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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...

    Science.gov (United States)

    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 PROTECTANTS...

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

    Science.gov (United States)

    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. Copyright © 2016 Elsevier Inc. All rights reserved.

  3. Baseline sensitivity of maize borers in India to the Bacillus thuringiensis insecticidal proteins Cry1A.105 and Cry2Ab2.

    Science.gov (United States)

    Jalali, Sushil K; Yadavalli, Lalitha; Ojha, Rakshit; Kumar, Pradyumn; Sulaikhabeevi, Suby B; Sharma, Reema; Nair, Rupa; Kadanur, Ravi C; Kamath, Subray P; Komarlingam, Mohan S

    2015-08-01

    Among the major pests of maize in India are two stem borers, Chilo partellus (Swinhoe) and Sesamia inferens (Walker), and an earworm, Helicoverpa armigera (Hübner). As a pest control strategy, transgenic Bacillus thuringiensis (Bt) maize hybrids are undergoing regulatory trials in India. We have determined the sensitivity of the target lepidopterans to the insecticidal Bt proteins expressed in Bt maize, as this determines product efficacy and the resistance management strategy to be adopted. Maize hybrids with event MON89034 express two insecticidal Bt proteins, Cry1A.105 and Cry2Ab2. Sensitivity profiles of 53 populations of C. partellus, 21 populations of S. inferens and 21 populations of H. armigera, collected between 2008 and 2013 from maize-growing areas in India, to Cry1A.105 and Cry2Ab2 proteins were generated through dose-response assays. Cry1A.105 protein was the most effective to neonates of C. partellus (mean MIC90 range 0.30-1.0 µg mL(-1) ) and H. armigera (mean MIC90 range 0.71-8.22 µg mL(-1) ), whereas Cry2Ab2 (mean MIC90 range 0.65-1.70 µg mL(-1) ) was the most effective to S. inferens. Populations of C. partellus, S. inferens and H. armigera were susceptible to the Bt proteins Cry1A.105 and Cry2Ab2. The Bt sensitivity data will serve as precommercialisation benchmarks for resistance monitoring purposes. © 2014 Society of Chemical Industry.

  4. Estimation of resistance allele frequency to maize incorporated Bacillus thuringiensis Cry2Ab2 protein in field populations of the fall army Spodoptera frugiperda (Lepidoptera: Noctuidae) from south region of the United State

    Science.gov (United States)

    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 Americas. In the falls of 2013 and 2014, a total of 215 F2 two-parent families of S. frugiperda were es...

  5. Cross-resistance to purified Bt proteins, Bt corn and Bt cotton in a Cry2Ab2-corn resistant strain of Spodoptera frugiperda.

    Science.gov (United States)

    Yang, Fei; Kerns, David L; Head, Graham P; Price, Paula; Huang, Fangneng

    2017-12-01

    Gene-pyramiding by combining two or more dissimilar Bacillus thuringiensis (Bt) proteins into a crop has been used to delay insect resistance. The durability of gene-pyramiding can be reduced by cross-resistance. Fall armyworm, Spodoptera frugiperda, is a major target pest of the Cry2Ab2 protein used in pyramided Bt corn and cotton. Here, we provide the first experimental evaluation of cross-resistance in S. frugiperda selected with Cry2Ab2 corn to multiple Bt sources including purified Bt proteins, Bt corn and Bt cotton. Concentration - response bioassays showed that resistance ratios for Cry2Ab2-resistant (RR) relative to Cry2Ab2-susceptible (SS) S. frugiperda were -1.4 for Cry1F, 1.2 for Cry1A.105, >26.7 for Cry2Ab2, >10.0 for Cry2Ae and -1.1 for Vip3A. Larvae of Cry2Ab2-heterozygous (RS), SS and RR S. frugiperda were all susceptible to Bt corn and Bt cotton containing Cry1 (Cry1F or Cry1A.105) and/or Vip3A proteins. Pyramided Bt cotton containing Cry1Ac + Cry2Ab2 or Cry1Ab + Cry2Ae were also effective against SS and RS, but not RR. These findings suggest that Cry2Ab2-corn-selected S. frugiperda is not cross-resistant to Cry1F, Cry1A.105 or Vip3A protein, or corn and cotton plants containing these Bt proteins, but it can cause strong cross-resistance to Cry2Ae and Bt crops expressing similar Bt proteins. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  6. Current research efforts with Bacillus thuringiensis

    Science.gov (United States)

    Normand R. Dubois

    1991-01-01

    The bioassay of 260 strains of Bacillus thuringiensis (Bt) and 70 commercial preparations show that regression coefficient estimates may be as critical as LC5O estimates when evaluating them for future consideration.

  7. Manejo de lepidópteros-praga na cultura do milho com o evento Bt piramidado Cry1A.105 e Cry2Ab2

    Directory of Open Access Journals (Sweden)

    José Magid Waquil

    2013-12-01

    Full Text Available O objetivo deste trabalho foi avaliar a eficácia do evento piramidado (MON 89034, que expressa as proteínas Cry1A.105 e Cry2Ab2, no controle dos principais lepidópteros-praga da cultura do milho no Brasil, Spodoptera frugiperda, Helicoverpa spp. e Diatraea saccharalis. Os ensaios foram conduzidos em quatro regiões do país, com o híbrido DKB 390, submetido a seis tratamentos: híbrido com o evento piramidado, híbrido com o evento que expressa apenas a proteína Cry1A(b (MON 810 e híbrido convencional (não Bt, todos com e sem manejo integrado de S. frugiperda. Para o evento piramidado, não foi necessário o controle químico em nenhum dos locais avaliados. Diferenças significativas foram observadas entre os tratamentos quanto aos danos e à presença de lagartas. Em geral, essas variáveis foram mais baixas no híbrido com o evento piramidado e mais altas no híbrido convencional, sem controle químico. Sob alta infestação, o controle químico reduziu os danos causados por S. frugiperda e D. saccharalis, tanto no evento que expressa apenas uma proteína, como no híbrido convencional. Com base nos danos causados pelos insetos, o evento piramidado Cry1A.105 e Cry2Ab2 é eficiente no controle dos principais lepidópteros-pragas do milho no Brasil.

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

    African Journals Online (AJOL)

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

  9. Bacillus thuringiensis and its application in agriculture

    African Journals Online (AJOL)

    USER

    2010-04-05

    Apr 5, 2010 ... Key words: Bacillus thuringiensis, endotoxins, crop plants. INTRODUCTION ..... of resistance in the pest and unfavorable interactions with beneficial .... with slower resistance evolution in North Carolina compared to .... level of 0.18% cross pollination in the experimental rice lines. .... Ecology and Safety.

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

    African Journals Online (AJOL)

    Bioassays were used to test the insecticidal activity of B. thuringiensis strains ... of crystal protein genes, 7 tested positive for cry 4, cry 11, and cyt toxin genes. ... mosquitocidal cry and cyt genes in Bacillus thuringiensis subsp. israelensis.

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

    International Nuclear Information System (INIS)

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

    2009-01-01

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

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

    African Journals Online (AJOL)

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

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

    African Journals Online (AJOL)

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

  14. BACILLUS THURINGIENSIS ELASTASES WITH INSECTICIDE ACTIVITY

    Directory of Open Access Journals (Sweden)

    E. V. Matseliukh

    2015-10-01

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

  15. Efektivitas Bacillus thuringiensis dalam Pengendalian Larva Nyamuk Anopheles sp.

    Directory of Open Access Journals (Sweden)

    Citra Inneke Wibowo

    2017-08-01

    Full Text Available Nyamuk Anopheles sp adalah vektor penyakit malaria. Pengendalian vektor penyakit malaria dapat dilakukan secara biologis yaitu dengan menggunakan Bacillus thuringiensis. Tujuan penelitian adalah untuk mengetahui efektivitas konsentrasi Bacillus thuringiensis dalam pengendalian larva nyamuk Anopheles sp.Penelitian ini dilakukan secara eksperimental menggunakan Rancangan Acak Lengkap Faktorial (RAL Faktorial yang terdiri atas dua faktor yaitu konsentrasi Bacillus thuringiensis dan stadia larva Anopheles dengan pengulangan tiga kali.Perlakuan yang dicobakan adalahkonsentrasi Bacillus thuringiensis (A yang terdiri atas 5 taraf:A0: konsentrasi B.thuringiensis 0 CFU.mL-1, A1: konsentrasi B.thuringiensis 102 CFU.mL-1, A2: konsentrasi B.thuringiensis 104 CFU.mL-1, A3: konsentrasi B.thuringiensis 106CFU.mL-1, A4: konsentrasi B.thuringiensis 108CFU.mL-1. Perlakuan tahapan instar larva Anopheles sp. (B adalah sebagai berikut:B1: stadia larva instar I, B2: stadia larva instar II, B3: stadia larva instar III, B4: stadia larva instar IVsehingga terdapat 60 satuan percobaan. Hasil penelitian  menunjukkan konsentrasi B. thuringiensis isolat CK dan IPB CC yang paling berpengaruh dalam pengendalian larva Anopheles sp adalah 108 CFU.mL-1 . Instar larva yang paling peka terhadap B. thuringiensis isolat IPB CC adalah instar I dan II sedangkan instar yang peka terhadap isolat CK adalah instar II, Perlakuan konsentrasi isolat B. thuringiensis dan tingkat instar larva yang paling baik dalam pengendalian larva Anopheles sp. adalah 108 CFU.mL-1, dan instar I dan II.

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

    African Journals Online (AJOL)

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

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

    African Journals Online (AJOL)

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

  18. A parasporin from Bacillus thuringiensis native to Peninsular India ...

    Indian Academy of Sciences (India)

    Thomas Chubicka

    2018-05-03

    May 3, 2018 ... Apoptosis; Bacillus thuringiensis; crystal protein; cytotoxicity; ... It acts by creating pores in the intestinal duct ... however diverse types of mechanisms of action have been ... parasporins that can be utilized in the cancer drug.

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

    Directory of Open Access Journals (Sweden)

    Lessandra Heck Paes Leme Ferreira

    2003-02-01

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

  20. Larval development of Spodoptera eridania and Spodoptera frugiperda fed on fresh ear of field corn expressing the Bt proteins (Cry1F and Cry1F + Cry1A.105 + Cry2Ab2)

    OpenAIRE

    Bortolotto,Orcial Ceolin; Bueno,Adeney de Freitas; Queiroz,Ana Paula de; Silva,Gabriela Vieira

    2016-01-01

    ABSTRACT: The objective of this study was to evaluate extent of larval period, larval survival (%), food consumption, and pupal biomass of Spodoptera eridania and Spodoptera frugiperda (Lepidoptera: Noctuidae ) fed on fresh ears of field corn expressing Bt proteins (Cry1F and Cry1F+Cry1A.105+Cry2Ab2). Larvae of Spodoptera spp. survived less than two days when they consumed Bt corncobs and showed 100% mortality. Spodoptera eridania reared on non-Bt corn cobs showed higher larval development (...

  1. Larval development of Spodoptera eridania and Spodoptera frugiperda fed on fresh ear of field corn expressing the Bt proteins (Cry1F and Cry1F + Cry1A.105 + Cry2Ab2

    Directory of Open Access Journals (Sweden)

    Orcial Ceolin Bortolotto

    Full Text Available ABSTRACT: The objective of this study was to evaluate extent of larval period, larval survival (%, food consumption, and pupal biomass of Spodoptera eridania and Spodoptera frugiperda (Lepidoptera: Noctuidae fed on fresh ears of field corn expressing Bt proteins (Cry1F and Cry1F+Cry1A.105+Cry2Ab2. Larvae of Spodoptera spp. survived less than two days when they consumed Bt corncobs and showed 100% mortality. Spodoptera eridania reared on non-Bt corn cobs showed higher larval development (21.6 days than S. frugiperda (18.4 days and lower viability (56.4% and 80.2% for S. eridania and S. frugiperda , respectively. A higher amount of corn grains was consumed by S. eridania (5.4g than by S. frugiperda (3.9g. In summary, this study demonstrated that the toxins Cry1F and Cry1F + Cry1A.105 + Cry2Ab2 expressed in fresh corn cobs contributed to protect ears of corn against S. frugiperda and the non-target pest S. eridania . However, itis important to monitor non-Bt cornfields because of the potential of both species to cause damage to ear sof corn.

  2. Larval development of Spodoptera eridania (Cramer fed on leaves of Bt maize expressing Cry1F and Cry1F + Cry1A.105 + Cry2Ab2 proteins and its non-Bt isoline

    Directory of Open Access Journals (Sweden)

    Orcial Ceolin Bortolotto

    2015-03-01

    Full Text Available This study aimed to evaluate, in controlled laboratory conditions (temperature of 25±2 °C, relative humidity of 60±10%, and 14/10 h L/D photoperiod, the larval development of Spodoptera eridania (Cramer, 1784 (Lepidoptera, Noctuidae fed with leaves of Bt maize expressing Cry1F and Cry1F + Cry1A.105 + Cry2Ab2 insecticide proteins and its non-Bt isoline. Maize leaves triggered 100% of mortality on S. eridania larvae independently of being Bt or non-Bt plants. However, it was observed that in overall Bt maize (expressing a single or pyramided protein slightly affects the larval development of S. eridania, even under reduced leaf consumption. Therefore, these results showed that Cry1F and Cry1F + Cry1A.105 + Cry2Ab2 can affect the larval development of S. eridania, although it is not a target pest of this plant; however, more research is needed to better understand this evidence. Finally, this study confirms that non-Bt maize leaves are unsuitable food source to S. eridania larvae, suggesting that they are not a potential pest in maize fields.

  3. Ultra-violet-resistant mutants of Bacillus thuringiensis

    Energy Technology Data Exchange (ETDEWEB)

    Jones, D R; Karunakaran, V [Polytechnic of Central London (UK). Faculty of Engineering and Science, School of Biological and Health Sciences; Burges, H D [Institute of Horticultural Research, Littlehampton (UK); Hacking, A J [Reading Univ. (UK). Dextra Labs.Ltd.

    1991-06-01

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

  4. Ultra-violet-resistant mutants of Bacillus thuringiensis

    International Nuclear Information System (INIS)

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

    1991-01-01

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

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

    International Nuclear Information System (INIS)

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

    1990-01-01

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

  6. Occurrence of Natural Bacillus thuringiensis Contaminants and Residues of Bacillus thuringiensis-Based Insecticides on Fresh Fruits and Vegetables

    Science.gov (United States)

    Frederiksen, Kristine; Rosenquist, Hanne; Jørgensen, Kirsten; Wilcks, Andrea

    2006-01-01

    A total of 128 Bacillus cereus-like strains isolated from fresh fruits and vegetables for sale in retail shops in Denmark were characterized. Of these strains, 39% (50/128) were classified as Bacillus thuringiensis on the basis of their content of cry genes determined by PCR or crystal proteins visualized by microscopy. Random amplified polymorphic DNA analysis and plasmid profiling indicated that 23 of the 50 B. thuringiensis strains were of the same subtype as B. thuringiensis strains used as commercial bioinsecticides. Fourteen isolates were indistinguishable from B. thuringiensis subsp. kurstaki HD1 present in the products Dipel, Biobit, and Foray, and nine isolates grouped with B. thuringiensis subsp. aizawai present in Turex. The commercial strains were primarily isolated from samples of tomatoes, cucumbers, and peppers. A multiplex PCR method was developed to simultaneously detect all three genes in the enterotoxin hemolysin BL (HBL) and the nonhemolytic enterotoxin (NHE), respectively. This revealed that the frequency of these enterotoxin genes was higher among the strains indistinguishable from the commercial strains than among the other B. thuringiensis and B. cereus-like strains isolated from fruits and vegetables. The same was seen for a third enterotoxin, CytK. In conclusion, the present study strongly indicates that residues of B. thuringiensis-based insecticides can be found on fresh fruits and vegetables and that these are potentially enterotoxigenic. PMID:16672488

  7. The colonization of Bacillus thuringiensis strains in bryophytes

    Czech Academy of Sciences Publication Activity Database

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

    2017-01-01

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

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

    African Journals Online (AJOL)

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

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

    African Journals Online (AJOL)

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

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

    African Journals Online (AJOL)

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

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

    African Journals Online (AJOL)

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

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

    African Journals Online (AJOL)

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

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

    Science.gov (United States)

    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...

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

    Science.gov (United States)

    2010-07-01

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

  15. [Potential of Bacillus thuringiensis israelensis Berliner for controlling Aedes aegypti].

    Science.gov (United States)

    Polanczyk, Ricardo Antonio; Garcia, Marcelo de Oliveira; Alves, Sérgio Batista

    2003-12-01

    The importance of the entomopathogenic bacterium Bacillus thuringiensis israelensis in the control of Aedes aegypti is presented. The use and potential of B. thuringiensis israelensis against the mosquito vector of dengue fever is described. Other aspects such as insect's resistance development against chemicals and advantages and constraints of using microbial control are discussed. Emphasis is given to the importance of the use of this bacterium in Brazil, which could contribute significantly to solving the mosquito problem without affecting the environment, humans and others invertebrate organisms in critical regions.

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

    OpenAIRE

    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

    2016-01-01

    Bacillus thuringiensis and its toxins are widely used for insect control. Notwithstanding the remarkable importance of this insect pathogen, its killing mechanism has yet to be fully elucidated. Here we show that the microbiota resident in the host midgut triggers a lethal septicemia. The infection process is enhanced by reducing the host immune response and its control on replication of midgut bacteria invading the body cavity through toxin-induced epithelial lesions. The experimental approa...

  17. Characterization of parasporin gene harboring Indian isolates of Bacillus thuringiensis

    OpenAIRE

    Lenina, N. K.; Naveenkumar, A.; Sozhavendan, A. E.; Balakrishnan, N.; Balasubramani, V.; Udayasuriyan, V.

    2013-01-01

    Bacillus thuringiensis (Bt) is popularly known as insecticidal bacterium. However, non-insecticidal Bt strains are more extensively available in natural environment than the insecticidal ones. Parasporin (PS) is a collection of genealogically heterogeneous Cry proteins synthesized in non-insecticidal isolates of Bt. An important character generally related with PS proteins is their strong cytocidal activity preferentially on human cancer cells of various origins. Identification and characteri...

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Alves Lúcia M. Carareto

    2000-01-01

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

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

    Science.gov (United States)

    Gomaa, Eman Zakaria

    2012-02-01

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

  1. TRANSGENIC PLANTS EXPRESSING BACILLUS THURINGIENSIS DELTA-ENDOTOXINS

    Institute of Scientific and Technical Information of China (English)

    Hua-rong,Li; BrendaOppert; KunYanZhu; RandallA.Higgins; Fang-nengHuang; LawrentL.Buschman

    2003-01-01

    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, modifications of the native Bt genes greatly enhanced expression levels. This is a review of the developments that made modem high-expression transgenic Bt plants possible, with an emphasis on the reasons for the low-level expression of native Bt genes in plant systems, and the techniques that have been used to improve plant expression of Bt toxin genes.

  2. Bacillus thuringiensis: the legacy to the XXI century

    Directory of Open Access Journals (Sweden)

    M. Realpe

    1998-01-01

    Full Text Available Bacillus thuringiensis-based insecticides are the main production line of the biopesticides world market. The research devoted to this area, promoted by the necessity to solve problems in agriculture and public health has resulted in an exhaustive knowledge of its biology. The diversity of the B. thuringiensis strains has permitted to develop several products mainly, but not exclusively, for insect control. With the new developments in the field of molecular biology, it has been possible to understand the molecular basis of the mode of action and to increase the range of activity as well. As a result of the broad use in several countries, resistant strains of some of the susceptible insects have appeared. The aim of this review is to elaborate a theoretical framework of the current state of research on B. thuringiensis, describing briefly the knowledge on this bacterium, with emphasis on biological phenomena that underlie its toxic activity and the problems that will be faced during the XXI century with the increasingly common resistance, all this analyzed from a biotechnological perspective.

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

    Directory of Open Access Journals (Sweden)

    Eitan Ben-Dov

    2014-03-01

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

  4. Systematic characterization of Bacillus Genetic Stock Center Bacillus thuringiensis strains using Multi-Locus Sequence Typing.

    Science.gov (United States)

    Wang, Kui; Shu, Changlong; Soberón, Mario; Bravo, Alejandra; Zhang, Jie

    2018-04-30

    The goal of this work was to perform a systematic characterization of Bacillus thuringiensis (Bt) strains from the Bacillus Genetic Stock Center (BGSC) collection using Multi-Locus Sequence Typing (MLST). Different genetic markers of 158 Bacillus thuringiensis (Bt) strains from 73 different serovars stored in the BGSC, that represented 92% of the different Bt serovars of the BGSC were analyzed, the 8% that were not analyzed were not available. In addition, we analyzed 72 Bt strains from 18 serovars available at the pubMLST bcereus database, and Bt strains G03, HBF18 and Bt185, with no H serovars provided by our laboratory. We performed a systematic MLST analysis using seven housekeeping genes (glpF, gmK, ilvD, pta, pur, pycA and tpi) and analyzed correlation of the results of this analysis with strain serovars. The 233 Bt strains analyzed were assigned to 119 STs from which 19 STs were new. Genetic relationships were established by phylogenetic analysis and showed that STs could be grouped in two major Clusters containing 21 sub-groups. We found that a significant number of STs (101 in total) correlated with specific serovars, such as ST13 that corresponded to nine Bt isolates from B. thuringiensis serovar kenyae. However, other serovars showed high genetic variability and correlated with multiple STs; for example, B. thuringiensis serovar morrisoni correlated with 11 different STs. In addition, we found that 16 different STs correlated with multiple serovars (2-4 different serovars); for example, ST12 correlated with B. thuringiensis serovar alesti, dakota, palmanyolensis and sotto/dendrolimus. These data indicated that only partial correspondence between MLST and serotyping can be established. Copyright © 2018 Elsevier Inc. All rights reserved.

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

    Science.gov (United States)

    2010-07-01

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

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

    DEFF Research Database (Denmark)

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

    2005-01-01

    Among 48,901 samples of ready-to-eat food products at the Danish retail market, 0.5% had counts of Bacillus cereus-like bacteria above 10(4) cfu g(-1). The high counts were most frequently found in starchy, cooked products, but also in fresh cucumbers and tomatoes. Forty randomly selected strains....../or content of cry genes. Thus, a large proportion of the B. cereus-like organisms present in food may belong to B. thuringiensis....

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

    Science.gov (United States)

    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...

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

    NARCIS (Netherlands)

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

    2010-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Sánchez-Yáñez Juan Manuel

    2016-08-01

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

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

    Directory of Open Access Journals (Sweden)

    Rose Gomes Monnerat

    2005-02-01

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

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

    Science.gov (United States)

    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.

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

    Directory of Open Access Journals (Sweden)

    Xavier, R.

    2007-01-01

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

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

    International Nuclear Information System (INIS)

    Majdoub, Nihed

    2010-01-01

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

  14. Evolution of Bacillus thuringiensis Cry toxins insecticidal activity.

    Science.gov (United States)

    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. © 2012 The Authors. Microbial Biotechnology © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.

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

    Directory of Open Access Journals (Sweden)

    Deise M. F Capalbo

    1995-02-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2010-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Viviane Zahner

    2013-02-01

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

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

    Science.gov (United States)

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

    2017-08-28

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

  19. Potencial de Bacillus thuringiensis israelensis Berliner no controle de Aedes aegypti Potential of Bacillus thuringiensis israelensis Berliner for controlling Aedes aegypti

    Directory of Open Access Journals (Sweden)

    Ricardo Antonio Polanczyk

    2003-12-01

    Full Text Available Relata-se a importância da bactéria entomopatogênica Bacillus thuringiensis israelensis para o controle de Aedes aegypti. São abordados a utilização e potencial de B. thuringiensis israelensis contra o mosquito vetor da dengue. Outros aspectos são discutidos como a evolução da resistência dos insetos em relação aos inseticidas químicos e as vantagens e desvantagens do controle microbiano como estratégia de controle. É dada ênfase à importância da utilização desta bactéria no Brasil como alternativa para resolver o problema em questão sem afetar o ambiente, o homem e outros vertebrados nas áreas de risco.The importance of the entomopathogenic bacterium Bacillus thuringiensis israelensis in the control of Aedes aegypti is presented. The use and potential of B. thuringiensis israelensis against the mosquito vector of dengue fever is described. Other aspects such as insect's resistance development against chemicals and advantages and constraints of using microbial control are discussed. Emphasis is given to the importance of the use of this bacterium in Brazil, which could contribute significantly to solving the mosquito problem without affecting the environment, humans and others invertebrate organisms in critical regions.

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

    Science.gov (United States)

    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.

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

    Directory of Open Access Journals (Sweden)

    Holt Jonathan

    2009-03-01

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

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

    Directory of Open Access Journals (Sweden)

    Gislayne T. Vilas-Bôas

    2005-09-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2016-01-01

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

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

    Science.gov (United States)

    Leah S. Bauer

    1995-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2015-01-01

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

  6. 40 CFR 180.1011 - Viable spores of the microorganism Bacillus thuringiensis Berliner; exemption from the...

    Science.gov (United States)

    2010-07-01

    ... EXEMPTIONS FOR PESTICIDE CHEMICAL RESIDUES IN FOOD Exemptions From Tolerances § 180.1011 Viable spores of the... characteristics of the parent strain or contamination by other microorganisms. (3) Each lot of spore preparation... production is a Bacillus thuringiensis strain which does not produce β-exotoxin under standard manufacturing...

  7. [Susceptibility of Aedes aegypti (L.) strains from Havana to a Bacillus thuringiensis var. israelensis].

    Science.gov (United States)

    Menéndez Díaz, Zulema; Rodríguez Rodríguez, Jinnay; Gato Armas, René; Companioni Ibañez, Ariamys; Díaz Pérez, Manuel; Bruzón Aguila, Rosa Yirian

    2012-01-01

    the integration of chemical and biological methods is one of the strategies for the vector control, due to the existing environmental problems and the concerns of the community as a result of the synthetic organic insecticide actions. The bacterium called Bacillus thuringiensis var. israelensis in liquid formulation has been widely used in the vector control programs in several countries and has shown high efficacy at lab in Cuba. to determine the susceptibility of Aedes aegypti collected in the municipalities of La Habana province to Bacillus thuringiensis var. israelensis. fifteen Aedes aegypti strains, one from each municipality, were used including larvae and pupas collected in 2010 and one reference strain known as Rockefeller. The aqueous formulation of Bacillus thuringiensis var. israelensis (Bactivec, Labiofam, Cuba) was used. The bioassays complied with the World Health Organization guidelines for use of bacterial larvicides in the public health sector. The larval mortality was read after 24 hours and the results were processed by the statistical system SPSS (11.0) through Probit analysis. the evaluated mosquito strains showed high susceptibility to biolarvicide, there were no significant differences in LC50 values of Ae. aegypti strains, neither in the comparison of these values with those of the reference strain. the presented results indicate that the use of Bacillus thuringiensis var. israelensis continues to be a choice for the control of Aedes aegypti larval populations in La Habana province.

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

    Directory of Open Access Journals (Sweden)

    Elham Moazamian

    2013-03-01

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

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

    DEFF Research Database (Denmark)

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

    2006-01-01

    The capability of two bioinsecticide strains of Bacillus thuringiensis (ssp. israelensis and ssp. kurstaki) to germinate and persist in vivo in the gastrointestinal tract of human-flora-associated rats was studied. Rats were dosed either with vegetative cells or spores of the bacteria for 4 conse...

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

    Czech Academy of Sciences Publication Activity Database

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

    2006-01-01

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

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

    Science.gov (United States)

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2013-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2014-01-01

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

  14. Activity of wild-type and hybrid Bacillus thuringiensis delta-endotoxins against Agrotis ipsilon

    NARCIS (Netherlands)

    Maagd, de R.A.; Weemen-Hendriks, M.; Molthoff, J.W.; Naimov, S.

    2003-01-01

    Twelve Cry1 and two Cry9 ?-endotoxins fromBacillus thuringiensis were tested for their activity against black cutworm (Agrotis ipsilon).A. ipsilon was not susceptible to many toxins, but three toxins had significant activity. Cry9Ca was the most toxic, followed by Cry1Aa and Cry1Fb. Hybrids between

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

    African Journals Online (AJOL)

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

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

    African Journals Online (AJOL)

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2014-01-01

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

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

    Science.gov (United States)

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

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

    Science.gov (United States)

    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...

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

    Science.gov (United States)

    Leah S. Bauer

    1990-01-01

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

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

    NARCIS (Netherlands)

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

    1995-01-01

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

  2. Ecology and diversity of Bacillus thuringiensis in soil environment

    African Journals Online (AJOL)

    SERVER

    2007-07-04

    Jul 4, 2007 ... B. thuringiensis isolates were circular, white, flat and undulate or entire. ... India exclusively by the multinational organizations (Sax- ena, 2000). However ... tive, reproductive and crystal structure determination. Physiological.

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

    Directory of Open Access Journals (Sweden)

    Broderick Nichole A

    2010-04-01

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

  4. An anionic defensin from Plutella xylostella with potential activity against Bacillus thuringiensis.

    Science.gov (United States)

    Xu, X-X; Zhang, Y-Q; Freed, S; Yu, J; Gao, Y-F; Wang, S; Ouyang, L-N; Ju, W-Y; Jin, F-L

    2016-12-01

    Insect defensins, are cationic peptides that play an important role in immunity against microbial infection. In the present study, an anionic defensin from Plutella xylostella, (designated as PxDef) was first cloned and characterized. Amino acid sequence analysis showed that the mature peptide owned characteristic six-cysteine motifs with predicted isoelectric point of 5.57, indicating an anionic defensin. Quantitative real-time polymerase chain reaction analysis showed that PxDef was significantly induced in epidermis, fat body, midgut and hemocytes after injection of heat-inactivated Bacillus thuringiensis, while such an induction was delayed by the injection of live B. thuringiensis in the 4th instar larvae of P. xylostella. Knocking down the expression of nuclear transcription factor Dorsal in P. xylostella by RNA interference significantly decreased the mRNA level of PxDef, and increased the sensitivity of P. xylostella larvae to the infection by live B. thuringiensis. The purified recombinant mature peptide (PxDef) showed higher activity against Gram-positive bacteria, with the minimum inhibition concentrations of 1.6 and 2.6 µM against B. thuringiensis and Bacillus subtilis, respectively. To our knowledge, this is the first report about an anionic PxDef, which may play an important role in the immune system of P. xylostella against B. thuringiensis.

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

    Czech Academy of Sciences Publication Activity Database

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

    2015-01-01

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

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

    NARCIS (Netherlands)

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

    2001-01-01

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

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

    Science.gov (United States)

    Daniel J. Krofcheck; Algimantas P. Valaitis

    2010-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Lílian Botelho Praça

    2004-01-01

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

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

    African Journals Online (AJOL)

    121, were effective in the reduction of R. solani infection. In addition, GM-23 increased the length of pepper seedlings. These results suggest that the B. thuringiensis strains studied have an excellent potential to be used as bio-control agents of R.

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

    DEFF Research Database (Denmark)

    Frederiksen, Kristine; Rosenquist, Hanne; Jørgensen, Kirsten

    2006-01-01

    . kurstaki HD1 present in the products Dipel, Biobit, and Foray, and nine isolates grouped with B. thuringiensis subsp. aizawai present in Turex. The commercial strains were primarily isolated from samples of tomatoes, cucumbers, and peppers. A multiplex PCR method was developed to simultaneously detect all...

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

    International Nuclear Information System (INIS)

    Khan, K. A.

    2006-01-01

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

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

    NARCIS (Netherlands)

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

    2006-01-01

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

  13. On-line Biomass Estimation in a Batch Biotechnological Process: Bacillus thuringiensis δ - endotoxins production.

    OpenAIRE

    Amicarelli, Adriana

    2010-01-01

    In this Chapter it has been addressed the problem of the biomass estimation in a batch biotechnological process: the Bacillus thuringiensis (Bt) δ-endotoxins production process. Different alternatives that can be successfully used in this sense were presented. It has been exposed the design of various biomass estimators, namely: a phenomenological biomass estimator, a standard EKF biomass estimator, a biomass estimator based on ANN, a decentralized Kalman Filter, and a biomass concentration ...

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

    Science.gov (United States)

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

    2015-12-01

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

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

    Directory of Open Access Journals (Sweden)

    C.P. Narkhede

    2017-04-01

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

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

    Directory of Open Access Journals (Sweden)

    Marzieh Soleimany

    2018-03-01

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

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

    International Nuclear Information System (INIS)

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

    2008-01-01

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

  18. Exploratory analysis for the optimization of culture media for Bacillus thuringiensis fermentation

    Directory of Open Access Journals (Sweden)

    Jenny M. Escobar

    2004-07-01

    Full Text Available Agrichemical involve health risks for producers and consumers; they can also affect beneficial insects, birds, fish and mammals. Bacillus thuringiensis-based biopesticides specific for insect plagues are available, which, represent an alternative to traditional chemically-based products. Culture media design is an impor-tant factor in the production of biopesticides based on Bacillus thuringiensis. The media have to be economi-cal and they must contain all the nutrition factors required by the bacteria. The yield of active ingredient, comprising the toxin-containing crystal protein and bacterial spores, must be high and with adequate toxi-city to formulate the biopesticide. In this work we studied different culture media and optimised a promi-sing culture medium for B. thuringiensis fermentation to obtain an active ingredient for use as raw material in manufacturing a biopesticide. The final concentration of active ingredient was 15 to 16 g/L and the raw material cost around US $0,30/kg product. Calculations for the experimental phase were done using the response surface technique in a central composite design (CCD, the optimisation phase used Derringer and Suich (1980 method for dual response. Key words: Biopesticides, culture media, surface response, dual response, contour plots, central composite design.

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

    Directory of Open Access Journals (Sweden)

    Óscar Enrique Guevara

    2004-01-01

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

  20. U.v.-induced and N-methyl-N'-nitrosoguanidine-induced mutagenesis in Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Auffray, Y.; Boutibonnes, P.

    1987-01-01

    The lethal and mutagenic effects of u.v. light and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) on Bacillus thuringiensis were investigated. Lethality studies demonstrated that B. thuringiensis was relatively sensitive to these agents. This bacterium was mutated at the rifampicin resistance marker by u.v. light and to a lesser extent by the direct acting alkylating agent MNNG. One mutant selected for its greater sensitivity to u.v. light expressed a higher frequency of mutagenesis after u.v. light treatment and appeared to be defective in an excision repair pathway. However, this mutant was only slightly mutable by MNNG in comparison with the wild-type strain. This unusual phenotype does not yet have a parallel among the radiation sensitive mutants described in other bacterial species. (author)

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

    Science.gov (United States)

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

    2016-10-01

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

  2. Characterisation of Bacillus thuringiensis kurstaki strains by toxicity ...

    African Journals Online (AJOL)

    SERVER

    2007-08-06

    Aug 6, 2007 ... Southern analysis of restricted plasmid combined with cryIA profile can provide an ... Abbreviations: ICP, Insecticidal Crystal Protein; H.a., .... 70 kDa ICP component were purified as per protocol of Harlow and .... They are in principal .... on the interaction of Bacillus thuriengensis subsp. kurstaki with the.

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

    DEFF Research Database (Denmark)

    Hendriksen, Niels Bohse; Hansen, Bjarne Munk

    2011-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Yuan Yihui

    2012-12-01

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

  5. A Novel Tenebrio molitor Cadherin Is a Functional Receptor for Bacillus thuringiensis Cry3Aa Toxin*

    OpenAIRE

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

    2009-01-01

    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 lepid...

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

    Science.gov (United States)

    Schesser, J H; Bulla, L A

    1979-05-01

    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.

  7. Commercial formulations of Bacillus thuringiensis for control of Indian meal moth.

    Science.gov (United States)

    Schesser, J H

    1976-10-01

    Doses of four commercial formulations and one experimental formulation of Bacillus thuringiensis Berliner were mixed with the diet used to rear colonies of the Indian meal moth Plodia interpunctella (Hübner). Indian meal moth eggs were introduced to the treated diet, and the resultant adult emergence was tabulated. The experimental formulations ranked as follows in efficacy in controlling the Indian meal moth: Dipel (50% lethal concentration [LC50], 25 mg/kg) greater than Bactospeine WP (LC50, 100 mg/kg) greater than Thuricide (LC50, 150 mg/kg) greater than IMC 90007 (LC30, 180 mg/kg) greater than Bactospeine Flowable (LC50, 440 mg/kg).

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

    Czech Academy of Sciences Publication Activity Database

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

    2011-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2010-01-01

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

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

    International Nuclear Information System (INIS)

    Hmaied, Ezzedine; Ben Mbarek, Wael

    2010-01-01

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

  11. Activity of a Brazilian strain of Bacillus thuringiensis israelensis against the cotton Boll Weevil Anthonomus grandis Boheman (Coleoptera: Tenebrionidae).

    Science.gov (United States)

    Monnerat, R; Martins, E; Praça, L; Dumas, V; Berry, C

    2012-02-01

    A Brazilian Bacillus thuringiensis subspecies israelensis, toxic to Diptera, including mosquitoes, was found also to show toxicity to the coleopteran boll weevil Anthonomus grandis Boheman at an equivalent level to that of the standard coleopteran-active B. thuringiensis subspecies tenebrionis T08017. Recombinant B. thuringiensis strains expressing the individual Cyt1Aa, Cry4Aa, Cry4Ba and Cry11Aa toxins from this strain were assessed to evaluate their potential contribution to the activity against A. grandis, either alone or in combination. Whilst individual toxins produced mortality, none was sufficiently potent to allow calculation of LC50 values. Combinations of toxins were unable to attain the same potency as the parental B. thuringiensis subsp. israelensis, suggesting a major role for other factors produced by this strain.

  12. Diversity of Bacillus thuringiensis strains in the maize and bean phylloplane and their respective soils in Colombia.

    Science.gov (United States)

    Jara, S; Maduell, P; Orduz, S

    2006-07-01

    To evaluate the distribution of Bacillus thuringiensis strains from maize and bean phylloplane and their respective soils. B. thuringiensis was isolated from the phylloplane and soil of maize and bean from three municipalities in Antioquia, Colombia. Ninety six samples of phylloplane and 24 of soil were analyzed. A total of 214 isolates were obtained from 96 phylloplane samples while 59 isolates were recovered from 24 soil samples. Sixty five per cent and 12% of the phylloplane and soil isolates, respectively, showed activity against Spodoptera frugiperda. These isolates contained delta-endotoxin proteins of 57 and 130 kDa. The most toxic isolates against S. frugiperda had the genotype cry1Aa, cry1Ac, cry1B, and cry1D. In contrast, 27% of the phylloplane isolates and 88% of the soil isolates were active against Culex quinquefasciatus and had protein profiles similar to B. thuringiensis serovar. medellin and B. thuringiensis serovar. israelensis. The most active isolates contain cry4 and cry11 genes. The predominant population of B. thuringiensis on the phylloplane harbored the cry1 gene and was active against S. frugiperda, whereas in soil, isolates harboring cry11 gene and active against C. quinquefasciatus were the majority. The predominance of specific B. thuringiensis populations, both on the leaves and in the soil, suggests the presence of selection in B. thuringiensis populations on the studied environment.

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

    Directory of Open Access Journals (Sweden)

    Naresh Arora

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

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

    Directory of Open Access Journals (Sweden)

    Wan-Fang Zhong

    2005-12-01

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

  15. Mob/oriT, a mobilizable site-specific recombination system for unmarked genetic manipulation in Bacillus thuringiensis and Bacillus cereus.

    Science.gov (United States)

    Wang, Pengxia; Zhu, Yiguang; Zhang, Yuyang; Zhang, Chunyi; Xu, Jianyi; Deng, Yun; Peng, Donghai; Ruan, Lifang; Sun, Ming

    2016-06-10

    Bacillus thuringiensis and Bacillus cereus are two important species in B. cereus group. The intensive study of these strains at the molecular level and construction of genetically modified bacteria requires the development of efficient genetic tools. To insert genes into or delete genes from bacterial chromosomes, marker-less manipulation methods were employed. We present a novel genetic manipulation method for B. thuringiensis and B. cereus strains that does not leave selection markers. Our approach takes advantage of the relaxase Mob02281 encoded by plasmid pBMB0228 from Bacillus thuringiensis. In addition to its mobilization function, this Mob protein can mediate recombination between oriT sites. The Mob02281 mobilization module was associated with a spectinomycin-resistance gene to form a Mob-Spc cassette, which was flanked by the core 24-bp oriT sequences from pBMB0228. A strain in which the wild-type chromosome was replaced with the modified copy containing the Mob-Spc cassette at the target locus was obtained via homologous recombination. Thus, the spectinomycin-resistance gene can be used to screen for Mob-Spc cassette integration mutants. Recombination between the two oriT sequences mediated by Mob02281, encoded by the Mob-Spc cassette, resulted in the excision of the Mob-Spc cassette, producing the desired chromosomal alteration without introducing unwanted selection markers. We used this system to generate an in-frame deletion of a target gene in B. thuringiensis as well as a gene located in an operon of B. cereus. Moreover, we demonstrated that this system can be used to introduce a single gene or an expression cassette of interest in B. thuringiensis. The Mob/oriT recombination system provides an efficient method for unmarked genetic manipulation and for constructing genetically modified bacteria of B. thuringiensis and B. cereus. Our method extends the available genetic tools for B. thuringiensis and B. cereus strains.

  16. Localization of Bacillus thuringiensis Cry1A toxin-binding molecules in gypsy moth larval gut sections using fluorescence microscopy

    Science.gov (United States)

    Algimantas P. Valaitis

    2011-01-01

    The microbial insecticide Bacillus thuringiensis (Bt) produces Cry toxins, proteins that bind to the brush border membranes of gut epithelial cells of insects that ingest it, disrupting the integrity of the membranes, and leading to cell lysis and insect death. In gypsy moth, Lymantria dispar, two toxin-binding molecules for the...

  17. Isolation and characterization of a novel native Bacillus thuringiensis strain BRC-HZM2 capable of degrading chlorpyrifos

    Czech Academy of Sciences Publication Activity Database

    Wu, S.; Peng, Y.; Huang, Z.; Huang, Z.; Xu, L.; Gelbič, Ivan; Guan, X.; Zhang, L.; Zou, S.

    2015-01-01

    Roč. 55, č. 3 (2015), s. 389-397 ISSN 0233-111X Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * biopesticide * biodegradation Subject RIV: EE - Microbiology, Virology Impact factor: 1.585, year: 2015 http://onlinelibrary.wiley.com/doi/10.1002/jobm.201300501/epdf

  18. Recovery of Bacillus thuringiensis and related spore-forming bacteria from soil after application for gypsy moth control

    Science.gov (United States)

    Phyllis A.W. Martin; Elizabeth A. Mongeon; Michael B. Blackburn; Dawn E. Gundersen-Rindal

    2011-01-01

    Bacillus thuringiensis Berliner (Bt) has been applied for gypsy moth (Lymantria dispar L.) control in forests in the northeastern U.S. for many years. The subspecies of Bt that is used (urstaki) is not common in U.S. soil. We attempted to recover Bt from...

  19. Survival of Bacillus thuringiensis strains in gypsy moth (Lymantria dispar) larvae is correlated with production of urease

    Science.gov (United States)

    Phyllis A.W. Martin; Robert R. Jr. Farrar; Michael B. Blackburn

    2011-01-01

    We tested 50 lepidopteran-toxic Bacillus thuringiensis Berliner (Bt) strains with diverse phenotypes for the ability to survive repeated passages through larvae of the gypsy moth, Lymantria dispar (L.), without intervening growth on artificial media. These experiments have revealed a remarkable correlation...

  20. Effect of proteolytic and detoxification enzyme inhibitors on Bacillus thuringiensis var. israelensis tolerance in the mosquito Aedes aegypti

    Czech Academy of Sciences Publication Activity Database

    Hu, X.; Guo, Y.; Wu, S.; Liu, Z.; Fu, T.; Shao, E.; Carballar-Lejarazú, R.; Zhao, G.; Huang, Z.; Gelbič, Ivan; Guan, X.; Zou, S.; Xu, L.; Zhang, L.

    2017-01-01

    Roč. 27, č. 2 (2017), s. 169-179 ISSN 0958-3157 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * Bti * Aedes aegypti Subject RIV: ED - Physiology OBOR OECD: Biology (theoretical, mathematical, thermal, cryobiology, biological rhythm), Evolutionary biology Impact factor: 0.919, year: 2016

  1. Characterization of eight Bacillus thuringiensis isolates originated from fecal samples of Fuzhou Zoo and Fuzhou Panda Center

    Czech Academy of Sciences Publication Activity Database

    Wu, Ch.; Wu, L.; Zhang, L.; Gelbič, Ivan; Xu, L.; Guan, X.

    2014-01-01

    Roč. 17, č. 3 (2014), s. 395-397 ISSN 1226-8615 Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis * faeces * microscophy Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection Impact factor: 0.946, year: 2014 http://www.sciencedirect.com/science/article/pii/S122686151400034X

  2. Bacillus thuringiensis Cry1Ca-resistant Spodoptera exigua lacks expression of one of four Aminopeptidase N genes

    NARCIS (Netherlands)

    Herrero, S.; Gechev, T.; Bakker, P.L.; Moar, W.; Maagd, de R.A.

    2005-01-01

    BACKGROUND: Insecticidal toxins from Bacillus thuringiensis bind to receptors on midgut epithelial cells of susceptible insect larvae. Aminopeptidases N (APNs) from several insect species have been shown to be putative receptors for these toxins. Here we report the cloning and expression analysis of

  3. Beetle-specific Bacillus thuringiensis Cry3Aa toxin reduces larval growth and curbs reproduction in Spodoptera littoralis (Boisd.)

    Czech Academy of Sciences Publication Activity Database

    Hussein, Hany; Habuštová, Oxana; Sehnal, František

    2005-01-01

    Roč. 61, - (2005), s. 1186-1192 ISSN 1526-498X R&D Projects: GA ČR(CZ) GA522/02/1507 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis * Spodoptera littoralis * Bt applications Subject RIV: ED - Physiology Impact factor: 1.175, year: 2005

  4. The introduction of integrated pest management in the Ethiopian horticultural sector : Bacillus thuringiensis strains and its toxicity

    NARCIS (Netherlands)

    Belder, den E.; Elderson, J.

    2012-01-01

    1 Introduction As hazards of conventional broad acting pesticides are documented, researchers, poli cymakers and growers look for pesticides that are toxic only to the target pest, have no impact on other such as beneficial species, and have fewer environmental effects. Bacillus thuringiensis (Bt)

  5. Bacillus thuringiensis delta-endotoxin Cry1 hybrid proteins with increased activity against the Colorado potato beetle

    NARCIS (Netherlands)

    Naimov, S.; Weemen-Hendriks, M.; Dukiandjiev, S.; Maagd, de R.A.

    2001-01-01

    Cry1 delta-endotoxins of Bacillus thuringiensis are generally active against lepidopteran insects, but Cry1Ba and Cry1Ia have additional, though low, levels of activity against coleopterans such as the Colorado potato beetle. Here we report the construction of Cry1Ba/Cry1Ia hybrid toxins which have

  6. A hybrid Bacillus thuringiensis delta-endotoxin gene gives resistance against a coleopteran and a lepidopteran pest in transgenic potato

    NARCIS (Netherlands)

    Naimov, S.; Dukiandjiev, S.; Maagd, de R.A.

    2003-01-01

    Expression of Bacillus thuringiensis delta-endotoxins has proven to be a successful strategy for obtaining insect resistance in transgenic plants. Drawbacks of expression of a single resistance gene are the limited target spectrum and the potential for rapid adaptation of the pest. Hybrid toxins

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

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry2Ae protein 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 PROTECTANTS...

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

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1A.105 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 and...

  9. 40 CFR 174.504 - Bacillus thuringiensis Cry1F protein in cotton; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1F protein 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 and...

  10. 40 CFR 174.509 - Bacillus thuringiensis Cry3A protein; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry3A protein; 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 Tolerance...

  11. 40 CFR 174.517 - Bacillus thuringiensis Cry9C protein in corn; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry9C protein in corn; exemption from the requirement of a tolerance. 174.517 Section 174.517 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS PROCEDURES AND REQUIREMENTS FOR PLANT-INCORPORATED PROTECTANTS Tolerances and...

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

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1F protein 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 and...

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

    Science.gov (United States)

    Hariram, Upasana; Labbé, Ronald

    2015-03-01

    Recent incidents of foodborne illness associated with spices as the vehicle of transmission prompted this examination of U.S. retail spices with regard to Bacillus cereus. This study focused on the levels of aerobic-mesophilic spore-forming bacteria and B cereus spores associated with 247 retail spices purchased from five states in the United States. Samples contained a wide range of aerobic-mesophilic bacterial spore counts (spices had high levels of aerobic spores (> 10(7) CFU/g). Using a novel chromogenic agar, B. cereus and B. thuringiensis spores were isolated from 77 (31%) and 11 (4%) samples, respectively. Levels of B. cereus were spice isolates to form spores, produce diarrheal toxins, and grow at moderately abusive temperatures makes retail spices an important potential vehicle for foodborne illness caused by B. cereus strains, in particular those that produce diarrheal toxins.

  14. New insights into the binding and catalytic mechanisms of Bacillus thuringiensis lactonase: insights into B. thuringiensis AiiA mechanism.

    Directory of Open Access Journals (Sweden)

    Marc N Charendoff

    Full Text Available The lactonase enzyme (AiiA produced by Bacillus thuringiensis serves to degrade autoinducer-1 (AI-1 signaling molecules in what is an evolved mechanism by which to compete with other bacteria. Bioassays have been previously performed to determine whether the AI-1 aliphatic tail lengths have any effect on AiiA's bioactivity, however, data to date are conflicting. Additionally, specific residue contributions to the catalytic activity of AiiA provide for some interesting questions. For example, it has been proposed that Y194 serves to provide an oxyanion hole to AI-1 which is curious given the fact the substrate spans two Zn(2+ ions. These ions might conceivably provide enough charge to promote both ligand stability and the carbonyl activation necessary to drive a nucleophilic attack. To investigate these questions, multiple molecular dynamics simulations were performed across a family of seven acylated homoserine lactones (AHL along with their associated intermediate and product states. Distance analyses and interaction energy analyses were performed to investigate current bioassay data. Our simulations are consistent with experimental studies showing that AiiA degrades AHLs in a tail length independent manner. However, the presence of the tail is required for activity. Also, the putative oxyanion hole function of Y194 toward the substrate is not observed in any of the reactant or product state simulation trajectories, but does seem to show efficacy in stabilizing the intermediate state. Last, we argue through ionization state analyses, that the proton shuttling necessary for catalytic activity might be mediated by both water and substrate-based intra-molecular proton transfer. Based on this argument, an alternate catalytic mechanism is proposed.

  15. Produccion por tecnologia de fermentacion de bacillus thuringiensis utilizando medios alternativos

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    Yaneth Amparo Muñoz-Peñalosa

    2000-01-01

    Full Text Available In the production by fermentation technology Bacillus thuringiensis of five alternative methods they were studied. The results of cell growth, working-level 100ml in static culture and temperature of 28 ° C, mostraronque the optimal substrate corresponded to the environment in which molasses and rice powder was added (alternative Medium No. 1] The cell development using 100 ml of substrate was studied with reciprocating shaking 110 rpm. in this test was determined filter curve creciemiento medio.The inoculum, allowed tiempode set the process in 6 hours. For the development of fermentations, are counted with the experimental equipment, glass bioreactor in two liters of capacity and aeration devices, mechanical stirring, sampling and output gases.The fermentation in the production of Bacillus thuringiensis is the type discontinuous submerged aerobic process and growth into account .Teniendo bibliographic information and preliminary results of the study, fixed working parameters were determined for production by fermentation of Bacillus thuringiensis, being alternative means No. 1, volume 1 liter temperature 28 ° C and cell concentration of the inoculum. To determine the optimum parameters of fermantacion was used a factorial design of experiments of the type 22, (two variables at two levels, with aeration (3.2-0.5 VVM and agitation (110-210 rpm .The fermentations performed 7, 4 design and 3 the average level of the variables. For monitoring fermentation sample was taken every 12 hours and cell concentration (Chamber of Neuvauer and pH was analyzed. The results of cell concentration measurement for fermentations at 60 hours shows that optimum working conditions and limitations correspond to the values ​​of the variable, 3.2 VVM aeration and agitation 210 Variable rpm.Significant was the aeration of pH in the fermentation media change neutral to acid and ended as a staple. A fermentations I were efectuo controlde microbiological quality, Gram

  16. Heat and desiccation are the predominant factors affecting inactivation of Bacillus licheniformis and Bacillus thuringiensis spores during simulated composting.

    Science.gov (United States)

    Stanford, K; Harvey, A; Barbieri, R; Xu, S; Reuter, T; Amoako, K K; Selinger, L B; McAllister, T A

    2016-01-01

    The suitability of composting for disposal of livestock mortalities due to Bacillus anthracis was assessed by measuring viability of surrogate spores from two strains each of Bacillus licheniformis and Bacillus thuringiensis after a heating cycle modelled on a cattle composting study. Sporulation was attempted from 10 to 37°C, but poor yields at lower temperatures resulted in 25, 30 and 37°C being selected to generate sufficient spores (8 log10  CFU ml(-1) ) for experiments. Spores were inoculated into 3 g autoclaved dried-ground compost rehydrated with 6 ml water or silica beads in a factorial design for each strain, sporulation temperature, matrix and sampling day (0, 25, 50, 100, 150). Maximum incubation temperature was 62°C, but spores were maintained at ≥55°C for 78 of 150 days. Although significant differences existed among Bacillus strains and sporulation temperatures, numbers of viable spores after 150 days averaged 1·3 log10  CFU g(-1) , a 5·2 log10 reduction from day 0. Spore inactivation was likely due to heat and desiccation as matrices were autoclaved prior to incubation, negating impacts of microflora. Results support composting for disposal of anthrax mortalities, provided long-term thermophillic heating is achieved. Due to limited sporulation at 10°C, livestock mortalities from anthrax at this or lower ambient temperatures would likely be of lower risk for disease transmission. © 2015 The Society for Applied Microbiology.

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

    Science.gov (United States)

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

    2012-01-01

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

  18. c-di-GMP Regulates Various Phenotypes and Insecticidal Activity of Gram-Positive Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Yang Fu

    2018-02-01

    Full Text Available C-di-GMP has been well investigated to play significant roles in the physiology of many Gram-negative bacteria. However, its effect on Gram-positive bacteria is less known. In order to more understand the c-di-GMP functions in Gram-positive bacteria, we have carried out a detailed study on the c-di-GMP-metabolizing enzymes and their physiological functions in Bacillus thuringiensis, a Gram-positive entomopathogenic bacterium that has been applied as an insecticide successfully. We performed a systematic study on the ten putative c-di-GMP-synthesizing enzyme diguanylate cyclases (DGCs and c-di-GMP-degrading enzyme phosphodiesterases (PDEs in B. thuringiensis BMB171, and artificially elevated the intracellular c-di-GMP level in BMB171 by deleting one or more pde genes. We found increasing level of intracellular c-di-GMP exhibits similar activities as those in Gram-negative bacteria, including altered activities in cell motility, biofilm formation, and cell-cell aggregation. Unexpectedly, we additionally found a novel function exhibited by the increasing level of c-di-GMP to promote the insecticidal activity of this bacterium against Helicoverpa armigera. Through whole-genome transcriptome profile analyses, we found that 4.3% of the B. thuringiensis genes were differentially transcribed when c-di-GMP level was increased, and 77.3% of such gene products are involved in some regulatory pathways not reported in other bacteria to date. In summary, our study represents the first comprehensive report on the c-di-GMP-metabolizing enzymes, their effects on phenotypes, and the transcriptome mediated by c-di-GMP in an important Gram-positive bacterium.

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

    Science.gov (United States)

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

    2009-01-01

    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

  20. Effects of Bacillus thuringiensis strains virulent to Varroa destructor on larvae and adults of Apis mellifera.

    Science.gov (United States)

    Alquisira-Ramírez, Eva Vianey; Peña-Chora, Guadalupe; Hernández-Velázquez, Víctor Manuel; Alvear-García, Andrés; Arenas-Sosa, Iván; Suarez-Rodríguez, Ramón

    2017-08-01

    The sublethal effects of two strains of Bacillus thuringiensis, which were virulent in vitro to Varroa destructor, were measured on Apis mellifera. The effects of five concentrations of total protein (1, 5, 25, 50 and 100μg/mL) from the EA3 and EA26.1 strains on larval and adult honey bees were evaluated for two and seven days under laboratory conditions. Based on the concentrations evaluated, total protein from the two strains did not affect the development of larvae, the syrup consumption, locomotor activity or proboscis extension response of adults. These same parameters were also tested for the effects of three concentrations (1, 10 and 15μg/kg) of cypermethrin as a positive control. Although no significant differences were observed after two days of treatment with cypermethrin, a dose-response relationship in syrup consumption and locomotor activity was observed. A significant reduction in the proboscis extension response of the bees treated with cypermethrin was also observed. Therefore, in contrast to cypermethrin, our results indicate that the EA3 and EA26.1 strains of B. thuringiensis can be used in beehives to control V. destructor and reduce the negative effects of this mite on colonies without adverse effects on the larvae and adults of A. mellifera. Additionally, the overuse of synthetic miticides, which produce both lethal and sublethal effects on bees, can be reduced. Copyright © 2017 Elsevier Inc. All rights reserved.

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

    Science.gov (United States)

    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.

  2. Immobilization of alginate-encapsulated Bacillus thuringiensis var. israelensis containing different multivalent counterions for mosquito control.

    Science.gov (United States)

    Prabakaran, G; Hoti, S L

    2008-08-01

    Immobilized techniques have been used widely for the controlled release formulation of mosquitoes. Among the microbial formulations, polymeric matrices play an important role in the controlled release of microbial pesticide at rates sufficiently effective to kill mosquitoes in the field. The advantage of these matrices is that they enhance the stability of both spores and toxin against pH, temperature variations, and UV irradiation. The disadvantage of using calcium alginate beads is that they are unstable upon contact with phosphate of potassium or sodium ions rich in the mosquito habitats. To overcome these problems, attempts were made to encapsulate Bacillus thuringiensis var. israelensis within alginate by using different multivalent counterions, namely, calcium chloride, zinc sulfate, copper sulfate, cobalt chloride, and ferric chloride, and the beads formed were tested for its mosquito larvicidal activity. Among all the beads tested, zinc alginate beads resulted in maximum larvicidal activity of 98% (+/-1.40 SE) against Culex quinquefasciatus IIIrd instar larvae and maximum spore count of 3.36 x 10(5) (+/-5291.50 SE) CFU/ml. Zinc alginate beads maintained their structure for up to 48 h when shaken vigorously on a rotary shaker at 180 rpm in the presence of 10 mM potassium phosphate buffer (pH 6.8 +/- 0.1). In conclusion, our results suggest that the use of zinc sulfate as counterions to encapsulate B. thuringiensis var. israelensis within alginate may be a potent mosquito control program in the habitats where more phosphate ions are present.

  3. Cytotoxicity analysis of three Bacillus thuringiensis subsp. israelensis δ-endotoxins towards insect and mammalian cells.

    Directory of Open Access Journals (Sweden)

    Roberto Franco Teixeira Corrêa

    Full Text Available Three members of the δ-endotoxin group of toxins expressed by Bacillus thuringiensis subsp. israelensis, Cyt2Ba, Cry4Aa and Cry11A, were individually expressed in recombinant acrystalliferous B. thuringiensis strains for in vitro evaluation of their toxic activities against insect and mammalian cell lines. Both Cry4Aa and Cry11A toxins, activated with either trypsin or Spodoptera frugiperda gastric juice (GJ, resulted in different cleavage patterns for the activated toxins as seen by SDS-PAGE. The GJ-processed proteins were not cytotoxic to insect cell cultures. On the other hand, the combination of the trypsin-activated Cry4Aa and Cry11A toxins yielded the highest levels of cytotoxicity to all insect cells tested. The combination of activated Cyt2Ba and Cry11A also showed higher toxic activity than that of toxins activated individually. When activated Cry4Aa, Cry11A and Cyt2Ba were used simultaneously in the same assay a decrease in toxic activity was observed in all insect cells tested. No toxic effect was observed for the trypsin-activated Cry toxins in mammalian cells, but activated Cyt2Ba was toxic to human breast cancer cells (MCF-7 when tested at 20 µg/mL.

  4. Isolation, characterization and toxicity of native Bacillus thuringiensis isolates from different hosts and habitats in Iran

    Directory of Open Access Journals (Sweden)

    Ghassemi-Kahrizeh Akbar

    2017-09-01

    Full Text Available Bacillus thuringiensis is a Gram-positive, aerobic, facultative anaerobic and endospore-forming bacterium. Different strains of this species have the ability to produce parasporal crystalline inclusions which are toxic to larvae of different insect orders and other invertebrates and cause rapid death of the host. To determine the importance of this species in microbial control, we collected native strains and studied their virulence on the diamondback moth, Plutella xylostella. More than 148 samples were collected from Alborz, Guilan and Mazandaran Provinces. Experimental samples, including soil samples from forests, fruit gardens, agricultural fields, diseased and dead larvae, were transferred to a laboratory in sterile plastic containers. For evaluating B. thuringiensis isolates virulence, a cabbage leaf dip method with 106 cell · ml−1 concentration of various Bt isolates was applied to diamondback moths. Larval mortality was recorded 72 h after treatment. Based on bioassay results, all isolates were classified into three high, medium and low virulence groups. Protein level characterization based on the SDS-PAGE gel analysis showed that two isolates from a high virulence group have proteins of high molecular masses of 121 and 109 kDa. Results revealed that there is a positive correlation between protein masses and virulence of isolates. In addition, this research introduced nine strains that are highly toxic to P. xylostella and would be valuable as insecticidal agents for controlling lepidopteran pests.

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

    Science.gov (United States)

    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.

  6. Bacillus thuringiensis in caterpillars and associated materials collected from protected tropical forests in northwestern Costa Rica.

    Science.gov (United States)

    Rodríguez-Sánchez, César; Sittenfeld, Ana; Janzen, Daniel H; Espinoza, Ana M

    2006-06-01

    Bacillus thuringiensis (Bt) synthesizes crystalline inclusions that are toxic to caterpillars (Lepidoptera) and other orders of invertebrates. Materials associated with 37 caterpillars from 16 species, collected while feeding on 15 different species of host plants in dry, cloud and rain forests located in the Area de Conservación Guanacaste in northwestern Costa Rica, were examined for the presence of Bt. From a total of 101 derived samples, 25 Bt isolates were cultured: 56% from host plant leaves, 8% from caterpillar guts and 36% from caterpillar fecal pellets. Bt was isolated from at least one sample in 38% of the systems constituted by the food plant, gut and fecal pellets corresponding to a single caterpillar. Four different morphologies of crystalline inclusions were observed, with bipyramidal and irregular crystal morphologies being the most prevalent.

  7. Microbiological ecology and association of Bacillus thuringiensis in chicken feces originating from feed

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Peng, Y.; Wu, S.; Sun, L.; Huang, E.; Huang, T.; Xu, L.; Wu, Ch.; Gelbič, Ivan; Guan, X.

