrogenous compounds, protein solubilizing agents, and organic acids were selected and tested. The chosen materials are low in cost and environmentally safe. Results show that many inorganic salts can increase the activity of B. thuringiensis in a range of 1.31- to 3.08-fold. These include calcium acetate, calcium chloride, calcium hydroxide, calcium sulfate, calcium carbonate, sodium carbonate, sodium acetate, potassium hydroxide, potassium carbonate, potassium acetate, magnesium chloride, magnesium sulfate, and zinc sulfate. Nitrogenous compounds, including peptone, sodium nitrate, and ammonium nitrate, can enhance the activity of B. thuringiensis 1.62-, 1.32-, and 1.37-fold, respectively. Among the protein solubilizing agents, EDTA, urea, mercaptoethanol and dipotassium hydrogen phosphate increased the activity of B. thuringiensis 1.62- to 2.34-fold. Among the organic acids, maleic and citric acids boosted the activity 1.45- and 1.55-fold, respectively. Meanwhile, sodium benzoate and resorcinol led to 1.74- and 1.44-fold activity gains, respectively. Use of appropriate additives could provide great benefit not only in reducing the costs for field applications of biological insecticides but also by boosting the efficacy of B. thuringiensis. PMID:23865169

  14. Bacillus thuringiensis colonises plant roots in a phylogeny-dependent manner.

    PubMed

    Vidal-Quist, J Cristian; Rogers, Hilary J; Mahenthiralingam, Eshwar; Berry, Colin

    2013-12-01

    Although much is known about the pathology of Bacillus thuringiensis against invertebrates, current understanding of its natural ecology is limited. This study evaluated the biodiversity of B. thuringiensis in relation to its interaction with plants. Phylogenetic relationships between 44 reference and field-collected strains, determined using 16S rRNA and gyrB gene sequences, revealed a high degree of variability, similar to that found in databases. An Arabidopsis thaliana in vitro inoculation model was developed to screen the ability of B. thuringiensis to colonise roots. Significant colonisation differences up to 91-fold were observed between strains, and correlation between strain phylogeny and colonisation was found. The genetics and biochemistry of auxin production; presence of the gene encoding indole pyruvate decarboxylase; and the abilities of Bt strains to swarm, grow in rich/minimal media and affect root growth differed between the strains, but only auxin production correlated significantly with ability to colonise roots. Co-inoculation with Burkholderia phytofirmans PsJN or Pseudomonas fluorescens SBW25 produced no effect on B. thuringiensis colonisation levels, regardless of the co-inoculant. Similarly, root colonisation of A. thaliana mutants impaired in plant defences was not significantly higher compared with controls. This is the first systematic and phylogenetic evaluation of B. thuringiensis interaction with plants. PMID:23822207

  15. Multi-method approach for characterizing the interaction between Fusarium verticillioides and Bacillus thuringiensis subsp. Kurstaki.

    PubMed

    Rocha, Liliana O; Tralamazza, Sabina Moser; Reis, Gabriela M; Rabinovitch, Leon; Barbosa, Cynara B; Corrêa, Benedito

    2014-01-01

    Bacterial antagonists used as biocontrol agents represent part of an integrated management program to reduce pesticides in the environment. Bacillus thuringiensis is considered a good alternative as a biocontrol agent for suppressing plant pathogens such as Fusarium. In this study, we used microscopy, flow cytometry, indirect immunofluorescence, and high performance liquid chromatography to determine the interaction between B. thuringiensis subsp. kurstaki LFB-FIOCRUZ (CCGB) 257 and F. verticillioides MRC 826, an important plant pathogen frequently associated with maize. B. thuringiensis showed a strong in vitro suppressive effect on F. verticillioides growth and inhibited fumonisin production. Flow cytometry analysis was found to be adequate for characterizing the fungal cell oscillations and death during these interactions. Further studies of the antagonistic effect of this isolate against other fungi and in vivo testing are necessary to determine the efficacy of B. thuringiensis subsp. kurstaki in controlling plant pathogens. This is the first report on the use of flow cytometry for quantifying living and apoptotic F. verticillioides cells and the B. thuringiensis Cry 1Ab toxin. PMID:24739804

  16. Multi-Method Approach for Characterizing the Interaction between Fusarium verticillioides and Bacillus thuringiensis Subsp. Kurstaki

    PubMed Central

    Rocha, Liliana O.; Tralamazza, Sabina Moser.; Reis, Gabriela M.; Rabinovitch, Leon; Barbosa, Cynara B.; Corrêa, Benedito

    2014-01-01

    Bacterial antagonists used as biocontrol agents represent part of an integrated management program to reduce pesticides in the environment. Bacillus thuringiensis is considered a good alternative as a biocontrol agent for suppressing plant pathogens such as Fusarium. In this study, we used microscopy, flow cytometry, indirect immunofluorescence, and high performance liquid chromatography to determine the interaction between B. thuringiensis subsp. kurstaki LFB-FIOCRUZ (CCGB) 257 and F. verticillioides MRC 826, an important plant pathogen frequently associated with maize. B. thuringiensis showed a strong in vitro suppressive effect on F. verticillioides growth and inhibited fumonisin production. Flow cytometry analysis was found to be adequate for characterizing the fungal cell oscillations and death during these interactions. Further studies of the antagonistic effect of this isolate against other fungi and in vivo testing are necessary to determine the efficacy of B. thuringiensis subsp. kurstaki in controlling plant pathogens. This is the first report on the use of flow cytometry for quantifying living and apoptotic F. verticillioides cells and the B. thuringiensis Cry 1Ab toxin. PMID:24739804

  17. The Pathogenomic Sequence Analysis of B. cereus and B. Thuringiensis isolates closely related to Bacillus anthracis

    SciTech Connect

    Han, C S; Xie, G; Challacombe, J F; Altherr, M R; Bhotika, S S; Bruce, D; Campbell, C S; Campbell, M L; Chen, J; Chertkov, O; Cleland, C; Dimitrijevic-Bussod, M; Doggett, N A; Fawcett, J J; Glavina, T; Goodwin, L A; Hill, K K; Hitchcock, P; Jackson, P J; Keim, P; Kewalramani, A R; Longmire, J; Lucas, S; Malfatti, S; McMurry, K; Meincke, L J; Misra, M; Moseman, B L; Mundt, M; Munk, A C; Okinaka, R T; Parson-Quintana, B; Reilly, L P; Richardson, P; Robinson, D L; Rubin, E; Saunders, E; Tapia, R; Tesmer, J G; Thayer, N; Thompson, L S; Tice, H; Ticknor, L O; Wills, P L; Gilna, P; Brettin, T S

    2005-10-12

    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 B anthracis. 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.

  18. Bacillus thuringiensis var. tenebrionis control of synanthropic mites (Acari: Acaridida) under laboratory conditions.

    PubMed

    Erban, Tomas; Nesvorna, Marta; Erbanova, Michaela; Hubert, Jan

    2009-12-01

    Bacillus thuringiensis (Bt) toxins present a potential for control of pest mites. Information concerning the effect of Bt and its possible application to the biocontrol of synathropic mites is rare. The toxic effect of Bacillus thuringiensis var. tenebrionis producing Cry3A toxin was tested on the mites Acarus siro L., Tyrophagus putrescentiae (Schrank), Dermatophagoides farinae Hughes, and Lepidoglyphus destructor (Schrank) via feeding tests. Fifty mites were reared on Bt additive diets in concentrations that ranged from 0 to 100 mg g(-1) under optimal conditions for their development. After 21 days, the mites were counted and the final populations were analyzed using a polynomial regression model. The Bt diet suppressed population growth of the four mite species. The fitted doses of Bt for 50% suppression of population growth were diets ranging from 25 to 38 mg g(-1). There were no remarkable differences among species. Possible applications of Bt for the control of synanthropic mites are discussed. PMID:19381844

  19. Cloning, Sequencing, and Expression of the Chitinase Gene chiA74 from Bacillus thuringiensis

    PubMed Central

    Barboza-Corona, J. Eleazar; Nieto-Mazzocco, Elizabeth; Velázquez-Robledo, Rocio; Salcedo-Hernandez, Rubén; Bautista, Mayela; Jiménez, Beatriz; Ibarra, Jorge E.

    2003-01-01

    The endochitinase gene chiA74 from Bacillus thuringiensis serovar kenyae strain LBIT-82 was cloned in Escherichia coli DH5αF′. A sequence of 676 amino acids was deduced when the gene was completely sequenced. A molecular mass of 74 kDa was estimated for the preprotein, which includes a putative 4-kDa signal sequence located at the N terminus. The deduced amino acid sequence showed high degree of identity with other chitinases such as ChiB from Bacillus cereus (98%) and ChiA71 from Bacillus thuringiensis serovar pakistani (70%). Additionally, ChiA74 showed a modular structure comprised of three domains: a catalytic domain, a fibronectin-like domain, and a chitin-binding domain. All three domains showed conserved sequences when compared to other bacterial chitinase sequences. A ca. 70-kDa mature protein expressed by the cloned gene was detected in zymograms, comigrating with a chitinase produced by the LBIT-82 wild-type strain. ChiA74 is active within a wide pH range (4 to 9), although a bimodal activity was shown at pH 4.79 and 6.34. The optimal temperature was estimated at 57.2°C when tested at pH 6. The potential use of ChiA74 as a synergistic agent, along with the B. thuringiensis insecticidal Cry proteins, is discussed. PMID:12571025

  20. Production of a Thermostable and Alkaline Chitinase by Bacillus thuringiensis subsp. kurstaki Strain HBK-51

    PubMed Central

    Kuzu, Secil Berna; Güvenmez, Hatice Korkmaz; Denizci, Aziz Akin

    2012-01-01

    This paper reports the isolation and identification of chitinase-producing Bacillus from chitin-containing wastes, production of a thermostable and alkaline chitinasese, and enzyme characterization. Bacillus thuringiensis subsp. kurstaki HBK-51 was isolated from soil and was identified. Chitinase was obtained from supernatant of B. thuringiensis HBK-51 strain and showed its optimum activity at 110°C and at pH 9.0. Following 3 hours of incubation period, the enzyme showed a high level of activity at 110°C (96% remaining activity) and between pH 9.0 and 12.0 (98% remaining activity). Considering these characteristics, the enzyme was described as hyperthermophile-thermostable and highly alkaline. Two bands of the enzyme weighing 50 and 125 kDa were obtained following 12% SDS-PAGE analyses. Among the metal ions and chemicals used, Ni2+ (32%), K+ (44%), and Cu2+ (56%) increased the enzyme activity while EDTA (7%), SDS (7%), Hg2+ (11%), and ethyl-acetimidate (20%) decreased the activity of the enzyme. Bacillus thuringiensis subsp. kurstaki HBK-51 is an important strain which can be used in several biotechnological applications as a chitinase producer. PMID:23304523

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

    PubMed

    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. PMID:26689874

  2. Genomic characterization and comparison of seven Myoviridae bacteriophage infecting Bacillus thuringiensis.

    PubMed

    Sauder, Amber Brooke; Quinn, McKenzie Rea; Brouillette, Alexis; Caruso, Steven; Cresawn, Steven; Erill, Ivan; Lewis, Lynn; Loesser-Casey, Kathryn; Pate, Morgan; Scott, Crystal; Stockwell, Stephanie; Temple, Louise

    2016-02-01

    Bacillus thuringiensis Kurstaki, a bacterium that is a source of biopesticides and a safe simulant for pathogenic Bacillus species, was used to isolate seven unique bacteriophages. The phage genomes were sequenced and ranged in size from 158,100 to 163,019 bp encoding 290-299 genes, and the GC content of ~38% was similar to that of the host bacterium. All phages had terminal repeats 2-3 kb long. Three of the phages encoded tRNAs and three contained a self-splicing intron in the DNA polymerase gene. They were categorized as a single cluster (>60% nucleotide conservation) containing three subclusters (>80% nucleotide conservation), supported by genomic synteny and phylogenetic analysis. Considering the published genomes of phages that infect the genus Bacillus and noting the ability of many of the Bacillus cereus group phages to infect multiple species, a clustering system based on gene content is proposed. PMID:26773385

  3. 40 CFR 180.1108 - Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Delta endotoxin of Bacillus... From Tolerances § 180.1108 Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta...

  4. 40 CFR 180.1108 - Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Delta endotoxin of Bacillus... From Tolerances § 180.1108 Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta...

  5. 40 CFR 180.1108 - Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Delta endotoxin of Bacillus... From Tolerances § 180.1108 Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta...

  6. 40 CFR 180.1108 - Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Delta endotoxin of Bacillus... From Tolerances § 180.1108 Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. The delta...

  7. The occurrence, biodiversity and toxicity of Bacillus thuringiensis strains isolated from the insect pest Lymantria dispar (Poland).

    PubMed

    Guz, Katarzyna; Bugla-Płoskońska, Gabriela; Doroszkiewicz, Włodzimierz

    2009-01-01

    The aim of this investigation was to survey the occurrence, biodiversity, and toxicity of Bacillus thuringiensis strains originating from dead caterpillars of the forest pest, Lymantria dispar (Lepidoptera). Morphological, biochemical, and microscopic identification of isolates from the insects showed the presence of five different Bacillus species, including 2% of B. thuringiensis. Based on the biochemical profiles, the B. thuringiensis were determined to be B. thuringiensis finitimus-like and B. thuringiensis alesti-like bacilli. Both produced spherical inclusions composed of three or five protoxins. The molecular weights of these proteins varied from 20 to ca. 64 kDa. Mixtures of spores/inclusions of the B. thuringiensis were tested for their toxicity against larvae of Drosophila melanogaster. The mortality levels of the larvae caused by these spores and crystalline inclusions varied from 5 to 15%. The lethal doses (LD50) of these isolates against D. melanogaster were 8.8 x 10(12) spores/ml for B. thuringiensis finitimus and 1.3 10(18) spores/ml for B. thuringiensis alesti. PMID:19824400

  8. Molecular cloning of the 130-kilodalton mosquitocidal delta-endotoxin gene of Bacillus thuringiensis subsp. israelensis in Bacillus sphaericus.

    PubMed Central

    Trisrisook, M; Pantuwatana, S; Bhumiratana, A; Panbangred, W

    1990-01-01

    A 3.7-kilobase (kb) XbaI fragment harboring the cryIVB gene (L. Thorne, F. Garduno, T. Thompson, D. Decker, M. A. Zounes, M. Wild, A. M. Walfield, and T. J. Pollock, J. Bacteriol. 166:801-811, 1986) which encoded a 130-kilodalton (kDa) mosquitocidal toxin from a 110-kb plasmid of Bacillus thuringiensis subsp. israelensis 4Q2-72 was cloned into pUC12 and transformed into Escherichia coli. The clone with a recombinant plasmid (designated pBT8) was toxic to Aedes aegypti larvae. The fragment (3.7 kb) was ligated into pBC16 (tetracycline resistant [Tcr]) and transformed by the method of protoplast transformation into Bacillus sphaericus 1593 and 2362, which were highly toxic to Anopheles and Culex mosquito larvae but less toxic to Aedes larvae. After cell regeneration on regeneration medium, the Tcr plasmids from transformants (pBTC1) of both strains of B. sphaericus were prepared and analyzed. The 3.7-kb XbaI fragment from the B. thuringiensis subsp. israelensis plasmid was shown to be present by agarose gel electrophoresis and Southern blot hybridization. In addition, B. sphaericus transformants produced a 130-kDa mosquitocidal toxin which was detected by Western (immuno-) blot analysis with antibody prepared against B. thuringiensis subsp. israelensis 130-kDa mosquitocidal toxin. The 50% lethal concentrations of the transformants of strains 1593 and 2362 against A. aegypti larvae were 2.7 X 10(2) and 5.7 X 10(2) cells per ml, respectively. This level of toxicity was comparable to the 50% lethal concentration of B. thuringiensis subsp. israelensis but much higher than that of B. sphaericus 1593 and 2362 (4.7 X 10(4) cells per ml) against A. aegypti larvae.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:2200339

  9. 40 CFR 174.510 - Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry1Ac protein... Cry1Ac protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ac protein in all plants are exempt from the requirement of a tolerance when used as...

  10. 40 CFR 174.518 - Bacillus thuringiensis Cry3Bb1 protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Cry3Bb1 protein... Cry3Bb1 protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry3Bb1 protein in corn are exempt from the requirement of a tolerance when used as...

  11. 40 CFR 174.510 - Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry1Ac protein... Cry1Ac protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ac protein in all plants are exempt from the requirement of a tolerance when used as...

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

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry3Bb1 protein... Cry3Bb1 protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry3Bb1 protein in corn are exempt from the requirement of a tolerance when used as...

  13. 40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as...

  14. 40 CFR 174.518 - Bacillus thuringiensis Cry3Bb1 protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry3Bb1 protein... Cry3Bb1 protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry3Bb1 protein in corn are exempt from the requirement of a tolerance when used as...

  15. 40 CFR 174.510 - Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry1Ac protein... Cry1Ac protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ac protein in all plants are exempt from the requirement of a tolerance when used as...

  16. 40 CFR 174.518 - Bacillus thuringiensis Cry3Bb1 protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry3Bb1 protein... Cry3Bb1 protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry3Bb1 protein in corn are exempt from the requirement of a tolerance when used as...

  17. 40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as...

  18. 40 CFR 174.510 - Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Cry1Ac protein... Cry1Ac protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ac protein in all plants are exempt from the requirement of a tolerance when used as...

  19. 40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as...

  20. 40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as...

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

    PubMed

    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. PMID:25686722

  2. Binding of Bacillus thuringiensis proteins to a laboratory-selected line of Heliothis virescens.

    PubMed

    MacIntosh, S C; Stone, T B; Jokerst, R S; Fuchs, R L

    1991-10-15

    A laboratory-selected colony of Heliothis virescens displaying a 20- to 70-fold level of resistance to Bacillus thuringiensis proteins was evaluated to identify mechanism(s) of resistance. Brush-border membrane vesicles were isolated from larval midgut epithelium from the susceptible and resistant strains of H. virescens. Two B. thuringiensis proteins, CryIA(b) and CryIA(c), were iodinated and shown to specifically bind to brush-border membrane vesicles of both insect strains. Multiple changes in the receptor-binding parameters were seen in the resistant strain as compared with the susceptible strain. A 2- to 4-fold reduction in binding affinity was accompanied by a 4- to 6-fold increase in binding-site concentration for both proteins. Although these two B. thuringiensis proteins competed for the same high-affinity binding site, competition experiments revealed different receptor specificity toward these proteins in the resistant H. virescens line. The H. virescens strains were not sensitive to a coleopteran-active protein, CryIIIA, nor did these proteins compete with the CryIA proteins for binding. Complexity of the mechanism of resistance is consistent with the complex mode of action of B. thuringiensis proteins. PMID:1924353

  3. Bacillus thuringiensis Metalloproteinase Bmp1 Functions as a Nematicidal Virulence Factor

    PubMed Central

    Luo, Xiaoxia; Chen, Ling; Huang, Qiong; Zheng, Jinshui; Zhou, Wei; Peng, Donghai; Ruan, Lifang

    2013-01-01

    Some Bacillus thuringiensis strains have high toxicity to nematodes. Nematicidal activity has been found in several families of crystal proteins, such as Cry5, Cry6, and Cry55. The B. thuringiensis strain YBT-1518 has three cry genes that have high nematicidal activity. The whole genome sequence of this strain contains multiple potential virulence factors. To evaluate the pathogenic potential of virulence factors, we focused on a metalloproteinase called Bmp1. It encompasses a consecutive N-terminal signal peptide, an FTP superfamily domain, an M4 neutral protease GluZincin superfamily, two Big-3 superfamily motifs, and a Gram-positive anchor superfamily motif as a C-terminal domain. Here, we showed that purified Bmp1 protein showed metalloproteinase activity and toxicity against Caenorhabditis elegans (the 50% lethal concentration is 610 ± 9.37 μg/ml). In addition, mixing Cry5Ba with Bmp1 protein enhanced the toxicity 7.9-fold (the expected toxicity of the two proteins calculated from their separate toxicities) against C. elegans. Confocal microscopic observation revealed that Bmp1 protein was detected from around the mouth and esophagus to the intestine. Striking microscopic images revealed that Bmp1 degrades intestine tissues, and the Cry5Ba causes intestinal shrinkage from the body wall. Thus, the B. thuringiensis Bmp1 metalloproteinase is a nematicidal virulence factor. These findings give a new insight into the relationship between B. thuringiensis and its host nematodes. PMID:23124228

  4. Activity of bacteriocins synthesized by Bacillus thuringiensis against Staphylococcus aureus isolates associated to bovine mastitis.

    PubMed

    Barboza-Corona, José Eleazar; de la Fuente-Salcido, Norma; Alva-Murillo, Nayeli; Ochoa-Zarzosa, Alejandra; López-Meza, Joel E

    2009-07-01

    Antimicrobial therapy is a useful tool to control bovine mastitis caused by Staphylococcus aureus, as consequence an increase in staphylococci resistant cases has been registered. Alternative strategies are desirable and bacteriocins represent attractive control agents to prevent bovine mastitis. The aim of this work was to evaluate the activity of five bacteriocins synthesized by Bacillus thuringiensis against S. aureus isolates associated to bovine mastitis. Fifty S. aureus isolates were recovered from milk composite samples of 26 Holstein lactating cows from one herd during September 2007 to February 2008 in México and susceptibility of those isolates to 12 antibiotics and 5 bacteriocins from B. thuringiensis was evaluated. S. aureus isolates were mainly resistant to penicillin (92%), dicloxacillin (86%), ampicillin (74%) and erythromycin (74%); whereas susceptibility to gentamicin, trimethoprim and tetracycline was detected at, respectively, 92%, 88%, and 72%. All S. aureus isolates showed susceptibility to the five bacteriocins synthesized by B. thuringiensis, mainly to morricin 269 and kurstacin 287 followed by kenyacin 404, entomocin 420 and tolworthcin 524. Our results showed that S. aureus isolates had differences in the antimicrobial resistance patterns and were susceptible to bacteriocins produced by B. thuringiensis, which could be useful as an alternative method to control bovine mastitis. PMID:19359107

  5. New variants of lepidoptericidal toxin genes encoding Bacillus thuringiensis Vip3Aa proteins.

    PubMed

    Sauka, Diego H; Rodriguez, Sonia E; Benintende, Graciela B

    2012-01-01

    Bacillus thuringiensis is an entomopathogenic bacterium characterized by producing parasporal proteinaceous insecticidal crystal inclusions during sporulation. Many strains are capable of also expressing other insecticidal proteins called Vip during the vegetative growing phase. Particularly, Vip3A proteins have activity against certain Lepidoptera species through a unique mechanism of action which emphasized their possible use in resistance management strategies against resistant pests. The aim of the work was to develop a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method that can distinguish between vip3A genes from B. thuringiensis strains. In addition, 4 novel vip3Aa genes were cloned and sequenced. The method was originally based on amplification of a single PCR amplicon and the use of 2 restriction enzymes with recognition sites that facilitate simultaneous detection. Subsequently, a third restriction enzyme was used to distinguish between vip3A variants. Thirteen vip3Aa genes were identified in strains belonging to 10 different B. thuringiensis serovars. Three intra-subclass variants of vip3Aa genes could be differentiated. The presented method can serve as an invaluable tool for the investigation of known and novel vip3A genes in B. thuringiensis strains. To the best of our knowledge, this is the first report where variants of a same subclass of insecticidal genes could be distinguished following PCR-RFLP. PMID:23307196

  6. Transduction of certain genes by an autonomously replicating Bacillus thuringiensis phage.

    PubMed Central

    Walter, T M; Aronson, A I

    1991-01-01

    A derivative of Bacillus thuringiensis subsp. kurstaki HD1 (HD1-9) released transducing phage (TP21) from late exponential cultures. Three of seven markers tested were transduced into Bacillus cereus, but only two of these (cysC and trpB/F) were transduced at a frequency of more than 100 times the reversion rates. A limited transduction capacity was given further support in that few chromosomal markers were carried in the HD1-9 lysate, as demonstrated by Southern hybridization. Restriction fragments from the phage DNA and from total B. thuringiensis DNA hybridized to an insertion sequence (IS231-like) probe, which may provide a region of homology for transduction. All of the B. cereus transductants contained the phage as a 44-kb plasmid, and each could transduce both the cys and trp genes to other B. cereus auxotrophs, albeit at lower frequencies than those for the B. thuringiensis transducing phage. In some cases, especially for cys, the transduced gene was integrated into the chromosome of the recipient, whereas the trp gene in many cases appeared to be lost with curing of the 44-kb plasmid. In addition, some B. cereus transductants lost prototrophy but retained a 44-kb plasmid, consistent with the presence of TP21 helper phage. These phage may mediate the subsequent transduction from B. cereus phototrophs. TP21 replicates as a plasmid and, at least under the conditions studied, selectively transfers markers to B. cereus. Images PMID:2059027

  7. Genes and environment interact to determine the fitness costs of resistance to Bacillus thuringiensis

    PubMed Central

    Raymond, Ben; Sayyed, Ali H; Wright, Denis J

    2005-01-01

    Genes which provide resistance to novel challenges such as pesticides, toxins or pathogens often impose fitness costs on individuals with a resistant phenotype. Studies of resistance to Bacillus thuringiensis and its insecticidal Cry toxins indicate that fitness costs may be variable and cryptic. Using two field populations (Karak and Serd4) of the diamondback moth, Plutella xylostella, we tested the hypothesis that the costs associated with resistance to the B. thuringiensis toxin Cry1Ac would be evident when insects were grown under poor environmental conditions, namely limited or poor quality resources. On a poor quality resource, a cultivar of Brassica oleracea var. capitata with varietal resistance to P. xylostella, only one resistant population, Karak, showed reduced fitness. Conversely, when we limited a high quality resource, Brassica pekinensis, by imposing larval competition, only resistant Serd4 insects had reduced survival at high larval densities. Furthermore, Cry1Ac resistance in Serd4 insects declined when reared at high larval densities while resistance at low densities fluctuated but did not decline significantly. These results confirm the hypothesis that resistance costs can appear under stressful conditions and demonstrate that the fitness cost of resistance to Bacillus thuringiensis can depend on the particular interaction between genes and the environment. PMID:16011928

  8. A purification and some properties of an insecticidal exotoxin from Bacillus thuringiensis Berliner

    PubMed Central

    Bond, R. P. M.; Boyce, C. B. C.; French, S. J.

    1969-01-01

    An insecticidal exotoxin from Bacillus thuringiensis var. thuringiensis (Berliner) has been purified. The efficiency of each stage of the purification has been ascertained and the yield of toxic material estimated by means of a quantitative bioassay. It is shown that the exotoxin is an adenine derivative substituted at position 9 and having a molecular weight of approximately 825. It can be dephosphorylated enzymically or chemically under conditions that define the exotoxin as a phosphomonoester. This results in loss of toxicity, both to insects and to mice. Spectroscopic and kinetic data are presented which suggest that a β-ribofuranosyl moiety may be attached to the adenine. Glucose and allomucic acid have been positively identified as hydrolysis fragments from the exotoxin. These results are discussed and compared with the results of others on similar (or possibly identical) compounds. PMID:5820635

  9. Neural networks applied to the prediction of fed-batch fermentation kinetics of Bacillus thuringiensis.

    PubMed

    Valdez-Castro, L; Baruch, I; Barrera-Cortés, J

    2003-01-01

    This paper proposes using a new recurrent neural network model (RNNM) to predict and control fed batch fermentations of Bacillus thuringiensis. The control variables are the limiting substrate and the feeding conditions. The multi-input multi-output RNNM proposed has twelve inputs, seven outputs, nineteen neurons in the hidden layer, and global and local feedbacks. The weight update learning algorithm designed is a version of the well known backpropagation through time algorithm directed to the RNNM learning. The error approximation for the last epoch of learning is 2% and the total learning time is 51 epochs, where the size of an epoch is 162 iterations. The RNNM generalization was carried out reproducing a B. thuringiensis fermentation not included in the learning process. It attains an error approximation of 1.8%. PMID:14505001

  10. Crystallization of parasporin-2, a Bacillus thuringiensis crystal protein with selective cytocidal activity against human cells.

    PubMed

    Akiba, Toshihiko; Abe, Yuichi; Kitada, Sakae; Kusaka, Yoshitomo; Ito, Akio; Ichimatsu, Tokio; Katayama, Hideki; Akao, Tetsuyuki; Higuchi, Kazuhiko; Mizuki, Eiichi; Ohba, Michio; Kanai, Ryuta; Harata, Kazuaki

    2004-12-01

    Bacillus thuringiensis is a valuable source of protein toxins that are specifically effective against certain insects and worms but harmless to mammals. In contrast, a protein toxin obtained from B. thuringiensis strain A1547, designated parasporin-2, is not insecticidal but has a strong cytocidal activity against human cells with markedly divergent target specificity. The 37 kDa inactive protein is proteolytically activated to a 30 kDa active form. The active form of the recombinant protein toxin was crystallized in the presence of ethylene glycol and polyethylene glycol 8000 at neutral pH. The crystals belong to the hexagonal space group P6(1) or P6(5), with unit-cell parameters a = b = 134.37, c = 121.24 A. Diffraction data from a native crystal were collected to 2.75 A resolution using a synchrotron-radiation source. PMID:15583389

  11. An ABC transporter from Bacillus thuringiensis is essential for beta-exotoxin I production.

    PubMed

    Espinasse, Sylvain; Gohar, Michel; Lereclus, Didier; Sanchis, Vincent

    2002-11-01

    beta-Exotoxin I is a nonspecific insecticidal metabolite secreted by some Bacillus thuringiensis strains. Several studies of B. thuringiensis strains that have lost the capacity to produce beta-exotoxin I have suggested that there is a strong correlation between high levels of beta-exotoxin I production and the ability to synthesize crystal proteins. In this study, we showed that a mutant strain, B. thuringiensis 407-1(Cry(-))(Pig(+)), with no crystal gene, produced considerable amounts of beta-exotoxin I together with a soluble brown melanin pigment. Therefore, beta-exotoxin I production can take place after a strain has lost the plasmids bearing the cry genes, which suggests that these curable plasmids probably contain determinants involved in the regulation of beta-exotoxin I production. Using a mini-Tn10 transposon, we constructed a library of strain 407-1(Cry(-))(Pig(+)) mutants. We screened for nonpigmented mutants with impaired beta-exotoxin I production and identified a genetic locus harboring two genes (berA and berB) essential for beta-exotoxin I production. The deduced amino acid sequence of the berA gene displayed significant similarity to the ATP-binding domains of the DRI (drug resistance and immunity) family of ATP-binding cassette (ABC) proteins involved in drug resistance and immunity to bacteriocins and lantibiotics. The berB gene encodes a protein with six putative transmembrane helices, which probably constitutes the integral membrane component of the transporter. The demonstration that berAB is required for beta-exotoxin I production and/or resistance in B. thuringiensis adds an adenine nucleotide analog to the wide range of substrates of the superfamily of ABC proteins. We suggest that berAB confers beta-exotoxin I immunity in B. thuringiensis, through active efflux of the molecule. PMID:12374817

  12. Constitutive activation of the midgut response to Bacillus thuringiensis in Bt-resistant Spodoptera exigua.

    PubMed

    Hernández-Martínez, Patricia; Navarro-Cerrillo, Gloria; Caccia, Silvia; de Maagd, Ruud A; Moar, William J; Ferré, Juan; Escriche, Baltasar; Herrero, Salvador

    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 involved in conferring resistance is of paramount importance. A colony of Spodoptera exigua (Lepidoptera: Noctuidae) was selected for 15 years in the laboratory for resistance to Xentari™, a B. thuringiensis-based insecticide, reaching a final resistance level of greater than 1,000-fold. Around 600 midgut ESTs were analyzed by DNA-macroarray in order to find differences in midgut gene expression between susceptible and resistant insects. Among the differentially expressed genes, repat and arylphorin were identified and their increased expression was correlated with B. thuringiensis resistance. We also found overlap among genes that were constitutively over-expressed in resistant insects with genes that were up-regulated in susceptible insects after exposure to Xentari™, suggesting a permanent activation of the response to Xentari™ in resistant insects. Increased aminopeptidase activity in the lumen of resistant insects in the absence of exposure to Xentari™ corroborated the hypothesis of permanent activation of response genes. Increase in midgut proliferation has been proposed as a mechanism of response to pathogens in the adult from several insect species. Analysis of S. exigua larvae revealed that midgut proliferation was neither increased in resistant insects nor induced by exposure of susceptible larvae to Xentari™, suggesting that mechanisms other than midgut proliferation are involved in the response to B. thuringiensis by S. exigua larvae. PMID:20862260

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

    PubMed Central

    Hernández-Martínez, Patricia; Navarro-Cerrillo, Gloria; Caccia, Silvia; de Maagd, Ruud A.; Moar, William J.; Ferré, Juan; Escriche, Baltasar; Herrero, Salvador

    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 involved in conferring resistance is of paramount importance. A colony of Spodoptera exigua (Lepidoptera: Noctuidae) was selected for 15 years in the laboratory for resistance to Xentari™, a B. thuringiensis-based insecticide, reaching a final resistance level of greater than 1,000-fold. Around 600 midgut ESTs were analyzed by DNA-macroarray in order to find differences in midgut gene expression between susceptible and resistant insects. Among the differentially expressed genes, repat and arylphorin were identified and their increased expression was correlated with B. thuringiensis resistance. We also found overlap among genes that were constitutively over-expressed in resistant insects with genes that were up-regulated in susceptible insects after exposure to Xentari™, suggesting a permanent activation of the response to Xentari™ in resistant insects. Increased aminopeptidase activity in the lumen of resistant insects in the absence of exposure to Xentari™ corroborated the hypothesis of permanent activation of response genes. Increase in midgut proliferation has been proposed as a mechanism of response to pathogens in the adult from several insect species. Analysis of S. exigua larvae revealed that midgut proliferation was neither increased in resistant insects nor induced by exposure of susceptible larvae to Xentari™, suggesting that mechanisms other than midgut proliferation are involved in the response to B. thuringiensis by S. exigua larvae. PMID:20862260

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

    PubMed Central

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

    PubMed

    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 σ(H) 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 σ(G). 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

  16. Resistance of Trichoplusia ni populations selected by Bacillus thuringiensis sprays to cotton plants expressing pyramided Bacillus thuringiensis toxins Cry1Ac and Cry2Ab.

    PubMed

    Kain, Wendy; Song, Xiaozhao; Janmaat, Alida F; Zhao, Jian-Zhou; Myers, Judith; Shelton, Anthony M; Wang, Ping

    2015-03-01

    Two populations of Trichoplusia ni that had developed resistance to Bacillus thuringiensis sprays (Bt sprays) in commercial greenhouse vegetable production were tested for resistance to Bt cotton (BollGard II) plants expressing pyramided Cry1Ac and Cry2Ab. The T. ni colonies resistant to Bacillus thuringiensis serovar kurstaki formulations were not only resistant to the Bt toxin Cry1Ac, as previously reported, but also had a high frequency of Cry2Ab-resistant alleles, exhibiting ca. 20% survival on BollGard II foliage. BollGard II-resistant T. ni strains were established by selection with BollGard II foliage to further remove Cry2Ab-sensitive alleles in the T. ni populations. The BollGard II-resistant strains showed incomplete resistance to BollGard II, with adjusted survival values of 0.50 to 0.78 after 7 days. The resistance to the dual-toxin cotton plants was conferred by two genetically independent resistance mechanisms: one to Cry1Ac and one to Cry2Ab. The 50% lethal concentration of Cry2Ab for the resistant strain was at least 1,467-fold that for the susceptible T. ni strain. The resistance to Cry2Ab in resistant T. ni was an autosomally inherited, incompletely recessive monogenic trait. Results from this study indicate that insect populations under selection by Bt sprays in agriculture can be resistant to multiple Bt toxins and may potentially confer resistance to multitoxin Bt crops. PMID:25480752

  17. Resistance of Trichoplusia ni Populations Selected by Bacillus thuringiensis Sprays to Cotton Plants Expressing Pyramided Bacillus thuringiensis Toxins Cry1Ac and Cry2Ab

    PubMed Central

    Kain, Wendy; Song, Xiaozhao; Janmaat, Alida F.; Zhao, Jian-Zhou; Myers, Judith; Shelton, Anthony M.

    2014-01-01

    Two populations of Trichoplusia ni that had developed resistance to Bacillus thuringiensis sprays (Bt sprays) in commercial greenhouse vegetable production were tested for resistance to Bt cotton (BollGard II) plants expressing pyramided Cry1Ac and Cry2Ab. The T. ni colonies resistant to Bacillus thuringiensis serovar kurstaki formulations were not only resistant to the Bt toxin Cry1Ac, as previously reported, but also had a high frequency of Cry2Ab-resistant alleles, exhibiting ca. 20% survival on BollGard II foliage. BollGard II-resistant T. ni strains were established by selection with BollGard II foliage to further remove Cry2Ab-sensitive alleles in the T. ni populations. The BollGard II-resistant strains showed incomplete resistance to BollGard II, with adjusted survival values of 0.50 to 0.78 after 7 days. The resistance to the dual-toxin cotton plants was conferred by two genetically independent resistance mechanisms: one to Cry1Ac and one to Cry2Ab. The 50% lethal concentration of Cry2Ab for the resistant strain was at least 1,467-fold that for the susceptible T. ni strain. The resistance to Cry2Ab in resistant T. ni was an autosomally inherited, incompletely recessive monogenic trait. Results from this study indicate that insect populations under selection by Bt sprays in agriculture can be resistant to multiple Bt toxins and may potentially confer resistance to multitoxin Bt crops. PMID:25480752

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

    PubMed Central

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

    2013-01-01

    Objective To investigate the toxicity of indigenous Bacillus thuringiensis (B. thuringiensis)isolates from Malang City for controlling Aedes aegypti (Ae. aegypti) larvae. Methods 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%. Results 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. Conclusions 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=2.3×108 cells/mL). PMID:23593589

  19. Acute toxicity and cytotoxicity of Bacillus thuringiensis and Bacillus sphaericus strains on fish and mouse bone marrow.

