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Sample records for bt cry1ac protein

  1. Acquisition of Cry1Ac protein by non-target arthropods in Bt soybean fields.

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    Huilin Yu

    Full Text Available Soybean tissue and arthropods were collected in Bt soybean fields in China at different times during the growing season to investigate the exposure of arthropods to the plant-produced Cry1Ac toxin and the transmission of the toxin within the food web. Samples from 52 arthropod species/taxa belonging to 42 families in 10 orders were analysed for their Cry1Ac content using enzyme-linked immunosorbent assay (ELISA. Among the 22 species/taxa for which three samples were analysed, toxin concentration was highest in the grasshopper Atractomorpha sinensis and represented about 50% of the concentration in soybean leaves. Other species/taxa did not contain detectable toxin or contained a concentration that was between 1 and 10% of that detected in leaves. These Cry1Ac-positive arthropods included a number of mesophyll-feeding Hemiptera, a cicadellid, a curculionid beetle and, among the predators, a thomisid spider and an unidentified predatory bug belonging to the Anthocoridae. Within an arthropod species/taxon, the Cry1Ac content sometimes varied between life stages (nymphs/larvae vs. adults and sampling dates (before, during, and after flowering. Our study is the first to provide information on Cry1Ac-expression levels in soybean plants and Cry1Ac concentrations in non-target arthropods in Chinese soybean fields. The data will be useful for assessing the risk of non-target arthropod exposure to Cry1Ac in soybean.

  2. Acquisition of Cry1Ac protein by non-target arthropods in Bt soybean fields.

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    Yu, Huilin; Romeis, Jörg; Li, Yunhe; Li, Xiangju; Wu, Kongming

    2014-01-01

    Soybean tissue and arthropods were collected in Bt soybean fields in China at different times during the growing season to investigate the exposure of arthropods to the plant-produced Cry1Ac toxin and the transmission of the toxin within the food web. Samples from 52 arthropod species/taxa belonging to 42 families in 10 orders were analysed for their Cry1Ac content using enzyme-linked immunosorbent assay (ELISA). Among the 22 species/taxa for which three samples were analysed, toxin concentration was highest in the grasshopper Atractomorpha sinensis and represented about 50% of the concentration in soybean leaves. Other species/taxa did not contain detectable toxin or contained a concentration that was between 1 and 10% of that detected in leaves. These Cry1Ac-positive arthropods included a number of mesophyll-feeding Hemiptera, a cicadellid, a curculionid beetle and, among the predators, a thomisid spider and an unidentified predatory bug belonging to the Anthocoridae. Within an arthropod species/taxon, the Cry1Ac content sometimes varied between life stages (nymphs/larvae vs. adults) and sampling dates (before, during, and after flowering). Our study is the first to provide information on Cry1Ac-expression levels in soybean plants and Cry1Ac concentrations in non-target arthropods in Chinese soybean fields. The data will be useful for assessing the risk of non-target arthropod exposure to Cry1Ac in soybean.

  3. Impact of water content and temperature on the degradation of Cry1Ac protein in leaves and buds of Bt cotton in the soil.

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    Mei-jun Zhang

    Full Text Available Determining the influence of soil environmental factors on degradation of Cry1Ac protein from Bt cotton residues is vital for assessing the ecological risks of this commercialized transgenic crop. In this study, the degradation of Cry1Ac protein in leaves and in buds of Bt cotton in soil was evaluated under different soil water content and temperature settings in the laboratory. An exponential model and a shift-log model were used to fit the degradation dynamics of Cry1Ac protein and estimate the DT50 and DT90 values. The results showed that Cry1Ac protein in the leaves and buds underwent rapid degradation in the early stage (before day 48, followed by a slow decline in the later stage under different soil water content and temperature. Cry1Ac protein degraded the most rapidly in the early stage at 35°C with 70% soil water holding capacity. The DT50 values were 12.29 d and 10.17 d and the DT90 values were 41.06 d and 33.96 d in the leaves and buds, respectively. Our findings indicated that the soil temperature was a major factor influencing the degradation of Cry1Ac protein from Bt cotton residues. Additionally, the relative higher temperature (25°C and 35°C was found to be more conducive to degradation of Cry1Ac protein in the soil and the greater water content (100%WHC retarded the process. These findings suggested that under appropriate soil temperature and water content, Cry1Ac protein from Bt cotton residues will not persist and accumulate in soil.