    2012-01-01

    Roč. 65, č. 6 (2012), s. 784-791 ISSN 0343-8651 R&D Projects: GA MŠk 2B08003 Grant - others:National Natural Science Foundation of China(CN) 31071745; Ministry of Education of China(CN) 20093515110010; Ministry of Education of China(CN) 20093515120010; Transformation Fund for Agricultural Science and Technology Achievements(CN) 2010GB2C400212; Education Department Foundation(CN) JA12092 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis Subject RIV: EH - Ecology, Behaviour Impact factor: 1.520, year: 2012 http://link.springer.com/content/pdf/10.1007%2Fs00284-012-0231-3

  8. Structure and distribution of the Bacillus thuringiensis Cry4Ba toxin in lipid membranes

    International Nuclear Information System (INIS)

    Puntheeranurak, Theeraporn; Stroh, Cordula; Zhu Rong; Angsuthanasombat, Chanan; Hinterdorfer, Peter

    2005-01-01

    Bacillus thuringiensis Cry δ-endotoxins cause death of susceptible insect larvae by forming lytic pores in the midgut epithelial cell membranes. The 65 kDa trypsin activated Cry4Ba toxin was previously shown to be capable of permeabilizing liposomes and forming ionic channels in receptor-free planar lipid bilayers. Here, magnetic ACmode (MACmode) atomic force microscopy (AFM) was used to characterize the lateral distribution and the native molecular structure of the Cry4Ba toxin in the membrane. Liposome fusion and the Langmuir-Blodgett technique were employed for supported lipid bilayer preparations. The toxin preferentially inserted in a self-assembled structure, rather than as a single monomeric molecule. In addition, the spontaneous insertion into receptor-free lipid bilayers lead to formation of characteristic pore-like structures with four-fold symmetry, suggesting that tetramers are the preferred oligomerization state of this toxin

  9. Growth of hydroxyapatite on the cellular membrane of the bacterium Bacillus thuringiensis for the preparation of hybrid biomaterials

    Energy Technology Data Exchange (ETDEWEB)

    Cervantes, Eric Reyes, E-mail: onomaeric@hotmail.com [Centro de Investigación en Ciencias Microbiológicas, Benemérita Universidad Autónoma de Puebla, Prolongación de la 24 Sur y Ave San Claudio, Ciudad Universitaria, Col San Manuel, C.P. 72570 Puebla, Pue (Mexico); Torres, Maykel González, E-mail: mikegcu@fata.unam.mx [Centro de Física Aplicada y Tecnología Avanzada, Universidad Nacional Autónoma de México, Campus Juriquilla, Boulevard Juriquilla 3001, Santiago de Querétaro, Querétaro C.P. 76230 (Mexico); Muñoz, Susana Vargas, E-mail: vmsu@unam.mx [Centro de Física Aplicada y Tecnología Avanzada, Universidad Nacional Autónoma de México, Campus Juriquilla, Boulevard Juriquilla 3001, Santiago de Querétaro, Querétaro C.P. 76230 (Mexico); Rosas, Efraín Rubio, E-mail: efrainrubio@yahoo.com [Centro de Investigación en Ciencias Microbiológicas, Benemérita Universidad Autónoma de Puebla, Prolongación de la 24 Sur y Ave San Claudio, Ciudad Universitaria, Col San Manuel, C.P. 72570 Puebla, Pue (Mexico); and others

    2016-01-01

    This study aimed to grow hydroxyapatite (HAp) crystals on the cellular wall of the Gram-positive bacterium Bacillus thuringiensis using a bio-mimetic method. Several strains were phenotypically and genotypically characterized using multilocus sequence typing (MLST) gene markers to differentiate the strains and confirm the identity of the isolated species to guarantee that the selected species was not harmful to human health or the environment. Three of the analyzed strains were selected because they exhibited the best nucleation and growth of HAp on the bacterial surface. This innovative method to grow HAp crystals on a cellular membrane helps to elucidate the mechanisms by which osseous tissue is formed in nature. The optimum concentration for the simulated physiological fluid (SPF) was 1.5 ×. The hybrid materials were characterized by optical microscopy, atomic force microscopy (AFM), scanning electron microscopy (SEM), X-ray powder diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR). - Highlights: • HAp crystals are grown on the cellular wall of a GP bacteria Bacillus thuringiensis. • The growing was carried out by using a bio-mimetic method. • Hybrid materials were characterized with morphological and spectroscopic techniques. • The reported method allows understanding the mechanisms to produce osseous tissue. • The membrane of Bacillus thuringiensis can grow more HAp than Bacillus halodurans.

  10. Selection of optimum conditions of medium acidity and aeration for submerget cultivation of Bacillus thuringiensis and Beauveria bassiana

    Directory of Open Access Journals (Sweden)

    O. A. Dregval

    2010-06-01

    Full Text Available The paper deals with the influence of medium pH and aeration rate on growth and sporulation of Bacillus thuringiensis and Вeauveria bassiana, which are main constituents of the complex microbial insecticide. It was established optimal medium pH for B. thuringiensis – 6.0 and for В. bassiana – 6.0–7.0. The maximum productivity of the studied microorganisms was observed in the same range of aeration – 7– 14 mmol O2/l/h. The selected conditions of cultivation are necessary for the production of complex biological insecticide based on the association of B. thuringiensis and B. bassiana.

  11. Isolation of Bacillus thuringiensis from the state of Amazonas, in Brazil, and screening against Aedes aegypti (Diptera, Culicidae

    Directory of Open Access Journals (Sweden)

    Joelma Soares-da-Silva

    2015-03-01

    Full Text Available We investigated the use of Bacillus thuringiensis isolated in the state of Amazonas, in Brazil, for the biological control of the dengue vector Aedes aegypti. From 25 soil samples collected in nine municipalities, 484 bacterial colonies were obtained, 57 (11.78% of which were identified as B. thuringiensis. Six isolates, IBt-03, IBt-06, IBt-07, IBt-28, IBt-30, and BtAM-27 showed insecticidal activity, and only BtAM-27 presents the five genes investigated cry4Aa, cry4Ba, cry10Aa, cry11Aa, and cry11Ba. The IBt-07 and IBt- 28, with lower LC50 values, showed equal toxicity compared to the standards. The isolates of B. thuringiensis from Amazonas constitute potential new means of biological control for A. aegypti, because of their larvicidal activity and the possibility that they may also contain new combinations of toxins.

  12. Survival and conjugal transfer between Bacillus thuringiensis strains in aquatic environment Sobrevivência e conjugação de Bacillus thuringiensis em ambiente aquático

    Directory of Open Access Journals (Sweden)

    Luciana Furlaneto

    2000-10-01

    Full Text Available Field and laboratory studies were conducted to assess the survival of cells and spores and plasmid transfer between Bacillus thuringienis strains in aquatic environment. Results indicated that cells and spores of B. thuringiensis can survive for 10 days in water, without altering their number. The sporulation process began after 12-15 hours of inoculation of water. B. thuringiensis was able to transfer conjugative plasmids in the aquatic environment.O presente trabalho é um estudo sobre a sobrevivência e a conjugação de linhagens de Bacillus thuringiensis em água. Os experimentos conduzidos no laboratório mostram que as células e os esporos de B. thuringiensis podem persistir pelo menos 10 dias na água. A esporulação inicia-se 12-15 horas após a inoculação. O processo de conjugação foi demonstrado em diferentes ambientes aquáticos, tanto em condições de laboratório quanto no meio ambiente.

  13. Isolation and characterization of radioresistant mutants in Bacillus subtilis and Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Kalinin, V.L.; Petrov, V.N.; Petrova, T.M.

    1981-01-01

    Vegetative cells of Bac. thuringiensis var. galleriae (the wild-type strain 351) are much more sensitive to lethal effects of UV light and 60 Co-γ-rays than those of Bac. subtilis (the wild-type strain 168). This difference is less pronounced for spores of these strains. By means of repeated γ-irradiation-regrowth cycles radioresistant mutants Bac. thuringiensis Gamsup(r) 14 and Bac. subtilis Gamsup(r) 9 were selected. The vegetative cells of these mutants are correspondingly 19 times and 3.9 times more resistant to lethal effects of γ-radiation than the cells of the parental strains. The resistance of the Gamsup(r) mutant cells to lethal effects of UV light and H 2 O 2 is also increased. The spores of the Gamsup(r) 14 mutant are 1.5-1.7 times more resistant to γ-radiation and UV light than the wild-type spores. The radioresistant mutants and the parental strains do not vary in their capacity for host-cell reactivation of UV- or γ-irradiated phages Tg13 and 105

  14. In vitro ovicidal and cestocidal effects of toxins from Bacillus thuringiensis on the canine and human parasite Dipylidium caninum.

    Science.gov (United States)

    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 μ g/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 μ g/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.

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

    Directory of Open Access Journals (Sweden)

    Guadalupe Peña

    2013-01-01

    Full Text Available 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.

  16. Decolorization of textile dye RB19 using volcanic rock matrix immobilized Bacillus thuringiensis cells with surface displayed laccase.

    Science.gov (United States)

    Wan, Juan; Sun, Xiaowen; Liu, Cheng; Tang, Mengjun; Li, Lin; Ni, Hong

    2017-06-01

    A triplicate volcanic rock matrix-Bacillus thuringiensis-laccase WlacD (VRMs-Bt-WlacD) dye decolorization system was developed. WlacD was displayed on the B. thuringiensis MB174 cell surface to prepare a whole-cell laccase biocatalyst by using two repeat N-terminal domains of autolysin Mbg (Mbgn) 2 as the anchoring motif. Immunofluorescence microscopic assays confirmed that the fusion protein (Mbgn) 2 -WlacD was anchored on the surface of the recombinant B. thuringiensis MB174. After optimization by a single factor test, L 9 (3 4 )-orthogonal test, Plackett-Burman test, steepest ascent method, and Box-Behnken response surface methodology, the whole-cell specific laccase activity of B. thuringiensis MB174 was improved to 555.2 U L -1 , which was 2.25 times than that of the primary culture condition. Optimized B. thuringiensis MB174 cells were further adsorbed by VRMs to prepare VRMs-Bt-WlacD, an immobilized whole-cell laccase biocatalyst. Decolorization capacity of as-prepared VRMs-Bt-WlacD toward an initial concentration of 500 mg L -1 of an textile dye reactive blue 19 (RB19) aqueous solution reached 72.36% at a solid-to-liquid ratio of 10 g-100 mL. Repeated decolorization-activation operations showed the high decolorization capacity of VRMs-Bt-WlacD and have the potential for large-scale or continuous operations.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2005-08-18

    The sequencing and analysis of two close relatives of Bacillus anthracis are reported. AFLP analysis of over 300 isolates of B.cereus, B. thuringiensis and B. anthracis identified two isolates as being very closely related to B. anthracis. One, a B. cereus, BcE33L, was isolated from a zebra carcass in Nambia; the second, a B. thuringiensis, 97-27, was isolated from a necrotic human wound. The B. cereus appears to be the closest anthracis relative sequenced to date. A core genome of over 3,900 genes was compiled for the Bacillus cereus group, including Banthracis. Comparative analysis of these two genomes with other members of the B. cereus group provides insight into the evolutionary relationships among these organisms. Evidence is presented that differential regulation modulates virulence, rather than simple acquisition of virulence factors. These genome sequences provide insight into the molecular mechanisms contributing to the host range and virulence of this group of organisms.

  18. Mannose Phosphate Isomerase Isoenzymes in Plutella xylostella Support Common Genetic Bases of Resistance to Bacillus thuringiensis Toxins in Lepidopteran Species

    OpenAIRE

    Herrero, Salvador; Ferré, Juan; Escriche, Baltasar

    2001-01-01

    A strong correlation between two mannose phosphate isomerase (MPI) isoenzymes and resistance to Cry1A toxins from Bacillus thuringiensis has been found in a Plutella xylostella population. MPI linkage to Cry1A resistance had previously been reported for a Heliothis virescens population. The fact that the two populations share similar biochemical, genetic, and cross-resistance profiles of resistance suggests the occurrence of homologous resistance loci in both species.

  19. Effect of ultraviolet and gamma rays on the activity of delta-endotoxin protein crystals of Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Burges, H.D.; Hillyer, S.; Chanter, D.O.

    1975-01-01

    Sensitive bioassays with larvae of Pieris brassicae revealed no reduction of insecticidal activity as a result of severe gamma or ultraviolet irradiation of crystals of Bacillus thuringiensis (serotype V). The measured response was the inhibition of larval feeding by the crystals over exposure periods short enough for the presence of live spores not to influence feeding. The results were analyzed using a logistic model. (U.S.)

  20. Spatial and temporal distribution of airborne Bacillus thuringiensis var. kurstaki during an aerial spray program for gypsy moth eradication.

    OpenAIRE

    Teschke, K; Chow, Y; Bartlett, K; Ross, A; van Netten, C

    2001-01-01

    We measured airborne exposures to the biological insecticide Bacillus thuringiensis var. kurstaki (Btk) during an aerial spray program to eradicate gypsy moths on the west coast of Canada. We aimed to determine whether staying indoors during spraying reduced exposures, to determine the rate of temporal decay of airborne concentrations, and to determine whether drift occurred outside the spray zone. During spraying, the average culturable airborne Btk concentration measured outdoors within the...

  1. Extraction and separation of water soluble proteins from Bacillus thuringiensis-transgenic and non-transgenic maize species by CZE

    Czech Academy of Sciences Publication Activity Database

    Sázelová, Petra; Kašička, Václav; Ibanez, E.; Cifuentes, A.

    2009-01-01

    Roč. 32, č. 21 (2009), s. 3801-3808 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GA203/08/1428 Grant - others:GA ČR(CZ) GA203/09/0675 Program:GA Institutional research plan: CEZ:AV0Z40550506 Keywords : Bacillus thuringiensis -transgenic maize * CZE-UV profiling * Maize proteins Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.551, year: 2009

  2. Micro-Etched Platforms for Thermal Inactivation of Bacillus Anthracis and Bacillus Thuringiensis Spores

    Science.gov (United States)

    2008-03-01

    slips was first coated with a detergent wash. Commercially available Ivory soap shavings were diluted with sterile Millipore® water in a...environments. This removed controllable variability between the Bacillus species and increased the confidence in continued use of such surrogacy

  3. Expression of the sigma35 and cry2AB genes involved in Bacillus thuringiensis virulence Expressão dos genes sigma35 e cry2AB envolvidos na virulência de Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Ana Maria Guidelli-Thuler

    2009-06-01

    Full Text Available There are several genes involved in Bacillus thuringiensis sporulation. The regulation and expression of these genes results in an upregulation in Cry protein production, and this is responsible for the death of insect larvae infected by Bacillus thuringiensis. Gene expression was monitored in Bacillus thuringiensis during three developmental phases. DNA macroarrays were constructed for selected genes whose sequences are available in the GenBank database. These genes were hybridized to cDNA sequences from B. thuringiensis var. kurstaki HD-1. cDNA probes were synthesized by reverse transcription from B. thuringiensis RNA templates extracted during the exponential (log growth, stationary and sporulation phases, and labeled with 33PadCTP. Two genes were differentially expressed levels during the different developmental phases. One of these genes is related to sigma factor (sigma35, and the other is a cry gene (cry2Ab. There were differences between the differential levels of expression of various genes and among the expression detected for different combinations of the sigma factor and cry2Ab genes. The maximum difference in expression was observed for the gene encoding sigma35 factor in the log phase, which was also expressed at a high level during the sporulation phase. The cry2Ab gene was only expressed at a high level in the log phase, but at very low levels in the other phases when compared to the sigma35.Muitos genes estão envolvidos nos mecanismos de esporulação da bactéria Bacillus thuringiensis. A regulação e expressão desses genes resultam em uma produção massiva da proteína Cry, responsável pela morte das larvas de muitos insetos. Neste trabalho monitorou-se a expressão de genes de Bacillus thuringiensis, ao longo de três fases de seu desenvolvimento. Foram construídos macroarrays de DNA dos genes selecionados, cujas seqüências estão disponibilizadas no GenBank. Estes genes foram hibridizados com cDNAs obtidos de B

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

    Science.gov (United States)

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

    2013-01-01

    In order to overproduce bioinsecticides production by a sporeless Bacillus thuringiensis strain, an optimal composition of a cheap medium was defined using a response surface methodology. In a first step, a Plackett-Burman design used to evaluate the effects of eight medium components on delta-endotoxin production showed that starch, soya bean and sodium chloride exhibited significant effects on bioinsecticides production. In a second step, these parameters were selected for further optimisation by central composite design. The obtained results revealed that the optimum culture medium for delta-endotoxin production consists of 30 g L(-1) starch, 30 g L(-1) soya bean and 9 g L(-1) sodium chloride. When compared to the basal production medium, an improvement in delta-endotoxin production up to 50% was noted. Moreover, relative toxin yield of sporeless Bacillus thuringiensis S22 was improved markedly by using optimised cheap medium (148.5 mg delta-endotoxins per g starch) when compared to the yield obtained in the basal medium (94.46 mg delta-endotoxins per g starch). Therefore, the use of optimised culture cheap medium appeared to be a good alternative for a low cost production of sporeless Bacillus thuringiensis bioinsecticides at industrial scale which is of great importance in practical point of view.

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

    Directory of Open Access Journals (Sweden)

    Saoussen Ben Khedher

    2013-09-01

    Full Text Available In order to overproduce bioinsecticides production by a sporeless Bacillus thuringiensis strain, an optimal composition of a cheap medium was defined using a response surface methodology. In a first step, a Plackett-Burman design used to evaluate the effects of eight medium components on delta-endotoxin production showed that starch, soya bean and sodium chloride exhibited significant effects on bioinsecticides production. In a second step, these parameters were selected for further optimisation by central composite design. The obtained results revealed that the optimum culture medium for delta-endotoxin production consists of 30 g L-1 starch, 30 g L-1 soya bean and 9g L-1 sodium chloride. When compared to the basal production medium, an improvement in delta-endotoxin production up to 50% was noted. Moreover, relative toxin yield of sporeless Bacillus thuringiensis S22 was improved markedly by using optimised cheap medium (148.5 mg delta-endotoxins per g starch when compared to the yield obtained in the basal medium (94.46 mg delta-endotoxins per g starch. Therefore, the use of optimised culture cheap medium appeared to be a good alternative for a low cost production of sporeless Bacillus thuringiensis bioinsecticides at industrial scale which is of great importance in practical point of view.

  6. Low translocation of Bacillus thuringiensis israelensis to inner organs in mice after pulmonary exposure to commercial biopesticide

    DEFF Research Database (Denmark)

    Barfod, Kenneth Klingenberg; Ørum-Smidt, Lasse; Krogfelt, Karen A.

    2010-01-01

    Translocation of viable cells from a Bacillus thuringiensis israelensis-based biopesticide to inner organs in a mouse model was studied. Mice were exposed to the originally formulated product through the lungs and gastrointestinal tract by intratracheal instillation. Colony forming units (CFU) were...... grown from lungs, caecum, spleen and liver on Bacillus cereus-specific agar (BCSA) after 24 h and finally determined to be biopesticide strain B. t. israelensis by large plasmid profile. No CFU were found in spleen or liver of the control mice or in any aerosol background or material. We have shown...

  7. Structural studies of {delta}-endotoxin Cry 1 C from Bacillus thuringiensis

    Energy Technology Data Exchange (ETDEWEB)

    Guimaraes, B.G.; Garratt, R.C.; Oliva, G. [Sao Paulo Univ., Sao Carlos, SP (Brazil). Inst. de Fisica; Lemos, M.V.F. [UNESP, Jaboticabal, SP (Brazil). Dept. de Biologia Aplicada Agropecuaria; Arantes, O.M.N. [Universidade Estadual de Londrina, PR (Brazil). Dept. de Biologia Geral

    1996-12-31

    Full text. The {delta}-endotoxins are a family of crystal protein by a soil bacterium, Bacillus thuringiensis. The study of these proteins has been of great interest due to their highly specific activity against insects of the orders Lepidoptera, Diptera and Coleoptera. Thus, the {delta}a-endotoxins have been used for more than two decades as biological insecticides to control agricultural pests and, more recently, insects vectors of some diseases. The knowledge of their three-dimensional structures is very important to understand their mechanism of action and their high specificity. To date, the structure of only three proteins of the {delta}-endotoxins family have been reported: Cry3A, a coleopteran-specific toxin (beetle toxin){sup 1}, Cry1Aa, a lepidopteran-specific toxin (butterfly toxin){sup 2} and CytB, a dipteran-specific toxin (mosquito toxin){sup 3} Our work is aimed at the determination of the crystallographic structure by X-ray diffraction of {delta}-endotoxin Cry1C, also toxic to insects of the Lepidoptera order but towards families other than those affected by Cry1Aa. A comparison between these structures may lead to important conclusions about the reasons for the specificity and would allow the planning of mutants with more efficient activity. The cry1C gene was cloned into an adequate vector and expressed in an acrystalliferous B. thuringiensis strain. After cell culture and sporulation the microcrystals of Cry1C were separated by ultra-centrifugation in sacharose. The protoxin inclusion bodies were activated by commercial trpsin and the protease-resistant core was purified by anion-exchange chromatography. Crystallization experiments are being conducted in order to obtain single crystals suitable for diffraction measurements. We intend to use the Protein Crystallograph Station of the LNLS to collect data as soon as it is available and we have suitable crystals. (author) 3 refs.

  8. Long lasting persistence of Bacillus thuringiensis Subsp. israelensis (Bti in mosquito natural habitats.

    Directory of Open Access Journals (Sweden)

    Mathieu Tilquin

    Full Text Available BACKGROUND: The detrimental effects of chemical insecticides on the environment and human health have lead to the call for biological alternatives. Today, one of the most promising solutions is the use of spray formulations based on Bacillus thuringiensis subsp. israelensis (Bti in insect control programs. As a result, the amounts of Bti spread in the environment are expected to increase worldwide, whilst the common belief that commercial Bti is easily cleared from the ecosystem has not yet been clearly established. METHODOLOGY/MAIN FINDINGS: In this study, we aimed to determine the nature and origin of the high toxicity toward mosquito larvae found in decaying leaf litter collected in several natural mosquito breeding sites in the Rhône-Alpes region. From the toxic fraction of the leaf litter, we isolated B. cereus-like bacteria that were further characterized as B. thuringiensis subsp. israelensis using PCR amplification of specific toxin genes. Immunological analysis of these Bti strains showed that they belong to the H14 group. We finally used amplified length polymorphism (AFLP markers to show that the strains isolated from the leaf litter were closely related to those present in the commercial insecticide used for field application, and differed from natural worldwide genotypes. CONCLUSIONS/SIGNIFICANCE: Our results raise the issue of the persistence, potential proliferation and environmental accumulation of human-spread Bti in natural mosquito habitats. Such Bti environmental persistence may lengthen the exposure time of insects to this bio-insecticide, thereby increasing the risk of resistance acquisition in target insects, and of a negative impact on non-target insects.

  9. Gamma Radiation to Increase Efficiency of Bacillus thuringiensis Thai Strain for Insect Pets Control

    International Nuclear Information System (INIS)

    Chanpaisaeng, Jariya; Keawsompong, Suttipun; Piadang, Nattaya; Tephan, Prakai; Keawchingduang, Wannapa

    2006-09-01

    Bacillus thuringiensis (Bt) isolates JCPT16 and JCPT68 were gamma-irradiated at 2, 4, 6 and 8 kGy. The efficiency of these Bt isolates on S. litura control was also undertaken. It was found that the 4 kGy irradiated JCPT16 isolate had lowest LC 50 of 6.6x10 3 spore/ml while the non-irradiated JCPT 16 isolate had LC 50 of 6.2x10 3 spore/ml. Whereas the irradiated JCPT68 isolate at 8 kGy was noticed to have the lowest LC 50 of 2.7 x 10 3 spores/ml, the non-irradiated JCPT68 had LC 50 of 1.8x10 3 spores/ml. The efficiency test of B. thuringiensis isolate on S. exigua showed that the 2 kGy irradiated JCPT16 isolate had the lowest LC 50 of 2.52x10 4 spores/ml while the non-irradiated JCPT16 isolate had LC 50 of 6.04x10 3 spores/ml. The irradiated JCPT68 isolate at 4 kGy had the lowest LC 50 of 5.41x10 4 spores/ml, the non irradiated JCPT68 had LC 50 of 1.51x10 4 spores/ml. According to LC 50 values, there were no significant differences of efficiency on S. litura and S. exigua control among Bt isolates irradiated at various concentrations. The isolate JCPT16, JCPT35, JCPT50 and JCPT68 irradiated at dose of 10 kGy showed higher UV tolerance. After expose by UV ray, most of irradiated isolates still displayed high efficiency of controlling S. litura, S. exigua and Plutella xylostell.

  10. Structural studies of δ-endotoxin Cry 1 C from Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Guimaraes, B.G.; Garratt, R.C.; Oliva, G.; Lemos, M.V.F.; Arantes, O.M.N.

    1996-01-01

    Full text. The δ-endotoxins are a family of crystal protein by a soil bacterium, Bacillus thuringiensis. The study of these proteins has been of great interest due to their highly specific activity against insects of the orders Lepidoptera, Diptera and Coleoptera. Thus, the δa-endotoxins have been used for more than two decades as biological insecticides to control agricultural pests and, more recently, insects vectors of some diseases. The knowledge of their three-dimensional structures is very important to understand their mechanism of action and their high specificity. To date, the structure of only three proteins of the δ-endotoxins family have been reported: Cry3A, a coleopteran-specific toxin (beetle toxin) 1 , Cry1Aa, a lepidopteran-specific toxin (butterfly toxin) 2 and CytB, a dipteran-specific toxin (mosquito toxin) 3 Our work is aimed at the determination of the crystallographic structure by X-ray diffraction of δ-endotoxin Cry1C, also toxic to insects of the Lepidoptera order but towards families other than those affected by Cry1Aa. A comparison between these structures may lead to important conclusions about the reasons for the specificity and would allow the planning of mutants with more efficient activity. The cry1C gene was cloned into an adequate vector and expressed in an acrystalliferous B. thuringiensis strain. After cell culture and sporulation the microcrystals of Cry1C were separated by ultra-centrifugation in sacharose. The protoxin inclusion bodies were activated by commercial trpsin and the protease-resistant core was purified by anion-exchange chromatography. Crystallization experiments are being conducted in order to obtain single crystals suitable for diffraction measurements. We intend to use the Protein Crystallograph Station of the LNLS to collect data as soon as it is available and we have suitable crystals. (author)

  11. Biochemical, immunological and toxicological characteristics of the crystal proteins of Bacillus thuringiensis subsp. medellin

    Directory of Open Access Journals (Sweden)

    Sergio Orduz

    1996-04-01

    Full Text Available Characterization of the insecticidal and hemolytic activity of solubilized crystal proteins of Bacillus thuringiensis (Bt subsp. medellin (Btmed was performed and compared to solubilized crystal proteins of isolates 1884 of B. thuringiensis subsp. israelensis (Bti and isolate PG-14 of B. thuringiensis subsp. morrisoni (Btm. In general, at acid pH values solubilization of the Bt crystalline parasporal inclusions (CPI was lower than at alkaline pH. The larvicidal activity demonstrated by the CPI of Btmed indicated that optimal solubilization of CPI takes place at a pH value of 11.3, in Bti at pH values from 5.03 to 11.3 and in Btm at pH values from 9.05 to 11.3. Hemolytic activity against sheep red blood cells was mainly found following extraction at pH 11.3 in all Bt strains tested. Polyacrylamide gel electrophoresis under denaturing conditions revealed that optimal solubilization of the CPI in all Bt strains takes place at the alkaline pH values from 9.05 to 11.3. An enriched preparation of Btmed crystals was obtained, solubilized and crystal proteins were separated on a size exclusion column (Sephacryl S-200. Three main protein peaks were observed on the chromatogram. The first peak had two main proteins that migrate between 90 to 100 kDa. These proteins are apparently not common to other Bt strains isolated to date. The second and third peaks obtained from the size exclusion column yielded polypeptides of 68 and 28-30 kDa, respectively. Each peak independently, showed toxicity against 1st instar Culex quinquefasciatus larvae. Interestingly, combinations of the fractions corresponding to the 68 and 30 kDa protein showed an increased toxicity. These results suggest that the 94 kDa protein is an important component of the Btmed toxins with the highest potency to kill mosquito larvae. When crystal proteins of Bti were probed with antisera raised independently against the three main protein fractions of Btmed, the only crystal protein that showed

  12. Bacillus thuringiensis in caterpillars and associated materials collected from protected tropical forests in northwestern Costa Rica

    Directory of Open Access Journals (Sweden)

    César Rodríguez-Sánchez

    2006-06-01

    Full Text Available Bacillus thuringiensis (Bt synthesizes crystalline inclusions that are toxic to caterpillars (Lepidoptera and other orders of invertebrates. Materials associated with 37 caterpillars from 16 species, collected while feeding on 15 different species of host plants in dry, cloud and rain forests located in the Área de Conservación Guanacaste in northwestern Costa Rica, were examined for the presence of Bt. From a total of 101 derived samples, 25 Bt isolates were cultured: 56% from host plant leaves, 8% from caterpillar guts and 36% from caterpillar fecal pellets. Bt was isolated from at least one sample in 38% of the systems constituted by the food plant, gut and fecal pellets corresponding to a single caterpillar. Four different morphologies of crystalline inclusions were observed, with bipyramidal and irregular crystal morphologies being the most prevalent. Rev. Biol. Trop. 54(2: 265-271. Epub 2006 Jun 01.Bacillus thuringiensis (Bt sintetiza inclusiones cristalinas que resultan tóxicas para algunas larvas de lepidópteros y otros órdenes de invertebrados. Su presencia fue examinada en materiales asociados a 37 orugas de mariposas de 16 especies, las cuales fueron colectadas mientras se alimentaban en 15 especies diferentes de plantas hospederas en bosques secos, nubosos y húmedos localizados dentro del Área de Conservación Guanacaste (ACG en el noroeste de Costa Rica. A partir de un total de 101 muestras se obtuvo 25 aislamientos de Bt: 56% a partir de material foliar de las plantas hospederas, 8% a partir del contenido intestinal de las larvas y 36% a partir de sus excrementos. Esta bacteria fue cultivada a partir de al menos uno de los 3 diferentes tipos de muestra asociados a una oruga particular (planta hospedera, intestino, excremento en 38% de los casos posibles. En la colección de aislamientos obtenida se observaron cuatro morfologías de inclusiones cristalinas, siendo aquellas bipiramidales e irregulares las más prevalentes.

  13. Susceptibility of The Asian Corn Borer, Ostrinia furnacalis, to Bacillus thuringiensis Toxin CRY1AC

    Directory of Open Access Journals (Sweden)

    Aye Kyawt Kyawt Ei

    2008-07-01

    Full Text Available The larval susceptibility of the Asian corn borer, Ostrinia furnacalis (Guenee (Lepidoptera: Crambidae, to a Bacillus thuringiensis protein (Cry1Ac was evaluated using insect feeding bioassays. The founding population of O. furnacalis was originally collected from the experimental station of UGM at Kalitirto and had been reared in the laboratory for three generations using an artificial diet “InsectaLf”. The tested instars were exposed on diets treated with a series of concentrations of Cry1Ac for one week. The LC50 values on the seventh day after treatment for 1st, 2nd, 3rd and 4th instars were 7.79, 21.12, 113.66, and 123.17 ppm, respectively, showing that the higher the instars the lesser the susceptibility to Cry1Ac. When the neonates were exposed to sublethal concentrations of Cry1Ac (0.0583, 0.116, and 0.5830 ppm, growth and development of the surviving larvae were inhibited. The fecundity and viability of females produced from treated larvae decreased with increasing the concentrations. These findings indicate that Cry1Ac is toxic to larva of O. furnacalis and has chronic effects to larvae surviving from Cry1Ac ingestion.   Kepekaan larva penggerek batang jagung Asia, Ostrinia furnacalis (Guenee (Lepidoptera: Crambidae, terhadap protein Bacillus thuringiensis Cry1Ac diuji dengan metode celup pakan. Larva berasal dari pertanaman jagung di KP-4, UGM di Kalitirto dan telah dikembangbiakkan di laboratorium menggunakan pakan buatan (InsectaLF selama tiga generasi sebelum digunakan untuk pengujian. Larva O. furnacalis yang diuji dipaparkan pada pakan buatan yang telah dicelupkan pada seri konsentrasi Cry1Ac. Nilai LC50 pada hari ketujuh setelah perlakukan untuk instar 1, 2, 3, dan 4 berturut-turut adalah 0,79; 21,12; 113,66; dan 123,17 ppm. Hal ini menunjukkan bahwa instar yang semakin tinggi tingkat kepekaannya terhadap Cry1Ac semakin menurun. Larva yang baru menetas dan diberi pakan yang telah dicelupkan pada konsentrasi sublethal Cry1Ac

  14. Resistance to Bacillus thuringiensis by the Indian meal moth, Plodia interpunctella: comparison of midgut proteinases from susceptible and resistant larvae.

    Science.gov (United States)

    Johnson, D E; Brookhart, G L; Kramer, K J; Barnett, B D; McGaughey, W H

    1990-03-01

    Midgut homogenates from susceptible and resistant strains of the Indian meal moth, Plodia interpunctella, were compared for their ability to activate the entomocidal parasporal crystal protein from Bacillus thuringiensis. The properties of midgut proteinases from both types of larvae were also examined. Electrophoretic patterns of crystal protein from B. thuringiensis subspecies kurstaki (HD-1) and aizawai (HD-133 and HD-144) were virtually unchanged following digestion by either type of midgut homogenate. Changes in pH (9.5 to 11.5) or midgut homogenate concentration during digestion failed to substantially alter protein electrophoretic patterns of B. thuringiensis HD-1 crystal toxin. In vitro toxicity of crystal protein activated by either type of midgut preparation was equal toward cultured insect cells from either Manduca sexta or Choristoneura fumiferana. Electrophoresis of midgut extracts in polyacrylamide gels containing gelatin as substrate also yielded matching mobility patterns of proteinases from both types of midguts. Quantitation of midgut proteolytic activity using tritiated casein as a substrate revealed variation between midgut preparations, but no statistically significant differences between proteolytic activities from susceptible and resistant Indian meal moth larvae. Inhibition studies indicated that a trypsin-like proteinase with maximal activity at pH 10 is a major constituent of Indian meal moth midguts. The results demonstrated that midguts from susceptible and resistant strains of P. interpunctella are similar both in their ability to activate B. thuringiensis protoxin and in their proteolytic activity.

  15. [Bacillus thuringiensis: general aspects. An approach to its use in the biological control of lepidopteran insects behaving as agricultural pests].

    Science.gov (United States)

    Sauka, Diego H; Benintende, Graciela B

    2008-01-01

    Bacillus thuringiensis is the most widely applied biological pesticide used to control insects that affect agriculture and forestry and which transmit human and animal pathogens. During the past decades B. thuringiensis has been the subject of intensive research. These efforts have yielded considerable data about the relationships between the structure, mechanism of action, and genetics of their pesticidal crystal proteins. As a result, a coherent picture of these relationships has emerged. Other studies have focused on the ecological role of the B. thuringiensis crystal proteins and their performance in agricultural and other natural settings. With this knowledge as background and the help of biotechnological tools, researchers are now reporting promising results in the development of more useful toxins, recombinant bacteria, new formulations and transgenic plants that express pesticidal activity, in order to assure that these products are utilized with the best efficiency and benefit. This article is an attempt to integrate all these recent developments in the study of B. thuringiensis into a context of biological control of lepidopteran insect pest of agricultural importance.

  16. Avaliação de produtos à base de Bacillus thuringiensis no controle da traça-das-crucíferas Evaluation of insecticides based on Bacillus thuringiensis in the control of the diamondback moth

    Directory of Open Access Journals (Sweden)

    Patrícia T Medeiros

    2006-06-01

    Full Text Available Avaliou-se em dois experimentos a suscetibilidade da traça-das-crucíferas a inseticidas à base de Bacillus thuringiensis em repolho cv. Itiban. O delineamento do primeiro experimento (de julho a setembro/03, em área de plantio comercial em Brazlândia (DF, foi de blocos casualizados, com seis tratamentos e dez repetições; os bioinseticidas utilizados foram B. thuringiensis kurstaki (S1450CO, B. thuringiensis aizawai comercial (Bta e três produtos formulados com as estirpes S1450BB, S811BB, S845BB de B. thuringiensis pertencentes ao Banco de Bacillus spp. da Embrapa Recursos Genéticos e Biotecnologia. No segundo experimento, realizado no campo experimental da Embrapa (DF, de outubro/03 a janeiro/04, o delineamento foi de blocos casualizados, com seis tratamentos e quatro repetições; os inseticidas utilizados foram os mesmos do primeiro experimento, com a adição de Spinosad, e retirado o tratamento S811BB. Os produtos foram aplicados quando foi atingido o nível de dano de seis furos nas quatro folhas centrais do repolho. O Bta comercial foi o produto mais eficaz no primeiro experimento, tendo sido aplicado cinco vezes e diferiu estaticamente dos demais produtos. Os formulados S845BB e S1450BB não apresentaram diferenças quando comparados ao produto comercial S1450 e foram aplicados seis vezes. O produto S811BB também foi aplicado seis vezes, mas sua eficácia foi inferior aos demais produtos e não diferiu da testemunha. Já no segundo experimento, o S1450 comercial foi aplicado cinco vezes e os demais produtos à base de Bt, seis vezes. Todos os produtos utilizados não diferiram entre si, diferindo apenas no número de aplicações.Two experiments were performed to evaluate the susceptibility of the diamondback moth to insecticides based on Bacillus thuringiensis in cabbage cv. Itiban. The first experiment was carried out from July to September 2003, in a production area in Brazlândia (DF, Brazil. Randomized blocks with six

  17. Elucidation of the mechanisms of CryIIIA overproduction in a mutagenized strain of Bacillus thuringiensis var. tenebrionis

    International Nuclear Information System (INIS)

    Adams, L.F.; Mathewes, S.; O'Hara, P.; Peterson, A.; Gürtler, H.

    1994-01-01

    NB176 is a Bacillus thuringiensis mutant derived by λ-irradiation of NB125 Bacillus thuringiensis var. tenebrionis (Krieg). It exhibits two interesting phenotypes: (i) oligosporogeny and (ii) twofold to threefold overproduction of the CryIIIA protein. Southern profiles of the NB176 strain showed an additional copy(s) of the cryIIIA gene located on a 4 kb HindIII fragment, in addition to the expected cryIIIA gene on a 3 kb HindIII fragment. Each cryIIIA gene-bearing HindIII fragment was cloned from NB176. The restriction map of the 3 kb HindIII fragment was identical to that published by Donovan and coworkers. Sequencing of the 4 kb HindIII fragment showed no alterations in the promoter region of the cryIIIA gene but did show replacement of the region immediately following the cryIIIA open reading frame with a sequence encoding a transposase with 50% amino acid homology to that of Tn 1000. These findings suggest that the overproduction phenotype of NB176 results from extra copies of the cryIIIA gene produced from a transposition event(s) induced or stabilized by γ-irradiation. Integration of additional copies of the cryIIIA gene into the native 90MDa plasmid of the wild-type B. thuringiensis var. tenebrionis strain resulted in strains that made enormous crystals, many possessing greatly enhanced insecticidal activity

  18. Susceptibility of Spodoptera frugiperda and S. exigua to Bacillus thuringiensis Vip3Aa insecticidal protein.

    Science.gov (United States)

    Chakroun, Maissa; Bel, Yolanda; Caccia, Silvia; Abdelkefi-Mesrati, Lobna; Escriche, Baltasar; Ferré, Juan

    2012-07-01

    The Vip3Aa protein is an insecticidal protein secreted by Bacillus thuringiensis during the vegetative stage of growth. The activity of this protein has been tested after different steps/protocols of purification using Spodoptera frugiperda as a control insect. The results showed that the Vip3Aa protoxin was stable and retained full toxicity after being subjected to common biochemical steps used in protein purification. Bioassays with the protoxin in S. frugiperda and S. exigua showed pronounced differences in LC(50) values when mortality was measured at 7 vs. 10d. At 7d most live larvae were arrested in their development. LC(50) values of "functional mortality" (dead larvae plus larvae remaining in the first instar), measured at 7d, were similar or even lower than the LC(50) values of mortality at 10d. This strong growth inhibition was not observed when testing the trypsin-activated protein (62 kDa) in either species. S. exigua was less susceptible than S. frugiperda to the protoxin form, with LC(50) values around 10-fold higher. However, both species were equally susceptible to the trypsin-activated form. Processing of Vip3Aa protoxin to the activated form was faster with S. frugiperda midgut juice than with S. exigua midgut juice. The results strongly suggest that the differences in the rate of activation of the Vip3Aa protoxin between both species are the basis for the differences in susceptibility towards the protoxin form. Copyright © 2012 Elsevier Inc. All rights reserved.

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

    Science.gov (United States)

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

    2017-12-01

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

  20. Isolation and characterization of Bacillus thuringiensis strains active against Elasmopalpus lignosellus (Zeller, 1848 (Lepidoptera, Pyralidae

    Directory of Open Access Journals (Sweden)

    Janaina Zorzetti

    2017-08-01

    Full Text Available Elasmopalpus lignosellus (Zeller, 1848 (Lepidoptera, Pyralidae is an insect pest of 60 economically important crops, including sugarcane, wheat, soybean, rice, beans, sorghum, peanuts, and cotton. The aim of this work was to select and characterize Bacillus thuringiensis isolates with insecticidal activity against E. Lignosellus that could be used as an alternative method of control. Selective bioassays were done to evaluate the toxicity of 47 isolates against first instar larvae of E. lignosellus. For the most toxic bacterial strains, the lethal concentration (LC50 was estimated and morphological, biochemical and molecular methods were used to characterize the isolates. Among the 47 isolates tested, 12 caused mortality above 85% and showed LC50 values from 0.038E+8 to 0.855E+8 spores mL-1. Isolates BR83, BR145, BR09, BR78, S1534, and S1302 had the lowest LC50 values and did not differ from the standard HD-1 strain; the exception was BR83.The protein profiles produced bands with molecular masses of 60-130 kDa. The genes cry1, cry2, cry3, and cry11 were identified in the molecular characterization. The morphological analysis identified three different crystal inclusions: bipyramidal, spherical and cuboidal. Among the tested isolates, 12 isolates have potential for biotechnological control of E. Lignosellus by development of new biopesticides or genetically modified plants.

  1. Proteomic Analysis of Bacillus thuringiensis Strain 4.0718 at Different Growth Phases

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    Xiaohui Li

    2012-01-01

    Full Text Available The growth process of Bacillus thuringiensis Bt4.0718 strain was studied using proteomic technologies. The proteins of Bt whole cells at three phases—middle vegetative, early sporulation, and late sporulation—were extracted with lysis buffer, followed with separation by 2-DE and identified by MALDI-TOF/TOF MS. Bioactive factors such as insecticidal crystal proteins (ICPs including Cry1Ac(3, Cry2Aa, and BTRX28, immune inhibitor (InhA, and InhA precursor were identified. InhA started to express at the middle vegetative phase, suggesting its contribution to the survival of Bt in the host body. At the early sporulation phase, ICPs started their expression. CotJC, OppA, ORF1, and SpoIVA related to the formation of crystals and spores were identified, the expression characteristics of which ensured the stable formation of crystals and spores. This study provides an important foundation for further exploration of the stable expression of ICPs, the smooth formation of crystals, and the construction of recombinant strains.

  2. A single type of cadherin is involved in Bacillus thuringiensis toxicity in Plutella xylostella.

    Science.gov (United States)

    Park, Y; Herrero, S; Kim, Y

    2015-12-01

    Cadherins have been described as one the main functional receptors for the toxins of the entomopathogenic bacterium, Bacillus thuringiensis (Bt). With the availability of the whole genome of Plutella xylostella, different types of cadherins have been annotated. In this study we focused on determining those members of the cadherin-related proteins that potentially play a role in the mode of action of Bt toxins. For this, we mined the genome of P. xylostella to identify these putative cadherins. The genome screening revealed 52 genes that were annotated as cadherin or cadherin-like genes. Further analysis revealed that six of these putative cadherins had three motifs common to all Bt-related cadherins: a signal peptide, cadherin repeats and a transmembrane domain. From the six selected cadherins, only P. xylostella cadherin 1 (PxCad1) was expressed in the larval midgut and only the silencing of this gene by RNA interference (double-stranded RNA feeding) reduce toxicity and binding to the midgut of the Cry1Ac type toxin from Bt. These results indicate that from the whole set of cadherin-related genes identified in P. xylostella, only PxCad1 is associated with the Cry1Ac mode of action. © 2015 The Royal Entomological Society.

  3. Toxicity of radiation-resistant strains of Bacillus thuringiensis (Berl.) to larval Plutella xylostella (L.)

    International Nuclear Information System (INIS)

    Jangi, M.S.; Ibrahim, Hasan

    1983-01-01

    A total of 24 isolates of Bacillus thuringiensis (Berliner), resistant to a γ-radiation dose of 100 krad, were screened for their toxicity to larval silkworms, Bombyxmori(L.), and 15 of them were subsequently tested for their toxicity to larval diamond-back moth, Plutella xylostella(L.). The LC 50 's of these isolates to B. mori ranged from 1.6 X 10 5 to 6.0 X 10 3 spores/mL or from 5.9 to 0.3 μg cellular protein/mL. The irradiation treatment produced isolates which were significantly more toxic to P. xylostella (LC 50 4 spores/mL or 3.7 μg cellular protein/mL) and/ or less toxic to B. mori (LC 50 > 2.3 X 10 4 spores/mL or 1.0 μg cellular protein/mL) than the parent commercial strain

  4. Novel genetic factors involved in resistance to Bacillus thuringiensis in Plutella xylostella.

    Science.gov (United States)

    Ayra-Pardo, C; Raymond, B; Gulzar, A; Rodríguez-Cabrera, L; Morán-Bertot, I; Crickmore, N; Wright, D J

    2015-12-01

    The widespread and sustainable exploitation of the entomopathogen Bacillus thuringiensis (Bt) in pest control is threatened by the evolution of resistance. Although resistance is often associated with loss of binding of the Bt toxins to the insect midgut cells, other factors have been implicated. Here we used suppressive subtractive hybridization and gene expression suppression to identify additional molecular components involved in Bt-resistance in Plutella xylostella. We isolated transcripts from genes that were differentially expressed in the midgut of larvae from a resistant population, following ingestion of a Bt kurstaki HD1 strain-based commercial formulation (DiPel), and compared with a genetically similar susceptible population. Quantitative real-time polymerase-chain reaction (RT-PCR) analysis confirmed the differential basal expression of a subset of these genes. Gene expression suppression of three of these genes (P. xylostella cyclin-dependent kinase 5 regulatory subunit associated protein 1-like 1, stromal cell-derived factor 2-like 1 and hatching enzyme-like 1) significantly increased the pathogenicity of HD1 to the resistant population. In an attempt to link the multitude of factors reportedly influencing resistance to Bt with the well-characterized loss of toxin binding, we also considered Bt-resistance models in P. xylostella and other insects. © 2015 The Royal Entomological Society.

  5. Extraction of flocculants from a strain of Bacillus thuringiensis and analysis of their properties

    Directory of Open Access Journals (Sweden)

    Jingrong Wu

    2017-07-01

    Full Text Available In a preliminary screening study, our laboratory isolated from the biofloc in aquaculture waters a strain of Bacillus thuringiensis, which produced highly efficient bio-flocculants. In the present study, we extracted the crude flocculants from this strain and analyzed their properties. Distribution analysis indicated that the flocculants were mainly distributed in the supernatant of the fermentation liquid. The flocculants were extracted using an ethanol extraction method, and the chemical compositions and morphology of the crude flocculants were analyzed using the Molish reaction, Fehling reaction, ninhydrin reaction, biuret reaction, phenol-sulfuric acid assay, Coomassie brilliant blue staining, ultraviolet scanning, infrared scanning and scanning electron microscopy. The carbohydrate composition of the polysaccharides in the flocculants was analyzed with thin layer chromatography. The results indicated that flocculants were solid substances with an ivory white color and their texture was loose and soft. Visualization under scanning electron microscopy revealed that their ultra-morphology consisted of small, long and fiber-like shapes. Chemical and physical analyses indicated that polysaccharides accounted for 34.5% of the components in the crude flocculants. The monosaccharides present in crude flocculants included mainly glucose, galactose and mannitose.

  6. Molecular Approaches to Improve the Insecticidal Activity of Bacillus thuringiensis Cry Toxins

    Directory of Open Access Journals (Sweden)

    Wagner A. Lucena

    2014-08-01

    Full Text Available Bacillus thuringiensis (Bt is a gram-positive spore-forming soil bacterium that is distributed worldwide. Originally recognized as a pathogen of the silkworm, several strains were found on epizootic events in insect pests. In the 1960s, Bt began to be successfully used to control insect pests in agriculture, particularly because of its specificity, which reflects directly on their lack of cytotoxicity to human health, non-target organisms and the environment. Since the introduction of transgenic plants expressing Bt genes in the mid-1980s, numerous methodologies have been used to search for and improve toxins derived from native Bt strains. These improvements directly influence the increase in productivity and the decreased use of chemical insecticides on Bt-crops. Recently, DNA shuffling and in silico evaluations are emerging as promising tools for the development and exploration of mutant Bt toxins with enhanced activity against target insect pests. In this report, we describe natural and in vitro evolution of Cry toxins, as well as their relevance in the mechanism of action for insect control. Moreover, the use of DNA shuffling to improve two Bt toxins will be discussed together with in silico analyses of the generated mutations to evaluate their potential effect on protein structure and cytotoxicity.

  7. Acute, sublethal, and combination effects of azadirachtin and Bacillus thuringiensis on the cotton bollworm, Helicoverpa armigera.

    Science.gov (United States)

    Abedi, Zahra; Saber, Moosa; Vojoudi, Samad; Mahdavi, Vahid; Parsaeyan, Ehsan

    2014-02-26

    The cotton bollworm, Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) is a polyphagous and cosmopolitan insect pest that causes damage to various plants. In this study, the lethal and sublethal effects of azadirachtin and Bacillus thuringiensis Berliner sub sp . kurstaki (Bacillales: Bacillaceae) were evaluated on third instar H. armigera under laboratory conditions. The LC50 values of azadirachtin and Bt were 12.95 and 96.8 µg a.i./mL, respectively. A total mortality of 56.7% was caused on third instar larvae when LC20 values of the insecticides were applied in combination with each other. The LT50 values of azadirachtin and Bt were 4.8 and 3.6 days, respectively. The results of the sublethal study showed that the application of LC30 value of azadirachtin and Bt reduced the larval and pupal weight and increased larval and pupal duration of H. armigera. The longevity and fecundity of female adults were affected significantly by the insecticides. Female fecundity was reduced by the treatments, respectively. The lowest adult emergence ratio and pupation ratio were observed in the azadirachtin treatment. The results indicated that both insecticides have high potential for controlling of the pest. This is an open access paper. We use the Creative Commons Attribution 3.0 license that permits unrestricted use, provided that the paper is properly attributed.

  8. Toxicity of radiation-resistant strains of Bacillus thuringiensis (Berl. ) to larval Plutella xylostella (L. )

    Energy Technology Data Exchange (ETDEWEB)

    Jangi, M.S.; Ibrahim, H. (Faculty of Health Sciences, Universiti Kebangsaan, Malysia, Bangi, Selangor)

    1983-05-01

    A total of 24 isolates of Bacillus thuringiensis (Berliner), resistant to a ..gamma..-radiation dose of 100 krad, were screened for their toxicity to larval silkworms, Bombyxmori(L.), and 15 of them were subsequently tested for their toxicity to larval diamond-back moth, Plutella xylostella(L.). The LC/sub 50/'s of these isolates to B. mori ranged from 1.6 X 10/sup 5/ to 6.0 X 10/sup 3/ spores/mL or from 5.9 to 0.3 ..mu..g cellular protein/mL. The irradiation treatment produced isolates which were significantly more toxic to P. xylostella (LC/sub 50/ < 8.1 X 10/sup 4/ spores/mL or 3.7 ..mu..g cellular protein/mL) and/ or less toxic to B. mori (LC/sub 50/ > 2.3 X 10/sup 4/ spores/mL or 1.0 ..mu..g cellular protein/mL) than the parent commercial strain.

  9. No adjuvant effect of Bacillus thuringiensis-maize on allergic responses in mice.

    Directory of Open Access Journals (Sweden)

    Daniela Reiner

    Full Text Available 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.

  10. SUSCETIBILIDADE DE Helicoverpa armigera Hübner A FORMULADOS À BASE DE Bacillus thuringiensis BERLINER

    Directory of Open Access Journals (Sweden)

    Victor Luiz de Souza Lima

    2017-01-01

    Full Text Available O inseto Helicoverpa armigera recentemente registrado no Brasil é uma das maiores pragas da agricultura mundial. Pode atacar mais de 200 espécies de plantas e possui populações resistentes a diversos inseticidas. A utilização de microrganismos com potencial patogênico contra insetos é uma alternativa aos inseticidas. Essa pesquisa foi realizada com o objetivo de determinar a suscetibilidade de lagartas de H. armigera à produtos formulados à base de Bacillus thuringiensis (Bt Para os experimentos, foram utilizados os produtos comerciais Dipel® e Agree®, os quais tiveram sua concentração ajustada para 108 conídios viáveis ml-1. Essa concentração foi aplicada sobre dieta artificial, a qual foi colocada em uma placa de Petri que continha 10 lagartas de primeiro instar. Foram realizadas cinco repetições para cada produto. As avaliações foram feitas a cada 24h durante sete dias. Os produtos comerciais Dipel® e Agree® causaram, respectivamente, 100% e 94% de mortalidade das lagartas de H. armigera. Esse resultado mostra o potencial de produtos à base de Bt sobre H. armigera.

  11. Crystallization and preliminary crystallographic analysis of poly(3-hydroxybutyrate) depolymerase from Bacillus thuringiensis.

    Science.gov (United States)

    Wang, Yung Lin; Lin, Yi Ting; Chen, Chia Lin; Shaw, Gwo Chyuan; Liaw, Shwu Huey

    2014-10-01

    Poly[(R)-3-hydroxybutyrate] (PHB) is a microbial biopolymer that has been commercialized as biodegradable plastics. The key enzyme for the degradation is PHB depolymerase (PhaZ). A new intracellular PhaZ from Bacillus thuringiensis (BtPhaZ) has been screened for potential applications in polymer biodegradation. Recombinant BtPhaZ was crystallized using 25% polyethylene glycol 3350, 0.2 M ammonium acetate, 0.1 M bis-tris pH 6.5 at 288 K. The crystals belonged to space group P1, with unit-cell parameters a = 42.97, b = 83.23, c = 85.50 Å, α = 73.45, β = 82.83, γ = 83.49°. An X-ray diffraction data set was collected to 1.42 Å resolution with an Rmerge of 6.4%. Unexpectedly, a molecular-replacement solution was obtained using the crystal structure of Streptomyces lividans chloroperoxidase as a template, which shares 24% sequence identity to BtPhaZ. This is the first crystal structure of an intracellular poly(3-hydroxybutyrate) depolymerase.

  12. Resistance and behavioural response of Plutella xylostella (Lepidoptera: Plutellidae) populations to Bacillus thuringiensis formulations.

    Science.gov (United States)

    Zago, Hugo B; Siqueira, Herbert Á A; Pereira, Eliseu J G; Picanço, Marcelo C; Barros, Reginaldo

    2014-03-01

    Insecticide resistance is probably the major cause of control failure of Plutella xylostella (L.) in Brazil. In most production regions, the use of chemicals has been the prevalent method of control, with reduced efficacy through cropping seasons, even for the most recent use of products based on Bacillus thuringiensis (Bt). The current status of the resistance to these products was assessed, as well as the behavioural response of P. xylostella populations to Bt sprays. Most populations of P. xylostella were resistant to Bt products, particularly to Xentari®WDG (2-54-fold). Differences in walking characteristics of larvae were variable for most populations, for both Dipel®WP and Xentari®WDG, but not associated with resistance. Most females preferred to lay eggs on untreated surfaces and showed a reduced proportion of oviposition on treated surfaces that only correlated with resistance to Dipel®WP (r = -0.74, P = 0.02). Broad and indiscriminate use of Bt-based products has selected Brazilian P. xylostella populations to resistance. Larval movement appears to be a resistance-independent mechanism. Most populations of P. xylostella preferred to lay eggs on Bt-free surfaces, which might be a result of growers' practice of spraying the cabbage head. Reduced oviposition on treated surfaces correlated with physiological resistance, suggesting a behavioural response among the Bt-resistant colonies to Dipel®WP. © 2013 Society of Chemical Industry.

  13. EFEKTIVITAS Bacillus thuringiensis H-14 STRAIN LOKAL DALAM BUAH KELAPA TERHADAP LARVA Anopheles sp dan Culex sp di KAMPUNG LAUT KABUPATEN CILACAP

    Directory of Open Access Journals (Sweden)

    Blondine Ch. P

    2013-07-01

    Full Text Available Abstrak Bacillus thuringiensis serotipe H-14 strain lokal adalah bakteri patogen bersifat target spesifiknya larva nyamuk, aman bagi mamalia dan lingkungan. Penelitian bertujuan menentukan efektivitas B. thuringiensis H-14 strain lokal yang dikembangbiakkan dalam buah kelapa untuk pengendalian larva Anopheles sp dan Culex sp. Rancangan eksperimental semu, terdiri dari kelompok perlakuan dan kontrol. Bacillus thuringiensis H-14 strain lokal dikembangbiakan dalam10 buah kelapa umur 6–8 bulan, dengan berat kira-kira 1 kg, telah berisi air kelapa sekitar 400-500 ml/buah kelapa yang diperoleh dari Desa Klaces, Kampung Laut, Kabupaten Cilacap. Diinkubasi selama 14 hari pada temperatur kamar dan ditebarkan di 6 kolam yang menjadi habitat perkembangbiakan larva nyamuk dengan luas berkisar 3–100 m2.Hasil yang diperoleh menunjukkan efektivitas B. thuringiensis H-14 strain lokal terhadap larva Anopheles sp dan Culex sp selama 1 hari sesudah penebaran kematian larva berturut-turut sebesar 80–100% dan 79,31–100%. Sedangkan pada hari ke-14 sebesar 69,30–76,71% dan 67,69–86,04%. Buah kelapa dapat digunakan sebagai media lokal alternatif untuk pengembangbiakan B. thuringiensis H-14 strain lokal Kata kunci: B. thuringiensis H-14,  strain  lokal, buah kelapa, pengendalian larva Abstract Bacillus thuringiensis serotype H-14 local strain is pathogenic bacteria which specific  target to mosquito larvae. It is safe for mammals and enviroment. The aims of this study was to determine the effectivity of B. thuringiensis H-14 local strain which culturing in thecoconut wates against Anopheles sp and Culex sp mosquito larvae. This research is quasi experiment which consist of treated  and control groups. Bacillus thuringiensis H-14 local strain was cultured in 10 coconuts with 6–8 months age with weight around 1 kg that contained were approximately 400-500 ml/coconut were taken from Klaces village, Kampung Laut. After that the coconuts incubated for 14

  14. Avaliação da eficiência de formulações de Bacillus thuringiensis para o controle de traça-das-crucíferas em repolho no Distrito Federal Efficiency of Bacillus thuringiensis formulations in controlling Diamondback Moth in cabbage in the Federal District

    Directory of Open Access Journals (Sweden)

    Marina Castelo Branco

    1999-11-01

    Full Text Available A traça-das-crucíferas (Plutella xylostella é a praga mais importante do repolho no Distrito Federal. Seu controle é feito basicamente com inseticidas. Novos produtos são constantemente avaliados para o controle da praga e neste trabalho duas novas formulações de Bacillus thuringiensis [Bacillus thuringiensis var. aizawai (350 e 500 g/hae B. thuringiensis var. kurstaki x B. thuringiensis var. aizawai (350 e 500 ml/ha] foram avaliadas no período de maio a outubro de 1995. Os tratamentos B. thuringiensis var. kurstaki (500 ml/ha, deltametrina (240 ml/ha e uma testemunha sem pulverização foram também incluídos no experimento. O delineamento foi blocos ao acaso, com sete tratamentos e quatro repetições. Os resultados mostraram que B. thuringiensis var. aizawai nas duas dosagens avaliadas e B. thuringiensis var. kurstaki x B. thuringiensis var. aizawai (500 ml/ha foram os produtos mais eficientes. Ao final do experimento larvas e pupas de traça-das-crucíferas foram coletadas no campo e a primeira geração foi submetida a um teste de laboratório onde discos de folhas de repolho foram tratados com as dosagens dos inseticidas a base de B. thuringiensis utilizadas no campo. Larvas de segundo estádio foram colocadas sobre os discos tratados e a mortalidade de larvas avaliada após 72 h. Todos os tratamentos causaram mais de 97% de mortalidade de larvas. O resultado do teste de laboratório sugere que a menor eficiência de B. thuringiensis var. kurstaki x B. thuringiensis var. aizawai (350 ml/ha e B. thuringiensis var. kurstaki (500 ml/ha no teste de campo quando comparada aos demais Bacillus, pode ser devido à mais rápida degradação destes produtos no ambiente.The Diamondback Moth (Plutella xylostella is the most important cabbage pest in the Federal District. New insecticides are frequently tested for its control and in this study two new Bacillus thuringiensis formulations [B. thuringiensis var. aizawai (350 and 500 g/ha and B

  15. Persistence of Toxic Activity of Fermentation Extracts from Bacillus thuringiensis var. israelensis after More Than Three Decades of Storage

    Directory of Open Access Journals (Sweden)

    Luis Jesús Galán-Wong

    2017-01-01

    Full Text Available This study was carried out to determine the persistence of toxicity of fermentation extracts of Bacillus thuringiensis var. israelensis after more than three decades of storage. For this purpose, a population of Aedes aegypti was established. The mortality rate of 20 spore-crystal extracts purified using the acetone-lactose coprecipitation method was measured and evaluated by bioassays according to a modified WHO protocol. The extracts with the highest mortality rate were determined in triplicate by their LD50 and LD98. All extracts showed toxicity at the highest tested dose (1000 ppm and some, such as strains 3260 and 3501, still killed larvae at doses as low as 0.01 ppm. These data are surprising because no study on the activity of B. thuringiensis toxic proteins after such a long storage time has been reported.

  16. Identification of new isolates of Bacillus thuringiensis using rep-PCR products and delta-endotoxin electron microscopy

    Directory of Open Access Journals (Sweden)

    A.S.G. Lima

    2002-01-01

    Full Text Available PCR has been used to analyze the distribution of REP (Repetitive Extragenic Palindromic and ERIC (Enterobacterial Repetitive Intergenic Consensus sequences (rep-PCR found within the genome of the bacterium Bacillus thuringiensis, with the purpose to analyze the genetic similarities among 56 subspecies samples and 95 field isolates. The PCR products were analyzed by EB-AGE (ethidium bromide-agarose electrophoresis and then submitted to banding comparisons, based on the Phyllip software algorithm. When the banding similarities were considered for comparison purposes among all the strains, the phylogenic tree patterns varied according to the rep-PCR primers considered, but, from a broader point of view, the ERIC sequences produced better results, which, together with electron microscopy analysis of the released parasporal bodies and colony morphology characteristics, allowed to detect two possible new subspecies of B. thuringiensis.

  17. Correspondence of High Levels of Beta-Exotoxin I and the Presence of cry1B in Bacillus thuringiensis

    Science.gov (United States)

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

    2002-01-01

    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 μg of β-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 μg/ml. However, these amounts of β-exotoxin I were below the threshold for detectable toxicity against this insect species. Secretion of large amounts of β-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 β-exotoxin I. PMID:12200263

  18. Larvicidal Activities of Indigenous Bacillus thuringiensis Isolates and Nematode Symbiotic Bacterial Toxins against the Mosquito Vector, Culex pipiens (Diptera: Culicidae

    Directory of Open Access Journals (Sweden)

    Ashraf M Ahmed

    2017-06-01

    Full Text Available Background: The incidence of mosquito-borne diseases and the resistance of mosquitoes to conventional pesticides have recently caused a panic to the authorities in the endemic countries. This study was conducted to identify native larvicidal biopesticides against Culex pipiens for utilization in the battle against mosquito-borne diseases.Methods: Larvicidal activities of new indigenous Bacillus thuringiensis isolates and crude toxin complexes (TCs of two nematode bacterial-symbionts, Photorhabdus luminescens akhurstii (HRM1 and Ph. luminescens akhurstii (HS1 that tested against Cx. pipiens. B. thuringiensis isolates were recovered from different environmental samples in Saudi Arabia, and the entomopathogenic nematodes, Heterorhabditis indica (HRM1 and He. sp (HS1 were iso­lated from Egypt. Larvicidal activities (LC50 and LC95 of the potentially active B. thuringiensis strains or TCs were then evaluated at 24 and 48h post-treatment.Results: Three B. thuringiensis isolates were almost as active as the reference B. thuringiensis israelensis (Bti-H14, and seven isolates were 1.6–5.4 times more toxic than Bti-H14. On the other hand, the TCs of the bacterial sym­bionts, HRM1 and HS1, showed promising larvicidal activities. HS1 showed LC50 of 2.54 folds that of HRM1 at 24h post-treatment. Moreover, histopathological examinations of the HS1-treated larvae showed deformations in midgut epithelial cells at 24h post-treatment.Conclusion: Synergistic activity and molecular characterization of these potentially active biocontrol agents are currently being investigated. These results may lead to the identification of eco-friend mosquito larvicidal product(s that could contribute to the battle against mosquito-borne diseases.