    PubMed

    Grisolia, Cesar Koppe; Oliveira-Filho, Eduardo Cyrino; Ramos, Felipe Rosa; Lopes, Madaí Cruz; Muniz, Daphne Heloisa Freitas; Monnerat, Rose Gomes

    2009-01-01

    The insecticidal properties of delta-endotoxins from Bacillus thuringiensis (Bt) serotypes kurstaki and israelensis and crystal proteins of Bacillus sphaericus (Bs) serotype H5 have been used in insect control for decades. The availability of microbial toxins in biopesticides as well as in plants with incorporated protection has been increasing the concerns about biosafety. Acute toxicity to Danio rerio and cytotoxicity on mouse bone marrow cells and peripheral erythrocytes of Oreochromis niloticus were tested with Bt israelensis, Bt kurstaki and Bs H5 strains. The concentration and dose tested were 10(6) and 10(8) spores/ml, respectively. Neither lethality nor effects on mouse bone marrow were promoted by any strain. In necrosis-apoptosis study on peripheral erythrocytes of O. niloticus an increased frequency of necrotic cells caused by exposure to strains of B. thuringiensis was found. Exposure to B. sphaericus did not show cytotoxic effects in either tested system. None of the strains studied induced apoptosis in contrast with the chemical controls. PMID:18670879

  20. Toxicity of parasporal crystals of Bacillus thuringiensis to the Indian meal moth, Plodia interpunctella.

    PubMed Central

    Schesser, J H; Bulla, L A

    1979-01-01

    Toxicity of Bacillus thuringiensis parasporal crystals to the Indian meal moth, Plodia interpunctella, is described. The numbers of insects killed were in relation to crystal dry weight. Mortality was determined by comparing adult emergence in diets treated with crystals to emergence in untreated diets. There was only a 30% survival at an application of 0.414 microgram/cm2, and the mean 50% lethal concentration value was found to be 0.299 microgram/cm2. The use of emergence data has provided a reliable and reproducible bioassay for comparing relative toxicities of crystals, spores, and other cellular components to this economically important insect. Images PMID:485134

  1. Relationship between poly-beta-hydroxybutyrate production and delta-endotoxin for Bacillus thuringiensis var. kurstaki.

    PubMed

    Navarro, A Karin; Farrera, Reynold R; López, Ruth; Pérez-Guevara, Fermín

    2006-05-01

    A linear relationship between total solid concentration (TSC), delta-endotoxin production [Cry = 0.2795(TSC)-0.2472, R2 = 0.8644] and poly-beta-hydroxybutyrate (PHB) accumulation [PHB = 0.1327(TSC) + 0.3974, R2 = 0.9877] in Bacillus thuringiensis var. kurstaki HD-73 was observed. A similar correlation between delta-endotoxin and PHB accumulation [Cry = 2.1573(PHB)-1.1248, R2 = 0.9181] was found. A minimum PHB accumulation of 0.52 mg l(-1) was required before the onset of delta-endotoxin production. PMID:16642302

  2. Influence of Lysogeny of Tectiviruses GIL01 and GIL16 on Bacillus thuringiensis Growth, Biofilm Formation, and Swarming Motility

    PubMed Central

    Gillis, Annika

    2014-01-01

    Bacillus thuringiensis is an entomopathogenic bacterium that has been used as an efficient biopesticide worldwide. Despite the fact that this bacterium is usually described as an insect pathogen, its life cycle in the environment is still largely unknown. B. thuringiensis belongs to the Bacillus cereus group of bacteria, which has been associated with many mobile genetic elements, such as species-specific temperate or virulent bacteriophages (phages). Temperate (lysogenic) phages are able to establish a long-term relationship with their host, providing, in some cases, novel ecological traits to the bacterial lysogens. Therefore, this work focuses on evaluating the potential influence of temperate tectiviruses GIL01 and GIL16 on the development of different life traits of B. thuringiensis. For this purpose, a B. thuringiensis serovar israelensis plasmid-cured (nonlysogenic) strain was used to establish bacterial lysogens for phages GIL01 and GIL16, and, subsequently, the following life traits were compared among the strains: kinetics of growth, metabolic profiles, antibiotics susceptibility, biofilm formation, swarming motility, and sporulation. The results revealed that GIL01 and GIL16 lysogeny has a significant influence on the bacterial growth, sporulation rate, biofilm formation, and swarming motility of B. thuringiensis. No changes in metabolic profiles or antibiotic susceptibilities were detected. These findings provide evidence that tectiviruses have a putative role in the B. thuringiensis life cycle as adapters of life traits with ecological advantages. PMID:25261525

  3. Influence of lysogeny of Tectiviruses GIL01 and GIL16 on Bacillus thuringiensis growth, biofilm formation, and swarming motility.

    PubMed

    Gillis, Annika; Mahillon, Jacques

    2014-12-01

    Bacillus thuringiensis is an entomopathogenic bacterium that has been used as an efficient biopesticide worldwide. Despite the fact that this bacterium is usually described as an insect pathogen, its life cycle in the environment is still largely unknown. B. thuringiensis belongs to the Bacillus cereus group of bacteria, which has been associated with many mobile genetic elements, such as species-specific temperate or virulent bacteriophages (phages). Temperate (lysogenic) phages are able to establish a long-term relationship with their host, providing, in some cases, novel ecological traits to the bacterial lysogens. Therefore, this work focuses on evaluating the potential influence of temperate tectiviruses GIL01 and GIL16 on the development of different life traits of B. thuringiensis. For this purpose, a B. thuringiensis serovar israelensis plasmid-cured (nonlysogenic) strain was used to establish bacterial lysogens for phages GIL01 and GIL16, and, subsequently, the following life traits were compared among the strains: kinetics of growth, metabolic profiles, antibiotics susceptibility, biofilm formation, swarming motility, and sporulation. The results revealed that GIL01 and GIL16 lysogeny has a significant influence on the bacterial growth, sporulation rate, biofilm formation, and swarming motility of B. thuringiensis. No changes in metabolic profiles or antibiotic susceptibilities were detected. These findings provide evidence that tectiviruses have a putative role in the B. thuringiensis life cycle as adapters of life traits with ecological advantages. PMID:25261525

  4. Chronic exposure of the European corn borer (Lepidoptera: Crambidae) to Cry1Ab Bacillus thuringiensis toxin.

    PubMed

    Chaufaux, J; Seguin, M; Swanson, J J; Bourguet, D; Siegfried, B D

    2001-12-01

    Transgenic corn expressing the insecticidal toxin from Bacillus thuringiensis Berliner is gaining support as an effective control technology for use against lepidopteran pests, particularly European corn borer, Ostrinia nubilalis Hübner (Lepidoptera: Crambidae). However, there is concern that widespread adoption of transgenic plants will rapidly lead to B. thuringiensis toxin resistance. Thus, long-term selection of O. nubilalis populations with the Cry1Ab B. thuringiensis toxin has been undertaken in several laboratories in the United States and in Europe. We present results from two independent selection experiments performed in laboratories at the University of Nebraska and at the Institut National de la Recherche Agronomique in France. Although the protocols and methods used by the two laboratories were different, the results were comparable. The highest level of resistance occurred at generation 7 (14-fold), generation 9 (13-fold), and generation 9 (32-fold) for three different strains. For each strain, the level of resistance fluctuated from generation to generation, although there were consistently significant decreases in toxin susceptibility across generations for all selected strains. These results suggest that low levels of resistance are common among widely distributed O. nubilalis populations. PMID:11777065

  5. Construction of Novel Bacillus thuringiensis Strains with Different Insecticidal Activities by Transduction and Transformation.

    PubMed

    Lecadet, M M; Chaufaux, J; Ribier, J; Lereclus, D

    1992-03-01

    The shuttle vector pHT3101 and its derivative pHT408, bearing a copy of a cryIA(a) delta-endotoxin gene, were transferred into several Bacillus thuringiensis subspecies through phage CP-54Ber-mediated transduction, with frequencies ranging from 5 x 10 to 2 x 10 transductant per CFU, depending on the strain and on the plasmid. In Cry and Cry native recipients, the introduction of the cryIA(a) gene resulted in the formation of large bipyramidal crystals that were active against the insect Plutella xylostella (order Lepidoptera). In both cases, high levels of gene expression were observed. Transductants displaying a dual specificity were constructed by using as recipients the new isolates LM63 and LM79, which have larvicidal activity against insects of the order Coleoptera. It was not possible, however, to introduce pHT7911 into B. thuringiensis subsp. entomocidus, aizawai, or israelensis by transduction. However, electrotransformation was successful, and transformants expressing the toxin gene cryIIIA, carried by pHT7911, were obtained. Again, high levels of expression of the cloned gene were observed. The results indicate that CP-54Ber-mediated transduction is a useful procedure for introducing cloned crystal protein genes into various B. thuringiensis recipients and thereby creating strains with new combinations of genes. Finally it was also shown that pHT3101 is a very good expression vector for the cloned delta-endotoxin genes in the different recipients. PMID:16348674

  6. Pathway and kinetics of cyhalothrin biodegradation by Bacillus thuringiensis strain ZS-19

    PubMed Central

    Chen, Shaohua; Deng, Yinyue; Chang, Changqing; Lee, Jasmine; Cheng, Yingying; Cui, Zining; Zhou, Jianuan; He, Fei; Hu, Meiying; Zhang, Lian-Hui

    2015-01-01

    Cyhalothrin is a common environmental pollutant which poses increased risks to non-target organisms including human beings. This study reported for the first time a newly isolated strain, Bacillus thuringiensis ZS-19 completely degraded cyhalothrin in minimal medium within 72 h. The bacterium transformed cyhalothrin by cleavage of both the ester linkage and diaryl bond to yield six intermediate products. Moreover, a novel degradation pathway of cyhalothrin in strain ZS-19 was proposed on the basis of the identified metabolites. In addition to degradation of cyhalothrin, this strain was found to be capable of degrading 3-phenoxybenzoic acid, a common metabolite of pyrethroids. Furthermore, strain ZS-19 participated in efficient degradation of a wide range of pyrethroids including cyhalothrin, fenpropathrinn, deltamethrin, beta-cypermethrin, cyfluthrin and bifenthrin. Taken together, our results provide insights into the mechanism of cyhalothrin degradation and also highlight the promising potentials of B.thuringiensis ZS-19 in bioremediation of pyrethroid-contaminated environment. This is the first report of (i) degradation of cyhalothrin and other pyrethroids by B.thuringiensis, (ii) identification of 3-phenoxyphenyl acetonitrile and N-(2-isoproxy-phenyl)-4-phenoxy-benzamide as the metabolites in the degradation pathway of pyrethroids, and (iii) a pathway of degradation of cyhalothrin by cleavage of both the ester linkage and diaryl bond in a microorganism. PMID:25740758

  7. Heme sensing in Bacillus thuringiensis: a supplementary HssRS-regulated heme resistance system.

    PubMed

    Schmidt, Rachel M; Carter, Micaela M; Chu, Michelle L; Latario, Casey J; Stadler, Sarah K; Stauff, Devin L

    2016-05-01

    Several Gram-positive pathogens scavenge host-derived heme to satisfy their nutritional iron requirement. However, heme is a toxic molecule capable of damaging the bacterial cell. Gram-positive pathogens within the phylum Firmicutes overcome heme toxicity by sensing heme through HssRS, a two-component system that regulates the heme detoxification transporter HrtAB. Here we show that heme sensing by HssRS and heme detoxification by HrtAB occur in the insect pathogen Bacillus thuringiensis We find that in B. thuringiensis, HssRS directly regulates an operon, hrmXY, encoding hypothetical membrane proteins that are not found in other Firmicutes with characterized HssRS and HrtAB systems. This novel HssRS-regulated operon or its orthologs BMB171_c3178 and BMB171_c3330 are required for maximal heme resistance. Furthermore, the activity of HrmXY is not dependent on expression of HrtAB. These results suggest that B. thuringiensis senses heme through HssRS and induces expression of separate membrane-localized systems capable of overcoming different aspects of heme toxicity. PMID:27030728

  8. Ingestion and Adsorption of Bacillus thuringiensis subsp. israelensis by Gammarus lacustris in the Laboratory

    PubMed Central

    Brazner, John C.; Anderson, Richard L.

    1986-01-01

    Several groups of Gammarus lacustris adults were exposed to solutions containing 0.5 and 5.0 mg of Bacillus thuringiensis subsp. israelensis per liter for 1- or 24-h periods by using traditional static bioassay exposure procedures. During a postexposure holding period, fecal pellets were removed and plated on tryptic soy agar to determine B. thuringiensis subsp. israelensis spore content. The experiments verified that traditional exposure procedures assure ingestion of B. thuringiensis subsp. israelensis spores and provided a mean dose estimate of 1,948 spores ingested per test animal with a 95% confidence interval ranging from 891 to 4,296 (1-h exposure, 5.0 mg/liter). It was also found that dose level is highly dependent upon both exposure duration and concentration and that relatively short exposures can result in a relatively long-term retention of spores postexposure (≥30 days). Body burden experiments established that large numbers of spores adsorb to the bodies of test animals during exposure and may in part explain the long-term retention of spores in the test system postexposure. These results imply that in field applications of microbial control agents, toxicologically unaffected but exposed organisms might transport the agent to untreated sites, expanding the effective treatment area and the number of organisms exposed. PMID:16347242

  9. Pathway and kinetics of cyhalothrin biodegradation by Bacillus thuringiensis strain ZS-19.

    PubMed

    Chen, Shaohua; Deng, Yinyue; Chang, Changqing; Lee, Jasmine; Cheng, Yingying; Cui, Zining; Zhou, Jianuan; He, Fei; Hu, Meiying; Zhang, Lian-Hui

    2015-01-01

    Cyhalothrin is a common environmental pollutant which poses increased risks to non-target organisms including human beings. This study reported for the first time a newly isolated strain, Bacillus thuringiensis ZS-19 completely degraded cyhalothrin in minimal medium within 72 h. The bacterium transformed cyhalothrin by cleavage of both the ester linkage and diaryl bond to yield six intermediate products. Moreover, a novel degradation pathway of cyhalothrin in strain ZS-19 was proposed on the basis of the identified metabolites. In addition to degradation of cyhalothrin, this strain was found to be capable of degrading 3-phenoxybenzoic acid, a common metabolite of pyrethroids. Furthermore, strain ZS-19 participated in efficient degradation of a wide range of pyrethroids including cyhalothrin, fenpropathrinn, deltamethrin, beta-cypermethrin, cyfluthrin and bifenthrin. Taken together, our results provide insights into the mechanism of cyhalothrin degradation and also highlight the promising potentials of B.thuringiensis ZS-19 in bioremediation of pyrethroid-contaminated environment. This is the first report of (i) degradation of cyhalothrin and other pyrethroids by B.thuringiensis, (ii) identification of 3-phenoxyphenyl acetonitrile and N-(2-isoproxy-phenyl)-4-phenoxy-benzamide as the metabolites in the degradation pathway of pyrethroids, and (iii) a pathway of degradation of cyhalothrin by cleavage of both the ester linkage and diaryl bond in a microorganism. PMID:25740758

  10. High-Salt Stress Conditions Increase the pAW63 Transfer Frequency in Bacillus thuringiensis

    PubMed Central

    Beuls, Elise; Modrie, Pauline; Deserranno, Cédric

    2012-01-01

    Conjugation experiments with Bacillus thuringiensis and transfer kinetics demonstrated that salt stress has a positive impact on plasmid transfer efficiency. Compared to standard osmotic conditions (0.5% NaCl), plasmid transfer occurred more rapidly, and at higher frequencies (>100-fold), when bacteria were exposed to a high-salt stress (5% NaCl) in liquid brain heart infusion (BHI). Under milder salt conditions (2.5% NaCl), only a 10-fold effect was observed in Luria-Bertani broth and no difference was detected in BHI. These observations are particularly relevant in the scope of potential gene exchanges among members of the Bacillus cereus group, which includes food-borne contaminants and pathogens. PMID:22820331

  11. Extraction of antibiotic zwittermicin A from Bacillus thuringiensis by macroporous resin and silica gel column chromatography.

    PubMed

    Hao, Zaibin; Yan, Li; Liu, Jianguo; Song, Fuping; Zhang, Jie; Li, Xia

    2015-01-01

    To establish a production process capable of providing refined zwittermicin A (ZwA) on a large scale, the macroporous resin and silica gel column chromatography were used to separate and purify the antibiotic ZwA from the fermentation broth of Bacillus thuringiensis HD-1. The result of high-performance liquid chromatography-mass spectrometry after purification suggests that the samples of ZwA were of high purity, 89%, and the average yield was 20 mg L(-1). Erwinia herbicola LS005, Escherichia coli, Staphylococcus aureus, and Bacillus subtilis were used to assess the toxicity of ZwA. The antibiotic had strong antibacterial activity against E. herbicola LS005 and a color reaction with ninhydrin. PMID:25099664

  12. Potential of sugarcane bagasse (agro-industrial waste) for the production of Bacillus thuringiensis israelensis.

    PubMed

    Poopathi, S; Mani, C; Rajeswari, G

    2013-09-01

    Sugarcane bagasse is a renewable resource that can be used to produce biopesticide for the control of mosquito vectors. In the present study, we demonstrated that cane processed bagasse could be used to produce Bacillus thuringiensis serovar israelensis (Bti) for control of mosquito vectors viz: Culex quinquefasciatus, Anopheles stephensi and Aedes aegypti. Biochemical studies indicated that the Bti spore/crystal toxins produced from the test culture medium (Bagasse, BG + Soybean, SB) are higher than that from the conventional medium (Nutrient Yeast Extract Salt Medium, NYSM). The bacteria produced in these media (NYSM, BG, SB, BG+SB) were bioassayed against the mosquito species and the toxic effect was found to be effective. Cost-effective analysis indicates that the use of BG and SB, as bacterial culture medium, is successful and economical, for production of this mosquito pathogenic bacillus. PMID:24189680

  13. Haematological, biochemical and histopathological alterations induced by abamectin and Bacillus thuringiensis in male albino rats.

    PubMed

    Eissa, F I; Zidan, N A

    2010-03-01

    The renal- and hepato-toxicity induced by abamectin pesticide (Vertimec) and a commercial form of a bio-insecticide Bacillus thuringiensis (Agerin) in male albino rats were evaluated. Blood picture and blood glucose level were investigated. Male albino rats were administered dietary doses each equivalent to 1/10 or 1/100 of the LD50 values of each toxicant for 30 consecutive days. Abamectin was found to pose risks of renal- and hepato-toxicity in rats, since the biochemical parameters of liver function (i.e. aspartate aminotransferase activity, alanine aminotransferase activity, acid phosphatase activity, albumin, and total protein levels) and kidney function (uric acid and creatinine concentration) were severely affected. These effects were verified by histopathological examination of liver and kidney tissues. Likewise, some haematological indices (i.e. erythrocyte count, leukocyte count and haemoglobin concentration) were also influenced; in addition abamectin might cause hypoglycaemia. On the other hand, the above-mentioned lesions were less pronounced in the case of Bacillus thuringiensis -treated rats. PMID:20194097

  14. Cloning and characterization of an insecticidal crystal protein gene from Bacillus thuringiensis subspecies kenyae.

    PubMed

    Misra, Hari S; Khairnar, Nivedita P; Mathur, Manjula; Vijayalakshmi, N; Hire, Ramesh S; Dongre, T K; Mahajan, S K

    2002-04-01

    A sporulating culture of Bacillus thuringiensis subsp. kenyae strain HD549 is toxic to larvae of lepidopteran insect species such as Spodoptera litura, Helicoverpa armigera and Phthorimaea operculella, and a dipteran insect, Culex fatigans. A 1.9-kb DNA fragment, PCR-amplified from HD549 using cryII-gene-specific primers, was cloned and expressed in E. coli. The recombinant protein produced 92% mortality in first-instar larvae of Spodoptera litura and 86% inhibition of adult emergence in Phthorimaea operculella, but showed very low toxicity against Helicoverpa armigera, and lower mortality against third-instar larvae of dipteran insects Culex fatigans, Anopheles stephensi and Aedes aegypti. The sequence of the cloned crystal protein gene showed almost complete homology with a mosquitocidal toxin gene from Bacillus thuringiensis var. kurstaki, with only five mutations scattered in different regions. Amino acid alignment with different insecticidal crystal proteins using the MUTALIN program suggested presence of the conserved block 3 region in the sequence of this protein. A mutation in codon 409 of this gene that changes a highly conserved phenylalanine residue to serine lies in this block. PMID:12357073

  15. Revision of the nomenclature for the Bacillus thuringiensis pesticidal crystal proteins.

    PubMed

    Crickmore, N; Zeigler, D R; Feitelson, J; Schnepf, E; Van Rie, J; Lereclus, D; Baum, J; Dean, D H

    1998-09-01

    The crystal proteins of Bacillus thuringiensis have been extensively studied because of their pesticidal properties and their high natural levels of production. The increasingly rapid characterization of new crystal protein genes, triggered by an effort to discover proteins with new pesticidal properties, has resulted in a variety of sequences and activities that no longer fit the original nomenclature system proposed in 1989. Bacillus thuringiensis pesticidal crystal protein (Cry and Cyt) nomenclature was initially based on insecticidal activity for the primary ranking criterion. Many exceptions to this systematic arrangement have become apparent, however, making the nomenclature system inconsistent. Additionally, the original nomenclature, with four activity-based primary ranks for 13 genes, did not anticipate the current 73 holotype sequences that form many more than the original four subgroups. A new nomenclature, based on hierarchical clustering using amino acid sequence identity, is proposed. Roman numerals have been exchanged for Arabic numerals in the primary rank (e.g., Cry1Aa) to better accommodate the large number of expected new sequences. In this proposal, 133 crystal proteins comprising 24 primary ranks are systematically arranged. PMID:9729610

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

    PubMed

    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 < 0.05) among the 5 rows, with mortality continuously decreasing with increasing distance from sprayer. Both sprayers provided on average >70% larval mortality 3.04-9.14 m from the spray line, and <60% mortality at 12.19 and 15.24 m. The data suggest that the SR450 and Superhawk XP may be comparable sprayers for use with Bti to control mosquito larvae. PMID:25843094

  17. Transcriptome profiling of the intoxication response of Tenebrio molitor larvae to Bacillus thuringiensis Cry3Aa protoxin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacillus thuringiensis (Bt) crystal (Cry) proteins are effective against some coleopteran pests, but improvements are needed in both efficacy and “time to kill” for Cry toxins to become valuable tools for use in integrated pest management. To gain insight into Bt intoxication of Coleoptera, we perfo...

  18. Detection of genes encoding antimicrobial peptides in Mexican strains of Trichoplusia ni (Hubner) exposed to Bacillus thuringiensis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The systemic immune response of Trichoplusia ni after Bacillus thuringiensis (Bt) exposure was evaluated by comparing the expression of genes encoding antimicrobial peptides (AMP) in Bt-susceptible and -resistant T. ni strains that were either exposed or not to XenTari® (Bt-XT). AMP genes were dete...

  19. Effects of four nematodes species on fitness costs of pink bollworm resistance to Bacillus thuringiensis toxin Cry1Ac

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Evolution of resistance by pests can reduce efficacy of transgenic crops that produce insecticidal toxins from the bacterium Bacillus thuringiensis (Bt). In conjunction with refuges of non-Bt host plants, fitness costs can delay the evolution of resistance. Furthermore, fitness costs often vary wit...

  20. Transcriptome of the lymantria dispar (gypsy moth) larval midgut and its response to infection by bacillus thuringiensis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transcriptomic profiles of the serious lepidopteran insect pest Lymantria dispar (gypsy moth) were characterized in the larval midgut in response to infection by Bacillus thuringiensis kurstaki, a biopesticide commonly used for its control in nature. RNA-Seq approaches were used to define a set of ...

  1. Reduced levels of membrane-bound alkaline phosphatase are common to lepidopteran strains resistant to Cry toxins from Bacillus thuringiensis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Development of insect resistance is one of the main concerns with the use of transgenic crops expressing Cry toxins from the bacterium Bacillus thuringiensis. Biomarkers for development of sensitive DNA-based methods to detect and monitor evolution of resistance to Bt toxins are currently needed. ...

  2. A PCR screen of field populations of Heliothis virescens for a retrotransposon insertion conferring resistance to Bacillus thuringiensis toxin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The evolution of pest resistance to transgenic crop plants producing insecticidal toxins from Bacillus thuringiensis Berliner (Bt) poses a continuing threat to their sustainable use in agriculture. One component of the USA-wide resistance management plan for Bt cotton involves monitoring the freque...

  3. 40 CFR 174.502 - Bacillus thuringiensis Cry1A.105 protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1A.105 protein; exemption from the requirement of a tolerance. 174.502 Section 174.502 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances...

  4. 40 CFR 180.1108 - Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Delta endotoxin of Bacillus thuringiensis variety San Diego encapsulated into killed Pseudomonas fluorescens; exemption from the requirement of a tolerance. 180.1108 Section 180.1108 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS TOLERANCES...

  5. 40 CFR 174.529 - Bacillus thuringiensis modified Cry1Ab protein as identified under OECD Unique Identifier SYN...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis modified Cry1Ab protein as identified under OECD Unique Identifier SYN-IR67B-1 in cotton; exemption from the requirement of a tolerance. 174.529 Section 174.529 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES...

  6. 40 CFR 174.504 - Bacillus thuringiensis Cry1F protein in cotton; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1F protein in cotton; exemption from the requirement of a tolerance. 174.504 Section 174.504 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances...

  7. 40 CFR 174.509 - Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance. 174.509 Section 174.509 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances and...

  8. 40 CFR 174.530 - Bacillus thuringiensis Cry2Ae protein in cotton; temporary exemption from the requirement of a...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry2Ae protein in cotton; temporary exemption from the requirement of a tolerance. 174.530 Section 174.530 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED...

  9. Expression of Bacillus thuringiensis Cytolytic Toxin (Cyt2Ca1) in citrus roots to control Diaprepes abbreviatus larvae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  10. 40 CFR 174.520 - Bacillus thuringiensis Cry1F protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1F protein in corn; exemption from the requirement of a tolerance. 174.520 Section 174.520 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances...

  11. Bacillus thuringiensis Cry3Aa protoxin intoxication of Tenebrio molitor induces widespread changes in the expression of serine peptidase transcripts

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  12. Isolation of transcripts from Diabrotica virgifera virgifera LeConte responsive to the Bacillus thuringiensis toxin Cry3Bb1

    EPA Science Inventory

    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...

  13. Isolation of transcripts from Diabrotica virgifera virgifera LeConte responsive to the Bacillus thuringiensis toxin Cry3Bb1

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Crystal (Cry) 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 Diabrotica). ...

  14. Cadherin is a functional receptor of bacillus thuringiensis toxin Cry2Aa in the beet armyworm, spodoptera exigua

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bacillus thuringiensis (Bt) insecticidal crystal (Cry) proteins are effective against some insect pests in sprays and transgenic crops, although the evolution of resistance could threaten the long-term efficacy of such Bt use. One strategy to delay resistance to Bt crops is to “pyramid” two or more ...

  15. Damage and survivorship of fall armyworm (Lepidoptera: noctuidae) on transgenic field corn expressing Bacillus thuringiensis cry proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Field corn, Zea mays L. plants expressing Cry1Ab and Cry1F insecticidal crystal (Cry) proteins of Bacillus thuringiensis (Bt) Berliner are planted on considerable acreage across the Southern region of the United States. The fall armyworm, Spodoptera frugiperda (J. E. Smith), is an economically impo...

  16. Genetic Variation for Resistance to Bacillus thuringiensis Toxins in Helicoverpa zea (Lepidoptera: Noctuidae) in Eastern North Carolina

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In order to evaluate resistance to Bacillus thuringiensis Berliner toxins, female bollworm moths, Helicoverpa zea (Boddie), were collected from four light trap locations in two eastern North Carolina counties from August-October during 2001 and 2002. Moths were allowed to oviposit, and upon hatch, ...

  17. Dipel-selected Ostrinia nubilalis larvae are not resistant to transgenic corn expressing Bacillus thuringiensis Cry1Ab

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The survival of KS-SC DipelTM-resistant and -susceptible European corn borer, Ostrinia nubilalis Hübner, was evaluated on different tissues from corn hybrids, including a non-transgenic and two transgenic corn plants (events MON810 and Bt11) expressing high doses of Bacillus thuringiensis (Bt) delta...

  18. Reduction of Bacillus thuringiensis Cry1Ac Toxicity Against Helicoverpa armigera by a Soluble Toxin-Binding Cadherin Fragment

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A cadherin-like protein has been identified as a putative receptor for Bacillus thuringiensis (Bt) Cry1Ac toxin in Helicoverpa armigera and plays a key role in Bt insecticidal action. In this study, we produced a fragment from this H. armigera Cry1Ac toxin-binding cadherin that included the predict...

  19. Bacillus thuringiensis Resistance Influences European Corn Borer (Lepidoptera: Crambidae) Larval Behavior after Exposure to Cry1Ab

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The behavior of target pests has been recognized as an important factor to appropriately define resistance management plans for Bacillus thuringiensis (Bt) crops. However, most data available do not include the possible impact resistance alleles may have on the behavior of pest larvae or adults. To ...

  20. Integrated use of Beauveria bassiana and Bacillus thuringiensis serovar. tenebrionis for microbial biocontrol of Colorado potato beetle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper reviews the potential for using insect pathogens to control the Colorado potato beetle, Leptinotarsa decemlineata, and summarizes results from nearly 10 years of research by USDA-ARS-PPRU scientists aimed at developing methods and strategies for integrated use of Bacillus thuringiensis an...

  1. 40 CFR 174.505 - Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... protein (mCry3A) in corn; exemption from the requirement of a tolerance. 174.505 Section 174.505... thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn are exempt from the requirement...

  2. 40 CFR 174.532 - Bacillus thuringiensis eCry3.1Ab protein in corn; temporary exemption from the requirement of a...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... protein in corn; temporary exemption from the requirement of a tolerance. 174.532 Section 174.532... thuringiensis eCry3.1Ab protein in corn; temporary exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis eCry3.1Ab protein in corn, in or on the food and feed commodities of corn;...

  3. 40 CFR 174.532 - Bacillus thuringiensis eCry3.1Ab protein in corn; temporary exemption from the requirement of a...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... protein in corn; temporary exemption from the requirement of a tolerance. 174.532 Section 174.532... thuringiensis eCry3.1Ab protein in corn; temporary exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis eCry3.1Ab protein in corn, in or on the food and feed commodities of corn;...

  4. 40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of...

  5. 40 CFR 174.505 - Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... protein (mCry3A) in corn; exemption from the requirement of a tolerance. 174.505 Section 174.505... thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn are exempt from the requirement...

  6. 40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of...

  7. 40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of...

  8. 40 CFR 174.505 - Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... protein (mCry3A) in corn; exemption from the requirement of a tolerance. 174.505 Section 174.505... thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn are exempt from the requirement...

  9. 40 CFR 174.505 - Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... protein (mCry3A) in corn; exemption from the requirement of a tolerance. 174.505 Section 174.505... thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn are exempt from the requirement...

  10. 40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of...

  11. Monitoring Bacillus thuringiensis-Susceptibility in Insect Pests That Occur in Large Geographies: How to Get the Best Information When Two Countries are Involved

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The adoption of Bacillus thuringiensis-expressing cotton around the world has been proven to be beneficial for growers and the environment. The effectiveness of this important genetically-modified crop can be jeopardized by the development of B. thuringiensis-resistance in pests, with the possibilit...

  12. Gut Bacteria Are Not Required for the Insecticidal Activity of Bacillus thuringiensis toward the Tobacco Hornworm, Manduca sexta▿ †

    PubMed Central

    Johnston, Paul R.; Crickmore, Neil

    2009-01-01

    It was recently proposed that gut bacteria are required for the insecticidal activity of the Bacillus thuringiensis-based insecticide, DiPel, toward the lepidopterans Manduca sexta, Pieris rapae, Vanessa cardui, and Lymantria dispar. Using a similar methodology, it was found that gut bacteria were not required for the toxicity of DiPel or Cry1Ac or for the synergism of an otherwise sublethal concentration of Cry1Ac toward M. sexta. The toxicities of DiPel and of B. thuringiensis HD73 Cry− spore/Cry1Ac synergism were attenuated by continuously exposing larvae to antibiotics before bioassays. Attenuation could be eliminated by exposing larvae to antibiotics only during the first instar without altering larval sterility. Prior antibiotic exposure did not attenuate Cry1Ac toxicity. The presence of enterococci in larval guts slowed mortality resulting from DiPel exposure and halved Cry1Ac toxicity but had little effect on B. thuringiensis HD73 Cry− spore/Cry1Ac synergism. B. thuringiensis Cry− cells killed larvae after intrahemocoelic inoculation of M. sexta, Galleria mellonella, and Spodoptera litura and grew rapidly in plasma from M. sexta, S. litura, and Tenebrio molitor. These findings suggest that gut bacteria are not required for B. thuringiensis insecticidal activity toward M. sexta but that B. thuringiensis lethality is reduced in larvae that are continuously exposed to antibiotics before bioassay. PMID:19525273

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

    PubMed Central

    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

  14. Isolation and characterization of a new Bacillus thuringiensis strain with a promising toxicity against Lepidopteran pests.

    PubMed

    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. PMID:27242138

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

    PubMed

    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

    2012-04-01

    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

  16. Efficiency of Intergeneric Recombinants Between Bacillus Thuringiensis and Bacillus Subtilis for Increasing Mortality Rate in Cotten Leaf Worm

    NASA Astrophysics Data System (ADS)

    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 .

  17. Mosquito biolarvicide production by sequential fermentation with dual strains of Bacillus thuringiensis subsp. israelensis and Bacillus sphaericus using sewage sludge.

    PubMed

    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. PMID:20855197

  18. Combining hexanoic acid plant priming with Bacillus thuringiensis insecticidal activity against Colorado potato beetle.