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

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    Yuan, Yiyang; Krogh, Paul Henning; Bai, Xue; Roelofs, Dick; Chen, Fajun; Zhu-Salzman, Keyan; Liang, Yuyong; Sun, Yucheng; Ge, Feng

    2014-01-01

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

  5. No impact of Bt soybean that express Cry1Ac protein on biological traits of Euschistus heros (Hemiptera, Pentatomidae and its egg parasitoid Telenomus podisi (Hymenoptera, Platygastridae

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    Gabriela Vieira Silva

    2014-09-01

    Full Text Available No impact of Bt soybean that express Cry1Ac protein on biological traits of Euschistus heros (Hemiptera, Pentatomidae and its egg parasitoid Telenomus podisi (Hymenoptera, Platygastridae. Biological traits of the stink bug Euschistus heros and its main biological control agent Telenomus podisi were evaluated under controlled environmental conditions (25 ± 2ºC; 60 ± 10% RH; and 14/10 h photoperiod by placing first instar nymphs into Petri dishes with pods originating from two soybean isolines (Bt-soybean MON 87701 × MON 89788, which expresses the Cry1Ac protein, and its near non-Bt isoline A5547 where they remained until the adult stage. Due to gregarious behavior exhibited by first instar nymphs, they were individualized only when at the second instar. Adults were separated by sex and weighed, and pronotum width of each individual was subsequently measured. They were placed into plastic boxes containing soybean grains of the same soybean isoline as food source. Egg viability and female fecundity were assessed in adult individuals. Adult females of T. podisi (up to 24h old were placed with eggs of E. heros from mothers reared on both soybean isolines. Nymphal development time, insect weight, pronotum width, sex ratio, female fecundity, and egg viability (% emergence of Euschistus heros did not differ between treatments. Eggto-adult development time, female longevity, sex ratio, and percentage of parasitized eggs were not impacted by the Bt-soybean (expressing Cry1Ac protein. Results indicate that the Bt-soybean, MON 87701 × MON 89788, has no direct significant impact on the two studied species.

  6. Synthetic fusion-protein containing domains of Bt Cry1Ac and Allium sativum lectin (ASAL) conferred enhanced insecticidal activity against major lepidopteran pests.

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    Tajne, Sunita; Boddupally, Dayakar; Sadumpati, Vijayakumar; Vudem, Dashavantha Reddy; Khareedu, Venkateswara Rao

    2014-02-10

    Different transgenic crop plants, developed with δ-endotoxins of Bacillus thuringiensis (Bt) and mannose-specific plant lectins, exhibited significant protection against chewing and sucking insects. In the present study, a synthetic gene (cry-asal) encoding the fusion-protein having 488 amino acids, comprising DI and DII domains from Bt Cry1Ac and Allium sativum agglutinin (ASAL), was cloned and expressed in Escherichia coli. Ligand blot analysis disclosed that the fusion-protein could bind to more number of receptors of brush border membrane vesicle (BBMV) proteins of Helicoverpa armigera. Artificial diet bioassays revealed that 0.025 μg/g and 0.50 μg/g of fusion-protein were sufficient to cause 100% mortality in Pectinophora gossypiella and H. armigera insects, respectively. As compared to Cry1Ac, the fusion-protein showed enhanced (8-fold and 30-fold) insecticidal activity against two major lepidopteran pests. Binding of fusion-protein to the additional receptors in the midgut cells of insects is attributable to its enhanced entomotoxic effect. The synthetic gene, first of its kind, appears promising and might serve as a potential candidate for engineering crop plants against major insect pests.

  7. 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

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    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.

  8. Increased frequency of pink bollworm resistance to Bt toxin Cry1Ac in China.

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    Wan, Peng; Huang, Yunxin; Wu, Huaiheng; Huang, Minsong; Cong, Shengbo; Tabashnik, Bruce E; Wu, Kongming

    2012-01-01

    Transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) kill some key insect pests, but evolution of resistance by pests can reduce their efficacy. The main approach for delaying pest adaptation to Bt crops uses non-Bt host plants as "refuges" to increase survival of susceptible pests. To delay evolution of pest resistance to transgenic cotton producing Bt toxin Cry1Ac, the United States and some other countries have required refuges of non-Bt cotton, while farmers in China have relied on "natural" refuges of non-Bt host plants other than cotton. The "natural" refuge strategy focuses on cotton bollworm (Helicoverpa armigera), the primary target of Bt cotton in China that attacks many crops, but it does not apply to another major pest, pink bollworm (Pectinophora gossypiella), which feeds almost entirely on cotton in China. Here we report data showing field-evolved resistance to Cry1Ac by pink bollworm in the Yangtze River Valley of China. Laboratory bioassay data from 51 field-derived strains show that the susceptibility to Cry1Ac was significantly lower during 2008 to 2010 than 2005 to 2007. The percentage of field populations yielding one or more survivors at a diagnostic concentration of Cry1Ac increased from 0% in 2005-2007 to 56% in 2008-2010. However, the median survival at the diagnostic concentration was only 1.6% from 2008 to 2010 and failure of Bt cotton to control pink bollworm has not been reported in China. The early detection of resistance reported here may promote proactive countermeasures, such as a switch to transgenic cotton producing toxins distinct from Cry1A toxins, increased planting of non-Bt cotton, and integration of other management tactics together with Bt cotton.