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

    Directory of Open Access Journals (Sweden)

    N. Knaak

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

  20. The Synergist Effect of The Henna Plant, Lawsonia alba on Bacillus thuringiensis var. kurstaki Against Third Larval Instar of Pistachio Leaf Borer, Ocneria terebinthina Strg. (Lep.: Lymanteriidae

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    Zahra Sheibani

    2017-12-01

    Full Text Available Ocneria terebinthina Strg. (Lep.: Lymanteriidae is a leaf borer pest in pistachio orchard. The toxicity of Bacillus thuringiensis var. kurstaki alone and in combination with henna powder was investigated on third larval instar of O. terebinthina under laboratory conditions. Bioassay was carried out using spray technique on pistachio offshoot. Probit analysis of concentration-mortality data was conducted to estimate the LC50 value. The LC50 value of B. thuringiensis on third instar of larvae was estimated as 2817.30 ppm. The synergist effect of henna powder on efficacy of B. thuringiensis was also evaluated. Three concentrations of henna powder were combined with minimum lethal dose of B. thuringiensis against third larval instar. The results revealed that the henna powder had synergist effect on B. thuringiensis. The combination of B. thuringiensis and henna powder (6000 ppm caused 80.68% mortality, whereas B. thuringiensis alone caused 15.91% mortality. Our results suggested that henna powder can increase the efficacy of B. thuringiensis in controlling O. terebinthina in pest integrated management.

  1. Uso de productos derivados de Bacillus thuringiensis como alternativa de control en nematodos de importancia veterinaria. Revisión

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    Alejandro Vázquez-Pineda

    2012-01-01

    Full Text Available La bacteria entomopatógena Bacillus thuringiensis produce cristales proteicos con actividad citotóxica en contra de insectos y nematodos. La toxicidad de B. thuringiensis en plagas agrícolas es ampliamente conocida, pero poco se conoce acerca de su actividad en contra de nematodos parásitos. Recientemente, la actividad nematicida de las proteínas derivadas de B. thuringiensis se demostró en parásitos de mamíferos como Haemonchus, Teladorsagia, Nippostrongylus, y Ancylostoma, y en nematodos de plantas, Globodera y Meloidogyne. Entre el grupo de B. thuringiensis con efecto nematicida, las proteínas de la cepa IB-16 han mostrado actividad letal de 50 a 100 % en contra de diferentes estadios del principal género de rumiantes, Haemonchus contortus. Asimismo, los géneros de nematodos de vida libre, Panagrellus redivivus y Caenorhabditis elegans han sido blanco de estudios de la acción nematicida de B. thuringiensis. Por ejemplo, el efecto tóxico de la proteína Cry5B de B. thuringiensis se observó en las células intestinales de C. elegans, además esta acción parece involucrar receptores celulares específicos, similares a los que se han notificado en contra de plagas agrícolas. Asimismo, la unión de la proteína Cry5B ocurre en receptores específicos, como moléculas de carbohidratos, las cuales están presentes en la membrana de las células de intestino de los nematodos, ocasionando daño y muerte. A través de este tipo de estudios, los derivados de B. thuringiensis podrían considerarse una alternativa de control en nematodos que afectan a los animales domésticos, como rumiantes, así como en contra de otros nematodos patógenos de mamíferos e incluso de plantas agrícolas.

  2. Enhancement of intrinsic antitumor activity in spore-endotoxin mixtures of Bacillus thuringiensis by exposure to ultraviolet radiation

    Energy Technology Data Exchange (ETDEWEB)

    Zamola, B; Karminski-Zamola, G; Fuks, Z; Kubovic, M [Zagreb Univ. (Yugoslavia); Wrishcer, M [Institut Rudjer Boskovic, Zagreb (Yugoslavia)

    1985-03-01

    Irradiation of spore-endotoxin mixtures from Bacillus thuringiensis cultures at 254 nm (60 ..mu..W cm/sup -2/) enhances their intrinsic antitumor potency as well as that of either component. The extent of enhancement depends on the length of exposure (optimum: 35 min) and may thus be due to photochemical changes of the endotoxin protein or/and to photoproduction of additional compounds with antitumor activity. Antitumor effects, expressed as survival rates of C57BL/6 mice inoculated with Lewis' mouse lung carcinoma and subjected to treatments 24 h later, depended on the number of doses of preparations administered (mixture, separated components).

  3. Preferential protection of domains ii and iii of bacillus thuringiensis cry1aa toxin by brush border membrane vesicles

    OpenAIRE

    Hussain, Syed-Rehan A.; Flórez, Álvaro M.; Dean, Donald H.; Alzate, Óscar

    2011-01-01

    Título español: Protección preferencial de los dominios II y III de la toxina Cry1Aa de Bacillus thuringiensis en Vesículas de Membrana de Borde de Cepillo Abstract The surface exposed Leucine 371 on loop 2 of domain II, in Cry1Aa toxin, was mutated to Lysine to generate the trypsin-sensitive mutant, L371K. Upon trypsin digestion L371K is cleaved into approximately 37 and 26 kDa fragments. These are separable on SDS-PAGE, but remain as a single molecule of 65 kDa upon purification by ...

  4. Preferential Protection of Domains II and III of Bacillus thuringiensis Cry1Aa Toxin by Brush Border Membrane Vesicles

    OpenAIRE

    Syed-Rehan A. Hussain; Álvaro M. Flórez; Donald H. Dean; Óscar Alzate

    2011-01-01

    Título español: Protección preferencial de los dominios II y III de la toxina Cry1Aa de Bacillus thuringiensis en Vesículas de Membrana de Borde de Cepillo Abstract The surface exposed Leucine 371 on loop 2 of domain II, in Cry1Aa toxin, was mutated to Lysine to generate the trypsin-sensitive mutant, L371K. Upon trypsin digestion L371K is cleaved into approximately 37 and 26 kDa fragments. These are separable on SDS-PAGE, but remain as a single molecule of 65 kDa upon purification by ...

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

    DEFF Research Database (Denmark)

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

    2008-01-01

    of growth and sporulation of vegetative cells. For the first time conjugative plasmid transfer in a mammalian intestinal tract was shown between two B. thuringiensis strains. Significance and Impact of the Study: Strains of B. thuringiensis are used worldwide to combat insect pests, and this study brings...

  6. Operational Evaluation Of Vectomax® WSP (Bacillus thuringiensis Subsp. israelensis+Bacillus sphaericus) Against Larval Culex pipiens in Septic Tanks (1).

    Science.gov (United States)

    Cetin, Huseyin; Oz, Emre; Yanikoglu, Atila; Cilek, James E

    2015-06-01

    The residual effectiveness of VectoMax® WSP (a water-soluble pouch formulation containing a combination of Bacillus thuringiensis subsp. israelensis strain AM65-52 and B. sphaericus strain ABTS 1743) when applied to septic tanks against 3rd- and 4th-stage larvae of Culex pipiens L. was evaluated in this study. This formulation was evaluated at operational application rates of 1 pouch (10 g) and 2 pouches (20 g) per septic tank. Both application rates resulted in >96% control of larvae for 24 days. Operationally, VectoMax WSP has proven to be a useful tool for the nonchemical control of Culex species in septic tank environments.

  7. Characterization of Bacillus thuringiensis soil isolates from Cuba, with insecticidal activity against mosquitoes

    Directory of Open Access Journals (Sweden)

    Aileen González

    2011-09-01

    Full Text Available 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. Rev. Biol. Trop. 59 (3: 1007-1016. Epub 2011 September 01.El uso prolongado de insecticidas ha conducido al desarrollo de resistencia en diferentes especies de mosquitos y al incremento de la degradación del ambiente. El control biológico de insectos ha devenido como una alternativa útil y de bajo impacto ambiental. En nuestro estudio fueron identificados, caracterizados tres aislamientos de suelos procedentes de diferentes regiones del archipiélago cubano y comparados con cepas de referencia: aisladas de los biolarvicidas Bactivec y Bactoculicida, además de IPS82. La diferenciación de los mismos se llevó a cabo mediante métodos morfol

  8. Selection of Bacillus thuringiensis strains toxic to cotton boll weevil (Anthonomus grandis, Coleoptera: Curculionidae) larvae.

    Science.gov (United States)

    Pérez, Melisa P; Sauka, Diego H; Onco, María I; Berretta, Marcelo F; Benintende, Graciela B

    Preliminary bioassays with whole cultures (WC) of 124 Bacillus thuringiensis strains were performed with neonate larvae of Anthonomus grandis, a major cotton pest in Argentina and other regions of the Americas. Three exotic and four native strains were selected for causing more than 50% mortality. All of them were β-exotoxin producers. The native strains shared similar morphology of parasporal crystals, similar protein pattern and identical insecticidal gene profiles. These features resembled Lepidoptera-toxic strains. Furthermore, these strains showed a Rep-PCR pattern identical to lepidoptericidal strain HD-1, suggesting that these strains may belong to serovar kurstaki. However, some differences were observed in the plasmid profiles and in the production of β-exotoxin. To determine the culture fractions where the insecticidal metabolites were present, bioassays including resuspended spore-crystal pellets, filtered supernatants (FS) were compared with those of WC. Both fractions tested showed some level of insecticidal activity. The results may suggest that the main toxic factors can be found in FS and could be directly correlated with the presence of β-exotoxin. Based on the bioassays with FS and autoclaved FS, the participation of thermolabile virulence factors such as Cry1I in toxicity is neither discarded. In the selected strains, β-exotoxin would be the major associated virulence factor; therefore, their use in biological control of A. grandis should be restricted. Nevertheless, these strains could be the source of genes (e.g., cry1Ia) to produce transgenic cotton plants resistant to this pest. Copyright © 2017 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

  9. pH-controlled Bacillus thuringiensis Cry1Ac protoxin loading and release from polyelectrolyte microcapsules.

    Directory of Open Access Journals (Sweden)

    Wenhui Yang

    Full Text Available Crystal proteins synthesized by Bacillus thuringiensis (Bt have been used as biopesticides because of their toxicity to the insect larval hosts. To protect the proteins from environmental stress to extend their activity, we have developed a new microcapsule formulation. Poly (acrylic acid (PAH and poly (styrene sulfonate (PSS were fabricated through layer-by-layer self-assembly based on a CaCO(3 core. Cry1Ac protoxins were loaded into microcapsules through layer-by-layer self-assembly at low pH, and the encapsulated product was stored in water at 4°C. Scanning electron microscopy (SEM was used to observe the morphology of the capsules. To confirm the successful encapsulation, the loading results were observed with a confocal laser scattering microscope (CLSM, using fluorescein-labeled Cry1Ac protoxin (FITC-Cry1Ac. The protoxins were released from the capsule under the alkaline condition corresponding to the midgut of certain insects, a condition which seldom exists elsewhere in the environment. The following bioassay experiment demonstrated that the microcapsules with Cry1Ac protoxins displayed approximately equivalent insecticidal activity to the Asian corn borer compared with free Cry1Ac protoxins, and empty capsules proved to have no effect on insects. Further result also indicated that the formulation could keep stable under the condition of heat and desiccation. These results suggest that this formulation provides a promising methodology that protects protoxins from the environment and releases them specifically in the target insects' midgut, which has shown potential as biopesticide in the field.

  10. An ABC transporter mutation is correlated with insect resistance to Bacillus thuringiensis Cry1Ac toxin.

    Directory of Open Access Journals (Sweden)

    Linda J Gahan

    2010-12-01

    Full Text Available Transgenic crops producing insecticidal toxins from Bacillus thuringiensis (Bt are commercially successful in reducing pest damage, yet knowledge of resistance mechanisms that threaten their sustainability is incomplete. Insect resistance to the pore-forming Cry1Ac toxin is correlated with the loss of high-affinity, irreversible binding to the mid-gut membrane, but the genetic factors responsible for this change have been elusive. Mutations in a 12-cadherin-domain protein confer some Cry1Ac resistance but do not block this toxin binding in in vitro assays. We sought to identify mutations in other genes that might be responsible for the loss of binding. We employed a map-based cloning approach using a series of backcrosses with 1,060 progeny to identify a resistance gene in the cotton pest Heliothis virescens that segregated independently from the cadherin mutation. We found an inactivating mutation of the ABC transporter ABCC2 that is genetically linked to Cry1Ac resistance and is correlated with loss of Cry1Ac binding to membrane vesicles. ABC proteins are integral membrane proteins with many functions, including export of toxic molecules from the cell, but have not been implicated in the mode of action of Bt toxins before. The reduction in toxin binding due to the inactivating mutation suggests that ABCC2 is involved in membrane integration of the toxin pore. Our findings suggest that ABC proteins may play a key role in the mode of action of Bt toxins and that ABC protein mutations can confer high levels of resistance that could threaten the continued utilization of Bt-expressing crops. However, such mutations may impose a physiological cost on resistant insects, by reducing export of other toxins such as plant secondary compounds from the cell. This weakness could be exploited to manage this mechanism of Bt resistance in the field.

  11. An ABC transporter mutation is correlated with insect resistance to Bacillus thuringiensis Cry1Ac toxin.

    Science.gov (United States)

    Gahan, Linda J; Pauchet, Yannick; Vogel, Heiko; Heckel, David G

    2010-12-16

    Transgenic crops producing insecticidal toxins from Bacillus thuringiensis (Bt) are commercially successful in reducing pest damage, yet knowledge of resistance mechanisms that threaten their sustainability is incomplete. Insect resistance to the pore-forming Cry1Ac toxin is correlated with the loss of high-affinity, irreversible binding to the mid-gut membrane, but the genetic factors responsible for this change have been elusive. Mutations in a 12-cadherin-domain protein confer some Cry1Ac resistance but do not block this toxin binding in in vitro assays. We sought to identify mutations in other genes that might be responsible for the loss of binding. We employed a map-based cloning approach using a series of backcrosses with 1,060 progeny to identify a resistance gene in the cotton pest Heliothis virescens that segregated independently from the cadherin mutation. We found an inactivating mutation of the ABC transporter ABCC2 that is genetically linked to Cry1Ac resistance and is correlated with loss of Cry1Ac binding to membrane vesicles. ABC proteins are integral membrane proteins with many functions, including export of toxic molecules from the cell, but have not been implicated in the mode of action of Bt toxins before. The reduction in toxin binding due to the inactivating mutation suggests that ABCC2 is involved in membrane integration of the toxin pore. Our findings suggest that ABC proteins may play a key role in the mode of action of Bt toxins and that ABC protein mutations can confer high levels of resistance that could threaten the continued utilization of Bt-expressing crops. However, such mutations may impose a physiological cost on resistant insects, by reducing export of other toxins such as plant secondary compounds from the cell. This weakness could be exploited to manage this mechanism of Bt resistance in the field.

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

    Science.gov (United States)

    Gao, Yulin; Hu, Yang; Fu, Qiang; Zhang, Jie; Oppert, Brenda; Lai, Fengxiang; Peng, Yufa; Zhang, Zhitao

    2010-09-01

    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 borer) and Chilo suppressalis (striped stem borer), was evaluated in the laboratory by feeding neonate larvae on artificial diets containing Bt protoxins. The results indicated that Cry1Ca exhibited the highest level of toxicity to both stem borers, with an LC(50) of 0.24 and 0.30 microg/g for C. suppressalis and S. inferens, respectively. However, S. inferens was 4-fold lower in susceptibility to Cry1Aa, and 6- and 47-fold less susceptible to Cry1Ab and Cry1Ba, respectively, compared to C. suppressalis. To evaluate interactions among Bt protoxins in stem borer larvae, toxicity assays were performed with mixtures of Cry1Aa/Cry1Ab, Cry1Aa/Cry1Ca, Cry1Ac/Cry1Ca, Cry1Ac/Cry1Ba, Cry1Ab/Cry1Ac, Cry1Ab/Cry1Ba, and Cry1Ab/Cry1Ca at 1:1 (w/w) ratios. All protoxin mixtures demonstrated significant synergistic toxicity activity against C. suppressalis, with values of 1.6- to 11-fold higher toxicity than the theoretical additive effect. Surprisingly, all but one of the Bt protoxin mixtures were antagonistic in toxicity to S. inferens. In mortality-time response experiments, S. inferens demonstrated increased tolerance to Cry1Ab and Cry1Ac compared to C. suppressalis when treated with low or high protoxin concentrations. The data indicate the utility of Cry1Ca protoxin and a Cry1Ac/Cry1Ca mixture to control both stem borer populations. Copyright 2010 Elsevier Inc. All rights reserved.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2008-07-01

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

  14. Lipopeptide biosurfactant from Bacillus thuringiensis pak2310: A potential antagonist against Fusarium oxysporum.

    Science.gov (United States)

    Deepak, R; Jayapradha, R

    2015-03-01

    The aims of the study were to evaluate the effects of a biosurfactant obtained from a novel Bacillus thuringiensis on Fusarium oxysporum to determine the morphological changes in the structure of the fungi and its biofilm in the presence of the biosurfactant and to evaluate the toxicity of the biosurfactant on HEp-2 human epithelial cell lines. The strain was screened and isolated from petroleum contaminated soil based on the E24 emulsification index. The biosurfactant was produced on glycerol, extracted using chloroform:methanol system and purified using HPLC. The purified fraction showing both surface activity (emulsification and oil-spread activity) and anti-fusarial activity (agar well diffusion method) was studied using FT-IR and MALDI-TOF MS, respectively. The minimum inhibitory concentration (MIC) and the biofilm inhibitory concentration (BIC) were determined using dilution method. The effect of biosurfactant on the morphology of Fusarium oxysporum was monitored using light microscopy and confocal laser scanning microscopy (for biofilm). The purified surfactant showed the presence of functional groups like that of surfactin in the FT-IR spectra and MALDI-TOF MS estimated the molecular weight as 700Da. The MIC and BIC were estimated to be 0.05 and 0.5mg/mL, respectively. The molecule was also non-toxic to HEp-2 cell lines at 10× MIC. A non-toxic and effective anti-Fusarium biosurfactant, that is both safe for human use and to the environment, has been characterized. The growth and metabolite production using glycerol (major byproduct of biodiesel and soap industries) also adds up to the efficiency and ecofriendly nature of this biosurfactant. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

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

    Directory of Open Access Journals (Sweden)

    Guangsheng Li

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

  16. Pathogenicity of Bacillus thuringiensis isolated from two species of Acromyrmex (Hymenoptera, Formicidae

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    L. M. N. Pinto

    Full Text Available The control of Acromyrmex leaf-cutting ants is necessary due to the severe damage they cause to diverse crops. A possibility was to control them using the bacterium Bacillus thuringiensis (Bt that characteristically produces insecticidal crystal proteins (ICPs. The ICPs have been effective in controlling lepidopterans, dipterans, and coleopterans, but their action against hymenopterans is unknown. This paper describes an attempt to isolate Bt from ants of two Acromyrmex species, to evaluate its pathogenicity towards these ants, and to test isolates by PCR. Bacterial isolates of Bt obtained from A. crassispinus and A. lundi have been assayed against A. lundi in the laboratory. The bioassays were carried out in BOD at 25°C, with a 12-hour photoperiod, until the seventh day after treatment. The Bt isolates obtained were submitted to total DNA extraction and tested by PCR with primers specific to cry genes. The results showed Bt presence in 40% of the assessed samples. The data from the in vivo assays showed a mortality rate higher than 50% in the target population, with the Bt HA48 isolate causing 100% of corrected mortality. The PCR results of Bt isolates showed a magnification of DNA fragments relative to cry1 genes in 22% of the isolates, and cry9 in 67%. Cry2, cry3, cry7, and cry8 genes were not detected in the tested samples, and 22% had no magnified DNA fragments corresponding to the assessed cry genes. The results are promising not only regarding allele identification in new isolates, but also fort the assays aimed at determining the Bt HA48 LC50's, which can eventually be applied in controlling of Acromyrmex leaf-cutting ants.

  17. Purification and Characterization of a Novel Cold Shock Protein-Like Bacteriocin Synthesized by Bacillus thuringiensis.

    Science.gov (United States)

    Huang, Tianpei; Zhang, Xiaojuan; Pan, Jieru; Su, Xiaoyu; Jin, Xin; Guan, Xiong

    2016-10-20

    Bacillus thuringiensis (Bt), one of the most successful biopesticides, may expand its potential by producing bacteriocins (thuricins). The aim of this study was to investigate the antimicrobial potential of a novel Bt bacteriocin, thuricin BtCspB, produced by Bt BRC-ZYR2. The results showed that this bacteriocin has a high similarity with cold-shock protein B (CspB). BtCspB lost its activity after proteinase K treatment; however it was active at 60 °C for 30 min and was stable in the pH range 5-7. The partial loss of activity after the treatments of lipase II and catalase were likely due to the change in BtCspB structure and the partial degradation of BtCspB, respectively. The loss of activity at high temperatures and the activity variation at different pHs were not due to degradation or large conformational change. BtCspB did not inhibit four probiotics. It was only active against B. cereus strains 0938 and ATCC 10987 with MIC values of 3.125 μg/mL and 0.781 μg/mL, and MBC values of 12.5 μg/mL and 6.25 μg/mL, respectively. Taken together, these results provide new insights into a novel cold shock protein-like bacteriocin, BtCspB, which displayed promise for its use in food preservation and treatment of B. cereus-associated diseases.

  18. Density Dependence and Growth Rate: Evolutionary Effects on Resistance Development to Bt (Bacillus thuringiensis).

    Science.gov (United States)

    Martinez, Jeannette C; Caprio, Michael A; Friedenberg, Nicholas A

    2018-02-09

    It has long been recognized that pest population dynamics can affect the durability of a pesticide, but dose remains the primary component of insect resistance management (IRM). For transgenic pesticidal traits such as Bt (Bacillus thuringiensis Berliner (Bacillales: Bacillaceae)), dose (measured as the mortality of susceptibles caused by a toxin) is a relatively fixed characteristic and often falls below the standard definition of high dose. Hence, it is important to understand how pest population dynamics modify durability and what targets they present for IRM. We used a deterministic model of a generic arthropod pest to examine how timing and strength of density dependence interacted with population growth rate and Bt mortality to affect time to resistance. As in previous studies, durability typically reached a minimum at intermediate doses. However, high population growth rates could eliminate benefits of high dose. The timing of density dependence had a more subtle effect. If density dependence operated simultaneously with Bt mortality, durability was insensitive to its strengths. However, if density dependence was driven by postselection densities, decreasing its strength could increase durability. The strength of density dependence could affect durability of both single traits and pyramids, but its influence depended on the timing of density dependence and size of the refuge. Our findings suggest the utility of a broader definition of high dose, one that incorporates population-dynamic context. That maximum growth rates and timing and strength of interactions causing density dependent mortality can all affect durability, also highlights the need for ecologically integrated approaches to IRM research. © The Author(s) 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  19. Persistence of detectable insecticidal proteins from Bacillus thuringiensis (Cry) and toxicity after adsorption on contrasting soils

    International Nuclear Information System (INIS)

    Hung, T.P.; Truong, L.V.; Binh, N.D.; Frutos, R.; Quiquampoix, H.; Staunton, S.

    2016-01-01

    Insecticidal Cry, or Bt, proteins are produced by the soil-endemic bacterium, Bacillus thuringiensis and some genetically modified crops. Their environmental fate depends on interactions with soil. Little is known about the toxicity of adsorbed proteins and the change in toxicity over time. We incubated Cry1Ac and Cry2A in contrasting soils subjected to different treatments to inhibit microbial activity. The toxin was chemically extracted and immunoassayed. Manduca sexta was the target insect for biotests. Extractable toxin decreased during incubation for up to four weeks. Toxicity of Cry1Ac was maintained in the adsorbed state, but lost after 2 weeks incubation at 25 °C. The decline in extractable protein and toxicity were much slower at 4 °C with no significant effect of soil sterilization. The major driving force for decline may be time-dependent fixation of adsorbed protein, leading to a decrease in the extraction yield in vitro, paralleled by decreasing solubilisation in the larval gut. - Graphical abstract: Biotest, presenting Cry-contaminated feed to Manduca sexta larvae in individual Perspex boxes. Display Omitted - Highlights: • Toxicity of Cry protein is initially conserved after adsorption on soil. • Toxicity and extractability decline with time, more rapidly at 25 °C than 4 °C. • Similar dynamics of Cry1AC and Cry2A on soil with varying texture and organic C. • Sterilization of soil does not change Cry dynamics or temperature effect in soil. • Cry decline is determined by progressive fixation on soil not microbial breakdown. - Toxicity was initially maintained after adsorption on soil and both extractable Cry and toxicity declined rapidly, more slowly at low temperature, due to different fixation dynamics. Toxicity of Cry protein is initially conserved after adsorption on soil.

  20. MORTALITY OF Spodoptera eridania (Cramer CATERPILLARS BY Bacillus thuringiensis (Berliner MORTALIDADE DE LAGARTAS DE Spodoptera eridania (Cramer PELA UTILIZAÇÃO DE Bacillus thuringiensis (Berliner

    Directory of Open Access Journals (Sweden)

    Carlos Brustolin

    2009-03-01

    Full Text Available

    This research evaluated the effects of two products based on Bacillus thuringiensis in the mortality rate of first and third instar caterpillars of Spodoptera eridania, in laboratory conditions, at 25±2°C, relative humidity 70±5%, and photoperiod of 12 hours. The treatments were B. thuringiensis kurstaki (Dipel SC, at 500 mL.ha-1

  1. Cloning and Characterization of a Unique Cytotoxic Protein Parasporin-5 Produced by Bacillus thuringiensis A1100 Strain

    Directory of Open Access Journals (Sweden)

    Keisuke Ekino

    2014-06-01

    Full Text Available 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.

  2. Poly-β-hydroxybutyrate Metabolism Is Unrelated to the Sporulation and Parasporal Crystal Protein Formation in Bacillus thuringiensis.

    Science.gov (United States)

    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.

  3. The mechanism of uranium transformation from U(VI) into nano-uramphite by two indigenous Bacillus thuringiensis strains

    International Nuclear Information System (INIS)

    Pan, Xiaohong; Chen, Zhi; Chen, Fanbing; Cheng, Yangjian; Lin, Zhang; Guan, Xiong

    2015-01-01

    Highlights: • Indigenous B. thuringiensis exhibited highly accumulation ability to U(VI) in the absence of additional nutrients. • The amorphous uranium compound would transformed into crystalline nano-uramphite by B. thuringiensis. • The chemical nature of formed U-species were monitored. • The cell-free extracts of B. thuringiensis had better uranium-immobilization ability than its cell debris. • Provided the understanding of the uranium transformation mechanism. - Abstract: The mechanism of uranium transformation from U(VI) into nano-uramphite by two indigenous Bacillus thuringiensis strains was investigated in the present work. Our data showed that the bacteria isolated from uranium mine possessed highly accumulation ability to U(VI), and the maximum accumulation capacity was around 400 mg U/g biomass (dry weight). X-ray powder diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FT-IR) analyzes indicated that the U(VI) was adsorbed on the bacterial surface firstly through coordinating with phosphate, −CH 2 and amide groups, and then needle-like amorphous uranium compounds were formed. With the extension of time, the extracellular crystalline substances were disappeared, but some particles were appeared in the intracellular region, and these particles were characterized as tetragonal-uramphite. Moreover, the disrupted experiment indicated that the cell-free extracts had better uranium-immobilization ability than cell debris. Our findings provided the understanding of the uranium transformation process from amorphous uranium to crystalline uramphite, which would be useful in the regulation of uranium immobilization process

  4. The mechanism of uranium transformation from U(VI) into nano-uramphite by two indigenous Bacillus thuringiensis strains

    Energy Technology Data Exchange (ETDEWEB)

    Pan, Xiaohong; Chen, Zhi [Key Lab of Biopesticide and Chemical Biology, Fujian Agriculture and Forestry University, Ministry of Education & Fujian–Taiwan Joint Center for Ecological Control of Crop Pests, Fuzhou, Fujian 350002 (China); Key Laboratory of Design and Assembly of Functional Nanostructures, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou, Fujian 350002 (China); Chen, Fanbing [Key Lab of Biopesticide and Chemical Biology, Fujian Agriculture and Forestry University, Ministry of Education & Fujian–Taiwan Joint Center for Ecological Control of Crop Pests, Fuzhou, Fujian 350002 (China); Cheng, Yangjian [Key Laboratory of Design and Assembly of Functional Nanostructures, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou, Fujian 350002 (China); Lin, Zhang, E-mail: zlin@fjirsm.ac.cn [Key Laboratory of Design and Assembly of Functional Nanostructures, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou, Fujian 350002 (China); School of Environment and Energy, South China University of Technology, Guangzhou 510006 (China); Guan, Xiong, E-mail: guanxfafu@126.com [Key Lab of Biopesticide and Chemical Biology, Fujian Agriculture and Forestry University, Ministry of Education & Fujian–Taiwan Joint Center for Ecological Control of Crop Pests, Fuzhou, Fujian 350002 (China)

    2015-10-30

    Highlights: • Indigenous B. thuringiensis exhibited highly accumulation ability to U(VI) in the absence of additional nutrients. • The amorphous uranium compound would transformed into crystalline nano-uramphite by B. thuringiensis. • The chemical nature of formed U-species were monitored. • The cell-free extracts of B. thuringiensis had better uranium-immobilization ability than its cell debris. • Provided the understanding of the uranium transformation mechanism. - Abstract: The mechanism of uranium transformation from U(VI) into nano-uramphite by two indigenous Bacillus thuringiensis strains was investigated in the present work. Our data showed that the bacteria isolated from uranium mine possessed highly accumulation ability to U(VI), and the maximum accumulation capacity was around 400 mg U/g biomass (dry weight). X-ray powder diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM) and Fourier transform infrared spectroscopy (FT-IR) analyzes indicated that the U(VI) was adsorbed on the bacterial surface firstly through coordinating with phosphate, −CH{sub 2} and amide groups, and then needle-like amorphous uranium compounds were formed. With the extension of time, the extracellular crystalline substances were disappeared, but some particles were appeared in the intracellular region, and these particles were characterized as tetragonal-uramphite. Moreover, the disrupted experiment indicated that the cell-free extracts had better uranium-immobilization ability than cell debris. Our findings provided the understanding of the uranium transformation process from amorphous uranium to crystalline uramphite, which would be useful in the regulation of uranium immobilization process.

  5. CARACTERIZACIÓN MOLECULAR MEDIANTE rep-PCR DE AISLADOS NATIVOS DE Bacillus thuringiensis, OBTENIDOS DE MUESTRAS DE SUELO

    Directory of Open Access Journals (Sweden)

    Fabián Galvis

    2014-01-01

    Full Text Available Bacillus thuringiensis es una bacteria Gram-positiva formadora de esporas, que produ - ce cristales parasporales de naturaleza proteica, tóxicos contra diferentes órdenes de insectos y biodegradables e inocuos para otras especies. Esta investigación empleó el modelo experimen - tal, que mediante técnicas de observación permi - tió, la identificación microbiológica y bioquímica de B. thuringiensis a partir de muestras de suelo de los municipios de Cúcuta, El Zulia, Los Patios, San Cayetano y Villa del Rosario, Norte de Santander, Colombia, y su posterior caracteri - zación con los marcadores moleculares Bc-Rep y MB1. Se identificaron microbiológica y bioquí - micamente 10 aislados como B. thuringiensis ; los resultados del análisis filogenético mostraron diferencias significativas en los agrupamientos obtenidos con los marcadores Bc-Rep y MB1. Con Bc-Rep se registró un índice de similaridad bajo (18%, mientras que con el marcador MB1 se obtuvo un índice mayor de similitud, 58%. En este trabajo se evidenció una gran variabilidad genética entre los aislados, que mostraron a los marcadores Bc-Rep y MB1 como altamente efectivos para diferenciar cepas estrechamente relacionadas, convirtiéndose en una herramienta genética de gran valor para estudios de identifi-cación y diversidad en B. thuringiensis.

  6. Exploration of Bacillus thuringiensis Berl. from soil and screening test its toxicity on insects of Lepidoptera order

    Science.gov (United States)

    Astuti, DT; Pujiastuti, Y.; Suparman, SHK; Damiri, N.; Nugraha, S.; Sembiring, ER; Mulawarman

    2018-01-01

    Bacillus thuringiensis is a gram-positive bacterium that produces crystal proteins toxic (ᴕ-endotoxin) specific to the target insect, but is not toxic to humans and non-target organisms. This study aims to explore the origin of the soil bacterium B. thuringiensis sub-district Sekayu, Banyuasin, South Sumatra and toxicity to larvae of lepidoptera. Fifty soil samples were taken from Musi Banyuasin District, namely 15 from Kayuare strip 2, 20 from Kayuare and 15 from Lumpatan. Isolation, characterization, identification and screening test were conducted in the laboratorium of Pest and Disease, Agricultural Faculty, Sriwijaya University. Isolat codes were given based on the area origin of the samples. Results of the study showed that from 50 isolates of bacteria that had been isolated, there were 15 bacterial isolates, characterized by morphology and physiology the same as B. thuringiensis, which has round colonies, white, wrinkled edges, slippery, elevation arise, aerobic and gram-positive. Of the 15 codes that contain positive isolates of B. thuringiensis, we have obtained several isolates of the following codes: KJ2D5, KJ2N1, KJ2N4, KJ2B3, KJ3R1, KJ3R2, KJ3R3, KJ3R5, KJ3J3, KJ3J4, KJ3P1, DLM5, DLKK12, and DLKK23. Results of screening tests on insects of the Lepidoptera Order showed that there were six isolates that had toxic to Plutella xylostella and Spodoptera litura insects, ie bacterial isolate codes DLM5, KJ3R3, KJ3R5, KJ3J4, KJ3P1, and DLKK23.

  7. Quorum Sensing in Bacillus thuringiensis Is Required for Completion of a Full Infectious Cycle in the Insect

    Directory of Open Access Journals (Sweden)

    Leyla Slamti

    2014-07-01

    Full Text Available 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.

  8. First report of detection of the putative receptor of Bacillus thuringiensis toxin Vip3Aa from black cutworm (Agrotis ipsilon

    Directory of Open Access Journals (Sweden)

    Gamal H. Osman

    2018-03-01

    Full Text Available Black cutworm (BCW Agrotis ipsilon, an economically important lepidopteran insect, has attracted a great attention. Bacillus thuringiensis (Bt is spore forming soil bacteria and is an excellent environment-friendly approach for the control of phytophagous and disease-transmitting insects. In fact, bio-pesticide formulations and insect resistant transgenic plants based on the bacterium Bt delta-endotoxin have attracted worldwide attention as a safer alternative to harmful chemical pesticides. The major objective of the current study was to understand the mechanism of interaction of Bt toxin with its receptor molecule(s. The investigation involved the isolation, identification, and characterization of a putative receptor – vip3Aa. In addition, the kinetics of vip toxin binding to its receptor molecule was also studied. The present data suggest that Vip3Aa toxin bound specifically with high affinity to a 48-kDa protein present at the brush border membrane vesicles (BBMV prepared from the midgut epithelial cells of BCW larvae. Keywords: Receptor, vip3Aa, Bacillus thuringiensis, BBMV

  9. Comparative analysis of Bacillus thuringiensis toxin binding to gypsy moth, browntail moth, and douglas-fir tussock moth midgut tissue sections using fluorescence microscopy

    Science.gov (United States)

    Algimantas P. Valaitis; John D. Podgwaite

    2011-01-01

    Many strains of Bacillus thuringiensis (Bt) produce insecticidal proteins, also referred to as Cry toxins, in crystal inclusions during sporulation. When ingested by insects, the Cry toxins bind to receptors on the brush border midgut epithelial cells and create pores in the epithelial gut membranes resulting in the death of...

  10. A differentially displayed mRNA related to restistance to Bacillus thuringiensis israelensis of Aedes albopictus selected in vitro-activated CYT1AA6

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Zhang, Q.; Huang, E.; Li, M.; Huang, T.; Xu, L.; Wu, Ch.; Guan, X.; Gelbič, Ivan

    2012-01-01

    Roč. 28, č. 4 (2012), s. 327-329 ISSN 8756-971X Institutional support: RVO:60077344 Keywords : Bacillus thuringiensis israelensis * resistence * Aedes albopictus Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.755, year: 2012 http://www.bioone.org/doi/pdf/10.2987/12-6263R.1

  11. Increased toxicity of Bacillus thuringiensis Cry3Aa against Crioceris quatuordecimpunctata, Phaedon brassicae and Colaphellus bowringi by a Tenebrio molitor cadherin fragment

    Science.gov (United States)

    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...

  12. Isolation and partial characterization of gypsy moth BTR-270, an anionic brush border membrane glycoconjugate that binds Bacillus thuringiensis Cry1A toxins with high affinity

    Science.gov (United States)

    Algimantas P. Valaitis; Jeremy L. Jenkins; Mi Kyong Lee; Donald H. Dean; Karen J. Garner

    2001-01-01

    BTR-270, a gypsy moth (Lymantria dispar) brush border membrane molecule that binds Bacillus thuringiensis (Bt) Cry1A toxins with high affinity, was purified by preparative gel electrophoresis. Rabbit antibodies specific for the Bt toxin-binding molecule were raised. Attempts to label BTR-270 by protein-directed techniques were...

  13. Draft Genome Sequence of Bacillus thuringiensis Strain BrMgv02-JM63, a Chitinolytic Bacterium Isolated from Oil-Contaminated Mangrove Soil in Brazil.

    Science.gov (United States)

    Marcon, Joelma; Taketani, Rodrigo Gouvêa; Dini-Andreote, Francisco; Mazzero, Giulia Inocêncio; Soares, Fabio Lino; Melo, Itamar Soares; Azevedo, João Lúcio; Andreote, Fernando Dini

    2014-01-30

    Here, we report the draft genome sequence and the automatic annotation of Bacillus thuringiensis strain BrMgv02-JM63. This genome comprises a set of genes involved in the metabolism of chitin and N-acetylglucosamine utilization, thus suggesting the possible role of this strain in the cycling of organic matter in mangrove soils.

  14. Draft genome sequence of Bacillus thuringiensis strain BrMgv02-JM63, a chitinolytic bacterium isolated from oil-contaminated mangrove soil in Brazil

    NARCIS (Netherlands)

    Marcon, Joelma; Taketani, Rodrigo Gouvêa; Dini-Andreote, Francisco; Mazzero, Giulia Inocêncio; Soares Junior, Fabio Lino; Melo, Itamar Soares; Azevedo, João Lúcio; Andreote, Fernando Dini

    2014-01-01

    Here, we report the draft genome sequence and the automatic annotation of Bacillus thuringiensis strain BrMgv02-JM63. This genome comprises a set of genes involved in the metabolism of chitin and N-acetylglucosamine utilization, thus suggesting the possible role of this strain in the cycling of

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

    NARCIS (Netherlands)

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

    2011-01-01

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

  16. Combined effect of seaweed (Sargassum wightii) and Bacillus thuringiensis var. israelensis on the coastal mosquito,Anopheles sundaicus, in Tamil Nadu, India

    Science.gov (United States)

    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...

  17. A synthetic cryIC gene, encoding a Bacillus thuringiensis δ-endotoxin, confers Spodoptera resistance in alfalfa and tobacco

    NARCIS (Netherlands)

    Strizhov, N.; Keller, M.; Mathur, J.; Koncz-Kaiman, Z.; Bosch, D.; Prudovksy, E.; Schell, J.; Sneh, B.; Koncz, C.; Zilberstein, A.

    1996-01-01

    Spodoptera species, representing widespread polyphagous insect pests, are resistant to Bacillus thuringiensis δ-endotoxins used thus far as insecticides in transgenic plants. Here we describe the chemical synthesis of a cryIC gene by a novel template directed ligation–PCR method. This simple and

  18. Field evaluation of the synergistic effects of neem oil with Beauveria bassiana (Hypocreales: Clavicipitaceae) and Bacillus thuringiensis var. kurstaki (Bacillales: Bacillaceae)

    NARCIS (Netherlands)

    Togbe, C.E.; Zannou, E.; Gbehounou, G.; Kossou, D.; Huis, van A.

    2014-01-01

    In the present study, the synergistic effects of Beauveria bassiana (Bals.-Criv. Vuill.) (isolate Bb11) and Bacillus thuringiensis var. kurstaki (Berliner) with neem oil were evaluated in three agroecological zones in Be´nin. Four bioinsecticide treatments (neem oil, neem oil and B. bassiana used

  19. Bacillus thuringiensis delta-endotoxin Cry1Ac domain III enhances activity against Heliothis virescens in some, but not all Cry1-Cry1Ac hybrids

    NARCIS (Netherlands)

    Karlova, R.B.; Weemen, W.M.J.; Naimov, S.; Ceron, J.; Dukiandjiev, S.; Maagd, de R.A.

    2005-01-01

    We investigated the role of domain III of Bacillus thuringiensis d-endotoxin Cry1Ac in determining toxicity against Heliothis virescens. Hybrid toxins, containing domain III of Cry1Ac with domains I and II of Cry1Ba, Cry1Ca, Cry1Da, Cry1Ea, and Cry1Fb, respectively, were created. In this way Cry1Ca,

  20. Changes in protease activity and Cry3Aa toxin binding in the Colorado potato beetle: implications for insect resistance to Bacillus thuringiensis toxins

    Science.gov (United States)

    Olga Loseva; Mohamed Ibrahim; Mehmet Candas; C. Noah Koller; Leah S. Bauer; Lee A. Jr. Bulla

    2002-01-01

    Widespread commercial use of Bacillus thuringiensis Cry toxins to control pest insects has increased the likelihood for development of insect resistance to this entomopathogen. In this study, we investigated protease activity profiles and toxin-binding capacities in the midgut of a strain of Colorado potato beetle (CPB) that has developed resistance...

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

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis modified Cry3A 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 PROTECTANTS...

  2. 40 CFR 174.501 - Bacillus thuringiensis Vip3Aa protein in corn and cotton; exemption from the requirement of a...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Vip3Aa protein 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 PROTECTANTS...

  3. 40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry34Ab1 and 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 PROTECTANTS...

  4. 40 CFR 174.518 - Bacillus thuringiensis Cry3Bb1 protein in corn; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry3Bb1 protein 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 and...

  5. 40 CFR 174.510 - Bacillus thuringiensis Cry1Ac protein in all plants; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1Ac protein 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 Tolerances...

  6. 40 CFR 174.532 - Bacillus thuringiensis eCry3.1Ab protein in corn; temporary exemption from the requirement of a...

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis eCry3.1Ab 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 PROTECTANTS...

  7. 40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1Ab protein 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 Tolerances...

  8. Recovery of commercially produced Bacillus thuringiensis var. israelensis and Bacillus sphaericus from tires and prevalence of bacilli in artificial and natural containers.

    Science.gov (United States)

    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.

  9. Potency of Bacillus thuringiensis isolates from bareng Tenes-Malang City as a biological control agent for suppressing third instar of Aedes aegypti larvae

    Science.gov (United States)

    Lutfiana, Nihayatul; Gama, Zulfaidah Penata

    2017-11-01

    Dengue is a mosquito-borne viral disease that is transmitted by the female Aedes species. The number of dengue fever cases has increased in many geographic regions including Indonesia and one of them occurred in Bareng Tenes, Malang City, East Java Province. The objective of this research was to identify the potency of B. thuringeinsis isolates from Bareng Tenes, Malang, as the biological agent to control third instar Ae. aegypti larvae and to identify the potential B. thuringiensis isolates based on 16S rDNA sequence. B. thuringiensis was isolated from water and soil from 12 sites in the Bareng Tenes area. Bacterial isolation was performed using B. thuringiensis selective media. Several isolates had similar phenotypic characters with B. thuringiensis used to toxicity test against third instar Ae. aegypti larvae. The LC50-96h value was determined using probit regression. The most effective isolate was identified based on the 16S rDNA sequence, then aligned to the reference isolate using the BLAST program. A phylogeny tree was constructed using the Maximum Likelihood method. This study showed that among 22 isolates of B. thuringiensis, only BA02b, BS04a, and BA03a isolates have similar phenotypic characters with B. thuringiensis. Based on the toxicity test of B. thuringiensis against the third instar of Ae. aegypti larvae, it was indicated that BA02b and BA03a isolates were the potential agents to control Ae. aegypti larvae. BA02b isolate was the most effective B. thuringiensis (LC50-96h = 2,75 x 107 cell/mL). Based on 16S rDNA sequence, BA02b was identified as Bacillus thuringiensis var. Israelensis BGSC4Q2 (99 % similarities).

  10. Proteome response of Tribolium castaneum larvae to Bacillus thuringiensis toxin producing strains.

    Directory of Open Access Journals (Sweden)

    Estefanía Contreras

    Full Text Available Susceptibility of Tribolium castaneum (Tc larvae was determined against spore-crystal mixtures of five coleopteran specific and one lepidopteran specific Bacillus thuringiensis Cry toxin producing strains and those containing the structurally unrelated Cry3Ba and Cry23Aa/Cry37Aa proteins were found toxic (LC(50 values 13.53 and 6.30 µg spore-crystal mixture/µL flour disc, respectively. Using iTRAQ combined with LC-MS/MS allowed the discovery of seven novel differentially expressed proteins in early response of Tc larvae to the two active spore-crystal mixtures. Proteins showing a statistically significant change in treated larvae compared to non-intoxicated larvae fell into two major categories; up-regulated proteins were involved in host defense (odorant binding protein C12, apolipophorin-III and chemosensory protein 18 and down-regulated proteins were linked to metabolic pathways affecting larval metabolism and development (pyruvate dehydrogenase Eα subunit, cuticular protein, ribosomal protein L13a and apolipoprotein LI-II. Among increased proteins, Odorant binding protein C12 showed the highest change, 4-fold increase in both toxin treatments. The protein displayed amino acid sequence and structural homology to Tenebrio molitor 12 kDa hemolymph protein b precursor, a non-olfactory odorant binding protein. Analysis of mRNA expression and mortality assays in Odorant binding protein C12 silenced larvae were consistent with a general immune defense function of non-olfactory odorant binding proteins. Regarding down-regulated proteins, at the transcriptional level, pyruvate dehydrogenase and cuticular genes were decreased in Tc larvae exposed to the Cry3Ba producing strain compared to the Cry23Aa/Cry37Aa producing strain, which may contribute to the developmental arrest that we observed with larvae fed the Cry3Ba producing strain. Results demonstrated a distinct host transcriptional regulation depending upon the Cry toxin treatment. Knowledge

  11. The persistence and ecological impacts of a cyanobacterium genetically engineered to express mosquitocidal Bacillus thuringiensis toxins.

    Science.gov (United States)

    Ketseoglou, Irene; Bouwer, Gustav

    2016-05-10

    The cyanobacterium Anabaena PCC 7120#11 has been genetically engineered to act as a delivery vehicle for Bacillus thuringiensis subspecies israelensis mosquitocidal toxins. To address ecological concerns about releasing this genetically engineered microorganism into the environment for mosquito larva control, the persistence and ecological impacts of PCC 7120#11 was evaluated using multi-species, standardized aquatic microcosms. The microcosms were set up as described in ASTM E1366-02 (Standard Practice for Standardized Aquatic Microcosms: Fresh Water), with a few modifications. The treatment group microcosms were inoculated with PCC 7120#11 and key water quality parameters and non-target effects were compared between the treatment and control groups over a period of 35 days. PCC 7120#11 decreased from a concentration of 4.50 × 10(6) cells/ml (at inoculation) to 1.32 × 10(3) cells/ml after 4 weeks and larvicidal activity against third instar larvae of Anopheles arabiensis was only evident for two weeks after treatment. Both treatment and the interaction of treatment and time had a significant effect on nitrate, phosphate and photosynthetic microorganism concentrations. Treatment with PCC 7120#11 caused a temporary spike in ammonia in the microcosms a week after treatment, but the concentrations were well below acute and chronic criteria values for ammonia in freshwater ecosystems. Cyprinotus vidua concentrations were not significantly different between PCC 7120#11 and control microcosms. In PCC 7120#11 microcosms, Daphnia pulex concentrations were significantly lower than control concentrations between days 18 and 25. By the end of the experiment, none of the measured variables were significantly different between the treatment groups. The standard aquatic microcosm experiments provided more data on the ecological impacts of PCC 7120#11 than single-organism assessments would have. On the basis of the relatively minor, short-term effects that PCC 7120

  12. Sub-chronic lung inflammation after airway exposures to Bacillus thuringiensis biopesticides in mice

    Directory of Open Access Journals (Sweden)

    Barfod Kenneth K

    2010-09-01

    Full Text Available Abstract Background The aim of the present study was to assess possible health effects of airway exposures to Bacillus thuringiensis (Bt based biopesticides in mice. Endpoints were lung inflammation evaluated by presence of inflammatory cells in bronchoalveolar lavage fluid (BALF, clearance of bacteria from the lung lumen and histological alterations of the lungs. Hazard identifications of the biopesticides were carried out using intratracheal (i.t. instillation, followed by an inhalation study. The two commercial biopesticides used were based on the Bt. subspecies kurstaki and israelensis, respectively. Groups of BALB/c mice were i.t instilled with one bolus (3.5 × 105 or 3.4 × 106 colony forming units (CFU per mouse of either biopesticide. Control mice were instilled with sterile water. BALFs were collected and the inflammatory cells were counted and differentiated. The BALFs were also subjected to CFU counts. Results BALF cytology showed an acute inflammatory response dominated by neutrophils 24 hours after instillation of biopesticide. Four days after instillation, the neutrophil number was normalised and inflammation was dominated by lymphocytes and eosinophils, whereas 70 days after instillation, the inflammation was interstitially located with few inflammatory cells present in the lung lumen. Half of the instilled mice had remaining CFU recovered from BALF 70 days after exposure. To gain further knowledge with relevance for risk assessment, mice were exposed to aerosols of biopesticide one hour per day for 2 × 5 days. Each mouse received 1.9 × 104 CFU Bt israelensis or 2.3 × 103 CFU Bt kurstaki per exposure. Seventy days after end of the aerosol exposures, 3 out of 17 mice had interstitial lung inflammation. CFU could be recovered from 1 out of 10 mice 70 days after exposure to aerosolised Bt kurstaki. Plethysmography showed that inhalation of Bt aerosol did not induce airway irritation. Conclusions Repeated low dose aerosol

  13. Characterization and selection of Bacillus thuringiensis isolates effective against Sitophilus oryzae Caracterização e seleção de isolados de Bacillus thuringiensis efetivos contra Sitophilus oryzae

    Directory of Open Access Journals (Sweden)

    Najara da Silva

    2010-08-01

    Full Text Available The entomopathogenic bacterium Bacillus thuringiensis is a control agent with toxic and environmental characteristics that allows the control of pest insects according to the Integrate Pest Management (IPM precepts. In order to find new strains, potentially toxic to Sitophilus oryzae L. 1763 (Coleoptera: Curculinidae, 1.073 strains of B. thuringiensis from parts of Brazil were used. Genetic material was extracted with InstaGene Matrix kit, used for the amplification of sequences in Polymerase chain reaction (PCR, and viewed in 1.5% agarose gel. The gene cry35Ba class was represented by 60 B. thuringiensis isolates (5.6%, which were then subjected to bioassays with S. oryzae larvae. Among the isolates studied, four caused more than 50% mortality in pathogenicity tests, and the isolates 544 and 622 were the most virulent, as determined by CL50 estimates. The four toxic isolates had spherical, bi-pyramidal and cuboid crystals, and a 44-kDa protein was found in sodium dodecyl sulphate - polyacrylamide gel electrophoresis (SDS-PAGE, which coded for the product of cry35Ba genes. These data demonstrate the potential of B. thuringiensis for the management of S. oryzae larvae.A bactéria entomopatogênica Bacillus thuringiensis (Bt é um agente de controle com características tóxicas e ambientais que permitem o controle de insetos-praga de acordo com as premissas do Manejo integrado de pragas (MIP. Com o objetivo de buscar novas linhagens potencialmente tóxicas para Sitophilus oryzae L. 1763 (Coleoptera: Curculinidae, caracterizaram-se molecularmente 1,073 isolados de B. thuringiensis de regiões do Brasil. O material genético foi extraído através do kit InstaGene Matrix, utilizado para a amplificação das seqüências através da técnica de Polymerase chain reaction PCR, sendo os resultados visualizados em gel de agarose 1,5%. A classe do gene cry35Ba foi representada por 60 isolados (5,6% de Bt, os quais foram submetidos a bioensaio com larvas

  14. Fatores de virulência de Bacillus thuringiensis: o que existe além das proteínas Cry

    Directory of Open Access Journals (Sweden)

    Gislayne Vilas-Bôas

    2012-03-01

    Virulence Factors of Bacillus thuringiensis Berliner: Something Beyond of Cry Proteins? Abstract. The Cry proteins produced by the entomopathogenic bacterium Bacillus thuringiensis Berliner are widely known due to its high toxicity against a variety of insects. The mode of action of these proteins is specific and becomes B. thuringiensis-based products the most used in biological control programs of insect pests in agriculture and of important human disease vectors. However, while the Cry proteins are the best-known insect-specific virulence factor, strains of B. thuringiensis show also a wide range of other virulence factors, which allow the bacteria to achieve the hemolymph and colonize efficiently the insect host. Among these factors, we highlight the Vip proteins, Cyt, enterotoxins, hemolysins, phospholipases, proteases and enzymes of degradation, in addition to the recently described parasporin. This review explores the action of these virulence factors, as well as, the characterization and control of expression of their genes. Additionally, we discuss aspects related to the ecological niche of the bacteria with emphasis on the characteristics involved in the biosafety of the use of B. thuringiensis-based products for biological control of target insects.

  15. Persistence of Bt Bacillus thuringiensis Cry1Aa toxin in various soils determined by physicochemical reactions

    Science.gov (United States)

    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

  16. Impact of UV radiation on activity of linear furanocoumarins and Bacillus thuringiensis var. kurstaki against Spodoptera exigua: Implications for tritrophic interactions

    Energy Technology Data Exchange (ETDEWEB)

    Trumble, J.T.; Moar, W.J.; Brewer, M.J.; Carson, W.G. (Univ. of California, Riverside (United States))

    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.

  17. Impact of UV radiation on activity of linear furanocoumarins and Bacillus thuringiensis var. kurstaki against Spodoptera exigua: Implications for tritrophic interactions

    International Nuclear Information System (INIS)

    Trumble, J.T.; Moar, W.J.; Brewer, M.J.; Carson, W.G.

    1991-01-01

    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 x 2 x 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

  18. Isolation and molecular characterization of Bacillus thuringiensis found in soils of the Cerrado region of Brazil, and their toxicity to Aedes aegypti larvae

    Directory of Open Access Journals (Sweden)

    Katiane dos Santos Lobo

    Full Text Available ABSTRACT This study investigated the potential of Bacillus thuringiensis isolates obtained in the Cerrado region of the Brazilian state of Maranhão for the biological control of Aedes aegypti larvae. The isolates were obtained from soil samples and the identification of the B. thuringiensis colonies was based on morphological characteristics. Bioassays were run to assess the pathogenicity and toxicity of the different strains of the B. thuringiensis against third-instar larvae of A. aegypti. Protein profiles were obtained by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Polymerase chain reaction assays were used to detect the toxin genes found in the bacterial isolates. Overall, 12 (4.0% of the 300 isolates obtained from 45 soil samples were found to present larvicidal activity, with the BtMA-104, BtMA-401 and BtMA-560 isolates causing 100% of mortality. The BtMA-401 isolate was the most virulent, with the lowest median lethal concentration (LC50 (0.004 × 107 spores/mL, followed by the Bacillus thuringiensis var. israelensis standard (0.32 × 107 spores/mL. The protein profiles of BtMA-25 and BtMA-401 isolates indicated the presence of molecular mass consistent with the presence of the proteins Cry4Aa, Cry11Aa and Cyt1, similar to the profile of Bacillus thuringiensis var. israelensis IPS-82. Surprisingly, however, none of the cry and cyt genes analyzed were amplified in the isolate BtMA-401. The results of the present study revealed the larvicidal potential of B. thuringiensis isolates found in the soils of the Cerrado region from Maranhão, although further research will be necessary to better elucidate and describe other genes associated with the production of insecticidal toxins in these isolates.

  19. Characterization of a Bacillus thuringiensis strain collection isolated from diverse Costa Rican natural ecosystems

    Directory of Open Access Journals (Sweden)

    Glen Arrieta

    2006-03-01

    Full Text Available 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 (Cararait 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 Río Medio Queso and Parque Marino las Baulas. Protein analyses of the crystal-spore preparations showed ä -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

  20. Presence and significance of Bacillus thuringiensis Cry proteins associated with the Andean weevil Premnotrypes vorax (Coleoptera: Curculionidae

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    SilvioAlejandro López-Pazos

    2009-12-01

    Full Text Available The Andean weevil Premnotrypes vorax represents an important cause of damage to Colombian potato crops. Due to the impact of this plague on the economy of the country, we searched for new alternatives for its biological control, based on the entomopathogenic bacteria Bacillus thuringiensis. A total of 300 B. thuringiensis strains obtained from potato plantations infested with P. vorax were analyzed through crystal morphology, SDS-PAGE, PCR and bioassays. We used site- directed mutagenesis to modify the Cry3Aa protein. Most of the B. thuringiensis isolates had a bipyramidal crystal morphology. SDS-PAGE analyses had seven strains groups with σ-endotoxins from 35 to 135 kDa. The genes cry 2 and cry 1 were significantly more frequent in the P. vorax habitat (PCR analyses. Three mutant toxins, 1 (D354E, 2 (R345A, ∆Y350, ∆Y351, and 3 (Q482A, S484A, R485A, were analyzed to assess their activity against P. vorax larvae. Toxicity was low, or absent, against P. vorax for isolates, wild type cry 3Aa and cry 3Aa mutants. The genetic characterization of the collection provides opportunities for the selection of strains to be tested in bioassays against other insect pests of agricultural importance, and for designing Cry proteins with improved insecticidal toxicity. Rev. Biol. Trop. 57 (4: 1235-1243. Epub 2009 December 01.El gorgojo andino Premnotrypes vorax es una causa importante de daño en los cultivos colombianos de este tubérculo. Debido al impacto que esta plaga tiene sobre la economía del país, nos interesamos en buscar alternativas nuevas para el control biológico de P. vorax, basadas en la bacteria entomopatógena Bacillus thuringiensis. Se recolectaron un total de 300 cepas de B. thuringiensis a partir de plantaciones de papa infestadas con P. vorax, las cuales fueron analizadas por medio de la morfología del cristal, SDS-PAGE, PCR y ensayos biológicos. La mayoría de los aislamientos de B. thuringiensis presentaron cristales

  1. Bacillus thuringiensis Cry1Ca-resistant Spodoptera exigua lacks expression of one of four Aminopeptidase N genes

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    Moar William J

    2005-06-01

    Full Text Available Abstract Background Insecticidal toxins from Bacillus thuringiensis bind to receptors on midgut epithelial cells of susceptible insect larvae. Aminopeptidases N (APNs from several insect species have been shown to be putative receptors for these toxins. Here we report the cloning and expression analysis of four APN cDNAs from Spodoptera exigua. Results Suppression Subtractive Hybridization (SSH was used to construct cDNA libraries of genes that are up-and down-regulated in the midgut of last instar larvae of beet armyworm, S. exigua exposed to B. thuringiensis Cry1Ca toxin. Among the clones from the SSH libraries, cDNA fragments coding for two different APNs were obtained (APN2 and APN4. A similar procedure was employed to compare mRNA differences between susceptible and Cry1Ca resistant S. exigua. Among the clones from this last comparison, cDNA fragments belonging to a third APN (APN1 were detected. Using sequences obtained from the three APN cDNA fragments and degenerate primers for a fourth APN (APN3, the full length sequences of four S. exigua APN cDNAs were obtained. Northern blot analysis of expression of the four APNs showed complete absence of APN1 expression in the resistant insects, while the other three APNs showed similar expression levels in the resistant and susceptible insects. Conclusion We have cloned and characterized four different midgut APN cDNAs from S. exigua. Expression analysis revealed the lack of expression of one of these APNs in the larvae of a Cry1Ca-resistant colony. Combined with previous evidence that shows the importance of APN in the mode of action of B. thuringiensis toxins, these results suggest that the lack of APN1 expression plays a role in the resistance to Cry1Ca in this S. exigua colony.

  2. Risk assessment of the biological plant protection product Turex 50 WG, with the organism Bacillus thuringiensis ssp. aizawai CG-91. Opinion of the Panel on Plant Protection Products of the Norwegian Scientific Committee for Food Safety

    OpenAIRE

    Källqvist, Torsten; Dirven, Hubert; Gjøen, Tor; Tronsmo, Arne; Yazdankhah, Siamak Pour; Rivedal, Edgar; Borgå, Katrine; Eklo, Ole Martin; Grung, Merete; Lyche, Jan Ludvig; Låg, Marit; Nilsen, Asbjørn Magne; Sverdrup, Line Emilie

    2016-01-01

    Bacillus thuringiensis are anaerobic, gram-positive bacteria that produce parasporal crystalline protein inclusions, δ-endotoxin, which are toxic to certain invertebrates, especially larvae belonging to the insect orders Coleoptera, Diptera and Lepidoptera. Different strains of Bacillus thuringiensis have therefore a long standing history as plant protective insecticides in many countries, but have not been approved for use in Norway. The Norwegian Scientific Committee for Food Safety (Vitens...

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

    Directory of Open Access Journals (Sweden)

    Neiva Knaak

    2007-09-01

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

  4. Use of Bacillus thuringiensis supernatant from a fermentation process to improve bioremediation of chlorpyrifos in contaminated soils.

    Science.gov (United States)

    Aceves-Diez, Angel E; Estrada-Castañeda, Kelly J; Castañeda-Sandoval, Laura M

    2015-07-01

    The aim of this research was to investigate the potential of a nutrient-rich organic waste, namely the cell-free supernatant of Bacillus thuringiensis (BtS) gathered from fermentation, as a biostimulating agent to improve and sustain microbial populations and their enzymatic activities, thereby assisting in the bioremediation of chlorpyrifos-contaminated soil at a high dose (70 mg kg(-1)). Experiments were performed for up to 80 d. Chlorpyrifos degradation and its major metabolic product, 3,5,6-trichloro-2-pyridinol (TCP), were quantified by high-performance liquid chromatography (HPLC); total microbial populations were enumerated by direct counts in specific medium; and fluorescein diacetate (FDA) hydrolysis was measured as an index of soil microbial activity. Throughout the experiment, there was higher chlorpyrifos degradation in soil supplemented with BtS (83.1%) as compared to non-supplemented soil. TCP formation and degradation occurred in all soils, but the greatest degradation (30.34%) was observed in soil supplemented with BtS. The total microbial populations were significantly improved by supplementation with BtS. The application of chlorpyrifos to soil inhibited the enzymatic activity; however, this negative effect was counteracted by BtS, inducing an increase of approximately 16% in FDA hydrolysis. These results demonstrate the potential of B. thuringiensis supernatant as a suitable biostimulation agent for enhancing chlorpyrifos and TCP biodegradation in chlorpyrifos-contaminated soils. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. PENGARUH BEBERAPA DOSIS BACILLUS THURINGIENSIS VAR ISRAELENSIS SEROTYPE H14 TERHADAP LARVA AEDES AEGYPTI DI KALIMANTAN BARAT

    Directory of Open Access Journals (Sweden)

    Dian Perwitasari

    2016-03-01

    Full Text Available One of the dengue control efforts is the use  of Bacillus thuringiensis in order to reduce the dengue vector of Aedes aegypti through its larvas. This was an experimental research using gram-positive bacteria B. thuringiensis var israelensis (Bactivec serotype H-14 which was applied with several concentrations (0.02 ml, 0.01 ml and 0.007 ml in 246 ml of water that has been filled with 25 larvas of the 3rd or 4th instars. Larvas were taken from the area of ​​West Kalimantan. Data were analyzed using a completely randomized design to examine the percentage of larval mortality within 3 hours, 9 hours and over 12 hours. The results showed that the concentration of 0,02 bactivec caused 89%  larval mortality, and  concentrations of 0,01 and 0,007 caused 88% and 87% larval mortality, respectively within the 9 hours exposure time. It can be concluded that the use of 0,07 ml of bactivec  is still effective to  control  Aedes aegypti larvae. To determine  the negative of  the use of bactivec,  further studies  are needed.  