    PubMed

    García-Robles, Inmaculada; Ochoa-Campuzano, Camila; Fernández-Crespo, Emma; Camañes, Gemma; Martínez-Ramírez, Amparo C; González-Bosch, Carmen; García-Agustín, Pilar; Rausell, Carolina; Real, María Dolores

    2013-01-01

    Interaction between insect herbivores and host plants can be modulated by endogenous and exogenous compounds present in the source of food and might be successfully exploited in Colorado potato beetle (CPB) pest management. Feeding tests with CPB larvae reared on three solanaceous plants (potato, eggplant and tomato) resulted in variable larval growth rates and differential susceptibility to Bacillus thuringiensis Cry3Aa toxin as a function of the host plant. An inverse correlation with toxicity was observed in Cry3Aa proteolytic patterns generated by CPB midgut brush-border membrane vesicles (BBMV) from Solanaceae-fed larvae, being the toxin most extensively proteolyzed on potato, followed by eggplant and tomato. We found that CPB cysteine proteases intestains may interact with Cry3Aa toxin and, in CPB BBMV from larvae fed all three Solanaceae, the toxin was able to compete for the hydrolysis of a papain substrate. In response to treatment with the JA-dependent plant inducer Hexanoic acid (Hx), we showed that eggplant reduced OPDA basal levels and both, potato and eggplant induced JA-Ile. CPB larvae feeding on Hx-induced plants exhibited enhanced Cry3Aa toxicity, which correlated with altered papain activity. Results indicated host-mediated effects on B. thuringiensis efficacy against CPB that can be enhanced in combination with Hx plant induction. PMID:23743826

  19. Intravital imaging of Bacillus thuringiensis Cry1A toxin binding sites in the midgut of silkworm.

    PubMed

    Li, Na; Wang, Jing; Han, Heyou; Huang, Liang; Shao, Feng; Li, Xuepu

    2014-02-15

    Identification of the resistance mechanism of insects against Bacillus thuringiensis Cry1A toxin is becoming an increasingly challenging task. This fact highlights the need for establishing new methods to further explore the molecular interactions of Cry1A toxin with insects and the receptor-binding region of Cry1A toxins for their wider application as biopesticides and a gene source for gene-modified crops. In this contribution, a quantum dot-based near-infrared fluorescence imaging method has been applied for direct dynamic tracking of the specific binding of Cry1A toxins, CrylAa and CrylAc, to the midgut tissue of silkworm. The in vitro fluorescence imaging displayed the higher binding specificity of CrylAa-QD probes compared to CrylAc-QD to the brush border membrane vesicles of midgut from silkworm. The in vivo imaging demonstrated that more CrylAa-QDs binding to silkworm midgut could be effectively and distinctly monitored in living silkworms. Furthermore, frozen section analysis clearly indicated the broader receptor-binding region of Cry1Aa compared to that of Cry1Ac in the midgut part. These observations suggest that the insecticidal activity of Cry toxins may depend on the receptor-binding sites, and this scatheless and visual near-infrared fluorescence imaging could provide a new avenue to study the resistance mechanism to maintain the insecticidal activity of B. thuringiensis toxins. PMID:24252542

  20. A novel Tenebrio molitor cadherin is a functional receptor for Bacillus thuringiensis Cry3Aa toxin.

    PubMed

    Fabrick, Jeff; Oppert, Cris; Lorenzen, Marcé D; Morris, Kaley; Oppert, Brenda; Jurat-Fuentes, Juan Luis

    2009-07-01

    Cry toxins produced by the bacterium Bacillus thuringiensis are effective biological insecticides. Cadherin-like proteins have been reported as functional Cry1A toxin receptors in Lepidoptera. Here we present data that demonstrate that a coleopteran cadherin is a functional Cry3Aa toxin receptor. The Cry3Aa receptor cadherin was cloned from Tenebrio molitor larval midgut mRNA, and the predicted protein, TmCad1, has domain structure and a putative toxin binding region similar to those in lepidopteran cadherin B. thuringiensis receptors. A peptide containing the putative toxin binding region from TmCad1 bound specifically to Cry3Aa and promoted the formation of Cry3Aa toxin oligomers, proposed to be mediators of toxicity in lepidopterans. Injection of TmCad1-specific double-stranded RNA into T. molitor larvae resulted in knockdown of the TmCad1 transcript and conferred resistance to Cry3Aa toxicity. These data demonstrate the functional role of TmCad1 as a Cry3Aa receptor in T. molitor and reveal similarities between the mode of action of Cry toxins in Lepidoptera and Coleoptera. PMID:19416969

  1. Combining Hexanoic Acid Plant Priming with Bacillus thuringiensis Insecticidal Activity against Colorado Potato Beetle

    PubMed Central

    García-Robles, Inmaculada; Ochoa-Campuzano, Camila; Fernández-Crespo, Emma; Camañes, Gemma; Martínez-Ramírez, Amparo C.; González-Bosch, Carmen; García-Agustín, Pilar; Rausell, Carolina; Real, María Dolores

    2013-01-01

    Interaction between insect herbivores and host plants can be modulated by endogenous and exogenous compounds present in the source of food and might be successfully exploited in Colorado potato beetle (CPB) pest management. Feeding tests with CPB larvae reared on three solanaceous plants (potato, eggplant and tomato) resulted in variable larval growth rates and differential susceptibility to Bacillus thuringiensis Cry3Aa toxin as a function of the host plant. An inverse correlation with toxicity was observed in Cry3Aa proteolytic patterns generated by CPB midgut brush-border membrane vesicles (BBMV) from Solanaceae-fed larvae, being the toxin most extensively proteolyzed on potato, followed by eggplant and tomato. We found that CPB cysteine proteases intestains may interact with Cry3Aa toxin and, in CPB BBMV from larvae fed all three Solanaceae, the toxin was able to compete for the hydrolysis of a papain substrate. In response to treatment with the JA-dependent plant inducer Hexanoic acid (Hx), we showed that eggplant reduced OPDA basal levels and both, potato and eggplant induced JA-Ile. CPB larvae feeding on Hx-induced plants exhibited enhanced Cry3Aa toxicity, which correlated with altered papain activity. Results indicated host-mediated effects on B. thuringiensis efficacy against CPB that can be enhanced in combination with Hx plant induction. PMID:23743826

  2. Microbial control of the cotton leafworm Spodoptera littoralis (Boisd.) by Egyptian Bacillus thuringiensis isolates.

    PubMed

    Alfazairy, Ahlam A; El-Ahwany, Amani M D; Mohamed, Eman A; Zaghloul, Heba A H; El-Helow, Ehab R

    2013-03-01

    Four local Bacillus thuringiensis (Bt) isolates that had been serologically identified as Bt var. kurstaki (Btk2, Btk3, and Btk66) and Bt var. mexicanensis (Btm27), in addition to two reference strains (4D20 and 4AC1), were laboratory assayed as microbial control agents against the Egyptian cotton leafworm Spodoptera littoralis (Boisd.). Polymerase chain reaction (PCR) amplification analysis revealed that each of the six experimental strains carries, at least, a cry1 type gene which expresses a protein toxin active against lepidopterous insects. Additionally, PCR amplification results demonstrated that 4D20 and Btk66 contain the Lepidoptera- and Diptera-active cry2 type gene and that Btk66 contains Coleoptera-active cry7 and cry8 genes. Among the six strains, Btk66 and Btm27 were the most promising microbial control agents against S. littoralis. The present findings were the first to report that Btm27 (classified as B. thuringiensis var. mexicanensis) is a very potent microbial control agent against S. littoralis-tested larvae. For more characterization of these two isolates, the sspO gene was investigated as a molecular chronometer. The DNA sequencing results proved that Btk66 and Btm27 carry sspO open reading frames with identical nucleotide sequences, suggesting a strong phylogenetic relationship between the two strains. PMID:22983675

  3. Preparation of nanoscale Bacillus thuringiensis chitinases using silica nanoparticles for nematicide delivery.

    PubMed

    Qin, Xu; Xiang, Xuemei; Sun, Xiaowen; Ni, Hong; Li, Lin

    2016-01-01

    A series of amino, carboxylic, and aldehydic surface-grafted silica nanoparticles (SNPs) was prepared based on SiO2 NYSi40 nanoparticles to develop an efficient, biocompatible, and cost-effective biopesticide delivery system. Bacillus thuringiensis chitinase (Chi9602) was immobilized onto SNP surface to prepare nanoscale chitinases (SNPCs) through electrostatic adsorption and covalent binding. The specimens were characterized by Fourier transform infrared, scanning electron microscopy, and zeta-potential analyses. The delivery capacity of the SNPs in Caenorhabditis elegans N2 was observed by immunofluorescence. Results demonstrated that amino-grafted SiO2 nanoparticles with Chi9602 electrostatically adsorbed onto their surface (SNPC2) exhibited a relatively high enzyme immobilization rate (80.2%) and the highest (94.1%) residual enzyme activity among all SNPCs. SNPC2 also showed wider pH tolerance and relatively higher thermostability and ultraviolet radiation resistance capacity than Chi9602. Bioassays further showed that SNPC2 synergistically enhanced the nematicidal effect of B. thuringiensis YBT-020 preparation against C. elegans, with a reduced LC50 of 8.35mg/mL and a shortened LT50 of 12.04h. Immunofluorescence assays showed that SNPC2 had considerable delivery capacity to carry a large protein into C. elegans. Therefore, SNP2 can serve as an efficient nanocarrier for the delivery of macromolecular proteic biopesticides or drugs, indicating potential agricultural or biotechnological applications. PMID:26476241

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

    PubMed

    Wu, Songqing; Lan, Yanjiao; Huang, Dongmei; Peng, Yan; Huang, Zhipeng; Xu, Lei; Gelbic, Ivan; Carballar-Lejarazu, Rebeca; Guan, Xiong; Zhang, Lingling; Zou, Shuangquan

    2014-02-01

    The aim of this study was to explore a cost-effective method for the mass production of Bacillus thuringiensis (Bt) by solid-state fermentation. As a locally available agroindustrial byproduct, spent mushroom substrate (SMS) was used as raw material for Bt cultivation, and four combinations of SMS-based media were designed. Fermentation conditions were optimized on the best medium and the optimal conditions were determined as follows: temperature 32 degrees C, initial pH value 6, moisture content 50%, the ratio of sieved material to initial material 1:3, and inoculum volume 0.5 ml. Large scale production of B. thuringiensis subsp. israelensis (Bti) LLP29 was conducted on the optimal medium at optimal conditions. High toxicity (1,487 international toxic units/milligram) and long larvicidal persistence of the product were observed in the study, which illustrated that SMS-based solid-state fermentation medium was efficient and economical for large scale industrial production of Bt-based biopesticides. The cost of production of 1 kg of Bt was approximately US$0.075. PMID:24665695

  5. Expression of Vitreoscilla hemoglobin in Bacillus thuringiensis improve the cell density and insecticidal crystal proteins yield.

    PubMed

    Liang, Feng; Shouwen, Chen; Ming, Sun; Ziniu, Yu

    2007-02-01

    The Vitreoscilla hemoglobin (VHb) gene (vgb) was integrated into the chromosome of Bacillus thuringiensis BMB171 using integrative vector pEG491. The production of VHb was confirmed by CO-difference spectra analysis. Fermentation experiments results showed that with the production of VHb, the critical oxygen concentration (COC) of the host strain was reduced from 18 to 12%. The maximum viable cell counts of the VHb+ strain in high, middle, and low aeration/agitation fermentations were 0.94-, 1.23-, and 1.59-fold of those of the VHb- strain, respectively. Under the same conditions, the yields of insecticidal crystal proteins (ICP) by VHb+ strain were 1.22-, 1.63-, and 3.13-fold of those of the VHb- strain. The production of VHb also accelerated the formation of ICP and spores. These results indicated that the production of VHb could improve the cell density and ICP yield of B. thuringiensis, especially under low aeration/agitation condition. PMID:17089120

  6. Specific oxygen uptake rate variations during batch fermentation of Bacillus thuringiensis subspecies kurstaki HD-1.

    PubMed

    Rowe, Gerald E; Margaritis, Argyrios; Wei, Ning

    2003-01-01

    The specific oxygen uptake rate (q(O)2, respiration rate) of Bacillus thuringiensis subsp. kurstaki HD-1 was very high at inoculation and was found to decrease essentially monotonically throughout both vegetative growth phase and transition phase under different batch culture conditions. Average q(O)2 values decreased from 8-10 mmol/g h at 1 h after inoculation to less than 2 mmol/g h by the time growth ended. The results are shown to be consistent with the few previous reports on q(O)2 in B. thuringiensis in the literature but also novel in that this pattern of monotonic decline has not been described previously. Both pH control and EDTA in low concentration shortened the vegetative growth phase and reduced the 10 h biomass concentration. Using plots of q(O)2 versus specific growth rate, mu, biomass yield based on the oxygen used for growth, was calculated for transition phase to be 0.041-0.047 g/mmol, consistent with literature values. The same plot also showed that the presence of EDTA resulted in an atypical q(O)2-mu trajectory and apparently much higher biomass yield from the oxygen consumed. PMID:14524704

  7. An overview of the safety and biological effects of Bacillus thuringiensis Cry toxins in mammals.

    PubMed

    Rubio-Infante, Néstor; Moreno-Fierros, Leticia

    2016-05-01

    Crystal proteins (Cry) produced during the growth and sporulation phases of Bacillus thuringiensis (Bt) bacterium are known as delta endotoxins. These toxins are being used worldwide as bioinsecticides to control pests in agriculture, and some Cry toxins are used against mosquitoes to control vector transmission. This review summarizes the relevant information currently available regarding the biosafety and biological effects that Bt and its insecticidal Cry proteins elicit in mammals. This work was performed because of concerns regarding the possible health impact of Cry toxins on vertebrates, particularly because Bt toxins might be associated with immune-activating or allergic responses. The controversial data published to date are discussed in this review considering earlier toxicological studies of B. thuringiensis, spores, toxins and Bt crops. We discussed the experimental studies performed in humans, mice, rats and sheep as well as in diverse mammalian cell lines. Although the term 'toxic' is not appropriate for defining the effects these toxins have on mammals, they cannot be considered innocuous, as they have some physiological effects that may become pathological; thus, trials that are more comprehensive are necessary to determine their effects on mammals because knowledge in this field remains limited. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26537666

  8. Isolation and characterization of a novel insecticidal crystal protein gene from Bacillus thuringiensis subsp. aizawai.

    PubMed

    Chambers, J A; Jelen, A; Gilbert, M P; Jany, C S; Johnson, T B; Gawron-Burke, C

    1991-07-01

    Bacillus thuringiensis subsp. aizawai EG6346, a novel grain dust isolate, was analyzed by Southern blot hybridization for its insecticidal crystal protein (ICP) gene profile. Strain EG6346 lacks previously characterized cryIA ICP genes yet does possess novel cryI-related gene sequences. A recombinant genomic plasmid library was constructed for strain EG6346 in Escherichia coli. One recombinant plasmid, pEG640, isolated from the library contained a novel ICP gene on a 5.7-kb Sau3A insert. The sequence of this gene, designated cryIF, was related to, but distinct from, the published sequences for other cryI genes. A second novel cryI-related sequence was also located on pEG640, approximately 500 bp downstream from cryIF. Introduction of cryIF into a Cry- B. thuringiensis recipient strain via electroporation enabled sufficient production of CryIF protein for quantitative bioassay analyses of insecticidal specificity. The CryIF crystal protein was selectively toxic to a subset of lepidopteran insects tested, including the larvae of Ostrinia nubilalis and Spodoptera exigua. PMID:2061280

  9. Characterization of Bacillus thuringiensis l-Isoleucine Dioxygenase for Production of Useful Amino Acids▿†

    PubMed Central

    Hibi, Makoto; Kawashima, Takashi; Kodera, Tomohiro; Smirnov, Sergey V.; Sokolov, Pavel M.; Sugiyama, Masakazu; Shimizu, Sakayu; Yokozeki, Kenzo; Ogawa, Jun

    2011-01-01

    We determined the enzymatic characteristics of an industrially important biocatalyst, α-ketoglutarate-dependent l-isoleucine dioxygenase (IDO), which was found to be the enzyme responsible for the generation of (2S,3R,4S)-4-hydroxyisoleucine in Bacillus thuringiensis 2e2. Depending on the amino acid used as the substrate, IDO catalyzed three different types of oxidation reactions: hydroxylation, dehydrogenation, and sulfoxidation. IDO stereoselectively hydroxylated several hydrophobic aliphatic l-amino acids, as well as l-isoleucine, and produced (S)-3-hydroxy-l-allo-isoleucine, 4-hydroxy-l-leucine, (S)-4-hydroxy-l-norvaline, 4-hydroxy-l-norleucine, and 5-hydroxy-l-norleucine. The IDO reaction product of l-isoleucine, (2S,3R,4S)-4-hydroxyisoleucine, was again reacted with IDO and dehydrogenated into (2S,3R)-2-amino-3-methyl-4-ketopentanoate, which is also a metabolite found in B. thuringiensis 2e2. Interestingly, IDO catalyzed the sulfoxidation of some sulfur-containing l-amino acids and generated l-methionine sulfoxide and l-ethionine sulfoxide. Consequently, the effective production of various modified amino acids would be possible using IDO as the biocatalyst. PMID:21821743

  10. Characterization of Bacillus thuringiensis L-isoleucine dioxygenase for production of useful amino acids.

    PubMed

    Hibi, Makoto; Kawashima, Takashi; Kodera, Tomohiro; Smirnov, Sergey V; Sokolov, Pavel M; Sugiyama, Masakazu; Shimizu, Sakayu; Yokozeki, Kenzo; Ogawa, Jun

    2011-10-01

    We determined the enzymatic characteristics of an industrially important biocatalyst, α-ketoglutarate-dependent l-isoleucine dioxygenase (IDO), which was found to be the enzyme responsible for the generation of (2S,3R,4S)-4-hydroxyisoleucine in Bacillus thuringiensis 2e2. Depending on the amino acid used as the substrate, IDO catalyzed three different types of oxidation reactions: hydroxylation, dehydrogenation, and sulfoxidation. IDO stereoselectively hydroxylated several hydrophobic aliphatic l-amino acids, as well as l-isoleucine, and produced (S)-3-hydroxy-l-allo-isoleucine, 4-hydroxy-l-leucine, (S)-4-hydroxy-l-norvaline, 4-hydroxy-l-norleucine, and 5-hydroxy-l-norleucine. The IDO reaction product of l-isoleucine, (2S,3R,4S)-4-hydroxyisoleucine, was again reacted with IDO and dehydrogenated into (2S,3R)-2-amino-3-methyl-4-ketopentanoate, which is also a metabolite found in B. thuringiensis 2e2. Interestingly, IDO catalyzed the sulfoxidation of some sulfur-containing l-amino acids and generated l-methionine sulfoxide and l-ethionine sulfoxide. Consequently, the effective production of various modified amino acids would be possible using IDO as the biocatalyst. PMID:21821743

  11. Shared binding sites in Lepidoptera for Bacillus thuringiensis Cry1Ja and Cry1A toxins.

    PubMed

    Herrero, S; González-Cabrera, J; Tabashnik, B E; Ferré, J

    2001-12-01

    Bacillus thuringiensis toxins act by binding to specific target sites in the insect midgut epithelial membrane. The best-known mechanism of resistance to B. thuringiensis toxins is reduced binding to target sites. Because alteration of a binding site shared by several toxins may cause resistance to all of them, knowledge of which toxins share binding sites is useful for predicting cross-resistance. Conversely, cross-resistance among toxins suggests that the toxins share a binding site. At least two strains of diamondback moth (Plutella xylostella) with resistance to Cry1A toxins and reduced binding of Cry1A toxins have strong cross-resistance to Cry1Ja. Thus, we hypothesized that Cry1Ja shares binding sites with Cry1A toxins. We tested this hypothesis in six moth and butterfly species, each from a different family: Cacyreus marshalli (Lycaenidae), Lobesia botrana (Tortricidae), Manduca sexta (Sphingidae), Pectinophora gossypiella (Gelechiidae), P. xylostella (Plutellidae), and Spodoptera exigua (Noctuidae). Although the extent of competition varied among species, experiments with biotinylated Cry1Ja and radiolabeled Cry1Ac showed that Cry1Ja and Cry1Ac competed for binding sites in all six species. A recent report also indicates shared binding sites for Cry1Ja and Cry1A toxins in Heliothis virescens (Noctuidae). Thus, shared binding sites for Cry1Ja and Cry1A occur in all lepidopteran species tested so far. PMID:11722929

  12. Shared Binding Sites in Lepidoptera for Bacillus thuringiensis Cry1Ja and Cry1A Toxins

    PubMed Central

    Herrero, Salvador; González-Cabrera, Joel; Tabashnik, Bruce E.; Ferré, Juan

    2001-01-01

    Bacillus thuringiensis toxins act by binding to specific target sites in the insect midgut epithelial membrane. The best-known mechanism of resistance to B. thuringiensis toxins is reduced binding to target sites. Because alteration of a binding site shared by several toxins may cause resistance to all of them, knowledge of which toxins share binding sites is useful for predicting cross-resistance. Conversely, cross-resistance among toxins suggests that the toxins share a binding site. At least two strains of diamondback moth (Plutella xylostella) with resistance to Cry1A toxins and reduced binding of Cry1A toxins have strong cross-resistance to Cry1Ja. Thus, we hypothesized that Cry1Ja shares binding sites with Cry1A toxins. We tested this hypothesis in six moth and butterfly species, each from a different family: Cacyreus marshalli (Lycaenidae), Lobesia botrana (Tortricidae), Manduca sexta (Sphingidae), Pectinophora gossypiella (Gelechiidae), P. xylostella (Plutellidae), and Spodoptera exigua (Noctuidae). Although the extent of competition varied among species, experiments with biotinylated Cry1Ja and radiolabeled Cry1Ac showed that Cry1Ja and Cry1Ac competed for binding sites in all six species. A recent report also indicates shared binding sites for Cry1Ja and Cry1A toxins in Heliothis virescens (Noctuidae). Thus, shared binding sites for Cry1Ja and Cry1A occur in all lepidopteran species tested so far. PMID:11722929

  13. Characterization of an Argentine isolate of Bacillus thuringiensis similar to the HD-1 strain.

    PubMed

    Sauka, Diego H; Basurto-Ríos, Regina E; Ibarra, Jorge E; Benintende, Graciela B

    2010-01-01

    We report the characterization of an Argentine isolate of Bacillus thuringiensis (INTA TA24-6) similar to the HD-1 strain, which harbors a cryptic cry2Ab gene that apparently is transcribed but not translated into a protein. INTA TA24-6 showed a Rep-PCR pattern identical to the HD-1 strain, a plasmid pattern that resembled that of this strain and cry1 and cry2 genes as HD-1. Screening of cry1 and cry2 genes showed that INTA TA24-6 harbors only cry1Ac and cry2Ab genes. Furthermore, crystalline inclusions of INTA TA24-6 exhibit a bipyramidal shape, typical of Lepidoptera-active B. thuringiensis strains, containing a major protein of ca. 130 kDa toxic to Epinotia aporema Wals. (Lepidoptera: Tortricidae) larvae. Neither the flat-square to cuboidal crystal nor a ca. 65 kDa protein typical of strains expressing Cry2 proteins were detected in INTA TA24-6. In agreement with this information, parasporal crystals of INTA TA24-6 did not show toxicity to Aedes aegypti L. (Diptera:Culicidae) larvae. Gene transcription analyses suggested that the cry2A gene might be cryptic in INTA TA24-6 despite its transcription at different sporulation stages. PMID:21120387

  14. Anthelmintic Effect of Bacillus thuringiensis Strains against the Gill Fish Trematode Centrocestus formosanus.

    PubMed

    Mendoza-Estrada, Luis Javier; Hernández-Velázquez, Víctor Manuel; Arenas-Sosa, Iván; Flores-Pérez, Fernando Iván; Morales-Montor, Jorge; Peña-Chora, Guadalupe

    2016-01-01

    Parasitic agents, such as helminths, are the most important biotic factors affecting aquaculture, and the fluke Centrocestus formosanus is considered to be highly pathogenic in various fish species. There have been efforts to control this parasite with chemical helminthicides, but these efforts have had unsuccessful results. We evaluated the anthelmintic effect of 37 strains of Bacillus thuringiensis against C. formosanus metacercariae in vitro using two concentrations of total protein, and only six strains produced high mortality. The virulence (CL50) on matacercariae of three strains was obtained: the GP308, GP526, and ME1 strains exhibited a LC50 of 146.2 μg/mL, 289.2 μg/mL, and 1721.9 μg/mL, respectively. Additionally, these six B. thuringiensis strains were evaluated against the cercariae of C. formosanus; the LC50 obtained from the GP526 strain with solubilized protein was 83.8 μg/mL, and it could be considered as an alternative control of the metacercariae and cercariae of this parasite in the productivity systems of ornamental fishes. PMID:27294137

  15. Anthelmintic Effect of Bacillus thuringiensis Strains against the Gill Fish Trematode Centrocestus formosanus

    PubMed Central

    Mendoza-Estrada, Luis Javier; Hernández-Velázquez, Víctor Manuel; Arenas-Sosa, Iván; Flores-Pérez, Fernando Iván; Morales-Montor, Jorge; Peña-Chora, Guadalupe

    2016-01-01

    Parasitic agents, such as helminths, are the most important biotic factors affecting aquaculture, and the fluke Centrocestus formosanus is considered to be highly pathogenic in various fish species. There have been efforts to control this parasite with chemical helminthicides, but these efforts have had unsuccessful results. We evaluated the anthelmintic effect of 37 strains of Bacillus thuringiensis against C. formosanus metacercariae in vitro using two concentrations of total protein, and only six strains produced high mortality. The virulence (CL50) on matacercariae of three strains was obtained: the GP308, GP526, and ME1 strains exhibited a LC50 of 146.2 μg/mL, 289.2 μg/mL, and 1721.9 μg/mL, respectively. Additionally, these six B. thuringiensis strains were evaluated against the cercariae of C. formosanus; the LC50 obtained from the GP526 strain with solubilized protein was 83.8 μg/mL, and it could be considered as an alternative control of the metacercariae and cercariae of this parasite in the productivity systems of ornamental fishes. PMID:27294137

  16. Influence of mutagenesis of Bacillus thuringiensis Cry1Aa toxin on larvicidal activity.

    PubMed

    Zhang, Chunyan; Xia, Liqiu; Ding, Xuezhi; Huang, Fan; Li, Huanfa; Sun, Yunjun; Yin, Jia

    2011-03-01

    Domain III of Bacillus thuringiensis Cry δ-endotoxins are considered to be related to the stability of the structure and avoidance of overdigestion by proteases. In this study, some residues of potential chymotrypsin and trypsin sites in Domain III of B. thuringiensis Cry1Aa were replaced individually with alanine by site-directed mutagenesis, in order to investigate their functional roles. Except F574A, all mutants F536A, R543A, F550A, F565A, R566A, F570A, F576A, F583A, and F590A were highly expressed the 130 kD protoxins at levels comparable to the wild-type tested by SDS-PAGE. In bioassays, F536A, R566A, and F590A increased toxicity against Spodoptera exigua Hüner larve by 20, 40, and 40%, respectively, as compared to the wild-type. F536A and F565A showed an increase of 6 and 10% in toxicity against Heliothis armigera Hubner than the wild-type. Toxicities of some mutants were altered greatly, and the same mutants were shown to have different toxicities against those two insects. Structural analyses showed that mutants R543A, F574A, F576A-affecting insecticidal activity might be relational to structural stability of toxin or decreased affinity for receptor binding. These results indicated that those residues were involved in the larvicidal activity of the Cry1Aa toxin. PMID:21082182

  17. Bacillus thuringiensis strain 199 can induce systemic resistance in tomato against Fusarium wilt

    PubMed Central

    Mahboob, Asrar; Javed, Asmat Ali

    2013-01-01

    The research work was performed to investigate the potential of Bacillus thuringiensis strain 199 to induce systemic resistance in tomato against Fusarium wilt. Roots of two-week-old seedlings of tomato plants were primed with bacterial strain. After 10 days of transplantation, some pots of tomato seedlings were provided with inoculum of Fusarium oxysporum lycopersici according to experimental design to induce disease. After 15 days of incubation period, plants challenged with F. oxysporum lycopersici alone were having obvious symptoms of Fusarium wilt. Plants that were treated with B. thuringiensis 199 + F. oxysporum lycopersici were having significant reduction of disease severity. Quantity of total phenolics increased 1.7-fold in bacterial-treated plants as compared to nontreated. Likewise, in case of defense-related enzymes, a significant increase of 1.3-, 1.8-, and 1.4-fold in polyphenol oxidase (PPO), phenyl ammonia lyase (PAL), and peroxidase (PO) was observed in comparison with untreated control. These results, hence, prove the potential of this bacterial strain for use as plant protection agent. PMID:24294498

  18. Synthetic cryIIIA gene from Bacillus thuringiensis improved for high expression in plants.

    PubMed

    Sutton, D W; Havstad, P K; Kemp, J D

    1992-09-01

    A 1974 bp synthetic gene was constructed from chemically synthesized oligonucleotides in order to improve transgenic protein expression of the cryIIIA gene from Bacillus thuringiensis var. tenebrionis in transgenic tobacco. The crystal toxin genes (cry) from B. thuringiensis are difficult to express in plants even when under the control of efficient plant regulatory sequences. We identified and eliminated five classes of sequence found throughout the cryIIIA gene that mimic eukaryotic processing signals and which may be responsible for the low levels of transcription and translation. Furthermore, the GC content of the gene was raised from 36% to 49% and the codon usage was changed to be more plant-like. When the synthetic gene was placed behind the cauliflower mosaic virus 35S promoter and the alfalfa mosaic virus translational enhancer, up to 0.6% of the total protein in transgenic tobacco plants was cryIIIA as measured from immunoblot analysis. Bioassay data using potato beetle larvae confirmed this estimate. PMID:1301214

  19. Mineralization of the Bacillus thuringiensis Cry1Ac endotoxin in soil.

    PubMed

    Accinelli, Cesare; Koskinen, William C; Becker, Joanna M; Sadowsky, Michael J

    2008-02-13

    Although a number of studies have been done describing the fate of Bacillus thuringiensis insecticidal endotoxins in soil, there is conflicting information on the persistence of this class of insecticidal toxins. This is partly due to methodological limitations in many of the previous studies. In the experiments reported here, 14C-labeled B. thuringiensis Cry1Ac endotoxin was used to study its mineralization in soil incubated under controlled conditions. Fifty-nine percent of the radiolabeled Cry1Ac was recovered as 14CO2 at the end of the 20 day incubation period. The addition of 4.5% corn residues stimulated mineralization of [14C]Cry1Ac toxin, and mineralization of glucose was 3.6 times faster than that of the Cry1Ac toxin, indicating that the soil was microbiologically and metabolically active. Because only low mineralization (approximately 6%) of the radiolabeled toxin was observed in autoclaved soil, the current findings indicate that microbial processes play a major role in the dissipation of the Cry1Ac endotoxin in soil. The results of this study suggest that there may be limited risk of the bioaccumulation of Cry1Ac in soil due to the eventual release of this insecticidal toxin by Bt-protected crops. PMID:18181567

  20. Effects of Two Varieties of Bacillus thuringiensis Maize on the Biology of Plodia interpunctella

    PubMed Central

    Gryspeirt, Aiko; Grégoire, Jean-Claude

    2012-01-01

    On the market since 1996, genetically modified plants expressing an insecticidal toxin (Cry toxin stemmed from Bacillus thuringiensis) target several lepidopteran and coleopteran pests. In this study, we assessed the impact of two varieties of Bt maize producing different toxins (Cry1Ab or Cry1Fa, respectively) on the biology of a storage pest: Plodia interpunctella (Hübner) (Lepidoptera: Pyralidae). The Indianmeal moths were susceptible to both toxins but showed an escape behavior only from Cry1Fa. The weight of females issued from larvae reared on Cry1Ab increased with increasing toxin concentration, but adults of both sexes reared on Cry1Fa had decreased weight. Both toxins increased development time from egg to adult regardless of sex and had no impact on the male adult lifespan. Finally, we recorded a time lag between metamorphosis from the non-Bt and the Bt diets, which increased proportionally to Cry concentration in the Bt diet. PMID:22778907

  1. A structured model for vegetative growth and sporulation in Bacillus thuringiensis

    SciTech Connect

    Starzak, M.; Bajpai, R.K.

    1991-12-31

    A mathematical model has been developed for the 6-endotoxin producing Bacillus thuringiensis. The structure of the model involves the processes taking place during vegetative growth, those leading to the initiation of sporulation under conditions of carbon and/or nitrogen limitation, and the sporulation events. The key features in the model are the pools of compounds, such as PRPP, IMP, ADP/ATP, GDP/GTP, pyrimidine nucleotides, NAD/NADH{sub 2}, amino acids, nucleic acids, cell wall, and vegetative and sporulation proteins. These, along with a-factors that control the nature of RNA-polymerase during the different phases, effectively stimulate the vegetative growth and sporulation. The initiation of sporulation is controlled by the intracellular concentration of GTP. Results of simulation of vegetative growth, initiation of sporulation, spore protein formation, and production of {delta}-endotoxin under C- or N-limitation are presented.

  2. Retargeting of the Bacillus thuringiensis toxin Cyt2Aa against hemipteran insect pests

    PubMed Central

    Chougule, Nanasaheb P.; Li, Huarong; Liu, Sijun; Linz, Lucas B.; Narva, Kenneth E.; Meade, Thomas; Bonning, Bryony C.

    2013-01-01

    Although transgenic crops expressing Bacillus thuringiensis (Bt) toxins have been used successfully for management of lepidopteran and coleopteran pest species, the sap-sucking insects (Hemiptera) are not particularly susceptible to Bt toxins. To overcome this limitation, we demonstrate that addition of a short peptide sequence selected for binding to the gut of the targeted pest species serves to increase toxicity against said pest. Insertion of a 12-aa pea aphid gut-binding peptide by adding to or replacing amino acids in one of three loops of the Bt cytolytic toxin, Cyt2Aa, resulted in enhanced binding and toxicity against both the pea aphid, Acyrthosiphon pisum, and the green peach aphid, Myzus persicae. This strategy may allow for transgenic plant-mediated suppression of other hemipteran pests, which include some of the most important pests of global agriculture. PMID:23650347

  3. The Entry Mechanism of Membrane-Containing Phage Bam35 Infecting Bacillus thuringiensis

    PubMed Central

    Gaidelytė, Aušra; Cvirkaitė-Krupovic, Virginija; Daugelavicius, Rimantas; Bamford, Jaana K. H.; Bamford, Dennis H.

    2006-01-01

    The temperate double-stranded DNA bacteriophage Bam35 infects gram-positive Bacillus thuringiensis cells. Bam35 has an icosahedral protein coat surrounding the viral membrane that encloses the linear 15-kbp DNA genome. The protein coat of Bam35 uses the same assembly principle as that of PRD1, a lytic bacteriophage infecting gram-negative hosts. In this study, we dissected the process of Bam35 entry into discrete steps: receptor binding, peptidoglycan penetration, and interaction with the plasma membrane (PM). Bam35 very rapidly adsorbs to the cell surface, and N-acetyl-muramic acid is essential for Bam35 binding. Zymogram analysis demonstrated that peptidoglycan-hydrolyzing activity is associated with the Bam35 virion. We showed that the penetration of Bam35 through the PM is a divalent-cation-dependent process, whereas adsorption and peptidoglycan digestion are not. PMID:16885461

  4. Occurrence and linkage between secreted insecticidal toxins in natural isolates of Bacillus thuringiensis.

    PubMed

    Espinasse, Sylvain; Chaufaux, Josette; Buisson, Christophe; Perchat, Stéphane; Gohar, Michel; Bourguet, Denis; Sanchis, Vincent

    2003-12-01

    Little is known about the occurrence and linkage between secreted insecticidal virulence factors in natural populations of Bacillus thuringiensis (Bt). We carried out a survey of 392 Bt strains isolated from various samples originating from 31 countries. The toxicity profile of the culture supernatants of these strains was determined individually against Anthonomus grandis (Coleoptera) and Spodoptera littoralis (Lepidoptera). We analyzed beta-exotoxin I production and searched for the genes encoding Vip1-2, Vip3, and Cry1I toxins in 125 of these strains. Our results showed that these insecticidal toxins were widespread in Bt but that their distribution was nonrandom, with significant linkage observed between vip3 and cry1I and between vip1-2 and beta-exotoxin I. Strains producing significant amounts of beta-exotoxin I were more frequently isolated from invertebrate samples than from dust, water, soil, or plant samples. PMID:14756535

  5. Toxicity of Bacillus thuringiensis crystal proteins against eri silkworm, Samia cynthia ricini (Lepidoptera: Saturniidae).

    PubMed

    Sandeep Kumar, Donthula; Tarakeswari, Muddanuru; Lakshminarayana, Maddukuri; Sujatha, Mulpuri

    2016-07-01

    Ten purified crystal proteins of Bacillus thuringiensis (Bt) were tested at concentrations ranging from 2.93 to 3000ng/cm(2) for their toxicity to eri silkworm through protein paint bioassays using castor leaves. Based on LC50 values, Cry1Aa (2.6ng/cm(2)) was highly toxic followed by Cry1Ac (29.3ng/cm(2)) and Cry1Ab (68.7ng/cm(2)). The Cry1Ca and Cry1Ea proteins were moderately toxic to eri silkworm larvae and resulted in 23% and 28% mortality, respectively at the highest concentration tested (3000ng/cm(2)). Only reduction in larval weight was observed with Cry2Aa, Cry1Da and Cry9Aa proteins while Cry3Aa and Cry1Ba proteins were found to be nontoxic. PMID:27377590

  6. Expression and characterization of a lytic polysaccharide monooxygenase from Bacillus thuringiensis.

    PubMed

    Zhang, Huiyan; Zhao, Yong; Cao, Hailong; Mou, Guangqing; Yin, Heng

    2015-08-01

    Lytic polysaccharide monooxygenases (LPMOs) are recently discovered oxidative enzymes that are capable of oxidative cleavage of recalcitrant polysaccharides such as chitin or cellulose. Despite the importance of LPMOs in biomass conversion and the large number of lpmo genes in microorganisms, only a few LPMOs have been well studied, and further characterization of these proteins is thus of interest. In this study, a chitin-active AA10 family LPMO from Bacillus thuringiensis subsp. kurstaki, BtLPMO10A, is described. This enzyme generates even-numbered oxidized oligosaccharides as the dominated products from crystalline chitin, however, interestingly, when colloidal chitin is used as the substrate, a ladder of oxidized oligosaccharides is observed. These results provide new insights into the action mode of LPMOs that may be affected by the substrates. PMID:25936286

  7. Growth, sporulation, delta-endotoxins synthesis, and toxicity during culture of bacillus thuringiensis H14.

    PubMed

    Sarrafzadeh, Mohammad H; Guiraud, Joseph P; Lagneau, Christophe; Gaven, Bruno; Carron, Alexandre; Navarro, Jean-Marie

    2005-08-01

    Growth, sporulation, synthesis of delta-endotoxins, and toxicity against the larvae of Aedes aegypti and Culex pipiens were studied during fermentation of Bacillus thuringiensis H14 in a 20-L fermentor. Measurements of optical density and dielectric permittivity for biomass determination suggest a highly promising technique for on-line evaluation of sporulation. The synthesis of 65-, 25- and 130-kDa proteins started at 16, 18, and 23 h, respectively. These proteins were enriched in different ways until the end of culture (48 h). Toxicity in the course of sporulation was significantly different for the larvae of both mosquito species. Maximal activity against Ae. aegypti was obtained at the end of culture, whereas for Cx. pipiens, the sample at 38 h was the most active. PMID:16059772

  8. Overcome of Carbon Catabolite Repression of Bioinsecticides Production by Sporeless Bacillus thuringiensis through Adequate Fermentation Technology.

    PubMed

    Ben Khedher, Saoussen; Jaoua, Samir; Zouari, Nabil

    2014-01-01

    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. PMID:25309756

  9. Optimization of medium composition for the production of mosquitocidal toxins from Bacillus thuringiensis subsp. israelensis.