  6. Characterization of lepidopteran-specific cry1 and cry2 gene harbouring native Bacillus thuringiensis isolates toxic against Helicoverpa armigera

    Directory of Open Access Journals (Sweden)

    Showkat Ahmad Lone

    2017-09-01

    Full Text Available Bacillus thuringiensis (Bt based biopesticides are feasible alternatives to chemical pesticides. Here, we present the distribution of lepidopteran-specific cry1 and cry2 genes in native B. thuringiensis. Forty four out of 86 colonies were found to harbour crystals by phase contrast microscopy exhibiting a Bt index of 0.51. PCR analysis resulted in the amplification of cry1 in 24 and cry2 in 14 isolates. Twelve of the isolates showed presence of both cry1 and cry2, while 18 isolates did not show presence of either of the genes. Toxicity screening using spore-crystal mixtures against 2nd instar larvae of Helicoverpa armigera revealed that the isolates (50% were either mildly toxic or not toxic (36.36%, and only 13.63% were toxic. The results are interesting, particularly so because the same isolates were previously reported to contain lepidopteran specific vip3A genes also, hence can complement the toxicity of the isolates harbouring vip3A genes.

  7. Use of endophytic diazotrophic bacteria as a vector to express the cry3A gene from Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Salles Joana Falcão

    2000-01-01

    Full Text Available The goal of this study was to evaluate the potential of endophytic diazotrophic bacteria as a vector to express a cry gene from Bacillus thuringiensis, envisaging the control of pests that attack sugarcane plants. The endophytic nitrogen-fixing bacteria Gluconacetobacter diazotrophicus strain BR11281 and Herbaspirillum seropedicae strain BR11335 were used as models. The cry3A gene was transferred by conjugation using a suicide plasmid and the recombinant strains were selected by their ability to fix nitrogen in semi-solid N-free medium. The presence of the cry gene was detected by Southern-blot using an internal fragment of 1.0 kb as a probe. The production of delta-endotoxin by the recombinant H. seropedicae strain was detected by dot blot while for G. diazotrophicus the Western-blot technique was used. In both cases, a specific antibody raised against the B. thuringiensis toxin was applied. The delta-endotoxin production showed by the G. diazotrophicus recombinant strain was dependent on the nitrogen fixing conditions since the cry3A gene was fused to a nif promoter. In the case of H. seropedicae the delta-endotoxin expression was not affected by the promoter (rhi used. These results suggest that endophytic diazotrophic bacteria can be used as vectors to express entomopathogenic genes envisaging control of sugarcane pests.

  8. Decontamination of materials contaminated with Bacillus anthracis and Bacillus thuringiensis Al Hakam spores using PES-Solid, a solid source of peracetic acid.

    Science.gov (United States)

    Buhr, T L; Wells, C M; Young, A A; Minter, Z A; Johnson, C A; Payne, A N; McPherson, D C

    2013-08-01

    To develop test methods and evaluate survival of Bacillus anthracis Ames, B. anthracis ∆Sterne and B. thuringiensis Al Hakam spores after exposure to PES-Solid (a solid source of peracetic acid), including PES-Solid formulations with bacteriostatic surfactants. Spores (≥ 7 logs) were dried on seven different test materials and treated with three different PES-Solid formulations (or preneutralized controls) at room temperature for 15 min. There was either no spore survival or less than 1 log (<10 spores) of spore survival in 56 of 63 test combinations (strain, formulation and substrate). Less than 2.7 logs (<180 spores) survived in the remaining seven test combinations. The highest spore survival rates were seen on water-dispersible chemical agent resistant coating (CARC-W) and Naval ship topcoat (NTC). Electron microscopy and Coulter analysis showed that all spore structures were intact after spore inactivation with PES-Solid. Three PES-Solid formulations inactivated Bacillus spores that were dried on seven different materials. A test method was developed to show that PES-Solid formulations effectively inactivate Bacillus spores on different materials. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.

  9. Controle de tortricídeos em macieira com duas formulações de Bacillus thuringiensis var. Kustaki em Fraiburgo-SC Tortricid moth control in apple with two formulation of Bacillus thuringiensis var. Kustaki in Fraiburgo-SC

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    Lino Bittencourt Monteiro

    2010-06-01

    Full Text Available Grapholita molesta e Bonagota cranaodes são duas importantes pragas de pomares de macieira controlada por inseticidas. O objetivo deste estudo foi testar duas formulações de Bacillus thuringiensis var. kurstaki (Btk para o controle desses tortricídeos. Parcelas de macieira 'Fuji' foram pulverizadas com Dipel PM e Dipel SC, nas concentrações de 100 ml por100 L de calda. A eficiência do Btk foi comparada com os inseticidas tebufenozide (Mimic 240 SC - 90 ml por 100 L e clorpyrifos (Lorsban 480 BR - 150 ml por 100 L. Duas vezes por semana, eram avaliadas as capturas de pragas em armadilha do tipo Delta. As avaliações de danos nos frutos foram realizadas antes e na colheita, sendo classificados os frutos em função do agrupamento em cachopa e a sua localização na planta. Os tratamentos com Btk tiveram mais danos do que os químicos. Houve uma tendência de maiores danos na parte interna, próximo ao tronco das macieiras e o agrupamento dos frutos não influenciou na presença de danos. Ambas as formulações de Dipel foram eficientes quando comparados com Mimic e Lorsban.Grapholita molesta and Bonagota cranaodes are two key pests in apple orchards controlled by insecticides. The objective of the present study was to test two formulas of Bacillus thuringiensis var. kurstaki to control these tortricid moths. Plot of 'Fuji' apple trees were sprayed with Dipel PM and Dipel SC, at a concentration of 100ml por 100 l liquid. The efficiency of Btk was compared with the insecticides tebuphenozide (Mimic 240 SC - 90 ml por 100 L and chlorpyrifos (Lorsban 480 BR - 150 ml por 100 L. The capture of pests in a Delta-type trap was assessed twice a week. Fruit damage was evaluated pre- and post- harvest and the fruit were classified by branch-end clustering and location on the plant. The treatments with B. thuringiensis were more damaged than those with chemical treatments. There tended to be more damage in the internal part, close to the apple tree

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

    DEFF Research Database (Denmark)

    Ke, X.; Krogh, P. H.

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

  11. Chitinase and cellulase activity from Bacillus thuringiensis strains - doi: 10.5102/ucs.v7i1.974

    Directory of Open Access Journals (Sweden)

    Vinícius Fiúza Dumas

    2009-12-01

    Full Text Available The present study aimed to analyze the production of chitinase and cellulase enzymes by strains of Bacillus thuringiensis toxic to Spodoptera frugiperda and Anthonomus grandis larvae. In order to evaluate the relationship between cellular growth and the chitinase and cellulase production, in vitro assays were carried through with bacteria cultures grown for 16h, 24h, 48h and 72h. Chitinase and cellulase activity was determined by a colorimetric method. The amount of N-acetylglucosamine (GlcNAc or its equivalent was measured by development of color in acid medium. All strains presented enzymatic production after 16h of cellular growth until 72h. However, a Kruskal-Wallis test detected no significant differences among the chitinase and cellulase activity during the cellular growth. According to these results, was not possible to associate chitinase and cellulose activity with the different level of toxicity of Bt strains against S. frugiperda and A. grandis larvae.

  12. Transmission dynamics of Bacillus thuringiensis infecting Plodia interpunctella: a test of the mass action assumption with an insect pathogen.

    Science.gov (United States)

    Knell, R J; Begon, M; Thompson, D J

    1996-01-22

    Central to theoretical studies of host-pathogen population dynamics is a term describing transmission of the pathogen. This usually assumes that transmission is proportional to the density of infectious hosts or particles and of susceptible individuals. We tested this assumption with the bacterial pathogen Bacillus thuringiensis infecting larvae of Plodia interpunctella, the Indian meal moth. Transmission was found to increase in a more than linear way with host density in fourth and fifth instar P. interpunctella, and to decrease with the density of infectious cadavers in the case of fifth instar larvae. Food availability was shown to play an important part in this process. Therefore, on a number of counts, the usual assumption was found not to apply in our experimental system.

  13. A theoretical model of the tridimensional structure of Bacillus thuringiensis subsp. medellin Cry 11Bb toxin deduced by homology modelling

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    Gutierrez Pablo

    2001-01-01

    Full Text Available Cry11Bb is an insecticidal crystal protein produced by Bacillus thuringiensis subsp. medellin during its stationary phase; this ¶-endotoxin is active against dipteran insects and has great potential for mosquito borne disease control. Here, we report the first theoretical model of the tridimensional structure of a Cry11 toxin. The tridimensional structure of the Cry11Bb toxin was obtained by homology modelling on the structures of the Cry1Aa and Cry3Aa toxins. In this work we give a brief description of our model and hypothesize the residues of the Cry11Bb toxin that could be important in receptor recognition and pore formation. This model will serve as a starting point for the design of mutagenesis experiments aimed to the improvement of toxicity, and to provide a new tool for the elucidation of the mechanism of action of these mosquitocidal proteins.

  14. Extraction and Characterization of Polyhydroxybutyrates (PHB from Bacillus thuringiensisKSADL127 Isolated from Mangrove Environments of Saudi Arabia

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    Abdullah A. Alarfaj

    2015-10-01

    Full Text Available ABSTRACTPolyhydroxybutyrate (PHB is a renowned biodegradable plastic that do not release any toxins or residues in the environment like petroleum based plastics. In the present study, 50 bacteria isolated from mangrove niche, Saudi Arabia, were screened for maximum PHB production. All the 50 strains showed positive for PHB production, of which one strain showed maximum of 137 mgL-1. The most PHB accumulated bacterium was selected and identified asBacillus thuringiensis KSADL127, based on phenotypic characterization and 16S rRNA sequence analysis. Characterization of extracted PHB was carried out by FT-IR, NMR, UV spectroscopy, DSC, TGA, and LC-MS, which later confirmed the presence of intracellular accumulated polymer and substantiated as PHB.

  15. Compatibility and survival of beneficial microorganisms for agricultural use (Beauveria bassiana, Paecilomyces lilacinus and Bacillus thuringiensis in compost

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    Villacís-Aldaz Luis Alfredo

    2016-11-01

    Full Text Available Compatibility and survivorship of beneficial microorganism in compost were evaluated in order to provide new alternatives to clean production in agriculture. The study was conducted at the Ecological Farm belonging to the Consejo Provincial de Tungurahua, Ecuador. Four treatments were considered related to dose of several beneficial microorganisms (EMs. After an interval of 30 or 60 days after composting process, analysis of nutritional biological content showed compatibility and survivorship of the EMs, which live together and reproduce on the organic fertilizer. Concerning physic-chemical analysis higher organic matter content, pH, electrical conductivity, nitrogen, phosphorous, potassium and calcium were evidenced in T1: 100 cc Beauveria bassiana - 100 cc Bacillus thuringiensis - 100 cc Paecilomyces lilacinus. From the results obtained will offer the possibility of using beneficial microorganisms to optimize their performance in obtaining compost of better nutritional and biological quality, so that gradually reduce the use of agrochemicals.

  16. Pesquisa de proteínas citotóxicas e nematicidas em isolados Bacillus thuringiensis dos Açores

    OpenAIRE

    Raimundo, Natacha Sofia Rodrigues

    2016-01-01

    Dissertação de Mestrado, Ciências Biomédicas, 25 de Maio de 2016, Universidade dos Açores. Bacillus thuringiensis (Bt), pertencente à família Bacillaceae, é classificado como gram-positiva e tem despertado elevado interesse em várias áreas, destacando-se a aplicações na agricultura e na saúde. Os isolados de Bt são caracterizados pela sua capacidade de produzir inclusões cristalinas de origem proteica durante a fase de esporulação, as quais são conhecidas como proteínas δ-endotoxinas ou pr...

  17. Amélioration des activités insecticides des protéines Vip3 de Bacillus thuringiensis

    Directory of Open Access Journals (Sweden)

    Sameh SELLAMI

    2016-07-01

    Full Text Available Vip3 proteins were considered as a second generation of biopesticides. They are synthesized and secreted by Bacillus thuringiensis during the vegetative growth phase. Vip3 proteins, which are discovered in 1990, are of great interest for the control of Lepidopteran insects pests such as Agrotis ipsilon, Spodoptera littoralis, Spodoptera exigua and Spodoptera frugiperda. Many researches were conducted on the Vip3 proteins in order to enlarge their spectrum, improve their insecticidal activities and resolve the problems of resistance that appeared recently after the massive use of δ-endotoxins, considered as first generation of biopesticides. In this review, we tried to summarize research studies interested in the improvement of the insecticidal activities of Vip3 proteins.

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

    Energy Technology Data Exchange (ETDEWEB)

    Kweon, Hyojin [Georgia Inst. of Technology, Atlanta, GA (United States); Yiacoumi, Sotira [Georgia Inst. of Technology, Atlanta, GA (United States); Tsouris, Costas [Georgia Inst. of Technology, Atlanta, GA (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)

    2011-11-07

    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. In conclusion, 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.

  19. Different toxicity of the novel Bacillus thuringiensis (Bacillales: Bacillaceae) strain LLP29 against Aedes albopictus and Culex quinquefasciatus (Diptera: Culicidae)

    Czech Academy of Sciences Publication Activity Database

    Zhang, L.; Tang, B.; Huang, E.; Huang, Z.; Liu, Z.; Huang, T.; Gelbič, Ivan; Guan, X.; Xu, L.

    2013-01-01

    Roč. 106, č. 3 (2013), s. 1098-1102 ISSN 0022-0493 Grant - others:National Natural Science Foundation of China(CN) 31071745; National Natural Science Foundation of China(CN) 31201574; Ministry of Education of China(CN) 20093515110010; Ministry of Education of China(CN) 20093515120010; Agricultural Science and Technology Achievements(CN) 2010GB2C400212; National High Technology Research and Development Program 863(CN) 2011AA10A203; Universities of Fujian Province(CN) JA12092; Fujian Agriculture and Forestry University(CN) xjq201203; Universities for the Development of the West Strait(CN) 0b08b005 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis * receptor binding * ELISA Subject RIV: ED - Physiology Impact factor: 1.605, year: 2013 http://www.bioone.org/doi/pdf/10.1603/EC12308

  20. Specificity of Bacillus thuringiensis endotoxins is correlated with the presence of high-affinity binding sites in the brush border membrane of target insect midguts

    International Nuclear Information System (INIS)

    Hofmann, C.; Vanderbruggen, H.; Hoefte, H.; Van Rie, J.; Jansens, S.; Van Mellaert, H.

    1988-01-01

    Binding studies were performed with two 125 I-labeled Bacillus thuringiensis δ-endotoxins on brush border membrane vesicles prepared from the larval midgut of the tobacco hornworm Manduca sexta or the cabbage butterfly Pieris brassicae. One δ-endotoxin, Bt2-protoxin, is a 130-kDa recombinant crystalline protein from B. thuringiensis subsp. berliner. It kills larvae of both insect species. The active Bt2-toxin is a 60-kDa proteolytic fragment of the Bt2-protoxin. It binds saturably and with high affinity to brush border membrane vesicles from the midgut of both species. The other δ-endotoxin, Bt4412-protoxin, is a 136-kDa crystalline protein from B. thuringiensis subsp. thuringiensis, which is highly toxic for P. brassicae, but not for M. sexta larvae. Bt4412-toxin, obtained after proteolytic activation of Bt4412-protoxin, shows high-affinity saturable binding to P. brassicae vesicles but not to M. sexta vesicles. The correlation between toxicity and specific binding is further strengthened by competition studies. Other B. thuringiensis δ-endotoxins active against M. sexta compete for binding of 125 I-labeled Bt2-toxin to M. sexta vesicles, whereas toxins active against dipteran or coleopteran larvae do not compete. Bt2-toxin and Bt4412-toxin bind to different sites on P. brassicae vesicles

  1. Native plant growth promoting bacteria Bacillus thuringiensis and mixed or individual mycorrhizal species improved drought tolerance and oxidative metabolism in Lavandula dentata plants.

    Science.gov (United States)

    Armada, E; Probanza, A; Roldán, A; Azcón, R

    2016-03-15

    This study evaluates the responses of Lavandula dentata under drought conditions to the inoculation with single autochthonous arbuscular mycorrhizal (AM) fungus (five fungal strains) or with their mixture and the effects of these inocula with a native Bacillus thuringiensis (endophytic bacteria). These microorganisms were drought tolerant and in general, increased plant growth and nutrition. Particularly, the AM fungal mixture and B. thuringiensis maximized plant biomass and compensated drought stress as values of antioxidant activities [superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase APX)] shown. The AMF-bacteria interactions highly reduced the plant oxidative damage of lipids [malondialdehyde (MDA)] and increased the mycorrhizal development (mainly arbuscular formation representative of symbiotic functionality). These microbial interactions explain the highest potential of dually inoculated plants to tolerate drought stress. B. thuringiensis "in vitro" under osmotic stress does not reduce its PGPB (plant growth promoting bacteria) abilities as indole acetic acid (IAA) and ACC deaminase production and phosphate solubilization indicating its capacity to improve plant growth under stress conditions. Each one of the autochthonous fungal strains maintained their particular interaction with B. thuringiensis reflecting the diversity, intrinsic abilities and inherent compatibility of these microorganisms. In general, autochthonous AM fungal species and particularly their mixture with B. thuringiensis demonstrated their potential for protecting plants against drought and helping plants to thrive in semiarid ecosystems. Copyright © 2015 Elsevier GmbH. All rights reserved.

  2. EFEKTIVITAS VECTOBAC 12 AS (Bt H-14 DAN Bacillus thuringiensis H-14 TERHADAP VEKTOR MALARIA An. maculatus DI KOBAKAN DESA HARGOTIRTO, KECAMATAN KOKAP, KABUPATEN KULON PROGO

    Directory of Open Access Journals (Sweden)

    Blondine Ch. P.

    2012-09-01

    Full Text Available A study using Vectobac 12 AS ( Bt H-14 and Bacillus thuringiensis H-14 local strain was conducted at laboratory Vector and Reservoir Control Research Unit and breeding ponds of Anopheles maculatus in Kokap regency and Kulon Progo district. The objectives of this study are : (1. to detect the efficacy of B. thuringiensis H-14 local strain toward An. maculatus larvae at the laboratory. (2. to measure the effectiveness of B. thuringiensis H-14 local strain dosages 1 x LC95. 5 x LC95 and 10 x LC95 toward An. maculatus at the field. The efficacy test of B. thuringiensis H-14 local strain toward An. maculatus based on to the method proposed by WHO in order to determine the LC50 and LC90 which is computed using the probit analysis at the laboratory. The methods used R thuringiensis H-14 local strain dosages of 2.145 ppm (1 x LC95, 10.724 ppm (5 x LC95 and 21.448 ppm (10 x LC95 respectively were applied 8 ponds with the width of ponds ranging from 0.08 to 0.45 m2, 0.29 to 0.64 m2 and from 0.08-0.79 m2.The results showed, the dosages after 24 hours were 10.22 ppm (LC50, 27.11 ppm (LC90 and 35.75 ppm (LC95. After 48 hours the dosages were needed 7.74 ppm (LC50, 17.06 ppm (LC90 and 21.34 ppm (LC95, The effectiveness o/R thuringiensis H-14 local strain dosages of 2.145ppm (1 x LC95 toward An. maculatus larvae until 50 % survive the same time (7.35 days as B. thuringiensis H-14 (8.14 days dosages of 10.724 ppm (5 x LC95. B. thuringiensis H-14 local strain dosages of 21.448 ppm (10 x LC95 toward An. maculatus larvae until 50 % survive longer time (16.21 days than B. thuringiensis H-14 local strain 1 x LC95 and 5 x LC95 The B. thuringiensis H-14 local strain is effective for controlling mosquitoes larvae

  3. Seleção e caracterização de estirpes de Bacillus thuringiensis eficientes contra a Diatraea saccharalis (Lepidoptera: Crambidae Selection and characterization of Bacillus thuringiensis efficient strains against Diatraea saccharalis (Lepidoptera: Crambidae

    Directory of Open Access Journals (Sweden)

    Cristina Lima de Macedo

    2012-12-01

    Full Text Available O objetivo deste trabalho foi selecionar e caracterizar estirpes nativas de Bacillus thuringiensis tóxicas a Diatraea saccharalis (Lepidoptera: Crambidae. Cento e seis estirpes pertencentes ao Banco de Bactérias de Invertebrados, da Embrapa Recursos Genéticos e Biotecnologia, foram testadas quanto à toxicidade a D. saccharalis, e, as mais tóxicas, caracterizadas por métodos bioquímicos e moleculares. Das 106 estirpes testadas, 16 causaram 100% de mortalidade em 24 horas. As três estirpes mais tóxicas apresentaram concentração letal média entre 8 e 43 ng cm-2. O perfil proteico das 16 estirpes mostrou a presença de proteínas de 130 e 65 kDa, e a caracterização molecular mostrou a presença dos genes tipo cry1 e cry2: cry1Aa, cry1Ab, cry1Ac e cry2Aa. A concentração letal média de esporos e cristais obtidos a partir de estirpes recombinantes, que expressavam individualmente os genes cry1Aa, cry1Ab, cry1Ac e cry2Aa, variou entre 222 e 610 ng cm-2, valores muito superiores aos das estirpes nativas mais tóxicas, que apresentavam possibilidade de expressão simultânea desses genes. Este resultado é indicativo de que há sinergia entre as toxinas. Há interação entre as toxinas de B. thuringiensis e seus receptores na broca-do-colmo da cana-de-açúcar.The objective of this work was to select and characterize native strains of Bacillus thuringiensis toxic to Diatraea saccharalis (Lepidoptera: Crambidae. A hundred-and-six strains, belonging to the bank of invertebrate bacteria (Brazil, of Embrapa Genetic Resources and Biotechnology, were tested as to their toxicity to D. saccharalis, and the most toxic ones were characterized by biochemical and molecular methods. Out of the 106 tested strains, 16 caused 100% mortality within 24 hours. The three most toxic strains showed median lethal concentrations between 8 and 43 ng cm-2. The protein profile of the 16 strains showed the presence of 130 and 65 kDa proteins, and the molecular

  4. Impacts of elevated CO2 on exogenous Bacillus thuringiensis toxins and transgene expression in transgenic rice under different levels of nitrogen

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    Jiang, Shoulin; Lu, Yongqing; Dai, Yang; Qian, Lei; Muhammad, Adnan Bodlah; Li, Teng; Wan, Guijun; Parajulee, Megha N.; Chen, Fajun

    2017-01-01

    Recent studies have highlighted great challenges of transgene silencing for transgenic plants facing climate change. In order to understand the impacts of elevated CO2 on exogenous Bacillus thuringiensis (Bt) toxins and transgene expression in transgenic rice under different levels of N-fertilizer supply, we investigated the biomass, exogenous Bt toxins, Bt-transgene expression and methylation status in Bt rice exposed to two levels of CO2 concentrations and nitrogen (N) supply (1/8, 1/4, 1/2...

  5. Field trials with tank mixtures of Bacillus thuringiensis subsp. israelensis and Bacillus sphaericus formulations against Culex pipiens larvae in septic tanks in Antalya, Turkey.

    Science.gov (United States)

    Cetin, Huseyin; Dechant, Peter; Yanikoglu, Atila

    2007-06-01

    Efficacy of tank mixtures of commercial Bacillus thuringiensis subsp. israelensis (Bti) and Bacillus sphaericus (Bs) water-dispersable granule (WDG) formulations was evaluated in septic tanks, against Culex pipiens L. (Diptera: Culicidae) larvae. VectoLex WDG (Bs) + VectoBac WDG (Bti) were evaluated at various ratios from 488 g/ha VectoLex + 250 g/ha VectoBac up to 3,000 g/ha of each. All applications and ratios delivering VectoLex WDG at a rate equal to or greater than 988 g/ha provided more than 90% control for 28 days after treatment. The lowest dose provided this level of control for at least 7 days after treatment, with greater than 80% control after 2 wk. These results suggest that a retreatment interval of 2 wk is recommended with the lowest dose and retreatment intervals of 4 or more wk are recommended with the doses equal to or higher than 988 g/ha VectoLex + 250 g/ha VectoBac.

  6. Analysis of the interaction between Bacillus coagulans and Bacillus thuringiensis S-layers and calcium ions by XRD, light microscopy, and FTIR.

    Science.gov (United States)

    Babolmorad, Ghazal; Emtiazi, Giti; Emamzadeh, Rahman

    2014-05-01

    S-layer is a self-assemble regularly crystalline surface that covers major cell wall component of many bacteria and archaea and exhibits a high metal-binding capacity. We have studied the effect of the calcium ions and type of solid support (glass or mica) on the structure of the S-layers from Bacillus coagulans HN-68 and Bacillus thuringiensis MH14 upon simple methods based on light microscopy and AFM. Furthermore, the Fourier transform infrared spectroscopy (FTIR) study is indicated that the calcium-S-layer interaction occurred mainly through the carboxylate groups of the side chains of aspartic acid (Asp) and glutamic acid (Glu) and nitrogen atoms of Lys, Asn, and histidine (His) amino acids and N-H groups of the peptide backbone. Studied FTIR revealed that inner faces of S-layer are mainly negative, and outer faces of S-layer are mainly positive. Probably, calcium ions with positive charges bound to the carboxyl groups of Glu and Asp. Accordingly, calcium ions are anchored in the space between the inner faces of S-layer with negative charge and the surface of mica with negative charge. This leads to regular arrangement of the S-layer subunits.

  7. Toxicity and residual control of Plutella xylostella L. (Lepidoptera: Plutellidae with Bacillus thuringiensis Berliner and insecticides Toxicidade e controle residual de Plutella xylostella L. (Lepidoptera: Plutellidae com Bacillus thuringiensis Berliner e inseticidas

    Directory of Open Access Journals (Sweden)

    Carla Pedroso de Moraes

    2012-08-01

    Full Text Available Plutella xylostella L. is the most important worldwide pest of cruciferous plants and indiscriminate use of insecticides has led to the resistance of the species to different groups. This research was conducted to compare the toxicity and persistence of two strains of Bacillus thuringiensis to P. xylostella larvae. Concentrations between 125 and 500g 100L-1 of water of the commercial products were evaluated and compared to the insect growth inhibitor diflubenzuron and to the neurotoxic pyrethroid deltamethrin. The efficacy of the insecticides was compared between treated plants kept indoor greenhouse and outdoor. Third instar larvae were more susceptible to B. thuringiensis than first instar ones. Agree and Dipel showed similar control rates of third instars until 10 days after treatment, but on the 15th day, Agree was significantly more efficient than Dipel. Both B. thuringiensis formulations showed a reduction in mortality after 10 days when the treated plants were exposed to natural weather conditions in comparison to the same treatments kept inside greenhouse. Dimilin (100g 100L-1 of water and deltamethrin (30ml of commercial product 100L-1 of water were not efficient to control third instar larvae of P. xylostella. This inefficiency cannot be attributed to a resistant population of P. xylostella since the larval population used in the experiments was not subjected to insecticide pressure, once the crop was organically cultivated all year round. The results showed that both formulations of B. thuringiensis are sound alternatives for the control of the diamondback moth in organically conducted cruciferous crops, considering the high residual control provided under subtropical weather conditions.Larvas de Plutella xylostella L. são as principais pragas de crucíferas cultivadas e o uso excessivo e indiscriminado de inseticidas tem levado a resistência da espécie para diferentes grupos de inseticidas. Este trabalho foi conduzido para

  8. Lethality of chlorine, chlorine dioxide, and a commercial fruit and vegetable sanitizer to vegetative cells and spores of Bacillus cereus and spores of Bacillus thuringiensis.

    Science.gov (United States)

    Beuchat, Larry R; Pettigrew, Charles A; Tremblay, Mario E; Roselle, Brian J; Scouten, Alan J

    2004-08-01

    Chlorine, ClO2, and a commercial raw fruit and vegetable sanitizer were evaluated for their effectiveness in killing vegetative cells and spores of Bacillus cereus and spores of Bacillus thuringiensis. The ultimate goal was to use one or both species as a potential surrogate(s) for Bacillus anthracis in studies that focus on determining the efficacy of sanitizers in killing the pathogen on food contact surfaces and foods. Treatment with alkaline (pH 10.5 to 11.0) ClO2 (200 microg/ml) produced by electrochemical technologies reduced populations of a five-strain mixture of vegetative cells and a five-strain mixture of spores of B. cereus by more than 5.4 and more than 6.4 log CFU/ml respectively, within 5 min. This finding compares with respective reductions of 4.5 and 1.8 log CFU/ml resulting from treatment with 200 microg/ml of chlorine. Treatment with a 1.5% acidified (pH 3.0) solution of Fit powder product was less effective, causing 2.5- and 0.4-log CFU/ml reductions in the number of B. cereus cells and spores, respectively. Treatment with alkaline ClO2 (85 microg/ml), acidified (pH 3.4) ClO2 (85 microg/ml), and a mixture of ClO2 (85 microg/ml) and Fit powder product (0.5%) (pH 3.5) caused reductions in vegetative cell/spore populations of more than 5.3/5.6, 5.3/5.7, and 5.3/6.0 log CFU/ml, respectively. Treatment of B. cereus and B. thuringiensis spores in a medium (3.4 mg/ml of organic and inorganic solids) in which cells had grown and produced spores with an equal volume of alkaline (pH 12.1) ClO2 (400 microg/ml) for 30 min reduced populations by 4.6 and 5.2 log CFU/ml, respectively, indicating high lethality in the presence of materials other than spores that would potentially react with and neutralize the sporicidal activity of ClO2.

  9. Caracterización de una delta endotoxina mutante de Bacillus thuringiensis con estabilidad y toxicidad aumentadas

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    A. Hussain Syed Rehan

    2012-06-01

    Full Text Available Título en ingles: Characterization of a Mutant Bacillus thuringiensis d-endotoxin With Enhanced Stability and Toxicity SummaryThe centrally located a-helix 5 of Bacillus thuringiensis d-endotoxins is critical for insect toxicity through ion-channel formation. We analyzed the role of the highly conserved residue Histidine 168 (H168 using molecular biology, electrophysiology and biophysical techniques. Toxin H168R was ~3-fold more toxic than the wild type (wt protein whereas H168Q was 3 times less toxic against Manduca sexta. Spectroscopic analysis revealed that the H168Q and H168R mutations did not produce gross structural alterations, and that H168R (Tm= 59 °C was more stable than H168Q (Tm= 57.5 °C or than the wt (Tm= 56 °C toxins. These three toxins had similar binding affinities for larval midgut vesicles (Kcom suggesting that the differences in toxicity did not result from changes in initial receptor binding. Dissociation binding assays and voltage clamping analysis suggest that the reduced toxicity of the H168Q toxin may result from reduced insertion and/or ion channel formation. In contrast, the H168R toxin had a greater inhibition of the short circuit current than the wt toxin and an increased rate of irreversible binding (kobs, consistent with its lower LC50 value.  Molecular modeling analysis suggested that both the H168Q and H168R toxins could form additional hydrogen bonds that could account for their greater thermal stability. In addition to this, it is likely that H168R has an extra positive charge exposed to the surface which could increase its rate of insertion into susceptible membranes. Key words: a-helix 5; Circular dichroism; molecular modeling; site-directed mutagenesis;  thermal stability; Bacillus thuringiensis ResumenLa a-Hélice 5 del domino I de las d-endotoxinas de Bacillus thuringiensis, es crítica para la toxicidad de las toxinas contra insectos al participar en la formación de canales iónicos. La

  10. Activity of vegetative insecticidal proteins Vip3Aa58 and Vip3Aa59 of Bacillus thuringiensis against lepidopteran pests.

    Science.gov (United States)

    Baranek, Jakub; Kaznowski, Adam; Konecka, Edyta; Naimov, Samir

    2015-09-01

    Vegetative insecticidal proteins (Vips) secreted by some isolates of Bacillus thuringiensis show activity against insects and are regarded as insecticides against pests. A number of B. thuringiensis strains harbouring vip3A genes were isolated from different sources and identified by using a PCR based approach. The isolates with the highest insecticidal activity were indicated in screening tests, and their vip genes were cloned and sequenced. The analysis revealed two polymorphic Vip protein forms, which were classified as Vip3Aa58 and Vip3Aa59. After expression of the vip genes, the proteins were isolated and characterized. The activity of both toxins was estimated against economically important lepidopteran pests of woodlands (Dendrolimus pini), orchards (Cydia pomonella) and field crops (Spodoptera exigua). Vip3Aa58 and Vip3Aa59 were highly toxic and their potency surpassed those of many Cry proteins used in commercial bioinsecticides. Vip3Aa59 revealed similar larvicidal activity as Vip3Aa58 against S. exigua and C. pomonella. Despite 98% similarity of amino acid sequences of both proteins, Vip3Aa59 was significantly more active against D. pini. Additionally the effect of proteolytic activation of Vip58Aa and Vip3Aa59 on toxicity of D. pini and S. exigua was studied. Both Vip3Aa proteins did not show any activity against Tenebrio molitor (Coleoptera) larvae. The results suggest that the Vip3Aa58 and Vip3Aa59 toxins might be useful for controlling populations of insect pests of crops and forests. Copyright © 2015 Elsevier Inc. All rights reserved.

  11. Eco-friendly microbial route to synthesize cobalt nanoparticles using Bacillus thuringiensis against malaria and dengue vectors.

    Science.gov (United States)

    Marimuthu, Sampath; Rahuman, Abdul Abdul; Kirthi, Arivarasan Vishnu; Santhoshkumar, Thirunavukkarasu; Jayaseelan, Chidambaram; Rajakumar, Govindasamy

    2013-12-01

    The developments of resistance and persistence to chemical insecticides and concerns about the non-target effects have prompted the development of eco-friendly mosquito control agents. The aim of this study was to investigate the larvicidal activities of synthesized cobalt nanoparticles (Co NPs) using bio control agent, Bacillus thuringiensis against malaria vector, Anopheles subpictus and dengue vector, Aedes aegypti (Diptera: Culicidae). The synthesized Co NPs were characterized by X-ray diffraction (XRD), Fourier transform infrared (FTIR), Field-emission scanning electron microscopy (FESEM) with energy dispersive X-ray spectroscopy, and Transmission electron microscopy (TEM). XRD analysis showed three distinct diffraction peaks at 27.03°, 31.00°, and 45.58° indexed to the planes 102, 122, and 024, respectively on the face-centered cubic cobalt acetate with an average size of 85.3 nm. FTIR spectra implicated role of the peak at 3,436 cm(-1) for O-H hydroxyl group, 2924 cm(-1) for methylene C-H stretch in the formation of Co NPs. FESEM analysis showed the topological and morphological appearance of NPs which were found to be spherical and oval in shape. TEM analysis showed polydispersed and clustered NPs with an average size of 84.81 nm. The maximum larvicidal mortality was observed in the cobalt acetate solution, B. thuringiensis formulation, and synthesized Co NPs against fourth instar larvae of A. subpictus and A. aegypti with LC50 values of 29.16, 8.12, 3.59 mg/L; 34.61, 6.94, and 2.87 mg/L; r (2) values of 0.986, 0.933, 0.942; 0.962, 0.957, and 0.922, respectively.

  12. Interação entre inimigos naturais: Trichogramma e Bacillus thuringiensis no controle biológico de pragas agrícolas = Interaction between natural enemies: Trichogramma and Bacillus thuringiensis in pest control

    Directory of Open Access Journals (Sweden)

    Ricardo Antonio Polanczyk

    2006-04-01

    Full Text Available O objetivo deste trabalho foi avaliar os efeitos adversos de Bacillus thuringiensis (Bt nos parasitóides de ovos Trichogramma pratissolii e Trichogramma pretiosum. Suspensões de 6 isolados de Bt: E-3, E-10, E-15, E-16, E-19, E-20 e Bt kurstaki foram misturadas com mel (1:1, como fonte de alimento, e cartelas com ovos de Anagasta kuehniella foram fornecidas para o parasitismo. O experimento foi mantido em BOD a 25 + 1ºC, umidade relativa de 70+ 10% e fotofase de 14 horas. Foram analisados a longevidade, os parasitismos diário, acumulado e total. Não houve influência dos tratamentos sobre a longevidade e todos ostratamentos em T. pratissolii e E-3, E-10, E-16 e E- 19 em T. pretiosum diminuíram o tempo necessário para os parasitóides atingirem 80% de parasitismo. Apesar dessa aceleração do parasitismo, o Bt não influenciou o total de ovos parasitados, mostrando-se como tática de controle que pode ser usada com Trichogramma em programas de MIP.The goal of this work was to detect the possible side-effectsprovoked by Bacillus thuringiensis (Bt strains on eggs parasitoids Thichogramma pratissolii and Trichogramma pretiosum. Six strains of Bt: E-3, E-10, E-15, E-16, E-19, and E-20 and Bt kurstaki were assayed in honey drop to stimulate the feeding of the parasitoids in each treatment and offered simultaneously displays with Anagasta kuehniella eggs. The experiment was maintained in a climatized chamber at 25+ 1ºC, RH 70+ 10%, and photophase of 14 hours. The longevity, the daily, the accumulated, and the total parasitism were observed. No treatments affected the longevity of both parasitoids. All treatments in T. pratissolii and E-3, E-10, E-16, and E-19 in T. pretiosum decreased the time necessary to reach 80% of parasitism. The lack of deleterious effects of Bt on these parasitoids suggests that these natural enemies can be used together in IPM programs.

  13. PENGEMBANGBIAKAN Bacillus thuringiensis H-14 GALUR LOKAL PADA BERBAGAI MACAM PH MEDIA AIR KELAPA DAN TOKSISITASNYA TERHADAP JENTIK NYAMUK VEKTOR Aedes aegypti DAN Anopheles aconitus

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    Blodine Cb. P

    2012-12-01

    Full Text Available The culture of bioinsecticide containing active Bacillus thuringiensis H-14 local strain on various kinds on coconut water pH and its toxicity against Aedes aegypti and Anopheles aconitus were carried out in the laboratory on Institute of Vector and Reservoir Control Research and Development Salatiga. The objectives of this study were : To determine the optimum pH from various kinds of coconut water pH forculturing of B. thuringiensis H-14 local strain. This study was using 20 coconuts with 6-8 months age coconut on average weight around 1 kg that contained water approximately 400-500 ml/coconut were taken from Kunir Rejo village, Butuh regency, Purworejo district. Fifteen out of 20 coconuts were used to culture cells and spores of B. thuringiensis H-14 local strain and 5 coconuts were used to analyze the contain of coconut water in the Institute of Health Laboratory Semarang. The results showed, that B. thuringiensis H-14 local strain can culture at ranges from pH 7 to pH 8.5 with the pH7 as the optimum pH. Total Viable Cell (TVC and a Total Viable Spore Count (TVSC were 3,5 x 10 degree 10 cells /ml and 3, 3 x 10 degree 10 spores/ml respectively. The Lethal Concentration (LC50 = 10.56 ppm and LC95 = 22. 13 ppm against Ae. agypti larvae and LC50 = 5 ppm and LC95 = 11 ppm against An.aconitus larvae. The result showed the analyze test contain of coconut water were 1.92 % carbohydrate, 0.01 % fat, 0.06 % protein and reduced glucose 1.87 %. Coconut water can be used asan alternative local media to culture B. thuringiensis H-14 local strain. Key words : B. thuringiensis H-14, pH, coconut water media.

  14. Diversity of Bacillus thuringiensis strains isolated from coffee plantations infested with the coffee berry borer Hypothenemus hampei

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    Glen Arrieta

    2004-09-01

    Full Text Available The coffee berry borer Hypothenemus hampei Ferrari (Coleoptera: Scolytidae was first reported infecting Costa Rican coffee plantations in the year 2000. Due to the impact that this plague has in the economy of the country, we were interested in seeking new alternatives for the biological control of H. hampei, based on the entomopathogenic bacteria Bacillus thuringiensis. Atotal of 202 B. thuringiensis isolates obtained from Costa Rican coffee plantations infested with H. hampei were analyzed through crystal morphology of the crystal inclusions and SDS-PAGE of d-endotoxins, while 105 strains were further evaluated by PCR for the presence cry, cyt and vip genes. Most of the Bt strains showed diverse crystal morphologies: pleomorphic (35%, oval (37%, bipyramidal (3%, bipyramidal and oval (12%, bipyramidal, oval and pleomorphic (10% and bipyramidal, oval and cubic (3%. The SDS-PAGE analyses of the crystal preparations showed five strains with delta -endotoxin from 20 to 40 kDa, six from 40 to 50 kDa, seven from 50 to 60 kDa, 19 from 60 to 70 kDa, 29 from 70 to 100 kDa and 39 from 100-145 kDa. PCR analyses demonstrated that the collection showed diverse cry genes profiles having several genes per strain: 78 strains contained the vip3 gene, 82 the cry2 gene, 45 the cry1 and 29 strains harbored cry3-cry7 genes. A total of 13 strains did not amplified with any of the cry primers used: cry1, cry2, cry37, cry5, cry11, cry12 and cry14. Forty-three different genetic profiles were found, mainly due to the combination of cry1A genes with other cry and vip genes. The genetic characterization of the collection provides opportunities for the selection of strains to be tested in bioassays against H. hampei and other insect pests of agricultural importance. Rev. Biol. Trop. 52(3: 757-764. Epub 2004 Dic 15.En el año 2000 se reportó por primera vez la principal plaga del cafeto, conocida como broca (Hypothenemus hampei Ferrari (Coleoptera: Scolitidae en

  15. Isolation and toxicity test of Bacillus thuringiensis from Sekayu region soil, South Sumatra on Spodopteralitura

    Science.gov (United States)

    Afriani, S. R.; Pujiastuti, Y.; Irsan, C.; Damiri, N.; Nugraha, S.; Sembiring, E. R.

    2018-01-01

    This study aimed to obtain bacterial isolates B. thuringiensis potential as a biological control against pests Spodoptera litura. The research was conducted at the Laboratory of Pest and Disease Department, Agricultural Faculty of Sriwijaya University, Campus Inderalaya Ogan Ilir, South Sumatera, from March to June 2017. The study was conducted with survey method and laboratory trial. The results showed that of the 50 soil samples from three villages selected through morphological observation, reaction staining, KOH test, catalase test, producing 13 bacterial isolates. Screening of the 13th toxicity of the isolates suspected B. thuringiensis against S. litura larvae was investigated. Based on the toxicity screening test the following facts were obtained: five isolates ie KJ2M2, KJ3E1, KJ3JB1, KJ3D3 and KJ3D5 were lower toxicity than Dipel, two isolates ie KJ3K4 and KJ3D3 which had the same toxicity to Dipel. Five isolates the KJ3E3, KJ3BW5, KJ3JB5, KJ3D1 and LC2, LC3 known to have effectiveness until the seventh day reached 40%. There was one isolate that is KJ3BW5 which was more effective compared to Dipel as comparison.

  16. Evaluation of alternative Plutella xylostella control by two Isaria fumosorosea conidial formulations - oil-based formulation and wettable powder - combined with Bacillus thuringiensis.

    Science.gov (United States)

    Nian, Xiao-Ge; He, Yu-Rong; Lu, Li-Hua; Zhao, Rui

    2015-12-01

    Entomopathogenic fungi are potential candidates for controlling the diamondback moth Plutella xylostella (L.) (Lepidoptera: Plutellidae). The control efficacy of two Isaria fumosorosea conidial formulations - wettable powder and oil-based formulation - combined with Bacillus thuringiensis against P. xylostella was tested. In the laboratory, the combined application of two pathogens increased larval mortality either in an additive or a synergistic way. P. xylostella larvae treated with oil-based formulation died sooner than larvae infected with wettable powder. For pot and field experiments, each formulation was applied alone or combined with B. thuringiensis 668 µg mL(-1) , and then larval mortality, pupation rate, adult emergence rate, female longevity and fecundity were recorded. In pot experiments there was no evidence of any antagonistic effects between the two pathogens. Combined application of B. thuringiensis and a high concentration of the two I. fumosorosea formulations resulted in higher mortality (84.4 and 86.2%) with minimum pupation (15.6 and 11.9%) and adult emergence rates (8.7 and 7.0%). Female longevity and fecundity were significantly reduced by the two formulations at high concentration compared with the control. Similar results were also observed in field experiments. The combined application of I. fumosorosea and B. thuringiensis is a promising alternative strategy for P. xylostella control. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

  17. Slow release formulations of Bacillus thuringiensis israelensis (AM 65-52 and spinosyns: effectiveness against the West Nile vector Culex pipiens in Saudi Arabia

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    Alaa Sulaiman Alsobhi

    2016-07-01

    Full Text Available Objective: To investigate the effectiveness of slow release formulations of Bacillus thuringiensis israelensis (AM 65-52 (B. thuringiensis israelensis and spinosyns against the West Nile vector Culex pipiens (Cx. pipiens in Saudi Arabia. Methods: We tested slow release insecticide formulations of Natular DT, Tap 60 and VectoBac granule against II instars of Cx. pipiens larvae in 50 L laboratory arenas. Results: Slow release formulations of B. thuringiensis israelensis and spinosyns gave continuous control against Cx. pipiens for several weeks. Natular DT was more effective over Tap 60 and VectoBac granule of about 1.3 and 5.8 times, respectively. Variations in the durations of effective control among the tested slow release formulations may reflect differences in their active ingredients and the mode of action. Conclusions: Our results highlighted the effectiveness of B. thuringiensis israelensis and spinosyns against an important West Nile vector, providing baseline data to develop ecofriendly mosquito control programs in Saudi Arabia.

  18. Expresión de la toxina Cry11Aa de Bacillus thuringiensis serovar. israelensis en Asticcacaulis excentricus, para el control de larvas acuáticas de dípteros de la familia Culicidae, vectores de enfermedades Expression of Bacillus thuringiensis serovar. israelensis toxins in Asticcacaulis excentricus to control dipteran larvae of vectors of diseases

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    Orduz Sergio

    2004-07-01

    Full Text Available Los genes cry de Bacillus thuringiensis codifican para un diverso grupo de proteínas formadoras de cristales que exhiben actividad insecticida contra larvas de dípteros, lepidópteros y coleópteros, entre otros. La efectividad de los insecticidas basados en formulaciones de proteínas de B. thuringiensis puede ser mejorada usando bacterias prostecadas acuáticas como portadoras alternativas de los genes cry, ya que no se sedimentan rápidamente; las proteínas expresadas en el citoplasma están protegidas de los rayos ultra violeta y, lo más importante, las larvas de los mosquitos se alimentan de ellas. Una cepa de referencia de Asticcacaulis excentricus fue transformada con el plásmido pSOD3, el cual contiene el gen que codifica para la proteína Cry1 1 Aa de B. thuringiensis serovar. israelensis. La expresión de la proteína recombinante fue evaluada por electroforesis de proteínas y por Western blot. El Western blot revelado con un anticuerpo policlonal anti-Cry1 1 Aa mostró una banda de 72 kDa correspondiente a la proteína Cry 11 Aa. La toxicidad de las cepas de A. excentricus transformadas fue evaluada en bioensayos con larvas de primer estadío del mosquito Culex quinquefasciatus. Se alcanzó un promedio de mortalidad del 50% de las larvas de primer instar a concentraciones de 23 ng/mL de la toxina. Otros bioensayos indican que A. excentricus recombinante es tóxica para larvas de primer instar de las especies Aedes aegypti y Anopheles albimanus. Los ensayos de flotabilidad indican que A. excentricus no sedimenta hasta pasados 7 días, mientras que B. thuringiensis serovar. israelensis sedimenta al cabo de algunas horas. Palabras clave: Asticcacaulis excentricus, Bacillus thuringiensis, bacterias prostecadas, dengue, malaria.Bacillus thuringiensis cry genes encode for a diverse group of crystal-forming proteins that exhibit insecticidal activity towards dipteran, lepidopteran and coleopteran larvae. The effectiveness of

  19. Transgenic loblolly pine (Pinus taeda L.) plants expressing a modified delta-endotoxin gene of Bacillus thuringiensis with enhanced resistance to Dendrolimus punctatus Walker and Crypyothelea formosicola Staud.

    Science.gov (United States)

    Tang, Wei; Tian, Yingchuan

    2003-02-01

    A synthetic version of the CRY1Ac gene of Bacillus thuringiensis has been used for the transformation of loblolly pine (Pinus taeda L.) using particle bombardment. Mature zygotic embryos were used to be bombarded and to generate organogenic callus and transgenic regenerated plants. Expression vector pB48.215 DNA contained a synthetic Bacillus thuringiensis (B.t.) CRY1Ac coding sequence flanked by the double cauliflower mosaic virus (CaMV) 35S promoter and nopaline synthase (NOS) terminator sequences, and the neomycin phosphotransferase II (NPTII) gene controlled by the promoter of the nopaline synthase gene was introduced into loblolly pine tissues by particle bombardment. The transformed tissues were proliferated and selected on media with kanamycin. Shoot regeneration was induced from the kanamycin-resistant calli, and transgenic plantlets were then produced. More than 60 transformed plants from independent transformation events were obtained for each loblolly pine genotype tested. The integration and expression of the introduced genes in the transgenic loblolly pine plants was confirmed by polymerase chain reactions (PCR) analysis, by Southern hybridization, by Northern blot analysis, and by Western blot analysis. Effective resistance of transgenic plants against Dendrolimus punctatus Walker and Crypyothelea formosicola Staud was verified in feeding bioassays with the insects. The transgenic plants recovered could represent a good opportunity to analyse the impact of genetic engineering of pine for sustainable resistance to pests using a B. thuringiensis insecticidal protein. This protocol enabled the routine transformation of loblolly pine plants that were previously difficult to transform.

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

    International Nuclear Information System (INIS)

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

    2015-01-01

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

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

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    Sudhanshu Kashyap

    2013-01-01

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

  2. Impact of routine Bacillus thuringiensis israelensis (Bti) treatment on the availability of flying insects as prey for aerial feeding predators.

    Science.gov (United States)

    Timmermann, Ute; Becker, Norbert

    2017-12-01

    Since 1980, mosquito breeding habitats in the Upper Rhine Valley were routinely treated with Bacillus thuringiensis var. israelensis (Bti). Bti is considered to significantly reduce the number of mosquitoes, and - especially when used in higher dosages - to be toxic to other Nematocera species, e.g. Chironomidae, which could be food sources for aerial feeding predators. To investigate direct and indirect effects of routine Bti treatment on food sources for aerial feeding predators, the availability of flying insects in treated and untreated areas was compared. A car trap was used for insect collection, which allowed their exact spatiotemporal assignment. The statistical analysis revealed that insect taxa abundance was influenced differently by the factors season, temperature and time of day. Nematocera (Diptera) were the most frequently collected insects in all areas. Chironomidae were the predominant aquatic Nematocera. The comparison of treated and untreated sites did not show significant differences that would indicate any direct or indirect effect of routine Bti treatment on the availability of flying insects. Additional to food availability, food selection must be considered when investigating food resources for aerial feeding predators. In this study, food selection of Delichon urbicum (House Martin) as an example was investigated with the help of neck ring samples. The preferred prey of the investigated D. urbicum colony consisted of diurnal insects with terrestrial larvae (Aphidina, Brachycera, Coleoptera). Chironomidae were consumed, but not preferred.

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

    Science.gov (United States)

    Oppert, Brenda; Martynov, Alexander G; Elpidina, Elena N

    2012-09-01

    The yellow mealworm, Tenebrio molitor, is a pest of stored grain products and is sensitive to the Bacillus thuringiensis (Bt) Cry3Aa toxin. As digestive peptidases are a determining factor in Cry toxicity and resistance, we evaluated the expression of peptidase transcripts in the midgut of T. molitor larvae fed either a control or Cry3Aa protoxin diet for 24 h (RNA-Seq), or in larvae exposed to the protoxin for 6, 12, or 24 h (microarrays). Cysteine peptidase transcripts (9) were similar to cathepsins B, L, and K, and their expression did not vary more than 2.5-fold in control and Cry3Aa-treated larvae. Serine peptidase transcripts (48) included trypsin, chymotrypsin and chymotrypsin-like, elastase 1-like, and unclassified serine peptidases, as well as homologs lacking functional amino acids. Highly expressed trypsin and chymotrypsin transcripts were severely repressed, and most serine peptidase transcripts were expressed 2- to 15-fold lower in Cry3Aa-treated larvae. Many serine peptidase and homolog transcripts were found only in control larvae. However, expression of a few serine peptidase transcripts was increased or found only in Cry3Aa-treated larvae. Therefore, Bt intoxication significantly impacted the expression of serine peptidases, potentially important in protoxin processing, while the insect maintained the production of critical digestive cysteine peptidases. Published by Elsevier Inc.

  4. Sodium Solute Symporter and Cadherin Proteins Act as Bacillus thuringiensis Cry3Ba Toxin Functional Receptors in Tribolium castaneum*

    Science.gov (United States)

    Contreras, Estefanía; Schoppmeier, Michael; Real, M. Dolores; Rausell, Carolina

    2013-01-01

    Understanding how Bacillus thuringiensis (Bt) toxins interact with proteins in the midgut of susceptible coleopteran insects is crucial to fully explain the molecular bases of Bt specificity and insecticidal action. In this work, aminopeptidase N (TcAPN-I), E-cadherin (TcCad1), and sodium solute symporter (TcSSS) have been identified by ligand blot as putative Cry3Ba toxin-binding proteins in Tribolium castaneum (Tc) larvae. RNA interference knockdown of TcCad1 or TcSSS proteins resulted in decreased susceptibility to Cry3Ba toxin, demonstrating the Cry toxin receptor functionality for these proteins. In contrast, TcAPN-I silencing had no effect on Cry3Ba larval toxicity, suggesting that this protein is not relevant in the Cry3Ba toxin mode of action in Tc. Remarkable features of TcSSS protein were the presence of cadherin repeats in its amino acid sequence and that a TcSSS peptide fragment containing a sequence homologous to a binding epitope found in Manduca sexta and Tenebrio molitor Bt cadherin functional receptors enhanced Cry3Ba toxicity. This is the first time that the involvement of a sodium solute symporter protein as a Bt functional receptor has been demonstrated. The role of this novel receptor in Bt toxicity against coleopteran insects together with the lack of receptor functionality of aminopeptidase N proteins might account for some of the differences in toxin specificity between Lepidoptera and Coleoptera insect orders. PMID:23645668

  5. Isolation and characterization of a novel native Bacillus thuringiensis strain BRC-HZM2 capable of degrading chlorpyrifos.

    Science.gov (United States)

    Wu, Songqing; Peng, Yan; Huang, Zhangmin; Huang, Zhipeng; Xu, Lei; Ivan, Gelbič; Guan, Xiong; Zhang, Lingling; Zou, Shuangquan

    2015-03-01

    Studies were carried out to isolate chlorpyrifos degrading Bacillus thuringiensis (Bt) strains from chlorpyrifos-contaminated samples. Six Bt strains (isolation rate 2.7%) were isolated by modified sodium acetate antibiotic heat treatment, and one novel strain (BRC-HZM2) was selected for further analysis. Phenotype and phylogeny analysis of this strain was conducted on the basis of biochemical reactions, antibiotic sensitivity, 16s rRNA genes, plasmid profile, insecticidal crystal protein profiles, and PCR-RFLP for cry and cyt genes. The degradation rate of chlorpyrifos in liquid culture was estimated during 48 h of incubation for the isolate BRC-HZM2. More than 50% of the initial chlorpyrifos concentration degraded within 12 h, 88.9% after 48 h. These results highlight the potential of the Bt strain for biological control and the bioremediation of environments contaminated with chlorpyrifos. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

    Directory of Open Access Journals (Sweden)

    Juan Luis Jurat-Fuentes

    Full Text Available Development of insect resistance is one of the main concerns with the use of transgenic crops expressing Cry toxins from the bacterium Bacillus thuringiensis. Identification of biomarkers would assist in the development of sensitive DNA-based methods to monitor evolution of resistance to Bt toxins in natural populations. We report on the proteomic and genomic detection of reduced levels of midgut membrane-bound alkaline phosphatase (mALP as a common feature in strains of Cry-resistant Heliothis virescens, Helicoverpa armigera and Spodoptera frugiperda when compared to susceptible larvae. Reduced levels of H. virescens mALP protein (HvmALP were detected by two dimensional differential in-gel electrophoresis (2D-DIGE analysis in Cry-resistant compared to susceptible larvae, further supported by alkaline phosphatase activity assays and Western blotting. Through quantitative real-time polymerase chain reaction (qRT-PCR we demonstrate that the reduction in HvmALP protein levels in resistant larvae are the result of reduced transcript amounts. Similar reductions in ALP activity and mALP transcript levels were also detected for a Cry1Ac-resistant strain of H. armigera and field-derived strains of S. frugiperda resistant to Cry1Fa. Considering the unique resistance and cross-resistance phenotypes of the insect strains used in this work, our data suggest that reduced mALP expression should be targeted for development of effective biomarkers for resistance to Cry toxins in lepidopteran pests.

  7. Selection and characterization of resistance to the Vip3Aa20 protein from Bacillus thuringiensis in Spodoptera frugiperda.

    Science.gov (United States)

    Bernardi, Oderlei; Bernardi, Daniel; Horikoshi, Renato J; Okuma, Daniela M; Miraldo, Leonardo L; Fatoretto, Julio; Medeiros, Fernanda Cl; Burd, Tony; Omoto, Celso

    2016-09-01

    Spodoptera frugiperda is one the main target pests of maize events expressing Vip3Aa20 protein from Bacillus thuringiensis (Bt) in Brazil. In this study, we selected a resistant strain of S. frugiperda on Bt maize expressing Vip3Aa20 protein and characterized the inheritance and fitness costs of the resistance. The resistance ratio of the Vip3Aa20-resistant strain of S. frugiperda was >3200-fold. Neonates of the Vip3Aa20-resistant strain were able to survive and emerge as fertile adults on Vip3Aa20 maize, while larvae from susceptible and heterozygous strains did not survive. The inheritance of Vip3Aa20 resistance was autosomal recessive and monogenic. Life history studies to investigate fitness cost revealed an 11% reduction in the survival rate until adult stage and a ∼50% lower reproductive rate of the Vip3Aa20-resistant strain compared with susceptible and heterozygous strains. This is the first characterization of S. frugiperda resistance to Vip3Aa protein. Our results provide useful information for resistance management programs designed to prevent or delay resistance evolution to Vip3Aa proteins in S. frugiperda. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  8. Monitoring resistance to Bacillus thuringiensis subsp. israelensis in the field by performing bioassays with each Cry toxin separately

    Directory of Open Access Journals (Sweden)

    Guillaume Tetreau

    2013-11-01

    Full Text Available Bacillus thuringiensis subsp. israelensis (Bti is increasingly used worldwide for mosquito control and is the only larvicide used in the French Rhône-Alpes region since decades. The artificial selection of mosquitoes with field-persistent Bti collected in breeding sites from this region led to a moderate level of resistance to Bti, but to relatively high levels of resistance to individual Bti Cry toxins. Based on this observation, we developed a bioassay procedure using each Bti Cry toxin separately to detect cryptic Bti-resistance evolving in field mosquito populations. Although no resistance to Bti was detected in none of the three mosquito species tested (Aedes rusticus, Aedes sticticus and Aedes vexans, an increased tolerance to Cry4Aa (3.5-fold and Cry11Aa toxins (8-fold was found in one Ae. sticticus population compared to other populations of the same species, suggesting that resistance to Bti may be arising in this population. This study confirms previous works showing a lack of Bti resistance in field mosquito populations treated for decades with this bioinsecticide. It also provides a first panorama of their susceptibility status to individual Bti Cry toxins. In combination with bioassays with Bti, bioassays with separate Cry toxins allow a more sensitive monitoring of Bti-resistance in the field.

  9. Isolation and purification of Bacillus thuringiensis var. israelensis IМV В-7465 peptidase with specificity toward elastin and collagen

    Directory of Open Access Journals (Sweden)

    N. А. Nidialkova

    2016-06-01

    Full Text Available Peptidase of Bacillus thuringiensis var. israelensis IМV В-7465 was isolated from culture supernatant using consecutive fractionations by an ammonium sulphate (60% saturation, ion-exchange chromatography and gel-filtration on the TSK-gels Toyoperl HW-55 and DEAE 650(M. Specific elastase (442 U∙mg of protein-1 and collagenase (212.7 U∙mg of protein-1 activities of the purified enzyme preparation were 8.0- and 6.1-fold, respectively higher than ones of the culture supernatant. Peptidase yields were 33.5% for elastase activity and 30.1% for collagenase activity. It was established that the enzyme is serine metal-dependent alkaline peptidase with Mr about 37 kDa. Maximal hydrolysis of elastin and collagen occurs at the optimum pH 8.0 and t° – 40 and 50 °С, respectively. The purified preparation has high stability at pH in the range of 7.0 to 10.0 and 40-50 °С.

  10. Mechanism of the Quorum-Quenching Lactonase (AiiA) from Bacillus thuringiensis. 1. Product-Bound Structures

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Dali; Momb, Jessica; Thomas, Pei W.; Moulin, Aaron; Petsko, Gregory A.; Fast, Walter; Ringe, Dagmar (Brandeis); (Texas)

    2008-08-06

    Enzymes capable of hydrolyzing N-acyl-l-homoserine lactones (AHLs) used in some bacterial quorum-sensing pathways are of considerable interest for their ability to block undesirable phenotypes. Most known AHL hydrolases that catalyze ring opening (AHL lactonases) are members of the metallo-{beta}-lactamase enzyme superfamily and rely on a dinuclear zinc site for catalysis and stability. Here we report the three-dimensional structures of three product complexes formed with the AHL lactonase from Bacillus thuringiensis. Structures of the lactonase bound with two different concentrations of the ring-opened product of N-hexanoyl-l-homoserine lactone are determined at 0.95 and 1.4 {angstrom} resolution and exhibit different product configurations. A structure of the ring-opened product of the non-natural N-hexanoyl-l-homocysteine thiolactone at 1.3 {angstrom} resolution is also determined. On the basis of these product-bound structures, a substrate-binding model is presented that differs from previous proposals. Additionally, the proximity of the product to active-site residues and observed changes in protein conformation and metal coordination provide insight into the catalytic mechanism of this quorum-quenching metalloenzyme.

  11. Improved insecticidal toxicity by fusing Cry1Ac of Bacillus thuringiensis with Av3 of Anemonia viridis.

    Science.gov (United States)

    Yan, Fu; Cheng, Xing; Ding, Xuezhi; Yao, Ting; Chen, Hanna; Li, Wenping; Hu, Shengbiao; Yu, Ziquan; Sun, Yunjun; Zhang, Youming; Xia, Liqiu

    2014-05-01

    Av3, a neurotoxin of Anemonia viridis, is toxic to crustaceans and cockroaches but inactive in mammals. In the present study, Av3 was expressed in Escherichia coli Origami B (DE3) and purified by reversed-phase liquid chromatography. The purified Av3 was injected into the hemocoel of Helicoverpa armigera, rendering the worm paralyzed. Then, Av3 was expressed alone or fusion expressed with the Cry1Ac in acrystalliferous strain Cry(-)B of Bacillus thuringiensis. The shape of Cry1Ac was changed by fusion with Av3. The expressed fusion protein, Cry1AcAv3, formed irregular rhombus- or crescent-shaped crystalline inclusions, which is quite different from the shape of original Cry1Ac crystals. The toxicity of Cry1Ac was improved by fused expression. Compared with original Cry1Ac expressed in Cry(-)B, the oral toxicity of Cry1AcAv3 to H. armigera was elevated about 2.6-fold. No toxicity was detected when Av3 was expressed in Cry(-)B alone. The present study confirmed that marine toxins could be used in bio-control and implied that fused expression with other insecticidal proteins could be an efficient way for their application.

  12. Synergistic Effect of Combining Plutella xylostella Granulovirus and Bacillus thuringiensis at Sublethal Dosages on Controlling of Diamondback Moth (Lepidoptera: Plutellidae).

    Science.gov (United States)

    Han, Guangjie; Li, Chuanming; Liu, Qin; Xu, Jian

    2015-10-01

    Plutella xylostella granulovirus (PxGV) and Bacillus thuringiensis (Bt) are both entomo-pathogens to the diamondback moth, Plutella xylostella (L.). The purpose of the present study was to measure the effect of the combination of PxGV and Bt at sublethal dosages on the development and mortality of diamondback moth in a laboratory setting. Bt and PxGV exhibited synergistic effect on diamondback moth larval mortality and effectively controlled diamondback moth populations with low dose combination treatment. The combination of three parts per million Bt and 1.3 × 10(3) occlusion bodies per milliliter of PxGV revealed a higher larval mortality compared with the treatment of Bt or PxGV alone. Combination of Bt and PxGV at sublethal concentrations also increased larval duration, reduced oviposition and decreased adult longevity remarkably, resulting in the lowest population trend index among the treatments. The results suggested that the combination of Bt and PxGV at sublethal dosages might provide a valuable way to improve the control efficacy of diamondback moth compared with treatment of Bt or PxGV alone. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  13. Baseline sensitivity of lepidopteran corn pests in India to Cry1Ab insecticidal protein of Bacillus thuringiensis.