    PubMed

    Poopathi, Subbiah; Archana, B

    2012-01-01

    Optimization of chicken feather (CF) based culture medium for the production of Bacillus thuringiensis subsp. israelensis (Bti) biomass in combination with the agro industrial by-product (coconut cake, CC) and manganese chloride (MnCl2) has been evaluated. The biomass yield of Bti spore/crystal toxin was highest (12.06 g/L) from the test medium (CF+CC+MnCl2) compared to the reference medium (Luria Bertani, LB). Toxicity assay with Bti produced from the test medium against mosquito vectors (Culex quinquefasciatus, Anopheles stephensi and Aedes aegypti) was also satisfactory and results were comparable with bacteria produced from LB. The results suggest that Bti can be produced to the maximum extent possible as a potential mosquitocidal activity as suggested by the test medium (CF+CC+MnCl2). PMID:22279944

  10. Flocculation of Bacillus thuringiensis var. israelensis suspension and its efficacy against mosquito larvae.

    PubMed

    Szepesszentgyörgyi, A; Bárány, S; Mécs, I

    2005-01-01

    Bacillus thuringiensis ssp. israelensis (Bti) is increasingly used as an ecologically friendly anti-mosquito agent. The bacterium cells undergo fermentation in dilute suspensions; before practical use, therefore it is necessary to concentrate the suspensions. Aggregation by polymers is a powerful tool with which to regulate the stability of suspensions. Typically, polymers at low concentrations destabilize and at high concentrations stabilize colloidal systems. Bti suspensions can be flocculated efficiently by either cationic or anionic polyelectrolytes. Cationic polyelectolytes were found to be the most efficient flocculants for bacterial suspensions. It was shown that the degree of toxicity of the flocculated Bti suspensions for biting mosquito larvae was in the same range than in non-flocculated suspension. PMID:15813223

  11. Cost-effective production of Bacillus thuringiensis by solid-state fermentation.

    PubMed

    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. PMID:15766933

  12. Friction and Adhesion Forces of Bacillus thuringiensis Spores on Planar Surfaces in Atmospheric Systems

    SciTech Connect

    Kweon, Hyojin; Yiacoumi, Sotira; Tsouris, Costas

    2011-01-01

    The kinetic friction force and the adhesion force of Bacillus thuringiensis spores on planar surfaces in atmospheric systems were studied using atomic force microscopy. The influence of relative humidity (RH) on these forces varied for different surface properties including hydrophobicity, roughness, and surface charge. The friction force of the spore was greater on a rougher surface than on mica, which is atomically flat. As RH increases, the friction force of the spores decreases on mica whereas it increases on rough surfaces. The influence of RH on the interaction forces between hydrophobic surfaces is not as strong as for hydrophilic surfaces. The friction force of the spore is linear to the sum of the adhesion force and normal load on the hydrophobic surface. The poorly defined surface structure of the spore and the adsorption of contaminants from the surrounding atmosphere are believed to cause a discrepancy between the calculated and measured adhesion forces.

  13. Control of pine processionary moth, Thaumetopoea pityocampa with Bacillus thuringiensis in Antalya, Turkey.

    PubMed

    Cebeci, H Huseyin; Oymen, R Tamer; Acer, Sabiha

    2010-05-01

    Taumetopoea pityocampa (Den. and Schiff) is one of the most common defoliator insects found in Turkey. Although several methods have been used in attempting to control this major forest pest up to now but the problem still remains largely unsolved in Turkey. There is an urgent need to control and minimize the damages caused by these defoliating caterpillars. Therefore, we planned and applied field treatments using by Foray 76B and VBC 60074 to put forward to the efficiency of these bioinsecticides against PPM. The bioinsecticides included in Bacillus thuringiensis subsp. kurstaki (Btk). The vulnerable performance of a single application has been observed in the field trial of these bioinsecticides with mortality rates ranging from 97 to 99% in Turkey's pine forests. PMID:21047011

  14. Effects of Bacillus thuringiensis kurstaki on Malpighian tubule cells of Thaumetopoea pityocampa (Lepidoptera: Thaumetopoeidae) larvae.

    PubMed

    Ogutchu, Ayşe; Suludere, Zekiye; Uzunhisarcikli, Meltem; Kalender, Yusuf

    2005-01-01

    In this study effects of Bacillus thuringiensis kurstaki (Btk) on Malpighian tubule cells of Thaumetopoea pityocampa (Lepidoptera: Thaumetopoeidae) larvae was investigated by electron microscopy. 3 mg/l Btk was given with food. After Btk administration, the Malpighian tubule cells were investigated and compared with a control group. 3 and 6 hrs after Btk administration swelling in Malpighian tubule cells was observed. Swelling of mitochondria and separation of their cristae was seen after 12 hrs. After 24 hrs dissolution of the basal cytoplasm, swelling and vacuolization of all mitochondria, partial dissolution of the nucleoplasm, and swelling and separation ofmicrovilli was documented. A membrane-body in the nucleus was seen after 48 hrs. The nucleoplasm was completely dissolved after 72 hrs and after 96 hrs large vacuoles appeared in the cytoplasm and shortening of microvilli was observed. PMID:16212102

  15. Norway spruce (Picea abies) genetic transformation with modified Cry3A gene of Bacillus thuringiensis.

    PubMed

    Bříza, Jindřich; Pavingerová, Daniela; Vlasák, Josef; Niedermeierová, Hana

    2013-01-01

    Modified versions of the Cry3A gene of Bacillus thuringiensis (Bt) were transferred into Norway spruce (Picea abies). Both the biolistic approach and Agrobacterium tumefaciens mediated procedure were employed for transformation of embryogenic tissue (ET) cultures. The latter method proved to be more efficient yielding 70 transgenic embryogenic tissue lines compared with 18 lines obtained by biolistics. The modified Cry3A genes were driven by a 35S promoter and the nptII screenable selection marker gene was used in all vectors. The transgenic ETs were molecularly characterized and converted into mature somatic embryos. Germinating embryos formed plantlets which were finally planted into perlite and their Cry3A gene transcription activities were demonstrated by RT-PCR. PMID:23888296

  16. Diversity of Bacillus thuringiensis in the rice field soils of different ecologies in India.

    PubMed

    Das, J; Dangar, T K

    2007-12-01

    Diversity of the Bacillus thuringiensis (Bt) in the rice field soils of different ecologies viz. the island (Port Blair), the Himalayan (Srinagar), brackish water (Mahe) and coastal mesophilic (Mangalore) habitats was analyzed by phenotypic characterization of 5, 66, 14 and 54 Bt isolates, respectively. The Bt isolates produced either monotypic (bipyramidal or spherical) or heterotypic (polymorphic-bipyramidal or bipyramidal-rhomboidal) crystals. The organisms were generally resistant to the penicillin group of antibiotics, tolerated 5-12% NaCl and 0.5M Na-acetate. The Bt isolates contained 1-5 plasmids of 0.89-58.61 kbp sizes. The plasmid profiles had no correlation with crystal morphology or salt tolerance of different bacteria. Each soil was inhabited by different types of Bt. Two Bt strains of Mangalore and one strain each of the other places were phenotypically similar. One Bt strain each of Port Blair and Srinagar was different from all other strains. PMID:23100691

  17. Frequency of resistance to Bacillus thuringiensis in field populations of pink bollworm

    PubMed Central

    Tabashnik, Bruce E.; Patin, Amanda L.; Dennehy, Timothy J.; Liu, Yong-Biao; Carrière, Yves; Sims, Maria A.; Antilla, Larry

    2000-01-01

    Strategies for delaying pest resistance to genetically modified crops that produce Bacillus thuringiensis (Bt) toxins are based primarily on theoretical models. One key assumption of such models is that genes conferring resistance are rare. Previous estimates for lepidopteran pests targeted by Bt crops seem to meet this assumption. We report here that the estimated frequency of a recessive allele conferring resistance to Bt toxin Cry1Ac was 0.16 (95% confidence interval = 0.05–0.26) in strains of pink bollworm (Pectinophora gossypiella) derived from 10 Arizona cotton fields during 1997. Unexpectedly, the estimated resistance allele frequency did not increase from 1997 to 1999 and Bt cotton remained extremely effective against pink bollworm. These results demonstrate that the assumptions and predictions of resistance management models must be reexamined. PMID:11087854

  18. Bacillus thuringiensis Cry1Aa toxin-binding region of Bombyx mori aminopeptidase N.

    PubMed

    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. PMID:10606725

  19. Bacillus thuringiensis as a specific, safe, and effective tool for insect pest control.

    PubMed

    Roh, Jong Yul; Choi, Jae Young; Li, Ming Shun; Jin, Byung Rae; Je, Yeon Ho

    2007-04-01

    Bacillus thuringiensis (Bt) was first described by Berliner [10] when he isolated a Bacillus species from the Mediterranean flour moth, Anagasta kuehniella, and named it after the province Thuringia in Germany where the infected moth was found. Although this was the first description under the name B. thuringiensis, it was not the first isolation. In 1901, a Japanese biologist, Ishiwata Shigetane, discovered a previously undescribed bacterium as the causative agent of a disease afflicting silkworms. Bt was originally considered a risk for silkworm rearing but it has become the heart of microbial insect control. The earliest commercial production began in France in, 1938, under the name Sporeine [72]. A resurgence of interest in Bt has been attributed to Edward Steinhaus [105], who obtained a culture in 1942 and attracted attention to the potential of Bt through his subsequent studies. In 1956, T. Angus [3] demonstrated that the crystalline protein inclusions formed in the course of sporulation were responsible for the insecticidal action of Bt. By the early 1980's, Gonzalez et al. [48] revealed that the genes coding for crystal proteins were localized on transmissible plasmids, using a plasmid curing technique, and Schnepf and Whiteley [103] first cloned and characterized the genes coding for crystal proteins that had toxicity to larvae of the tobacco hornworm, from plasmid DNA of Bt subsp. kurstaki HD-1. This first cloning was followed quickly by the cloning of many other cry genes and eventually led to the development of Bt transgenic plants. In the 1980s, several scientists successively demonstrated that plants can be genetically engineered, and finally, Bt cotton reached the market in 1996 [104]. PMID:18051264

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

    PubMed Central

    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

  1. Distinct clpP genes control specific adaptive responses in Bacillus thuringiensis.

    PubMed

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

    2002-10-01

    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 approximately 88 and 67% amino acid sequence identity with ClpP of B. subtilis, respectively. Inactivation of clpC in B. thuringiensis impaired sporulation efficiency. The clpP1 and clpP2 mutants were both slightly susceptible to salt stress, whereas disruption of clpP2 negatively affected sporulation and abolished motility. Virulence of the clp mutants was assessed by injecting bacteria into the hemocoel of Bombyx mori larvae. The clpP1 mutant displayed attenuated virulence, which appeared to be related to its inability to grow at low temperature (25 degrees C), suggesting an essential role for ClpP1 in tolerance of low temperature. Microscopic examination of clpP1 mutant cells grown at 25 degrees C showed altered bacterial division, with cells remaining attached after septum formation. Analysis of lacZ transcriptional fusions showed that clpP1 was expressed at 25 and 37 degrees C during the entire growth cycle. In contrast, clpP2 was expressed at 37 degrees C but not at 25 degrees C, suggesting that ClpP2 cannot compensate for the absence of ClpP1 in the clpP1 mutant cells at low temperature. Our study demonstrates that ClpP1 and ClpP2 control distinct cellular regulatory pathways in B. thuringiensis. PMID:12270812

  2. Novel Bacillus thuringiensis Binary Insecticidal Crystal Proteins Active on Western Corn Rootworm, Diabrotica virgifera virgifera LeConte

    PubMed Central

    Ellis, R. Tracy; Stockhoff, Brian A.; Stamp, Lisa; Schnepf, H. Ernest; Schwab, George E.; Knuth, Mark; Russell, Josh; Cardineau, Guy A.; Narva, Kenneth E.

    2002-01-01

    A new family of insecticidal crystal proteins was discovered by screening sporulated Bacillus thuringiensis cultures for oral activity against western corn rootworm (WCR) larvae. B. thuringiensis isolates PS80JJ1, PS149B1, and PS167H2 have WCR insecticidal activity attributable to parasporal inclusion bodies containing proteins with molecular masses of ca. 14 and 44 kDa. The genes encoding these polypeptides reside in apparent operons, and the 14-kDa protein open reading frame (ORF) precedes the 44-kDa protein ORF. Mutagenesis of either gene in the apparent operons dramatically reduced insecticidal activity of the corresponding recombinant B. thuringiensis strain. Bioassays performed with separately expressed, biochemically purified 14- and 44-kDa polypeptides also demonstrated that both proteins are required for WCR mortality. Sequence comparisons with other known B. thuringiensis insecticidal proteins failed to reveal homology with previously described Cry, Cyt, or Vip proteins. However, there is evidence that the 44-kDa polypeptide and the 41.9- and 51.4-kDa binary dipteran insecticidal proteins from Bacillus sphaericus are evolutionarily related. The 14- and 44-kDa polypeptides from isolates PS80JJ1, PS149B1, and PS167H2 have been designated Cry34Aa1, Cry34Ab1, and Cry34Ac1, respectively, and the 44-kDa polypeptides from these isolates have been designated Cry35Aa1, Cry35Ab1, and Cry35Ac1, respectively. PMID:11872461

  3. Novel Bacillus thuringiensis binary insecticidal crystal proteins active on western corn rootworm, Diabrotica virgifera virgifera LeConte.

    PubMed

    Ellis, R Tracy; Stockhoff, Brian A; Stamp, Lisa; Schnepf, H Ernest; Schwab, George E; Knuth, Mark; Russell, Josh; Cardineau, Guy A; Narva, Kenneth E

    2002-03-01

    A new family of insecticidal crystal proteins was discovered by screening sporulated Bacillus thuringiensis cultures for oral activity against western corn rootworm (WCR) larvae. B. thuringiensis isolates PS80JJ1, PS149B1, and PS167H2 have WCR insecticidal activity attributable to parasporal inclusion bodies containing proteins with molecular masses of ca. 14 and 44 kDa. The genes encoding these polypeptides reside in apparent operons, and the 14-kDa protein open reading frame (ORF) precedes the 44-kDa protein ORF. Mutagenesis of either gene in the apparent operons dramatically reduced insecticidal activity of the corresponding recombinant B. thuringiensis strain. Bioassays performed with separately expressed, biochemically purified 14- and 44-kDa polypeptides also demonstrated that both proteins are required for WCR mortality. Sequence comparisons with other known B. thuringiensis insecticidal proteins failed to reveal homology with previously described Cry, Cyt, or Vip proteins. However, there is evidence that the 44-kDa polypeptide and the 41.9- and 51.4-kDa binary dipteran insecticidal proteins from Bacillus sphaericus are evolutionarily related. The 14- and 44-kDa polypeptides from isolates PS80JJ1, PS149B1, and PS167H2 have been designated Cry34Aa1, Cry34Ab1, and Cry34Ac1, respectively, and the 44-kDa polypeptides from these isolates have been designated Cry35Aa1, Cry35Ab1, and Cry35Ac1, respectively. PMID:11872461

  4. Evidence of Bacillus thuringiensis intra-serovar diversity revealed by Bacillus cereus group-specific repetitive extragenic palindromic sequence-based PCR genomic fingerprinting.

    PubMed

    Sauka, Diego H; Basile, Juan I; Benintende, Graciela

    2011-01-01

    Bacillus thuringiensis is classified into serovars on the basis of H-flagellar antigens. Several alternative typing methods have been described. Among them, a B. cereus group-specific repetitive extragenic palindromic (Rep)-PCR fingerprinting technique was shown to be discriminative and able to identify B. thuringiensis serovars. The aim of this study was to investigate the genomic diversity and relationship among B. thuringiensis strains collected from different Argentinean ecosystems. Thirty-seven B. thuringiensis reference strains and 131 Argentinean isolates were analyzed using a B. cereus group-specific Rep-PCR. Fourteen different patterns were identified among the Argentinean isolates. Eight could not be associated to any pattern obtained from a reference strain. The pattern identical to the serovar kurstaki HD-1 strain was the most frequently identified in 68 native isolates. The profiles allowed tracing a single dendrogram with two groups and eight main lineages. Some strains showed distinctive patterns despite belonging to the same serovar. An intraspecific diversity resulted from this analysis that was highlighted by this technique since strains from a given serovar showed distinct profiles. This study may help to establish a system of B. thuringiensis classification with a higher discrimination level than established by the H antigen serotyping. PMID:22286045

  5. Isolation and Identification of novel toxins from a new mosquitocidal isolate from Malaysia, Bacillus thuringiensis subsp. jegathesan.

    PubMed

    Kawalek, M D; Benjamin, S; Lee, H L; Gill, S S

    1995-08-01

    A new mosquitocidal Bacillus thuringiensis subsp., jegathesan, has recently been isolated from Malaysia. Parasporal crystal inclusions were purified from this strain and bioassayed against fourth-instar larvae of Culex quinquefasciatus, Aedes aegypti, Aedes togoi, Aedes albopictus, Anopheles maculatus, and Mansonia uniformis. The 50% lethal concentration of crystal inclusions for each species was 0.34, 8.08, 0.34, 17.59, 3.91, and 120 ng/ml, respectively. These values show that parasporal inclusions from this new subspecies have mosquitocidal toxicity comparable to that of inclusions isolated from B. thuringiensis subsp. israelensis. Solubilized and chymotrypsin-activated parasporal inclusions possessed low-level hemolytic activity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that the crystals were composed of polypeptides of 77, 74, 72, 68, 55, 38, 35, 27, and 23 kDa. Analysis by Western blotting (immunoblotting) with polyclonal antisera raised against toxins purified from B. thuringiensis subsp. israelensis reveals that proteins in parasporal inclusions of subsp. jegathesan are distinct, because little cross-reactivity was shown. Analysis of the plasmid content of B. thuringiensis subsp. jegathesan indicates that the genes for toxin production may be located on 105- to 120-kb plasmids. Cry- clones that have been cured of these plasmids are nontoxic. Southern blot analysis of plasmid and chromosomal DNA from subsp. jegathesan showed little or low homology to the genes coding for CryIVA, CryIVB, and CryIVD from B. thuringiensis subsp. israelensis. PMID:7487029

  6. Effect of fermentation conditions on the enterotoxigenicity, cytotoxicity and pesticidal activity of Bacillus thuringiensis strains isolated in Taiwan.

    PubMed

    Pang, Jen-Chieh; Chen, Ming-Lun; Ho, Yi-Cheng; Yang, Chi-Yea; Tzeng, Ching-Chou; Kao, Suey-Sheng; Tsen, Hau-Yang

    2010-03-01

    A total of 75 Bacillus thuringiensis strains, among them 62 of Taiwan's microbiota, were screened for their enterotoxin genes, hemolysin BL activity and cytotoxicity. All the strains harbored enterotoxin genes and were cytotoxic to the cultivated Chinese hamster ovary (CHO) cells. The hemolysin BL and cytotoxicity titers of the B. thuringiensis culture in casitone yeast sucrose (CYS) broth were lower than those in brain heart infusion (BHI) broth, and when the B. thuringiensis strains were cultivated in CYS broth for 5 days, no cytotoxicity was detected. The spores and crystal toxins collected from 40 isolates showed high levels of insecticidal activity against Plutella xylostella. All strains exhibiting low cytotoxicity also had low pesticidal activity. Our study demonstrated that it is difficult to find B. thuringiensis strains that are both effective against insect targets and do not produce enterotoxins or cytotoxic effects in CHO cells. However, it is possible to avoid or reduce unwanted properties, but not the insecticidal activity, of some B. thuringiensis preparations by alteration of culture media and conditions. PMID:19880313

  7. Efficacy of Bacillus thuringiensis, Paecilomyces marquandii,and Streptomyces costaricanus with and without Organic Amendments against Meloidogyne hapla Infecting Lettuce.

    PubMed

    Chen, J; Abawi, G S; Zuckerman, B M

    2000-03-01

    Chitin, wheat mash, or brewery compost were incorporated into unfumigated and methyl bromide-fumigated organic soils placed in microplots formed from cylindrical drainage tiles (0.25 m-diam. clay tile). After 3 weeks, Meloidogyne hapla and cell or spore suspensions of Bacillus thuringiensis, Paecilomyces marquandii, and Streptomyces costaricanus were individually added to the soils of designated microplots. A B. thuringiensis + S. costaricanus combination was also tested. Lettuce seedlings, cv. Montello, were transplanted into the soils 3 to 4 days later. All the bacterial and fungal antagonists applied without a soil amendment, except the B. thuringiensis + S. costaricanus treatment, reduced root galling and increased lettuce head weight in the unfumigated organic soil, but not in the fumigated soil. All three amendments were also effective against M. hapla and reduced root galling in fumigated and unfumigated soils. Wheat mash amendment increased lettuce head weight in the unfumigated soil. In general, no antagonist x amendment interaction was detected. Soil populations of B. thuringiensis were maintained at >/=4.0 log10 colony-forming units/g organic soil during the first 14 days after planting. However, viable cells of B. thuringiensis were not detected after 49 days. PMID:19270951

  8. Efficacy of Bacillus thuringiensis, Paecilomyces marquandii,and Streptomyces costaricanus with and without Organic Amendments against Meloidogyne hapla Infecting Lettuce

    PubMed Central

    Chen, J.; Abawi, G. S.; Zuckerman, B. M.

    2000-01-01

    Chitin, wheat mash, or brewery compost were incorporated into unfumigated and methyl bromide-fumigated organic soils placed in microplots formed from cylindrical drainage tiles (0.25 m-diam. clay tile). After 3 weeks, Meloidogyne hapla and cell or spore suspensions of Bacillus thuringiensis, Paecilomyces marquandii, and Streptomyces costaricanus were individually added to the soils of designated microplots. A B. thuringiensis + S. costaricanus combination was also tested. Lettuce seedlings, cv. Montello, were transplanted into the soils 3 to 4 days later. All the bacterial and fungal antagonists applied without a soil amendment, except the B. thuringiensis + S. costaricanus treatment, reduced root galling and increased lettuce head weight in the unfumigated organic soil, but not in the fumigated soil. All three amendments were also effective against M. hapla and reduced root galling in fumigated and unfumigated soils. Wheat mash amendment increased lettuce head weight in the unfumigated soil. In general, no antagonist × amendment interaction was detected. Soil populations of B. thuringiensis were maintained at ≥4.0 log10 colony-forming units/g organic soil during the first 14 days after planting. However, viable cells of B. thuringiensis were not detected after 49 days. PMID:19270951

  9. Identification and characterization of a novel cytotoxic protein, parasporin-4, produced by Bacillus thuringiensis A1470 strain.

    PubMed

    Okumura, Shiro; Saitoh, Hiroyuki; Ishikawa, Tomoyuki; Mizuki, Eiichi; Inouye, Kuniyo

    2008-01-01

    In 1901, a unique bacterium was isolated as a pathogen of the sotto disease of the silkmoth larvae, and later in 1915, the organism was described as Bacillus thuringiensis. Since the discovery, this bacterium has widely attracted attention of not only insect pathologists but many other scientists who are interested in strong and specific insecticidal activity associated with inclusion bodies of B. thuringiensis. This has led to the recent worldwide development of B. thuringiensis-based microbial insecticides and insect-resistant transgenic plants, as well as the epoch-making discovery of parasporin, a cancer cell-specific cytotoxin. In the review, we introduce a detection study of interaction between inclusion proteins of B. thuringiensis and brush border membrane of insects using surface plasmon resonance-based biosensor, and then identification and cloning of parasporin-4, a latest cancer cell-killing protein produced by B. thuringiensis A1470 strain. Inclusion bodies of the parasporin-4 produced by recombinant Escherichia coli were solubilized and activated with a new method and purified by an anion-exchange chromatography. At last the characterization of the recombinant parasporin-4 was shown. PMID:18606366

  10. Evidence of two mechanisms involved in Bacillus thuringiensis israelensis decreased toxicity against mosquito larvae: Genome dynamic and toxins stability.

    PubMed

    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. PMID:26070692

  11. In Vitro Ovicidal and Cestocidal Effects of Toxins from Bacillus thuringiensis on the Canine and Human Parasite Dipylidium caninum

    PubMed Central

    Peña, Guadalupe; Aguilar Jiménez, Fortino Agustín; Hallal-Calleros, Claudia; Morales-Montor, Jorge; Hernández-Velázquez, Víctor Manuel; Flores-Pérez, Fernando Iván

    2013-01-01

    Bacillus thuringiensis is a gram-positive soil-dwelling bacterium that is commonly used as a biological pesticide. This bacterium may also be used for biological control of helminth parasites in domestic animals. In this study, we evaluated the possible ovicidal and cestocidal effects of a total protein extract of B. thuringiensis native strains on the zoonotic cestode parasite of dogs, Dipylidium caninum (D. caninum). Dose and time response curves were determined by coincubating B. thuringiensis proteins at concentration ranging from 100 to 1000 μg/mL along with 4000 egg capsules of D. caninum. Egg viability was evaluated using the trypan blue exclusion test. The lethal concentration of toxins on eggs was 600 μg/ml, and the best incubation time to produce this effect was 3 h. In the adult stage, the motility and the thickness of the tegument were used as indicators of damage. The motility was inhibited by 100% after 8 hours of culture compared to the control group, while the thickness of the cestode was reduced by 34%. Conclusively, proteins of the strain GP526 of B. thuringiensis directly act upon D. caninum showing ovicidal and cestocidal effects. Thus, B. thuringiensis is proposed as a potential biological control agent against this zoonosis. PMID:23484087

  12. The Pathogenomic Sequence Analysis of B. cereus and B.thuringiensis Isolates Closely Related to Bacillus anthracis

    SciTech Connect

    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.

  13. [A new resolution vector with cry1Ac10 gene based on Bacillus thuringiensis transposon Tn4430].

    PubMed

    Wu, L; Sun, M; Yu, Z

    2000-06-01

    A new resolution vector with cry1Ac10 gene based on TnpI-mediated site-specific recombination system of Bacillus thuringiensis(Bt) transposon Tn4430 was developed. The gene cry1Ac10, encoding a protoxin against plutella xylostella larvae, and the gene ori1030, from a plasmid of wide type Bacillus thuringiensis, were inserted into two copy sets of RES sites, named pBMB801. When pBMB801 was introduced into crystal negative Bt host BMB171, antibiotic resistance genes and other non-Bt DNA can be selectively eliminated. This recombinant plasmid was found very stable without antibiotic selection. The resulting strain only contained Bt DNA and is free of antibiotic resistance genes. This strategy should facilitate regulatory approval for its development as a commercial biopesticide. PMID:12548990

  14. Identification of a Bacillus thuringiensis Cry11Ba toxin-binding aminopeptidase from the mosquito, Anopheles quadrimaculatus

    PubMed Central

    Abdullah, Mohd Amir F; Valaitis, Algimantas P; Dean, Donald H

    2006-01-01

    Background Aminopeptidase N (APN) type proteins isolated from several species of lepidopteran insects have been implicated as Bacillus thuringiensis (Bt) toxin-binding proteins (receptors) for Cry toxins. We examined brush border membrane vesicle (BBMV) proteins from the mosquito Anopheles quadrimaculatus to determine if APNs from this organism would bind mosquitocidal Cry toxins that are active to it. Results A 100-kDa protein with APN activity (APNAnq 100) was isolated from the brush border membrane of Anopheles quadrimaculatus. Native state binding analysis by surface plasmon resonance shows that APNAnq 100 forms tight binding to a mosquitocidal Bt toxin, Cry11Ba, but not to Cry2Aa, Cry4Ba or Cry11Aa. Conclusion An aminopeptidase from Anopheles quadrimaculatus mosquitoes is a specific binding protein for Bacillus thuringiensis Cry11Ba. PMID:16716213

  15. Purification and identification of a novel leucine aminopeptidase from Bacillus thuringiensis israelensis.

    PubMed

    Cahan, Rivka; Hetzroni, Efrat; Nisnevitch, Marina; Nitzan, Yeshayahu

    2007-11-01

    A novel leucine aminopeptidase was purified from a Bacillus thuringiensis israelensis (Bti) culture. The purification stages included heating the concentrated supernatant to 65 degrees C for 90 min, anion-exchange chromatography by DEAE cellulose, and hydrophobic chromatography by phenyl Sepharose. The specific activity of leucine aminopeptidase after the hydrophobic chromatography increased by 215.5-fold and the yield was 16%. The molecular weight of the active enzyme was 59 kDa. Mass spectrometry analysis of the 59-kDa leucine aminopeptidase revealed that this protein has at least 41% homology with the cytosol leucine aminopeptidase produced by Bacillus cereus. Maximal leucine aminopeptidase activity occurred at 65 degrees C, pH 10 toward leucine as the amino acid terminus. The enzyme was strongly inhibited by bestatin, dithiothreitol, and 1,10-phenanthroline, indicating that the enzyme might be considered as a metallo-aminopeptidase that has disulfide bonds at the catalytic site or at a region that influences its configuration. Examination of the purified leucine aminopeptidase's effect on the activation of the protoxin Cyt1Aa from Bti revealed that when it acts synergistically with Bti endogenous proteases, it has only a minor role in the processing of Cyt1Aa into an active toxin. PMID:17682820

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

    EPA Science Inventory

    Abstract

    The 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...

  17. [The effect of a soil extract on the development of Bacillus thuringiensis and on its synthesis of an insecticidal endotoxin].

    PubMed

    Dregval', O A; Cherevach, N V; Andrienko, O E; Vinnikov, A I

    1999-01-01

    Selection of effective and inexpensive nutrient medium for cultivation of entomopathogenic bacteria Bacillus thuringiensis was carried out. The medium with molasses [correction of patoka], corn extract and mineral salts has been chosen. Addition of a soil extract to the medium enhanced growth of microorganisms, increased the rate of culture development, the yield of spore-crystalline material and quantity of synthesized endotoxin. Apparently the strengthening effect belongs to humic substances contained in the soil extract. PMID:10565149

  18. Chromosome-Directed PCR-Based Detection and Quantification of Bacillus cereus Group Members with Focus on B. thuringiensis Serovar israelensis Active against Nematoceran Larvae

    PubMed Central

    Hendriksen, Niels B.; Melin, Petter; Lundström, Jan O.; Sundh, Ingvar

    2015-01-01

    Bacillus thuringiensis serovar israelensis is a wide-spread soil bacterium affiliated with the B. cereus group (Bcg) and is widely used in biocontrol products applied against mosquito and black fly larvae. For monitoring and quantification of applied B. thuringiensis serovar israelensis and its effect on indigenous B. thuringiensis serovar israelensis and Bcg assemblages, efficient and reliable tools are essential. The abundance and properties of B. thuringiensis serovar israelensis strains in the environment traditionally have been investigated with cultivation-dependent techniques, which are hampered by low sensitivity and the morphological similarity between B. cereus and B. thuringiensis. Currently available PCR-based detection and quantification tools target markers located on plasmids. In this study, a new cultivation-independent PCR-based method for efficient and specific quantification of B. thuringiensis serovar israelensis and Bcg is presented, utilizing two sets of PCR primers targeting the bacterial chromosome. Sequence database searches and empirical tests performed on target and nontarget species, as well as on bulk soil DNA samples, demonstrated that this diagnostic tool is specific for B. thuringiensis serovar israelensis and Bcg. The method will be useful for comparisons of Bcg and B. thuringiensis serovar israelensis abundances in the same samples. Moreover, the effect of B. thuringiensis serovar israelensis-based insecticide application on the total Bcg assemblages, including indigenous populations, can be investigated. This type of information is valuable in risk assessment and policy making for use of B. thuringiensis serovar israelensis in the environment. PMID:25979887

  19. Application of statistical experimental design for optimisation of bioinsecticides production by sporeless Bacillus thuringiensis strain on cheap medium

    PubMed Central

    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. PMID:24516462

  20. [Expression of gene aiiA carrying the promoter of gene cry3Aa in Bacillus thuringiensis].

    PubMed

    Zhu, Chen-Guang; Sun, Ming; Yu, Zi-Niu

    2003-07-01

    N-acyl-homoserine lactones (AHLs), are widely conserved signal molecules present in quorum-sensing systems of many Gram-negative bacteria. AHLs molecules mediate the expression of virulence genes of a range of bacterial pathogens. Recently, it has been reported that AiiA protein, which widely exists in Bacillus species, can inactivate the AHLs by hydrolyzing the lactone bond of AHLs, thus attenuate the diseases caused by the expression of virulence genes of bacterial pathogens. Bacillus thuringiensis, a type of Gram-positive bacteria, has been used extensively as a microbial insecticide in the last few decades. However, most of important insecticidal B. thuringiensis strains have not been exploited for bacterial disease control because they usually do not produce antibiotics that are effective against bacteria and fungi. The discovery of AiiA protein in B. thuringiensis shows the application potential of B. thuringiensis on biocontrol against bacterial diseases. In this study, in order to construct the B. thuringiensis recombinant strain that has high expression of AiiA protein, the promoter of insecticidal crystal protein coding gene cry3Aa of B. thuringiensis was selected. The promoter of gene cry3Aa is a non-sporulation promoter, it promotes the transcription earlier and longer than the promoters of other cry genes. The promoter of AiiA protein coding gene aiiA was replaced with the promoter of gene cry3Aa by overlapping PCR, resulting fusion gene pro3A-aiiA. The gene pro3A-aiiA was inserted into shuttle vector pHT304 at site BamH I / Sph I , resulting recombinant plasmid pBMB686. The plasmid pBMB686 was introduced into B. thuringiensis acrystalliferous strain BMB171, the resulting strain BMB686 had a higher and more stable expression level of protein AiiA comparing with the parental strain BMB171. Furthermore, the strain BMB686 exhibited stronger ability of AHLs inactivation and much more effective restraint to the potato's soft rot disease caused by Erwinia

  1. Plasmid Capture by the Bacillus thuringiensis Conjugative Plasmid pXO16▿

    PubMed Central

    Timmery, Sophie; Modrie, Pauline; Minet, Olivier; Mahillon, Jacques

    2009-01-01

    Conjugation, mobilization, and retromobilization are three related mechanisms of horizontal gene transfer in bacteria. They have been extensively studied in gram-negative species, where retromobilization, the capture of DNA from a recipient by a donor cell, was shown to result from two successive steps: the transfer of the conjugative plasmid from the donor to the recipient followed by the retrotransfer of the mobilizable plasmid to the donor. This successive model was established for gram-negative bacteria but was lacking experimental data from the gram-positive counterparts. In the present work, the mobilization and retromobilization abilities of the conjugative plasmid pXO16 from Bacillus thuringiensis subsp. israelensis were studied using the mobilizable plasmids pUB110 and pE194 and the “nonmobilizable” element pC194 lacking the mob and oriT features (all from Staphylococcus aureus). Experimental data suggested a successive model, since different retromobilization frequencies were observed between the small plasmids. More importantly, retromobilization was shown to be delayed by 50 and 150 min for pUB110 and pE194, respectively, compared to pXO16 conjugation. Natural liquid foods (cow milk, soy milk, and rice milk) were used to evaluate the putative ecological impact of these transfers. In cow and soy milk, conjugation, mobilization, and retromobilization were shown to occur at frequencies of 8.0 × 10−1, 1.0 × 10−2, and 1.2 × 10−4 transconjugants per recipient, respectively. These data are comparable to those obtained with LB medium and about 10-fold lower than in the case of rice milk. Taken together, these results emphasize the potential role of plasmid capture played by B. thuringiensis in natural environments. PMID:19181805

  2. Potato flour mediated solid-state fermentation for the enhanced production of Bacillus thuringiensis-toxin.

    PubMed

    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. PMID:23773700

  3. Identification and Characterization of Three Previously Undescribed Crystal Proteins from Bacillus thuringiensis subsp. jegathesan

    PubMed Central

    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

  4. Molecular characterization and genetic diversity of insecticidal crystal protein genes in native Bacillus thuringiensis isolates.

    PubMed

    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. PMID:23207696

  5. Effect of midgut proteolytic activity on susceptibility of lepidopteran larvae to Bacillus thuringiensis subsp. Kurstaki

    PubMed Central

    Talaei-Hassanloui, Reza; Bakhshaei, Raziyeh; Hosseininaveh, Vahid; Khorramnezhad, Ayda

    2014-01-01

    Bacillus thuringiensis (Bt) is the most effective microbial control agent for controlling numerous species from different insect orders. All subspecies and strains of B. thuringiensis can produce a spore and a crystalline parasporal body. This crystal which contains proteinaceous protoxins is dissolved in the alkaline midgut, the resulting molecule is then cleaved and activated by proteolytic enzymes and acts as a toxin. An interesting aspect of this activation process is that variations in midgut pH and protease activity have been shown to account for the spectrum of some Bt proteins activity. Thus, an important factor that could be a determinant of toxin activity is the presence of proteases in the midgut microenvironment of susceptible insects. Reciprocally, any alteration in the midgut protease composition of the host can result in resistance to Bt. Here in this paper, we reviewed this processes in general and presented our assays to reveal whether resistance mechanism to Bt in Diamondback Moth (DbM) larvae could be due to the function of the midgut proteases? We estimated LC50 for both probable susceptible and resistant populations in laboratory and greenhouse tests. Then, the midgut protease activities of the B. thuringiensis induced-resistant and susceptible populations of the DbM were assayed on Hemoglubin and on N-alpha-benzoyl-DL-arginine-p-nitroanilide (BapNA) for total and tryptic activities, respectively. Six hours after feeding on Bt treated and untreated canola leaves, the midguts of instar larvae of both populations were isolated. Following related protocols, peptides released through the activity of proteinases on Hemoglubin and BApNA were recorded using microplate reader. Control (Blank) was also considered with adding TCA to reaction mix before adding enzymatic extract. Data analysis indicated that there are significant differences for tryptic activity on BApNA and also for total proteolytic activity on Hemoglubin between susceptible and

  6. Transcriptome analysis of Bacillus thuringiensis spore life, germination and cell outgrowth in a vegetable-based food model.

    PubMed

    Bassi, Daniela; Colla, Francesca; Gazzola, Simona; Puglisi, Edoardo; Delledonne, Massimo; Cocconcelli, Pier Sandro

    2016-05-01

    Toxigenic species belonging to Bacillus cereus sensu lato, including Bacillus thuringiensis, cause foodborne outbreaks thanks to their capacity to survive as spores and to grow in food matrixes. The goal of this work was to assess by means of a genome-wide transcriptional assay, in the food isolate B. thuringiensis UC10070, the gene expression behind the process of spore germination and consequent outgrowth in a vegetable-based food model. Scanning electron microscopy and Energy Dispersive X-ray microanalysis were applied to select the key steps of B. thuringiensis UC10070 cell cycle to be analyzed with DNA-microarrays. At only 40 min from heat activation, germination started rapidly and in less than two hours spores transformed in active growing cells. A total of 1646 genes were found to be differentially expressed and modulated during the entire B. cereus life cycle in the food model, with most of the significant genes belonging to transport, transcriptional regulation and protein synthesis, cell wall and motility and DNA repair groups. Gene expression studies revealed that toxin-coding genes nheC, cytK and hblC were found to be expressed in vegetative cells growing in the food model. PMID:26742618

  7. Effects of algae on the efficacy of Bacillus thuringiensis var. israelensis against larval black flies.

    PubMed

    Stephens, Marianne S; Overmyer, Jay P; Gray, Elmer W; Noblet, Ray

    2004-06-01

    Personnel from several black fly control programs have reported that the efficacy of Bacillus thuringiensis var. israelesis (Bti) is reduced during periods when algal concentrations are high in the waterways. Although the reduction in Bti-induced mortality in black fly larvae is presumed to be related to the presence of algae, no scientific data support this theory. In this study, 4 genera of algae (Microcytis, Scenedesmus, Dictrosphaerium, and Chlorella) commonly detected in Pennsylvania rivers where Bti-induced mortality in black fly larvae has been reduced were assessed to determine their respective effects on Bti-induced mortality by using an orbital shaker bioassay with laboratory-reared black fly larvae (Simulium vittatum cytospecies IS-7). A significant reduction in Bti-induced mortality was observed when Scenedesmus was present in the flasks at concentrations > or = 16,000 cells/ml. The Bti-induced mortality of larvae was not significantly reduced when Chlorella, Dictyosphaerium, or Microcytis was present in the flasks, even at concentrations > or = 250,000 cells/ml. These results indicate that the presence of certain types of algae can reduce the mortality of black flies exposed to Bti. Although not clearly defined, the mechanisms involved may be related to algal morphology due to overall size and structures associated with certain types of algae, and possible interference with feeding. PMID:15264627

  8. Toxicity and Affecting Factors of Bacillus thuringiensis var. Israelensis on Chironomus kiiensis Larvae

    PubMed Central

    Cao, Chuan-Wang; Sun, Li-Li; Wen, Rong-Rong; Li, Xiao-Peng; Wu, Hong-Qu; Wang, Zhi-Ying

    2012-01-01

    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. PMID:23465075

  9. Ultralow volume application of Bacillus thuringiensis ssp. israelensis for the control of mosquitoes.

    PubMed

    Lee, H L; Gregorio, E R; Khadri, M S; Seleena, P

    1996-12-01

    Evaluation of the effectiveness of Bacillus thuringiensis ssp. israelensis (B.t.i.) against mosquito larvae dispersed by ultralow volume (ULV) spraying was conducted in simulated field trials. Effectiveness was measured using 3 different indicators: larval mortality, colony-forming unit enumeration, and droplet analysis. B.t.i. was dispersed with a ULV generator using 2 different flow rates: 0.3 and 0.5 liter/min on 2 different days. Based on the results of this study, it can be concluded that an output of 0.3 liter/min is effective for controlling Aedes aegypti. although a dosage of 0.5 liter/min can be used when high residual activity is desired. For Culex quinquefasciatus control, both dosages were effective but with low residual activity. For Anopheles maculatus control, only a discharge rate of 0.5 liter/min was effective with low residual activity. B.t.i. application at both dosages penetrated tires well, indicating that B.t.i. ULV application is an effective method for controlling container-inhabiting mosquitoes. Good coverage of target area and penetration were attributed to satisfactory droplet profiles. PMID:9046471

  10. Host–Pathogen Coevolution: The Selective Advantage of Bacillus thuringiensis Virulence and Its Cry Toxin Genes

    PubMed Central

    Papkou, Andrei; Laehnemann, David; Guenther, Patrick S.; Prahl, Swantje; Saebelfeld, Manja; Hollensteiner, Jacqueline; Liesegang, Heiko; Brzuszkiewicz, Elzbieta; Daniel, Rolf; Michiels, Nicolaas K.; Schulte, Rebecca D.; Kurtz, Joachim; Rosenstiel, Philip; Telschow, Arndt; Bornberg-Bauer, Erich; Schulenburg, Hinrich

    2015-01-01

    Reciprocal coevolution between host and pathogen is widely seen as a major driver of evolution and biological innovation. Yet, to date, the underlying genetic mechanisms and associated trait functions that are unique to rapid coevolutionary change are generally unknown. We here combined experimental evolution of the bacterial biocontrol agent Bacillus thuringiensis and its nematode host Caenorhabditis elegans with large-scale phenotyping, whole genome analysis, and functional genetics to demonstrate the selective benefit of pathogen virulence and the underlying toxin genes during the adaptation process. We show that: (i) high virulence was specifically favoured during pathogen–host coevolution rather than pathogen one-sided adaptation to a nonchanging host or to an environment without host; (ii) the pathogen genotype BT-679 with known nematocidal toxin genes and high virulence specifically swept to fixation in all of the independent replicate populations under coevolution but only some under one-sided adaptation; (iii) high virulence in the BT-679-dominated populations correlated with elevated copy numbers of the plasmid containing the nematocidal toxin genes; (iv) loss of virulence in a toxin-plasmid lacking BT-679 isolate was reconstituted by genetic reintroduction or external addition of the toxins. We conclude that sustained coevolution is distinct from unidirectional selection in shaping the pathogen's genome and life history characteristics. To our knowledge, this study is the first to characterize the pathogen genes involved in coevolutionary adaptation in an animal host–pathogen interaction system. PMID:26042786

  11. Stable Bacillus thuringiensis transgene introgression from Brassica napus to wild mustard B. juncea.

    PubMed

    Cao, Di; Stewart, C Neal; Zheng, Min; Guan, Zhengjun; Tang, Zhi-Xi; Wei, Wei; Ma, Ke-Ping

    2014-10-01

    Transgenic canola (Brassica napus) with a Bacillus thuringiensis cry1Ac gene and a green fluorescent protein (GFP) marker gene was used in hybridization experiments with wild Brassica juncea. Hybrid F1 and successive five backcross generations were obtained. The pod-set frequency on backcrossed B. juncea plants was over 66%, which suggested relatively high crossing compatibility between the hybrids and wild species. The seed setting in BC1 was the least of all generations tested, and then increased at the BC2 generation for which the thousand-seed weight was the highest of all generations. Seed size in backcrossed generations eventually approached that of the wild parent. The plants in all backcrossed generations were consistent with the expected 1:1 segregation ratio of the transgenes. The Bt Cry1Ac protein concentrations at bolting and flowering stages was higher compared to the 4-5-leaf and pod-formation stages. Nonetheless, the Bt toxin in the fifth backcrossing generation (BC5) was sufficient to kill both polyphagous (Helicoverpa armigera) and oligophagous (Plutella xylostella) Lepidoptera. As a consequence, the subsequent generations harboring the transgene from F1 to BC5 could have selection advantage against insect pests. The result is useful in understanding gene flow from transgenic crops and the followed transgene introgression into wild. PMID:25219305

  12. Transcriptome of the Lymantria dispar (gypsy moth) larval midgut in response to infection by Bacillus thuringiensis.

    PubMed

    Sparks, Michael E; Blackburn, Michael B; Kuhar, Daniel; Gundersen-Rindal, Dawn E

    2013-01-01

    Transcriptomic profiles of the serious lepidopteran insect pest Lymantria dispar (gypsy moth) were characterized in the larval midgut in response to infection by Bacillus thuringiensis kurstaki, a biopesticide commonly used for its control. RNA-Seq approaches were used to define a set of 49,613 assembled transcript sequences, of which 838, 1,248 and 3,305 were respectively partitioned into high-, mid- and low-quality tiers on the basis of homology information. Digital gene expression profiles suggested genes differentially expressed at 24 hours post infection, and qRT-PCR analyses were performed for verification. The differentially expressed genes primarily associated with digestive function, including α-amylase, lipase and carboxypeptidase; immune response, including C-type lectin 4; developmental genes such as arylphorin; as well as a variety of binding proteins: cellular retinoic acid binding protein (lipid-binding), insulin-related peptide binding protein (protein-binding) and ovary C/EBPg transcription factor (nucleic acid-binding). This is the first study conducted to specifically investigate gypsy moth response to a bacterial infection challenge using large-scale sequencing technologies, and the results highlight important genes that could be involved in biopesticide resistance development or could serve as targets for biologically-based control mechanisms of this insect pest. PMID:23658687

  13. Evaluation of Bt (Bacillus thuringiensis) corn on mouse testicular development by dual parameter flow cytometry.

    PubMed

    Brake, Denise G; Thaler, Robert; Evenson, Donald P

    2004-04-01

    The health safety of Bt (Bacillus thuringiensis) corn (Zea mays L.) was studied using mouse testes as a sensitive biomonitor of potential toxic effects. Pregnant mice were fed a Bt corn or a nontransgenic (conventional) diet during gestation and lactation. After they were weaned, young male mice were maintained on the respective diets. At 8, 16, 26, 32, 63, and 87 days after birth, three male mice and an adult reference mouse were killed, the testes were surgically removed, and the percentage of germ cell populations was measured by flow cytometry. Multigenerational studies were conducted in the same manner. There were no apparent differences in percentages of testicular cell populations (haploid, diploid, and tetraploid) between the mice fed the Bt corn diet and those fed the conventional diet. Because of the high rate of cell proliferation and extensive differentiation that makes testicular germ cells highly susceptible to some toxic agents, it was concluded that the Bt corn diet had no measurable or observable effect on fetal, postnatal, pubertal, or adult testicular development. If data from this study were extrapolated to humans, Bt corn is not harmful to human reproductive development. PMID:15053558

  14. Response Surface Methodology: Optimisation of Antifungal Bioemulsifier from Novel Bacillus thuringiensis

    PubMed Central

    Venkatachalam, Ponnusami

    2014-01-01

    An antifungal bioemulsifier compound was produced from a novel strain of Bacillus thuringiensis pak2310. To accentuate the production and as the first step to improve the yield, a central composite design (CCD) was used to study the effect of various factors like minimal salts (1X and 3X), glycerol concentration (2% and 4%), beef extract concentration (1% and 3%), and sunflower oil concentration (2% and 4%) on the production of bioemulsifier molecule and to optimize the conditions to increase the production. The E24 emulsification index was used as the response variable as the increase in surfactant production was seen to be proportional to increased emulsification. A quadratic equation was employed to express the response variable in terms of the independent variables. Statistical tools like student's t-test, F-test, and ANOVA were employed to identify the important factors and to test the adequacy of the model. Under optimum conditions (1X concentration of minimal salts (MS), 2.6% glycerol (v/v), 1% beef extract (w/v), and 2% sunflower oil (v/v)) a 65% increase in yield was produced. PMID:25379529

  15. Bacillus thuringiensis (Bt) toxin susceptibility and isolation of resistance mutants in the nematode Caenorhabditis elegans.

    PubMed Central

    Marroquin, L D; Elyassnia, D; Griffitts, J S; Feitelson, J S; Aroian, R V

    2000-01-01

    The protein toxins produced by Bacillus thuringiensis (Bt) are the most widely used natural insecticides in agriculture. Despite successful and extensive use of these toxins in transgenic crops, little is known about toxicity and resistance pathways in target insects since these organisms are not ideal for molecular genetic studies. To address this limitation and to investigate the potential use of these toxins to control parasitic nematodes, we are studying Bt toxin action and resistance in Caenorhabditis elegans. We demonstrate for the first time that a single Bt toxin can target a nematode. When fed Bt toxin, C. elegans hermaphrodites undergo extensive damage to the gut, a decrease in fertility, and death, consistent with toxin effects in insects. We have screened for and isolated 10 recessive mutants that resist the toxin's effects on the intestine, on fertility, and on viability. These mutants define five genes, indicating that more components are required for Bt toxicity than previously known. We find that a second, unrelated nematicidal Bt toxin may utilize a different toxicity pathway. Our data indicate that C. elegans can be used to undertake detailed molecular genetic analysis of Bt toxin pathways and that Bt toxins hold promise as nematicides. PMID:10924467

  16. Quantitative spectral light scattering polarimetry for monitoring fractal growth pattern of Bacillus thuringiensis bacterial colonies

    NASA Astrophysics Data System (ADS)

    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.

  17. Isolation, characterization and biological role of camelysin from Bacillus thuringiensis subsp. israelensis.

    PubMed

    Nisnevitch, Marina; Sigawi, Sasi; Cahan, Rivka; Nitzan, Yeshayahu

    2010-09-01

    The present study reports a simple rapid method for isolating the zinc-containing metalloprotease camelysin from Bacillus thuringiensis subsp. israelensis (Bti) by extraction from intact bacterial cells with egg L-alpha-phosphatidylcholine containing monolamellar liposomes, followed by separation on a sucrose gradient. Characterization of the isolated camelysin revealed a molecular weight of 23 kDa and a pI of 6.2. The camelysin exhibited maximal activity against the substrate azocasein at a temperature of 37 degrees C and pH 7.5. However, the enzyme's activity remained high also at basic pH values (8-10). In a rich growth medium (LB), camelysin appeared at the late logarithmic phase of Bti growth and reached its maximum in the stationary phase. Camelysin was shown to activate the protoxins Cyt1Aa and Cyt2Ba produced by Bti. The hemolytic activity of Cyt1Aa increased from 40 to 70% and that of Cyt2Ba from 6 to 50% in the presence of 50% (w/w) camelysin. It is concluded that these protoxins can be activated not only by insect gut proteases, but also by the endogeneous metalloprotease camelysin of the Bti bacterium. PMID:20127334

  18. Mode of action of Bacillus thuringiensis Cry and Cyt toxins and their potential for insect control.

    PubMed

    Bravo, Alejandra; Gill, Sarjeet S; Soberón, Mario

    2007-03-15

    Bacillus thuringiensis Crystal (Cry) and Cytolitic (Cyt) protein families are a diverse group of proteins with activity against insects of different orders--Lepidoptera, Coleoptera, Diptera and also against other invertebrates such as nematodes. Their primary action is to lyse midgut epithelial cells by inserting into the target membrane and forming pores. Among this group of proteins, members of the 3-Domain Cry family are used worldwide for insect control, and their mode of action has been characterized in some detail. Phylogenetic analyses established that the diversity of the 3-Domain Cry family evolved by the independent evolution of the three domains and by swapping of domain III among toxins. Like other pore-forming toxins (PFT) that affect mammals, Cry toxins interact with specific receptors located on the host cell surface and are activated by host proteases following receptor binding resulting in the formation of a pre-pore oligomeric structure that is insertion competent. In contrast, Cyt toxins directly interact with membrane lipids and insert into the membrane. Recent evidence suggests that Cyt synergize or overcome resistance to mosquitocidal-Cry proteins by functioning as a Cry-membrane bound receptor. In this review we summarize recent findings on the mode of action of Cry and Cyt toxins, and compare them to the mode of action of other bacterial PFT. Also, we discuss their use in the control of agricultural insect pests and insect vectors of human diseases. PMID:17198720

  19. Development of photoperiod- and thermo-sensitive male sterility rice expressing transgene Bacillus thuringiensis

    PubMed Central

    Liu, Xin; Zhang, Jiwen; Zhang, Cuicui; Wang, Liangchao; Chen, Hao; Zhu, Zengrong; Tu, Jumin

    2015-01-01

    Stem borers and leaffolders are the main pests that cause severe damage in rice (Oryza sativa L.) production worldwide. We developed the first photoperiod- and thermo-sensitive male sterility (PTSMS) rice 208S with the cry1Ab/1Ac Bacillus thuringiensis (Bt) gene, through sexual crossing with Huahui 1 (elite line with the cry1Ab/1Ac gene). The novel 208S and its hybrids presented high and stable resistance to stem borers and leaffolders, and the content of Cry1Ab/1Ac protein in chlorophyllous tissues achieved the identical level as donor and showed little accumulation in non-chlorophyllous tissue. No dominant dosage effect in the Bt gene was observed in 208S and its derived hybrids. An analysis of fertility transition traits indicated that 208S was completely sterile under long day length/high temperature, but partially fertile under short day length/low temperature. With fine grain quality and favorable combining ability, 208S had no observed negative effects on fertility and agronomic traits from Bt (cry1Ab/1Ac). Additionally, 208S as a male sterile line showed no fertility decrease caused by Bt transgenic process, as it is the case in Huahui 1. Thus, 208S has great application value in two-line hybrid production for insect resistance, and can also be used as a bridge material in rice Bt transgenic breeding. PMID:26366116

  20. Response surface methodology: optimisation of antifungal bioemulsifier from novel Bacillus thuringiensis.

    PubMed

    Rajendran, Deepak; Venkatachalam, Ponnusami; Ramakrishnan, Jayapradha

    2014-01-01

    An antifungal bioemulsifier compound was produced from a novel strain of Bacillus thuringiensis pak2310. To accentuate the production and as the first step to improve the yield, a central composite design (CCD) was used to study the effect of various factors like minimal salts (1X and 3X), glycerol concentration (2% and 4%), beef extract concentration (1% and 3%), and sunflower oil concentration (2% and 4%) on the production of bioemulsifier molecule and to optimize the conditions to increase the production. The E 24 emulsification index was used as the response variable as the increase in surfactant production was seen to be proportional to increased emulsification. A quadratic equation was employed to express the response variable in terms of the independent variables. Statistical tools like student's t-test, F-test, and ANOVA were employed to identify the important factors and to test the adequacy of the model. Under optimum conditions (1X concentration of minimal salts (MS), 2.6% glycerol (v/v), 1% beef extract (w/v), and 2% sunflower oil (v/v)) a 65% increase in yield was produced. PMID:25379529

  1. Toxicity of Bacillus thuringiensis Cry proteins to Helicoverpa armigera (Lepidoptera: Noctuidae) in South Africa.

    PubMed

    Li, Hua; Bouwer, Gustav

    2012-01-01

    The susceptibility of one of the most important pests in southern Africa, Helicoverpa armigera (Lepidoptera: Noctuidae), to Bacillus thuringiensis Cry proteins was evaluated by bioassay. Cry proteins were produced in Escherichia coli BL21 cells that were transformed with plasmids containing one of six cry genes. The toxicity of each Cry protein to H. armigera larvae was determined by the diet contamination method for second instar larvae and the droplet feeding method for neonate larvae. For each of the proteins, dose-mortality and dose-growth inhibition responses were analyzed and the median lethal dose (LD(50)) and median inhibitory dose (ID(50)) determined. Second instar larvae were consistently less susceptible to the evaluated Cry proteins than neonate larvae. The relative toxicity of Cry proteins ranked differently between neonate larvae and second instar larvae. On the basis of the LD(50) and ID(50) values, Cry1Ab, Cry1Ac, and Cry2Aa were the most toxic of the evaluated proteins to H. armigera larvae. The study provides an initial benchmark of the toxicity of individual Cry proteins to H. armigera in South Africa. PMID:22019386

  2. Division of labour and terminal differentiation in a novel Bacillus thuringiensis strain.

    PubMed

    Deng, Chao; Slamti, Leyla; Raymond, Ben; Liu, Guiming; Lemy, Christelle; Gominet, Myriam; Yang, Jingni; Wang, Hengliang; Peng, Qi; Zhang, Jie; Lereclus, Didier; Song, Fuping

    2015-02-01

    A major challenge in bacterial developmental biology has been to understand the mechanisms underlying cell fate decisions. Some differentiated cell types display cooperative behaviour. Cooperation is one of the greatest mysteries of evolutionary biology and microbes have been considered as an excellent system for experimentally testing evolution theories. Bacillus thuringiensis (Bt) is a spore-forming bacterium, which is genetically closely related to B. anthracis, the agent of anthrax, and to B. cereus, an opportunistic human pathogen. The defining feature that distinguishes Bt from its relatives is its ability to produce crystal inclusions in the sporulating cells. These toxins are solubilized after ingestion and are cooperative public goods in insect hosts. In this study, we describe a Bt strain LM1212 that presents the unique ability to terminally differentiate into crystal producers and spore formers. Transcriptional analysis based on lacZ and gfp reporter genes suggested that this phenotype is the consequence of a new type of cell differentiation associated with a novel regulation mode of cry gene expression. The differentiating crystal-producer phenotype has higher spore productivity than a typical Bt strain and is better able to compete with Cry toxin null 'cheaters'. Potentially, this division of labour provides additional fitness benefits in terms of spore viability or durability of Cry toxin. PMID:25083932

  3. A two-year field study with transgenic Bacillus thuringiensis maize: effects on soil microorganisms.

    PubMed

    Oliveira, Adília P; Pampulha, Maria E; Bennett, James P

    2008-11-01

    We evaluated the changes of some soil microbiological characteristics due to the use of transgenic maize expressing Bacillus thuringiensis (Bt) toxin. A two-year field experiment was conducted (2003 and 2004). Two lines of transgenic Bt maize that express the Cry1Ab protein (event 176 and MON 810) and their near-isogenic non-Bt lines were used. Rhizosphere and non-rhizosphere soils were collected and measurements were performed during the maize cultural cycle and immediately at pre-harvest. Key soil microbiological parameters measured included the numbers of culturable aerobic bacteria, including actinomycetes, and fungi, the activity of dehydrogenase and nitrogenase enzymes and ATP content. There were clear seasonal effects in the microbial parameters as evidenced by the consistent changes in sampling dates across the two years. Differences in the measured variables were also observed between rhizosphere and non-rhizosphere soils. However, under our field conditions, the presence of Bt maize did not cause, in a general way, changes in the microbial populations of the soil or in the activity of the microbial community. PMID:18656246

  4. A novel delta-endotoxin gene cryIM from Bacillus thuringiensis ssp. wuhanensis.

    PubMed

    Shevelev, A B; Kogan YaN; Bushueva, A M; Voronina, E J; Rebrikov, D V; Novikova, S I; Chestukhina, G G; Kuvshinov, V; Pehu, E; Stepanov, V M

    1997-03-10

    A new cryI-related sequence designated cryIM was cloned using an immunoscreening technique from ssp. wuhanensis of Bacillus thuringiensis (BT), previously reported to produce multiple Cry proteins [Chestukhina et al. (1994) Can. J. Microbiol. 240, 1026-1034]. Analysis of the cryIM nucleotide sequence revealed an ORF, BTII-type promoter-like sequence, peculiar for such genes, a translation initiation element and a putative transcription terminator. Nevertheless, its product was not previously found in the crystals of the host strain [Chestukhina et al. (1994) Can. J. Microbiol. 240, 1026-1034] which shows its weak or absent natural expression. The amino acid sequence of 1151 residues encoded by the continuous reading frame of cryIM is not identical but is essentially similar to the other delta-endotoxins of the CryI class. An IS231-like sequence was found 400 bp downstream of the cryIM stop codon and a fragment of the cryIAb gene was located 3 kb upstream of its initiator codon in the same orientation. Artificial expression of the cloned gene in E. coli under the control of the lacZ promoter allowed us to obtain its hypothetical protein product. PMID:9119053

  5. Proteomic analysis of Bacillus thuringiensis strain 4.0718 at different growth phases.

    PubMed

    Li, Xiaohui; Ding, Xuezhi; Xia, Liqiu; Sun, Yunjun; Yuan, Can; Yin, Jia

    2012-01-01

    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. PMID:22649324

  6. Genetic resistance to Bacillus thuringiensis alters feeding behaviour in the cabbage looper, Trichoplusia ni.

    PubMed

    Shikano, Ikkei; Cory, Jenny S

    2014-01-01

    Evolved resistance to xenobiotics and parasites is often associated with fitness costs when the selection pressure is absent. Resistance to the widely used microbial insecticide Bacillus thuringiensis (Bt) has evolved in several insect species through the modification of insect midgut binding sites for Bt toxins, and reports of costs associated with Bt resistance are common. Studies on the costs of Bt-resistance restrict the insect to a single artificial diet or host-plant. However, it is well documented that insects can self-select appropriate proportions of multiple nutritionally unbalanced foods to optimize life-history traits. Therefore, we examined whether Bt-resistant and susceptible cabbage loopers Trichoplusia ni differed in their nutrient intake and fitness costs when they were allowed to compose their own protein:carbohydrate diet. We found that Bt-resistant T. ni composed a higher ratio of protein to carbohydrate than susceptible T. ni. Bt-resistant males exhibited no fitness cost, while the fitness cost (reduced pupal weight) was present in resistant females. The absence of the fitness cost in resistant males was associated with increased carbohydrate consumption compared to females. We demonstrate a sex difference in a fitness cost and a new behavioural outcome associated with Bt resistance. PMID:24465656

  7. Bacillus thuringiensis fermentation of hydrolyzed sludge--rheology and formulation studies.

    PubMed

    Brar, Satinder K; Verma, M; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2007-03-01

    Rheology of Bacillus thuringiensis fermentation of hydrolyzed sludge was investigated in bench scale fermenter. Stable liquid formulations were developed and optimized for two-year based studies comprising various physical/chemical (viscosity, particle size, corrosion and suspendibility) and biological (microbial contamination, viable spores and entomotoxicity) parameters at different pHs and temperatures. The hydrolyzed sludge depicted non-Newtonian and pseudoplastic behaviour during fermentation with 90% to 96% confidence of fits into Casson, Power and IPC paste models. Higher values of consistency and flow index during exponential growth and stationary phase, respectively, affected downstream processing. The power law was also followed by stable formulations. Sorbitol, sodium monophosphate and sodium metabisulfite (2.2:1:1) as suspending agents produced suspendibility ranging from 69% to 94%. The stable formulation (FH-4) comprising sorbitol, sodium monophosphate and sodium metabisulfite deteriorated at pHs 6, 6.5 and temperatures, 40 and 50 degrees C, with no signs of corrosion and microbial contamination. The viscosity of FH-4 formulations decreased with shear rate which could improve handling and consequent spraying. PMID:17184817

  8. Photostabilization of Bacillus thuringiensis fermented wastewater and wastewater sludge based biopesticides using additives.

    PubMed

    Adjalle, K D; Brar, S K; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2009-07-01

    Photoprotection (against UV-A and UV-B radiations) of the active components of Bacillus thuringiensis var. kurstaki obtained from the fermentation of various culture media was investigated. The culture media comprised: starch industry wastewater; secondary wastewater sludge (non-hydrolyzed and hydrolyzed) and soya (used as a reference). Photoprotection was carried out by using various UV-protection additives, namely, para-aminobenzoic acid, lignosulfonic acid and molasses at different concentrations (0.1%, 0.15% and 0.2%, w/w). In the absence of UV-protection agents, secondary sludge demonstrated natural UV protection with half-lives ranging from 3.25 to 3.4 d. The half-life for soya and starch industry wastewater was 1.9 and 1.8 d, respectively. Para-amino-benzoic acid as a UV-protection agent at 0.20% (w/w) gave excellent UV-protection for soya and starch industry wastewater with half-lives being 5.9 and 7 d, respectively. Likewise, lignosulfonic acid at 0.20% (w/w) was an effective photostabilizer for hydrolyzed and non-hydrolyzed secondary sludge with half-lives of 7.25 and 8 d, respectively. Hence, when similar concentration of the UV-protection additives was used, photoprotection was higher for the alternative media than the conventional soya medium, validating the technical feasibility of using three additives. PMID:19100704

  9. Efficient centrifugal recovery of Bacillus thuringiensis biopesticides from fermented wastewater and wastewater sludge.

    PubMed

    Brar, Satinder K; Verma, M; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2006-03-01

    Studies were conducted on harvesting of Bacillus thuringiensis (Bt)-based biopesticides from fermented broths of starch industry wastewater (SIW), wastewater sludge (raw and hydrolyzed-NH and TH, respectively) and semi-synthetic soyameal to enhance entomotoxicity (Tx) by centrifugation. Pertinent factors influencing Tx, solids concentration, pH, temperature and centrifugal force were investigated. The centrifugate solids concentration beyond 100 g/l did not enhance Tx, instead caused pellet formation. Centrifugation efficiency (Tx recovery) was higher at pH 4, and temperature 20 degrees C for starch wastewater (98%), wastewater sludge (98% and 97.8% for non-hydrolyzed and hydrolyzed, respectively) and soya broth (83%). For maximum Tx recovery (SIW-95%; NH-90%; TH-98% and soya-78%), the centrifugal force and time required was 48,000 g and 30 min, respectively. Losses in recovery efficiency were lower for SIW and wastewater sludge in comparison to soya on adopting commercially recommended centrifugal force of 9000 g. The settling velocity computations for different fermented broths enabled calculation of Sigma factor for continuous commercial centrifuge of a given capacity and hence simulation of power requirements. It was established that power requirements for a given Tx recovery efficiency were highest for conventional medium (soya) in comparison to other waste-based fermented broths. PMID:16515801

  10. Bacillus thuringiensis fermentation of wastewater and wastewater sludge--presence and characterization of chitinases.

    PubMed

    Brar, S K; Verma, M; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2008-02-01

    This study investigated the presence of chitinases in Bacillus thuringiensis ssp kurstaki HD-1 (Bt) fermented broths of wastewater sludge (non-hydrolyzed and hydrolyzed); starch industry wastewater and soyameal. Chitinase activity was absent in soyameal and present in others. Chitinase demonstrated peaks at pH 4.0 and temperatures 40 and 50 degrees C with higher activity between pH 4-5 and 10-11. The chitinase band on SDS-PAGE was found to be between 36 and 45 kDa for non-hydrolyzed (NH) and hydrolyzed sludge (TH) and starch industry wastewater. The chitinase profile during fermentation showed peaks at 15 and 30 h for non-hydrolyzed and hydrolyzed sludge and 15 and 24 h for starch industry wastewater. Chitinase retained 96-99 % activity after two weeks incubation at room temperature and pH 4. Bioassays with supplementation of Bt chitinases showed 1.2 fold increase in entomotoxicity of wastewater sludge and a small increase in starch industry wastewater. This study sheds light on production of Bt chitinases in alternative media which will have a long term effect on entomotoxicity of these formulations. PMID:18613615

  11. Entomotoxicity, protease and chitinase activity of Bacillus thuringiensis fermented wastewater sludge with a high solids content.

    PubMed

    Brar, Satinder K; Verma, M; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2009-10-01

    This study investigated the production of biopesticides, protease and chitinase activity by Bacillus thuringiensis grown in raw wastewater sludge at high solids concentration (30 g/L). The rheology of wastewater sludge was modified with addition of Tween-80 (0.2% v/v). This addition resulted in 1.6 and 1.3-fold increase in cell and spore count, respectively. The maximum specific growth rate (micro(max)) augmented from 0.17 to 0.22 h(-1) and entomotoxicity (Tx) increased by 29.7%. Meanwhile, volumetric mass transfer coefficient (k(L)a) showed marked variations during fermentation, and oxygen uptake rate (OUR) increased 2-fold. The proteolytic activity increased while chitinase decreased for Tween amended wastewater sludge, but the entomotoxicity increased. The specific entomotoxicity followed power law when plotted against spore concentration and the relation between Tx and protease activity was linear. The viscosity varied and volume percent of particles increased in Tween-80 amended wastewater sludge and particle size (D(50)) decreased at the end of fermentation. Thus, there was an increase in entomotoxicity at higher suspended solids (30 g/L) as Tween addition improved rheology (viscosity, particle size, surface tension); enhanced maximum growth rate and OUR. PMID:19447031

  12. Batch and fed-batch fermentation of Bacillus thuringiensis using starch industry wastewater as fermentation substrate.

    PubMed

    Vu, Khanh Dang; Tyagi, Rajeshwar Dayal; Valéro, José R; Surampalli, Rao Y

    2010-08-01

    Bacillus thuringiensis var. kurstaki biopesticide was produced in batch and fed-batch fermentation modes using starch industry wastewater as sole substrate. Fed-batch fermentation with two intermittent feeds (at 10 and 20 h) during the fermentation of 72 h gave the maximum delta-endotoxin concentration (1,672.6 mg/L) and entomotoxicity (Tx) (18.5 x 10(6) SBU/mL) in fermented broth which were significantly higher than maximum delta-endotoxin concentration (511.0 mg/L) and Tx (15.8 x 10(6) SBU/mL) obtained in batch process. However, fed-batch fermentation with three intermittent feeds (at 10, 20 and 34 h) of the fermentation resulted in the formation of asporogenous variant (Spo-) from 36 h to the end of fermentation (72 h) which resulted in a significant decrease in spore and delta-endotoxin concentration and finally the Tx value. Tx of suspended pellets (27.4 x 10(6) SBU/mL) obtained in fed-batch fermentation with two feeds was the highest value as compared to other cases. PMID:19888605

  13. Determination of spore concentration in Bacillus thuringiensis through the analysis of dipicolinate by capillary zone electrophoresis.

    PubMed

    He, Jin; Luo, Xiaofeng; Chen, Shouwen; Cao, Lili; Sun, Ming; Yu, Ziniu

    2003-04-25

    A new capillary zone electrophoresis (CZE) method for the analysis of dipicolinic acid, a specific component found in spores but not in vegetative cells, was used to determine spore concentration in Bacillus thuringiensis according to the relationship between the spore concentration and the content of dipicolinate. The quantitative relationship was established by using purified spores. Electrolyte conditions that affected the separation efficiency of dipicolinate and the reproducibility were investigated. With 10 mM phosphate, 10 mM ethylenediaminetetraacetic acid and 0.25 mM tetradecyltrimethylammonium bromide at pH 6.2 as the carrier electrolyte, dipicolinate can be determined within 8 min at an applied voltage of -25 kV (anode at detector) and a capillary temperature of 25 degrees C. The method has a high separation efficiency with which the number of theoretical plates is above 300,000 plates m(-1). The relative standard deviations for migration time and peak area are less than 0.5% and 2.0%, respectively. The detection limit for dipicolinate was 10 ng ml(-1), which corresponds to 7.2 x 10(5) spores ml(-1). The method was used to determine spores in fermentation broths, and the results obtained agreed well with the values obtained by plate counting. PMID:12779231

  14. Kinetics of Bacillus thuringiensis var. israelensis growth on high glucose concentrations.

    PubMed

    Berbert-Molina, M A; Prata, A M R; Pessanha, L G; Silveira, M M

    2008-11-01

    The kinetic and general growth features of Bacillus thuringiensis var. israelensis were evaluated. Initial glucose concentration (S0) in fermentation media varied from 10 to 152 g/l. The results afforded to characterize four morphologically and physiologically well-defined culture phases, independent of S0 values: Phase I, vegetative growth; Phase II, transition to sporulation; Phase III, sporulation; and Phase IV, spores maturation and cell lysis. Important process parameters were also determined. The maximum specific growth rates (microX,m) were not affected with S0 up to 75 g/l (1.0-1.1 per hour), but higher glucose concentrations resulted in growth inhibition by substrate, revealed by a reduction in microX,m values. These higher S0 values led to longer Phases III and IV and delayed sporulation. Similar biomass concentrations (Xm=15.2-15.9 g/l) were achieved with S0 over 30.8 g/l, with increasing residual substrate, suggesting a limitation in some other nutrients and the use of glucose to form other metabolites. In this case, with S0 from 30.8 to 152 g/l, cell yield (YX/S) decreased from 0.58 to 0.41 g/g. On the other hand, with S0=10 g/l growth was limited by substrate, and YX/S has shown its maximum value (0.83 g/g). PMID:18712542

  15. Bioconversion of industrial wastewater and wastewater sludge into Bacillus thuringiensis based biopesticides in pilot fermentor.