    Science.gov (United States)

    Jalali, Sushil K; Lalitha, Yadavalli; Kamath, Subray P; Mohan, Komarlingam S; Head, Graham P

    2010-08-01

    Genetically engineered corn (Bt corn) expressing Bacillus thuringiensis Berliner insecticidal protein Cry1Ab is a biotechnological option being considered for management of lepidopteran corn pests in India. As a resistance management practice it was essential to determine the sensitivity of multiple populations of the stalk borer Chilo partellus (Swinhoe), pink borer Sesamia inferens (Walker) and the cob borer Helicoverpa armigera (Hübner) to Cry1Ab protein through bioassays. The insect populations were collected during growing seasons of Rabi 2005 (October 2005 to February 2006) and Kharif 2006 (May to September 2006). Multiple populations of the three lepidopteran corn pests were found to be susceptible to Cry1Ab. Median lethal concentrations (LC(50)) ranged between 0.008 and 0.068 microg Cry1Ab mL(-1) diet for 18 populations of C. partellus (across two seasons), between 0.12 and 1.99 microg mL(-1) for seven populations of H. armigera and between 0.46 and 0.56 microg mL(-1) for two populations of S. inferens. Dose-response concentrations for lethality and growth inhibition have been determined to mark baseline sensitivity of multiple populations of key lepidopteran corn pests in India to Cry1Ab protein. These benchmark values will be referenced while monitoring resistance to Cry1Ab should Bt corn hybrids expressing Cry1Ab be approved for commercial cultivation in India. Copyright (c) 2010 Society of Chemical Industry.

  14. Bombyx mori ABC transporter C2 structures responsible for the receptor function of Bacillus thuringiensis Cry1Aa toxin.

    Science.gov (United States)

    Tanaka, Shiho; Endo, Haruka; Adegawa, Satomi; Iizuka, Ami; Imamura, Kazuhiro; Kikuta, Shingo; Sato, Ryoichi

    2017-12-01

    Because Bombyx mori ABC transporter C2 (BmABCC2) has 1000-fold higher potential than B. mori cadherin-like protein as a receptor for Bacillus thuringiensis Cry1Aa toxin (Tanaka et al., 2013), the gate-opening ability of the latent pore under six extracellular loops (ECLs) of BmABCC2 was expected to be the reason for its higher potential (Heckel, 2012). In this study, cell swelling assays in Sf9 cells showed that BmABCC2 mutants lacking substrate-excreting activity retained receptor activity, indicating that the gate-opening activity of BmABCC2 is not responsible for Cry1Aa toxicity. The analysis of 29 BmABCC2 mutants demonstrated that 770 DYWL 773 of ECL 4 comprise a putative binding site to Cry1Aa. This suggests that specific toxicity of Cry1Aa toxin to a restricted range of lepidopteran insects is dependent on conservation and variation in the amino acid residues around 770 DYWL 773 of ECL 4 in the ABCC2. Copyright © 2017 Elsevier Ltd. All rights reserved.

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

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    Jenkins Jeremy L

    2001-10-01

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

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

    Science.gov (United States)

    Jenkins, Jeremy L; Dean, Donald H

    2001-01-01

    Background To better understand the molecular interactions of Bt toxins with non-target insects, we have examined the real-time binding specificity and affinity of Cry1 toxins to native silkworm (Bombyx mori) midgut receptors. Previous studies on B. mori receptors utilized brush border membrane vesicles or purifed receptors in blot-type assays. Results The Bombyx mori (silkworm) aminopeptidase N (APN) and cadherin-like receptors for Bacillus thuringiensis insecticidal Cry1Aa toxin were purified and their real-time binding affinities for Cry toxins were examined by surface plasmon resonance. Cry1Ab and Cry1Ac toxins did not bind to the immobilized native receptors, correlating with their low toxicities. Cry1Aa displayed moderate affinity for B. mori APN (75 nM), and unusually tight binding to the cadherin-like receptor (2.6 nM), which results from slow dissociation rates. The binding of a hybrid toxin (Aa/Aa/Ac) was identical to Cry1Aa. Conclusions These results indicate domain II of Cry1Aa is essential for binding to native B. mori receptors and for toxicity. Moreover, the high-affinity binding of Cry1Aa to native cadherin-like receptor emphasizes the importance of this receptor class for Bt toxin research. PMID:11722800

  17. Characterisation of a radiation-resistant strain of bacillus thuringiensis subsp. Aizawai with improved toxicity to larval plutella xylostella

    International Nuclear Information System (INIS)

    Mahadi, N.M.; Boo, J.M.L.; Jangi, M.S.

    1989-01-01

    A radiation-resistant strain of Bacillus thuringiensis subsp. Aizawai which was previously shown to be more toxic against larval Plutell xylostella was further characterized. Some of the growth characteristics of the mutant strain were quite different from those of the parent strain. In shake flask culture, its lag period was shorter and its cell yield was lower. The growth rate, however, was the same as that of the parent. Electron microscope studies show that the insecticidal parasporal crystals from the mutant strain are significantly bigger than those produced by the parent strain. The average length and width of the crystals were 1.25 and 0.53 um respectively whereas those of the parent were 0.87 and 0.35 um, respectively. The crystals from the mutant strain were also more toxic. The LC 50 was 0.30 ug crystal protein per ml as against 0.66 ug crystal protein per ml for those from the parent strain. Protein profile of the crystals obtained with SDS-PA gel electrophoresis showed that the mutant strain produced an additional polypeptide of 143 KDa polypeptide. The mutant strain also has an additional high molecular weight plasmid. The improved toxicity may have been brought about by a number of factors including an alteration in the regulatory mechanism that control the synthesis of the polypeptides that make up the crystals. (Auth.). 5 figs.; 21 refs.; 2 tabs

  18. Identification and Characterization of Hyphantria cunea Aminopeptidase N as a Binding Protein of Bacillus thuringiensis Cry1Ab35 Toxin

    Directory of Open Access Journals (Sweden)

    Yakun Zhang

    2017-11-01

    Full Text Available The fall webworm, Hyphantria cunea (Drury is a major invasive pest in China. Aminopeptidase N (APN isoforms in lepidopteran larvae midguts are known for their involvement in the mode of action of insecticidal crystal (Cry proteins from Bacillus thuringiensis. In the present work, we identified a putative Cry1Ab toxin-binding protein, an APN isoform designated HcAPN3, in the midgut of H. cunea by ligand blot and mass spectrometry. HcAPN3 was highly expressed throughout all larval developmental stages and was abundant in the midgut and hindgut tissues. HcAPN3 was down-regulated at 6 h, then was up-regulated significantly at 12 h and 24 h after Cry1Ab toxin treatment. We expressed HcAPN3 in insect cells and detected its interaction with Cry1Ab toxin by ligand blot assays. Furthermore, RNA interference (RNAi against HcAPN3 using oral delivery and injection of double-stranded RNA (dsRNA resulted in a 61–66% decrease in transcript level. Down-regulating of the expression of HcAPN3 was closely associated with reduced susceptibility of H. cunea to Cry1Ab. In addition, the HcAPN3E fragment peptide expressed in Escherichia coli enhanced Cry1Ab toxicity against H. cunea larvae. This work represents the first evidence to suggest that an APN in H. cunea is a putative binding protein involved in Cry1Ab susceptibility.

  19. EFEITO DE BACILLUS THURINGIENSIS (BT SOBRE LAGARTAS DE SPODOPTERA FRUGIPERDA E ANTICARSIA GEMMATALIS EM LABORATÓRIO

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    Fabrício Oliveira Fernandes

    2017-02-01

    Full Text Available Anticarsia gemmatalis (lagarta-da-soja e Spodoptera frugiperda (lagarta-do-cartucho são insetos polífagos e desfolhadores, responsáveis por prejuízos em diversas culturas. O objetivo deste trabalho é avaliar a mortalidade desses insetos através do consumo de dieta artificial com a adição de cepas de Bacillus thuringiensis (Bt. Para a realização dos experimentos foram utilizadas 30 lagartas de A. gemmatalis (3º instar e S. frugiperda (1º instar para cada tratamento, as quais foram acondicionadas em tubos de vidro esterilizados contendo 1 cm3 de dieta artificial. Posteriormente foram adicionadas alíquotas de 150µl de Bt var israelensis (Bti, Bt var kurstaki (Btk, Bt var oswaldo cruz (Bto e água destilada para o controle. O tratamento com Btk resultou em maior mortalidade ambas espécies, seguidas por Bto para A. gemmatalis e Bti para S. frugiperda.

  20. Mechanism of the Quorum-Quenching Lactonase (AiiA) from Bacillus thuringiensis. 2. Substrate Modeling and Active Site Mutations

    Energy Technology Data Exchange (ETDEWEB)

    Momb, Jessica; Wang, Canhui; Liu, Dali; Thomas, Pei W.; Petsko, Gregory A.; Guo, Hua; Ringe, Dagmar; Fast, Walter (UNM); (Brandeis); (Texas)

    2008-12-02

    The N-acyl-l-homoserine lactone hydrolases (AHL lactonases) have attracted considerable attention because of their ability to quench AHL-mediated quorum-sensing pathways in Gram-negative bacteria and because of their relation to other enzymes in the metallo-{beta}-lactamase superfamily. To elucidate the detailed catalytic mechanism of AHL lactonase, mutations are made on residues that presumably contribute to substrate binding and catalysis. Steady-state kinetic studies are carried out on both the wild-type and mutant enzymes using a spectrum of substrates. Two mutations, Y194F and D108N, present significant effects on the overall catalysis. On the basis of a high-resolution structural model of the enzyme-product complex, a hybrid quantum mechanical/molecular mechanical method is used to model the substrate binding orientation and to probe the effect of the Y194F mutation. Combining all experimental and computational results, we propose a detailed mechanism for the ring-opening hydrolysis of AHL substrates as catalyzed by the AHL lactonase from Bacillus thuringiensis. Several features of the mechanism that are also found in related enzymes are discussed and may help to define an evolutionary thread that connects the hydrolytic enzymes of this mechanistically diverse superfamily.

  1. Solid-phase extraction of copper, iron and zinc ions on Bacillus thuringiensis israelensis loaded on Dowex optipore V-493

    Energy Technology Data Exchange (ETDEWEB)

    Tuzen, Mustafa; Melek, Esra [Gaziosmanpasa University, Faculty of Science and Arts, Chemistry Department, 60250 Tokat (Turkey); Soylak, Mustafa [Erciyes University, Faculty of Science and Arts, Chemistry Department, 38039 Kayseri (Turkey)], E-mail: msoylak@gmail.com

    2008-11-30

    Bacillus thuringiensis israelensis loaded on Dowex optipore V-493 as new adsorbent for the separation-preconcentration of heavy metal ions has been proposed. The analytical conditions for the quantitative recoveries of copper(II), iron(III) and zinc(II) including pH, amounts of adsorbent, sample volume, etc. were investigated. The influences of alkaline and earth alkaline ions were also reported. The recovery values for the analytes are generally higher than 95%. The preconcentration factor was 37. The limit of detections of the analyte ions (k = 3, N = 21) were 1.14 {mu}g L{sup -1} for copper, 2.01 {mu}g L{sup -1} for iron and 0.14 {mu}g L{sup -1} for zinc. The relative standard deviations of the determinations were found to be lower than 9%. The procedure was validated by analyzing copper, iron and zinc contents in two certified reference materials, NRCC-SLRS-4 Riverine water and NIST SRM 1515 Apple leaves. Agreements between the obtained results and the certified values were achieved. The developed preconcentration method was applied in the flame atomic absorption spectrometric determination of copper, iron and zinc in several samples including a multivitamin-multimineral tablet, dialysis solutions, natural waters and some food samples.

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

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    Helena Porta

    2011-01-01

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

  3. Equilibrium, kinetic and thermodynamic studies on the adsorption of the toxins of Bacillus thuringiensis subsp. kurstaki by clay minerals

    Energy Technology Data Exchange (ETDEWEB)

    Fu Qingling; Deng Yali; Li Huishu; Liu Jie [Key Laboratory of Subtropical Agricultural Resource and Environment, Ministry of Agriculture, Huazhong Agricultural University, Wuhan, 430070 (China); Hu Hongqing, E-mail: hqhu@mail.hzau.edu.cn [Key Laboratory of Subtropical Agricultural Resource and Environment, Ministry of Agriculture, Huazhong Agricultural University, Wuhan, 430070 (China); Chen Shouwen [Key Laboratory of Subtropical Agricultural Resource and Environment, Ministry of Agriculture, Huazhong Agricultural University, Wuhan, 430070 (China); Sa Tongmin [Department of Agricultural Chemistry, College of Agriculture, Chungbuk National University, Cheongju, 361-763 (Korea, Republic of)

    2009-02-01

    The persistence of Bacillus thuringiensis (Bt) toxins in soil is further enhanced through association with soil particles. Such persistence may improve the effectiveness of controlling target pests, but impose a hazard to non-target organisms in soil ecosystems. In this study, the equilibrium adsorption of the Bt toxin by four clay minerals (montmorillonite, kaolinite, goethite, and silicon dioxide) was investigated, and the kinetic and thermodynamic parameters were calculated. The results showed that Bt toxin could be adsorbed easily by minerals, and the adsorption was much easier at low temperature than at high temperature at the initial concentration varying from 0 to 1000 mg L{sup -1}. The adsorption fitted well to both Langmuir and Freundlich isotherm models, but the Freundlich equation was more suitable. The pseudo-second-order (PSO) was the best application model to describe the adsorption kinetic. The adsorption process appeared to be controlled by chemical process, and the intra-particle diffusion was not the only rate-controlling step. The negative standard free energy ({Delta}{sub r}G{sub m}{sup {theta}}) values of the adsorption indicated that the adsorption of the Bt toxin by the minerals was spontaneous, and the changes of the standard enthalpy ({Delta}{sub r}H{sub m}{sup {theta}}) showed that the adsorption of the Bt toxin by montmorillonite was endothermic while the adsorption by the other three minerals was exothermic.

  4. Characterization of a highly toxic strain of Bacillus thuringiensis serovar kurstaki very similar to the HD-73 strain.

    Science.gov (United States)

    Reinoso-Pozo, Yaritza; Del Rincón-Castro, Ma Cristina; Ibarra, Jorge E

    2016-09-01

    The LBIT-1200 strain of Bacillus thuringiensis was recently isolated from soil, and showed a 6.4 and 9.5 increase in toxicity, against Manduca sexta and Trichoplusia ni, respectively, compared to HD-73. However, LBIT-1200 was still highly similar to HD-73, including the production of bipyramidal crystals containing only one protein of ∼130 000 kDa, its flagellin gene sequence related to the kurstaki serotype, plasmid and RepPCR patterns similar to HD-73, no production of β-exotoxin and no presence of VIP genes. Sequencing of its cry gene showed the presence of a cry1Ac-type gene with four amino acid differences, including two amino acid replacements in domain III, compared to Cry1Ac1, which may explain its higher toxicity. In conclusion, the LBIT-1200 strain is a variant of the HD-73 strain but shows a much higher toxicity, which makes this new strain an important candidate to be developed as a bioinsecticide, once it passes other tests, throughout its biotechnological development. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  5. Specificity and combinatorial effects of Bacillus thuringiensis Cry toxins in the context of GMO environmental risk assessment

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    Angelika eHilbeck

    2015-11-01

    Full Text Available Stacked GM crops expressing up to six Cry toxins from Bacillus thuringiensis are today replacing the formerly grown single- transgene GM crop varieties. Stacking of multiple Cry toxins not only increase the environmental load of toxins but also raise the question on how possible interactions of the toxins can be assessed for risk assessment, which is mandatory for GM crops. However, no operational guidelines for a testing strategy or testing procedures exist. From the developers point of view, little data testing for combinatorial effects of Cry toxins is necessary as the range of affected organisms is focused on pest species and no evidence is claimed to exists pointing to combinatorial effects on nontarget organisms. We have examined this rationale critically using information reported in the scientific literature. To do so we address the hypothesis of narrow specificity of Cry toxins subdivided into three underlying different conceptual conditions i 'efficacy' in target pests as indicator for 'narrow specificity', ii lack of reported adverse effects of Cry toxins on nontarget organisms, and iii proposed modes of action of Cry toxins (or the lack thereof as mechanisms underlying the reported activity/efficacy/specificity of Cry toxins. Complementary to this information we evaluate reports about outcomes of combinatorial effect testing of Cry toxins in the scientific literature and relate those findings to the practice of the environmental risk assessment of Bt-corps in general and of stacked Bt-events in particular.

  6. Pilot-scale biopesticide production by Bacillus thuringiensis subsp. kurstaki using starch industry wastewater as raw material.

    Science.gov (United States)

    Ndao, Adama; Sellamuthu, Balasubramanian; Gnepe, Jean R; Tyagi, Rajeshwar D; Valero, Jose R

    2017-09-02

    Pilot-scale Bacillus thuringiensis based biopesticide production (2000 L bioreactor) was conducted using starch industry wastewater (SIW) as a raw material using optimized operational parameters obtained in 15 L and 150 L fermenters. In pilot scale fermentation process the oxygen transfer rate is a major limiting factor for high product yield. Thus, the volumetric mass transfer coefficient (K L a) remains a tool to determine the oxygen transfer capacity [oxygen utilization rate (OUR) and oxygen transfer rate (OTR)] to obtain better bacterial growth rate and entomotoxicity in new bioreactor process optimization and scale-up. This study results demonstrated that the oxygen transfer rate in 2000 L bioreactor was better than 15 L and 150 L fermenters. The better oxygen transfer in 2000 L bioreactor augmented the bacterial growth [total cell (TC) and viable spore count (SC)] and delta-endotoxin yield. Prepared a stable biopesticide formulation for field use and its entomotoxicity was also evaluated. This study result corroborates the feasibility of industrial scale operation of biopesticide production using starch industry wastewater as raw material.

  7. The Vip3Ag4 Insecticidal Protoxin from Bacillus thuringiensis Adopts A Tetrameric Configuration That Is Maintained on Proteolysis

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    Leopoldo Palma

    2017-05-01

    Full Text Available The Vip3 proteins produced during vegetative growth by strains of the bacterium Bacillus thuringiensis show insecticidal activity against lepidopteran insects with a mechanism of action that may involve pore formation and apoptosis. These proteins are promising supplements to our arsenal of insecticidal proteins, but the molecular details of their activity are not understood. As a first step in the structural characterisation of these proteins, we have analysed their secondary structure and resolved the surface topology of a tetrameric complex of the Vip3Ag4 protein by transmission electron microscopy. Sites sensitive to proteolysis by trypsin are identified and the trypsin-cleaved protein appears to retain a similar structure as an octomeric complex comprising four copies each of the ~65 kDa and ~21 kDa products of proteolysis. This processed form of the toxin may represent the active toxin. The quality and monodispersity of the protein produced in this study make Vip3Ag4 a candidate for more detailed structural analysis using cryo-electron microscopy.

  8. Improving SERS Detection of Bacillus thuringiensis Using Silver Nanoparticles Reduced with Hydroxylamine and with Citrate Capped Borohydride

    International Nuclear Information System (INIS)

    Felix-Rivera, H.; Gonzalez, R.; Rodriguez, G.D.M.; Oliva, M. P.; Hernandez-Rivera, S.P.; Rios-Velazquez, C.

    2011-01-01

    The development of techniques that could be useful in fields other than biological warfare agents countermeasures such as medical diagnostics, industrial microbiology, and environmental applications have become a very important subject of research. Raman spectroscopy can be used in near field or at long distances from the sample to obtain fingerprinting information of chemical composition of microorganisms. In this research, biochemical components of the cell wall and endospores of Bacillus thuringiensis (Bt) were identified by surface-enhanced Raman scattering (SERS) spectroscopy using silver (Ag) nanoparticles (NPs) reduced by hydroxylamine and borohydride capped with sodium citrate. Activation of hot spots, aggregation and surface charge modification of the NPs, was studied and optimized to obtain signal enhancements from Bt by SERS. Slight aggregation of the NPs as well as surface charge modification to a more acidic ambient was induced using small-size borohydride-reduced NPs in the form of metallic suspensions aimed at increasing the Ag NP-Bt interactions. Hydroxylamine-reduced NPs required slight aggregation and no pH modifications in order to obtain high spectral quality results in bringing out SERS signatures of Bt.

  9. Equilibrium, kinetic and thermodynamic studies on the adsorption of the toxins of Bacillus thuringiensis subsp. kurstaki by clay minerals

    International Nuclear Information System (INIS)

    Fu Qingling; Deng Yali; Li Huishu; Liu Jie; Hu Hongqing; Chen Shouwen; Sa Tongmin

    2009-01-01

    The persistence of Bacillus thuringiensis (Bt) toxins in soil is further enhanced through association with soil particles. Such persistence may improve the effectiveness of controlling target pests, but impose a hazard to non-target organisms in soil ecosystems. In this study, the equilibrium adsorption of the Bt toxin by four clay minerals (montmorillonite, kaolinite, goethite, and silicon dioxide) was investigated, and the kinetic and thermodynamic parameters were calculated. The results showed that Bt toxin could be adsorbed easily by minerals, and the adsorption was much easier at low temperature than at high temperature at the initial concentration varying from 0 to 1000 mg L -1 . The adsorption fitted well to both Langmuir and Freundlich isotherm models, but the Freundlich equation was more suitable. The pseudo-second-order (PSO) was the best application model to describe the adsorption kinetic. The adsorption process appeared to be controlled by chemical process, and the intra-particle diffusion was not the only rate-controlling step. The negative standard free energy (Δ r G m θ ) values of the adsorption indicated that the adsorption of the Bt toxin by the minerals was spontaneous, and the changes of the standard enthalpy (Δ r H m θ ) showed that the adsorption of the Bt toxin by montmorillonite was endothermic while the adsorption by the other three minerals was exothermic.

  10. Improving SERS Detection of Bacillus thuringiensis Using Silver Nanoparticles Reduced with Hydroxylamine and with Citrate Capped Borohydride

    Directory of Open Access Journals (Sweden)

    Hilsamar Félix-Rivera

    2011-01-01

    Full Text Available The development of techniques that could be useful in fields other than biological warfare agents countermeasures such as medical diagnostics, industrial microbiology, and environmental applications have become a very important subject of research. Raman spectroscopy can be used in near field or at long distances from the sample to obtain fingerprinting information of chemical composition of microorganisms. In this research, biochemical components of the cell wall and endospores of Bacillus thuringiensis (Bt were identified by surface-enhanced Raman scattering (SERS spectroscopy using silver (Ag nanoparticles (NPs reduced by hydroxylamine and borohydride capped with sodium citrate. Activation of “hot spots”, aggregation and surface charge modification of the NPs, was studied and optimized to obtain signal enhancements from Bt by SERS. Slight aggregation of the NPs as well as surface charge modification to a more acidic ambient was induced using small-size borohydride-reduced NPs in the form of metallic suspensions aimed at increasing the Ag NP-Bt interactions. Hydroxylamine-reduced NPs required slight aggregation and no pH modifications in order to obtain high spectral quality results in bringing out SERS signatures of Bt.

  11. ISOLASI BACILLUS THURINGIENSIS DARI LARVA DAN PENGUJIAN PATOGENISITASNYA TERHADAP LARVA NYAMUK VEKTOR

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    Blondine Ch. P.

    2012-09-01

    Full Text Available A study to evaluate pathogenic organisms as cause of mosquito larvae death was conducted at Wonokerto and Pabelan villages, Salatiga Luar Kota subdistrict, Semarang regency in Central Java from May 1991 through December 1991. Bacterial isolation from dead larvae showed that 31 B. thuringicnsis isolates were obtained from 31 larvae samples collected from 2 location e.g Wonokerto village (3 samples, Pabelan village (28 samples. Nineteen isolates (61,3% showed a pathogenicity of more than 50% to third toward instar larvae of Aedes aegypti and Culex quinquefasciatus respectively 24 hours after exposure. This study shows the possible use of B. thuringiensis for biologic control of mosquitoes which can act as vectors for human diseases.

  12. Susceptibilidade de larvas de Cerotoma arcuata Olivier (Coleoptera: Chrysomelidae a Beauveria bassiana (Bals. Vuillemin, Metarhizium anisopliae (Metsch. Sorokin e Bacillus thuringiensis Berliner Susceptibility of Cerotoma arcuata Olivier (Coleoptera: Chrysomelidae larvae to Beauveria bassiana (Bals. Vuillemin, Metarhizium anisopliae (Metsch. Sorokin and Bacillus thuringiensis Berliner

    Directory of Open Access Journals (Sweden)

    Maria Lucia França Teixeira

    2007-02-01

    Full Text Available Larvas de 2° instar de Cerotoma arcuata foram avaliadas em relação à susceptibilidade aos fungos entomopatogênicos Beauveria bassiana, Metarhizium anisopliae e a bactéria Bacillus thuringiensis com as toxinas Cry3. Os insetos adultos foram mantidos em gaiolas e alimentados com plântulas de feijão (Phaseolus vulgaris L. e as larvas em "gerbox" com cotilédones de plântulas de feijão recém-germinadas. Das oito estirpes de B. bassiana avaliadas, CG 156 e CG 213 causaram 100% de mortalidade das larvas, as duas estirpes de M. anisopliae CG 210 e CG 321 foram patogênicas, eliminando 80 e 100% das larvas de C. arcuata, e, das cinco estirpes de B. thuringiensis testadas, o isolado CG 940 causou 70% de mortalidade das larvas.Second instar larvae of Cerotoma arcuata were evaluated concerning the susceptibility to fungi Beauveria bassiana and Metarhizium anisopliae and Bacillus thuringiensis strains containing Cry3 toxin. Adults of C. arcuata were kept in large cages and fed on bean seedlings and the larvae were reared in ‘gearbox’ feeding on germinated Phaseolus bean cotyledons. Strains CG 156 and CG 213 of B. bassiana killed 100% of the insect larvae and strains CG 210 and CG 321 of M. anisopliae killed 80 and 100% of the insect larvae. Strain CG 940 of B. thuringiensis killed 70% of the insect larvae.

  13. Sequence and expression of two cry8 genes from Bacillus thuringiensis INTA Fr7-4, a native strain from Argentina.

    Science.gov (United States)

    Navas, Laura E; Berretta, Marcelo F; Pérez, Melisa P; Amadio, Ariel F; Ortiz, Elio M; Sauka, Diego H; Benintende, Graciela B; Zandomeni, Rubén O

    2014-01-01

    We found and characterized two cry8 genes from the Bacillus thuringiensis strain INTA Fr7-4 isolated in Argentina. These genes, cry8Kb3 and cry8Pa3, are located in a tandem array within a 13,200-bp DNA segment sequenced from a preparation of total DNA. They encode 1,169- and 1,176-amino-acid proteins, respectively. Both genes were cloned with their promoter sequences and the proteins were expressed separately in an acrystalliferous strain of B. thuringiensis leading to the formation of ovoid crystals in the recombinant strains. The toxicity against larvae of Anthonomus grandis Bh. (Coleoptera: Curculionidae) of a spore-crystal suspension from the recombinant strain containing cry8Pa3 was similar to that of the parent strain INTA Fr7-4. © 2014 S. Karger AG, Basel.

  14. Análisis del Klap como parámetro de escalado en fermentaciones de Bacillus thuringiensis subesp. Medellín

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    Sergio Ordúz

    2000-02-01

    Full Text Available

    Los biopesticidas basados en Bacillus thuringiensis son los de mayor producción y uso a nivel mundial. B. thuringiensis es una bacteria gram positiva que sintetiza proteínas d-endotoxinas que son tóxicas contra larvas de insectos de los géneros Díptera, Lepidóptera y Coleóptera. Las ventajas que tienen los biopesticidas en comparación a los agroquímicos convencionales son: 1 alta selectividad por los insectos blancos, 2 ningún efecto adverso al medio ambiente y 3 bajo índice de resistencia.

  15. Effect ob Bacillus Thuringiensis Aegypti and Gamma Irradiation on the Ultrastructure of the Mid Gut of Arenipses Sabella Larvae Hampson (pyralidae:Lepidoptera)

    International Nuclear Information System (INIS)

    Mikhaiel, A.A.; Abul Fadl, H.A.A.

    2011-01-01

    Laboratory experiments were carried out to investigate the effect of Egyptian Bacillus thuringiensis aegypti on the third larval instar of Arenipses sabella at five concentrations (0.125, 0.25, 0.50, 1 and 2 g/100 ml water). The results showed that the percentages of larval mortality were increased by increasing the applied concentration. A combination of irradiation and B. thuringiensis led to higher mortality in F1 progeny of A. sabella while F1 progeny of irradiated parents with 175 Gy was more susceptible to B. thuringiensis than that of non-irradiated parents. The LC 50 was 0.721 g/100 ml for F1 progeny of non-irradiated parents but 0.237 g/100 ml for those of irradiated parents. There was an inverse relation between the concentration and emergence percentage in non-irradiated larvae and there was a great reduction in the emergence of F1 progeny of irradiated parents as compared to non-irradiated parents. Such reduction was increased by applying a higher concentration of B. thuringiensis. The histopathological studies using ultrastructure microscopy were carried out on the mid gut of the 3rd larval instar after the treatment with LC 50 of the agerin, F1 larvae resulted from irradiated parents with 125 Gy and combined effect of both treatments. Electron microscope observations revealed that the most characteristic effects were mid gut columnar cell vacuolisation, microvilli damages, epithelium cell contents passing into the mid gut lumen and finally the cell death. It could be concluded that the integrated of inherited sterility technique and B. thuringiensis application gave a good control result against A. sabella.

  16. Toxicidad biológica de cepas nativas de Bacillus thuringiensis Berliner en larvas de Tecia solanivora Povolny

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    Paola Martínez

    2011-11-01

    Full Text Available La biodiversidad microbiológica de los suelos del departamento de Boyacá aún no ha sido explorada en toda su magnitud y existen microorganismos, como en el caso de Bacillus thuringiensis Berliner (Bt, que pueden emplearse para el desarrollo de estrategias biológicas de control de plagas en el futuro. Por lo anterior, el presente trabajo evaluó la actividad biológica, expresada como  toxicidad, de cepas nativas de B. thuringiensis en la Polilla Guatemalteca  de  la  papa  Tecia  solanivora Povolny, una de  las plagas más  limitantes en el cultivo de papa en la región andina colombiana. Esta  evaluación  se  realizó con  aislados de  Bt obtenidos y conservados por el Grupo de Manejo Biológico de Cultivos (GMBC en el Laboratorio de Control Biológico de la UPTC, colectados de muestras de  suelo en  la Provincia Centro de Boyacá y  previamente  caracterizados  macro  y microscópicamente.  La   toxicidad  se  evaluó mediante bioensayos con larvas de primer instar de  T.  solanivora,  utilizando  el método  decontaminación superficial de cubos de papa. Se evaluó  la mortalidad  ocho  días  después  de montado  el  bioensayo  y  se  determinó, igualmente,  la Concentración letal media (Cl 50 de las cepas que presentaron  la mayor actividad  tóxica en  los  bioensayos  iniciales.  Los  aislamientosGMBC-B054, GMBC-B071, GMBC-B076, GMBC- B098, GMBC-B111 y GMBC-B117 fueron  los más activos, con Cl 50 de 1.08x106, 4.24x106, 5.12x106, 4.36x103, 3.56x103  y  1.19x104  esporas  · mL-1,respectivamente.

  17. Seleção e caracterização de estirpes de Bacillus thuringiensis efetivas no controle da traça-das-crucíferas Plutella xylostella Selection and characterization of Bacillus thuringiensis strains effective to control the diamondback moth Plutella xylostella

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    Patrícia Teles Medeiros

    2005-11-01

    Full Text Available O objetivo deste trabalho foi selecionar e caracterizar, no Banco de Germoplasma de Bacillus spp., da Empresa Brasileira de Pesquisa Agropecuária, as estirpes de Bacillus thuringiensis mais tóxicas à Plutella xylostella, por métodos morfológicos, bioquímicos e moleculares. Das 203 estirpes testadas, sete causaram 100% de mortalidade e foram semelhantes à estirpe padrão utilizada, B. thuringiensis subsp. kurstaki. Elas apresentaram proteínas de 130 kDa e 65 kDa, presença de genes cry1 e cry2 e cristais bipiramidais, cubóides e redondos. As estirpes selecionadas oferecem novas perspectivas de controle de P. xylostella.The aim of this work was to select and characterize the most toxic Bacillus thuringiensis strains, from the Germplasm Bank of Bacillus spp. of Empresa Brasileira de Pesquisa Agropecuária, against Plutella xylostella. Strains were characterized by morphological, biochemical and molecular methods. It was observed that seven out of the 203 strains tested showed high toxicity compared to the standard used B. thuringiensis subsp. kurstaki (HD-1, which showed 100% mortality. Selected strains showed features described for lepidoptera regarding the protein of 130 kDa and 65 kDa; profile and features were obtained through the PCR reactions, making possible to identify the presence of cry1 and cry2 genes. Moreover, the scanning electron microscopy showed the bipiramydal, cubed and round crystal forms. The selected strains offer new perspectives to control P. xylostella.

  18. Histopathology of Anticarsia gemmatalis Hübner (Lepidoptera; Noctuidae treated with Nucleopolyhedrovirus and Bacillus thuringiensis serovar kurstaki Histopatologia de Anticarsia gemmatalis Hübner (Lepidoptera; Noctuidae tratadas com Virus de Poliedrose Nuclear e Bacillus thuringiensis sorovar kurstaki

    Directory of Open Access Journals (Sweden)

    Neiva Knaak

    2005-06-01

    Full Text Available The Anticarsia gemmatalis is responsible for the use of chemical insecticides in the soybean culture, causing a significant increase in the costs of farming and a great unbalance in the ecosystem. The use of microbial agents, like Bacillus thuringiensis serovar kurstaki (Btk and Anticarsia gemmatalis nucleopolyhedrovirus (AgNPV, they are an alternative to chemical control of the pest insects. In the interaction analysis of the entomopathogenic bacteria and virus it is considered important the in vitro action mode of these microbiology control agents. Therefore, the present study aims the histopathological analysis of the A. gemmatalis larvae digestive system after the interaction in vivo of the entomopathogenic Btk and AgNPV, represented the Dipel and Baculovirus anticarsia formulations, respectively. The evaluations were realized in larvae of 2nd instar, in which the mortality was evaluated daily, and a histopathology was done with collected larvae in time of 1, 3, 6, 12 and 24 hours after the treatments application. The results of the in vivo assays reveal that the treatment using the association of AgNPV-Btk (98.68% of mortality was more efficient than using AgNPV isolatedly (81.28% of mortality, but the Btk when used isolatedly had a mortality of 100%. The treatments showed significant (PA Anticarsia gemmatalis é responsável pelo uso de inseticidas químicos na cultura da soja, ocasionando um significativo aumento nos custos das lavouras e um grande desequilíbrio no ecossistema. O uso de agentes microbianos, como Bacillus thuringiensis sorovar kurstaki (Btk e Vírus de Poliedrose Nuclear de Anticarsia gemmatalis (VPNAg, é uma alternativa para o controle químico de insetos-praga. Na análise da interação de bactérias e vírus entomopatogênicos, considera-se importante o modo de ação in vitro desses agentes de controle microbiano. Assim, o presente trabalho objetiva a análise histopatológica do sistema digestivo das lagartas de A

  19. Bacillus thuringiensis: generalidades: Un acercamiento a su empleo en el biocontrol de insectos lepidópteros que son plagas agrícolas Bacillus thuringiensis: general aspects: An approach to its use in the biological control of lepidopteran insects behaving as agricultural pests

    Directory of Open Access Journals (Sweden)

    Diego H. Sauka

    2008-06-01

    Full Text Available Bacillus thuringiensis es el insecticida biológico más aplicado en el mundo y se utiliza para controlar diversos insectos que afectan la agricultura, la actividad forestal y que transmiten patógenos humanos y animales. B. thuringiensis constituyó durante las últimas décadas un tema de investigación intensiva. Estos esfuerzos brindaron datos importantes sobre las relaciones entre la estructura, el mecanismo de acción y la genética de sus proteínas cristalinas pesticidas, y una visión más clara y coherente sobre estas relaciones ha emergido gracias a ellos. Otros estudios se centraron en el rol ecológico de las proteínas cristalinas de B. thuringiensis, su funcionamiento en sistemas agrícolas y en otros sistemas naturales. Teniendo como base todo el conocimiento generado y las herramientas de la biotecnología, los investigadores están ahora divulgando resultados prometedores sobre el desarrollo de toxinas más útiles, bacterias recombinantes, formulaciones nuevas y plantas transgénicas que expresan actividad pesticida, con el objetivo de asegurar que estos productos sean utilizados con un mayor beneficio y eficacia. Este artículo constituye una tentativa de integrar todos estos progresos recientes sobre el estudio de B. thuringiensis en un contexto de control biológico de plagas de insectos lepidópteros de importancia agrícola.Bacillus thuringiensis is the most widely applied biological pesticide used to control insects that affect agriculture and forestry and which transmit human and animal pathogens. During the past decades B. thuringiensis has been the subject of intensive research. These efforts have yielded considerable data about the relationships between the structure, mechanism of action, and genetics of their pesticidal crystal proteins. As a result, a coherent picture of these relationships has emerged. Other studies have focused on the ecological role of the B. thuringiensis crystal proteins and their performance

  20. Histopathological effects of Bacillus Thuringiensis and gamma irradiation on F1 Larvae of the greater Wax Moth, Galleria Mellonella L

    International Nuclear Information System (INIS)

    Mohamed, H.F.; Mikhaiel, A.A.; Abul-Fadl, H.A.

    2006-01-01

    Full grown male pupae of the greater wax moth, Galleria mellonella L., were gamma irradiated with 50, 100, 200, 300 and 400 Gy. The resulting F1 larvae were treated at the fourth instar with different concentrations (0, 5, 10, 20, and 40 %) of Bacillus thuringiensis (Bt.) var. kurstaki. Combined effects of the two doses of gamma radiation (50 and 100 Gy) and / or Bt. (LC 50 ) on certain biological aspects in addition to histological effects on larval mid gut were studied. The obtained results indicated that Bt. or irradiation treatments either alone or in combination decreased the number of F1 larvae that reached the adult stage as compared to the control. Also, the reduction in survived individuals was obvious at dose level 400 Gy than 50, 100 and 200 Gy (the lower doses). The larval mortality, percent pupation, percent emergence and adult survival were decreased gradually by increasing the concentration of Bt. especially at the combined treatments. The sex ratio was altered in favour of males at either Bt. and / or irradiation treatments. Certain histological changes through longitudinal sections of the mid gut of F1 larvae due to irradiation and / or Bt. treatments were detected. The damage of tissues was increased by increasing the dose of irradiation and / or concentration of Bt. The cytoplasmic extrusion was appeared as the apical margin of cells as a confluent mass and the muscular layers were broken in some parts, large amount of secretions was released in the lumen of the mid gut while a few amounts were attached to the apical margin of the cells. Much destruction of the mid gut took place when the Bt. treatments were combined with gamma irradiation where large number of epithelial cells became vacuolated and the cytoplasm was appeared as confluent masses because of the hydropic analysis of the epithelium

  1. Histopathological effects of cypermethrin and Bacillus thuringiensis var. israelensis on midgut of Chironomus calligraphus larvae (Diptera: Chironomidae).

    Science.gov (United States)

    Lavarías, Sabrina; Arrighetti, Florencia; Siri, Augusto

    2017-06-01

    Pesticides are extensively used for the control of agricultural pests and disease vectors, but they also affect non-target organisms. Cypermethrin (CYP) is a synthetic pyrethroid used worldwide. Otherwise, bioinsecticides like Bacillus thuringiensis var. israelensis (Bti) have received great attention as an environmentally benign and desirable alternative. In order to evaluate the toxicity of those pesticides, Chironomus calligraphus was selected due to its high sensitivity to some toxicants. Third and fourth instars larvae were exposed to serial dilutions of CYP and Bti to determine LC 50 values. In order to evaluate the potentially histopathological alterations as biomarkers, after 96-h of exposure, live larvae were fixed for histological analysis of the mid region of digestive tract. The 96-h LC 50 values were 0.52 and 1.506μg/L for CYP and Bti, respectively. Midgut histological structure of the control group showed a single layer of cubical cells with microvilli in their apical surface and a big central nucleus. The midgut epithelium of larvae exposed to a low concentration of CYP (0.037μg/L) showed secretion activity and vacuolization while at high concentration (0.3μg/L) cells showed a greater disorganization and a more developed fat body. On the other hand, Bti caused progressive histological damage in this tissue. Chironomus calligraphus is sensitive to Bti and CYP toxicity like other Chironomus species. The histopathological alterations could be a valuable tool to assess toxicity mechanism of different pesticides. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Individual and Combined Effects of Bacillus Thuringiensis and Azadirachtin on Plodia Interpunctella Hübner (Lepidoptera: Pyralidae).

    Science.gov (United States)

    Nouri-Ganbalani, Gadir; Borzoui, Ehsan; Abdolmaleki, Arman; Abedi, Zahra; George Kamita, Shizuo

    2016-01-01

    The Indianmeal moth, Plodia interpunctella Hübner (Lepidoptera: Pyralidae), is a major stored product pest that is found throughout the world. In this study, the effect of oral exposure to Bacillus thuringiensis (Berliner) subsp. kurstaki (Bacillales: Bacillaceae) and azadirachtin was evaluated in third instar P. interpunctella under laboratory conditions. The median lethal concentration (LC50) of Bt and azadirachtin on third instars was 490 and 241 μg a.i./ml, respectively. The median lethal time (LT50) of these insecticides was the same (4.5 d following exposure to 750 or 400 μg a.i./ml of Bt or azadirachtin, respectively). When the larvae fed on diet containing LC30 concentrations of both Bt and azadirachtin an additive interaction in terms of mortality was found. A synergistic interaction was found when the larvae fed on diet containing LC50 concentrations of both insecticides. Larvae that fed on insecticide-containing diet (either Bt or azadirachtin at an LC30 concentration, or both insecticides at LC30 or LC50 concentrations) showed lower glycogen and lipid levels, and generally lower protein content in comparison to control larvae. Larvae that fed on diet containing both Bt and azadirachtin showed reduced weight gain and nutritional indices in comparison to control larvae or larvae fed on the diet containing only one of the insecticides. Finally, exposure to both insecticides, either individually or in combination, reduced the level of digestive enzymes found in the midgut. Our findings indicate that both Bt and azadirachtin, either individually or in combination have significant potential for use in controlling of P. interpunctella. © The Author 2016. Published by Oxford University Press on behalf of the Entomological Society of America.

  3. Comparative evaluation of phenoloxidase activity in different larval stages of four lepidopteran pests after exposure to Bacillus thuringiensis.

    Science.gov (United States)

    Valadez-Lira, J A; Alcocer-Gonzalez, J M; Damas, G; Nuñez-Mejía, G; Oppert, B; Rodriguez-Padilla, C; Tamez-Guerra, P

    2012-01-01

    Microbial entomopathogen-based bioinsecticides are recognized as alternatives to synthetic pesticides. Insects defend themselves against microbial pathogens by innate mechanisms, including increased phenoloxidase (PO) activity, but its relationship with microbial bioinsecticides efficacy is little known. This study evaluated the differences in PO activity at different developmental stages of the tobacco budworm Heliothis virescens Fabricius (Lepidoptera: Noctuidae), Indian meal moth Plodia interpunctella (Hübner) (Pyralidae), beet armyworm Spodoptera exigua (Hübner) (Noctuidae), and cabbage looper Trichoplusia ni (Hübner) (Noctuidae). Additionally, 2(nd)- and 4(th)-instars were exposed to the LC(50) value of the commercial Bacillus thuringiensis (Bt) spray, Biobit(®). The percentage of insecticidal activity (IA%) on 2(nd)-instar Biobit-exposed larvae was approximately the predicted 50 % mortality for all species except S. exigua. With all 4(th) instar Biobit-exposed larvae, mortality was not significantly different from that of unexposed larvae. Unexposed insects had a significantly higher PO activity in pre-pupae and pupae than early-instar larvae and adults, whereas PO activity was higher in adult females than in males. Correlation analysis between IA% and PO activity revealed significant r-values (p < 0.01) in 2(nd) instar H. virescens (r = 0.979) and P. interpunctella (r = 0.930). Second instar Biobit-exposed P. interpunctella had 10 times more PO activity than unexposed larvae. Similarly, the amount of total protein was lower in 4(th) instar Biobit-exposed H. virescens and higher in S. exigua. Therefore, the results indicated a relationship between Biobit susceptibility and PO activity in some cases. This information may be useful if the Biobit application period is timed for a developmental stage with low PO activity. However, more studies are needed to determine the correlation of each insect with a particular bioinsecticide.

  4. Bacillus thuringiensis Cry1Ia10 and Vip3Aa protein interactions and their toxicity in Spodoptera spp. (Lepidoptera).

    Science.gov (United States)

    Bergamasco, V B; Mendes, D R P; Fernandes, O A; Desidério, J A; Lemos, M V F

    2013-02-01

    The polyphagous pests belonging to the genus Spodoptera are considered to be among the most important causes of damage and are widely distributed throughout the Americas'. Due to the extensive use of genetically modified plants containing Bacillus thuringiensis genes that code for insecticidal proteins, resistant insects may arise. To prevent the development of resistance, pyramided plants, which express multiple insecticidal proteins that act through distinct mode of actions, can be used. This study analyzed the mechanisms of action for the proteins Cry1Ia10 and Vip3Aa on neonatal Spodoptera frugiperda, Spodoptera albula, Spodoptera eridania and Spodoptera cosmioides larvae. The interactions of these toxins with receptors on the intestinal epithelial membrane were also analyzed by binding biotinylated toxins to brush border membrane vesicles (BBMVs) from the intestines of these insects. A putative receptor of approximately 65 kDa was found by ligand blotting in all of these species. In vitro competition assays using biotinylated proteins have indicated that Vip3Aa and Cry1Ia10 do not compete for the same receptor for S. frugiperda, S. albula and S. cosmioides and that Vip3Aa was more efficient than Cry1Ia10 when tested individually, by bioassays. A synergistic effect of the toxins in S. frugiperda, S. albula and S. cosmioides was observed when they were combined. However, in S. eridania, Cry1Ia10 and Vip3Aa might compete for the same receptor and through bioassays Cry1Ia10 was more efficient than Vip3Aa and showed an antagonistic effect when the proteins were combined. These results suggest that using these genes to develop pyramided plants may not prove effective in preventing the development of resistance in S. eridiana. Copyright © 2012 Elsevier Inc. All rights reserved.

  5. Comparison and validation of methods to quantify Cry1Ab toxin from Bacillus thuringiensis for standardization of insect bioassays.

    Science.gov (United States)

    Crespo, André L B; Spencer, Terence A; Nekl, Emily; Pusztai-Carey, Marianne; Moar, William J; Siegfried, Blair D

    2008-01-01

    Standardization of toxin preparations derived from Bacillus thuringiensis (Berliner) used in laboratory bioassays is critical for accurately assessing possible changes in the susceptibility of field populations of target pests. Different methods were evaluated to quantify Cry1Ab, the toxin expressed by 80% of the commercially available transgenic maize that targets the European corn borer, Ostrinia nubilalis (Hübner). We compared three methods of quantification on three different toxin preparations from independent sources: enzyme-linked immunosorbent assay (ELISA), sodium dodecyl sulfate-polyacrylamide gel electrophoresis and densitometry (SDS-PAGE/densitometry), and the Bradford assay for total protein. The results were compared to those obtained by immunoblot analysis and with the results of toxin bioassays against susceptible laboratory colonies of O. nubilalis. The Bradford method resulted in statistically higher estimates than either ELISA or SDS-PAGE/densitometry but also provided the lowest coefficients of variation (CVs) for estimates of the Cry1Ab concentration (from 2.4 to 5.4%). The CV of estimates obtained by ELISA ranged from 12.8 to 26.5%, whereas the CV of estimates obtained by SDS-PAGE/densitometry ranged from 0.2 to 15.4%. We standardized toxin concentration by using SDS-PAGE/densitometry, which is the only method specific for the 65-kDa Cry1Ab protein and is not confounded by impurities detected by ELISA and Bradford assay for total protein. Bioassays with standardized Cry1Ab preparations based on SDS-PAGE/densitometry showed no significant differences in LC(50) values, although there were significant differences in growth inhibition for two of the three Cry1Ab preparations. However, the variation in larval weight caused by toxin source was only 4% of the total variation, and we conclude that standardization of Cry1Ab production and quantification by SDS-PAGE/densitometry may improve data consistency in monitoring efforts to identify changes in

  6. Spatial and temporal distribution of airborne Bacillus thuringiensis var. kurstaki during an aerial spray program for gypsy moth eradication.

    Science.gov (United States)

    Teschke, K; Chow, Y; Bartlett, K; Ross, A; van Netten, C

    2001-01-01

    We measured airborne exposures to the biological insecticide Bacillus thuringiensis var. kurstaki (Btk) during an aerial spray program to eradicate gypsy moths on the west coast of Canada. We aimed to determine whether staying indoors during spraying reduced exposures, to determine the rate of temporal decay of airborne concentrations, and to determine whether drift occurred outside the spray zone. During spraying, the average culturable airborne Btk concentration measured outdoors within the spray zone was 739 colony-forming units (CFU)/m3 of air. Outdoor air concentrations decreased over time, quickly in an initial phase with a half time of 3.3 hr, and then more slowly over the following 9 days, with an overall half-time of about 2.4 days. Inside residences during spraying, average concentrations were initially 2-5 times lower than outdoors, but at 5-6 hr after spraying began, indoor concentrations exceeded those outdoors, with an average of 244 CFU/m3 vs. 77 CFU/m3 outdoors, suggesting that the initial benefits of remaining indoors during spraying may not persist as outside air moves indoors with normal daily activities. There was drift of culturable Btk throughout a 125- to 1,000-meter band outside the spray zone where measurements were made, a consequence of the fine aerosol sizes that remained airborne (count median diameters of 4.3 to 7.2 microm). Btk concentrations outside the spray zone were related to wind speed and direction, but not to distance from the spray zone.

  7. Mapping of residues of fibrinogen cleaved by protease II of Bacillus thuringiensis var. israelensis IMV B-7465

    Directory of Open Access Journals (Sweden)

    E. M. Stohniy

    2016-04-01

    Full Text Available The limited proteolysis of macromolecules allows obtaining the fragments that preserve the structure and functional properties of the whole molecule and could be used in the study of proteins structure and function. Proteases targeted to fibrinogen and fibrin are of interest as the tool for obtaining of functionally active fragments of fibrin(ogen and for the direct defibrination in vivo. That is why the aim of the present work was to study the proteolytic action of Protease II (PII purified from Bacillus thuringiensis var. israelensis IMV B-7465 on fibrinogen. Hydrolysis products of fibrinogen by PII were analysed by SDS-PAGE under reducing conditions with further immunoprobing using the mouse monoclonal 1-5A (anti-Aα509-610 and ІІ-5С (anti-Aα20-78 antibodies. It was shown that PII cleaved preferentially the Aα-chain of fibrinogen splitting off the peptide with apparent molecular weight of 10 kDa that corresponded the C-terminal part of Aα-chain of fibrinogen molecule. MALDI-TOF analysis of hydrolysis of fibrinogen was performed using a Voyager-DE. Results analyzed by Data Explorer 4.0.0.0 allowed to detect the main peak occurring at mass/charge (M/Z ratio of 11 441 Da. According to “Peptide Mass Calculator” this peptide corresponded to fragment Аα505-610 of fibrinogen molecule. The result showed that PII cleaves the peptide bond AαAsp-Thr-Ala504-Ser505. Thus, PII can be used for the obtaining of unique fragments of fibrinogen molecule. As far as αC-domain contains numerous sites of fibrin intermolecular interactions we can consider PII as a prospective agent for their study and for the defibrination.

  8. Acute, sublethal and combination effects of azadirachtin and Bacillus thuringiensis toxins on Helicoverpa armigera (Lepidoptera: Noctuidae) larvae.

    Science.gov (United States)

    Singh, G; Rup, P J; Koul, Opender

    2007-08-01

    The efficacy of neem (1500 ppm azadirachtin (AI)), Delfin WG, a biological insecticide based on selected strain of Bacillus thuringiensis Berliner (Bt) subspecies kurstaki, and Cry1Ac protein, either individually or in combination, were examined against first to fourth instar Helicoverpa armigera (Hübner) larvae. Using an oral administration method, various growth inhibitory concentrations (EC) and lethal concentrations (LC) were determined for each bioagent. Combinations of sublethal concentrations of Bt spray formulation with azadirachtin at EC50 or EC95 levels not only enhanced the toxicity, but also reduced the duration of action when used in a mixture. The LC20 and LC50 values for Cry1Ac toxin were 0.06 and 0.22 microg ml-1, respectively. Bt-azadirachtin combinations of LC50+EC20 and LC50+EC50 result in 100% mortality. The mortality also was significant in LC20+EC20 and LC20+EC50 mixtures. These studies imply that the combined action is not synergistic but complimentary, with azadirachtin particularly facilitating the action of Bt. The Bt spray-azadirachtin combination is more economical than combinations that involve isolating the toxic protein, as the Bt spray formulations can be combined in a spray mixture with neem. These combinations may be useful for controlling bollworm populations that have acquired resistance to Bt as they may not survive the effect of mixture. Azadirachtin may be useful as a means of reducing the endotoxin concentrations in a mixture, to promote increased economic savings and further reduce the probability of resistance development to either insect control agent.

  9. Induction of rapid and selective cell necrosis in Drosophila using Bacillus thuringiensis Cry toxin and its silkworm receptor.

    Science.gov (United States)

    Obata, Fumiaki; Tanaka, Shiho; Kashio, Soshiro; Tsujimura, Hidenobu; Sato, Ryoichi; Miura, Masayuki

    2015-07-08

    Genetic ablation of target cells is a powerful tool to study the origins and functions of cells, tissue regeneration, or pathophysiology in a human disease model in vivo. Several methods for selective cell ablation by inducing apoptosis have been established, using exogenous toxins or endogenous proapoptotic genes. However, their application is limited to cells with intact apoptotic machinery. Herein, we established a method for inducing rapid and selective cell necrosis by the pore-forming bacterial toxin Cry1Aa, which is specifically active in cells expressing the Cry1Aa receptor (CryR) derived from the silkworm Bombyx mori. We demonstrated that overexpressing CryR in Drosophila melanogaster tissues induced rapid cell death of CryR-expressing cells only, in the presence of Cry1Aa toxin. Cry/CryR system was effective against both proliferating cells in imaginal discs and polyploid postmitotic cells in the fat body. Live imaging analysis of cell ablation revealed swelling and subsequent osmotic lysis of CryR-positive cells after 30 min of incubation with Cry1Aa toxin. Osmotic cell lysis was still triggered when apoptosis, JNK activation, or autophagy was inhibited, suggesting that Cry1Aa-induced necrotic cell death occurred independently of these cellular signaling pathways. Injection of Cry1Aa into the body cavity resulted in specific ablation of CryR-expressing cells, indicating the usefulness of this method for in vivo cell ablation. With Cry toxins from Bacillus thuringiensis, we developed a novel method for genetic induction of cell necrosis. Our system provides a "proteinous drill" for killing target cells through physical injury of the cell membrane, which can potentially be used to ablate any cell type in any organisms, even those that are resistant to apoptosis or JNK-dependent programmed cell death.

  10. Monoclonal Antibody Analysis and Insecticidal Spectrum of Three Types of Lepidopteran-Specific Insecticidal Crystal Proteins of Bacillus thuringiensis

    Science.gov (United States)

    Höfte, Herman; Van Rie, Jeroen; Jansens, Stefan; Van Houtven, Annemie; Vanderbruggen, Hilde; Vaeck, Mark

    1988-01-01

    We have investigated the protein composition and the insecticidal spectrum of crystals of 29 Bacillus thuringiensis strains active against lepidopteran larvae. All crystals contained proteins of 130 to 140 kilodaltons (kDa) which could be grouped into three types by the molecular weight of the protoxin and the trypsin-activated core fragment. Proteins of the three types showed a characteristic insecticidal spectrum when tested against five lepidopteran species. Type A crystal proteins were protoxins of 130 or 133 kDa, which were processed into 60-kDa toxins by trypsin. Several genes encoding crystal proteins of this type have been cloned and sequenced earlier. They are highly conserved in the N-terminal half of the toxic fragment and were previously classified in three subtypes (the 4.5-, 5.3-, and 6.6-kilobase subtypes) based on the restriction map of their genes. The present study shows that different proteins of these three subtypes were equally toxic against Manduca sexta and Pieris brassicae and had no detectable activity against Spodoptera littoralis. However, the 4.5-, 5.3-, and 6.6-kilobase subtypes differed in their toxicity against Heliothis virescens and Mamestra brassicae. Type B crystal proteins consisted of 140-kDa protoxins with a 55-kDa tryptic core fragment. These were only active against one of the five insect species tested (P. brassicae). The protoxin and the trypsin-activated toxin of type C were 135- and 63-kDa proteins, respectively. Proteins of this type were associated with high toxicity against S. littoralis and M. brassicae. A panel of 35 monoclonal antibodies was used to compare the structural characteristics of crystal proteins of the three different types and subtypes. Each type of protein could be associated with a typical epitope structure, indicating an unambiguous correlation between antigenic structure and insect specificity. Images PMID:16347711

  11. Modulation of Bacillus thuringiensis Phosphatidylinositol-Specific Phospholipase C Activity by Mutations in the Putative Dimerization Interface

    Energy Technology Data Exchange (ETDEWEB)

    Shi, X.; Shao, C; Zhang, X; Zambonelli, C; Redfield, A; Head, J; Seaton, B; Roberts, M

    2009-01-01

    Cleavage of phosphatidylinositol (PI) to inositol 1,2-(cyclic)-phosphate (cIP) and cIP hydrolysis to inositol 1-phosphate by Bacillus thuringiensis phosphatidylinositol-specific phospholipase C are activated by the enzyme binding to phosphatidylcholine (PC) surfaces. Part of this reflects improved binding of the protein to interfaces. However, crystallographic analysis of an interfacially impaired phosphatidylinositol-specific phospholipase (W47A/W242A) suggested protein dimerization might occur on the membrane. In the W47A/W242A dimer, four tyrosine residues from one monomer interact with the same tyrosine cluster of the other, forming a tight dimer interface close to the membrane binding regions. We have constructed mutant proteins in which two or more of these tyrosine residues have been replaced with serine. Phospholipid binding and enzymatic activity of these mutants have been examined to assess the importance of these residues to enzyme function. Replacing two tyrosines had small effects on enzyme activity. However, removal of three or four tyrosine residues weakened PC binding and reduced PI cleavage by the enzyme as well as PC activation of cIP hydrolysis. Crystal structures of Y247S/Y251S in the absence and presence of myo-inositol as well as Y246S/Y247S/Y248S/Y251S indicate that both mutant proteins crystallized as monomers, were very similar to one another, and had no change in the active site region. Kinetic assays, lipid binding, and structural results indicate that either (i) a specific PC binding site, critical for vesicle activities and cIP activation, has been impaired, or (ii) the reduced dimerization potential for Y246S/Y247S/Y248S and Y246S/Y247S/Y248S/Y251S is responsible for their reduced catalytic activity in all assay systems.

  12. Dynamics of Bacillus thuringiensis var. israelensis and Lysinibacillus sphaericus spores in urban catch basins after simultaneous application against mosquito larvae.

    Directory of Open Access Journals (Sweden)

    Valeria Guidi

    Full Text Available Bacillus thuringiensis var. israelensis (Bti and Lysinibacillus sphaericus (Lsph are extensively used in mosquito control programs. These biocides are the active ingredients of a commercial larvicide. Quantitative data on the fate of both Bti and Lsph applied together for the control of mosquitoes in urban drainage structures such as catch basins are lacking. We evaluated the dynamics and persistence of Bti and Lsph spores released through their concomitant application in urban catch basins in southern Switzerland. Detection and quantification of spores over time in water and sludge samples from catch basins were carried out using quantitative real-time PCR targeting both cry4A and cry4B toxin genes for Bti and the binA gene for Lsph. After treatment, Bti and Lsph spores attained concentrations of 3.76 (± 0.08 and 4.13 (± 0.09 log ml(-1 in water, then decreased progressively over time, reaching baseline values. For both Bti and Lsph, spore levels in the order of 10(5 g(-1 were observed in the bottom sludge two days after the treatment and remained constant for the whole test period (275 days. Indigenous Lsph strains were isolated from previously untreated catch basins. A selection of those was genotyped using pulsed field gel electrophoresis of SmaI-digested chromosomal DNA, revealing that a subset of isolates were members of the clonal population of strain 2362. No safety issues related to the use of this biopesticide in the environment have been observed during this study, because no significant increase in the number of spores was seen during the long observation period. The isolation of native Lysinibacillus sphaericus strains belonging to the same clonal population as strain 2362 from catch basins never treated with Lsph-based products indicates that the use of a combination of Bti and Lsph for the control of mosquitoes does not introduce non-indigenous microorganisms in this area.

  13. Effect of transgenic Bacillus thuringiensis rice lines on mortality and feeding behavior of rice stem borers (Lepidoptera: Crambidae).

    Science.gov (United States)

    Chen, Hao; Zhang, Guoan; Zhang, Qifa; Lin, Yongjun

    2008-02-01

    Ten transgenic Bacillus thuringiensis Bt rice, Oryza sativa L., lines with different Bt genes (two Cry1Ac lines, three Cry2A lines, and five Cry9C lines) derived from the same variety Minghui 63 were evaluated in both the laboratory and the field. Bioassays were conducted by using the first instars of two main rice lepidopteran insect species: yellow stem borer, Scirpophaga incertulas (Walker) and Asiatic rice borer, Chilo suppressalis (Walker). All transgenic lines exhibited high toxicity to these two rice borers. Field evaluation results also showed that all transgenic lines were highly insect resistant with both natural infestation and manual infestation of the neonate larvae of S. incertulas compared with the nontransformed Minghui63. Bt protein concentrations in leaves of 10 transgenic rice lines were estimated by the sandwich enzyme-linked immunosorbent assay. The cry9C gene had the highest expression level, next was cry2A gene, and the cry1Ac gene expressed at the lowest level. The feeding behavior of 7-d-old Asiatic rice borer to three classes of Bt transgenic rice lines also was detected by using rice culm cuttings. The results showed that 7-d-old larvae of Asiatic rice borer have the capacity to distinguish Bt and non-Bt culm cuttings and preferentially fed on non-Bt cuttings. When only Bt culm cuttings with three classes of different Bt proteins (CrylAc, Cry2A, and Cry9C) were fed, significant distribution difference of 7-d-old Asiatic rice borer in culm cuttings of different Bt proteins also was found. In the current study, we evaluate different Bt genes in the same rice variety in both the laboratory and the field, and also tested feeding behavior of rice insect to these Bt rice. These data are valuable for the further development of two-toxin Bt rice and establishment of appropriate insect resistance management in the future.

  14. Intracellular and Extracellular Expression of Bacillus thuringiensis Crystal Protein Cry5B in Lactococcus lactis for Use as an Anthelminthic

    Science.gov (United States)

    Durmaz, Evelyn; Hu, Yan; Aroian, Raffi V.

    2015-01-01

    The Bacillus thuringiensis crystal (Cry) protein Cry5B (140 kDa) and a truncated version of the protein, tCry5B (79 kDa), are lethal to nematodes. Genes encoding the two proteins were separately cloned into a high-copy-number vector with a strong constitutive promoter (pTRK593) in Lactococcus lactis for potential oral delivery against parasitic nematode infections. Western blots using a Cry5B-specific antibody revealed that constitutively expressed Cry5B and tCry5B were present in both cells and supernatants. To increase production, cry5B was cloned into the high-copy-number plasmid pMSP3535H3, carrying a nisin-inducible promoter. Immunoblotting revealed that 3 h after nisin induction, intracellular Cry5B was strongly induced at 200 ng/ml nisin, without adversely affecting cell viability or cell membrane integrity. Both Cry5B genes were also cloned into plasmid pTRK1061, carrying a promoter and encoding a transcriptional activator that invoke low-level expression of prophage holin and lysin genes in Lactococcus lysogens, resulting in a leaky phenotype. Cry5B and tCry5B were actively expressed in the lysogenic strain L. lactis KP1 and released into cell supernatants without affecting culture growth. Lactate dehydrogenase (LDH) assays indicated that Cry5B, but not LDH, leaked from the bacteria. Lastly, using intracellular lysates from L. lactis cultures expressing both Cry5B and tCry5B, in vivo challenges of Caenorhabditis elegans worms demonstrated that the Cry proteins were biologically active. Taken together, these results indicate that active Cry5B proteins can be expressed intracellularly in and released extracellularly from L. lactis, showing potential for future use as an anthelminthic that could be delivered orally in a food-grade microbe. PMID:26682852

  15. Midgut GPI-anchored proteins with alkaline phosphatase activity from the cotton boll weevil (Anthonomus grandis) are putative receptors for the Cry1B protein of Bacillus thuringiensis.