    PubMed

    Yezza, A; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2006-10-01

    Starch industry wastewater (SWW), slaughterhouse wastewater (SHWW) and secondary sludges from three different wastewater treatment plants (Jonquière--JQS, Communauté Urbaine de Québec--CUQS and Black lake-BLS) were used as raw materials for the production of Bacillus thuringiensis (Bt) based biopesticides in a pilot scale fermentor (100 L working volume). The slaughterhouse wastewater exhibited the lowest Bt growth and entomotoxcity (Tx) potential (measured against spruce budworm) due to low availability of carbon, nitrogen and other nutrients. Performance variation (growth, sporulation, proteolytic activity and Tx potential) within the three types of sludges was directly related to the availability of nitrogen and carbohydrates, which could change with sludge origin and methods employed for its generation. The Tx potential of Bt obtained in different secondary sludges (JQS: 12 x 10(9) SBU/L; CUQS: 13 x 10(9) SBU/L and BLS: 16 x 10(9) SBU/L) and SWW (18 x 10(9) SBU/L) was higher than the soybean based synthetic medium (10 x 10(9) SBU/L). The maximum protease activity was obtained in CUQ secondary sludge (4.1 IU/mL) due to its high complex protein concentration. Nevertheless, high carbohydrate concentration in SWW repressed enzyme production. The secondary sludges and SWW were found to be suitable raw materials for high potency Bt biopesticide production. PMID:16242319

  16. Division of labour and terminal differentiation in a novel Bacillus thuringiensis strain

    PubMed Central

    Deng, Chao; Slamti, Leyla; Raymond, Ben; Liu, Guiming; Lemy, Christelle; Gominet, Myriam; Yang, Jingni; Wang, Hengliang; Peng, Qi; Zhang, Jie; Lereclus, Didier; Song, Fuping

    2015-01-01

    A major challenge in bacterial developmental biology has been to understand the mechanisms underlying cell fate decisions. Some differentiated cell types display cooperative behaviour. Cooperation is one of the greatest mysteries of evolutionary biology and microbes have been considered as an excellent system for experimentally testing evolution theories. Bacillus thuringiensis (Bt) is a spore-forming bacterium, which is genetically closely related to B. anthracis, the agent of anthrax, and to B. cereus, an opportunistic human pathogen. The defining feature that distinguishes Bt from its relatives is its ability to produce crystal inclusions in the sporulating cells. These toxins are solubilized after ingestion and are cooperative public goods in insect hosts. In this study, we describe a Bt strain LM1212 that presents the unique ability to terminally differentiate into crystal producers and spore formers. Transcriptional analysis based on lacZ and gfp reporter genes suggested that this phenotype is the consequence of a new type of cell differentiation associated with a novel regulation mode of cry gene expression. The differentiating crystal-producer phenotype has higher spore productivity than a typical Bt strain and is better able to compete with Cry toxin null ‘cheaters'. Potentially, this division of labour provides additional fitness benefits in terms of spore viability or durability of Cry toxin. PMID:25083932

  17. Wastewater treatment sludge as a raw material for the production of Bacillus thuringiensis based biopesticides.

    PubMed

    Montiel, M D; Tyagi, R D; Valero, J R

    2001-11-01

    Seven wastewater sludges of different origins and types were used as an alternate culture medium for producing Bacillus thuringiensis variety kurstaki HD-1. The sludge samples were used under three different preparations: without pre-treatment, with acid treatment (hydrolysed sludge) and the supernatant obtained after centrifugation of the hydrolysed sludge. The sludge composition varied widely with origin and the type of sludge. Growth and sporulation were evaluated by the total viable cell count and spore count of the preparations. Growth, sporulation and endotoxin production were affected by the sludge origin. Hydrolysed sludge gave the highest viable cell and spore counts while the liquid phase (supernatant) gave the lowest. Non-hydrolysed primary sludge from Valcartier was unable to sustain bacterial growth because of its low pH. Bioassays were conducted against larvae of spruce budworm to evaluate entomotoxic potential of the preparations obtained. In general, sludge hydrolysis increased the entomotoxicity yields. Similar entomotoxicity was observed in Black Lake secondary sludge (4100 IU/microL) as that obtained in the reference soya medium (3800 IU/microL). The use of the sludge supernatant (liquid phase) was not recommended due to the low entomotoxic potential obtained. PMID:12230163

  18. Novel actin filaments from Bacillus thuringiensis form nanotubules for plasmid DNA segregation

    PubMed Central

    Jiang, Shimin; Narita, Akihiro; Popp, David; Ghoshdastider, Umesh; Lee, Lin Jie; Srinivasan, Ramanujam; Balasubramanian, Mohan K.; Oda, Toshiro; Koh, Fujiet; Larsson, Mårten; Robinson, Robert C.

    2016-01-01

    Here we report the discovery of a bacterial DNA-segregating actin-like protein (BtParM) from Bacillus thuringiensis, which forms novel antiparallel, two-stranded, supercoiled, nonpolar helical filaments, as determined by electron microscopy. The BtParM filament features of supercoiling and forming antiparallel double-strands are unique within the actin fold superfamily, and entirely different to the straight, double-stranded, polar helical filaments of all other known ParMs and of eukaryotic F-actin. The BtParM polymers show dynamic assembly and subsequent disassembly in the presence of ATP. BtParR, the DNA-BtParM linking protein, stimulated ATP hydrolysis/phosphate release by BtParM and paired two supercoiled BtParM filaments to form a cylinder, comprised of four strands with inner and outer diameters of 57 Å and 145 Å, respectively. Thus, in this prokaryote, the actin fold has evolved to produce a filament system with comparable features to the eukaryotic chromosome-segregating microtubule. PMID:26873105

  19. Bacillus thuringiensis peptidoglycan hydrolase SleB171 involved in daughter cell separation during cell division.

    PubMed

    Li, Hua; Hu, Penggao; Zhao, Xiuyun; Yu, Ziniu; Li, Lin

    2016-04-01

    Whole-genome analyses have revealed a putative cell wall hydrolase gene (sleB171) that constitutes an operon with two other genes (ypeBandyhcN) of unknown function inBacillus thuringiensisBMB171. The putative SleB171 protein consists of 259 amino acids and has a molecular weight of 28.3 kDa. Gene disruption ofsleB171in the BMB171 genome causes the formation of long cell chains during the vegetative growth phase and delays spore formation and spore release, although it has no significant effect on cell growth and the ultimate release of the spores. The inseparable vegetative cells were nearly restored through the complementation ofsleB171expression. Real-time quantitative polymerase chain reaction analysis revealed thatsleB171is mainly active in the vegetative growth phase, with a maximum activity at the early stationary growth phase. Western blot analysis also confirmed thatsleB171is preferentially expressed during the vegetative growth phase. These results demonstrated that SleB171 plays an essential role in the daughter cell separation during cell division. PMID:26922318

  20. Effect of resistance to Bacillus thuringiensis cotton on pink bollworm (Lepidoptera: Gelechiidae) response to sex pheromone.

    PubMed

    Carrière, Yves; Nyboer, Megan E; Ellers-Kirk, Christa; Sollome, James; Colletto, Nick; Antilla, Larry; Dennehy, Timothy J; Staten, Robert T; Tabashnik, Bruce E

    2006-06-01

    Fitness costs associated with resistance to transgenic crops producing toxins from Bacillus thuringiensis (Bt) could reduce male response to pheromone traps. Such costs would cause underestimation of resistance frequency if monitoring was based on analysis of males caught in pheromone traps. To develop a DNA-based resistance monitoring program for pink bollworm, Pectinophora gossypiella (Saunders) (Lepidoptera: Gelechiidae), we compared the response to pheromone traps of males with and without cadherin alleles associated with resistance to Bt cotton (Gossypium hirsutum L.). When irradiated males from two hybrid laboratory strains with an intermediate frequency of resistance alleles were released in large field cages, the probability of capture in pheromone traps was not lower for males with resistance alleles than for males without resistance alleles. These results suggest that analysis of trapped males would not underestimate the frequency of resistance. As the time males spent in traps in the field increased from 3 to 15 d, the success of DNA amplification declined from 100 to 30%. Thus, the efficiency of a DNA-based resistance monitoring program would be improved by analyzing males remaining in traps for 3 d or less. PMID:16813335

  1. Effect of Insect Larval Midgut Proteases on the Activity of Bacillus thuringiensis Cry Toxins▿

    PubMed Central

    Fortier, Mélanie; Vachon, Vincent; Frutos, Roger; Schwartz, Jean-Louis; Laprade, Raynald

    2007-01-01

    To test the possibility that proteolytic cleavage by midgut juice enzymes could enhance or inhibit the activity of Bacillus thuringiensis insecticidal toxins, once activated, the effects of different toxins on the membrane potential of the epithelial cells of isolated Manduca sexta midguts in the presence and absence of midgut juice were measured. While midgut juice had little effect on the activity of Cry1Aa, Cry1Ac, Cry1Ca, Cry1Ea, and R233A, a mutant of Cry1Aa from which one of the four salt bridges linking domains I and II of the toxin was eliminated, it greatly increased the activity of Cry1Ab. In addition, when tested in the presence of a cocktail of protease inhibitors or when boiled, midgut juice retained almost completely its capacity to enhance Cry1Ab activity, suggesting that proteases were not responsible for the stimulation. On the other hand, in the absence of midgut juice, the cocktail of protease inhibitors also enhanced the activity of Cry1Ab, suggesting that proteolytic cleavage by membrane proteases could render the toxin less effective. The lower toxicity of R233A, despite a similar in vitro pore-forming ability, compared with Cry1Aa, cannot be accounted for by an increased susceptibility to midgut proteases. Although these assays were performed under conditions approaching those found in the larval midgut, the depolarizing activities of the toxins correlated only partially with their toxicities. PMID:17693568

  2. Identification of Bombyx mori midgut receptor for Bacillus thuringiensis insecticidal CryIA(a) toxin.

    PubMed

    Nagamatsu, Y; Toda, S; Yamaguchi, F; Ogo, M; Kogure, M; Nakamura, M; Shibata, Y; Katsumoto, T

    1998-04-01

    As part of a study of the mechanism by which Bacillus thuringiensis insecticidal crystal protein acts, a Bombyx mori receptor to the CryIA(a) toxin specific for lepidopterans was examined. Histological examination showed that the toxin acted on the brush-border membrane of the midgut columnar cells and broke its infolding structure, causing cell lysis. The membrane vesicles were purified, and a 175-kDa protein binding the toxin was found that accounted for some 0.015% of membrane proteins. The protein, designated BtR175, was a glycoprotein that reacted with concanavalin A. Anti-BtR antibodies inhibited the binding of toxin to membrane vesicles in vitro and decreased the effect of the toxin to silkworms in vivo. BtR175, although found in the gut, was not found in fat bodies, integument, or silk glands. These results indicated that BtR175 was the receptor protein for the insecticidal toxin. Proteins (137 and 107 kDa) binding the CryIA(a) toxin also were found in the gut membranes of Tenebrio moritor larvae, a coleopteran not sensitive to the toxin. The specificity of the toxin could not be explained only in term of the existence of its binding protein. PMID:9614702

  3. Optimization of photobioreactor growth conditions for a cyanobacterium expressing mosquitocidal Bacillus thuringiensis Cry proteins.

    PubMed

    Ketseoglou, Irene; Bouwer, Gustav

    2013-08-10

    An Anabaena strain (PCC 7120#11) that was genetically engineered to express Bacillus thuringiensis subsp. israelensis cry genes has shown good larvicidal activity against Anopheles arabiensis, a major vector of malaria in Africa. Response surface methodology was used to evaluate the relationship between key growth factors and the volumetric productivity of PCC 7120#11 in an indoor, flat-plate photobioreactor. The interaction of input CO₂ concentration and airflow rate had a statistically significant effect on the volumetric productivity of PCC 7120#11, as did the interaction of airflow rate and photosynthetic photon flux density. Model-based numerical optimization indicated that the optimal factor level combination for maximizing PCC 7120#11 volumetric productivity was a photosynthetic photon flux density of 154 μmol m⁻² s⁻¹ and air enriched with 3.18% (v/v) CO₂ supplied at a flow rate of 1.02 vessel volumes per minute. At the levels evaluated in the study, none of the growth factors had a significant effect on the median lethal concentration of PCC 7120#11 against An. arabiensis larvae. This finding is important because loss of mosquitocidal activity under growth conditions that maximize volumetric productivity would impact on the feasibility of using PCC 7120#11 in malaria vector control programs. The study showed the usefulness of response surface methodology for determination of the optimal growth conditions for a cyanobacterium that is genetically engineered to have larvicidal activity against malaria vectors. PMID:23732832

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

    PubMed Central

    Hurley, J M; Bulla, L A; Andrews, R E

    1987-01-01

    Two proteins from parasporal crystals of Bacillus thuringiensis subsp. israelensis were purified to electrophoretic homogeneity by gel filtration and anion-exchange chromatography. The larger of the two proteins (molecular weight, 68,000) was not cytolytic, whereas the smaller protein (molecular weight, 28,000) was highly cytolytic when assayed against rat erythrocytes. When these proteins were assayed against larvae of the yellow fever mosquito, Aedes aegypti, the larger protein was at least 100-fold more toxic than the smaller protein. Although proteolytic activity was not detected in solubilized crystals nor in purified protein preparations, the toxin (molecular weight, 68,000) was readily degraded to smaller, nontoxic molecules, even when maintained at 4 degrees C. Mixtures of the two purified proteins were significantly more toxic to mosquito larvae than was either protein alone. Thus, it is likely that both the mosquitocidal and the cytolytic protein play roles in the overall insecticidal action of the parasporal crystal produced by this bacterium. Images PMID:3606108

  5. Construction of an environmental safe Bacillus thuringiensis engineered strain against Coleoptera.

    PubMed

    Yu, Yajun; Yuan, Yihui; Gao, Meiying

    2016-05-01

    Cloning of new toxic genes from Bacillus thuringiensis (Bt) and construction of Bt engineered strains are two key strategies for bio-control of coleopteran pests in agriculture and forestry. In this study, we cloned a new cry3Aa-type gene, cry3Aa8, from wild Bt strain YC-03 against coleopteran, and constructed a Bt engineered strain, ACE-38, containing insecticidal protein-encoding gene cry3Aa8. The engineered strain, with almost four times of Cry3Aa yield compared with strain YC-03, was an antibiotic marker-free strain. Though no selective pressure was presented in the medium, cry3Aa8 in the engineered strain ACE-38 remained stable. The yield of Cry3Aa by strain ACE-38 reached 2.09 mg/ml in the optimized fermentation medium. The activity of strain ACE-38 against Plagiodera versicolora was tested, and the LC50 of ACE-38 cultures in the optimized fermentation medium was 1.13 μl/ml. Strain ACE-38 is a non-antibiotic Bt engineered strain with high Chrysomelidae toxicity and exhibits good fermentation property. The modified indigenous site-specific recombination system constructed in this study might be useful for the construction of Bt engineered strains containing genes that cannot be expressed in the indigenous site-specific recombination system using plasmid pBMB1205R. PMID:26767987

  6. The impact of strain diversity and mixed infections on the evolution of resistance to Bacillus thuringiensis

    PubMed Central

    Raymond, Ben; Wright, Denis J.; Crickmore, Neil; Bonsall, Michael B.

    2013-01-01

    Pesticide mixtures can reduce the rate at which insects evolve pesticide resistance. However, with live biopesticides such as the naturally abundant pathogen Bacillus thuringiensis (Bt), a range of additional biological considerations might affect the evolution of resistance. These can include ecological interactions in mixed infections, the different rates of transmission post-application and the impact of the native biodiversity on the frequency of mixed infections. Using multi-generation selection experiments, we tested how applications of single and mixed strains of Bt from diverse sources (natural isolates and biopesticides) affected the evolution of resistance in the diamondback moth, Plutella xylostella, to a focal strain. There was no significant difference in the rate of evolution of resistance between single and mixed-strain applications although the latter did result in lower insect populations. The relative survivorship of Bt-resistant genotypes was higher in the mixed-strain treatment, in part owing to elevated mortality of susceptible larvae in mixtures. Resistance evolved more quickly with treatments that contained natural isolates, and biological differences in transmission rate may have contributed to this. Our data indicate that the use of mixtures can have unexpected consequences on the fitness of resistant and susceptible insects. PMID:24004937

  7. Strategies to improve the insecticidal activity of Cry toxins from Bacillus thuringiensis.

    PubMed Central

    Pardo-López, L.; Muñoz-Garay, C.; Porta, H.; Rodríguez-Almazán, C.; Soberón, M.; Bravo, A.

    2009-01-01

    Bacillus thuringiensis Cry toxins have been widely used in the control of insect pests either as spray products or expressed in transgenic crops. These proteins are pore forming toxins with a complex mechanism of action that involves the sequential interaction with several toxin-receptors. Cry toxins are specific against susceptible larvae and although they are often highly effective, some insect pests are not affected by them or show low susceptibility. In addition, the development of resistance threatens their effectiveness, so strategies to cope with all these problems are necessary. In this review we will discuss and compare the different strategies that have been used to improve insecticidal activity of Cry toxins. The activity of Cry toxins can be enhanced by using additional proteins in the bioassay like serine protease inhibitors, chitinases, Cyt toxins, or a fragment of cadherin receptor containing a toxin-binding site. On the other hand, different modifications performed in the toxin gene such as site directed mutagenesis, introduction of cleavage sites in specific regions of the protein, and deletion of small fragments from the amino-terminal region lead to improved toxicity or overcome resistance, representing interesting alternatives for insect pest control. PMID:18773932

  8. Persistence of detectable insecticidal proteins from Bacillus thuringiensis (Cry) and toxicity after adsorption on contrasting soils.

    PubMed

    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. PMID:26549751

  9. Alteration in Bacillus thuringiensis toxicity by curing gut flora: novel approach for mosquito resistance management.

    PubMed

    Patil, Chandrashekhar D; Borase, Hemant P; Salunke, Bipinchandra K; Patil, Satish V

    2013-09-01

    Mosquitoes are known for acquiring resistance against insecticides in many ways, namely target side mutation, enzyme modification, sequestration, quick elimination, etc. But, the role of microflora present in abundance in the larval midgut is less explored with respect to their role in insecticide resistance. During the course of their development, mosquitoes are continuously exposed to microbes and have naturally acquired midgut microbial flora. This midgut flora can modulate the mosquito's susceptibility to Bacillus thuringiensis (Bt) infection by degrading toxic Bt protein forms through an unknown mechanism. In this study, we show that microbe-free aseptic mosquito larvae displayed an increased susceptibility to Bt toxicity compared to larvae harboring natural microbial flora. Fourth instar larvae of Anopheles stephensi were treated separately with penicillin, streptomycin, erythromycin (100 μg/ml), and mixtures of all three antibiotics and then analyzed for Bt toxicity. We have also examined the influence of the mosquito's midgut microbial flora under microaerophilic condition on the Bt protein degradation through plate, broth, TLC, and UV-vis spectrophotometric assay. A better understanding of the roles of microbiota in preventing Bt toxicity to mosquitoes could potentially lead to the development of new sustainable mosquito control strategies. PMID:23820604

  10. Occurrence, characterization and insecticidal activity of Bacillus thuringiensis strains isolated from argan fields in Morocco.

    PubMed

    Aboussaid, H; Vidal-Quist, J C; Oufdou, K; El Messoussi, S; Castañera, P; González-Cabrera, J

    2011-01-01

    Soils collected from five locations in the argan forest (an endemic plant) in Morocco were used to form the first collection of Bacillus thuringiensis (Bt) strains from this area (58 strains). Here we found that the argan forest is a major source of Bt, as 90.62% of the samples contained Bt strains. These strains produced mainly spherical or irregular crystals that in some cases remained adhered to the spore after cell lysis. There was no strain producing bipyramidal crystals, suggesting the absence of strains bearing crv1 genes. This was confirmed by PCR analysis using eight primer pairs that can potentially detect 13 different groups of cry and cyt genes. Strains containing cry7/8 were the most abundant (25.53%), followed by strains harbouring cry9A (14.89%), cry11 (8.51%) and cry4 (4.25%). The mixtures of spores and crystals as well as culture supernatants were assayed for toxicity towards Ceratitis capitata (Medfly), showing up to 30% mortality. Our findings suggest that the argan region is a suitable target for future and wider screening programmes looking for strains bearing toxins or combinations of them to develop more efficient Bt-based formulates. PMID:21970180

  11. Contamination of refuges by Bacillus thuringiensis toxin genes from transgenic maize.

    PubMed

    Chilcutt, Charles F; Tabashnik, Bruce E

    2004-05-18

    Transgenic crops producing insecticidal toxins from Bacillus thuringiensis (Bt) are widely used to control pests, but their benefits will be lost if pests evolve resistance. The mandated high-dose/refuge strategy for delaying pest resistance requires planting refuges of toxin-free crops near Bt crops to promote survival of susceptible pests. We report that pollen-mediated gene flow up to 31 m from Bt maize caused low to moderate Bt toxin levels in kernels of non-Bt maize refuge plants. Immunoassays of non-Bt maize sampled from the field showed that the mean concentration of Bt toxin Cry1Ab in kernels and the percentage of kernels with Cry1Ab decreased with distance from Bt maize. The highest Bt toxin concentration in pooled kernels of non-Bt maize plants was 45% of the mean concentration in kernels from adjacent Bt maize plants. Most previous work on gene flow from transgenic crops has emphasized potential effects of transgene movement on wild relatives of crops, landraces, and organic plantings, whereas implications for pest resistance have been largely ignored. Variable Bt toxin production in seeds of refuge plants undermines the high-dose/refuge strategy and could accelerate pest resistance to Bt crops. Thus, guidelines should be revised to reduce gene flow between Bt crops and refuge plants. PMID:15136739

  12. No Adjuvant Effect of Bacillus thuringiensis-Maize on Allergic Responses in Mice

    PubMed Central

    Dekan, Gerhard; Epstein, Michelle M.

    2014-01-01

    Genetically modified (GM) foods are evaluated carefully for their ability to induce allergic disease. However, few studies have tested the capacity of a GM food to act as an adjuvant, i.e. influencing allergic responses to other unrelated allergens at acute onset and in individuals with pre-existing allergy. We sought to evaluate the effect of short-term feeding of GM Bacillus thuringiensis (Bt)-maize (MON810) on the initiation and relapse of allergic asthma in mice. BALB/c mice were provided a diet containing 33% GM or non-GM maize for up to 34 days either before ovalbumin (OVA)-induced experimental allergic asthma or disease relapse in mice with pre-existing allergy. We observed that GM-maize feeding did not affect OVA-induced eosinophilic airway and lung inflammation, mucus hypersecretion or OVA-specific antibody production at initiation or relapse of allergic asthma. There was no adjuvant effect upon GM-maize consumption on the onset or severity of allergic responses in a mouse model of allergic asthma. PMID:25084284

  13. Characterization of Bacillus thuringiensis soil isolates from Cuba, with insecticidal activity against mosquitoes.

    PubMed

    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. PMID:22017108

  14. Transgenic organisms expressing genes from Bacillus thuringiensis to combat insect pests.

    PubMed

    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. PMID:21326834

  15. Sample preparation for beta-exotoxin determination in Bacillus thuringiensis cultures by reversed-phase high-performance liquid chromatography.

    PubMed

    Gohar, M; Perchat, S

    2001-11-01

    Beta-exotoxin is a nucleotide analogue produced by the entomopathogenic bacterium Bacillus thuringiensis. We have defined two new HPLC procedures for quantification of this exotoxin in culture supernatants of B. thuringiensis grown in poor or rich medium. The sample is prepared either by precipitation in solvent or by solid-phase extraction. Solvent precipitation is achieved treating the sample with acetone and acetonitrile. Solid-phase extraction is performed with a C18 and an anion-exchange cartridge. Reversed-phase HPLC with gradient elution of the prepared samples gives a limit of quantitation of 2 microg/ml for samples prepared by solvent precipitation and of 0.3 microg/ml for samples prepared by solid-phase extraction. PMID:11673902

  16. Correspondence of high levels of beta-exotoxin I and the presence of cry1B in Bacillus thuringiensis.

    PubMed

    Espinasse, Sylvain; Gohar, Michel; Chaufaux, Josette; Buisson, Christophe; Perchat, Stéphane; Sanchis, Vincent

    2002-09-01

    Examination of 640 natural isolates of Bacillus thuringiensis showed that the 58 strains (9%) whose supernatants were toxic to Anthonomus grandis (Coleoptera: Curculionidae) produced between 10 and 175 micro g of beta-exotoxin I per ml. We also found that 55 (46%) of a sample of 118 strains whose culture supernatants were not toxic to A. grandis nevertheless produced between 2 and 5 micro g/ml. However, these amounts of beta-exotoxin I were below the threshold for detectable toxicity against this insect species. Secretion of large amounts of beta-exotoxin I was strongly associated with the presence of cry1B and vip2 genes in the 640 natural B. thuringiensis isolates studied. We concluded that strains carrying cry1B and vip2 genes also possess, on the same plasmid, genetic determinants necessary to promote high levels of production of beta-exotoxin I. PMID:12200263

  17. Novel Isolate of Bacillus thuringiensis subsp. thuringiensis That Produces a Quasicuboidal Crystal of Cry1Ab21 Toxic to Larvae of Trichoplusia ni▿

    PubMed Central

    Swiecicka, Izabela; Bideshi, Dennis K.; Federici, Brian A.

    2008-01-01

    A new isolate (IS5056) of Bacillus thuringiensis subsp. thuringiensis that produces a novel variant of Cry1Ab, Cry1Ab21, was isolated from soil collected in northeastern Poland. Cry1Ab21 was composed of 1,155 amino acids and had a molecular mass of 130.5 kDa, and a single copy of the gene coding for this endotoxin was located on a ∼75-kbp plasmid. When synthesized by the wild-type strain, Cry1Ab21 produced a unique, irregular, bipyramidal crystal whose long and short axes were both approximately 1 μm long, which gave it a cuboidal appearance in wet mount preparations. In diet incorporation bioassays, the 50% lethal concentrations of the crystal-spore complex were 16.9 and 29.7 μg ml−1 for second- and fourth-instar larvae of the cabbage looper, Trichoplusia ni, respectively, but the isolate was essentially nontoxic to larvae of the beet armyworm, Spodoptera exigua. A bioassay of autoclaved spore-crystal preparations showed no evidence of β-exotoxin activity, indicating that toxicity was due primarily to Cry1Ab21. Studies of the pathogenesis of isolate IS5056 in second-instar larvae of T. ni showed that after larval death the bacterium colonized and subsequently sporulated extensively throughout the cadaver, suggesting that other bacteria inhabiting the midgut lumen played little if any role in mortality. As T. ni is among the most destructive pests of vegetable crops in North America and has developed resistance to B. thuringiensis, this new isolate may have applied value. PMID:18083867

  18. Postflight analyses of Bacillus thuringiensis organisms exposed to space flight conditions

    NASA Technical Reports Server (NTRS)

    Wrenn, R. T.; Simmonds, R. C.; Heimpel, A. M.

    1973-01-01

    Cultures of B. thuringiensis returned from space flight appeared to be normal to slightly affected adversely in their ability to produce three toxins that affect insects. In addition, it can be stated that B. thuringiensis spores are very resistant to ultraviolet irradiation at the individual wavelengths and energy levels previously described. Full sunlight, however, does have a detrimental effect on the viability of B. thuringiensis spores.

  19. Response surface modeling for hot, humid air decontamination of materials contaminated with Bacillus anthracis ∆Sterne and Bacillus thuringiensis Al Hakam spores

    PubMed Central

    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

  20. Characterization of native Bacillus thuringiensis strains and selection of an isolate active against Spodoptera frugiperda and Peridroma saucia.

    PubMed

    Alvarez, Analía; Virla, Eduardo G; Pera, Licia M; Baigorí, Mario D

    2009-12-01

    Twelve Bacillus thuringiensis (Bt) strains, isolated from larvae and soil samples in Argentina, were molecularly and phenotypically characterized and their insecticidal activities against Spodoptera frugiperda and Peridroma saucia were determined. One isolate--Bt RT--produced more than 93% mortality on first instar larvae of both species, which was higher than that produced by the reference strain Bt 4D1. Bt RT carried a different cry gene profile than Bt 4D1. Scanning electron microscopy showed the presence of bipyramidal and cuboidal crystals. Phenotypic characterization revealed lytic enzymes that could contribute to Bt pathogenicity. PMID:19693442

  1. Comparison of susceptibility of Chilo suppressalis and Bombyx mori to five Bacillus thuringiensis proteins.

    PubMed

    Jiao, Yaoyu; Yang, Yan; Meissle, Michael; Peng, Yufa; Li, Yunhe

    2016-05-01

    Transformation of rice with genes encoding insecticidal Cry proteins from Bacillus thuringiensis (Bt) should confer high resistance to target lepidopteran pests, such as Chilo suppressalis, and low toxicity to non-target organisms, such as silkworm Bombyx mori. Five purified Cry proteins that have been used for plant transformation were tested using dietary exposure assays. The susceptibility of C. suppressalis larvae to the five insecticidal proteins in the decreasing order was: Cry1Ca>Cry1Ab>Cry1Ac>Cry2Aa>Cry1Fa. However, the toxicities of the Cry proteins to B. mori were in the order: Cry1Fa>Cry1Ca>Cry2Aa>Cry1Ab>Cry1Ac. The Cry1Ca, Cry1Ab and Cry1Ac proteins exhibited relatively high toxicity to C. suppressalis larvae, with EC50 values of 16.4, 45.8 and 89.6ng/g, respectively. The toxicities of the three Cry proteins to B. mori larvae were 8, 14, and 22times lower, with EC50 values of 138.3, 628.4 and 1939.2ng/g, respectively. The Cry1Fa and Cry2Aa proteins showed high toxicity to B. mori larvae, with EC50 values of 135.7 and 373.9ng/g, respectively, but low toxicity to C. suppressalis larvae, with EC50 values of 6092.1 and 1208.5ng/g, respectively. We thus conclude that Cry1Ab, Cry1Ac and Cry1Ca are appropriate for transforming rice to control lepidopteran rice pests. In contrast, Cry1Fa and Cry2Aa are not appropriate due to their high toxicity to silkworm larvae and low activity against the target pest. PMID:26994840

  2. An ABC Transporter Mutation Is Correlated with Insect Resistance to Bacillus thuringiensis Cry1Ac Toxin

    PubMed Central

    Gahan, Linda J.; Pauchet, Yannick; Vogel, Heiko; Heckel, David G.

    2010-01-01

    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. PMID:21187898

  3. Diversity and Toxicity of Bacillus thuringiensis from Shifting Cultivation (Jhum) Habitat.

    PubMed

    Zothansanga, Ralte; Senthilkumar, Nachimuthu; Gurusubramanian, Guruswami

    2016-01-01

    Bacillus thuringiensis (Bt) strains were isolated from jhum-agriculture, jhum-forest, aquatic and fallow soil samples from Mizoram by acetate selection method. Isolates were characterized for biochemical typing, cry gene and protein profiling, growth curve study and toxicity against Culex tritaeniorhynchus. Bt frequency was high in jhum-agriculture land (69.56%) whereas low in jhum-forest soils (31.57%). Bt was found to be abundant in jhum shifting cultivation soil with an index ranging between 0.010 and 0.015. Majority of the isolates from jhum soils produced oval and spherical crystals and showed eleven types of crystal proteins groups. PCR analysis revealed predominance of dipteran-active cry genes (cry4 and cry9). The variations in crystal morphology, cry genes and Cry protein (s) from the isolates of Bt revealed molecular diversity. Higher mortality, lower lethal dose, and lesser time to kill were observed in Bt isolates from jhum soils than aquatic and fallow habitats. Based on the toxicity test, SC1 and HP7 isolates containing cry 4 and cry 9 genes showed higher activity. Growth curve analysis showed significant variations among Bt isolates to reach the sporulating stage. Higher growth index and lower mean generation time were observed in SC1 and HP7 Bt isolates. Bt strains express different endotoxin genes and crystal proteins and their harvesting time also varied from strain to strain. Significant variation was found in Bt isolates from jhum habitats in relation to the cry gene composition, protein profiling and toxicity. Results from this study suggest that novel Bt entomopathogens may complement for regulating mosquito vectors. PMID:27350428

  4. Broad-spectrum resistance to Bacillus thuringiensis toxins by western corn rootworm (Diabrotica virgifera virgifera).

    PubMed

    Jakka, Siva R K; Shrestha, Ram B; Gassmann, Aaron J

    2016-01-01

    The evolution of resistance and cross-resistance threaten the sustainability of genetically engineered crops that produce insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt). Western corn rootworm, Diabrotica virgifera virgifera LeConte, is a serious pest of maize and has been managed with Bt maize since 2003. We conducted laboratory bioassays with maize hybrids producing Bt toxins Cry3Bb1, mCry3A, eCry3.1Ab, and Cry34/35Ab1, which represent all commercialized Bt toxins for management of western corn rootworm. We tested populations from fields where severe injury to Cry3Bb1 maize was observed, and populations that had never been exposed to Bt maize. Consistent with past studies, bioassays indicated that field populations were resistant to Cry3Bb1 maize and mCry3A maize, and that cross-resistance was present between these two types of Bt maize. Additionally, bioassays revealed resistance to eCry3.1Ab maize and cross-resistance among Cry3Bb1, mCry3A and eCry3.1Ab. However, no resistance or cross-resistance was detected for Cry34/35Ab1 maize. This broad-spectrum resistance illustrates the potential for insect pests to develop resistance rapidly to multiple Bt toxins when structural similarities are present among toxins, and raises concerns about the long-term durability of Bt crops for management of some insect pests. PMID:27297953

  5. Bacillus thuringiensis delta-endotoxin binding to brush border membrane vesicles of rice stem borers.

    PubMed

    Alcantara, Edwin P; Aguda, Remedios M; Curtiss, April; Dean, Donald H; Cohen, Michael B

    2004-04-01

    The receptor binding step in the molecular mode of action of five delta-endotoxins (Cry1Ab, Cry1Ac, Cry1C, Cry2A, and Cry9C) from Bacillus thuringiensis was examined to find toxins with different receptor sites in the midgut of the striped stem borer (SSB) Chilo suppressalis (Walker) and yellow stem borer (YSB) Scirpophaga incertulas (Walker) (Lepidoptera: Pyralidae). Homologous competition assays were used to estimate binding affinities (K(com)) of (125)I-labelled toxins to brush border membrane vesicles (BBMV). The SSB BBMV affinities in decreasing order was: Cry1Ab = Cry1Ac > Cry9C > Cry2A > Cry1C. In YSB, the order of decreasing affinities was: Cry1Ac > Cry1Ab > Cry9C = Cry2A > Cry1C. The number of binding sites (B(max)) estimated by homologous competition binding among the Cry toxins did not affect toxin binding affinity (K(com)) to both insect midgut BBMVs. Results of the heterologous competition binding assays suggest that Cry1Ab and Cry1Ac compete for the same binding sites in SSB and YSB. Other toxins bind with weak (Cry1C, Cry2A) or no affinity (Cry9C) to Cry1Ab and Cry1Ac binding sites in both species. Cry2A had the lowest toxicity to 10-day-old SSB and Cry1Ab and Cry1Ac were the most toxic. Taken together, the results of this study show that Cry1Ab or Cry1Ac could be combined with either Cry1C, Cry2A, or Cry9C for more durable resistance in transgenic rice. Cry1Ab should not be used together with Cry1Ac because a mutation in one receptor site could diminish binding of both toxins. PMID:15027071

  6. An ABC transporter mutation is correlated with insect resistance to Bacillus thuringiensis Cry1Ac toxin.

    PubMed

    Gahan, Linda J; Pauchet, Yannick; Vogel, Heiko; Heckel, David G

    2010-12-01

    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. PMID:21187898

  7. Heterologous expression, purification and biochemical characterization of endochitinase ChiA74 from Bacillus thuringiensis.

    PubMed

    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. PMID:25478931

  8. Field-evolved resistance by western corn rootworm to multiple Bacillus thuringiensis toxins in transgenic maize.

    PubMed

    Gassmann, Aaron J; Petzold-Maxwell, Jennifer L; Clifton, Eric H; Dunbar, Mike W; Hoffmann, Amanda M; Ingber, David A; Keweshan, Ryan S

    2014-04-01

    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. PMID:24639498

  9. Novel Bacillus thuringiensis insecticidal crystal protein with a silent activity against coleopteran larvae.

    PubMed Central

    Lambert, B; Höfte, H; Annys, K; Jansens, S; Soetaert, P; Peferoen, M

    1992-01-01

    A novel Bacillus thuringiensis crystal protein with a silent activity against the Colorado potato beetle is described. The crystal proteins are produced as bipyramidal crystals. These crystals contain a protein of 129 kDa with a trypsin-resistant core fragment of 72 kDa. Neither a spore-crystal mixture nor in vitro-solubilized crystals are toxic to any of several Lepidoptera and Coleoptera species tested. In contrast, a trypsin-treated solution containing the 72-kDa tryptic core fragment of the protoxin is highly toxic to Colorado potato beetle larvae. The crystal protein-encoding gene was cloned and sequenced. The inferred amino acid sequence of the putative toxic fragment has 37, 32, and 33% homology to the CryIIIA, CryIIIB, and CryIIID toxins, respectively. Interestingly, the 501 C-terminal amino acids show 41 to 48% amino acid identity with corresponding C-terminal amino acid sequences of other crystal proteins. Because of the toxicity of the fragment to the Colorado potato beetle and because of the distinct similarities of the toxic fragment with the other CryIII proteins, this gene was given a new subclass name (cryIIIC) within the CryIII class of coleopteran-active crystal proteins. CryIIIC represents the first example of a crystal protein with a silent activity towards coleopteran insect larvae. Natural CryIIIC crystals are not toxic. Toxicity is revealed only after an in vitro solubilization and activation step. Images PMID:1514800

  10. Biphasic fermentation is an efficient strategy for the overproduction of δ-endotoxin from Bacillus thuringiensis.

    PubMed

    Jisha, Veloorvalappil Narayanan; Smitha, Robinson Babysarojam; Priji, Prakasan; Sajith, Sreedharan; Benjamin, Sailas

    2015-02-01

    This study illustrates a biphasic solid-state fermentation (SSF) strategy for the overproduction of δ-endotoxin from Bacillus thuringiensis subsp. kurstaki (Btk) and also purification of δ-endotoxin from the solid-fermented medium. The fermentation strategy had two phases (biphasic); i.e., the first short phase was semisolid state (12 h), and the remaining long phase was strict SSF. To achieve the biphasic SSF, after 12 h (150 rpm, 37 °C) fermentation of the medium [Luria-Bertani (LB) supplemented with 30 % (w/v) raw soybean flour (phase I)], the supernatant in it was completely centrifuged out (1,000 × g, 10 min) aseptically for harvesting the extracellular enzymes as by-product. The resultant wet solid matter without free-flowing liquid but with embedded Btk was incubated 60 h more (phase II) for enhancing δ-endotoxin production at static condition (37 °C). Coupled with this, δ-endotoxin was purified by the modified phase separation method, and its purity was physically confirmed by both staining and microscopic techniques. The maximum δ-endotoxin yield from solid medium (48 h) was 15.8 mg/mL (recovery was 55-59 %) LB-equivalent, while that of LB control (recovery was 95 %) was only 0.43 mg/mL (72 h), i.e., thus, in comparison, 36.74-fold more yield in solid medium obtained by 24 h less gestation period. The purified crystal proteins showed apparent molecular weights (MWs) of 45, 35, and 6 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Briefly, this unique study physically demonstrates how Btk δ-endotoxin is purified (95-99 % purity) from solid-fermented matter for the first time, coupled with its overproduction at the expense of only 21.5 % higher production cost. PMID:25410805

  11. Screening of different adjuvants for wastewater/wastewater sludge-based Bacillus thuringiensis formulations.

    PubMed

    Brar, Satinder K; Verma, M; Tyagi, R D; Valéro, J R; Surampalli, R Y

    2006-08-01

    Screening of different adjuvants, namely, suspending agents, phagostimulants, stickers, antimicrobial agents, and UV screens to develop aqueous biopesticidal suspensions of Bacillus thuringiensis (Bt) variety kurstaki HD-1 fermented broths, specifically, nonhydrolyzed sludge, hydrolyzed sludge, starch industry wastewater, and soya (commercial medium), were investigated. The selected suspending agents [20% (wt:vol)] included sorbitol, sodium monophosphate, and sodium metabisulfite with corresponding suspendibility of 74-92, 69-85, and 71-82%, respectively. Molasses [0.2% (wt:vol)] increased adherence by 84-90% for all fermented broths. The optimal phagostimulants [0.5% (wt:vol)], namely, soya and molasses, caused entomotoxicity increase of 3-13 and 7-13%, respectively. Sorbic and propionic acids showed high antimicrobial action [0.5% (wt:vol)], irrespective of fermentation medium. Sodium lignosulfonate, molasses, and Congo red, when used as UV screens [0.2% (wt:vol)], showed percent corresponding entomotoxicity losses of 3-5, 0.5-5 and 2-16, respectively. The Bt formulations, when exposed to UV radiation, showed higher half-lives (with and without UV screens) than the fermented broths or semisynthetic soya medium and commercial Bt formulation. UV screen-amended nonhydrolyzed, hydrolyzed, and starch industry wastewater formulations showed 1.3-1.5-fold higher half-lives than commercial Bt formulation. Thus, the recommended formulation comprises sorbitol, sodium monophosphate, sodium metabisulfite (suspending agents); molasses, soya flour (phagostimulants); molasses and skimmed milk powder (rainfasteners); sorbic and propionic acids (antimicrobial agents) and sodium lignosulfate; and molasses and Congo red (UV screens). These waste-based Bt formulations offer better UV resistance in comparison with commercial formulation. PMID:16937657

  12. Cell Differentiation in a Bacillus thuringiensis Population during Planktonic Growth, Biofilm Formation, and Host Infection

    PubMed Central

    Verplaetse, Emilie; Slamti, Leyla; Gohar, Michel

    2015-01-01

    ABSTRACT Bacillus thuringiensis (Bt) is armed to complete a full cycle in its insect host. During infection, virulence factors are expressed under the control of the quorum sensor PlcR to kill the host. After the host’s death, the quorum sensor NprR controls a necrotrophic lifestyle, allowing the vegetative cells to use the insect cadaver as a bioincubator and to survive. Only a part of the Bt population sporulates in the insect cadaver, and the precise composition of the whole population and its evolution over time are unknown. Using fluorescent reporters to record gene expression at the single-cell level, we have determined the differentiation course of a Bt population and explored the lineage existing among virulent, necrotrophic, and sporulating cells. The dynamics of cell differentiation were monitored during growth in homogenized medium, biofilm formation, and colonization of insect larvae. We demonstrated that in the insect host and in planktonic culture in rich medium, the virulence, necrotrophism, and sporulation regulators are successively activated in the same cell. In contrast, in biofilms, activation of PlcR is dispensable for NprR activation and we observed a greater heterogeneity than under the other two growth conditions. We also showed that sporulating cells arise almost exclusively from necrotrophic cells. In biofilm and in the insect cadaver, we identified an as-yet-uncharacterized category of cells that do not express any of the reporters used. Overall, we showed that PlcR, NprR, and Spo0A act as interconnected integrators to allow finely tuned adaptation of the pathogen to its environment. PMID:25922389

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

    PubMed

    Li, Guangsheng; Wang, Yongmo; Liu, Biao; Zhang, Guoan

    2014-01-01

    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. PMID:25105299

  14. Effects of bacillus thuringiensis transgenic corn on corn earworm and fall armyworm (Lepidoptera: Noctuidae) densities.

    PubMed

    Chilcutt, Charles F; Odvody, Gary N; Correa, J Carlos; Remmers, Jeff

    2007-04-01

    We examined 17 pairs of near-isogenic hybrids of Bacillus thuringiensis (Bt) (176, Mon810, and Bt11) and non-Bt corn, Zea mays L., to examine the effects of Bt on larval densities of Helicoverpa zea (Boddie) and Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) during 2 yr. During ear formation, instar densities of H. zea and S. frugiperda were recorded for each hybrid. We found that H. zea first, second, and fifth instar densities were each affected by Mon810 and Bt11 Bt corn but not by 176 corn. Surprisingly, first and second instars were found in higher numbers on ears of Mon810 and Bt11 corn than on non-Bt corn. Densities of third and fourth instars were equal on Bt and non-Bt hybrids, whereas densities of fifth instars were lower on Bt plants. S. frugiperda larval densities were only affected during 1 yr when second, and fourth to sixth instars were lower on ears of Mon810 and Bt11 hybrids compared with their non-Bt counterparts. Two likely explanations for early instar H. zea densities being higher on Bt corn than non-Bt corn are that (1) Bt toxins delay development, creating a greater abundance of early instars that eventually die, and (2) reduced survival of H. zea to later instars on Bt corn decreased the normal asymmetric cannibalism or H. zea-S. frugiperda intraguild predation of late instars on early instars. Either explanation could explain why differences between Bt and non-Bt plants were greater for H. zea than S. frugiperda, because H. zea is more strongly affected by Bt toxins and more cannibalistic. PMID:17461054

  15. Improvement and efficient display of Bacillus thuringiensis toxins on M13 phages and ribosomes.

    PubMed

    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. PMID:26606918

  16. Comparative Analysis of Genomics and Proteomics in Bacillus thuringiensis 4.0718

    PubMed Central

    Rang, Jie; He, Hao; Wang, Ting; Ding, Xuezhi; Zuo, Mingxing; Quan, Meifang; Sun, Yunjun; Yu, Ziquan; Hu, Shengbiao; Xia, Liqiu

    2015-01-01

    Bacillus thuringiensis is a widely used biopesticide that produced various insecticidal active substances during its life cycle. Separation and purification of numerous insecticide active substances have been difficult because of the relatively short half-life of such substances. On the other hand, substances can be synthetized at different times during development, so samples at different stages have to be studied, further complicating the analysis. A dual genomic and proteomic approach would enhance our ability to identify such substances, and particularily using mass spectrometry-based proteomic methods. The comparative analysis for genomic and proteomic data have showed that not all of the products deduced from the annotated genome could be identified among the proteomic data. For instance, genome annotation results showed that 39 coding sequences in the whole genome were related to insect pathogenicity, including five cry genes. However, Cry2Ab, Cry1Ia, Cytotoxin K, Bacteriocin, Exoenzyme C3 and Alveolysin could not be detected in the proteomic data obtained. The sporulation-related proteins were also compared analysis, results showed that the great majority sporulation-related proteins can be detected by mass spectrometry. This analysis revealed Spo0A~P, SigF, SigE(+), SigK(+) and SigG(+), all known to play an important role in the process of spore formation regulatory network, also were displayed in the proteomic data. Through the comparison of the two data sets, it was possible to infer that some genes were silenced or were expressed at very low levels. For instance, found that cry2Ab seems to lack a functional promoter while cry1Ia may not be expressed due to the presence of transposons. With this comparative study a relatively complete database can be constructed and used to transform hereditary material, thereby prompting the high expression of toxic proteins. A theoretical basis is provided for constructing highly virulent engineered bacteria and for

  17. Role of Bacillus thuringiensis Cry1 δ Endotoxin Binding in Determining Potency during Lepidopteran Larval Development

    PubMed Central

    Gilliland, Androulla; Chambers, Catherine E.; Bone, Eileen J.; Ellar, David J.

    2002-01-01

    Five economically important crop pests, Manduca sexta, Pieris brassicae, Mamestra brassicae, Spodoptera exigua, and Agrotis ipsilon, were tested at two stages of larval development for susceptibility to Bacillus thuringiensis toxins Cry1Ac, Cry1Ca, Cry1J, and Cry1Ba. Bioassay results for M. sexta showed that resistance to all four Cry toxins increased from the neonate stage to the third-instar stage; the increase in resistance was most dramatic for Cry1Ac, the potency of which decreased 37-fold. More subtle increases in resistance during larval development were seen in M. brassicae for Cry1Ca and in P. brassicae for Cry1Ac and Cry1J. By contrast, the sensitivity of S. exigua did not change during development. At both larval stages, A. ipsilon was resistant to all four toxins. Because aminopeptidase N (APN) is a putative Cry1 toxin binding protein, APN activity was measured in neonate and third-instar brush border membrane vesicles (BBMV). With the exception of S. exigua, APN activity was found to be significantly lower in neonates than in third-instar larvae and thus inversely correlated with increased resistance during larval development. The binding characteristics of iodinated Cry1 toxins were determined for neonate and third-instar BBMV. In M. sexta, the increased resistance to Cry1Ac and Cry1Ba during larval development was positively correlated with fewer binding sites in third-instar BBMV than in neonate BBMV. The other species-instar-toxin combinations did not reveal positive correlations between potency and binding characteristics. The correlation between binding and potency was inconsistent for the species-instar-toxin combinations used in this study, reaffirming the complex mode of action of Cry1 toxins. PMID:11916662

  18. Bacillus thuringiensis: mechanism of action, resistance, and new applications: a review.

    PubMed

    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. PMID:25264571

  19. Bacillus thuringiensis-derived Cry5B Has Potent Anthelmintic Activity against Ascaris suum

    PubMed Central

    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. PMID:23818995

  20. Broad-spectrum resistance to Bacillus thuringiensis toxins by western corn rootworm (Diabrotica virgifera virgifera)

    PubMed Central

    Jakka, Siva R. K.; Shrestha, Ram B.; Gassmann, Aaron J.

    2016-01-01

    The evolution of resistance and cross-resistance threaten the sustainability of genetically engineered crops that produce insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt). Western corn rootworm, Diabrotica virgifera virgifera LeConte, is a serious pest of maize and has been managed with Bt maize since 2003. We conducted laboratory bioassays with maize hybrids producing Bt toxins Cry3Bb1, mCry3A, eCry3.1Ab, and Cry34/35Ab1, which represent all commercialized Bt toxins for management of western corn rootworm. We tested populations from fields where severe injury to Cry3Bb1 maize was observed, and populations that had never been exposed to Bt maize. Consistent with past studies, bioassays indicated that field populations were resistant to Cry3Bb1 maize and mCry3A maize, and that cross-resistance was present between these two types of Bt maize. Additionally, bioassays revealed resistance to eCry3.1Ab maize and cross-resistance among Cry3Bb1, mCry3A and eCry3.1Ab. However, no resistance or cross-resistance was detected for Cry34/35Ab1 maize. This broad-spectrum resistance illustrates the potential for insect pests to develop resistance rapidly to multiple Bt toxins when structural similarities are present among toxins, and raises concerns about the long-term durability of Bt crops for management of some insect pests. PMID:27297953

  1. Field-evolved resistance by western corn rootworm to multiple Bacillus thuringiensis toxins in transgenic maize

    PubMed Central

    Gassmann, Aaron J.; Petzold-Maxwell, Jennifer L.; Clifton, Eric H.; Dunbar, Mike W.; Hoffmann, Amanda M.; Ingber, David A.; Keweshan, Ryan S.

    2014-01-01

    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. PMID:24639498

  2. Transgenic Bacillus thuringiensis (Bt) Rice Is Safer to Aquatic Ecosystems than Its Non-Transgenic Counterpart

    PubMed Central

    Li, Guangsheng; Wang, Yongmo; Liu, Biao; Zhang, Guoan

    2014-01-01

    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. PMID:25105299

  3. Absence of toxicity of Bacillus thuringiensis pollen to black swallowtails under field conditions.

    PubMed

    Wraight, C L; Zangerl, A R; Carroll, M J; Berenbaum, M R

    2000-07-01

    A single laboratory study on monarch butterflies has prompted widespread concern that corn pollen, engineered to express Bacillus thuringiensis (Bt) endotoxin, might travel beyond corn fields and cause mortality in nontarget lepidopterans. Among the lepidopterans at high potential risk from this technology is the black swallowtail butterfly, Papilio polyxenes, whose host plants in the midwestern U. S. are located principally in narrow strips between roads and crop fields. A field study was performed to assess whether mortality of early instar black swallowtails was associated either with proximity to a field of Bt corn or by levels of Bt pollen deposition on host plants. Potted host plants were infested with first instar black swallowtails and placed at intervals from the edge of a field of Bt corn (Pioneer 34R07 containing Monsanto event 810) at the beginning of anthesis. We confirmed by ELISA that pollen from these plants contained Cry1Ab endotoxin (2.125 +/- 0.289 ng/g). Although many of the larvae died during the 7 days that the experiments were run, there was no relationship between mortality and proximity to the field or pollen deposition on host plants. Moreover, pollen from these same plants failed to cause mortality in the laboratory at the highest pollen dose tested (10,000 grains/cm(2)), a level that far exceeded the highest pollen density observed in the field (200 grains/cm(2)). We conclude that Bt pollen of the variety tested is unlikely to affect wild populations of black swallowtails. Thus, our results suggest that at least some potential nontarget effects of the use of transgenic plants may be manageable. PMID:10840067

  4. Cloning and characterization of a unique cytotoxic protein parasporin-5 produced by Bacillus thuringiensis A1100 strain.

    PubMed

    Ekino, Keisuke; Okumura, Shiro; Ishikawa, Tomoyuki; Kitada, Sakae; Saitoh, Hiroyuki; Akao, Tetsuyuki; Oka, Takuji; Nomura, Yoshiyuki; Ohba, Michio; Shin, Takashi; Mizuki, Eiichi

    2014-06-01

    Parasporin is the cytocidal protein present in the parasporal inclusion of the non-insecticidal Bacillus thuringiensis strains, which has no hemolytic activity but has cytocidal activities, preferentially killing cancer cells. In this study, we characterized a cytocidal protein that belongs to this category, which was designated parasporin-5 (PS5). PS5 was purified from B. thuringiensis serovar tohokuensis strain A1100 based on its cytocidal activity against human leukemic T cells (MOLT-4). The 50% effective concentration (EC₅₀) of PS5 to MOLT-4 cells was approximately 0.075 μg/mL. PS5 was expressed as a 33.8-kDa inactive precursor protein and exhibited cytocidal activity only when degraded by protease at the C-terminal into smaller molecules of 29.8 kDa. Although PS5 showed no significant homology with other known parasporins, a Position Specific Iterative-Basic Local Alignment Search Tool (PSI-BLAST) search revealed that the protein showed slight homology to, not only some B. thuringiensis Cry toxins, but also to aerolysin-type β-pore-forming toxins (β-PFTs). The recombinant PS5 protein could be obtained as an active protein only when it was expressed in a precursor followed by processing with proteinase K. The cytotoxic activities of the protein against various mammalian cell lines were evaluated. PS5 showed strong cytocidal activity to seven of 18 mammalian cell lines tested, and low to no cytotoxicity to the others. PMID:24945755

  5. Microdroplet application of mosquitocidal Bacillus thuringiensis using ultra-low-volume generator for the control of mosquitos.

    PubMed

    Seleena, P; Lee, H L; Nazni, W A; Rohani, A; Kadri, M S

    1996-09-01

    In an effort to develop a more effective technique in dispersing a microbial control agent, Bacillus thuringiensis (Bt), a truck-mounted ultra low volume (ULV) generator (Scorpion) was used to disperse B. thuringiensis israelensis (Bti) and Bti with malathion. Complete larval and adult mortalities for all tested mosquito species within the first 70-80 feet from the ULV generator were achieved. Beyond that distance less than 50% mortality was achieved as insufficient sprayed particles reached the area. A minimum of 10(3) Bti colony forming units per ml is required to cause 100% larval mortality. The sprayed Bti larvicidal toxins were persistent in the test water 7 days post ULV. The effectiveness of B. thuringiensis jegathesan (Btj), a new mosquitocidal Bt serotype was also evaluated. Similar mortality results as Bti were achieved except that the Btj toxins underwent degradation in the test water, since less than 50% less in larval mortality was observed in 7 days post ULV samples. This ULV method has the potential to disperse Bt and malathion effectively for a simultaneous control of mosquito adults and larvae. PMID:9185282

  6. Production of polyclonal and monoclonal antibodies against the Bacillus thuringiensis vegetative insecticidal protein Vip3Aa16.

    PubMed

    Ben Hamadou-Charfi, Dorra; Sauer, Annette Juliane; Abdelkafi-Mesrati, Lobna; Jaoua, Samir; Stephan, Dietrich

    2015-03-01

    The aim of this study is to establish a quantitative determination of the vegetative insecticidal protein Vip3A from the culture supernatant of Bacillus thuringiensis either by ELISA or by the conventional quantification method of the Western blot band. The Vip3A protein was produced by fermentation of the B. thuringiensis reference strain BUPM95 in 3 L. By Western blot, the Vip3Aa16 toxin was detected in the culture supernatant during the exponential growth phase of B. thuringiensis BUPM95. However, the detection of Vip3Aa16 on Western blot showed in addition to the toxin two other strips (62 and 180 kDa) recognized by the anti-Vip3Aa16 polyclonal antibodies prepared at the Centre of Biotechnology of Sfax Tunisia. For that reason and in order to develop a technique for reliable quantification of the toxin, we have considered the production of polyclonal antibodies at the Julius Kühn Institute, Germany. These antibodies were the basis for the production of monoclonal antibodies directed against the protein produced by the Vip3Aa16 recombinant strain Escherichia coli BL21 (DE3). These monoclonal antibodies were tested by plate-trapped antigen (PTA) and triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA). The selection of hybridoma supernatants gave us four positive clones producing monoclonal antibodies. PMID:25492687

  7. The Metabolic Regulation of Sporulation and Parasporal Crystal Formation in Bacillus thuringiensis Revealed by Transcriptomics and Proteomics*

    PubMed Central

    Wang, Jieping; Mei, Han; Zheng, Cao; Qian, Hongliang; Cui, Cui; Fu, Yang; Su, Jianmei; Liu, Ziduo; Yu, Ziniu; He, Jin

    2013-01-01

    Bacillus thuringiensis is a well-known entomopathogenic bacterium used worldwide as an environmentally compatible biopesticide. During sporulation, B. thuringiensis accumulates a large number of parasporal crystals consisting of insecticidal crystal proteins (ICPs) that can account for nearly 20–30% of the cell's dry weight. However, the metabolic regulation mechanisms of ICP synthesis remain to be elucidated. In this study, the combined efforts in transcriptomics and proteomics mainly uncovered the following 6 metabolic regulation mechanisms: (1) proteases and the amino acid metabolism (particularly, the branched-chain amino acids) became more active during sporulation; (2) stored poly-β-hydroxybutyrate and acetoin, together with some low-quality substances provided considerable carbon and energy sources for sporulation and parasporal crystal formation; (3) the pentose phosphate shunt demonstrated an interesting regulation mechanism involving gluconate when CT-43 cells were grown in GYS medium; (4) the tricarboxylic acid cycle was significantly modified during sporulation; (5) an obvious increase in the quantitative levels of enzymes and cytochromes involved in energy production via the electron transport system was observed; (6) most F0F1-ATPase subunits were remarkably up-regulated during sporulation. This study, for the first time, systematically reveals the metabolic regulation mechanisms involved in the supply of amino acids, carbon substances, and energy for B. thuringiensis spore and parasporal crystal formation at both the transcriptional and translational levels. PMID:23408684

  8. The glycoprotein toxin of Bacillus thuringiensis subsp. israelensis indicates a lectinlike receptor in the larval mosquito gut.

    PubMed Central

    Muthukumar, G; Nickerson, K W

    1987-01-01

    The mosquito-active protein crystals produced by Bacillus thuringiensis subsp. israelensis contain covalently attached aminosugars which are critical for their larvicidal activity. The 50% lethal concentrations toward Aedes aegypti larvae were increased up to 10-fold by mild periodate treatment, up to 40-fold by forming the protein crystals in the presence of tunicamycin, and up to 7-fold by the presence during the mosquito bioassays of N-acetylglucosamine or its trimer, triacetylchitotriose. Periodate-treated crystals and crystals formed in the presence of tunicamycin had greatly reduced binding capacities for wheat germ agglutinin, an N-acetylglucosamine-specific lectin. These results suggest that the B. thuringiensis subsp. israelensis glycoprotein toxin binds to a lectinlike receptor in the larval mosquito gut. Furthermore, the distinct lectin-binding patterns exhibited by diptera-active versus lepidoptera-active B. thuringiensis crystals suggest that host specificity for the microbial insecticides is determined, in part, by the carbohydrate portion of their glycoprotein crystals. Images PMID:2827571

  9. Low persistence of Bacillus thuringiensis serovar israelensis spores in four mosquito biotopes of a salt marsh in southern France.

    PubMed

    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). PMID:16328650

  10. Complete sequence of three plasmids from Bacillus thuringiensis INTA-FR7-4 environmental isolate and comparison with related plasmids from the Bacillus cereus group.

    PubMed

    Amadio, Ariel F; Benintende, Graciela B; Zandomeni, Rubén O

    2009-11-01

    Bacillus thuringiensis is an insect pathogen used worldwide as a bioinsecticide. It belongs to the Bacillus cereus sensu lato group as well as Bacillus anthracis and B. cereus. Plasmids from this group of organisms have been implicated in pathogenicity as they carry the genes responsible for different types of diseases that affect mammals and insects. Some plasmids, like pAW63 and pBT9727, encode a functional conjugation machinery allowing them to be transferred to a recipient cell. They also share extensive homology with the non-functional conjugation apparatus of pXO2 from B. anthracis. In this study we report the complete sequence of three plasmids from an environmental B. thuringiensis isolate from Argentina, obtained by a shotgun sequencing method. We obtained the complete nucleotide sequence of plasmids pFR12 (12,095bp), pFR12.5 (12,459bp) and pFR55 (55,712bp) from B. thuringiensis INTA-FR7-4. pFR12 and pFR12.5 were classified as cryptic as they do not code for any obvious functions besides replication and mobilization. Both small plasmids were classified as RCR plasmids due to similarities with the replicases they encode. Plasmid pFR55 showed a structural organization similar to that observed for plasmids pAW63, pBT9727 and pXO2. pFR55 also shares a tra region with these plasmids, containing genes related to T4SS and conjugation. A comparison between pFR55 and conjugative plasmids led to the postulation that pFR55 is a conjugative plasmid. Genes related to replication functions in pFR55 are different to those described for plasmids with known complete sequences. pFR55 is the first completely sequenced plasmid with a replication machinery related to that of ori44. The analysis of the complete sequence of plasmids from an environmental isolate of B. thuringiensis permitted the identification of a near complete conjugation apparatus in pFR55, resembling those of plasmids pAW63, pBT9727 and pXO2. The availability of this sequence is a step forward in the study

  11. Poly-β-hydroxybutyrate Metabolism Is Unrelated to the Sporulation and Parasporal Crystal Protein Formation in Bacillus thuringiensis

    PubMed Central

    Wang, Xun; Li, Zhou; Li, Xin; Qian, Hongliang; Cai, Xia; Li, Xinfeng; He, Jin

    2016-01-01

    Poly-3-hydroxybutyrate (PHB) is a natural polymer synthesized by many bacteria as a carbon-energy storage material. It was accumulated maximally prior to the spore formation but was degraded during the process of sporulation in Bacillus thuringiensis. Intriguingly, B. thuringiensis also accumulates large amounts of insecticidal crystal proteins (ICPs) during sporulation, which requires considerable input of carbon and energy sources. How PHB accumulation affects sporulation and ICP formation remains unclear to date. Intuitively, one would imagine that accumulated PHB provides the energy required for ICP formation. Yet our current data indicate that this is not the case. First, growth curves of the deletion mutants of phaC (encoding the PHB synthase) and phaZ (encoding the PHB depolymerase) were found to be similar to the parent strain BMB171; no difference in growth rate could be observed. In addition we further constructed the cry1Ac10 ICP gene overexpression strains of BMB171 (BMB171-cry), as well as its phaC and phaZ deletion mutants ΔphaC-cry and ΔphaZ-cry to compare their spore and ICP production rates. Again, not much change of ICP production was observed among these strains either. In fact, PHB was still degraded in most ΔphaZ-cry cells as observed by transmission electron microscopy. Together these results indicated that there is no direct association between the PHB accumulation and the sporulation and ICP formation in B. thuringiensis. Some other enzymes for PHB degradation or other energy source may be responsible for the sporulation and/or ICP formation in B. thuringiensis. PMID:27379025

  12. Poly-β-hydroxybutyrate Metabolism Is Unrelated to the Sporulation and Parasporal Crystal Protein Formation in Bacillus thuringiensis.

    PubMed

    Wang, Xun; Li, Zhou; Li, Xin; Qian, Hongliang; Cai, Xia; Li, Xinfeng; He, Jin

    2016-01-01

    Poly-3-hydroxybutyrate (PHB) is a natural polymer synthesized by many bacteria as a carbon-energy storage material. It was accumulated maximally prior to the spore formation but was degraded during the process of sporulation in Bacillus thuringiensis. Intriguingly, B. thuringiensis also accumulates large amounts of insecticidal crystal proteins (ICPs) during sporulation, which requires considerable input of carbon and energy sources. How PHB accumulation affects sporulation and ICP formation remains unclear to date. Intuitively, one would imagine that accumulated PHB provides the energy required for ICP formation. Yet our current data indicate that this is not the case. First, growth curves of the deletion mutants of phaC (encoding the PHB synthase) and phaZ (encoding the PHB depolymerase) were found to be similar to the parent strain BMB171; no difference in growth rate could be observed. In addition we further constructed the cry1Ac10 ICP gene overexpression strains of BMB171 (BMB171-cry), as well as its phaC and phaZ deletion mutants ΔphaC-cry and ΔphaZ-cry to compare their spore and ICP production rates. Again, not much change of ICP production was observed among these strains either. In fact, PHB was still degraded in most ΔphaZ-cry cells as observed by transmission electron microscopy. Together these results indicated that there is no direct association between the PHB accumulation and the sporulation and ICP formation in B. thuringiensis. Some other enzymes for PHB degradation or other energy source may be responsible for the sporulation and/or ICP formation in B. thuringiensis. PMID:27379025

  13. Biosorption of heavy metals by Bacillus thuringiensis strain OSM29 originating from industrial effluent contaminated north Indian soil

    PubMed Central

    Oves, Mohammad; Khan, Mohammad Saghir; Zaidi, Almas

    2012-01-01

    The study was navigated to examine the metal biosorbing ability of bacterial strain OSM29 recovered from rhizosphere of cauliflower grown in soil irrigated consistently with industrial effluents. The metal tolerant bacterial strain OSM29 was identified as Bacillus thuringiensis following 16S rRNA gene sequence analysis. In the presence of the varying concentrations (25–150 mgl−1) of heavy metals, such as cadmium, chromium, copper, lead and nickel, the B. thuringiensis strain OSM29 showed an obvious metal removing potential. The effect of certain physico-chemical factors such as pH, initial metal concentration, and contact time on biosorption was also assessed. The optimum pH for nickel and chromium removal was 7, while for cadmium, copper and lead, it was 6. The optimal contact time was 30 min. for each metal at 32 ± 2 °C by strain OSM29. The biosorption capacity of the strain OSM29 for the metallic ions was highest for Ni (94%) which was followed by Cu (91.8%), while the lowest sorption by bacterial biomass was recorded for Cd (87%) at 25 mgl−1 initial metal ion concentration. The regression coefficients obtained for heavy metals from the Freundlich and Langmuir models were significant. The surface chemical functional groups of B. thuringiensis biomass identified by Fourier transform infrared (FTIR) were amino, carboxyl, hydroxyl, and carbonyl groups, which may be involved in the biosorption of heavy metals. The biosorption ability of B. thuringiensis OSM29 varied with metals and was pH and metal concentration dependent. The biosorption of each metal was fairly rapid which could be an advantage for large scale treatment of contaminated sites. PMID:24115905

  14. Effects of physical culture parameters on bacteriocin production by Mexican strains of Bacillus thuringiensis after cellular induction.

    PubMed

    Martínez-Cardeñas, Janeth Adriana; de la Fuente-Salcido, Norma M; Salcedo-Hernández, Rubén; Bideshi, Dennis K; Barboza-Corona, J Eleazar

    2012-01-01

    We have shown previously that in the presence of inducer Bacillus cereus 183, significant increases in bacteriocin production and bactericidal activity of B. thuringiensis occur when the latter is cultivated at pH 7.2, 28°C, and 180 rpm. Here we show that this activity can be further improved when B. thuringiensis is induced with B. cereus 183 and then cultivated with modification of pH, temperature, and agitation. Five native strains of B. thuringiensis, LBIT 269, LBIT 287, LBIT 404, LBIT 420, and LBIT 524 which synthesize, respectively, morricin 269, kurstacin 287, kenyacin 404, entomocin 420, and tolworthcin 524, were cultivated in four different fermentation media. Of these, fermentation in tryptic soy broth (TSB) yielded the highest level of bacteriocin activity (~100-133 FU). Bacteria grown in TSB were induced with B. cereus 183 and cultivated at different pH (6.0, 7.2, 8.0), temperature (26, 28, 30°C), and agitation (150, 180, 210 rpm). Full factorial design was performed and results were analyzed with analysis of variance (ANOVA) and Tukey multiple comparison tests at significant level of α ≤ 0.05 to study the influence of the three variables on bacterial growth and bacteriocin production. Our data show that the highest bacteriocin activity was found with LBIT 269 and LBIT 404 with an increase of ~95-100% compared with induced B. thuringiensis strains cultivated under fixed conditions (pH 7.2, 28°C, 180 rpm), for which the data were set at 0%. The optimal conditions for morricin 269 and kenyacin 404 production were, respectively, pH 8, 30°C, 210 rpm and pH 7.2, 26°C, 210 rpm. PMID:21743998

  15. Recovery of commercially produced Bacillus thuringiensis var. israelensis and Bacillus sphaericus from tires and prevalence of bacilli in artificial and natural containers.

    PubMed

    Siegel, J P; Smith, A R; Novak, R J

    2001-03-01

    We conducted surveys to identify the species of spore-forming bacteria present in natural and artificial containers. Most of our samples came from Illinois. Identification was based on the cellular fatty acid composition of the bacterial cell wall. In addition, we utilized a custom database for commercially produced strains of Bacillus thuringiensis var. israelensis (Bti) and B. sphaericus, to differentiate between larvicidal isolates with commercial or native origin. Native Bti was present at low levels in almost all habitats but was not recovered from bromeliads and metal containers. In temporary woodland pools, 27.9% of the colonies recovered were native Bti. We did not recover larvicidal B. sphaericus in untreated habitats. VectoBac and VectoLex were applied to tires containing water and the tires were sampled 3 months and 9 months after treatment. Isolates of Bti and B. sphaericus with commercial origin were recovered as long as 9 months after application. We noticed numerous cadavers of Aedes triseriatus in several tires 9 months after treatment with VectoBac. We could not determine if this mortality resulted from recycling of Bti in these tires or whether insecticidal crystal proteins from the original treatment were resuspended. Bacillus thuringiensis var. israelensis isolates with commercial ancestry were recovered from untreated tires 9 months after application. Isolates of larvicidal B. sphaericus that differed from the bacteria in VectoLex were also recovered from untreated tires. PMID:11345416

  16. [A prediction model for the activity of insecticidal crystal proteins from Bacillus thuringiensis based on support vector machine].

    PubMed

    Lin, Yi; Cai, Fu-Ying; Zhang, Guang-Ya

    2007-01-01

    A quantitative structure-property relationship (QSPR) model in terms of amino acid composition and the activity of Bacillus thuringiensis insecticidal crystal proteins was established. Support vector machine (SVM) is a novel general machine-learning tool based on the structural risk minimization principle that exhibits good generalization when fault samples are few; it is especially suitable for classification, forecasting, and estimation in cases where small amounts of samples are involved such as fault diagnosis; however, some parameters of SVM are selected based on the experience of the operator, which has led to decreased efficiency of SVM in practical application. The uniform design (UD) method was applied to optimize the running parameters of SVM. It was found that the average accuracy rate approached 73% when the penalty factor was 0.01, the epsilon 0.2, the gamma 0.05, and the range 0.5. The results indicated that UD might be used an effective method to optimize the parameters of SVM and SVM and could be used as an alternative powerful modeling tool for QSPR studies of the activity of Bacillus thuringiensis (Bt) insecticidal crystal proteins. Therefore, a novel method for predicting the insecticidal activity of Bt insecticidal crystal proteins was proposed by the authors of this study. PMID:17366901

  17. Quorum sensing in Bacillus thuringiensis is required for completion of a full infectious cycle in the insect.

    PubMed

    Slamti, Leyla; Perchat, Stéphane; Huillet, Eugénie; Lereclus, Didier

    2014-08-01

    Bacterial cell-cell communication or quorum sensing (QS) is a biological process commonly described as allowing bacteria belonging to a same pherotype to coordinate gene expression to cell density. In Gram-positive bacteria, cell-cell communication mainly relies on cytoplasmic sensors regulated by secreted and re-imported signaling peptides. The Bacillus quorum sensors Rap, NprR, and PlcR were previously identified as the first members of a new protein family called RNPP. Except for the Rap proteins, these RNPP regulators are transcription factors that directly regulate gene expression. QS regulates important biological functions in bacteria of the Bacillus cereus group. PlcR was first characterized as the main regulator of virulence in B. thuringiensis and B. cereus. More recently, the PlcR-like regulator PlcRa was characterized for its role in cysteine metabolism and in resistance to oxidative stress. The NprR regulator controls the necrotrophic properties allowing the bacteria to survive in the infected host. The Rap proteins negatively affect sporulation via their interaction with a phosphorelay protein involved in the activation of Spo0A, the master regulator of this differentiation pathway. In this review we aim at providing a complete picture of the QS systems that are sequentially activated during the lifecycle of B. cereus and B. thuringiensis in an insect model of infection. PMID:25089349

  18. Improvement of Bacillus sphaericus toxicity against dipteran larvae by integration, via homologous recombination, of the Cry11A toxin gene from Bacillus thuringiensis subsp. israelensis.

    PubMed Central

    Poncet, S; Bernard, C; Dervyn, E; Cayley, J; Klier, A; Rapoport, G

    1997-01-01

    Integrative plasmids were constructed to enable integration of foreign DNA into the chromosome of Bacillus sphaericus 2297 by in vivo recombination. Integration of the aphA3 kanamycin resistance gene by a two-step procedure demonstrated that this strategy was applicable with antibiotic resistance selection. Hybridization experiments evidenced two copies of the operon encoding the binary toxin from B. sphaericus in the recipient strain. The Bacillus thuringiensis subsp. israelensis cry11Aal gene (referred to as cry11A), encoding a delta-endotoxin with toxicity against Culex, Aedes, and Anopheles larvae, was integrated either by a single crossover event [strain 2297 (::pHT5601), harboring the entire recombinant plasmid] or by two successive crossover events [strain 2297 (::cry11A)]. The level of the Cry11A production in B. sphaericus was high; two crystalline inclusions were produced in strain 2297 (::pHT5601). Synthesis of the Cry11A toxin conferred toxicity to the recombinant strains against Aedes aegypti larvae, for which the parental strain was not toxic. Interestingly, the level of larvicidal activity of strain 2297 (::pHT5601) against Anopheles stephensi was as high as that of B. thuringiensis subsp. israelensis and suggested synergy between the B. thuringiensis and B. sphaericus toxins. The toxicities of parental and recombinant B. sphaericus strains against Culex quinquefasciatus were similar, but the recombinant strains killed the larvae more rapidly. The production of the Cry11A toxin in B. sphaericus also partially restored toxicity for C. quinquefasciatus larvae from a population resistant to B. sphaericus 1593. In vivo recombination therefore appears to be a promising approach to the creation of new B. sphaericus strains for vector control. PMID:9361428

  19. Combined effect of seaweed (Sargassum wightii) and Bacillus thuringiensis var. israelensis on the coastal mosquito,Anopheles sundaicus, in Tamil Nadu, India

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Studies were made of the extract of Sargassum wightii combined with Bacillus thuringiensis var. israelensis (Bti) for control of the malaria vector Anopheles sundaicus. Treatment of mosquito larvae with 0.001% S. wightii extract indicated median lethal concentrations (LC50) of 88, 73, 134, 156, and...