    Science.gov (United States)

    Martins, Erica Soares; Monnerat, Rose Gomes; Queiroz, Paulo Roberto; Dumas, Vinicius Fiuza; Braz, Shélida Vasconcelos; de Souza Aguiar, Raimundo Wagner; Gomes, Ana Cristina Menezes Mendes; Sánchez, Jorge; Bravo, Alejandra; Ribeiro, Bergmann Morais

    2010-02-01

    Cry toxins from Bacillus thuringiensis (Bt) are used for insect control. They interact with specific receptors located on the host cell surface and are activated by host proteases following receptor binding resulting in midgut epithelial cells lysis. In this work we had cloned, sequenced and expressed a cry1Ba toxin gene from the B thuringiensis S601 strain which was previously shown to be toxic to Anthonomus grandis, a cotton pest. The Cry1Ba6 protein expressed in an acrystaliferous B. thuringiensis strain was toxic to A. grandis in bioassays. The binding of Cry1Ba6 toxin to proteins located in the midgut brush border membrane of A. grandis was analyzed and we found that Cry1Ba6 binds to two proteins (62 and 65kDa) that showed alkaline phosphatase (ALP) activity. This work is the first report that shows the localization of Cry toxin receptors in the midgut cells of A. grandis. 2009. Published by Elsevier Ltd.

  16. Bacillus thuringiensis (Bt) transgenic crop: an environment friendly insect-pest management strategy.

    Science.gov (United States)

    Kumar, Suresh; Chandra, Amaresh; Pandey, K C

    2008-09-01

    Introduction of DDT (dichloro-diphenyl-trichloroethane) and following move towards indiscriminate use of synthetic chemical insecticides led to the contamination of water and food sources, poisoning of non-target beneficial insects and development of insect-pests resistant to the chemical insecticides. Increased public concems about the adverse environmental effects of indiscriminate use of chemical insecticides prompted search of altemative methods for insect-pest control. One of the promising alternatives has been the use of biological control agents. There is well-documented history of safe application of Bt (B. thuringiensis, a gram positive soil bacterium) as effective biopesticides and a number of reports of expression of delta-endotoxin gene(s) in crop plants are available. Only a few insecticidal sprays are required on Bt transgenic crops, which not only save cost and time, but also reduce health risks. Insects exhibit remarkable ability to develop resistance to different insecticidal compounds, which raises concern about the unsystematic use of Bt transgenic technology also. Though resistance to Bt products among insect species under field conditions has been rare, laboratory studies show that insects are capable of developing high levels of resistance to one ormore Cry proteins. Now it is generally agreed that 'high-dose/refuge strategy' is the most promising and practical approach to prolong the effectiveness of Bt toxins. Although manybiosafety concerns, ethical and moral issues exist, area under Bt transgenic crops is rapidly increasing and they are cultivated on more than 32 million hectares world over Even after reservation of European Union (EU) for acceptance of geneticaly modified (GM) crops, 6 out of 25 countries have already adopted Bt crops and many otherindustrial countries will adopt Bt transgenic crops in near future. While the modem biotechnology has been recognized to have a great potential for the promotion of human well-being, adoption

  17. Deletion of aprA and nprA genes for alkaline protease A and neutral protease A from bacillus thuringiensis: effect on insecticidal crystal proteins.

    Science.gov (United States)

    Tan, Y; Donovan, W P

    2001-11-17

    The aprA gene encoding alkaline protease A (AprA) was cloned from Bacillus thuringiensis subsp. kurstaki, and the cloned gene was used to construct aprA-deleted (aprA1) strains of B. thuringiensis. An aprA1 strain of B. thuringiensis that contained the wild-type gene for neutral protease A (nprA(+)) displayed levels of extracellular proteolytic activity that were similar to those of an aprA(+)nprA(+) strain. However, when EDTA was included in the protease assay to inhibit NprA activity the aprA1nprA(+) strain displayed only 2% of the extracellular proteolytic activity of the aprA(+)nprA(+) strain. A strain that was deleted for both aprA and nprA (aprA1nprA3 strain) failed to produce detectable levels of proteolytic activity either in the presence or absence of EDTA in the assay. Compared with the aprA(+)nprA(+) strain the aprA1nprA(+) strain yielded 10% more full-length Cry1Bb crystal protein and the aprA1nprA3 strain yielded 25% more full-length Cry1Bb protein. No significant differences were seen in the 50% lethal dose of Cry1Bb protein from aprA(+)nprA(+) and aprA1nprA3 strains against three species of lepidopteran insects. These results suggest that enhanced yield of certain crystal proteins can be obtained by deletion of the genes aprA and nprA which are the major extracellular proteases of B. thuringiensis.

  18. Proteomic analysis of Bacillus thuringiensis at different growth phases by using an automated online two-dimensional liquid chromatography-tandem mass spectrometry strategy.

    Science.gov (United States)

    Huang, Shaoya; Ding, Xuezhi; Sun, Yunjun; Yang, Qi; Xiao, Xiuqing; Cao, Zhenping; Xia, Liqiu

    2012-08-01

    The proteome of a new Bacillus thuringiensis subsp. kurstaki strain, 4.0718, from the middle vegetative (T(1)), early sporulation (T(2)), and late sporulation (T(3)) phases was analyzed using an integrated liquid chromatography (LC)-based protein identification system. The system comprised two-dimensional (2D) LC coupled with nanoscale electrospray ionization (ESI) tandem mass spectrometry (MS/MS) on a high-resolution hybrid mass spectrometer with an automated data analysis system. After deletion of redundant proteins from the different batches and B. thuringiensis subspecies, 918, 703, and 778 proteins were identified in the respective three phases. Their molecular masses ranged from 4.6 Da to 477.4 Da, and their isoelectric points ranged from 4.01 to 11.84. Function clustering revealed that most of the proteins in the three phases were functional metabolic proteins, followed by proteins participating in cell processes. Small molecular and macromolecular metabolic proteins were further classified according to the Kyoto Encyclopedia of Genes and Genome and BioCyc metabolic pathway database. Three protoxins (Cry2Aa, Cry1Aa, and Cry1Ac) as well as a series of potential intracellular active factors were detected. Many significant proteins related to spore and crystal formation, including sporulation proteins, help proteins, chaperones, and so on, were identified. The expression patterns of two identified proteins, CotJc and glutamine synthetase, were validated by Western blot analysis, which further confirmed the MS results. This study is the first to use shotgun technology to research the proteome of B. thuringiensis. Valuable experimental data are provided regarding the methodology of analyzing the B. thuringiensis proteome (which can be used to produce insecticidal crystal proteins) and have been added to the related protein database.

  19. Evaluation of the time-concentration-mortality responses of Plutella xylostella larvae to the interaction of Isaria fumosorosea with the insecticides beta-cypermethrin and Bacillus thuringiensis.

    Science.gov (United States)

    Nian, Xiao-ge; He, Yu-rong; Lu, Li-hua; Zhao, Rui

    2015-02-01

    Entomopathogenic fungi are potential candidates for controlling Plutella xylostella, a cosmopolitan pest of crucifers. In this study, bioassays were conducted to evaluate the interaction between Isaria fumosorosea and sublethal doses of two insecticides, beta-cypermethrin and Bacillus thuringiensis, against P. xylostella. Data of each assay were in good agreement with the time-concentration-mortality model, indicating a strong dependence of the fungus and insecticide interaction on both concentration and post-exposure time. Using beta-cypermethrin 58-116 µg mL(-1) or B. thuringiensis 222.5-890 µg mL(-1) with the fungus significantly enhanced fungal efficacy. The LC50 values of the fungus declined over a 1-7 day period after exposure, and the LT50 values decreased with increasing concentration. Based on LC50 or LC90 estimates, synergism between the fungus and beta-cypermethrin resulted in a 2.7-28.3-fold reduction in LC50 values and a 12.1-19.6-fold reduction in LC90 values, while synergism of the fungus with B. thuringiensis led to a 2.4-385.0-fold reduction in LC50 values and a 4.4-151.7-fold reduction in LC90 values. Results show that sublethal doses of B. thuringiensis and beta-cypermethrin can synergise I. fumosorosea activity on P. xylostella, suggesting that combination of I. fumosorosea with the two insecticides might offer an integrated approach to controlling P. xylostella in practice. © 2014 Society of Chemical Industry. © 2014 Society of Chemical Industry.

  20. Consequences for Protaphorura armata (Collembola: Onychiuridae) following exposure to genetically modified Bacillus thuringiensis (Bt) maize and non-Bt maize

    DEFF Research Database (Denmark)

    Heckmann, Lars-Henrik; Griffiths, Bryan S; Caul, Sandra

    2006-01-01

    Studies on the effect of genetically modified Bacillus thuringiensis (Bt) crops on true soil dwelling non-target arthropods are scarce. The objective of this study was to assess the influence of a 4-week exposure to two Bt maize varieties (Cry1Ab) Cascade and MEB307 on the collembolan Protaphorur...

  1. Bacillus thuringiensis var. tenebrionis’in Leptinotarsa decemlineata (Say.) (Coleoptera:Chrysomelidae) larvalarının ortabarsağina etik sürecinin histolojik yöntemlerle belirlenmesi

    Czech Academy of Sciences Publication Activity Database

    Turanli, F.; Cabuk, M.; Kismali, S.; Gelbič, Ivan

    2006-01-01

    Roč. 30, č. 2 (2006), s. 137-150 ISSN 1010-6960 Institutional research plan: CEZ:AV0Z50070508 Keywords : Bacillus thuringiensis var. tenebrionis * Leptinotarsa decemlineata * mode of action Subject RIV: GF - Plant Pathology, Vermin, Weed, Plant Protection

  2. [Genes of insecticidal crystal proteins with dual specificity in Bacillus thuringiensis strains, isolated in the Crimea territory].

    Science.gov (United States)

    Rymar, S Iu; Isakova, I A; Kuznietsova, L M; Kordium, V A

    2006-01-01

    The insecticidal crystal proteins of 15 B. thuringiensis strains, isolated in the Crimea territory that are toxical for some Lepidoptera and Colorado potato beetle larvae were identified by PAGE electrophoresis. Ten strains produced the crystal proteins with high molecular weight (> 120 kD). PCR with use of broad specificity primers and DNA of these B. thuringiensis strains as template demonstrated the specific PCR products (1000 bp). Amplified DNA fragments were cloned and sequenced. The nucleotide sequence analysis revealed that the genomes of ten strains of B. thuringiensis carried Cry1B genes, which are responsible for production of the insecticidal crystal proteins with dual specificity. The influence of the solubilization conditions on the structure and toxicity of Cry1B protein for Colorado potato beetle larvae was shown. The dual toxicity of studied B. thuringiensis strains is explained by the Cry1B genes presence in their genomes. These strains may be used to develop the broad specificity bioinsecticides.

  3. Conjugative plasmid pAW63 brings new insights into the genesis of the Bacillus anthracis virulence plasmid pXO2 and of the Bacillus thuringiensis plasmid pBT9727

    Directory of Open Access Journals (Sweden)

    Mahillon Jacques

    2005-07-01

    Full Text Available Abstract Background Bacillus cereus, Bacillus anthracis and Bacillus thuringiensis belong to the genetically close-knit Bacillus cereus sensu lato group, a family of rod-shaped Gram-positive bacteria. pAW63 is the first conjugative plasmid from the B. cereus group to be completely sequenced. Results The 71,777 bp nucleotide sequence of pAW63 reveals a modular structure, including a 42 kb tra region encoding homologs of the Type IV secretion systems components VirB11, VirB4 and VirD4, as well as homologs of Gram-positive conjugation genes from Enterococcus, Lactococcus, Listeria, Streptococcus and Staphylococcus species. It also firmly establishes the existence of a common backbone between pAW63, pXO2 from Bacillus anthracis and pBT9727 from the pathogenic Bacillus thuringiensis serovar konkukian strain 97-27. The alignment of these three plasmids highlights the presence of well conserved segments, in contrast to distinct regions of high sequence plasticity. The study of their specific differences has provided a three-point reference framework that can be exploited to formulate solid hypotheses concerning the functionalities and the molecular evolution of these three closely related plasmids. This has provided insight into the chronology of their divergence, and led to the discovery of two Type II introns on pAW63, matching copies of the mobile element IS231L in different loci of pXO2 and pBT9727, and the identification on pXO2 of a 37 kb pathogenicity island (PAI containing the anthrax capsule genes. Conclusion The complete sequence determination of pAW63 has led to a functional map of the plasmid yielding insights into its conjugative apparatus, which includes T4SS-like components, as well as its resemblance to other large plasmids of Gram-positive bacteria. Of particular interest is the extensive homology shared between pAW63 and pXO2, the second virulence plasmid of B. anthracis, as well as pBT9727 from the pathogenic strain B. thuringiensis

  4. Evaluación de dos formulaciones de Bacillus thuringiensis H-14 para el control de larvas de Aedes aegypti

    Directory of Open Access Journals (Sweden)

    Marco F. Suárez

    1987-06-01

    Full Text Available Se evaluaron en el laboratorio y en condiciones naturales, dos formulaciones de Bacillus thuringiensis H-14, el cual es un insecticida microbiológico contra larvas de Aedes aegypti Se utilizó polvo humectable (3.500 AA/mg y líquido concentrado (1.000 AA/mg (AA = Unidades Tóxicas Internacionales en A. aegypti. Se calcularon las dosis letales (DL 99,9 para cada formulación. En una estación de laboratorio que simulaba las condiciones naturales de cría de A. aegypti, se realizaron pruebas con cuatro dosis diferentes de cada una de las dos formulaciones. Con una dosis de 1.050 AA/I, se encontró una acción residual de 11 días con el polvo humectable y de 9,9 días con el líquido concentrado. La acción residual no aumentó en proporción directa con la dosis. La tasa de emergencia diaria de adultos tardó 21,7 días en recuperarse hasta el 30% de su nivel de pretratamiento, cuando se aplicó polvo humectable y 18 días cuando se aplicó Iíquido concentrado. Aunque en las pruebas en condiciones seminaturales, se encontraron diferencias estadísticamente significativas entre las diferentes dosis y los diferentes tipos de criaderos, operacionalmente estas diferencias no tienen importancia práctica porque no aumenta el rendimiento de la medida de control. Los ensayos en condiciones naturales se realizaron aplicando una dosis de 1.050 AA/I a las albercas, toneles y llantas. El B. t. H-14 demostró ser efectivo, al causar 100% de mortalidad en todos los estadios larvarios de A. aegyptien los criaderos tratados. Se obtuvo una acción residual de 9 días en los criaderos cuando se aplicó polvo humectable y 7,2 días cuando se aplicó líquido concentrado. El uso del B. t. H-14. como larvicida alterno, tiene posibilidades en situaciones especiales, donde los métodos químicosson indeseadoso imprácticos.

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

    Science.gov (United States)

    Oppert, Brenda; Dowd, Scot E; Bouffard, Pascal; Li, Lewyn; Conesa, Ana; Lorenzen, Marcé D; Toutges, Michelle; Marshall, Jeremy; Huestis, Diana L; Fabrick, Jeff; Oppert, Cris; Jurat-Fuentes, Juan Luis

    2012-01-01

    Bacillus thuringiensis (Bt) crystal (Cry) proteins are effective against a select number of insect pests, but improvements are needed to increase efficacy and decrease time to mortality for coleopteran pests. To gain insight into the Bt intoxication process in Coleoptera, we performed RNA-Seq on cDNA generated from the guts of Tenebrio molitor larvae that consumed either a control diet or a diet containing Cry3Aa protoxin. Approximately 134,090 and 124,287 sequence reads from the control and Cry3Aa-treated groups were assembled into 1,318 and 1,140 contigs, respectively. Enrichment analyses indicated that functions associated with mitochondrial respiration, signalling, maintenance of cell structure, membrane integrity, protein recycling/synthesis, and glycosyl hydrolases were significantly increased in Cry3Aa-treated larvae, whereas functions associated with many metabolic processes were reduced, especially glycolysis, tricarboxylic acid cycle, and fatty acid synthesis. Microarray analysis was used to evaluate temporal changes in gene expression after 6, 12 or 24 h of Cry3Aa exposure. Overall, microarray analysis indicated that transcripts related to allergens, chitin-binding proteins, glycosyl hydrolases, and tubulins were induced, and those related to immunity and metabolism were repressed in Cry3Aa-intoxicated larvae. The 24 h microarray data validated most of the RNA-Seq data. Of the three intoxication intervals, larvae demonstrated more differential expression of transcripts after 12 h exposure to Cry3Aa. Gene expression examined by three different methods in control vs. Cry3Aa-treated larvae at the 24 h time point indicated that transcripts encoding proteins with chitin-binding domain 3 were the most differentially expressed in Cry3Aa-intoxicated larvae. Overall, the data suggest that T. molitor larvae mount a complex response to Cry3Aa during the initial 24 h of intoxication. Data from this study represent the largest genetic sequence dataset for T. molitor

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

    Directory of Open Access Journals (Sweden)

    Brenda Oppert

    Full Text Available Bacillus thuringiensis (Bt crystal (Cry proteins are effective against a select number of insect pests, but improvements are needed to increase efficacy and decrease time to mortality for coleopteran pests. To gain insight into the Bt intoxication process in Coleoptera, we performed RNA-Seq on cDNA generated from the guts of Tenebrio molitor larvae that consumed either a control diet or a diet containing Cry3Aa protoxin. Approximately 134,090 and 124,287 sequence reads from the control and Cry3Aa-treated groups were assembled into 1,318 and 1,140 contigs, respectively. Enrichment analyses indicated that functions associated with mitochondrial respiration, signalling, maintenance of cell structure, membrane integrity, protein recycling/synthesis, and glycosyl hydrolases were significantly increased in Cry3Aa-treated larvae, whereas functions associated with many metabolic processes were reduced, especially glycolysis, tricarboxylic acid cycle, and fatty acid synthesis. Microarray analysis was used to evaluate temporal changes in gene expression after 6, 12 or 24 h of Cry3Aa exposure. Overall, microarray analysis indicated that transcripts related to allergens, chitin-binding proteins, glycosyl hydrolases, and tubulins were induced, and those related to immunity and metabolism were repressed in Cry3Aa-intoxicated larvae. The 24 h microarray data validated most of the RNA-Seq data. Of the three intoxication intervals, larvae demonstrated more differential expression of transcripts after 12 h exposure to Cry3Aa. Gene expression examined by three different methods in control vs. Cry3Aa-treated larvae at the 24 h time point indicated that transcripts encoding proteins with chitin-binding domain 3 were the most differentially expressed in Cry3Aa-intoxicated larvae. Overall, the data suggest that T. molitor larvae mount a complex response to Cry3Aa during the initial 24 h of intoxication. Data from this study represent the largest genetic sequence

  7. Potencial para desarrollar tolerancia a las deltaendotoxinas de Bacillus thuringiensis subsp. kurstaki en Plutella xylostella (Lepidoptera: Plutellidae

    Directory of Open Access Journals (Sweden)

    M. A. Silva-Flores

    2005-01-01

    Full Text Available El presente estudio demuestra el potencial de Plutella xylostella L. para desarrollar tolerancia a las entomotoxinas de Bacillus thuringiensis subsp. kurstaki (Btk. Las poblaciones bajo estudio fueron una susceptible y otra sometida a presión de selección. La primera se crió durante ocho generaciones sin ningún tratamiento de insecticida, a la novena generación se le estimó la concentración letal mediana y la 65; con la segunda se inició la selección de la población tratada con Btk. Cuatro generaciones continuas de la población resistente se expusieron a la concentración de Btk correspondiente a la CL65 estimada en la población susceptible de P. xylostella. La entomotoxina utilizada en el experimento correspondió a Btk formulada al 6.4 % (Javelin® WG, Novartis México. En los bioensayos, se cortaron discos de hojas de col (Brassica oleracea L. de 90 mm de diámetro, se sumergieron durante 10 segundos en diferentes concentraciones de entomotoxinas y en cajas petri se infestaron con larvas de tercer instar; se trataron 10 larvas por concentración con tres repeticiones respectivas, el porcentaje de mortalidad se cuantificó 48 horas después. El análisis de los datos se realizó asumiendo el modelo Probit, mediante regresión lineal y usando el programa PcProbit; con él se estimó la CL50, CL65, y la CL95. Además se calculó la resistencia relativa. Los resultados indican que el porcentaje de mortalidad de P. xylostella disminuye más del 50 % después de ser sometida a presión de selección y el factor de tolerancia obtenido fue de aproximadamente 14 veces

  8. Eficiência e Persistência de Três Produtos Comerciais à Base de Bacillus thuringiensis israelensis e Bacillus sphaericus no controle de Culicidae (Diptera em Lagoas de Tratamento de Efluentes

    Directory of Open Access Journals (Sweden)

    Jose Lopes

    2010-11-01

    Abstract. The hematofagic effect caused by females belonging to some species of Culicidae on humans and animals can be directly related to pathogen transmission, allergic reactions and uneasiness. The emergence of populations resistant to chemical insecticides has fostered the use of alternative methods, mainly biological control. The trials were conducted in three effluent treatment lagoons, on larvae of Culicidae to test the efficiency and persistence of commercial products whose active principles are based on Bacillus thuringiensis israelensis Berliner and Bacillus sphaericus Neide. The products tested were Vectolex (a granulated formulation of B. sphaericus, Sphaericus (a liquid formulation of B. sphaericus and Bt-horus (a liquid formulation of B. thuringiensis. The products were applied biweekly and evaluations were conducted 0, 24, 48, 72, and 120 hours after each application. The lagoons were colonized by Culex nigripalpus Theobald (1.5%, Culex saltanensis Dyar (2.25%, and Culex quinquefasciatus Say (96.25%. Bt-horus reduced larvae by 89.06%, 83.97% and 89.96% at 24, 48 and 72 hours after product application, respectively. The granulated and the liquid formulations containing B. sphaericus reduced larvae by 98.89 % and 98.34% 24 hours after application, and by 99.79% and 99.78% after 48 hours, respectively. The products and the different formulations were effective in controlling larvae of all three Culicidae species in lagoons with high levels of organic matter, but the persistence was recorded in two and three days for products containing respectively B. sphaericus and B. thuringiensis israelensis.

  9. Eficiência do Bacillus thuringiensis var. kurstaki (Berliner, 1915 no controle da traça da cera Galleria mellonella (Linnaeus, 1758 (Lepidoptera: Pyralidae Efficiency of Bacillus thuringiensis var. kurstaki (Berliner, 1915 for control of the greater wax moth Galleria mellonella (Linnaeus, 1758 (Lepidoptera: Pyralidae

    Directory of Open Access Journals (Sweden)

    Deodoro Magno Brighenti

    2005-02-01

    Full Text Available Objetivou-se avaliar a eficiência do Bacillus thuringiensis var. kurstaki (Berliner no controle de Galleria mellonella (Linnaeus. Os experimentos foram realizados no Laboratório de Biologia de Insetos do Departamento de Entomologia da Universidade Federal de Lavras - UFLA, Lavras, MG, a 28±2ºC, UR 70±10% e fotofase de 12 horas. Aplicou-se formulação comercial de B. thuringiensis var. kurstaki por meio de pulverização, imersão dos favos e também foi incorporada à dieta artificial fornecida às lagartas de terceiro ínstar da traça da cera. A aplicação do produto fitossanitário por meio da pulverização dos favos mostrou-se eficiente no controle de lagartas, atingindo níveis iguais ou superiores a 85% de mortalidade quando foram utilizados 5 g/100 mL de água. Na aplicação por imersão dos favos, todas as dosagens testadas foram eficientes atingindo até 100% de mortalidade. Adicionada à dieta artificial, a concentração com maior porcentagem de mortalidade foi de 10g/60g de dieta. Sintomas de infecção pelo B. thuringiensis foram identificados nas lagartas e o isolamento da bactéria, por meio de uma cultura de Bacillus, comprovou a causa da morte desses insetos em todas as dosagens da formulação comercial do B. thuringiensis var. kurstaki utilizadas.This research aim at evaluating the efficiency of Bacillus thruringiensis var. kurstaki (Berliner in the control of Galleria mellonella (Linnaeus.The experiments were conducted in the Insect Biology Laboratory of the Entomology Department of the Universidade Federal de Lavras - UFLA, Lavras, MG, Brazil, at 28±2ºC, RH 70±10% and 12-hour photophase. Commercial formulation of B. thuringiensis. var. kurstaki was applied through spraying, comb soaking and also incorporated into the artificial diet fed to third instar caterpillars of the greater wax moth. Application of B. thuringiensis var. kurstaki through the spraying of the combs was efficient for control, reaching levels

  10. Recombinant Bacillus thuringiensis subsp. kurstaki HD73 strain that synthesizes Cry1Ac and chimeric ChiA74∆sp chitinase inclusions.

    Science.gov (United States)

    González-Ponce, Karen S; Casados-Vázquez, Luz E; Salcedo-Hernández, Rubén; Bideshi, Dennis K; Del Rincón-Castro, María C; Barboza-Corona, José E

    2017-05-01

    In this study, the endochitinase chiA74 gene lacking its secretion signal peptide sequence (chiA74∆sp) was fused in frame with the sequence coding for the C-terminal crystallization domain and transcription terminator of cry1Ac. The chimeric gene was expressed under the strong pcytA-p/STAB-SD promoter system in an acrystalliferous Cry - B strain of Bacillus thuringiensis and B. thuringiensis subsp. kurstaki HD73. We showed that the chimeric ChiA74∆sp produced amorphous inclusions in both Cry - B and HD73. In addition to the amorphous inclusions putatively composed of the chimera, bipyramidal Cry1Ac crystals, smaller than the wild-type crystal, were observed in recombinant HD73, and chitinase activity was remarkably higher (75-fold) in this strain when compared with parental HD73. Moreover, we observed that lyophilized samples of a mixture containing Cry1Ac, amorphous inclusions, and spores maintained chitinase activity. Amorphous inclusions could not be separated from Cry1Ac crystals by sucrose gradient centrifugation. Interestingly, the chitinase activity of purified Cry1Ac/amorphous inclusions was 51-fold higher compared to purified Cry1Ac inclusions of parental HD73, indicating that the increased enzymatic activity was due primarily to the presence of the atypical amorphous component. The possibility that the chimera is occluded with the Cry1Ac crystal, thereby contributing to the increased endochitinolytic activity, cannot be excluded. Finally, bioassays against larvae of Spodoptera frugiperda with spore/crystals of HD73 or spore-crystal ChiA74∆sp chimeric inclusions of recombinant HD73 strain showed LC 50 s of 396.86 and 290.25 ng/cm 2 , respectively. Our study suggests a possible practical application of the chimera in formulations of B. thuringiensis-based lepidopteran larvicides.

  11. Purification and characterization of the bacteriocin Thuricin Bn1 produced by Bacillus thuringiensis subsp. kurstaki Bn1 isolated from a hazelnut pest.

    Science.gov (United States)

    Ugras, Serpil; Sezen, Kazim; Kati, Hatice; Demirbag, Zihni

    2013-02-01

    A novel bioactive molecule produced by Bacillus thuringiensis subsp. kurstaki Bn1 (Bt-Bn1), isolated from a common pest of hazelnut, Balaninus nucum L. (Coleoptera: Curculionidae), was determined, purified, and characterized in this study. The Bt-Bn1 strain was investigated for antibacterial activity with an agar spot assay and well diffusion assay against B. cereus, B. weinhenstephenensis, L. monocytogenes, P. savastanoi, P. syringae, P. lemoignei, and many other B. thuringiensis strains. The production of bioactive molecule was determined at the early logarithmic phase in the growth cycle of strain Bt-Bn1 and its production continued until the beginning of the stationary phase. The mode of action of this molecule displayed bacteriocidal or bacteriolytic effect depending on the concentration. The bioactive molecule was purified 78-fold from the bacteria supernatant with ammonium sulfate precipitation, dialysis, ultrafiltration, gel filtration chromatography, and HPLC, respectively. The molecular mass of this molecule was estimated via SDS-PAGE and confirmed by the ESI-TOFMS as 3,139 Da. The bioactive molecule was also determined to be a heat-stable, pH-stable (range 6-8), and proteinase K sensitive antibacterial peptide, similar to bacteriocins. Based on all characteristics determined in this study, the purified bacteriocin was named as thuricin Bn1 because of the similarities to the previously identified thuricin-like bacteriocin produced by the various B. thuringiensis strains. Plasmid elution studies showed that gene responsible for the production of thuricin Bn1 is located on the chromosome of Bt-Bn1. Therefore, it is a novel bacteriocin and the first recorded one produced by an insect originated bacterium. It has potential usage for the control of many different pathogenic and spoilage bacteria in the food industry, agriculture, and various other areas.

  12. 78 FR 69849 - Issuance of an Experimental Use Permit

    Science.gov (United States)

    2013-11-21

    ... http://www.regulations.gov or at the Office of Pesticide Programs Regulatory Public Docket (OPP Docket... Dv49 double stranded RNA (dsRNA) suppression cassette in combination with Bacillus thuringiensis (Bt....105, Cry2Ab2, Vip3Aa20, Dv49 dsRNA, Cry3Bb1, Cry34Ab1, Cry35Ab1, eCry3.1Ab, respectively) are to be...

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

    International Nuclear Information System (INIS)

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

    2007-01-01

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

  14. Combined remediation of Cd-phenanthrene co-contaminated soil by Pleurotus cornucopiae and Bacillus thuringiensis FQ1 and the antioxidant responses in Pleurotus cornucopiae.

    Science.gov (United States)

    Jiang, Juan; Liu, Hongying; Li, Qiao; Gao, Ni; Yao, Yuan; Xu, Heng

    2015-10-01

    Remediation of soil co-contaminated with heavy metals and PAHs by mushroom and bacteria is a novel technique. In this study, the combined remediation effect of mushroom (Pleurotus cornucopiae) and bacteria (FQ1, Bacillus thuringiensis) on Cd and phenanthrene co-contaminated soil was investigated. The effect of bacteria (B. thuringiensis) on mushroom growth, Cd accumulation, phenanthrene degradation by P. cornucopiae and antioxidative responses of P. cornucopiae were studied. P. cornucopiae could adapt easily and grow well in Cd-phenanthrene co-contaminated soil. It was found that inoculation of FQ1 enhanced mushroom growth (biomass) and Cd accumulation with the increment of 26.68-43.58% and 14.29-97.67% respectively. Up to 100% and 95.07% of phenanthrene were removed in the bacteria-mushroom (B+M) treatment respectively spiked with 200mg/kg and 500mg/kg phenanthrene. In addition, bacterial inoculation alleviated oxidative stress caused by co-contamination with relative decreases in lipid peroxidation and enzyme activity, including malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD). This study demonstrated that the integrated remediation strategy of bacteria and mushroom is an effective and promising method for Cd-phenanthrene co-contaminated soil bioremediation. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Construction and characterization of the interdomain chimeras using Cry11Aa and Cry11Ba from Bacillus thuringiensis and identification of a possible novel toxic chimera.

    Science.gov (United States)

    Sun, Yunjun; Zhao, Qiang; Zheng, Dasheng; Ding, Xuezhi; Wang, Jingfang; Hu, Quanfang; Yuan, Zhiming; Park, Hyun-Woo; Xia, Liqiu

    2014-01-01

    Three structural domains of mosquitocidal Cry11Aa and Cry11Ba from Bacillus thuringiensis were exchanged to produce interdomain chimeras [BAA (11Ba/11Aa/11Aa), ABA (11Aa/11Ba/11Aa), AAB (11Aa/11Aa/11Ba), ABB (11Aa/11Ba/11Ba), BAB (11Ba/11Aa/11Ba), BBA (11Ba/11Ba/11Aa]. Chimeras BAB, BAA, BBA, and AAB formed inclusion bodies in the crystal-negative B. thuringiensis host and produced expected protein bands on SDS-PAGE gel. However, no inclusion body or target protein could be found for chimeras ABA and ABB. In bioassays using the fourth-instar larvae of Culex quinquefasciatus and Aedes aegypti, AAB had ~50 % lethal concentrations of 4.8 and 2.2 μg ml(-1), respectively; however, the rest of chimeras were not toxic. This study thus helps to understand the domain-function relationships of the Cry11Aa and Cry11Ba toxins. The toxic chimera, AAB, might be a candidate for mosquito control as its amino acid sequence is different from the two parental toxins.

  16. Three-dimensional (3D) structure prediction and function analysis of the chitin-binding domain 3 protein HD73_3189 from Bacillus thuringiensis HD73.

    Science.gov (United States)

    Zhan, Yiling; Guo, Shuyuan

    2015-01-01

    Bacillus thuringiensis (Bt) is capable of producing a chitin-binding protein believed to be functionally important to bacteria during the stationary phase of its growth cycle. In this paper, the chitin-binding domain 3 protein HD73_3189 from B. thuringiensis has been analyzed by computer technology. Primary and secondary structural analyses demonstrated that HD73_3189 is negatively charged and contains several α-helices, aperiodical coils and β-strands. Domain and motif analyses revealed that HD73_3189 contains a signal peptide, an N-terminal chitin binding 3 domains, two copies of a fibronectin-like domain 3 and a C-terminal carbohydrate binding domain classified as CBM_5_12. Moreover, analysis predicted the protein's associated localization site to be the cell wall. Ligand site prediction determined that amino acid residues GLU-312, TRP-334, ILE-341 and VAL-382 exposed on the surface of the target protein exhibit polar interactions with the substrate.

  17. Caracterização de novos isolados de Bacillus thuringiensis para o controle de importantes insetos-praga da agricultura

    Directory of Open Access Journals (Sweden)

    Emeline Boni Campanini

    2012-01-01

    Full Text Available A bactéria Bacillus thuringiensis Berliner produz um corpo de inclusão paraesporal (cristal de natureza proteica, formado durante a esporulação, que atua de forma eficiente no controle de insetos-praga de culturas economicamente importantes. Esse cristal é constituído de proteínas Cry, que são codificadas pelos genes cry; um isolado pode ser caracterizado pelo conteúdo de genes cry que apresenta. Visando caracterizar novos isolados no combate de insetos-praga pertencentes às ordens Lepidoptera e Coleoptera, 76 isolados bacterianos foram analisados molecularmente e tiveram seu potencial de controle avaliado por meio de bioensaios com larvas de Spodoptera frugiperda (J.E. Smith, Sphenophorus levis Vaurie e Tenebrio molitor Linnaeus. As análises moleculares indicaram 11 isolados (14,5% da coleção, contendo genes lepidóptero-específicos e 17 (22,37% com genes coleóptero-específicos. As análises de patogenicidade revelaram dois isolados com alto potencial de controle para lagartas de S. frugiperda, um para larvas de S. levis e seis prejudiciais ao desenvolvimento das larvas de T. molitor. Esses isolados de B. thuringiensis podem ser promissores no controle biológico das referidas pragas.

  18. A Tenebrio molitor GPI-anchored alkaline phosphatase is involved in binding of Bacillus thuringiensis Cry3Aa to brush border membrane vesicles.

    Science.gov (United States)

    Zúñiga-Navarrete, Fernando; Gómez, Isabel; Peña, Guadalupe; Bravo, Alejandra; Soberón, Mario

    2013-03-01

    Bacillus thuringiensis Cry toxins recognizes their target cells in part by the binding to glycosyl-phosphatidyl-inositol (GPI) anchored proteins such as aminopeptidase-N (APN) or alkaline phosphatases (ALP). Treatment of Tenebrio molitor brush border membrane vesicles (BBMV) with phospholipase C that cleaves out GPI-anchored proteins from the membranes, showed that GPI-anchored proteins are involved in binding of Cry3Aa toxin to BBMV. A 68 kDa GPI-anchored ALP was shown to bind Cry3Aa by toxin overlay assays. The 68 kDa GPI-anchored ALP was preferentially expressed in early instar larvae in comparison to late instar larvae. Our work shows for the first time that GPI-anchored ALP is important for Cry3Aa binding to T. molitor BBMV suggesting that the mode of action of Cry toxins is conserved in different insect orders. Copyright © 2012 Elsevier Inc. All rights reserved.

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

    International Nuclear Information System (INIS)

    Wu Dianxing; Ye Qingfu; Wang Zhonghua; Xia Yingwu

    2004-01-01

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

  20. [Susceptibility of populations of Simulium (Chirostilbia) pertinax Kollar, 1832 (Culicomorpha, Simuliidae) to eemephos and to Bacillus thuringiensis var. israelensis-based formulation].

    Science.gov (United States)

    de Andrade, C F; Castello Branco Júnior, A

    1991-10-01

    The use of wooden troughs on stream beds, artificially colonized by blackfly larvae, is proposed for larvicide evaluations. Mortality was recorded 3 or 4 hours after treatment. Larval susceptibility was also evaluated utilizing the LT50 criterion. In there field assays Simulium (C.) pertinax populations from the litoral of S. Paulo and Rio de Janeiro States were shown to be resistant to temephos, even when subjected to high concentrations. Vectobac 12 AS, a Bacillus thuringiensis var. israelensis product, was shown to be more potent against late instar larvae and efficient in concentrations higher than 7,200 ITU/l (10 min). The LT50 to 3,744 ITU/l (10 min) was calculated as 70.9 min.

  1. Detection of the mosquitocidal toxin genes encoding Cry11 proteins from Bacillus thuringiensis using a novel PCR-RFLP method Detección de genes que codifican proteínas mosquitocidas Cry11 de Bacillus thuringiensis mediante un método de PCR-RFLP novedoso

    Directory of Open Access Journals (Sweden)

    D. H. Sauka

    2010-02-01

    Full Text Available A polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP method for detection of cry11 genes from Bacillus thuringiensis was established. Based on the analysis of conserved regions of the cry11 genes, 2 oligonucleotide primers were designed to amplify a 1459-bp fragment of the cry11Aa gene, and a 1471-bp of the cry11Ba and cry11Bb genes. The amplification products were digested with restriction endonuclease HinfI. Exotic B. thuringiensis strains and native isolates collected from soils, leaves and stored product dust of Argentina were analyzed to study the distribution of cry11 genes. The PCR-RFLP patterns revealed the detection of cry11 genes in 3 of 64 exotic strains and in 10 of 107 native B. thuringiensis isolates tested. Just the cry11Aa gene subclass was detected among these bacteria. Since the methodology was also developed to detect cry11Ba and cry11Bb genes, an experimental future confirmation will be required. Based on the results obtained, the PCR-RFLP method presented may be a valuable tool for specific detection of the mosquitocidal toxin genes encoding Cry11 proteins from B. thuringiensis.En el presente estudio se estableció una estrategia basada en la amplificación génica (PCR y el posterior análisis de restricción (RFLP para detectar todos los genes cry11 de Bacillus thuringiensis informados hasta ahora. De acuerdo con el análisis de las regiones conservadas en los genes cry11, se diseñaron dos cebadores para amplificar un fragmento de 1459 pb de los genes cry11Aa y un fragmento de 1471 pb de los genes cry11Ba y cry11Bb. Los productos de la amplificación fueron digeridos con la enzima de restricción HinfI. Se analizaron cepas exóticas de B. thuringiensis y aislamientos nativos de Argentina obtenidos a partir de muestras de suelos, hojas y polvillo de silos, para estudiar la distribución de los genes cry11. Los patrones de PCR-RFLP revelaron la presencia de genes cry11 en 3 de las 64 cepas ex

  2. Routine implementation costs of larviciding with Bacillus thuringiensis israelensis against malaria vectors in a district in rural Burkina Faso.

    Science.gov (United States)

    Dambach, Peter; Schleicher, Michael; Stahl, Hans-Christian; Traoré, Issouf; Becker, Norbert; Kaiser, Achim; Sié, Ali; Sauerborn, Rainer

    2016-07-22

    The key tools in malaria control are early diagnosis and treatment of cases as well as vector control. Current strategies for malaria vector control in sub-Saharan Africa are largely based on long-lasting insecticide-treated nets (LLINs) and to a much smaller extent on indoor residual spraying (IRS). An additional tool in the fight against malaria vectors, larval source management (LSM), has not been used in sub-Saharan Africa on a wider scale since the abandonment of environmental spraying of DDT. Increasing concerns about limitations of LLINs and IRS and encouraging results from large larvicide-based LSM trials make a strong case for using biological larviciding as a complementary tool to existing control measures. Arguments that are often quoted against such a combined approach are the alleged high implementation costs of LSM. This study makes the first step to test this argument. The implementation costs of larval source management based on Bacillus thuringiensis israelensis (Bti) (strain AM65-52) spraying under different implementation scenarios were analysed in a rural health district in Burkina Faso. The analysis draws on detailed cost data gathered during a large-scale LSM intervention between 2013 and 2015. All 127 villages in the study setup were assigned to two treatment arms and one control group. Treatment either implied exhaustive spraying of all available water collections or targeted spraying of the 50 % most productive larval sources via remote-sensing derived and entomologically validated risk maps. Based on the cost reports from both intervention arms, the per capita programme costs were calculated under the assumption of covering the whole district with either intervention scenario. Cost calculations have been generalized by providing an adaptable cost formula. In addition, this study assesses the sensitivity of per capita programme costs with respect to changes in the underlying cost components. The average annual per capita costs of

  3. Effect Bacillus thuringiensis gekwantificeerd

    NARCIS (Netherlands)

    Elderson, J.; Belder, den E.

    2001-01-01

    PRI onderzocht in veldproeven de effecten van het biologische middel Dipel WP voor de bestrijding van rupsen (koolmot e.a.) in de biologische spruitkoolteelt. Na behandeling loopt het aantal koolmotten fors terug; de opbrengst van de behandelde en onbehandelde velden was gelijk, maar de kwaliteit

  4. Sequence Analysis of Inducible Prophage phIS3501 Integrated into the Haemolysin II Gene of Bacillus thuringiensis var israelensis ATCC35646

    Directory of Open Access Journals (Sweden)

    Bouziane Moumen

    2012-01-01

    Full Text Available Diarrheic food poisoning by bacteria of the Bacillus cereus group is mostly due to several toxins encoded in the genomes. One of them, cytotoxin K, was recently identified as responsible for severe necrotic syndromes. Cytotoxin K is similar to a class of proteins encoded by genes usually annotated as haemolysin II (hlyII in the majority of genomes of the B. cereus group. The partially sequenced genome of Bacillus thuringiensis var israelensis ATCC35646 contains several potentially induced prophages, one of them integrated into the hlyII gene. We determined the complete sequence and established the genomic organization of this prophage-designated phIS3501. During induction of excision of this prophage with mitomycin C, intact hlyII gene is formed, thus providing to cells a genetic ability to synthesize the active toxin. Therefore, this prophage, upon its excision, can be implicated in the regulation of synthesis of the active toxin and thus in the virulence of bacterial host. A generality of selection for such systems in bacterial pathogens is indicated by the similarity of this genetic arrangement to that of Staphylococcus aureus  β-haemolysin.

  5. Cry1A(b)16 toxin from Bacillus thuringiensis: Theoretical refinement of three-dimensional structure and prediction of peptides as molecular markers for detection of genetically modified organisms.

    Science.gov (United States)

    Plácido, Alexandra; Coelho, Andreia; Abreu Nascimento, Lucas; Gomes Vasconcelos, Andreanne; Fátima Barroso, Maria; Ramos-Jesus, Joilson; Costa, Vladimir; das Chagas Alves Lima, Francisco; Delerue-Matos, Cristina; Martins Ramos, Ricardo; Marani, Mariela M; Roberto de Souza de Almeida Leite, José

    2017-07-01

    Transgenic maize produced by the insertion of the Cry transgene into its genome became the second most cultivated crop worldwide. Cry gene from Bacillus thuringiensis kurstaki expresses protein derivatives of crystalline endotoxins which confer insect resistance onto the maize crop. Mandatory labeling of processed food containing or made by genetically modified organisms is in force in many countries, so, it is very urgent to develop fast and practical methods for GMO identification, for example, biosensors. In the absence of an available empirical structure of Cry1A(b)16 protein, a theoretical model was effectively generated, in this work, by homology modeling and molecular dynamics simulations based on two available homologous protein structures. Molecular dynamics simulations were carried out to refine the selected model, and an analysis of its global structure was performed. The refined models of Cry1A(b)16 showed a standard fold and structural characteristics similar to those seen in Bacillus thuringiensis Cry1A(a) insecticidal toxin and Bacillus thuringiensis serovar kurstaki Cry1A(c) toxin. After in silico analysis of Cry1A(b)16, two immunoreactive candidate peptides were selected and specific polyclonal antibodies were produced resulting in antibody-peptide interaction. Biosensing devices are expected to be developed for detection of the Cry1A(b) protein as a marker of transgenic maize in food. Proteins 2017; 85:1248-1257. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  6. Use of by-products rich in carbon and nitrogen as a nutrient source to produce Bacillus thuringiensis (Berliner)-based bio pesticide

    International Nuclear Information System (INIS)

    Valicente, Fernando H.; Mourao, Andre H.C.

    2008-01-01

    The amount and sources of carbon and nitrogen used to produce Bacillus thuringiensis (Berliner)-based biopesticide may influence the quality of the fi nal product. The objective of this research was to test different levels of carbon and nitrogen: medium 1 - 1.5% maize glucose + 0.5% soy fl our, medium 2 - 3.0% maize glucose + 1.0% soy flour, medium 3 - 1.0% maize glucose + 3.0% soy fl our and medium 4 - Luria Bertani (LB) + salts (FeSO 4 , ZnSO 4 , MnSO 4 , MgSO 4 ). The seed culture was produced in LB medium plus salt, under agitation (200 rpm) for 18h at 30 deg C. The strain 344 of Bt was used (B. thuringiensis var tolworthi - belonging to the EMBRAPA's Bt Bank). The pH was measured at regular intervals, and After culturing for 96h, the pH of the four tested media was basified (6.91 and 8.15), the number of spores yielded 4.39 x 10 9 spores/ml in medium 3, where the amount of protein is high. The dry biomass weight accumulated in media 3 was 39.3 g/l. Mortality of 2-day-old larvae Spodoptera frugiperda (J.E. Smith) was 100% when using Bt produced in media 3 and 4. CL 50 for medium 3 was 8.4 x 10 6 spores/ml. All tested media were satisfactory to Bt growth, and medium 3 was the most promising to be used on a large scale Bt-based biopesticide production. (author)

  7. Characterization of the synergistic interaction between Beauveria bassiana strain GHA and Bacillus thuringiensis morrisoni strain tenebrionis applied against Colorado potato beetle larvae.

    Science.gov (United States)

    Wraight, S P; Ramos, M E

    2017-03-01

    Studies were undertaken to further characterize the previously identified synergistic activity of Bacillus thuringiensis- and Beauveria bassiana-based biopesticides against Colorado potato beetle (CPB). A flowable concentrate of B. thuringiensis morrisoni strain tenebrionis (Bt) (Novodor® FC) and a wettable powder of B. bassiana strain GHA (Bb) (Mycotrol® 22WP) were applied against CPB larval populations infesting potato in field plots. Novodor FC and an oil-dispersion formulation of Bb (Mycotrol ES) were applied against second-instar CPB larvae on potted potato plants in greenhouse tests under low relative humidity (RH), variable-temperature conditions. Each pathogen was applied alone and in combination (tank-mixed) with the other pathogen. In the field tests, each biopesticide was also combined with the spray-carrier (formulation without active ingredient) of the other pathogen. Results from the greenhouse tests showed that under warm, dry conditions, low activity of Mycotrol was counterbalanced by high activity of the Novodor, and under cool, somewhat more humid conditions, low Novodor activity was balanced by high activity of Mycotrol, with the result being a constant level of synergism (CPB mortality ca. 20 percentage points higher than predicted by independent action). Similar levels of synergism were observed under the markedly different conditions of the field and greenhouse environments, and the synergism was confirmed as arising from interaction of the two micobes, as the Bt spray carrier had no significant effect on efficacy of the Mycotrol product and the Bb spray carrier had no effect on the efficacy of Novodor. The great capacity of these two control agents to act in concert to control CPB is well documented (the fast-acting, toxic Bt acting to protect potato crops from defoliation and the slow-acting Bb reducing survival to the adult stage). These finding further underscore the strong complementary action of these agents applied jointly against CPB

  8. Seleção e caracterização de estirpes de Bacillus thuringiensis eficientes contra a Diatraea saccharalis (Lepidoptera: Crambidae

    Directory of Open Access Journals (Sweden)

    Cristina Lima de Macedo

    2012-12-01

    Full Text Available O objetivo deste trabalho foi selecionar e caracterizar estirpes nativas de Bacillus thuringiensis tóxicas a Diatraea saccharalis (Lepidoptera: Crambidae. Cento e seis estirpes pertencentes ao Banco de Bactérias de Invertebrados, da Embrapa Recursos Genéticos e Biotecnologia, foram testadas quanto à toxicidade a D. saccharalis, e, as mais tóxicas, caracterizadas por métodos bioquímicos e moleculares. Das 106 estirpes testadas, 16 causaram 100% de mortalidade em 24 horas. As três estirpes mais tóxicas apresentaram concentração letal média entre 8 e 43 ng cm-2. O perfil proteico das 16 estirpes mostrou a presença de proteínas de 130 e 65 kDa, e a caracterização molecular mostrou a presença dos genes tipo cry1 e cry2: cry1Aa, cry1Ab, cry1Ac e cry2Aa. A concentração letal média de esporos e cristais obtidos a partir de estirpes recombinantes, que expressavam individualmente os genes cry1Aa, cry1Ab, cry1Ac e cry2Aa, variou entre 222 e 610 ng cm-2, valores muito superiores aos das estirpes nativas mais tóxicas, que apresentavam possibilidade de expressão simultânea desses genes. Este resultado é indicativo de que há sinergia entre as toxinas. Há interação entre as toxinas de B. thuringiensis e seus receptores na broca-do-colmo da cana-de-açúcar.

  9. Use of by-products rich in carbon and nitrogen as a nutrient source to produce Bacillus thuringiensis (Berliner)-based bio pesticide

    Energy Technology Data Exchange (ETDEWEB)

    Valicente, Fernando H. [EMBRAPA Milho e Sorgo, Sete Lagoas, MG (Brazil)]. E-mail: valicent@cnpms.embrapa.br; Mourao, Andre H.C. [Curso de Meio Ambiente, Sete Lagoas, MG (Brazil)

    2008-11-15

    The amount and sources of carbon and nitrogen used to produce Bacillus thuringiensis (Berliner)-based biopesticide may influence the quality of the fi nal product. The objective of this research was to test different levels of carbon and nitrogen: medium 1 - 1.5% maize glucose + 0.5% soy fl our, medium 2 - 3.0% maize glucose + 1.0% soy flour, medium 3 - 1.0% maize glucose + 3.0% soy fl our and medium 4 - Luria Bertani (LB) + salts (FeSO{sub 4}, ZnSO{sub 4}, MnSO{sub 4}, MgSO{sub 4}). The seed culture was produced in LB medium plus salt, under agitation (200 rpm) for 18h at 30 deg C. The strain 344 of Bt was used (B. thuringiensis var tolworthi - belonging to the EMBRAPA's Bt Bank). The pH was measured at regular intervals, and After culturing for 96h, the pH of the four tested media was basified (6.91 and 8.15), the number of spores yielded 4.39 x 10{sup 9} spores/ml in medium 3, where the amount of protein is high. The dry biomass weight accumulated in media 3 was 39.3 g/l. Mortality of 2-day-old larvae Spodoptera frugiperda (J.E. Smith) was 100% when using Bt produced in media 3 and 4. CL{sub 50} for medium 3 was 8.4 x 10{sup 6} spores/ml. All tested media were satisfactory to Bt growth, and medium 3 was the most promising to be used on a large scale Bt-based biopesticide production. (author)

  10. Development of a Low Cost Bioprocess for Endotoxin Production by Bacillus thuringiensis var israelensis Intended for Biological Control of Aedes aegypti

    Directory of Open Access Journals (Sweden)

    Carlos Ricardo Soccol

    2009-11-01

    Full Text Available Aedes aegypti is the vector of Dengue disease, responsible for 20,000 deaths/year worldwide. Bacillus thuringiensis var israelensis - Bti releases selective and effective toxins (crystal proteins against A. aegypti larvae. We present a low cost bioprocess for toxin production, accomplished by a selected Brazilian strain Bti (BR-LPB01 and employment of low cost substrates. Soybean meal and sugarcane molasses lead to high toxic effectiveness after 2L bioreactor fermentation (LD50=26ng/mL, near to the reference strain IPS82 (LD50=17.3 ng/mL. The pH ranged between 5.8 and 7.0 during the exponential growth period and between 7.0 and 8.4 during the stationary phase, with low activity. Thus, control of foam and pH 7.0 were started and proved to be crucial for high activity. It was verified that the fermentation could be discontinued after 20 hours, when the highest activity was present.A dengue é transmitida pelo Aedes aegypti, doença responsável por 20.000 mortes/ano no mundo. Bacillus thuringiensis var israelensis libera toxinas seletivas e eficazes (proteínas cristal contra larvas de A. aegypti. Propõe-se um bioprocesso de baixo custo para a produção da toxina, pelo emprego de uma cepa brasileira selecionada de Bti (BR-LPB01 e de substratos de baixo custo. Farelo de soja e melaço de cana levaram a eficácia tóxica alta após fermentação em biorreator 2L (DL50=26ng/mL, valor próximo a estirpe de referência IPS82 (DL50=17,3 ng/mL. O pH variou entre 5,8 e 7,0 durante o período de crescimento exponencial e entre 7,0 e 8,4 durante a fase estacionária, com baixa atividade larvicida. Assim, controles de espuma e de pH 7,0 foram iniciados e demonstraram serem cruciais para alta atividade. Verificou-se que a fermentação deve ser interrompida após vinte horas, quando se obtém a maior atividade.

  11. Cry64Ba and Cry64Ca, Two ETX/MTX2-Type Bacillus thuringiensis Insecticidal Proteins Active against Hemipteran Pests.

    Science.gov (United States)

    Liu, Yonglei; Wang, Yinglong; Shu, Changlong; Lin, Kejian; Song, Fuping; Bravo, Alejandra; Soberón, Mario; Zhang, Jie

    2018-02-01

    Genetically modified crops that express insecticidal Bacillus thuringiensis (Bt) proteins have become a primary approach for control of lepidopteran (moth) and coleopteran (beetle) pests that feed by chewing the plants. However, the sap-sucking insects (Hemiptera) are not particularly susceptible to Bt toxins. In this study, we describe two Cry toxins (Cry64Ba and Cry64Ca) from Bt strain 1012 that showed toxicity against two important hemipteran rice pests, Laodelphax striatellus and Sogatella furcifera Both of these proteins contain an ETX/MTX2 domain and share common sequence features with the β-pore-forming toxins. Coexpression of cry64Ba and cry64Ca genes in the acrystalliferous Bt strain HD73 - resulted in high insecticidal activity against both hemipteran pests. No toxicity was observed on other pests such as Ostrinia furnacalis , Plutella xylostella , or Colaphellus bowringi Also, no hemolytic activity or toxicity against cancer cells was detected. Binding assays showed specific binding of the Cry64Ba/Cry64Ca toxin complex to brush border membrane vesicles isolated from L. striatellus Cry64Ba and Cry64Ca are Bt Cry toxins highly effective against hemipteran pests and could provide a novel strategy for the environmentally friendly biological control of rice planthoppers in transgenic plants. IMPORTANCE In Asia, rice is an important staple food, whose production is threatened by rice planthoppers. To date, no effective Bacillus thuringiensis (Bt) protein has been shown to have activity against rice planthoppers. We cloned two Bt toxin genes from Bt strain 1012 that showed toxicity against small brown planthoppers ( Laodelphax striatellus ) and white-backed planthoppers ( Sogatella furcifera ). To our knowledge, the proteins encoded by the cry64Ba and cry64Ca genes are the most efficient insecticidal Bt Cry proteins with activity against hemipteran insects reported so far. Cry64Ba and Cry64Ca showed no toxicity against some lepidopteran or coleopteran pests

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2007-09-15

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

  13. Characterization of Bacillus thuringiensis Berl. indigenous from soil and its potency as biological agents of Spodoptera litura (Lepidoptera: Noctuidae)

    Science.gov (United States)

    Pujiastuti, Y.; Astuti, D. T.; Afriyani, S. R.; Suparman, S.; Irsan, C.; Sembiring, E. R.; Nugraha, S.; Mulawarman; Damiri, N.

    2018-01-01

    The objective of the study was to characterize the molecular weight of protein in order to be utilized as biological agent of S. litura and their cell or spores production. To investigate the molecular weight of protein was done by SDS-Page electrophoresis. Growth media used for producing B. thuringiensis were T3, LB broth and agricultural waste. The results showed that the molecular weight of protein ranged from 37 to 140 kDa. In DLM and DLKK23 isolates were found ranging from 37 to 40 kDa and from 110 to 130 kDa, respectively. KJ3R5 and KJ3P1 isolates were obtained having three protein bands ranging from 43 to 45, 73 to 80 and 110 to 130 kDa and 45-50, 75-80 and 130-140 kDa, respectively. It was predicted that isolates B. thuringiensis were belonging to Cry IA, Cry IIA, Cry IVC and Cry15c. These crystal proteins were toxic to S. litura. There was no protein bands found in the two last isolates (KJ3R3 and KJ3J4). Production of spore after sporulation in agricultural waste media ranged from 0.5 to 106 - 2.67 x 107 spores/ml showing medium level of toxicity to S. litura.

  14. Susceptibilidade de populações de Simulium (Chirostilbia pertinax Kollar, 1832 (Culicomorpha, Simuliidae ao temephos e a um formulado à base de Bacillus thuringiensis var. israelensis Susceptibility of population of Simulium (Chirostilbia pertinax Kollar, 1832 (Culicomorpha, Simuliidae to temephos and Bacillus thuringiensis var. israelensis formulation

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    Carlos Fernando S. de Andrade

    1991-10-01

    Full Text Available O uso de pequenas calhas de madeira no leito de riachos, manualmente colonizadas com borrachudos, é proposto para avaliações da eficiência de larvicidas 3 ou 4 horas após as aplicações. A susceptibilidade larval também foi avaliada pelo critério de TL50. Em três ensaios no litoral do Estado de São Paulo e Rio de Janeiro, populações de Simulium (C. pertinax Kollar, 1832 mostraram-se resistentes ao temephos, mesmo em altas concentrações. Vectobac 12 AS, à base de Bacillus thuringiensis var. israelensis mostrou-se mais potente contra larvas dos últimos estádios e eficiente para controle em concentrações a partir de 7.200 UIP/1 (10 min.. O TL50para 3.744 UIP/1 (10 min. foi de 70,9 minutos.The use of wooden troughs on stream beds, artificially colonized by blackfly larvae, is proposed for larvicide evaluations. Mortality was recorded 3 or 4 hours after treatment. Larval susceptibility was also evaluated utilizing the LT50 criterion. In there field assays Simulium (C. pertinax populations from the litoral of S. Paulo and Rio de Janeiro States were shown to be resistant to temephos, even when subjected to high concentrations. Vectobac 12 AS, a Bacillus thuringiensis var. israelensis product, was shown to be more potent against late instar larvae and efficient in concentrations higher than 7,200 ITU/1 (10 min. The LT50 to 3,744 ITU/1 (10 min was calculated as 70.9 min.

  15. Identificación de los Genes cry en Cepas Mexicanas de Bacillus thuringiensis con Potencial Insecticida Identificación de los Genes cry en Cepas Mexicanas de Bacillus thuringiensis con Potencial Insecticida

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    J. E. Ibarra

    2012-02-01

    Full Text Available En el presente estudio se determinó la morfología y la composición proteica de los cristales de las cepas LBIT-499 y LBIT-504 de B. thuringiensis, ambas nativas de Guanajuato, México. La primera mostró sólo cristales bipiramidales mientras que la segunda presentó cristales tanto bipiramidales como cúbicos. Estos cristales estuvieron compuestos de proteínas de aproximadamente 130 y 60 kDa, respectivamente. En ambas cepas, además de la LBIT-500 y LBIT-544, se detectó una gran variedad de genes cry1. En éstas últimas se encontraron los genes cry2A y cry2B, en la LBIT- 504 sólo el cry2B y en la LBIT-499 no se detectó ningún cry2. This report revealed the crystal morphology and protein composition of strains LBIT-499 and LBIT-504 of B. thuringiensis, native to Guanajuato, Mexico. LBIT-499 showed only bipyramidal crystals, while LBIT-504 showed both bipyramidal and cubical crystals. These crystals were composed by proteins of ca. 130 and 60 kDa, respectively. Both strains as well as LBIT-500 and LBIT-544, showed a great variety of cry1 genes, while cry2A and cry2B were identified in LBT-500 and LBIT-544, LBIT-504 showed only the cry2B gene, and no cry2 was detected in LBIT-499.

  16. Identification, cloning and lactonase activity of recombinant protein of N-acyl homoserine lactonase (AiiA from Bacillus thuringiensis 147-115-16 strain.

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    Alvaro Mauricio Florez Escobar

    2014-01-01

    Short title: N-acyl homoserine lactonase (AiiA from Bacillus thuringiensis  Abstract: The quorum-quenching N-acyl homoserine lactonases are a family of bacterial metalloenzymes that participate in degradation of N-acyl homoserine lactones (AHLs, disrupting the quorum sensing system of gram negative bacterial species. From a collection of Bacillus thuringiensis strains isolated in Colombia from plants and exhibiting toxic activity against lepidopteran insects, 310 bacterial isolates were tested to determine lactonase activity by using biosensor systems in presence of synthetic N-hexanoyl-L-homoserine lactone (C6-HSL and N-octanoyl-L-homoserine lactone (C8-HSL. From them, 251 strains showed degrading activity to both C6-HSL and C8-HSL, 57% exhibited degrading activity to C6-HSL and 43% to C8-HSL. One B. thuringiensis strain, denoted as 147-115-16, that exhibit high degrading activity to C6-HSL and C8-HSL, was able to attenuate soft rot symptoms in infected potato slices with Pectobacterium carotovorum. This strain contains an homologous of the aiiA gene that was cloned, sequenced and expressed in Esherichia coli DE3. The recombinant protein AiiA147-11516 displays activity to C6-HSL, C8-HSL, N-(β-ketocaproyl (3-O-C6-HSL and N-3-oxo-dodecanoyl (3-O-C12-HSL. The recombinant strain in the presence of P. caratovorum cultures was able to attenuate the infection, suggesting that it interferes either with the accumulation or with the response to the AHLs signals. Acording to this data and based on previous report from recombinant AiiA147-11516, this enzyme exhibits activity to a wide range of catalytic substrates suggesting its industrial application in the disease control programs through plants transformation. Key words: lactones, Quorum sensing, Quorum quenching, Lactonases, Pectobacterium caratovorum Resumen: Las N-acíl homoserina lactonasas son una familia de metaloenzimas bacterianas que participan en la degradación de N-acil homoserina lactonas (AHLs

  17. Large-Scale Purification, Characterization, and Spore Outgrowth Inhibitory Effect of Thurincin H, a Bacteriocin Produced by Bacillus thuringiensis SF361.

    Science.gov (United States)

    Wang, Gaoyan; Manns, David C; Guron, Giselle K; Churey, John J; Worobo, Randy W

    2014-06-01

    Large-scale purification of the highly hydrophobic bacteriocin thurincin H was accomplished via a novel and simple two-step method: ammonia sulfate precipitation and C18 solid-phase extraction. The inhibition spectrum and stability of thurincin H as well as its antagonistic activity against Bacillus cereus F4552 spores were further characterized. In the purification method, secreted proteins contained in the supernatant of a 40 h incubated culture of B. thuringiensis SF361 were precipitated by 68 % ammonia sulfate and purified by reverse-phase chromatography, with a yield of 18.53 mg/l of pure thurincin H. Silver-stained SDS-PAGE, high-performance liquid chromatography, and liquid chromatography-mass spectrometry confirmed the high purity of the prepared sample. Thurincin H exhibited a broad antimicrobial activity against 22 tested bacterial strains among six different genera including Bacillus, Carnobacterium, Geobacillus, Enterococcus, Listeria, and Staphylococcus. There was no detectable activity against any of the selected yeast or fungi. The bacteriocin activity was stable for 30 min at 50 °C and decreased to undetectable levels within 10 min at temperatures above 80 °C. Thurincin H is also stable from pH 2-7 for at least 24 h at room temperature. Thurincin H is germicidal against B. cereus spores in brain heart infusion broth, but not in Tris-NaCl buffer. The efficient purification method enables the large-scale production of pure thurincin H. The broad inhibitory spectrum of this bacteriocin may be of interest as a potential natural biopreservative in the food industry, particularly in post-processed and ready-to-eat food.

  18. Intraguild competition and enhanced survival of western bean cutworm (Lepidoptera: Noctuidae) on transgenic Cry1Ab (MON810) Bacillus thuringiensis corn.

    Science.gov (United States)

    Dorhout, David L; Rice, Marlin E

    2010-02-01

    The effect of genetically modified corn (event MON810, YieldGard Corn Borer) expressing the Bacillus thuringiensis sp. kurstaki (Berliner) (Bt) endotoxin, Cry1Ab, on the survival of western bean cutworm, Striacosta albicosta (Smith), larvae was examined during intraguild competition studies with either European corn borer, Ostrinia nubilalis (Hübner), or corn earworm, Helicoverpa zea (Boddie), larvae. Competition scenarios were constructed by using either a laboratory or field competition arena containing one of five different diets and one of 13 different larval size-by-species scenarios. The survival of western bean cutworms competing with corn earworms in the laboratory arenas on either a meridic diet or isoline corn silk diet was significantly lower (P corn earworm on a Cry1Ab-MON810 corn silk diet was significant higher (P corn borers generally did not alter the outcomes observed in the western bean cutworm and corn earworm only two-way competitions. These data suggest that Cry1Ab-MON810 corn may confer a competitive advantage to western bean cutworm larvae during intraguild competition, particularly from corn earworms, and that western bean cutworms become equal competitors only when they are of equal or larger size and the diet is Cry1Ab-MON810 corn.

  19. Impacts of elevated CO2 on exogenous Bacillus thuringiensis toxins and transgene expression in transgenic rice under different levels of nitrogen.

    Science.gov (United States)

    Jiang, Shoulin; Lu, Yongqing; Dai, Yang; Qian, Lei; Muhammad, Adnan Bodlah; Li, Teng; Wan, Guijun; Parajulee, Megha N; Chen, Fajun

    2017-11-07

    Recent studies have highlighted great challenges of transgene silencing for transgenic plants facing climate change. In order to understand the impacts of elevated CO 2 on exogenous Bacillus thuringiensis (Bt) toxins and transgene expression in transgenic rice under different levels of N-fertilizer supply, we investigated the biomass, exogenous Bt toxins, Bt-transgene expression and methylation status in Bt rice exposed to two levels of CO 2 concentrations and nitrogen (N) supply (1/8, 1/4, 1/2, 1 and 2 N). It is elucidated that the increased levels of global atmospheric CO 2 concentration will trigger up-regulation of Bt toxin expression in transgenic rice, especially with appropriate increase of N fertilizer supply, while, to some extent, the exogenous Bt-transgene expression is reduced at sub-N levels (1/4 and 1/2N), even though the total protein of plant tissues is reduced and the plant growth is restricted. The unpredictable and stochastic occurrence of transgene silencing and epigenetic alternations remains unresolved for most transgenic plants. It is expected that N fertilization supply may promote the expression of transgenic Bt toxin in transgenic Bt rice, particularly under elevated CO 2 .

  20. Identification of Bacillus thuringiensis Cry3Aa toxin domain II loop 1 as the binding site of Tenebrio molitor cadherin repeat CR12.

    Science.gov (United States)

    Zúñiga-Navarrete, Fernando; Gómez, Isabel; Peña, Guadalupe; Amaro, Itzel; Ortíz, Ernesto; Becerril, Baltazar; Ibarra, Jorge E; Bravo, Alejandra; Soberón, Mario

    2015-04-01

    Bacillus thuringiensis Cry toxins exert their toxic effect by specific recognition of larval midgut proteins leading to oligomerization of the toxin, membrane insertion and pore formation. The exposed domain II loop regions of Cry toxins have been shown to be involved in receptor binding. Insect cadherins have shown to be functionally involved in toxin binding facilitating toxin oligomerization. Here, we isolated a VHH (VHHA5) antibody by phage display that binds Cry3Aa loop 1 and competed with the binding of Cry3Aa to Tenebrio molitor brush border membranes. VHHA5 also competed with the binding of Cry3Aa to a cadherin fragment (CR12) that was previously shown to be involved in binding and toxicity of Cry3Aa, indicating that Cry3Aa binds CR12 through domain II loop 1. Moreover, we show that a loop 1 mutant, previously characterized to have increased toxicity to T. molitor, displayed a correlative enhanced binding affinity to T. molitor CR12 and to VHHA5. These results show that Cry3Aa domain II loop 1 is a binding site of CR12 T. molitor cadherin. Copyright © 2015 Elsevier Ltd. All rights reserved.

  1. Effects of Bacillus thuringiensis israelensis and spinosad on adult emergence of the non-biting midges Polypedilum nubifer (Skuse) and Tanytarsus curticornis Kieffer (Diptera: Chironomidae) in coastal wetlands.

    Science.gov (United States)

    Duchet, Claire; Franquet, Evelyne; Lagadic, Laurent; Lagneau, Christophe

    2015-05-01

    To optimize their efficacy, some insecticides used for mosquito control are introduced into aquatic ecosystems where mosquito larvae develop (marshes, ponds, sanitation devices) and cannot escape from the treated water. However, this raises the question of possible effects of mosquito larvicides on non-target aquatic species. Bacillus thuringiensis var. israelensis (Bti), which is well-known for its selectivity for Nematocera dipterans, is widely used for mosquito control all over the world. Spinosad, a mixture of spinosyns A and D known as fermentation products of a soil actinomycete (Saccharopolyspora spinosa), is a biological neurotoxic insecticide with a broader action spectrum. It is a candidate larvicide for mosquito control, but some studies showed that it may be toxic to beneficial or non-target species, including non-biting midges. The present study was therefore undertaken to assess the impact of Bti and spinosad on natural populations of Polypedilum nubifer (Skuse) and Tanytarsus curticornis Kieffer (Diptera: Chironomidae) in field enclosures implemented in Mediterranean coastal wetlands. Unlike Bti, spinosad had a strong lethal effect on P. nubifer and seems to affect T. curticornis at presumed recommended rates for field application. Differences in the sensitivity of these two species to spinosad confirm that population dynamics need to be known for a proper assessment of the risk encountered by chironomids in wetlands where larvicide-based mosquito control occurs. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. Actividad biológica de Bacillus thuringiensis sobre la polilla guatemalteca de la papa, Tecia solanivora Povolny (Lepidoptera: Gelechiidae

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    Adriana Carolina Rojas Arias

    2013-12-01

    Full Text Available La papa (Solanum tuberosum es uno de los cultivos más importantes de Colombia. Las larvas de la polilla guatemalteca de la papa, Tecia solanivora Povolny (Lepidoptera: Gelechiidae, causan daños directos a los tubérculos, produciendo pérdidas económicas e incremento en el uso de agroquímicos. Bacillus thuringiensis (Bt es una alternativa en el manejo de insectos plaga gracias a su especificidad. Su actividad depende de proteínas denominadas Cry, que cuando son ingeridas por un insecto susceptible forman poros en sus células intestinales que producen la muerte del insecto. Esta revisión presenta los estudios sobre el efecto de Bt hacia T. solanivora. Se ha encontrado que las toxinas Cry1Ac (base para variedades transgénicas y Cry1B tienen importante actividad tóxica. Igualmente se destaca la experiencia del diseño y evaluación de una toxina híbrida (Cry1B-Cry1I que resultó en una importante letalidad hacia T. solanivora.

  3. Two conformational states of the membrane-associated Bacillus thuringiensis Cry4Ba δ-endotoxin complex revealed by electron crystallography: Implications for toxin-pore formation

    International Nuclear Information System (INIS)

    Ounjai, Puey; Unger, Vinzenz M.; Sigworth, Fred J.; Angsuthanasombat, Chanan

    2007-01-01

    The insecticidal nature of Cry δ-endotoxins produced by Bacillus thuringiensis is generally believed to be caused by their ability to form lytic pores in the midgut cell membrane of susceptible insect larvae. Here we have analyzed membrane-associated structures of the 65-kDa dipteran-active Cry4Ba toxin by electron crystallography. The membrane-associated toxin complex was crystallized in the presence of DMPC via detergent dialysis. Depending upon the charge of the adsorbed surface, 2D crystals of the oligomeric toxin complex have been captured in two distinct conformations. The projection maps of those crystals have been generated at 17 A resolution. Both complexes appeared to be trimeric; as in one crystal form, its projection structure revealed a symmetrical pinwheel-like shape with virtually no depression in the middle of the complex. The other form revealed a propeller-like conformation displaying an obvious hole in the center region, presumably representing the toxin-induced pore. These crystallographic data thus demonstrate for the first time that the 65-kDa activated Cry4Ba toxin in association with lipid membranes could exist in at least two different trimeric conformations, conceivably implying the closed and open states of the pore

  4. Histopathological and ultrastructural effects of delta-endotoxins of Bacillus thuringiensis serovar israelensis in the midgut of Simulium pertinax larvae (Diptera, Simuliidae

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    CFG Cavados

    2004-08-01

    Full Text Available The bacterium Bacillus thuringiensis (Bt produces parasporal crystals containing delta-endotoxins responsible for selective insecticidal activity on larvae. Upon ingestion, these crystals are solubilized in the midgut lumen and converted into active toxins that bind to receptors present on the microvilli causing serious damage to the epithelial columnar cells. We investigated the effect of these endotoxins on larvae of the Simulium pertinax, a common black fly in Brazil, using several concentrations during 4 h of the serovar israelensis strain IPS-82 (LFB-FIOCRUZ 584, serotype H-14 type strain of the Institute Pasteur, Paris. Light and electron microscope observations revealed, by time and endotoxin concentration, increasing damages of the larvae midgut epithelium. The most characteristic effects were midgut columnar cell vacuolization, microvilli damages, epithelium cell contents passing into the midgut lumen and finally the cell death. This article is the first report of the histopathological effects of the Bti endotoxins in the midgut of S. pertinax larvae and the data obtained may contribute to a better understanding of the mode of action of this bacterial strain used as bioinsecticide against black fly larvae.

  5. Expression of a Synthetic Gene for the Major Cytotoxin (Cyt1Aa of Bacillus thuringiensis subsp. israelensis in the Chloroplast of Wild-Type Chlamydomonas

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    Seongjoon Kang

    2018-05-01

    Full Text Available Chlamydomonas reinhardtii (Chlamydomonas strains that are toxic to mosquito larvae because they express chloroplast transgenes that are based on the mosquitocidal proteins of Bacillus thuringiensis subsp. israelensis (Bti could be very useful in mosquito control. Chlamydomonas has several advantages for this approach, including genetic controls not generally available with industrial algae. The Bti toxin is produced by sporulating bacteria and has been used for mosquito control for >30 years without creating highly resistant mosquito populations. The suite of toxins is four main proteins: three Cry proteins and the cytotoxic Cyt1Aa (27 kDa. Cyt1Aa is not very toxic to mosquitoes by itself, but it prevents the development of resistance. The production of Cyt1Aa in other microbes, however, has been challenging due to its affinity for certain membrane phospholipids. Here we report on the production of recombinant Cyt1Aa (rCyt1A in the chloroplast of photosynthetic Chlamydomonas at levels of at least 0.3% total protein. Live cell bioassays demonstrated toxicity of the rCyt1Aa Chlamydomonas to larvae of Aedes aegypti. We also expressed the chloroplast cyt1Aa gene in a wild-type Chlamydomonas strain (21 gr that can grow on nitrate. These results have implications for developing a Chlamydomonas strain that will be toxic to mosquito larvae but will not induce strongly resistant populations.

  6. The Replacement of five Consecutive Amino Acids in the Cyt1A Protein of Bacillus thuringiensis Enhances its Cytotoxic Activity against Lung Epithelial Cancer Cells

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    Kavita Nair

    2018-03-01

    Full Text Available Cyt1A protein is a cytolytic protein encoded by the cyt gene of Bacillus thuringiensis subsp. israelensis (Bti as part of the parasporal crystal proteins produced during the sporulation. Cyt1A protein is unique compared to the other endotoxins present in these parasporal crystals. Unlike δ-endotoxins, Cyt1A protein does not require receptors to bind to the target cell and activate the toxicity. It has the ability to affect a broad range of cell types and organisms, due to this characteristic. Cyt1A has been recognized to not only target the insect cells directly, but also recruit other endotoxins by acting as receptors. Due to these mode of actions, Cyt1A has been studied for its cytolytic activity against human cancer cell lines, although not extensively. In this study, we report a novel Cyt1A protein produced by a Bti strain QBT229 isolated from Qatar. When tested for its cytotoxicity against lung cancer cells, this local strain showed considerably higher activity compared to that of the reference Bti and other strains tested. The possible reasons for such enhanced activity were explored at the gene and protein levels. It was evidenced that five consecutive amino acid replacements in the β8 sheet of the Cyt1A protein enhanced the cytotoxicity against the lung epithelial cancer cells. Such novel Cyt1A protein with high cytotoxicity against lung cancer cells has been characterized and reported through this study.

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

    Science.gov (United States)

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

    2015-01-01

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

  8. Short communication. First field assessment of Bacillus thuringiensis subsp. kurstaki aerial application on the colony performance of Apis mellifera L. (Hymenoptera: Apidae

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    Maria del Mar Leza Salord

    2014-05-01

    Full Text Available Honeybee populations around the world are experiencing a decrease in colony numbers probably due to a combination of different causes, such as diseases, poor nutrition and frequent applications of insecticides to control pests. Previous studies about the effect of pesticide Bacillus thuringiensis subsp. kurstaki (Btk on Apis mellifera L. report different results. The aim of this study was to analyze the effect of field aerial applications of Btk on bee colony performance, specifically on the brood cell percentage evolution, which can be used as an indicator of queen health and brood development breeding rates. To achieve it, the brood cell surface was photographed in every sampling, and data were analyzed using a method based on image treatment software. A total of 480 pictures were examined from two groups of four nucleus hives in two areas, one receiving aerial spraying with Btk and the other without treatment. A mixed factorial design was realized to analyse the data showing no differences in colony performance between the two groups of colonies either before the treatment, during and at the end of the assay. Furthermore, the brood surface ratio of Btk-treated/untreated increased along the experiment. Therefore, the results of the present study suggest that Btk aerial applications did not affect the brood development of honeybees under natural conditions. Nevertheless further field studies are required to ascertain a safe use of Btk in forest pest management.

  9. Toxicidad de δ-endotoxinas recombinantes de Bacillus thuringiensis sobre larvas de la polilla guatemalteca (Tecia solanivora (Lepidóptera: Gelechiidae

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    Javier Hernández-Fernández

    2008-07-01

    Full Text Available Con el objetivo de determinar la actividad tóxica específica de las proteínas recombinantes Cry1Aa, Cry1Ac, Cry1B y Cry1C de Bacillus thuringiensis (Bt, sobre larvas de primer instar de Tecia solanivora se estableció la CL50 para las toxinas. Para este fin se implementó la cría masiva de este insecto bajo condiciones de laboratorio, 58±5% de humedad relativa, 18±5ºC de temperatura y un fotoperiodo de 23 h oscuridad y 1 h luz. Se utilizó una dieta seminatural consistente en láminas de papa variedad parda pastusa autoclavada con solución preservante (ácido ascórbico 7 g/L y metilparabeno 5 g/L, ya que fue estable en el tiempo, garantizó la reproducibilidad de los resultados y fue de fácil evaluación. Las proteínas recombinantes se evaluaron a una concentración de 0,1 μg/cm2. Los resultados obtenidos de porcentaje de mortalidad indicaron que no había diferencias entre las delta-endotoxinas recombinantes Cry1Aa, Cry1Ac, Cry1B y Cry1C de Bt (P

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

    Science.gov (United States)

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

    2018-04-01

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

  11. Lack of fitness costs and inheritance of resistance to Bacillus thuringiensis Cry1Ac toxin in a near-isogenic strain of Plutella xylostella (Lepidoptera: Plutellidae).

    Science.gov (United States)

    Zhu, Xun; Yang, Yanjv; Wu, Qingjun; Wang, Shaoli; Xie, Wen; Guo, Zhaojiang; Kang, Shi; Xia, Jixing; Zhang, Youjun

    2016-02-01

    Resistance to Bacillus thuringiensis (Bt) formulations in insects may be associated with fitness costs. A lack of costs enables resistance alleles to persist, which may contribute to the rapid development and spread of resistance in populations. To assess the fitness costs associated with Bt Cry1Ac resistance in Plutella xylostella, life tables were constructed for a near-isogenic resistant strain (NIL-R) and a susceptible strain in this study. No fitness costs associated with Cry1Ac resistance in NIL-R were detected, based on the duration of egg and larval stages, the survival of eggs and larvae, adult longevity, fecundity, net reproductive rate, gross reproduction rate, finite rate of increase and mean generation time. Based on log dose-probit lines, resistance in NIL-R is incompletely recessive and results from a single, autosomal, recessive locus; the degree of dominance was estimated to be -0.74 and -0.71 for F1 (resistant ♀ × susceptible ♂) and F1 ' (susceptible ♀ × resistant ♂) progeny respectively. Assessment of near-isogenic Cry1Ac-resistant and Cry1Ac-susceptible strains of P. xylostella indicated that resistance is not accompanied with fitness costs, and that resistance is incompletely recessive. These findings should be useful in managing the development of Bt Cry1Ac resistance. © 2015 Society of Chemical Industry.

  12. The midgut cadherin-like gene is not associated with resistance to Bacillus thuringiensis toxin Cry1Ac in Plutella xylostella (L.).

    Science.gov (United States)

    Guo, Zhaojiang; Kang, Shi; Zhu, Xun; Wu, Qingjun; Wang, Shaoli; Xie, Wen; Zhang, Youjun

    2015-03-01

    The Gram-positive bacterium Bacillus thuringiensis (Bt) produces Cry toxins that have been used to control important agricultural pests. Evolution of resistance in target pests threatens the effectiveness of these toxins when used either in sprayed biopesticides or in Bt transgenic crops. Although alterations of the midgut cadherin-like receptor can lead to Bt Cry toxin resistance in many insects, whether the cadherin gene is involved in Cry1Ac resistance of Plutella xylostella (L.) remains unclear. Here, we present experimental evidence that resistance to Cry1Ac or Bt var. kurstaki (Btk) in P. xylostella is not due to alterations of the cadherin gene. The bona fide P. xylostella cadherin cDNA sequence was cloned and analyzed, and comparisons of the cadherin cDNA sequence among susceptible and resistant P. xylostella strains confirmed that Cry1Ac resistance was independent of mutations in this gene. In addition, real-time quantitative PCR (qPCR) indicated that cadherin transcript levels did not significantly differ among susceptible and resistant P. xylostella strains. RNA interference (RNAi)-mediated suppression of cadherin gene expression did not affect larval susceptibility to Cry1Ac toxin. Furthermore, genetic linkage assays using four cadherin gDNA allelic biomarkers confirmed that the cadherin gene is not linked to resistance against Cry1Ac in P. xylostella. Taken together, our findings demonstrate that Cry1Ac resistance of P. xylostella is independent of the cadherin gene. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. In vitro uptake of 140 kDa Bacillus thuringiensis nematicidal crystal proteins by the second stage juvenile of Meloidogyne hapla.

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    Fengjuan Zhang

    Full Text Available Plant-parasitic nematodes (PPNs are piercing/sucking pests, which cause severe damage to crops worldwide, and are difficult to control. The cyst and root-knot nematodes (RKN are sedentary endoparasites that develop specialized multinucleate feeding structures from the plant cells called syncytia or giant cells respectively. Within these structures the nematodes produce feeding tubes, which act as molecular sieves with exclusion limits. For example, Heterodera schachtii is reportedly unable to ingest proteins larger than 28 kDa. However, it is unknown yet what is the molecular exclusion limit of the Meloidogyne hapla. Several types of Bacillus thuringiensis crystal proteins showed toxicity to M. hapla. To monitor the entry pathway of crystal proteins into M. hapla, second-stage juveniles (J2 were treated with NHS-rhodamine labeled nematicidal crystal proteins (Cry55Aa, Cry6Aa, and Cry5Ba. Confocal microscopic observation showed that these crystal proteins were initially detected in the stylet and esophageal lumen, and subsequently in the gut. Western blot analysis revealed that these crystal proteins were modified to different molecular sizes after being ingested. The uptake efficiency of the crystal proteins by the M. hapla J2 decreased with increasing of protein molecular mass, based on enzyme-linked immunosorbent assay analysis. Our discovery revealed 140 kDa nematicidal crystal proteins entered M. hapla J2 via the stylet, and it has important implications in designing a transgenic resistance approach to control RKN.

  14. A novel strategy for producing compost with enhanced biopesticide properties through solid-state fermentation of biowaste and inoculation with Bacillus thuringiensis.

    Science.gov (United States)

    Ballardo, Cindy; Barrena, Raquel; Artola, Adriana; Sánchez, Antoni

    2017-12-01

    In the framework of a circular economy, organic solid wastes are considered to be resources useful for obtaining value-added products. Among other potential uses, biodegradable wastes from agricultural, industrial, and domestic sources are being studied to obtain biopesticides through solid-state fermentation (SSF), mainly at the laboratory scale. The suitability of biowaste (source-selected organic fraction of municipal solid waste) for use as a substrate for Bacillus thuringiensis (Bt) growth under non-sterile conditions in a 10 L SSF reactor was determined in this study. An operational strategy for setting up a semi-continuous process yielding a stabilised organic compost-like material enriched with Bt suitable for use as a soil amendment was developed. Concentrations of 1.7·10 7 -2.2·10 7 and 1.3·10 7 -2.1·10 7  CFU g -1 DM for Bt viable cells and spores, respectively, were obtained in the final material. As the results confirmed, Bt-enriched compost-like material with potential biopesticide properties can be produced from non-sterile biowaste. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. The diverse nematicidal properties and biocontrol efficacy of Bacillus thuringiensis Cry6A against the root-knot nematode Meloidogyne hapla.

    Science.gov (United States)

    Yu, Ziquan; Xiong, Jing; Zhou, Qiaoni; Luo, Haiyan; Hu, Shengbiao; Xia, Liqiu; Sun, Ming; Li, Lin; Yu, Ziniu

    2015-02-01

    Cry6A toxin from Bacillus thuringiensis is a representative nematicidal crystal protein with a variety of nematicidal properties to free-living nematode Caenorhabditis elegans. Cry6A shares very low homology and different structure with Cry5B, another representative nematicidal crystal protein, and probably acts in a distinct pathway. All these strongly indicate that Cry6A toxin is likely a potent candidate for nematicide. The present study dealt with global investigation to determine the detrimental impacts of Cry6Aa2 toxin on Meloidogyne hapla, a root-knot nematode, and evaluated its biocontrol efficacy in pot experiment. Obtained results indicated that Cry6Aa2 toxin exhibits obvious toxicity to second-stage juvenile of M. hapla, and significantly inhibits egg hatch, motility, and penetration to host plant. Pot experiment suggested that soil drenching with spore-crystal mixture of Cry6Aa2 can clearly lighten the disease of root-knot nematode, including reduction of galling index and egg masses on host plant root, decreasing final population of nematode in soil. Moreover, application of Cry6Aa2 can obviously promote plant growth. These results demonstrated that Cry6Aa2 toxin is a promising nematicidal agent, and possesses great potential in plant-parasitic nematode management and construction of transgenic crop with constant resistance to nematode. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. In vivo binding of the Cry11Bb toxin of Bacillus thuringiensis subsp. medellin to the midgut of mosquito larvae (Diptera: Culicidae

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    Ruiz Lina María

    2004-01-01

    Full Text Available Bacillus thuringiensis subsp. medellin produces numerous proteins among which 94 kDa known as Cry11Bb, has mosquitocidal activity. The mode of action of the Cry11 proteins has been described as similar to those of the Cry1 toxins, nevertheless, the mechanism of action is still not clear. In this study we investigated the in vivo binding of the Cry11Bb toxin to the midgut of the insect species Anopheles albimanus, Aedes aegypti, and Culex quinquefasciatus by immunohistochemical analysis. Spodoptera frugiperda was included as negative control. The Cry11Bb protein was detected on the apical microvilli of the midgut epithelial cells, mostly on the posterior midgut and gastric caeca of the three mosquito species. Additionally, the toxin was detected in the Malpighian tubules of An. albimanus, Ae. aegypti, Cx. quinquefasciatus, and in the basal membrane of the epithelial cells of Ae. aegypti midgut. No toxin accumulation was observed in the peritrophic membrane of any of the mosquito species studied. These results confirm that the primary site of action of the Cry11 toxins is the apical membrane of the midgut epithelial cells of mosquito larvae.

  17. Further research on the biological function of inclusion bodies of Anomala cuprea entomopoxvirus, with special reference to the effect on the insecticidal activity of a Bacillus thuringiensis formulation.

    Science.gov (United States)

    Mitsuhashi, Wataru; Asano, Shoji; Miyamoto, Kazuhisa; Wada, Sanae

    2014-01-01

    Entomopoxviruses (EVs) form two types of inclusion body: spheroids, which contain virions, and spindles, which do not. The authors tested whether the spindles from a coleopteran EV, Anomala cuprea EV (ACEV), enhanced the insecticidal activity of a commercial Bacillus thuringiensis (Bt) formulation and the susceptibility of scarabaeid pest species in Japan to the virus's spheroids, to assess whether ACEV inclusion bodies are potential biological control agents for pest insects. Peroral inoculation with both ACEV spindles and the Bt toxin only or the complete Bt formulation shortened the survival and increased the mortality of treated insects compared with those of insects inoculated with Bt without the spindles (8-38 h of decrease in LT50 values among assays). ACEV showed high infectivity to a major scarabaeid pest species in Japanese sugar cane fields. The results suggest that spindles or the constituent protein fusolin can be used as a coagent with Bt formulations, and that fusolin coexpression with a Bt toxin in crops might improve the insecticidal efficacy. In addition, the spheroids are potential biocontrol agents for some scarabaeid pests that are not easy to control because of their underground habitation. © 2013 Society of Chemical Industry.

  18. Development of Monoclonal Antibodies Recognizing Linear Epitope: Illustration by Three Bacillus thuringiensis Crystal Proteins of Genetically Modified Cotton, Maize, and Tobacco.

    Science.gov (United States)

    Cao, Zhen; Zhang, Wei; Ning, Xiangxue; Wang, Baomin; Liu, Yunjun; Li, Qing X

    2017-11-22

    Bacillus thuringiensis Cry1Ac, Cry1Ia1, and Cry1Ie are δ-endotoxin insecticidal proteins widely implemented in genetically modified organisms (GMO), such as cotton, maize, and potato. Western blot assay integrates electrophoresis separation power and antibody high specificity for monitoring specific exogenous proteins expressed in GMO. Procedures for evoking monoclonal antibody (mAb) for Western blot were poorly documented. In the present study, Cry1Ac partially denatured at 100 °C for 5 min was used as an immunogen to develop mAbs selectively recognizing a linear epitope of Cry1Ac for Western blot. mAb 5E9C6 and 3E6E2 selected with sandwich ELISA strongly recognized the heat semidenatured Cry1Ac. Particularly, 3E6E2 recognized both E. coli and cotton seed expressed Cry1Ac in Western blot. Such strategy of using partially denatured proteins as immunogens and using sandwich ELISA for mAb screening was also successfully demonstrated with production of mAbs against Cry1Ie for Western blot assay in maize.

  19. Detection of cry1 genes in Bacillus thuringiensis isolates from South of Brazil and activity against Aanticarsia gemmatalis (Lepidoptera:Noctuidae

    Directory of Open Access Journals (Sweden)

    Bobrowski Vera Lucia

    2001-01-01

    Full Text Available The bacterium Bacillus thuringiensis (Bt is characterized by its ability to produce proteic crystalline inclusions during sporulation. Cry1 protein has insecticidal activity and is highly specific to certain insects and not toxic to unrelated insects, plants or vertebrates. In this work, the patogenicity of twelve Bt isolates was tested against Anticarsia gemmatalis, one of the most important insect pests of soybeans. Spore-crystal complex was applied to the surface of artificial diets and the mortality of A. gemmatalis larvae was assessed seven days after each treatment. When compared to a control Bt isolate known by its high toxicity to A. gemmatalis larvae, four novel Bt isolates exhibited even higher toxic activities against the insect, resulting in more than 90% mortality. PCR was used to amplify DNA fragments related to known cry1 genes. Bt strains with high toxicity produced expected PCR products of around 280 bp, whereas non-toxic or low toxic strains did not produce any PCR product or showed amplified fragments of different sizes. Toxic Bt isolates also exhibited an expected protein profile when total protein extracts were evaluated by SDS-PAGE.

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

    Science.gov (United States)

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

    2015-10-01

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

  1. Multiple linear regression and artificial neural networks for delta-endotoxin and protease yields modelling of Bacillus thuringiensis.

    Science.gov (United States)

    Ennouri, Karim; Ben Ayed, Rayda; Triki, Mohamed Ali; Ottaviani, Ennio; Mazzarello, Maura; Hertelli, Fathi; Zouari, Nabil

    2017-07-01

    The aim of the present work was to develop a model that supplies accurate predictions of the yields of delta-endotoxins and proteases produced by B. thuringiensis var. kurstaki HD-1. Using available medium ingredients as variables, a mathematical method, based on Plackett-Burman design (PB), was employed to analyze and compare data generated by the Bootstrap method and processed by multiple linear regressions (MLR) and artificial neural networks (ANN) including multilayer perceptron (MLP) and radial basis function (RBF) models. The predictive ability of these models was evaluated by comparison of output data through the determination of coefficient (R 2 ) and mean square error (MSE) values. The results demonstrate that the prediction of the yields of delta-endotoxin and protease was more accurate by ANN technique (87 and 89% for delta-endotoxin and protease determination coefficients, respectively) when compared with MLR method (73.1 and 77.2% for delta-endotoxin and protease determination coefficients, respectively), suggesting that the proposed ANNs, especially MLP, is a suitable new approach for determining yields of bacterial products that allow us to make more appropriate predictions in a shorter time and with less engineering effort.

  2. Domain wise docking analyses of the modular chitin binding protein CBP50 from Bacillus thuringiensis serovar konkukian S4.

    Science.gov (United States)

    Sehar, Ujala; Mehmood, Muhammad Aamer; Hussain, Khadim; Nawaz, Salman; Nadeem, Shahid; Siddique, Muhammad Hussnain; Nadeem, Habibullah; Gull, Munazza; Ahmad, Niaz; Sohail, Iqra; Gill, Saba Shahid; Majeed, Summera

    2013-01-01

    This paper presents an in silico characterization of the chitin binding protein CBP50 from B. thuringiensis serovar konkukian S4 through homology modeling and molecular docking. The CBP50 has shown a modular structure containing an N-terminal CBM33 domain, two consecutive fibronectin-III (Fn-III) like domains and a C-terminal CBM5 domain. The protein presented a unique modular structure which could not be modeled using ordinary procedures. So, domain wise modeling using MODELLER and docking analyses using Autodock Vina were performed. The best conformation for each domain was selected using standard procedure. It was revealed that four amino acid residues Glu-71, Ser-74, Glu-76 and Gln-90 from N-terminal domain are involved in protein-substrate interaction. Similarly, amino acid residues Trp-20, Asn-21, Ser-23 and Val-30 of Fn-III like domains and Glu-15, Ala-17, Ser-18 and Leu-35 of C-terminal domain were involved in substrate binding. Site-directed mutagenesis of these proposed amino acid residues in future will elucidate the key amino acids involved in chitin binding activity of CBP50 protein.

  3. Procesamiento proteolítico de la toxina Cyt1Ab1 producida por Bacillus thuringiensis subespecie Medellín

    Directory of Open Access Journals (Sweden)

    Sergio Ordúz

    2000-02-01

    Full Text Available

    Bacillus thuringiensis produce d1 endotoxinas que requieren activación proteolítica para ser activas. La activación de la toxina citolítica de 28 kDa (Cyt1Ab1 de B. thuringiensis subesp. Medellín con tripsina, quimotripsina y extractos intestinales de larvas del mosquito Culex quinquefasciatus fue analizada. La toxina Cyt1Ab1 de B. thuringiensis subsp. medellin fue procesada por todas las proteasa evaluadas a fragmentes entre 23 y 25 kDa, mientras que el procesamiento de la toxina Cyt1Aa1 produjo fragmentos entre 22.5 y 24.5 kDa. La toxina Cyt1Ab1 fue procesada preferencialmente in vitro en un pH de 12, produciendo un fragmento de 25 kDa cuando se trató con extractos intestinales de larvas de mosquito, y resultados similares se obtuvieron cuando la toxina fue activada in vivo por larvas de C. quinquefasciatus. La toxina solubilizada y los fragmentos resistentes a la acción de las proteasas generados en los experimentos in vitro, demostraron actividad hemolítica, pero no mosquitocida. La secuencia amino terminal del fragmento tratado con extractos intestinales de C. quinquefasciatus indica que el sitio de corte está localizado entre Lys31 y Asp32, generando una proteína con secuencia amino Terminal de DDPNEKNNHNS; mientras que la toxina tratada con tripsina mostró un sitio de corte entre Leu29 y Arg30, y la tratada con

  4. Synthetic Polymer Affinity Ligand for Bacillus thuringiensis ( Bt) Cry1Ab/Ac Protein: The Use of Biomimicry Based on the Bt Protein-Insect Receptor Binding Mechanism.

    Science.gov (United States)

    Liu, Mingming; Huang, Rong; Weisman, Adam; Yu, Xiaoyang; Lee, Shih-Hui; Chen, Yalu; Huang, Chao; Hu, Senhua; Chen, Xiuhua; Tan, Wenfeng; Liu, Fan; Chen, Hao; Shea, Kenneth J

    2018-05-24

    We report a novel strategy for creating abiotic Bacillus thuringiensis ( Bt) protein affinity ligands by biomimicry of the recognition process that takes place between Bt Cry1Ab/Ac proteins and insect receptor cadherin-like Bt-R 1 proteins. Guided by this strategy, a library of synthetic polymer nanoparticles (NPs) was prepared and screened for binding to three epitopes 280 FRGSAQGIEGS 290 , 368 RRPFNIGINNQQ 379 and 436 FRSGFSNSSVSIIR 449 located in loop α8, loop 2 and loop 3 of domain II of Bt Cry1Ab/Ac proteins. A negatively charged and hydrophilic nanoparticle (NP12) was found to have high affinity to one of the epitopes, 368 RRPFNIGINNQQ 379 . This same NP also had specific binding ability to both Bt Cry1Ab and Bt Cry1Ac, proteins that share the same epitope, but very low affinity to Bt Cry2A, Bt Cry1C and Bt Cry1F closely related proteins that lack epitope homology. To locate possible NP- Bt Cry1Ab/Ac interaction sites, NP12 was used as a competitive inhibitor to block the binding of 865 NITIHITDTNNK 876 , a specific recognition site in insect receptor Bt-R 1 , to 368 RRPFNIGINNQQ 379 . The inhibition by NP12 reached as high as 84%, indicating that NP12 binds to Bt Cry1Ab/Ac proteins mainly via 368 RRPFNIGINNQQ 379 . This epitope region was then utilized as a "target" or "bait" for the separation and concentration of Bt Cry1Ac protein from the extract of transgenic Bt cotton leaves by NP12. This strategy, based on the antigen-receptor recognition mechanism, can be extended to other biotoxins and pathogen proteins when designing biomimic alternatives to natural protein affinity ligands.

  5. A Large Scale Biorational Approach Using Bacillus thuringiensis israeliensis (Strain AM65-52 for Managing Aedes aegypti Populations to Prevent Dengue, Chikungunya and Zika Transmission.

    Directory of Open Access Journals (Sweden)

    Catherine A Pruszynski

    Full Text Available Aedes aegypti is a container-inhabiting mosquito and a vector of dengue, chikungunya, and Zika viruses. In 2009 several cases of autochthonous dengue transmission were reported in Key West, Florida, USA prompting a comprehensive response to control A. aegypti. In Key West, larvae of this mosquito develop in containers around human habitations which can be numerous and labor intensive to find and treat. Aerial applications of larvicide covering large areas in a short time can be an efficient and economical method to control A. aegypti. Bacillus thuringiensis israelensis (Bti is a bacterial larvicide which is highly target specific and appropriate for wide area spraying over urban areas, but to date, there are no studies that evaluate aerial spraying of Bti to control container mosquitoes like A. aegypti.This paper examines the effectiveness of aerial larvicide applications using VectoBac® WG, a commercially available Bti formulation, for A. aegypti control in an urban setting in the USA. Droplet characteristics and spray drop deposition were evaluated in Key West, Florida, USA. The mortality of A. aegypti in containers placed under canopy in an urban environment was also evaluated. Efficacy of multiple larvicide applications on adult female A. aegypti population reduction was compared between an untreated control and treatment site.Droplet characteristics showed that small droplets can penetrate through dense canopy to reach small containers. VectoBac WG droplets reached small containers under heavy canopy in sufficient amounts to cause > 55% mortality on all application days and >90% mortality on 3 of 5 application days while controls had 50%. Aerial larvicide applications using VectoBac WG can cover wide areas in a short period of time and can be effective in controlling A. aegypti and reducing A. aegypti-borne transmission in urban areas similar to Key West, Florida, USA.

  6. An Intramolecular Salt Bridge in Bacillus thuringiensis Cry4Ba Toxin Is Involved in the Stability of Helix α-3, Which Is Needed for Oligomerization and Insecticidal Activity.

    Science.gov (United States)

    Pacheco, Sabino; Gómez, Isabel; Sánchez, Jorge; García-Gómez, Blanca-Ines; Soberón, Mario; Bravo, Alejandra

    2017-10-15

    Bacillus thuringiensis three-domain Cry toxins kill insects by forming pores in the apical membrane of larval midgut cells. Oligomerization of the toxin is an important step for pore formation. Domain I helix α-3 participates in toxin oligomerization. Here we identify an intramolecular salt bridge within helix α-3 of Cry4Ba (D111-K115) that is conserved in many members of the family of three-domain Cry toxins. Single point mutations such as D111K or K115D resulted in proteins severely affected in toxicity. These mutants were also altered in oligomerization, and the mutant K115D was more sensitive to protease digestion. The double point mutant with reversed charges, D111K-K115D, recovered both oligomerization and toxicity, suggesting that this salt bridge is highly important for conservation of the structure of helix α-3 and necessary to promote the correct oligomerization of the toxin. IMPORTANCE Domain I has been shown to be involved in oligomerization through helix α-3 in different Cry toxins, and mutations affecting oligomerization also elicit changes in toxicity. The three-dimensional structure of the Cry4Ba toxin reveals an intramolecular salt bridge in helix α-3 of domain I. Mutations that disrupt this salt bridge resulted in changes in Cry4Ba oligomerization and toxicity, while a double point reciprocal mutation that restored the salt bridge resulted in recovery of toxin oligomerization and toxicity. These data highlight the role of oligomer formation as a key step in Cry4Ba toxicity. Copyright © 2017 American Society for Microbiology.

  7. Susceptibility of Anthonomus grandis (cotton boll weevil) and Spodoptera frugiperda (fall armyworm) to a cry1ia-type toxin from a Brazilian Bacillus thuringiensis strain.

    Science.gov (United States)

    Grossi-de-Sa, Maria Fatima; Quezado de Magalhaes, Mariana; Silva, Marilia Santos; Silva, Shirley Margareth Buffon; Dias, Simoni Campos; Nakasu, Erich Yukio Tempel; Brunetta, Patricia Sanglard Felipe; Oliveira, Gustavo Ramos; Neto, Osmundo Brilhante de Oliveira; Sampaio de Oliveira, Raquel; Soares, Luis Henrique Barros; Ayub, Marco Antonio Zachia; Siqueira, Herbert Alvaro Abreu; Figueira, Edson L Z

    2007-09-30

    Different isolates of the soil bacterium Bacillus thuringiensis produce multiple crystal (Cry) proteins toxic to a variety of insects, nematodes and protozoans. These insecticidal Cry toxins are known to be active against specific insect orders, being harmless to mammals, birds, amphibians, and reptiles. Due to these characteristics, genes encoding several Cry toxins have been engineered in order to be expressed by a variety of crop plants to control insectpests. The cotton boll weevil, Anthonomus grandis, and the fall armyworm, Spodoptera frugiperda, are the major economically devastating pests of cotton crop in Brazil, causing severe losses, mainly due to their endophytic habit, which results in damages to the cotton boll and floral bud structures. A cry1Ia-type gene, designated cry1Ia12, was isolated and cloned from the Bt S811 strain. Nucleotide sequencing of the cry1Ia12 gene revealed an open reading frame of 2160 bp, encoding a protein of 719 amino acid residues in length, with a predicted molecular mass of 81 kDa. The amino acid sequence of Cry1Ia12 is 99% identical to the known Cry1Ia proteins and differs from them only in one or two amino acid residues positioned along the three domains involved in the insecticidal activity of the toxin. The recombinant Cry1Ia12 protein, corresponding to the cry1Ia12 gene expressed in Escherichia coli cells, showed moderate toxicity towards first instar larvae of both cotton boll weevil and fall armyworm. The highest concentration of the recombinant Cry1Ia12 tested to achieve the maximum toxicities against cotton boll weevil larvae and fall armyworm larvae were 230 microg/mL and 5 microg/mL, respectively. The herein demonstrated insecticidal activity of the recombinant Cry1Ia12 toxin against cotton boll weevil and fall armyworm larvae opens promising perspectives for the genetic engineering of cotton crop resistant to both these devastating pests in Brazil.

  8. Synergism and Antagonism between Bacillus thuringiensis Vip3A and Cry1 Proteins in Heliothis virescens, Diatraea saccharalis and Spodoptera frugiperda

    Science.gov (United States)

    Lemes, Ana Rita Nunes; Davolos, Camila Chiaradia; Legori, Paula Cristina Brunini Crialesi; Fernandes, Odair Aparecido; Ferré, Juan; Lemos, Manoel Victor Franco; Desiderio, Janete Apparecida

    2014-01-01

    Second generation Bt crops (insect resistant crops carrying Bacillus thuringiensis genes) combine more than one gene that codes for insecticidal proteins in the same plant to provide better control of agricultural pests. Some of the new combinations involve co-expression of cry and vip genes. Because Cry and Vip proteins have different midgut targets and possibly different mechanisms of toxicity, it is important to evaluate possible synergistic or antagonistic interactions between these two classes of toxins. Three members of the Cry1 class of proteins and three from the Vip3A class were tested against Heliothis virescens for possible interactions. At the level of LC50, Cry1Ac was the most active protein, whereas the rest of proteins tested were similarly active. However, at the level of LC90, Cry1Aa and Cry1Ca were the least active proteins, and Cry1Ac and Vip3A proteins were not significantly different. Under the experimental conditions used in this study, we found an antagonistic effect of Cry1Ca with the three Vip3A proteins. The interaction between Cry1Ca and Vip3Aa was also tested on two other species of Lepidoptera. Whereas antagonism was observed in Spodoptera frugiperda, synergism was found in Diatraea saccharalis. In all cases, the interaction between Vip3A and Cry1 proteins was more evident at the LC90 level than at the LC50 level. The fact that the same combination of proteins may result in a synergistic or an antagonistic interaction may be an indication that there are different types of interactions within the host, depending on the insect species tested. PMID:25275646

  9. Characterization of field-evolved resistance to Bacillus thuringiensis-derived Cry1F δ-endotoxin in Spodoptera frugiperda populations from Argentina.

    Science.gov (United States)

    Chandrasena, Desmi I; Signorini, Ana M; Abratti, Gustavo; Storer, Nicholas P; Olaciregui, Magdalena L; Alves, Analiza P; Pilcher, Clinton D

    2018-03-01

    Transgenic maize (Zea mays L.) event TC1507 (Herculex ® I insect protection), expressing Cry1F δ-endotoxin derived from Bacillus thuringiensis var. aizawai, was commercialized in 2003 in the Americas. Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) susceptibility to Cry1F was monitored annually across several regions in Argentina using diagnostic concentration bioassays. Reduced performance of TC1507 maize against S. frugiperda was reported in 2013. A resistant population was established in the laboratory and the dominance of Cry1F resistance was characterized. During 2012-2015, high-survivorship of several populations was observed in the resistance monitoring program. Reciprocal crosses of a Cry1F-resistant population with a Cry1F-susceptible population were evaluated to calculate effective dominance (D ML ) based on mortality levels observed at 100 µg/ml Cry1F. Two additional dominance levels (D LC and D EC ) were calculated using lethal (LC 50 ) or effective concentration (EC 50 ) derived from concentration-response bioassays. Estimates indicated that Cry1F resistance in S. frugiperda in Argentina was either highly recessive (D ML = 0.005) or incompletely recessive (D LC frugiperda Cry1F field-evolved resistance in Argentina. The resistance to Cry1F in S. frugiperda populations collected in Argentina, is autosomal and incompletely recessive similar to the resistance reported in Brazil. © 2017 The Authors. Pest Management Science published by John Wiley © Sons Ltd on behalf of Society of Chemical Industry. © 2017 The Authors. Pest Management Science published by John Wiley © Sons Ltd on behalf of Society of Chemical Industry.

  10. Gypsy moth (Lepidoptera: Lymantriidae) control with ground-based hydraulic applications of Gypchek, in vitro-produced virus, and Bacillus thuringiensis

    International Nuclear Information System (INIS)

    Thorpe, K.W.; Podgwaite, J.D.; Slavicek, J.M.; Webb, R.E.

    1998-01-01

    Gypchek, a registered microbial insecticide for aerial and ground-based application against the gypsy moth, Lymantris dispar L., was field-tested in 1996 and 1997 at 2 doses (10(11) and 10(12) polyhedral inclusion bodies (PIB) per 379 liters (100 gallons)) and with and without a sunscreen. An in vitro-produced strain of gypsy moth virus was tested in 1997 at the 10(11)-PIB dose; however, the effective dose was lower because in vitro-produced PIBs contained 5-fold fewer virions than Gypchek PIBs. Treatments were applied to overstory oak, Quercus spp., trees with a truck-mounted hydraulic sprayer. Larval mortality was significantly greater, and larval density was significantly lower, on trees treated with both Gypchek and in vitro-produced virus than on untreated trees. Treatment with in vitro-produced virus resulted in significantly lower mortality and higher larval density than Gypchek treatment. Defoliation was significantly reduced by all treatments in 1996 but not in 1997, when overall gypsy moth population density dropped to very low levels. Mortality was significantly higher among larvae treated at the 10(12)-PIB dose in 1996, but there was no significant dose effect on larval density or defoliation. Mortality, density, and defoliation were not dose dependent in the 1997 test. The addition of sunscreen to Gypchek applied at the 10(12)-PIB dose did not have a significant effect on any of the measured parameters in 1996 or 1997. In 1996, the addition of an enhancer, Blankophor BBH, to the 10(11)-PIB dose increased mortality to a level that was not significantly different from that at the 10(12)-PIB dose. The level of control with the 10(12)-PIB treatment was not significantly different from that achieved with an application of Bacillus thuringiensis subsp. kurstaki Berliner (Foray 48B) at 36 billion international units (BIU) per 379 liters

  11. A Large Scale Biorational Approach Using Bacillus thuringiensis israeliensis (Strain AM65-52) for Managing Aedes aegypti Populations to Prevent Dengue, Chikungunya and Zika Transmission.

    Science.gov (United States)

    Pruszynski, Catherine A; Hribar, Lawrence J; Mickle, Robert; Leal, Andrea L

    2017-01-01

    Aedes aegypti is a container-inhabiting mosquito and a vector of dengue, chikungunya, and Zika viruses. In 2009 several cases of autochthonous dengue transmission were reported in Key West, Florida, USA prompting a comprehensive response to control A. aegypti. In Key West, larvae of this mosquito develop in containers around human habitations which can be numerous and labor intensive to find and treat. Aerial applications of larvicide covering large areas in a short time can be an efficient and economical method to control A. aegypti. Bacillus thuringiensis israelensis (Bti) is a bacterial larvicide which is highly target specific and appropriate for wide area spraying over urban areas, but to date, there are no studies that evaluate aerial spraying of Bti to control container mosquitoes like A. aegypti. This paper examines the effectiveness of aerial larvicide applications using VectoBac® WG, a commercially available Bti formulation, for A. aegypti control in an urban setting in the USA. Droplet characteristics and spray drop deposition were evaluated in Key West, Florida, USA. The mortality of A. aegypti in containers placed under canopy in an urban environment was also evaluated. Efficacy of multiple larvicide applications on adult female A. aegypti population reduction was compared between an untreated control and treatment site. Droplet characteristics showed that small droplets can penetrate through dense canopy to reach small containers. VectoBac WG droplets reached small containers under heavy canopy in sufficient amounts to cause > 55% mortality on all application days and >90% mortality on 3 of 5 application days while controls had treated sites was >50%. Aerial larvicide applications using VectoBac WG can cover wide areas in a short period of time and can be effective in controlling A. aegypti and reducing A. aegypti-borne transmission in urban areas similar to Key West, Florida, USA.

  12. Evaluating Liquid and Granular Bacillus thuringiensis var. israelensis Broadcast Applications for Controlling Vectors of Dengue and Chikungunya Viruses in Artificial Containers and Tree Holes.

    Science.gov (United States)

    Harwood, James F; Farooq, Muhammad; Turnwall, Brent T; Richardson, Alec G

    2015-07-01

    The principal vectors of chikungunya and dengue viruses typically oviposit in water-filled artificial and natural containers, including tree holes. Despite the risk these and similar tree hole-inhabiting mosquitoes present to global public health, surprisingly few studies have been conducted to determine an efficient method of applying larvicides specifically to tree holes. The Stihl SR 450, a backpack sprayer commonly utilized during military and civilian vector control operations, may be suitable for controlling larval tree-hole mosquitoes, as it is capable of delivering broadcast applications of granular and liquid dispersible formulations of Bacillus thuringiensis var. israelensis (Bti) to a large area relatively quickly. We compared the application effectiveness of two granular (AllPro Sustain MGB and VectoBac GR) and two liquid (Aquabac XT and VectoBac WDG) formulations of Bti in containers placed on bare ground, placed beneath vegetative cover, and hung 1.5 or 3 m above the ground to simulate tree holes. Aedes aegypti (L.) larval mortality and Bti droplet and granule density data (when appropriate) were recorded for each formulation. Overall, granular formulations of Bti resulted in higher mortality rates in the simulated tree-hole habitats, whereas applications of granular and liquid formulations resulted in similar levels of larval mortality in containers placed on the ground in the open and beneath vegetation. Published by Oxford University Press on behalf of Entomological Society of America 2015. This work is written by US Government employees and is in the public domain in the US.

  13. MAPK Signaling Pathway Alters Expression of Midgut ALP and ABCC Genes and Causes Resistance to Bacillus thuringiensis Cry1Ac Toxin in Diamondback Moth

    Science.gov (United States)

    Wu, Qingjun; Wang, Shaoli; Xie, Wen; Zhu, Xun; Baxter, Simon W.; Zhou, Xuguo; Jurat-Fuentes, Juan Luis; Zhang, Youjun

    2015-01-01

    Insecticidal crystal toxins derived from the soil bacterium Bacillus thuringiensis (Bt) are widely used as biopesticide sprays or expressed in transgenic crops to control insect pests. However, large-scale use of Bt has led to field-evolved resistance in several lepidopteran pests. Resistance to Bt Cry1Ac toxin in the diamondback moth, Plutella xylostella (L.), was previously mapped to a multigenic resistance locus (BtR-1). Here, we assembled the 3.15 Mb BtR-1 locus and found high-level resistance to Cry1Ac and Bt biopesticide in four independent P. xylostella strains were all associated with differential expression of a midgut membrane-bound alkaline phosphatase (ALP) outside this locus and a suite of ATP-binding cassette transporter subfamily C (ABCC) genes inside this locus. The interplay between these resistance genes is controlled by a previously uncharacterized trans-regulatory mechanism via the mitogen-activated protein kinase (MAPK) signaling pathway. Molecular, biochemical, and functional analyses have established ALP as a functional Cry1Ac receptor. Phenotypic association experiments revealed that the recessive Cry1Ac resistance was tightly linked to down-regulation of ALP, ABCC2 and ABCC3, whereas it was not linked to up-regulation of ABCC1. Silencing of ABCC2 and ABCC3 in susceptible larvae reduced their susceptibility to Cry1Ac but did not affect the expression of ALP, whereas suppression of MAP4K4, a constitutively transcriptionally-activated MAPK upstream gene within the BtR-1 locus, led to a transient recovery of gene expression thereby restoring the susceptibility in resistant larvae. These results highlight a crucial role for ALP and ABCC genes in field-evolved resistance to Cry1Ac and reveal a novel trans-regulatory signaling mechanism responsible for modulating the expression of these pivotal genes in P. xylostella. PMID:25875245

  14. MAPK signaling pathway alters expression of midgut ALP and ABCC genes and causes resistance to Bacillus thuringiensis Cry1Ac toxin in diamondback moth.

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    Zhaojiang Guo

    2015-04-01

    Full Text Available Insecticidal crystal toxins derived from the soil bacterium Bacillus thuringiensis (Bt are widely used as biopesticide sprays or expressed in transgenic crops to control insect pests. However, large-scale use of Bt has led to field-evolved resistance in several lepidopteran pests. Resistance to Bt Cry1Ac toxin in the diamondback moth, Plutella xylostella (L., was previously mapped to a multigenic resistance locus (BtR-1. Here, we assembled the 3.15 Mb BtR-1 locus and found high-level resistance to Cry1Ac and Bt biopesticide in four independent P. xylostella strains were all associated with differential expression of a midgut membrane-bound alkaline phosphatase (ALP outside this locus and a suite of ATP-binding cassette transporter subfamily C (ABCC genes inside this locus. The interplay between these resistance genes is controlled by a previously uncharacterized trans-regulatory mechanism via the mitogen-activated protein kinase (MAPK signaling pathway. Molecular, biochemical, and functional analyses have established ALP as a functional Cry1Ac receptor. Phenotypic association experiments revealed that the recessive Cry1Ac resistance was tightly linked to down-regulation of ALP, ABCC2 and ABCC3, whereas it was not linked to up-regulation of ABCC1. Silencing of ABCC2 and ABCC3 in susceptible larvae reduced their susceptibility to Cry1Ac but did not affect the expression of ALP, whereas suppression of MAP4K4, a constitutively transcriptionally-activated MAPK upstream gene within the BtR-1 locus, led to a transient recovery of gene expression thereby restoring the susceptibility in resistant larvae. These results highlight a crucial role for ALP and ABCC genes in field-evolved resistance to Cry1Ac and reveal a novel trans-regulatory signaling mechanism responsible for modulating the expression of these pivotal genes in P. xylostella.

  15. Bacterial Larvicide, Bacillus thuringiensis israelensis Strain AM 65-52 Water Dispersible Granule Formulation Impacts Both Dengue Vector, Aedes aegypti (L.) Population Density and Disease Transmission in Cambodia.

    Science.gov (United States)

    Setha, To; Chantha, Ngan; Benjamin, Seleena; Socheat, Doung

    2016-09-01

    A multi-phased study was conducted in Cambodia from 2005-2011 to measure the impact of larviciding with the bacterial larvicide, Bacillus thuringiensis israelensis (Bti), a water dispersible granule (WG) formulation on the vector, Aedes aegypti (L.) and the epidemiology. In our studies, all in-use containers were treated at 8 g/1000 L, including smaller containers and animal feeders which were found to contribute 23% of Ae aegypti pupae. The treated waters were subjected to routine water exchange activities. Pupal production was suppressed by an average 91% for 8 weeks. Pupal numbers continued to remain significantly lower than the untreated commune (UTC) for 13 weeks post treatment in the peak dengue vector season (ptreated commune. An average 70% of the household harbored 0-5 Ae aegypti mosquitoes per home for 8 weeks post treatment, but in the same period of time >50% of the household in the UTC harbored ≥11 mosquitoes per home. The adult population continued to remain at significantly much lower numbers in the Bti treated commune than in the UTC for 10-12 weeks post treatment (ptreatment in the 6 months monsoon season with complete coverage of the target districts achieved an overall dengue case reduction of 48% in the 6 treated districts compared to the previous year, 2010. Five untreated districts in the same province had an overwhelming increase of 352% of dengue cases during the same period of time. The larvicide efficacy, treatment of all in-use containers at the start of the monsoon season, together with treatment coverage of entire districts interrupted disease transmission in the temephos resistant province.

  16. A Large Scale Biorational Approach Using Bacillus thuringiensis israeliensis (Strain AM65-52) for Managing Aedes aegypti Populations to Prevent Dengue, Chikungunya and Zika Transmission

    Science.gov (United States)

    Pruszynski, Catherine A.; Hribar, Lawrence J.; Mickle, Robert; Leal, Andrea L.

    2017-01-01

    Background Aedes aegypti is a container-inhabiting mosquito and a vector of dengue, chikungunya, and Zika viruses. In 2009 several cases of autochthonous dengue transmission were reported in Key West, Florida, USA prompting a comprehensive response to control A. aegypti. In Key West, larvae of this mosquito develop in containers around human habitations which can be numerous and labor intensive to find and treat. Aerial applications of larvicide covering large areas in a short time can be an efficient and economical method to control A. aegypti. Bacillus thuringiensis israelensis (Bti) is a bacterial larvicide which is highly target specific and appropriate for wide area spraying over urban areas, but to date, there are no studies that evaluate aerial spraying of Bti to control container mosquitoes like A. aegypti. Methodology This paper examines the effectiveness of aerial larvicide applications using VectoBac® WG, a commercially available Bti formulation, for A. aegypti control in an urban setting in the USA. Droplet characteristics and spray drop deposition were evaluated in Key West, Florida, USA. The mortality of A. aegypti in containers placed under canopy in an urban environment was also evaluated. Efficacy of multiple larvicide applications on adult female A. aegypti population reduction was compared between an untreated control and treatment site. Conclusions Droplet characteristics showed that small droplets can penetrate through dense canopy to reach small containers. VectoBac WG droplets reached small containers under heavy canopy in sufficient amounts to cause > 55% mortality on all application days and >90% mortality on 3 of 5 application days while controls had 50%. Aerial larvicide applications using VectoBac WG can cover wide areas in a short period of time and can be effective in controlling A. aegypti and reducing A. aegypti-borne transmission in urban areas similar to Key West, Florida, USA. PMID:28199323

  17. Bacillus thuringiensis Cry5B protein is highly efficacious as a single-dose therapy against an intestinal roundworm infection in mice.

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    Yan Hu

    2010-03-01

    Full Text Available Intestinal parasitic nematode diseases are one of the great diseases of our time. Intestinal roundworm parasites, including hookworms, whipworms, and Ascaris, infect well over 1 billion people and cause significant morbidity, especially in children and pregnant women. To date, there is only one drug, albendazole, with adequate efficacy against these parasites to be used in mass drug administration, although tribendimidine may emerge as a second. Given the hundreds of millions of people to be treated, the threat of parasite resistance, and the inadequacy of current treatments, new anthelmintics are urgently needed. Bacillus thuringiensis (Bt crystal (Cry proteins are the most common used biologically produced insecticides in the world and are considered non-toxic to vertebrates.Here we study the ability of a nematicidal Cry protein, Cry5B, to effect a cure in mice of a chronic roundworm infection caused by the natural intestinal parasite, Heligmosomoides bakeri (formerly polygyrus. We show that Cry5B produced from either of two Bt strains can act as an anthelmintic in vivo when administered as a single dose, achieving a approximately 98% reduction in parasite egg production and approximately 70% reduction in worm burdens when delivered per os at approximately 700 nmoles/kg (90-100 mg/kg. Furthermore, our data, combined with the findings of others, suggest that the relative efficacy of Cry5B is either comparable or superior to current anthelmintics. We also demonstrate that Cry5B is likely to be degraded quite rapidly in the stomach, suggesting that the actual dose reaching the parasites is very small.This study indicates that Bt Cry proteins such as Cry5B have excellent anthelmintic properties in vivo and that proper formulation of the protein is likely to reveal a superior anthelmintic.

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

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    B.M. Ribeiro

    1998-06-01

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

  19. Study of the allergenic potential of Bacillus thuringiensis Cry1Ac toxin following intra-gastric administration in a murine model of food-allergy.

    Science.gov (United States)

    Santos-Vigil, Karla I; Ilhuicatzi-Alvarado, Damaris; García-Hernández, Ana L; Herrera-García, Juan S; Moreno-Fierros, Leticia

    2018-06-07

    Cry1Ac toxin, from Bacillus thuringiensis, is widely used as a biopesticide and expressed in genetically modified (GM) plants used for human and animal consumption. Since Cry1Ac is also immunogenic and able to activate macrophages, it is crucial to thoroughly evaluate the immunological effects elicited after intra-gastric administration. The allergenic potential of purified Cry1Ac was assessed and compared with that induced in a murine model of food-allergy to ovalbumin (OVA), in which animals are sensitized with the adjuvant Cholera toxin (CT). Mice were weekly intragastrically administered with: i) vehicle phosphate-buffered saline (PBS), ii) OVA, iii) OVA plus CT iv) Cry1Ac or v) OVA plus Cry1Ac. Seven weeks after, mice were intragastrically challenged and allergic reactions along with diverse allergy related immunological parameters were evaluated at systemic and intestinal level. The groups immunized with, Cry1Ac, OVA/Cry1Ac or OVA/CT developed moderate allergic reactions, induced significant IgE response and increased frequencies of intestinal granulocytes, IgE+ eosinophils and IgE+ lymphocytes. These same groups also showed colonic lymphoid hyperplasia, notably in humans, this has been associated with food allergy and intestinal inflammation. Although the adjuvant and allergenic potential of CT were higher than the effects of Cry1Ac, the results show that applied intra-gastrically at 50 μg doses, Cry1Ac is immunogenic, moderately allergenic and able to provoke intestinal lymphoid hyperplasia. Moreover, Cry1Ac is also able to induce anaphylaxis, since when mice were intragastrically sensitized with increasing doses of Cry1Ac, with every dose tested, a significant drop in rectal temperature was recorded after intravenous challenge. Copyright © 2018 Elsevier B.V. All rights reserved.