  20. Susceptibility of northern corn rootworm Diabrotica barberi Smith & Lawrence (Coleoptera: Chrysomelidae) to mCry3A and eCry3.1Ab Bacillus thuringiensis proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Susceptibility of the northern corn rootworm (NCR), to mCry3A and eCry3.1Ab proteins derived from Bacillus thuringiensis (Bt) was determined using a diet bioassay. Northern corn rootworm neonates were exposed to different concentrations of mCry3A and eCry3.1Ab, incorporated into artificial diet. Lar...

  1. Evaluation of a Granulovirus (PoGV) and Bacillus Thuringiensis subsp. kurstaki for Control of the Potato Tuberworm in Stored Tubers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Currently no insecticides are registered for postharvest use on potato tubers in North America. We evaluated liquid suspensions and dry formulations of a granulovirus (PoGV) and bacterium Bacillus thuringiensis subsp. kurstaki Berliner (Btk) for control of the potato tuber moth (PTM), Phthorimaea op...

  2. Geographical susceptibility of Louisiana and Texas populations of sugarcane borer, Diatraea saccharalis (F.) (Lepidopetera: Crambidae) to Bacillus thuringiensis Cry1Ab protein

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The susceptibility of 18 field populations of the sugarcane borer, Diatraea saccharalis (F.) to two sources of Bacillus thuringiensis Cry1Ab protein was determined by laboratory bioassays. Fifteen of the 18 field populations were collected from seven locations across Louisiana and the other 3 popula...

  3. Infection of Tribolium castaneum with Bacillus thuringiensis: Quantification of Bacterial Replication within Cadavers, Transmission via Cannibalism, and Inhibition of Spore Germination

    PubMed Central

    Milutinović, Barbara; Höfling, Christina; Futo, Momir; Scharsack, Jörn P.

    2015-01-01

    Reproduction within a host and transmission to the next host are crucial for the virulence and fitness of pathogens. Nevertheless, basic knowledge about such parameters is often missing from the literature, even for well-studied bacteria, such as Bacillus thuringiensis, an endospore-forming insect pathogen, which infects its hosts via the oral route. To characterize bacterial replication success, we made use of an experimental oral infection system for the red flour beetle Tribolium castaneum and developed a flow cytometric assay for the quantification of both spore ingestion by the individual beetle larvae and the resulting spore load after bacterial replication and resporulation within cadavers. On average, spore numbers increased 460-fold, showing that Bacillus thuringiensis grows and replicates successfully in insect cadavers. By inoculating cadaver-derived spores and spores from bacterial stock cultures into nutrient medium, we next investigated outgrowth characteristics of vegetative cells and found that cadaver-derived bacteria showed reduced growth compared to bacteria from the stock cultures. Interestingly, this reduced growth was a consequence of inhibited spore germination, probably originating from the host and resulting in reduced host mortality in subsequent infections by cadaver-derived spores. Nevertheless, we further showed that Bacillus thuringiensis transmission was possible via larval cannibalism when no other food was offered. These results contribute to our understanding of the ecology of Bacillus thuringiensis as an insect pathogen. PMID:26386058

  4. An Empirical Test of the F2 Screen for Detection of Bacillus thuringiensis-Resistance Alleles in Tobacco Budworm (Lepidoptera: Noctuidae)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Insects exposed to genetically-modified crops expressing Bacillus thuringiensis (Bt) toxins are under intense selection pressure that could result in widespread Bt resistance. Screening for early indications of Bt resistance developing in targeted Lepidoptera is conducted in many of the regions wher...

  5. Linkage of an ABCC transporter to a single QTL that controls Ostrinia nubilalis larval resistance to the Bacillus thuringiensis Cry1Fa toxin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Field evolved resistance of insect populations to Bacillus thuringiensis (Bt) crystalline (Cry) toxins expressed by crop plants has resulted in reduced control of insect feeding damage to field crops, and threatens the sustainability of Bt transgenic technologies. A single quantitative trait locus ...

  6. 40 CFR 174.510 - Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance. 174.510 Section 174.510 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS...

  7. Bacillus thuringiensis toxin resistance mechanisms among Lepidoptera: progress on genomic approaches to uncover causal mutations in the European corn borer, Ostrinia nubilalis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenic plants that expressed Bacillus thuringiensis (Bt) crystalline (Cry) protein toxins can suffer feeding damage from a small number of lepidopteran insect species under field conditions, which has heightened concerns about the durability of pest control tactics. Genomics research has provid...

  8. 40 CFR 174.532 - Bacillus thuringiensis eCry3.1Ab protein in corn; temporary exemption from the requirement of a...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis eCry3.1Ab protein in corn; temporary exemption from the requirement of a tolerance. 174.532 Section 174.532 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED...

  9. 40 CFR 174.505 - Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis modified Cry3A protein (mCry3A) in corn; exemption from the requirement of a tolerance. 174.505 Section 174.505 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED...

  10. 40 CFR 174.518 - Bacillus thuringiensis Cry3Bb1 protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry3Bb1 protein in corn; exemption from the requirement of a tolerance. 174.518 Section 174.518 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances...

  11. 40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED...

  12. 40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance. 174.511 Section 174.511 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS...

  13. 40 CFR 174.501 - Bacillus thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a tolerance. 174.501 Section 174.501 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED...

  14. Expressed sequence tags from larval gut of the european corn borer (Ostrinia nubilalis): exploring candidate genes potenially involved in Bacillus thuringiensis toxicity and resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Background: 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) toxin and for discovering new targets for novel toxins for use in pest management. This study analyzed the ES...

  15. Unlinked genetic loci control the reduced transcription of aminopeptidase N 1 and 3 in the European corn borer and determine tolerance to Bacillus thuringiensis Cry1Ab toxin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Crystalline (Cry) toxins from Bacillus thuringiensis (Bt) control insect feeding damage on crop plants via foliar applications or by expression within transgenic plants, but continued Bt use is threatened by the buildup of insect resistance traits. Aminopeptidase N (apn) gene family members encode m...

  16. SEMI-FIELD EVALUATION OF THE POTATO TUBER MOTH, PHTHORIMAEA OPERCULELLA ZELLER, GRANULOVIRUS AND BACILLUS THURINGIENSIS VAR. KURSTAKI FOR SEASON-LONG CONTROL OF P. OPERCULELLA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In 2006 and 2007, scientists at YARL evaluated two low risk biological pesticides, an insect-specific virus (PoGV) and a bacterium [Bacillus thuringiensis var. kurstaki (Btk)], that would provide alternative tools to control PTM. Compared with controls, 10 weekly applications of PoGV (standard rate)...

  17. Increased toxicity of Bacillus thuringiensis Cry3Aa against Crioceris quatuordecimpunctata, Phaedon brassicae and Colaphellus bowringi by a Tenebrio molitor cadherin fragment

    Technology Transfer Automated Retrieval System (TEKTRAN)

    BACKGROUND: Biopesticides containing Cry insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) are effective against many lepidopteran pests, but there is a lack of Bt-based pesticides to efficiently control important coleopteran pests. Based on the reported increase of Bt toxin olig...

  18. 40 CFR 174.532 - Bacillus thuringiensis eCry3.1Ab protein in corn; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... protein in corn; exemption from the requirement of a tolerance. 174.532 Section 174.532 Protection of...Cry3.1Ab protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis eCry3.1Ab protein in corn, in or on the food and feed commodities of corn; corn, field;...

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

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... protein in corn; exemption from the requirement of a tolerance. 174.532 Section 174.532 Protection of...Cry3.1Ab protein in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis eCry3.1Ab protein in corn, in or on the food and feed commodities of corn; corn, field;...

  20. Infection of Tribolium castaneum with Bacillus thuringiensis: quantification of bacterial replication within cadavers, transmission via cannibalism, and inhibition of spore germination.

    PubMed

    Milutinović, Barbara; Höfling, Christina; Futo, Momir; Scharsack, Jörn P; Kurtz, Joachim

    2015-12-01

    Reproduction within a host and transmission to the next host are crucial for the virulence and fitness of pathogens. Nevertheless, basic knowledge about such parameters is often missing from the literature, even for well-studied bacteria, such as Bacillus thuringiensis, an endospore-forming insect pathogen, which infects its hosts via the oral route. To characterize bacterial replication success, we made use of an experimental oral infection system for the red flour beetle Tribolium castaneum and developed a flow cytometric assay for the quantification of both spore ingestion by the individual beetle larvae and the resulting spore load after bacterial replication and resporulation within cadavers. On average, spore numbers increased 460-fold, showing that Bacillus thuringiensis grows and replicates successfully in insect cadavers. By inoculating cadaver-derived spores and spores from bacterial stock cultures into nutrient medium, we next investigated outgrowth characteristics of vegetative cells and found that cadaver-derived bacteria showed reduced growth compared to bacteria from the stock cultures. Interestingly, this reduced growth was a consequence of inhibited spore germination, probably originating from the host and resulting in reduced host mortality in subsequent infections by cadaver-derived spores. Nevertheless, we further showed that Bacillus thuringiensis transmission was possible via larval cannibalism when no other food was offered. These results contribute to our understanding of the ecology of Bacillus thuringiensis as an insect pathogen. PMID:26386058

  1. Bacillus thuringiensis plants expressing Cry1Ac, Cry2Ab and Cry1F are not toxic to the assassin bug, Zelus renardii

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cotton and maize delivering insecticidal crystal (Cry) proteins from the bacterium, Bacillus thuringiensis (Bt), have been commercialized since 1996. Bt plants are subjected to environmental risk assessments for non-target organisms, especially natural enemies that suppress pest populations. In th...

  2. Molecular characterization and RNA interference of three midgut aminopeptidase N isozymes from bacillus thuringiensis-susceptible and -resistant strains of sugarcane borer diatraea saccharalis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aminopeptidase N (APN) proteins located at the midgut epithelium of some lepidopterous species have been implicated as receptors for insecticidal proteins from Bacillus thuringiensis. cDNAs of three APN isoforms, DsAPN1, DsAPN2, and DsAPN3, from Cry1Ab-susceptible (Cry1Ab-SS) and -resistant (Cry1Ab-...

  3. Susceptibility of Northern Corn Rootworm Diabrotica barberi Smith (Coleoptera: Chrysomelidae) to mCry3A and eCry3.1Ab Bacillus thuringiensis proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Susceptibility of the northern corn rootworm (NCR), to mCry3A and eCry3.1Ab proteins derived from Bacillus thuringiensis (Bt) was determined using a diet bioassay. Northern corn rootworm neonates were exposed to different concentrations of mCry3A and eCry3.1Ab, incorporated into artificial diet. Lar...

  4. Integrated applications of Bacillus thuringiensis serovar. tenebrionis and Beauveria bassiana for biologically-based integrated pest management of Colorado potato beetle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Research conducted over the past decade has indicated a low level of synergism and potentially high degree of complementarity between Bacillus thuringiensis (Bt)- and Beauveria bassiana (Bb)-based biopesticides applied for management of the Colorado potato beetle, Leptinotarsa decemlineata. In view...

  5. Genome Sequence of the Mosquitocidal Bacillus thuringiensis Strain BR58, a Biopesticide Product Effective against the Coffee Berry Borer (Hypothenemus hampei)

    PubMed Central

    Zorzetti, Janaina; Ricietto, Ana P. S.; da Silva, Carlos R. M.; Wolf, Ivan R.; Neves, Pedro M. O. J.; Meneguim, Ana M.; Vilas-Boas, Laurival A.

    2015-01-01

    Bacillus thuringiensis is an important microbial control agent against insect pests. The draft genome sequence of the Brazilian strain BR58 described here contains the insecticidal genes cry4A, cry4B, cry10A, cry11A, cry60A, cry60B, and cyt1A, which show toxicity to both Aedes aegypti and Hypothenemus hampei larvae. PMID:26659669

  6. Cross-Resistance Responses of Cry1Ac-Selected Heliothis virescens (Lepidoptera: Noctuidae) to the Bacillus thuringiensis Protein Vip3A

    Technology Transfer Automated Retrieval System (TEKTRAN)

    One susceptible and three Cry1Ac-resistant strains of tobacco budworm, Heliothis virescens (F.), were used in laboratory studies to determine the level of cross-resistance between the Bacillus thuringiensis (Berliner) toxins Cry1Ac and Vip3A using concentration-mortality and leaf tissue experiments....

  7. Potential Prepore Trimer Formation by the Bacillus thuringiensis Mosquito-specific Toxin

    PubMed Central

    Sriwimol, Wilaiwan; Aroonkesorn, Aratee; Sakdee, Somsri; Kanchanawarin, Chalermpol; Uchihashi, Takayuki; Ando, Toshio; Angsuthanasombat, Chanan

    2015-01-01

    The insecticidal feature of the three-domain Cry δ-endotoxins from Bacillus thuringiensis is generally attributed to their capability to form oligomeric pores, causing lysis of target larval midgut cells. However, the molecular description of their oligomerization process has not been clearly defined. Here a stable prepore of the 65-kDa trypsin-activated Cry4Ba mosquito-specific toxin was established through membrane-mimetic environments by forming an ∼200-kDa octyl-β-d-glucoside micelle-induced trimer. The SDS-resistant trimer caused cytolysis to Sf9 insect cells expressing Aedes-mALP (a Cry4Ba receptor) and was more effective than a toxin monomer in membrane perturbation of calcein-loaded liposomes. A three-dimensional model of toxin trimer obtained by negative-stain EM in combination with single-particle reconstruction at ∼5 nm resolution showed a propeller-shaped structure with 3-fold symmetry. Fitting the three-dimensional reconstructed EM map with a 100-ns molecular dynamics-simulated Cry4Ba structure interacting with an octyl-β-d-glucoside micelle showed relative positioning of individual domains in the context of the trimeric complex with a major protrusion from the pore-forming domain. Moreover, high-speed atomic force microscopy imaging at nanometer resolution and a subsecond frame rate demonstrated conformational transitions from a propeller-like to a globularly shaped trimer upon lipid membrane interactions, implying prepore-to-pore conversion. Real-time trimeric arrangement of monomers associated with l-α-dimyristoylphosphatidylcholine/3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonic acid bicelle membranes was also envisaged by successive high-speed atomic force microscopy imaging, depicting interactions among three individual subunits toward trimer formation. Together, our data provide the first pivotal insights into the structural requirement of membrane-induced conformational changes of Cry4Ba toxin monomers for the

  8. Compatibility of Bacillus thuringiensis serovar israelensis and chemical insecticides for the control of Aedes mosquitoes.

    PubMed

    Seleena, P; Lee, H L; Chiang, Y F

    1999-12-01

    The compatibility of the commercial aqueous Bacillus thuringiensis serovar israelensis (B.t.i.) formulation, Vectobac 12AS, with the chemical insecticides Actellic 50EC, Aqua Resigen, Resigen, and Fendona SC, for the simultaneous control of Aedes larvae and adults was studied by dispersing nine different formulations using a portable mist blower, in single story half-brick houses. The effectiveness of the treatment was evaluated by measuring the larval mortality, adult mortality, and droplet analysis at varying distances from the sprayer. Persistence of the larvicidal activity of the chemical insecticides and B.t.i was also determined by measuring the larval mortality in the test samples 7 days posttreatment. The sprayed particles in all the trials were 50-60 microns in size, indicating that the particles were those of mist spray. Test samples placed within 3 m from the sprayer gave the maximum larval and adult mortality. Chemical insecticides exhibited maximum larval mortality in the 1 h posttreatment test samples and it was comparable to the larvicidal activity of B.t.i. The larvicidal toxins of B.t.i were more stable and were able to affect sufficient larval mortality for 7 days posttreatment. The larvicidal activity of the mixtures, i.e., chemical insecticides with B.t.i, in the 1 h posttreatment test samples was not significantly different from the larvicidal activity of the chemical insecticides and it was comparable to the larvicidal activity of B.t.i alone. However, the larvicidal activity of the mixtures was significantly more than the chemical insecticides alone in the 7 days posttreatment test samples except for the Actellic 50EC and Vectobac 12AS mixture. In all the trials, with or without B.t.i, there was no significant difference in adult mortality, indicating that this B.t.i formulation, Vectobac 12AS, was not antagonistic to the adulticidal activity of the chemical insecticides. From this study, it can be concluded that chemical insecticides can be

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

    PubMed

    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

  10. Safety and advantages of Bacillus thuringiensis-protected plants to control insect pests.

    PubMed

    Betz, F S; Hammond, B G; Fuchs, R L

    2000-10-01

    Plants modified to express insecticidal proteins from Bacillus thuringiensis (referred to as Bt-protected plants) provide a safe and highly effective method of insect control. Bt-protected corn, cotton, and potato were introduced into the United States in 1995/1996 and grown on a total of approximately 10 million acres in 1997, 20 million acres in 1998, and 29 million acres globally in 1999. The extremely rapid adoption of these Bt-protected crops demonstrates the outstanding grower satisfaction of the performance and value of these products. These crops provide highly effective control of major insect pests such as the European corn borer, southwestern corn borer, tobacco budworm, cotton bollworm, pink bollworm, and Colorado potato beetle and reduce reliance on conventional chemical pesticides. They have provided notably higher yields in cotton and corn. The estimated total net savings to the grower using Bt-protected cotton in the United States was approximately $92 million in 1998. Other benefits of these crops include reduced levels of the fungal toxin fumonisin in corn and the opportunity for supplemental pest control by beneficial insects due to the reduced use of broad-spectrum insecticides. Insect resistance management plans are being implemented to ensure the prolonged effectiveness of these products. Extensive testing of Bt-protected crops has been conducted which establishes the safety of these products to humans, animals, and the environment. Acute, subchronic, and chronic toxicology studies conducted over the past 40 years establish the safety of the microbial Bt products, including their expressed insecticidal (Cry) proteins, which are fully approved for marketing. Mammalian toxicology and digestive fate studies, which have been conducted with the proteins produced in the currently approved Bt-protected plant products, have confirmed that these Cry proteins are nontoxic to humans and pose no significant concern for allergenicity. Food and feed derived

  11. Characterization of a Bacillus thuringiensis strain collection isolated from diverse Costa Rican natural ecosystems.

    PubMed

    Arrieta, Glen; Espinoza, Ana M

    2006-03-01

    Costa Rican natural ecosystems are among the most diverse in the world. For this reason, we isolated strains of the entomopathogenic bacteria Bacillus thuringiensis (Bt) to determine their diversity, distribution and abundance. A total of 146 Bt strains were obtained from environmental samples collected from diverse natural ecosystems and life zones of Costa Rica. We recovered Bt strains from 71%, 63%, 61% and 54% of soil samples, fresh leaves, other substrates and leaf litter respectively. Bt was isolated in 65% of the samples collected in the humid tropical forest in national parks (Braulio Carrillo, Gandoca Manzanillo, Sierpe, Hitoy Cerere, and Cahuita), and in 59% of the samples collected in the dry tropical forest (Parque Nacional Marino las Baulas, Palo Verde and Santa Rosa). In the very humid tropical forest (Tortuguero) Bt was isolated in 75% of the samples and in the very humid tropical forest transition perhumid (Carara) it was found in 69% of the samples. The strains exhibit a diverse number, size and morphology of parasporal inclusion bodies: irregular (47%), oval (20%), bipyramidal (3%), bipyramidal and cubic (1%), bipyramidal, oval and irregular (5%) and bipyramidal, oval and cubic crystals (2%). Strains isolated from Braulio Carrillo, Tortuguero and Cahuita, presented predominantly irregular crystals. On the other hand, more than 60% of the isolates from Térraba-Sierpe and Hitoy-Cerere had medium oval crystals. Strains from Gandoca-Manzanillo, Palo Verde and Carara presented mainly combinations of oval and irregular crystals. Nevertheless, the greatest diversity in crystal morphology was observed in those from Santa Rosa, Llanos del Rio Medio Queso and Parque Marino las Baulas. Protein analyses of the crystal-spore preparations showed delta-endotoxin with diverse electrophoretic patterns, with molecular weights in the range of 20 to 160 kDa. Fifty six percent of the strains amplified with the cry2 primer, 54% with vip3, 20% with cry1, 9% with cry3

  12. Secondary effects of the pesticide Bacillus thuringiensis kurstaki on chicks of spruce grouse (Dendragapus canadensis).

    PubMed

    Norton, M L; Bendell, J F; Bendell-Young, L I; LeBlanc, C W

    2001-10-01

    The objective of this study was to determine if application of the biological pesticide Bacillus thuringiensis kurstaki (B.t.k.) had secondary effects on spruce grouse (Dendragapus canadensis) chicks in the southern boreal forest of Ontario, Canada. B.t.k. kills larvae of Lepidoptera that might otherwise be food. This may affect types of food ingested and influence chick growth rates by reducing the availability of a high-protein food. Amount of ingested grit, which is diet-dependent, may also be influenced, which could alter exposure to anthropogenic toxic metals, such as cadmium (Cd) and lead (Pb), associated with the grit. Chick growth rate from 0 to 14 days of age was determined, and gizzard contents were analyzed for types of foods and levels of macronutrients and trace metals in chicks raised in a plot treated with B.t.k. compared to chicks raised on a control plot. Chick growth rates were significantly different (p < 0.05); chicks from the treatment plot increased body mass 30% less as compared to chicks raised on the control plot. Control chicks contained on average 2.5-fold the mass (in g) of grit and nearly twofold the mass of food, which included larvae, in their gizzards compared to those exposed to B.t.k. whose diets were dominated by spiders and ants. Concentrations of magnesium (Mg), calcium (Ca), phosphorous (P), iron (Fe), manganese (Mn), sulfur (S) and the trace metals zinc (Zn), copper (Cu), nickel (Ni), and chromium (Cr) did not differ between experimental and control birds (p > 0.05). In contrast, control chicks had higher levels of aluminum (Al), Cd, cobalt (Co), and Pb in their gizzards than chicks from the sprayed area (p < 0.05), attributed to greater amounts of consumed grit. Although birds from the sprayed area had lower levels of the toxic metals Pb and Cd in their gizzards, a protein-deficient diet could have been a factor that contributed to decreased rates of growth in B.t.k.-exposed birds as compared to those birds held on nonsprayed

  13. Immune responses in farm workers after exposure to Bacillus thuringiensis pesticides.

    PubMed Central

    Bernstein, I L; Bernstein, J A; Miller, M; Tierzieva, S; Bernstein, D I; Lummus, Z; Selgrade, M K; Doerfler, D L; Seligy, V L

    1999-01-01

    Although health risks to pesticides containing Bacillus thuringiensis (Bt) have been minimal, the potential allergenicity of these organisms has not been evaluated. Therefore, a health survey was conducted in farm workers before and after exposure to Bt pesticides. Farm workers who picked vegetables that required Bt pesticide spraying were evaluated before the initial spraying operation (n = 48) and 1 and 4 months after (n = 32 and 20, respectively). Two groups of low- (n = 44) and medium- (n = 34) exposure workers not directly exposed to Bt spraying were also assessed. The investigation included questionnaires, nasal/mouth lavages, ventilatory function assessment, and skin tests to indigenous aeroallergens and to a variety of Bt spore and vegetative preparations. To authenticate exposure to the organism present in the commercial preparation, isolates from lavage specimens were tested for Bt genes by DNA-DNA hybridization. Humoral immunoglobulin G (IgG) and immunoglobulin E (IgE) antibody responses to spore and vegetative Bt extracts were assayed. There was no evidence of occupationally related respiratory symptoms. Positive skin-prick tests to several spore extracts were seen chiefly in exposed workers. In particular, there was a significant (p < 0.05) increase in the number of positive skin tests to spore extracts 1 and 4 months after exposure to Bt spray. The number of positive skin test responses was also significantly higher in high (p < 0.05) than in low- or medium-exposure workers. The majority of nasal lavage cultures from exposed workers was positive for the commercial Bt organism, as demonstrated by specific molecular genetic probes. Specific IgE antibodies were present in more high-exposure workers (p < 0.05) than in the low and medium groups. Specific IgG antibodies occurred more in the high (p < 0.05) than in the low-exposure group. Specific IgG and IgE antibodies to vegetative organisms were present in all groups of workers. Exposure to Bt sprays may

  14. Interactions of Bacillus thuringiensis Cry1Ac toxin in genetically engineered cotton with predatory heteropterans.

    PubMed

    Torres, Jorge B; Ruberson, John R

    2008-06-01

    A number of cotton varieties have been genetically transformed with genes from Bacillus thuringiensis (Bt) to continuously produce Bt endotoxins, offering whole plant and season-long protection against many lepidopteran larvae. Constant whole-plant toxin expression creates a significant opportunity for non-target herbivores to acquire and bio-accumulate the toxin for higher trophic levels. In the present study we investigated movement of Cry1Ac toxin from the transgenic cotton plant through specific predator-prey pairings, using omnivorous predators with common cotton pests as prey: (1) the beet armyworm, Spodoptera exigua (Lepidoptera: Noctuidae), with the predator Podisus maculiventris (Heteroptera: Pentatomidae); (2) the two-spotted spider mite, Tetranychus urticae (Acarina: Tetranychidae), with the predatory big-eyed bug Geocoris punctipes (Heteroptera: Geocoridae) and (3) with the predatory damsel bug Nabis roseipennis (Heteropera: Nabidae); and (4) the thrips Frankliniella occidentalis (Thysanoptera: Thripidae) with the predatory pirate bug Orius insidiosus (Heteroptera: Anthocoridae). We quantified Cry1Ac toxin in the cotton plants, and in the pests and predators, and the effects of continuous feeding on S. exigua larvae fed either Bt or non-Bt cotton on life history traits of P. maculiventris. All three herbivores were able to convey Cry1Ac toxin to their respective predators. Among the herbivores, T. urticae exhibited 16.8 times more toxin in their bodies than that expressed in Bt-cotton plant, followed by S. exigua (1.05 times), and F. occidentalis immatures and adults (0.63 and 0.73 times, respectively). Of the toxin in the respective herbivorous prey, 4, 40, 17 and 14% of that amount was measured in the predators G. punctipes, P. maculiventris, O. insidiosus, and N. roseipennis, respectively. The predator P. maculiventris exhibited similar life history characteristics (developmental time, survival, longevity, and fecundity) regardless of the prey's food

  15. Use of Bacillus thuringiensis Toxins for Control of the Cotton Pest Earias insulana (Boisd.) (Lepidoptera: Noctuidae)

    PubMed Central

    Ibargutxi, María A.; Estela, Anna; Ferré, Juan; Caballero, Primitivo

    2006-01-01

    Thirteen of the most common lepidopteran-specific Cry proteins of Bacillus thuringiensis have been tested for their efficacy against newly hatched larvae of two populations of the spiny bollworm, Earias insulana. At a concentration of 100 μg of toxin per milliliter of artificial diet, six Cry toxins (Cry1Ca, Cry1Ea, Cry1Fa, Cry1Ja, Cry2Aa, and Cry2Ab) were not toxic at all. Cry1Aa, Cry1Ja, and Cry2Aa did not cause mortality but caused significant inhibition of growth. The other Cry toxins (Cry1Ab, Cry1Ac, Cry1Ba, Cry1Da, Cry1Ia, and Cry9Ca) were toxic to E. insulana larvae. The 50% lethal concentration values of these toxins ranged from 0.39 to 21.13 μg/ml (for Cry9Ca and Cry1Ia, respectively) for an E. insulana laboratory colony originating from Egypt and from 0.20 to 4.25 μg/ml (for Cry9Ca and Cry1Da, respectively) for a laboratory colony originating from Spain. The relative potencies of the toxins in the population from Egypt were highest for Cry9Ca and Cry1Ab, and they were both significantly more toxic than Cry1Ac and Cry1Ba, followed by Cry1Da and finally Cry1Ia. In the population from Spain, Cry9Ca was the most toxic, followed in decreasing order by Cry1Ac and Cry1Ba, and the least toxic was Cry1Da. Binding experiments were performed to test whether the toxic Cry proteins shared binding sites in this insect. 125I-labeled Cry1Ac and Cry1Ab and biotinylated Cry1Ba, Cry1Ia, and Cry9Ca showed specific binding to the brush border membrane vesicles from E. insulana. Competition binding experiments among these toxins showed that only Cry1Ab and Cry1Ac competed for the same binding sites, indicating a high possibility that this insect may develop cross-resistance to Cry1Ab upon exposure to Cry1Ac transgenic cotton but not to the other toxins tested. PMID:16391075

  16. Efficacy of olive mill wastewater for protecting Bacillus thuringiensis formulation from UV radiations.

    PubMed

    Jallouli, Wafa; Sellami, Sameh; Sellami, Maissa; Tounsi, Slim

    2014-12-01

    The effectiveness of 10 low-cost UV-absorbers in protecting Bacillus thuringiensis subsp. kurstaki BLB1 toxins against inactivation by UV-A and UV-B irradiation was evaluated in this study. Among them, two by-products, molasses and olive mill wastewater (OMW) were selected for further studies. They were tested at different concentrations of 0.05, 0.1, 0.15 and 0.2% using the para-aminobenzoic acid (PABA) as a common UV protectant. Interestingly, addition of PABA and OMW to BLB1 formulations was found to be most effective in protecting BLB1 spores at 90.8 and 76.4% respectively and in preserving delta-endotoxin concentration at a level of 81.7 and 72.2%, respectively when used at a concentration of 0.2%. The lowest preserved spores (46.3%) and delta-endotoxin level (12.4%) was found using molasses. In contrast, spore count and delta-endotoxin concentration were completely reduced after an exposure of unprotected Bt strain BLB1 to UV radiations up to 96h. SDS-PAGE analysis of protected and unprotected samples revealed that delta-endotoxin bands (130, 65-70kDa) were conserved until 96h of UV exposure in presence of PABA or OMW compared with their disappearance in presence of molasses after 72h of exposure and their dramatically decline from 8h of exposure in unprotected mixture. A complete loss of larvicidal toxicity against Ephestia kuehniella was found after 24h of exposure in absence of any UV-absorber. Addition of OMW or PABA offered the highest levels of insecticidal activity with 63.2 and 74.7% of residual toxicity, respectively. Whereas, molasses addition, as UV protectant retained only 26.3% of residual activity after 96h of exposure. Therefore, addition of OMW by-product to Bt formulation may be a suitable alternative to others synthetic chemical compounds. OMW may also provided added value, be environmentally friendly and less hazardous, when used at low concentration. PMID:25093915

  17. Integrative Model for Binding of Bacillus thuringiensis Toxins in Susceptible and Resistant Larvae of the Diamondback Moth (Plutella xylostella)

    PubMed Central

    Ballester, Victoria; Granero, Francisco; Tabashnik, Bruce E.; Malvar, Thomas; Ferré, Juan

    1999-01-01

    Insecticidal crystal proteins from Bacillus thuringiensis in sprays and transgenic crops are extremely useful for environmentally sound pest management, but their long-term efficacy is threatened by evolution of resistance by target pests. The diamondback moth (Plutella xylostella) is the first insect to evolve resistance to B. thuringiensis in open-field populations. The only known mechanism of resistance to B. thuringiensis in the diamondback moth is reduced binding of toxin to midgut binding sites. In the present work we analyzed competitive binding of B. thuringiensis toxins Cry1Aa, Cry1Ab, Cry1Ac, and Cry1F to brush border membrane vesicles from larval midguts in a susceptible strain and in resistant strains from the Philippines, Hawaii, and Pennsylvania. Based on the results, we propose a model for binding of B. thuringiensis crystal proteins in susceptible larvae with two binding sites for Cry1Aa, one of which is shared with Cry1Ab, Cry1Ac, and Cry1F. Our results show that the common binding site is altered in each of the three resistant strains. In the strain from the Philippines, the alteration reduced binding of Cry1Ab but did not affect binding of the other crystal proteins. In the resistant strains from Hawaii and Pennsylvania, the alteration affected binding of Cry1Aa, Cry1Ab, Cry1Ac, and Cry1F. Previously reported evidence that a single mutation can confer resistance to Cry1Ab, Cry1Ac, and Cry1F corresponds to expectations based on the binding model. However, the following two other observations do not: the mutation in the Philippines strain affected binding of only Cry1Ab, and one mutation was sufficient for resistance to Cry1Aa. The imperfect correspondence between the model and observations suggests that reduced binding is not the only mechanism of resistance in the diamondback moth and that some, but not all, patterns of resistance and cross-resistance can be predicted correctly from the results of competitive binding analyses of susceptible

  18. Persistence of Bt Bacillus thuringiensis Cry1Aa toxin in various soils determined by physicochemical reactions

    NASA Astrophysics Data System (ADS)

    Helassa, N.; Noinville, S.; Déjardin, P.; Janot, J. M.; Quiquampoix, H.; Staunton, S.

    2009-04-01

    Insecticidal Cry proteins from the soil bacterium, Bacillus thuringiensis (Bt) are produced by a class of genetically modified (GM) crops, and released into soils through root exudates and upon decomposition of residues. In contrast to the protoxin produced by the Bacillus, the protein produced in GM crops does not require activation in insect midguts and thereby potentially looses some of its species specificity. Although gene transfer and resistance emergence phenomena are well documented, the fate of these toxins in soil has not yet been clearly elucidated. Cry proteins, in common with other proteins, are adsorbed on soils and soil components. Adsorption on soil, and the reversibility of this adsorption is an important aspect of the environmental behaviour of these toxins. The orientation of the molecule and conformational changes on surfaces may modify the toxicity and confer some protection against microbial degradation. Adsorption will have important consequences for both the risk of exposition of non target species and the acquisition of resistance by target species. We have adopted different approaches to investigate the fate of Cry1Aa in soils and model minerals. In each series of experiments we endeavoured to maintain the protein in a monomeric form (pH above 6.5 and a high ionic strength imposed with 150 mM NaCl). The adsorption and the desorbability of the Cry1Aa Bt insecticidal protein were measured on two different homoionic clays: montmorillonite and kaolinite. Adsorption isotherms obtained followed a low affinity interaction for both clays and could be fitted using the Langmuir equation. Binding of the toxin decreased as the pH increased from 6.5 (close to the isoelectric point) to 9. Maximum adsorption was about 40 times greater on montmorillonite (1.71 g g-1) than on kaolinite (0.04 g g-1) in line with the contrasting respective specific surface areas of the minerals. Finally, some of the adsorbed toxin was desorbed by water and more, about 36

  19. Impact of UV radiation on activity of linear furanocoumarins and Bacillus thuringiensis var. kurstaki against Spodoptera exigua: Implications for tritrophic interactions

    SciTech Connect

    Trumble, J.T.; Moar, W.J.; Brewer, M.J.; Carson, W.G. )

    1991-05-01

    Acidic fogs with a pH of 2.0 and duration of 2 hr did not reduce the efficacy of Bacillus thuringiensis var. kurstaki. Therefore, the impact of UV radiation was investigated on the interactions between (1) levels of the antibacterial linear furanocoumarins psoralen, bergapten, and xanthotoxin in Apium graveolens (L.) occurring following a 2.0 pH acidic fog episode, (2) the noctuid Spodoptera exigua, and (3) a sublethal dosage of the microbial pathogen B. thuringiensis var. kurstaki. Mean time to pupation in the absence of UV radiation was significantly extended by the addition of either psoralens or B. thuringiensis. Larvae developing on diets containing B. thuringiensis plus psoralens required nearly 40% longer to pupate than controls, but their effects were additive as the interaction was not significant. Mean time to mortality, a weighted average time of death, was not significantly affected by any of the treatments. In a 2 {times} 2 {times} 2 factorial analysis, all main effects reduced survival significantly, as did the three-way interaction. Thus, antagonistic interactions with psoralens that would reduce the effectiveness of B. thuringiensis in the field were not observed. When pairs of main effects were nested within the two levels of the third factor, several two-way interactions were found. Interestingly, the activity of B. thuringiensis and the psoralens, individually or in combination, was enhanced by exposure to UV radiation. Implications of this research are discussed for both natural and agricultural ecosystems.

  20. Optimization of process parameters for maximum poly(-beta-)hydroxybutyrate (PHB) production by Bacillus thuringiensis IAM 12077.

    PubMed

    Pal, Adwitiya; Prabhu, Ashwini; Kumar, Avinash Arun; Rajagopal, Badri; Dadhe, Kajal; Ponnamma, Vomsi; Shivakumar, Srividya

    2009-01-01

    The study aimed at screening and identifying a potential poly-beta-hydroxybutyrate (PHB) accumulating Bacillus strain and optimization of media parameters for increased PHB production by the strain. A Gram-positive bacterium that accumulated PHB was isolated from local garden soil of Bangalore. Based on morphological and physiological properties, and nucleotide sequence (about 1.5 kb) of its 16S rDNA it was identified as Bacillus thuringiensis IAM 12077. PHB production was found to be comparable to most of the Bacillus sp. reported to date. PHB production by this strain was dependent on nutrient limitation. Cell dry weight and PHB accumulation increased significantly under biphasic growth condition (from nutrient broth to nitrogen-deficient medium) as compared with growth in nutrient broth alone (from 0.32 g/l to 2.76 g/l cell dry weight; 24% to 43.37% PHB accumulation; 0.2 g/l to 1.2 g/l PHB production), with maximum accumulation at 24 h in nitrogen-deficient medium. Time course study of growth and PHB production by this strain in the nitrogen deficient medium showed that PHB production was associated with the stationary phase of growth. All the tested media containing different carbon and nitrogen sources supported growth and PHB production. Ultraviolet spectrum of the extracted polymer showed a characteristic peak at 235 nm. PMID:19824399