  20. A reliable bioassay procedure to evaluate per os toxicity of Bacillus thuringiensis strains against the rice delphacid, Tagosodes orizicolus (Homoptera: Delphacidae

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    Rebeca Mora

    2007-06-01

    Full Text Available A reliable bioassay procedure was developed to test ingested Bacillus thuringiensis (Bt toxins on the rice delphacid Tagosodes orizicolus. Initially, several colonies were established under greenhouse conditions, using rice plants to nurture the insect. For the bioassay, an in vitro feeding system was developed for third to fourth instar nymphs. Insects were fed through Parafilm membranes on sugar (10 % sucrose and honey bee (1:48 vol/vol solutions, observing a natural mortality of 10-15 % and 0-5 %, respectively. Results were reproducible under controlled conditions during the assay (18±0.1 °C at night and 28±0.1 °C during the day, 80 % RH and a 12:12 day:light photoperiod. In addition, natural mortality was quantified on insect colonies, collected from three different geographic areas of Costa Rica, with no significant differences between colonies under controlled conditions. Finally, bioassays were performed to evaluate the toxicity of a Bt collection on T. orizicolus. A preliminary sample of twenty-seven Bt strains was evaluated on coarse bioassays using three loops of sporulated colonies in 9 ml of liquid diet, the strains that exhibited higher percentages of T. orizicolus mortality were further analyzed in bioassays using lyophilized spores and crystals (1 mg/ml. As a result, strains 26-O-to, 40-X-m, 43S-d and 23-O-to isolated from homopteran insects showed mortalities of 74, 96, 44 and 82 % respectively while HD-137, HD-1 and Bti showed 19, 83 and 95 % mortalities. Controls showed mortalities between 0 and 10 % in all bioassays. This is the first report of a reliable bioassay procedure to evaluate per os toxicity for a homopteran species using Bacillus thuringiensis strains. Rev. Biol. Trop. 55 (2: 373-383. Epub 2007 June, 29.Se desarrolló una metodología de bioensayo para evaluar toxinas de Bacillus thuringiensis (Bt ingeridas por Tagosodes orizicolus, plaga del arroz y vector del virus de la hoja blanca. Se establecieron colonias

  1. LSSP-PCR para la identificación de polimorfismos en el gen cry1B en cepas nativas de Bacillus thuringiensis

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    Martha Ilce Orozco Mera

    2012-01-01

    Full Text Available Título en ingles: LSSP-PCR to identify polymorphisms in the gene cry1B of Bacillus thuringiensis native strain Resumen: Se estandarizó la técnica LSSP-PCR (reacción en cadena de la polimerasa con un único oligonucleótido en condiciones de baja astringencia, para identificar polimorfismos del gen cry1B en aislamientos nativos de Bacillus thuringiensis (Bt. Se evaluaron 164 aislamientos nativos colombianos identificándose el gen cry1Ba en 11 de estos aislamientos. Los 11 fragmentos amplificados, junto con el de la cepa de referencia Bt subsp. aizawai HD137, se analizaron por LSSP-PCR y los patrones electroforéticos obtenidos se compararon cualitativamente. Con los productos amplificados mediante el oligonucleótido directo se construyó un dendrograma utilizando UPGMA que  mostró tres agrupamientos con similitud de 83, 79 y 68%. La agrupación con 68% de similaridad correspondió al aislamiento nativo BtGC120 que presentó el patrón de bandas más variable. Con el oligonucleótido reverso el aislamiento BtGC120 mostró una menor variabilidad (43%. La secuencia nucleotidica obtenida de este fragmento de 806 pares de bases mostró una identidad de 93% con la secuencia de los genes cry1Bc1 de Bt morrisoni y cry1Bb1 de la cepa BT-EG5847. Se predijo del marco de lectura +3 una proteína de 268 residuos aminoácidicos, con 88% de identidad con la proteína Cry1Bc. Esta  secuencia reveló dos dominios, una endotoxina N implicada en la formación del poro y otra endotoxina M relacionada en el reconocimiento del receptor. La evaluación biológica del aislamiento BtGC120 sobre larvas de primer instar del insecto plaga Spodoptera frugiperda, mostró una CL50 de 1,896 ng de proteína total por cm2. Este estudio muestra que la LSSP-PCR es una técnica que permite identificar de una manera específica variaciones en las secuencias de los genes cry de Bt, con potencialidad de encontrar nuevos genes con novedosas actividades biológicas. Abstract

  2. Larval gut pH profile in pestiferous .i.Chironomus crassicaudatus./i. and .i.Glyptotendipes paripes./i. (Chironomidae: Diptera) in reference to the toxicity potential of .i.Bacillus thuringiensis./i. serovar .i.israelensis./i

    Czech Academy of Sciences Publication Activity Database

    Frouz, Jan; Lobinske, R.J.; Yaqub, A.; Ali, A.

    2007-01-01

    Roč. 23, č. 3 (2007), s. 355-358 ISSN 8756-971X Institutional research plan: CEZ:AV0Z60660521 Keywords : chironomidae larvae * gut pH * Bacillus thuringiensis serovar israelensis Subject RIV: EH - Ecology, Behaviour Impact factor: 0.706, year: 2007

  3. Toxicity of Bacillus thuringiensis var. israelensis in aqueous suspension on the South American common frog Leptodactylus latrans (Anura: Leptodactylidae) tadpoles

    International Nuclear Information System (INIS)

    Lajmanovich, Rafael C.; Junges, Celina M.; Cabagna-Zenklusen, Mariana C.; Attademo, Andrés M.; Peltzer, Paola M.; Maglianese, Mariana; Márquez, Vanina E.; Beccaria, Alejandro J.

    2015-01-01

    The effects of commercial formulations of Bacillus thuringiensisvar.israelensis (Bti) on non-target organisms are still a matter of debate; in amphibians, the risks of Bti are little known. To evaluate the toxicity of a commercial liquid (aqueous suspension, AS) formulation of Bti (Introban ® ) on Leptodactylus latrans tadpoles, including median lethal concentration (LC 50 ) and no-and lowest–observed-effect concentrations (NOEC and LOEC, respectively), as well as the possible effects of Bti on oxidative responses, erythrocytes genotoxicity, and histology of the intestines. In the laboratory, tadpoles were exposed to nominal concentrations of 0 (control), 2.5, 5, 10, 20 and 40 mg/L of formulated Bti-AS. Glutathione S-transferase (GST) and catalase (CAT) activities, as well as formation of erythrocyte nuclear abnormalities (ENAs), and histological effect were measured in tadpoles displaying survival rates >85%. L. latrans tadpoles were sensitive to exposure to Bti-AS, reaching 100% mortality after 48 h of exposure at the highest concentration. Bti-AS induced GST and CAT enzymes and genotoxicity (erythrocyte's nuclear abnormalities), and caused intestine's histopathology. Our results demonstrate that toxicity of Bti-AS is dose-dependent for L. latrans tadpoles and that sublethal exposure alters enzymes of oxidative stress, induces genotoxicity, and causes intestine damage. Further research is needed to evaluate the ecotoxicological risk of the massive use of Bti formulations on amphibian populations that commonly used suburban wastewater or urban waterbodies to reproduce and where this biopesticide is frequently applied. - Highlights: • An ecotoxicological evaluation of a Bti formulation on amphibian was conducted. • Toxicity of Bti-AS was dose-dependent for Leptodactylus latrans tadpoles. • Sublethal exposure altered the enzymes of oxidative stress (GST and CAT). • Bti-AS was genotoxic because increased MN frequencies (CO: 0.82–2.74‰).

  4. Identification of Glyceraldehyde-3-phosphate dehydrogenase (GAPDH as a binding protein for a 68-kDa Bacillus thuringiensis parasporal protein cytotoxic against leukaemic cells

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    Nadarajah Vishna

    2010-11-01

    Full Text Available Abstract Background Bacillus thuringiensis (Bt, an ubiquitous gram-positive spore-forming bacterium forms parasporal proteins during the stationary phase of its growth. Recent findings of selective human cancer cell-killing activity in non-insecticidal Bt isolates resulted in a new category of Bt parasporal protein called parasporin. However, little is known about the receptor molecules that bind parasporins and the mechanism of anti-cancer activity. A Malaysian Bt isolate, designated Bt18 produces parasporal protein that exhibit preferential cytotoxic activity for human leukaemic T cells (CEM-SS but is non-cytotoxic to normal T cells or other cancer cell lines such as human cervical cancer (HeLa, human breast cancer (MCF-7 and colon cancer (HT-29 suggesting properties similar to parasporin. In this study we aim to identify the binding protein for Bt18 in human leukaemic T cells. Methods Bt18 parasporal protein was separated using Mono Q anion exchange column attached to a HPLC system and antibody was raised against the purified 68-kDa parasporal protein. Receptor binding assay was used to detect the binding protein for Bt18 parasporal protein in CEM-SS cells and the identified protein was sent for N-terminal sequencing. NCBI protein BLAST was used to analyse the protein sequence. Double immunofluorescence staining techniques was applied to localise Bt18 and binding protein on CEM-SS cell. Results Anion exchange separation of Bt18 parasporal protein yielded a 68-kDa parasporal protein with specific cytotoxic activity. Polyclonal IgG (anti-Bt18 for the 68-kDa parasporal protein was successfully raised and purified. Receptor binding assay showed that Bt18 parasporal protein bound to a 36-kDa protein from the CEM-SS cells lysate. N-terminal amino acid sequence of the 36-kDa protein was GKVKVGVNGFGRIGG. NCBI protein BLAST revealed that the binding protein was Glyceraldehyde-3-phosphate dehydrogenase (GAPDH. Double immunofluorescence staining showed

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

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    Yingying Wang

    2015-12-01

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

  6. Brasicáceas y perspectivas de control biológico del insecto plaga Plutella xylostella (Lepidóptera: Plutellidae utilizando Bacillus thuringiensis

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    Julieth Mena Guerrero

    2017-09-01

    Full Text Available La Plutella xylostella (L. es un insecto plaga que afecta principalmente los cultivos de brasicáceas o crucíferas como el repollo, coliflor, brócoli y rábano, entre otros. Las limitaciones del rendimiento y calidad de estos cultivos se deben principalmente al difícil manejo de esta plaga. En la actualidad, para controlar la P. xylostella se utilizan insecticidas de síntesis química, como piretroides, carbamatos u organofosforados, los cuales tienen acción inmediata pero causan efectos adversos en la salud y en el ambiente. Adicionalmente, la ausencia de enemigos naturales capaces de controlar a sus poblaciones, y su resistencia a numerosos insecticidas convencionales hacen que este insecto se establezca rápidamente en áreas productoras de brasicáceas. Se calcula que los costos mundiales asociados al control de la P. xylostella, sumados a las pérdidas en la producción agrícola, están entre cuatro y cinco billones de dólares anuales. Se han propuesto alternativas seguras, efectivas y de menor impacto ambiental, como el control biológico, que permite la producción sostenible de los cultivos de brasicáceas. La industria agrícola y forestal reconoce a la bacteria entomopatógena Bacillus thuringiensis (Bt como una buena alternativa biológica a los insecticidas químicos, puesto que es inocua sobre el ambiente y su toxicidad es altamente selectiva, ligada a su estrecho rango de especificidad sobre diferentes insectos plaga de los órdenes Lepidóptera, Coleóptera y Díptera. En esta revisión se presentan las posibilidades actuales que se pueden emplear para el control de la P. xylostella utilizando modelos investigativos basados en ensayos biológicos con Bt. Dichas posibilidades buscan superar las desventajas existentes en relación con la plaga P. xylostella y los cultivos de brasicáceas, además de entender la fisiología de Bt bajo condiciones que permitan incrementar la eficacia en el control biológico.

  7. LC and LD50 values of Bacillus thuringiensis Serovar japonensis strain buibui toxin to Oriental beetle and northern masked chafer larvae (Coleoptera: Scarabaeidae).

    Science.gov (United States)

    Mashtoly, Tamer A; El-Zemaity, Mohamed El-Said; Hussien, Mohamed I; Alm, Steven R

    2009-10-01

    Bacillus thuringiensis serovar japonensis strain Buibui has the potential to be an important control agent for pest scarabs. Bioassays were designed to test B. t. japonensis against two of the major turf and ornamental scarab pests infesting turfgrasses and ornamentals and to serve as a basis for further tests against other scarab pests. LC and LD50 values of B. t. serovarjaponensis strain Buibui toxin and spores were determined by four different bioassays for the oriental beetle, Anomala orientalis (Waterhouse), and northern masked chafer, Cyclocephala borealis Arrow. Oriental beetle larvae were bioassayed in autoclaved and nonautoclaved soil from where they were collected (Kingston, RI [native]), in nonautoclaved soil from where the northern masked chafer larvae were collected (Groton, CT [foreign]), and per os. Northern masked chafer larvae were bioassayed in autoclaved and nonautoclaved soil from where they were collected (Groton, CT [native]), in nonautoclaved soil from where the oriental beetle larvae were collected (Kingston, RI [foreign]) and per os. LC50 values of 3.93 microg toxin/g autoclaved native soil, 1.80 microg toxin/g nonautoclaved native soil, and 0.42 microg toxin/g nonautoclaved foreign soil and an LD50 value of 0.41 microg per os were determined at 14 d forA. orientalis. LC50 values of 588.28 microg toxin/g autoclaved native soil, 155.10 microg toxin/g nonautoclaved native soil, 265.32 microg toxin/g nonautoclaved foreign soil, and LD50 of 5.21 microg per os were determined at 14 d (soils) and 10 d (per os) for C. borealis. There were significant differences in LC50 values for oriental beetles in autoclaved, nonautoclaved native soil and nonautoclaved foreign soil. There were significant differences in LCo values for northern masked chafers in autoclaved and nonautoclaved native soil. B. t. japonensis can be applied now for control of oriental beetles at rates that are economically competitive with synthetic chemicals. If we can determine the

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

    Directory of Open Access Journals (Sweden)

    Yueqin Wang

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

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

    Directory of Open Access Journals (Sweden)

    Silvia Caccia

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2016-04-01

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

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

    International Nuclear Information System (INIS)

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

    2016-01-01

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

  12. Community-based biological control of malaria mosquitoes using Bacillus thuringiensis var. israelensis (Bti) in Rwanda: community awareness, acceptance and participation.

    Science.gov (United States)

    Ingabire, Chantal Marie; Hakizimana, Emmanuel; Rulisa, Alexis; Kateera, Fredrick; Van Den Borne, Bart; Muvunyi, Claude Mambo; Mutesa, Leon; Van Vugt, Michelle; Koenraadt, Constantianus J M; Takken, Willem; Alaii, Jane

    2017-10-03

    Targeting the aquatic stages of malaria vectors via larval source management (LSM) in collaboration with local communities could accelerate progress towards malaria elimination when deployed in addition to existing vector control strategies. However, the precise role that communities can assume in implementing such an intervention has not been fully investigated. This study investigated community awareness, acceptance and participation in a study that incorporated the socio-economic and entomological impact of LSM using Bacillus thuringiensis var. israelensis (Bti) in eastern Rwanda, and identified challenges and recommendations for future scale-up. The implementation of the community-based LSM intervention took place in Ruhuha, Rwanda, from February to July 2015. The intervention included three arms: control, community-based (CB) and project-supervised (PS). Mixed methods were used to collect baseline and endline socio-economic data in January and October 2015. A high perceived safety and effectiveness of Bti was reported at the start of the intervention. Being aware of malaria symptoms and perceiving Bti as safe on other living organisms increased the likelihood of community participation through investment of labour time for Bti application. On the other hand, the likelihood for community participation was lower if respondents: (1) perceived rice farming as very profitable; (2) provided more money to the cooperative as a capital; and, (3) were already involved in rice farming for more than 6 years. After 6 months of implementation, an increase in knowledge and skills regarding Bti application was reported. The community perceived a reduction in mosquito density and nuisance biting on treated arms. Main operational, seasonal and geographical challenges included manual application of Bti, long working hours, and need for transportation for reaching the fields. Recommendations were made for future scale-up, including addressing above-mentioned concerns and

  13. Toxicity of Bacillus thuringiensis var. israelensis in aqueous suspension on the South American common frog Leptodactylus latrans (Anura: Leptodactylidae) tadpoles

    Energy Technology Data Exchange (ETDEWEB)

    Lajmanovich, Rafael C., E-mail: lajmanovich@hotmail.com [National Council for Scientific and Technical Research (CONICET), Buenos Aires (Argentina); Faculty of Biochemistry and Biological Sciences (FBCB-UNL), Ciudad Universitaria Paraje el Pozo s/n, 3000 Santa Fe (Argentina); Junges, Celina M. [National Council for Scientific and Technical Research (CONICET), Buenos Aires (Argentina); Faculty of Biochemistry and Biological Sciences (FBCB-UNL), Ciudad Universitaria Paraje el Pozo s/n, 3000 Santa Fe (Argentina); Cabagna-Zenklusen, Mariana C. [Faculty of Biochemistry and Biological Sciences (FBCB-UNL), Ciudad Universitaria Paraje el Pozo s/n, 3000 Santa Fe (Argentina); Attademo, Andrés M.; Peltzer, Paola M. [National Council for Scientific and Technical Research (CONICET), Buenos Aires (Argentina); Faculty of Biochemistry and Biological Sciences (FBCB-UNL), Ciudad Universitaria Paraje el Pozo s/n, 3000 Santa Fe (Argentina); Maglianese, Mariana; Márquez, Vanina E.; Beccaria, Alejandro J. [Faculty of Biochemistry and Biological Sciences (FBCB-UNL), Ciudad Universitaria Paraje el Pozo s/n, 3000 Santa Fe (Argentina)

    2015-01-15

    The effects of commercial formulations of Bacillus thuringiensisvar.israelensis (Bti) on non-target organisms are still a matter of debate; in amphibians, the risks of Bti are little known. To evaluate the toxicity of a commercial liquid (aqueous suspension, AS) formulation of Bti (Introban{sup ®}) on Leptodactylus latrans tadpoles, including median lethal concentration (LC{sub 50}) and no-and lowest–observed-effect concentrations (NOEC and LOEC, respectively), as well as the possible effects of Bti on oxidative responses, erythrocytes genotoxicity, and histology of the intestines. In the laboratory, tadpoles were exposed to nominal concentrations of 0 (control), 2.5, 5, 10, 20 and 40 mg/L of formulated Bti-AS. Glutathione S-transferase (GST) and catalase (CAT) activities, as well as formation of erythrocyte nuclear abnormalities (ENAs), and histological effect were measured in tadpoles displaying survival rates >85%. L. latrans tadpoles were sensitive to exposure to Bti-AS, reaching 100% mortality after 48 h of exposure at the highest concentration. Bti-AS induced GST and CAT enzymes and genotoxicity (erythrocyte's nuclear abnormalities), and caused intestine's histopathology. Our results demonstrate that toxicity of Bti-AS is dose-dependent for L. latrans tadpoles and that sublethal exposure alters enzymes of oxidative stress, induces genotoxicity, and causes intestine damage. Further research is needed to evaluate the ecotoxicological risk of the massive use of Bti formulations on amphibian populations that commonly used suburban wastewater or urban waterbodies to reproduce and where this biopesticide is frequently applied. - Highlights: • An ecotoxicological evaluation of a Bti formulation on amphibian was conducted. • Toxicity of Bti-AS was dose-dependent for Leptodactylus latrans tadpoles. • Sublethal exposure altered the enzymes of oxidative stress (GST and CAT). • Bti-AS was genotoxic because increased MN frequencies (CO: 0.82–2.74

  14. Desempenho de Trichogramma pratissolii Querino & Zucchi (Hymenoptera, Trichogrammatidae em ovos de Anagasta kuehniella (Zeller (Lepidoptera, Pyralidae sob efeito de Bacillus thuringiensis Berliner Trichogramma pratissolii Querino & Zucchi (Hymenoptera, Trichogrammatidae performance in Anagasta kuehniella (Zeller (Lepidoptera, Pyralidae eggs affected by Bacillus thuringiensis Berliner

    Directory of Open Access Journals (Sweden)

    Dirceu Pratissoli

    2006-04-01

    Full Text Available Visando estudar o efeito da bactéria entomopatogênica Bacillus thuringiensis (Bt sobre fêmeas adultas de Trichogramma pratissolii e sua progênie, este trabalho foi realizado com diferentes isolados de Bt e Bt kurstaki misturados ao alimento fornecido para o parasitóide antes de parasitar ovos de Anagasta kuehniella. Para tanto, suspensões de seis isolados de Bt E-3, E-10, E-15, E-16, E-19, E-20 e o Bt kurstaki foram misturados em gotícula de mel (1:1, como fonte de alimento e mel puro como testemunha, e, em seguida, foram oferecidos simultaneamente cartelas com ovos do hospedeiro para o parasitismo. Foram utilizadas 20 repetições por tratamento. O experimento foi mantido em câmara climatizada a 25 ± 1ºC, UR 70 ± 10% e fotofase de 14h. Foram avaliados os parasitismos diários, totais e acumulados; sobrevivência; emergência e razão sexual da progênie. Os tratamentos não afetaram o parasitismo, razão sexual e a longevidade, porém em alguns tratamentos (E-3, E-10, E-16 e E-20 foi observado efeito indireto sobre a emergência da progênie, o que implicaria a necessidade de mais liberações massais do parasitóide para alcançar os resultados esperados. Por outro lado, a aceleração do parasitismo verificada em todos os tratamentos sugere que adultos de T. pratissolii, quando submetidos à pressão de algum fator externo tendem a parasitar o mais rápido possível para assegurar a sobrevivência da progênie. Portanto, a combinação Bt + T. pratissolii pode favorecer a atuação do parasitóide em campo, principalmente em casos em que é necessária uma rápida redução dos níveis populacionais da praga.This work was carried out to detect at laboratory the possible side-effects provoked by Bacillus thuringiensis (Bt strains on egg parasitoid Thichogramma pratissolii and its progeny. Six strains of Bt E-3, E-10, E-15, E-16, E-19, E-20 and Bt kurstaki were assayed in honey drop to stimulate the feeding of the parasitoids in each

  15. Field and semi-field evaluation of Bacillus thuringiensis var. israelensis versus Temephos® in Aedes aegypti controlAvaliação de campo e simulado de campo de Bacillus thuringiensis var. israelensis versus Temephos® no controle de Aedes aegypti

    Directory of Open Access Journals (Sweden)

    Jose Bento Pereira Lima

    2016-06-01

    Full Text Available Introduction: Aedes aegypti  is a vector of the important arboviruses worldwide. Vector control continues to rely mainly on fighting immature stages. Resistance to the larvicide Temephos® was detected in many regions of Brazil since 2000 what led control programs to search for alternative products, such as Bacillus thuringiensis var. israelensis (Bti. Caicó municipality (Rio Grande do Norte State, Brazil was one of the first cities to use Bti. However, after some time, Bti low persistence was noticed as jeopardizing effective vector control. Objective: To compare the efficacy of two Bti granulate formulations, Vectobac G® and Vectobac WDG® and Temephos® against Ae. aegypti in field and semi-field conditions. Methods: Field tests were carried out in two neighbouring  areas which presented Ae. aegypti infestation indices >3%: Walfredo Gurgel and Boa Passagem, Caicó, RGN, Brazil, in 2004. Semi- field tests were performed in the patio of a building. Results: For the field conditions, mortality rates >80% were maintained for 14 days, average. After nine weeks, positive containers for Ae. aegypti in the field were >10% in the area of application of Bti and <1% in the area where Temephos® was applied. In the semi-field conditions Ae. aegypti larval mortality >80% was maintained for up to 56 days for Temephos®, 35 days for Vectobac G® and 49 days for Vectobac WDG®. Conclusions: The results point out to low Bti persistence in the field, mainly for containers exposed to sunlight. Local climatic and environmental conditions should be regarded when new products are tested due to high regional variability prevailing in Brazil. 

  16. Evaluation of Cytotoxicity, Genotoxicity and Hematotoxicity of the Recombinant Spore-Crystal Complexes Cry1Ia, Cry10Aa and Cry1Ba6 from Bacillus thuringiensis in Swiss Mice

    Directory of Open Access Journals (Sweden)

    Ingrid de Souza Freire

    2014-09-01

    Full Text Available The insecticidal properties of Cry-endotoxins from Bacillus thuringiensis (Bt have long been used as spore-crystals in commercial spray formulations for insect control. Recently, some Bt-endotoxin genes have been cloned in many different plants. Toxicological evaluations of three spore-crystal endotoxins, BtCry1Ia, BtCry10Aa and BtCry1Ba6 from B. thuringiensis, were carried out on mice to understand their adverse effects on hematological systems and on genetic material. These three spore-crystals have shown toxic activity to the boll weevil, which is one of the most aggressive pests of the cotton crop. Cry1Ia, Cry10Aa and Cry1Ba6 did not increase the micronucleus frequency in the peripheral erythrocytes of mice and did not cause changes in the frequency of polychromatic erythrocytes. However, some hematologic disburbances were observed, specifically related to Cry1Ia and Cry1Ba6, respectively, for the erythroid and lymphoid lineage. Thus, although the profile of such adverse side effects can be related to their high level of exposure, which is not commonly found in the environment, results showed that these Bt spore-crystals were not harmless to mice, indicating that each spore-crystal endotoxin presents a characteristic profile of toxicity and might be investigated individually.

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

    DEFF Research Database (Denmark)

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

    2005-01-01

    Field trials were established at three European sites (Denmark, Eastern France, South-West France) of genetically modified maize (Zea mays L.) expressing the CryIAb Bacillus thuringiensis toxin (Bt), the near-isogenic non-Bt cultivar, another conventional maize cultivar and grass. Soil from Denmark......) and phospholipid fatty acid analysis (PLFA), and protozoa and nematodes in all samples. Individual differences within a site resulted from: greater nematode numbers under grass than maize on three occasions; different nematode populations under the conventional maize cultivars once; and two occasions when...... there was a reduced protozoan population under Bt maize compared to non-Bt maize. Microbial community structure within the sites only varied with grass compared to maize, with one occurrence of CLPP varying between maize cultivars (Bt versus a conventional cultivar). An overall comparison of Bt versus non-Bt maize...

  18. Effect of sunlight radiation, rainfall and droplet spectra of sprays on persistence of Bacillus thuringiensis deposits after application of DiPel 76AF formulation onto conifers

    International Nuclear Information System (INIS)

    Sundaram, A.; Sundaram, K.M.S.

    1996-01-01

    The Effect of sunlight radiation, rainfall and droplet spectra of sprays on per ‐sistence of a Bacillus thuringiensis subspp. kurstaki (Btk) formulation, DiPel? 76AF, was examined after application onto spruce [Picea glauca (Moench) Voss] foliage. The investigation consisted of three studies: (i) Study I: a laboratory microcosm study to examine the photostability of DiPel 76AF deposits on foliage after different periods of exposure to two radiation intensities, (ii) Study II: a laboratory microcosm study to examine the rainfastness of foliar deposits after exposure to different amounts of rainfall consisting of two separate droplet spectra, and (iii) Study III: a field microcosm study to investigate the influence of two different droplet spectra of DiPel 76AF sprays on foliar persistence of Btk under natural weathering conditions. In all studies, persistence of Btk was investigated both by bioassay [using spruce budworm (Choristoneura fumiferana Clemens)] and total protein assay.The findings of Study I indicated that bioactivity of foliar deposits decreased with increasing duration of exposure to radiation, and with increasing radiation intensity. The half‐life (DT 50 , the exposure period required for 50% of the initial bioactivity to disappear) was 5.1 d for the low intensity, and 3.9 d for the higher intensity. In contrast with the bioassay results, the total protein levels [determined by the bicinchoninic acid (BCA) method] showed no decrease with increasing duration of exposure, or with increasing radiation intensity.The findings of Study II indicated that bioactivity of foliar deposits decreased with increasing cumulative rainfall. A new term, RF 50 [the amount of rain (in mm) required to washoff 50% of the initial deposit], was introduced to understand the relationship between rainfall intensity and reduction in bioactivity. When the same amount of rain was applied in different droplet sizes, the RF 50 value was high (5.2 mm) for the small rain droplets

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

    Directory of Open Access Journals (Sweden)

    Crespo Andre LB

    2009-06-01

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

  20. 75 FR 34040 - Bacillus thuringiensis

    Science.gov (United States)

    2010-06-16

    ... is the appropriate level of sensitivity to detect possible IgE epitopes without high false positive... standards that would require Agency consideration of voluntary consensus standards pursuant to section 12(d...

  1. 76 FR 14289 - Bacillus thuringiensis

    Science.gov (United States)

    2011-03-16

    ... sensitivity to detect possible IgE epitopes without high false positive rates. 3. Prevalence in food... any technical standards that would require Agency consideration of voluntary consensus standards...

  2. Controle integrado de espécies de Simulium (Diptera, Simuliidae por Bacillus thuringiensis e manejos mecânicos no riacho e nos vertedouros de tanques de piscicultura, Almirante Tamandaré, Paraná, Brasil Integrated pest control of Simulium (Diptera, Simuliidae with Bacillus thuringiensis and mechanical handling in creek and pisciculture spillway, Almirante Tamandaré, Paraná, Brazil

    Directory of Open Access Journals (Sweden)

    Fabiane Petry

    2004-03-01

    Full Text Available No Riacho dos Padres na Chácara Evíssima, numa microempresa de agronegócio com criação de peixes e animais domésticos, área de lazer e salão de eventos, localizada na Região Metropolitana de Curitiba, os simulídeos em grande proliferação causaram sério impacto com as picadas a tal ponto que dificultaram o convívio habitual. De janeiro a agosto de 2002, coletou-se 24.021 pupas por meio das quais foram identificadas as espécies: Simulium inaequale, Simulium perflavum, Simulium pertinax com 85,98% e Simulium orbitale, Simulium subnigrum, Simulium distinctum e Simulium incrustatum com somente 14,02% de freqüência. O combate a estas populações iniciado em 28 de fevereiro terminou em 08 de agosto de 2002. O controle integrado consistiu na aplicação de biopesticida Bacillus thuringiensis var. israelensis (Bti e em manejos mecânicos não só removendo os imaturos (ovos, larvas e pupas por meio de escovação de vertedouros, mas também pela retirada de substratos naturais e antrópicos do leito. A densidade larvária média inicial, antes da aplicação de Bti no leito do riacho (75 m e no último vertedouro, verificada em substratos antrópicos fitilhos (área 13.500 cm², foi de 6,7 e 11,5 larvas/cm², respectivamente. Nestes habitats, o controle integrado com Bti de 1,77 a 2,09 mg/litro do ingrediente ativo, suplementado com manejos mecânicos, monitorado em substratos fitilhos (área 20.250 cm², resultou em redução larvar de 72,61% a 99,97% e de 74,91% a 99,45%, respectivamente, nas vazões de 0,39 a 0,45 e de 0,38 a 0,43 m³/min. No primeiro vertedouro (área 5.110 cm², porém, só com manejos mecânicos, atingiu-se a redução larvária de 53,81% a 99,59%. O controle efetuado reduziu as picadas hematófagas ao nível esperado, surtindo o efeito positivo almejado.At Riacho dos Padres, Chácara Evíssima, a small agribusiness venture featuring fish, poultry and cattle farming; a recreation outdoor facility and Event Hall

  3. Evaluation of the impact of Bacillus thuringiensis serovar israelensis and Temephos, used for the control of Simulium (Chirostilbia pertinax Kollar, 1832 (Diptera, Simuliidae on the associated entomofauna, Paraty, state of Rio de Janeiro, Brazil

    Directory of Open Access Journals (Sweden)

    Araújo-Coutinho Carlos José Pereira da Cunha de

    2003-01-01

    Full Text Available The study was set up to evaluate the impact of two commercial larvicide formulations, Bacillus thuringiensis serovar israelensis base (Bti at 15 ppm/1 min and temephos at 0.03 ppm of active ingredient, used to control Simulium pertinax populations, on associated non-target entomofauna occupying the same breeding sites. The experiments were carried out on the Pedra Branca and Muricana rivers, on the slopes of Serra do Mar massif, municipality of Paraty, state of Rio de Janeiro, Brazil. Bti was applied to the river Pedra Branca and temephosto theriver Muricana. On both rivers, treatment and control sections were labeled as such, each one with two observation posts: slow moving water and fast water regions respectively. Artificial substrata was used to evaluate the abundance of associated entomofauna. Attached immature stages of arthropods were removed from both of its surfaces fortnightly. Were collected, from the two rivers, 28 477 specimens of the entomofauna associated with S. pertinax. The families Hydropsychidae, Chironomidae, Bactidae, Simuliidae, Blephariceridae and Megapodagrionidae were represented. These was an impact of temephos on the entomofauna associated with S. pertinax only in Simuliidae and Chironomidae, and to Bti only in Simuliidae. However, the reduction in their numbers was not statistically significant.

  4. Evaluation of the impact of Bacillus thuringiensis serovar israelensis and Temephos, used for the control of Simulium (Chirostilbia) pertinax Kollar, 1832 (Diptera, Simuliidae) on the associated entomofauna, Paraty, state of Rio de Janeiro, Brazil.

    Science.gov (United States)

    de Araújo-Coutinho, Carlos José Pereira da Cunha; Cunha, Andrea de Barros Pinto Viviani; Serra-Freire, Nicolau Maués; de Mello, Rubens Pinto

    2003-07-01

    The study was set up to evaluate the impact of two commercial larvicide formulations, Bacillus thuringiensis serovar israelensis base (Bti) at 15 ppm/1 min and temephos at 0.03 ppm of active ingredient, used to control Simulium pertinax populations, on associated non-target entomofauna occupying the same breeding sites. The experiments were carried out on the Pedra Branca and Muricana rivers, on the slopes of Serra do Mar massif, municipality of Paraty, state of Rio de Janeiro, Brazil. Bti was applied to the river Pedra Branca and temephosto theriver Muricana. On both rivers, treatment and control sections were labeled as such, each one with two observation posts: slow moving water and fast water regions respectively. Artificial substrata was used to evaluate the abundance of associated entomofauna. Attached immature stages of arthropods were removed from both of its surfaces fortnightly. Were collected, from the two rivers, 28 477 specimens of the entomofauna associated with S. pertinax. The families Hydropsychidae, Chironomidae, Bactidae, Simuliidae, Blephariceridae and Megapodagrionidae were represented. These was an impact of temephos on the entomofauna associated with S. pertinax only in Simuliidae and Chironomidae, and to Bti only in Simuliidae. However, the reduction in their numbers was not statistically significant.

  5. Evaluation of two different culture media for the development of biopesticides based on Bacillus thuringiensis and their application in larvae of Aedes aegypti - doi: 10.4025/actascitechnol.v35i1.13831

    Directory of Open Access Journals (Sweden)

    Samara Ernandes

    2013-01-01

    Full Text Available Normal 0 21 false false false MicrosoftInternetExplorer4 The bacteria Bacillus thuringiensis var. israelensis (Bti generates certain toxins with pesticide action, which can be used on the control of transmissible diseases by culicides, specially Aedes aegypti, the dengue vector. This biopesticide has been produced by submerged fermentation and, in Brazil, this production has been made by very little research centers and, more recently, by a unique small enterprise. For the implementation of a viable vectors control program through biopesticides, some studies about culture media are essential in order to join efficiency and low costs. In this way, agroindustrial wastes or by-products have been used as a nutrient source for the culture media production. In this study, corn steep liquor, a corn industrial processing by-product and tryptose, both with / without sugar addition, were compared as culture media. Cellular growth was evaluated by optical density at 620 nm, spore production by total viable cell count and LC50 by bioassays against 4th instar larvae. Among the four examined substrates, the medium composed by glucose plus corn steep liquor presented the best spore production and bioassay results.

  6. Seleção e caracterização de estirpes de Bacillus thuringiensis tóxicas a Agrotis ipsilon - doi: 10.5102/ucs.v8i2.1142

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    Rafael Silva Menezes

    2011-04-01

    Full Text Available Agrotis ipsilon (Lepidoptera: Noctuidae, conhecida como lagarta rosca, é uma praga polífaga e cosmopolita, causadora de sérios danos em cultivos hortícolas e em sistemas de produção de grãos. Uma das alternativas para o combate a esta praga pode ser a utilização de produtos a base de Bacillus thuringiensis (Bt, bactéria aeróbica, Gram positiva, caracterizada pela produção proteínas tóxicas a insetos. A Embrapa Recursos Genéticos e Biotecnologia possui uma coleção de cerca de 2.300 estirpes de Bt. Neste trabalho cem estirpes desta coleção foram testadas para controle de A. ipsilon e destas, nove foram bastante tóxicas. Essas estirpes pertencem aos sorotipos kurstaki, aizawai, sotto e galleriae. A análise molecular e protéica mostrou a presença dos genes cry1Aa, cry1Ab, cry1Ac, cry1B, cry1C, cry1F, cry2 e cry11 e suas respectivas proteínas, indicando serem elas as proteínas envolvidas na atividade tóxica das estirpes selecionadas.

  7. Suscetibilidade da traça-das-crucíferas a Bacillus thuringiensis em repolho no Distrito Federal - doi: 10.5102/ucs.v8i1.995

    Directory of Open Access Journals (Sweden)

    Lílian Botelho Praça

    2010-10-01

    Full Text Available Plutella xylostella é a mais importante praga das brássicas em todo o mundo. Com o objetivo de se estudar um método de controle desta praga mais eficiente e menos danoso ao meio ambiente, avaliou-se a eficiência de dois bioinseticidas a base de Bacillus thuringiensis (Bt e um inseticida químico a base de deltametrina no controle de P. xylostella em repolho no Distrito Federal. O delineamento experimental foi de blocos ao acaso, com quatro tratamentos e cinco repetições. Cada parcela foi formada por quatro linhas com 20 plantas. Os bioinseticidas a base de Bt foram os mais eficientes no controle de P. xylostella. A testemunha e o tratamento químico não diferiram entre si e apresentaram resultados significativamente inferiores em relação ao percentual de cabeças comercializáveis quando comparado com os bioinseticidas. Este trabalho comprovou a eficiência dos bioinseticidas a base de Bt no controle de P. xylostella em repolho no Distrito Federal.

  8. Elevated atmospheric ozone increases concentration of insecticidal Bacillus thuringiensis (Bt) Cry1Ac protein in Bt Brassica napus and reduces feeding of a Bt target herbivore on the non-transgenic parent

    International Nuclear Information System (INIS)

    Himanen, Sari J.; Nerg, Anne-Marja; Nissinen, Anne; Stewart, C. Neal; Poppy, Guy M.; Holopainen, Jarmo K.

    2009-01-01

    Sustained cultivation of Bacillus thuringiensis (Bt) transgenic crops requires stable transgene expression under variable abiotic conditions. We studied the interactions of Bt toxin production and chronic ozone exposure in Bt cry1Ac-transgenic oilseed rape and found that the insect resistance trait is robust under ozone elevations. Bt Cry1Ac concentrations were higher in the leaves of Bt oilseed rape grown under elevated ozone compared to control treatment, measured either per leaf fresh weight or per total soluble protein of leaves. The mean relative growth rate of a Bt target herbivore, Plutella xylostella L. larvae was negative on Bt plants in all ozone treatments. On the non-transgenic plants, larval feeding damage was reduced under elevated ozone. Our results indicate the need for monitoring fluctuations in Bt toxin concentrations to reveal the potential of ozone exposure for altering dosing of Bt proteins to target and non-target herbivores in field environments experiencing increasing ozone pollution. - Elevated atmospheric ozone can induce fluctuations in insecticidal protein concentrations in transgenic plants

  9. Elevated atmospheric ozone increases concentration of insecticidal Bacillus thuringiensis (Bt) Cry1Ac protein in Bt Brassica napus and reduces feeding of a Bt target herbivore on the non-transgenic parent

    Energy Technology Data Exchange (ETDEWEB)

    Himanen, Sari J. [University of Kuopio, Department of Environmental Science, P.O. Box 1627, FIN-70211 Kuopio (Finland)], E-mail: sari.himanen@uku.fi; Nerg, Anne-Marja [University of Kuopio, Department of Environmental Science, P.O. Box 1627, FIN-70211 Kuopio (Finland); Nissinen, Anne [University of Kuopio, Department of Environmental Science, P.O. Box 1627, FIN-70211 Kuopio (Finland); MTT Agrifood Research Finland, Plant Protection, FIN-31600 Jokioinen (Finland); Stewart, C. Neal [University of Tennessee, Department of Plant Sciences, Knoxville, TN 37996-4561 (United States); Poppy, Guy M. [University of Southampton, School of Biological Sciences, Southampton SO16 7PX (United Kingdom); Holopainen, Jarmo K. [University of Kuopio, Department of Environmental Science, P.O. Box 1627, FIN-70211 Kuopio (Finland)

    2009-01-15

    Sustained cultivation of Bacillus thuringiensis (Bt) transgenic crops requires stable transgene expression under variable abiotic conditions. We studied the interactions of Bt toxin production and chronic ozone exposure in Bt cry1Ac-transgenic oilseed rape and found that the insect resistance trait is robust under ozone elevations. Bt Cry1Ac concentrations were higher in the leaves of Bt oilseed rape grown under elevated ozone compared to control treatment, measured either per leaf fresh weight or per total soluble protein of leaves. The mean relative growth rate of a Bt target herbivore, Plutella xylostella L. larvae was negative on Bt plants in all ozone treatments. On the non-transgenic plants, larval feeding damage was reduced under elevated ozone. Our results indicate the need for monitoring fluctuations in Bt toxin concentrations to reveal the potential of ozone exposure for altering dosing of Bt proteins to target and non-target herbivores in field environments experiencing increasing ozone pollution. - Elevated atmospheric ozone can induce fluctuations in insecticidal protein concentrations in transgenic plants.

  10. Toxicological Evaluation of a Potential Immunosensitizer for Use as a Mucosal Adjuvant—Bacillus thuringiensis Cry1Ac Spore-Crystals: A Possible Inverse Agonist that Deserves Further Investigation

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    Bélin Poletto Mezzomo

    2015-12-01

    Full Text Available In addition to their applicability as biopesticides, Bacillus thuringiensis (Bt Cry1Ac spore-crystals are being researched in the immunology field for their potential as adjuvants in mucosal and parenteral immunizations. We aimed to investigate the hematotoxicity and genotoxicity of Bt spore-crystals genetically modified to express Cry1Ac individually, administered orally (p.o. or with a single intraperitoneal (i.p. injection 24 h before euthanasia, to simulate the routes of mucosal and parenteral immunizations in Swiss mice. Blood samples were used to perform hemogram, and bone marrow was used for the micronucleus test. Cry1Ac presented cytotoxic effects on erythroid lineage in both routes, being more severe in the i.p. route, which also showed genotoxic effects. The greater severity noted in this route, mainly at 6.75 mg/kg, as well as the intermediate effects at 13.5 mg/kg, and the very low hematotoxicity at 27 mg/kg, suggested a possible inverse agonism. The higher immunogenicity for the p.o. route, particularly at 27 mg/kg, suggested that at this dose, Cry 1Ac could potentially be used as a mucosal adjuvant (but not in parenteral immunizations, due to the genotoxic effects observed. This potential should be investigated further, including making an evaluation of the proposed inverse agonism and carrying out cytokine profiling.

  11. 40 CFR 180.1154 - CryIA(c) and CryIC derived delta-endotoxins of Bacillus thuringiensis var. kurstaki encapsulated...

    Science.gov (United States)

    2010-07-01

    ... expression plasmid and cloning vector genetic constructs. 180.1154 Section 180.1154 Protection of Environment... RESIDUES IN FOOD Exemptions From Tolerances § 180.1154 CryIA(c) and CryIC derived delta-endotoxins of... plasmid and cloning vector genetic constructs. CryIA(c) and CryIC derived delta-endotoxins of Bacillus...

  12. Cloning and expression of 130-kd mosquito-larvicidal delta-endotoxin gene of Bacillus thuringiensis var. Israelensis in Escherichia coli.

    Science.gov (United States)

    Angsuthanasombat, C; Chungjatupornchai, W; Kertbundit, S; Luxananil, P; Settasatian, C; Wilairat, P; Panyim, S

    1987-07-01

    Five recombinant E. coli clones exhibiting toxicity to Aedes aegypti larvae were obtained from a library of 800 clones containing XbaI DNA fragments of 110 kb plasmid from B. thuringiensis var. israelensis. All the five clones (pMU 14/258/303/388/679) had the same 3.8-kb insert and encoded a major protein of 130 kDa which was highly toxic to A. aegypti larvae. Three clones (pMU 258/303/388) transcribed the 130 kD a gene in the same direction as that of lac Z promoter of pUC12 vector whereas the transcription of the other two (pMU 14/679) was in the opposite direction. A 1.9-kb fragment of the 3.8 kb insert coded for a protein of 65 kDa. Partial DNA sequence of the 3.8 kb insert, corresponding to the 5'-terminal of the 130 kDa gene, revealed a continuous reading frame, a Shine-Dalgarno sequence and a tentative 5'-regulatory region. These results demonstrated that the 3.8 kb insert is a minimal DNA fragment containing a regulatory region plus the coding sequence of the 130 kDa protein that is highly toxic to mosquito larvae.

  13. Evaluation of methylated soy oil and water-based formulations of Bacillus thuringiensis var. Israelensis and Golden Bear Oil (GB-1111) against anopheles quadrimaculatus larvae in small rice plots.

    Science.gov (United States)

    Dennett, J A; Lampman, R L; Novak, R J; Meisch, M V

    2000-12-01

    The efficacy of formulations containing methylated soybean oil (MSO) alone and with technical-grade Bacillus thuringiensis var. israelensis (Bti) were compared to Golden Bear Oil (GB-1111) and a water-based Bti formulation against 3rd- to 4th-stage Anopheles quadrimaculatus larvae confined to sentinel cages in small rice plots. Three replicates each of MSO with 2% Pyroter added as a surfactant (MSO + PYR), MSO with 2% Pyroter and 4 g of Bti technical powder (MSO + PYR + Bti), GB-1111, a water-based formulation with 4 g of Bti technical powder (Bti + water), and untreated controls were performed. Mosquito larvae were introduced on the 1st day of treatment and at 4 days posttreatment. Mortality was recorded at 24 and 48 h posttreatment for the 1st installation and at 5 days posttreatment for the 2nd installation. The Bti + water formulation provided 71% control and the MSO + PYR + Bti formulation achieved 64% control, whereas MSO + PYR and GB-1111 produced 16 and 18% control, respectively, at 24 h posttreatment. With the exception of MSO + PYR + Bti, which decreased by 2%, the mean percent control increased slightly at 48h posttreatment across remaining treatments, with Bti + water obtaining 72% control. This was significantly higher than GB-1111, which achieved 23% control at 48 h posttreatment. The MSO + PYR and MSO + PYR + Bti formulations yielded 56 and 62% control, respectively, during the same interval and were not significantly different from one another. Formulations containing MSO + PYR exhibited delayed activity similar to GB-1111, with all formulations except MSO + PYR + Bti providing greatest control at 48 h posttreatment. Both MSO formulations (MSO + PYR + Bti and MSO + PYR) were statistically comparable to Bti + water and GB-1111, respectively, at 24 and 48 h posttreatment. None of the formulations exhibited a residual activity adequate enough to control An. quadrimaculatus larvae for up to 5 days.

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

    Science.gov (United States)

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

    2012-10-01

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

  15. Biocontrol of the Sugarcane Borer Eldana saccharina by Expression of the Bacillus thuringiensis cry1Ac7 and Serratia marcescens chiA Genes in Sugarcane-Associated Bacteria

    Science.gov (United States)

    Downing, Katrina J.; Leslie, Graeme; Thomson, Jennifer A.

    2000-01-01

    The cry1Ac7 gene of Bacillus thuringiensis strain 234, showing activity against the sugarcane borer Eldana saccharina, was cloned under the control of the tac promoter. The fusion was introduced into the broad-host-range plasmid pKT240 and the integration vector pJFF350 and without the tac promoter into the broad-host-range plasmids pML122 and pKmM0. These plasmids were introduced into a Pseudomonas fluorescens strain isolated from the phylloplane of sugarcane and the endophytic bacterium Herbaspirillum seropedicae found in sugarcane. The ptac-cry1Ac7 construct was introduced into the chromosome of P. fluorescens using the integration vector pJFF350 carrying the artificial interposon Omegon-Km. Western blot analysis showed that the expression levels of the integrated cry1Ac7 gene were much higher under the control of the tac promoter than under the control of its endogenous promoter. It was also determined that multicopy expression in P. fluorescens and H. seropedicae of ptac-cry1Ac7 carried on pKT240 caused plasmid instability with no detectable protein expression. In H. seropedicae, more Cry1Ac7 toxin was produced when the gene was cloned under the control of the Nmr promoter on pML122 than in the opposite orientation and bioassays showed that the former resulted in higher mortality of E. saccharina larvae than the latter. P. fluorescens 14::ptac-tox resulted in higher mortality of larvae than did P. fluorescens 14::tox. An increased toxic effect was observed when P. fluorescens 14::ptac-tox was combined with P. fluorescens carrying the Serratia marcescens chitinase gene chiA, under the control of the tac promoter, integrated into the chromosome. PMID:10877771

  16. Producción de Bacillus thuringiensis H-14 var. Israelensis utilizando espárrago (Asparagus officinalis y su uso potencial para el control de la Malaria en la Libertad- Perú

    Directory of Open Access Journals (Sweden)

    Franklin Vargas V

    2001-07-01

    Full Text Available Objetivo: Estandarizar un medio, de cultivo utilizando la infusión de espárrago (Asparagus officinalis para la producción masiva de Bacillus thuringiensis (Bti H-14 var. israelensis y determinar el efecto biolarvicida del Bti sobre Anopheles sp en criaderos naturales del distrito Laredo durante los meses de Enero a Diciembre del 2000. Materiales y métodos: Se ensayaron 3 medios a base de infusión de espárrago blanco: M1: 100 mL de la infusión, pH 9; M2: 50 mL de la infusión con 50 mL de buffer fosfato, pH 7; y M3: 25 mL de la infusión con 75 mL de agua destilada, pH 9. Como control se utilizó el medio estándar TPH. La producción de Bti en los medios de cultivo fueron evaluadas para determinar la efectividad biolarvicida a través del LC50 y LC90. El medio de cultivo óptimo (menor LC50 y LC90 sirvió para la producción masiva del Bti, el cual se sometió a bioensayos de laboratorio y aplicaciones en criaderos naturales. La efectividad fue determinada mediante la densidad larvaria pre y post aplicación del Bti. Resultados: El medio de cultivo óptimo para la producción de Bti fue M1 mostrando alta efectividad, con 100% de mortalidad en condiciones de laboratorio y 71-97% de mortalidad en el campo a las 24 horas de exposición con 3 aplicaciones realizadas semanalmente. Conclusiones: M1 es el medio optimo para cultivar Bti, con alta efectividad para controlar larvas de Anopheles en el laboratorio y en el campo.

  17. Evaluación de la toxicidad de proteínas de Bacillus thuringiensis Berliner sobre el picudo del algodonero Anthonomus grandis Boheman

    Directory of Open Access Journals (Sweden)

    Gómez Sylvia

    2006-12-01

    Full Text Available

    El picudo del algodonero, Anthonomus grandis Boheman (Coleoptera: Curculionidae, es considerado la principal plaga del cultivo de algodón en Colombia, no sólo por los daños económicos y sociales que causa, sino porque con sus apariciones tempranas altera e interrumpe el desarrollo de los programas de manejo integrado de plagas. La importancia de un manejo inteligente radica en el hecho de que este insecto se  comporta como plaga ‘clave’, con una capacidad de daño en el cultivo entre 50% y 90%, si no se controla. En el presente estudio se estableció una metodología de bioensayo que determinó la actividad tóxica de  proteínas Cry3Aa y Cry1Ia de cepas de B. thuringiensis sobre larvas de primer instar de A. grandis. En los bioensayos se empleó una dieta artificial mezclada con los extractos bacterianos que contenían el complejo espora-cristal de cada cepa. Los resultados indicaron que la cepa de referencia variedad san diego presentó toxicidad sobre larvas de primer instar de A. grandis, en condiciones de laboratorio, con una concentración letal 50 (CL50 de 147,6143 μg de proteína total por mililitro de volumen final de dieta artificial, representando una alternativa potencial para el manejo de este insecto plaga del cultivo de algodón.

  18. Seleção e caracterização molecular de isolados de Bacillus thuringiensis para o controle de Spodoptera spp.

    Directory of Open Access Journals (Sweden)

    Kelly Christiane Constanski

    2015-08-01

    Full Text Available Resumo:O objetivo deste trabalho foi selecionar e caracterizar molecularmente isolados de Bacillus thuringiensistóxicos a Spodoptera eridaniaeS. frugiperda. Trinta e quatro isolados foram submetidos ao bioensaio, dos quais três foram selecionados e usados para a estimativa da CL50. Os isolados selecionados não diferiram da linhagem padrão HD-1. Na caracterização molecular, identificou-se a presença dos genes cry1 e cry2, nos isolados BR37 e BR94, e dos genes cry4A, cry4B, cry10, cry11 e cyt1 no isolado BR58, o que confirmou o perfil proteico obtido de 130, 70 e 65 kDa. Foram identificados cristais bipiramidais e esféricos. O isolado BR58, apesar de não conter os genes relacionados à toxicidade a Lepidoptera, causa mortalidade em ambas as espécies

  19. Comprehensive analysis of gene expression profiles of the beet armyworm Spodoptera exigua larvae challenged with Bacillus thuringiensis Vip3Aa toxin.

    Directory of Open Access Journals (Sweden)

    Yolanda Bel

    Full Text Available Host-pathogen interactions result in complex relationship, many aspects of which are not completely understood. Vip proteins, which are Bacillus thuringensis (Bt insecticidal toxins produced during the vegetative stage, are selectively effective against specific insect pests. This new group of Bt proteins represents an interesting alternative to the classical Bt Cry toxins because current data suggests that they do not share the same mode of action. We have designed and developed a genome-wide microarray for the beet armyworm Spodoptera exigua, a serious lepidopteran pest of many agricultural crops, and used it to better understand how lepidopteran larvae respond to the treatment with the insecticidal protein Vip3Aa. With this approach, the goal of our study was to evaluate the changes in gene expression levels caused by treatment with sublethal doses of Vip3Aa (causing 99% growth inhibition at 8 and 24 h after feeding. Results indicated that the toxin provoked a wide transcriptional response, with 19% of the microarray unigenes responding significantly to treatment. The number of up- and down-regulated unigenes was very similar. The number of genes whose expression was regulated at 8 h was similar to the number of genes whose expression was regulated after 24 h of treatment. The up-regulated sequences were enriched for genes involved in innate immune response and in pathogen response such as antimicrobial peptides (AMPs and repat genes. The down-regulated sequences were mainly unigenes with homology to genes involved in metabolism. Genes related to the mode of action of Bt Cry proteins were found, in general, to be slightly overexpressed. The present study is the first genome-wide analysis of the response of lepidopteran insects to Vip3Aa intoxication. An insight into the molecular mechanisms and components related to Vip intoxication will allow designing of more effective management strategies for pest control.

  20. substances by Bacillus thuringiensis BAR 3

    African Journals Online (AJOL)

    DR CHARLES O.NWUCHE;Prof. A. R. Popoola

    substances were achieved when the basal medium (3 g/L beef extract, 5g/L peptone and .... room temperature with constant agitation at 160 ... the culture medium was tested by a modification ... to 100oC for 2 min for enzyme inactivation.

  1. Isolamento e caracterização de estirpes de Bacillus thuringiensis coletadas em solos do oeste baiano - doi: 10.5102/ucs.v7i2.999

    Directory of Open Access Journals (Sweden)

    Lílian Botelho Praça

    2010-06-01

    Full Text Available A partir de 21 amostras de solos da região oeste da Bahia, foram isoladas nove estirpes de B. thuringiensis. As estirpes de B. thuringiensis foram testadas contra lagartas de Spodoptera frugiperda, Anticarsia gemmatalis, Plutella xylostella e Anthonomus grandis. Das nove estirpes, duas apresentaram efetividade, a S2183 contra S. frugiperda e a S2186 contra S. frugiperda, A. gemmatalis e P. xylostella. Com relação ao A. grandis, nenhuma das estirpes apresentou mortalidade acima de 70%. Estas estirpes foram submetidas à bioensaios para cálculo da CL50 e a caracterizações bioquímicas e moleculares. Em bioensaio, a S2183 não apresentou efetividade nas doses recomendadas contra os insetos testados e S2186 apresentou uma CL50 de 375 µg/mL apenas para P. xylostella. As estirpes apresentaram duas proteínas principais de 100 e 70 kDa. Somente S2186 apresentou produtos de PCR para o gene cry2 e cristais bipiramidais, apresentando-se semelhante ao padrão B. thuringiensis subespécie kurstaki.

  2. Composition of forage and grain from second-generation insect-protected corn MON 89034 is equivalent to that of conventional corn (Zea mays L.).

    Science.gov (United States)

    Drury, Suzanne M; Reynolds, Tracey L; Ridley, William P; Bogdanova, Natalia; Riordan, Susan; Nemeth, Margaret A; Sorbet, Roy; Trujillo, William A; Breeze, Matthew L

    2008-06-25

    Insect-protected corn hybrids containing Cry insecticidal proteins derived from Bacillus thuringiensis have protection from target pests and provide effective management of insect resistance. MON 89034 hybrids have been developed that produce both the Cry1A.105 and Cry2Ab2 proteins, which provide two independent modes of insecticidal action against the European corn borer ( Ostrinia nubilalis ) and other lepidopteran insect pests of corn. The composition of MON 89034 corn was compared to conventional corn by measuring proximates, fiber, and minerals in forage and by measuring proximates, fiber, amino acids, fatty acids, vitamins, minerals, antinutrients, and secondary metabolites in grain collected from 10 replicated field sites across the United States and Argentina during the 2004-2005 growing seasons. Analyses established that the forage and grain from MON 89034 are compositionally comparable to the control corn hybrid and conventional corn reference hybrids. These findings support the conclusion that MON 89034 is compositionally equivalent to conventional corn hybrids.

  3. The identification of a tetracycline resistance gene tet(M), on a Tn916-like transposon, in the Bacillus cereus group

    DEFF Research Database (Denmark)

    Agersø, Yvonne; Jensen, Lars Bogø; Givskov, Michael Christian

    2002-01-01

    In order to investigate whether resistance genes present in bacteria in manure could transfer to indigenous soil bacteria, resistant isolates belonging to the Bacillus cereus group (Bacillus cereus, Bacillus anthracis and Bacillus thuringiensis) were isolated from farm soil (72 isolates) and manure...

  4. Molecular characterization of Lepidopteran specific Bacillus ...

    African Journals Online (AJOL)

    Guest

    2013-05-15

    May 15, 2013 ... Bacillus thuringiensis (Bt) strains pathogenic to Lepidopteran insects and native to hilly zone soils of. Karnataka (India) were explored. 19 strains were isolated from the soils and identified by morphological and microscopic characters. Toxicity level of the Bt isolates was tested by treating third Instar larvae ...

  5. Production of milk-clotting enzyme by Bacillus subtilis B1 from wheat ...

    African Journals Online (AJOL)

    Three strains, Bacillus subtilis B1, B. subtilis B18 and Bacillus thuringiensis B12, were screened from wheat bran to produce milk-clotting enzyme. Among them, B. subtilis B1 exhibited considerable milkclotting activity with low proteolytic activity. After response surface methodology optimization, milkclotting activity was ...

  6. COMPLEXES OF BISCITRATOGERMANATES AND BISCITRATOSTANATES WITH METALS ARE MODIFIERS OF ACTIVITY OF Bacillus thuringiensis var. іsraelensis PEPTIDASES AND α-Penicillium canescens, Cladosporium cladosporioides AND Aspergillus niger GALACTOIDASES

    Directory of Open Access Journals (Sweden)

    L. D. Varbanets

    2016-06-01

    Full Text Available The aim of the research was to study the effect of a number of coordination compounds of stanum and germanium (compounds 1‒8 as modifiers of activity of peptidases and α-galactosidases. The coordination compounds with the same type structure of were investigated as enzymes effectors. Two types of complexes: 1 [M(H2O6][Ge(НCitr2]4H2O (M = Mg(1, Mn(2, Co(3, Ni(4, Zn(5, containing biscitrate-stanate anion ([Ge(НCitr2]2-, and 2 [M(H2O6][Sn(НCitr2]4H2O (M = Mg(6, Co(7, Ni(8, containing biscitratostanate anion and various hexaaquacations ([M(H2O6]2+, М= Mg, Mn, Co, Ni, Zn were studied. It is shown that the compound 6 which is a biscitratostanate complex containing magnesium ions as metal, can be used for stimulation on 20‒25% of collagenase activity of B. thuringiensis var. israelensis IMV B-7465 peptidase 1 and peptidase 2. Compound 1 (biscitrate-stanate complex containing magnesium ions as metal and 7 (biscitratostanate complex containing cobalt ions as metal and compound 6 in a concentration of 0.001% are able to increase elastolytic activity of peptidase 1 on 55‒58%. However compound 7 has shown the greatest activating effect. It increased the elastolytic activity of peptidase 2 on 100‒140% (for both tested concentrations. This indicates that the compound 7 can further be used as of peptidase 2 elastolytic activity effector. In the study of effect of the considered coordination compounds on the activity of α-galactosidase of Penicillium canescens, Cladosporium cladosporioides and Aspergillus niger was found that when using a number of complexes (1‒2 and 4‒8, there is a slight increase on 12‒20% of enzyme activity of P. canescens, and the maximum effect (~20%, concentration 0.01% was provided by complex 6.

  7. Cross-Resistance between Cry1 Proteins in Fall Armyworm (Spodoptera frugiperda) May Affect the Durability of Current Pyramided Bt Maize Hybrids in Brazil.

    Science.gov (United States)

    Bernardi, Daniel; Salmeron, Eloisa; Horikoshi, Renato Jun; Bernardi, Oderlei; Dourado, Patrick Marques; Carvalho, Renato Assis; Martinelli, Samuel; Head, Graham P; Omoto, Celso

    2015-01-01

    Genetically modified plants expressing insecticidal proteins from Bacillus thuringiensis (Bt) offer valuable options for managing insect pests with considerable environmental and economic benefits. Despite the benefits provided by Bt crops, the continuous expression of these insecticidal proteins imposes strong selection for resistance in target pest populations. Bt maize (Zea mays) hybrids have been successful in controlling fall armyworm (Spodoptera frugiperda), the main maize pest in Brazil since 2008; however, field-evolved resistance to the protein Cry1F has recently been reported. Therefore it is important to assess the possibility of cross-resistance between Cry1F and other Cry proteins expressed in Bt maize hybrids. In this study, an F2 screen followed by subsequent selection on MON 89034 maize was used to select an S. frugiperda strain (RR) able to survive on the Bt maize event MON 89034, which expresses the Cry1A.105 and Cry2Ab2 proteins. Field-collected insects from maize expressing the Cry1F protein (event TC1507) represented most of the positive (resistance allele-containing) (iso)families found. The RR strain showed high levels of resistance to Cry1F, which apparently also conferred high levels of cross resistance to Cry1A.105 and Cry1Ab, but had only low-level (10-fold) resistance to Cry2Ab2. Life history studies to investigate fitness costs associated with the resistance in RR strain revealed only small reductions in reproductive rate when compared to susceptible and heterozygous strains, but the RR strain produced 32.2% and 28.4% fewer females from each female relative to the SS and RS (pooled) strains, respectively. Consistent with the lack of significant resistance to Cry2Ab2, MON 89034 maize in combination with appropriate management practices continues to provide effective control of S. frugiperda in Brazil. Nevertheless, the occurrence of Cry1F resistance in S. frugiperda across Brazil, and the cross-resistance to Cry1Ab and Cry1A.105

  8. Field-evolved resistance to Bt maize in sugarcane borer (Diatraea saccharalis) in Argentina.

    Science.gov (United States)

    Grimi, Damián A; Parody, Betiana; Ramos, María Laura; Machado, Marcos; Ocampo, Federico; Willse, Alan; Martinelli, Samuel; Head, Graham

    2018-04-01

    Maize technologies expressing Bacillus thuringiensis (Bt) insecticidal proteins are widely used in Argentina to control sugarcane borer (Diatraea saccharalis Fabricius). Unexpected D. saccharalis damage was observed to Bt maize events TC1507 (expressing Cry1F) and MON 89034 × MON 88017 (expressing Cry1A.105 and Cry2Ab2) in an isolated area of San Luis Province. Diatraea saccharalis larvae were sampled from MON 89034 × MON 88017 fields in the area to generate a resistant strain (RR), which was subsequently characterized in plant and diet bioassays. Survivorship of the RR strain was high on TC1507 leaf tissue, intermediate on MON 89034 × MON 88017, and low on MON 810 (expressing Cry1Ab). The RR strain had high resistance to Cry1A.105 (186.74-fold) and no resistance to Cry2Ab2 in diet bioassays. These results indicate resistance to Cry1F and Cry1A.105 (and likely cross-resistance between them) but not to Cry1Ab or Cry2Ab2. Resistance to MON 89034 × MON 88017 was functionally recessive. Reviews of grower records suggest that resistance initially evolved to Cry1F, conferring cross-resistance to Cry1A.105, with low refuge compliance as the primary cause. A mitigation plan was implemented in San Luis that included technology rotation, field monitoring, and grower education on best management practices (BMPs) including refuges. In the affected area, the resistance to Cry1F and Cry1A.105 is being managed effectively through use of MON 89034 × MON 88017 and MON 810 in combination with BMPs, and no spread of resistance to other regions has been observed. © 2017 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry. © 2017 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

  9. DNA fingerprinting of spore-forming bacterial isolates, using Bacillus ...

    African Journals Online (AJOL)

    Bc-repetitive extragenic palindromic polymerase chain reaction (Bc-Rep PCR) analysis was conducted on seven Bacillus thuringiensis isolates accessed from the Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ) culture collection and on five local isolates of entomopathogenic spore-forming bacteria.