  9. Increased frequency of pink bollworm resistance to Bt toxin Cry1Ac in China.

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    Peng Wan

    Full Text Available Transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt kill some key insect pests, but evolution of resistance by pests can reduce their efficacy. The main approach for delaying pest adaptation to Bt crops uses non-Bt host plants as "refuges" to increase survival of susceptible pests. To delay evolution of pest resistance to transgenic cotton producing Bt toxin Cry1Ac, the United States and some other countries have required refuges of non-Bt cotton, while farmers in China have relied on "natural" refuges of non-Bt host plants other than cotton. The "natural" refuge strategy focuses on cotton bollworm (Helicoverpa armigera, the primary target of Bt cotton in China that attacks many crops, but it does not apply to another major pest, pink bollworm (Pectinophora gossypiella, which feeds almost entirely on cotton in China. Here we report data showing field-evolved resistance to Cry1Ac by pink bollworm in the Yangtze River Valley of China. Laboratory bioassay data from 51 field-derived strains show that the susceptibility to Cry1Ac was significantly lower during 2008 to 2010 than 2005 to 2007. The percentage of field populations yielding one or more survivors at a diagnostic concentration of Cry1Ac increased from 0% in 2005-2007 to 56% in 2008-2010. However, the median survival at the diagnostic concentration was only 1.6% from 2008 to 2010 and failure of Bt cotton to control pink bollworm has not been reported in China. The early detection of resistance reported here may promote proactive countermeasures, such as a switch to transgenic cotton producing toxins distinct from Cry1A toxins, increased planting of non-Bt cotton, and integration of other management tactics together with Bt cotton.

  10. Cross-resistance and interactions between Bt toxins Cry1Ac and Cry2Ab against the cotton bollworm.

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    Wei, Jizhen; Guo, Yuyuan; Liang, Gemei; Wu, Kongming; Zhang, Jie; Tabashnik, Bruce E; Li, Xianchun

    2015-01-14

    To delay evolution of pest resistance to transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt), the "pyramid" strategy uses plants that produce two or more toxins that kill the same pest. We conducted laboratory diet experiments with the cotton bollworm, Helicoverpa armigera, to evaluate cross-resistance and interactions between two toxins in pyramided Bt cotton (Cry1Ac and Cry2Ab). Selection with Cry1Ac for 125 generations produced 1000-fold resistance to Cry1Ac and 6.8-fold cross-resistance to Cry2Ab. Selection with Cry2Ab for 29 generations caused 5.6-fold resistance to Cry2Ab and 61-fold cross-resistance to Cry1Ac. Without exposure to Bt toxins, resistance to both toxins decreased. For each of the four resistant strains examined, 67 to 100% of the combinations of Cry1Ac and Cry2Ab tested yielded higher than expected mortality, reflecting synergism between these two toxins. Results showing minor cross-resistance to Cry2Ab caused by selection with Cry1Ac and synergism between these two toxins against resistant insects suggest that plants producing both toxins could prolong the efficacy of Bt cotton against this pest in China. Including toxins against which no cross-resistance occurs and integrating Bt cotton with other control tactics could also increase the sustainability of management strategies.

  11. A toxin-binding alkaline phosphatase fragment synergizes Bt toxin Cry1Ac against susceptible and resistant Helicoverpa armigera.

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    Chen, Wenbo; Liu, Chenxi; Xiao, Yutao; Zhang, Dandan; Zhang, Yongdong; Li, Xianchun; Tabashnik, Bruce E; Wu, Kongming

    2015-01-01

    Evolution of resistance by insects threatens the continued success of pest control using insecticidal crystal (Cry) proteins from the bacterium Bacillus thuringiensis (Bt) in sprays and transgenic plants. In this study, laboratory selection with Cry1Ac yielded five strains of cotton bollworm, Helicoverpa armigera, with resistance ratios at the median lethal concentration (LC50) of activated Cry1Ac ranging from 22 to 1700. Reduced activity and reduced transcription of an alkaline phosphatase protein that binds Cry1Ac was associated with resistance to Cry1Ac in the four most resistant strains. A Cry1Ac-binding fragment of alkaline phosphatase from H. armigera (HaALP1f) was not toxic by itself, but it increased mortality caused by Cry1Ac in a susceptible strain and in all five resistant strains. Although synergism of Bt toxins against susceptible insects by toxin-binding fragments of cadherin and aminopeptidase N has been reported previously, the results here provide the first evidence of synergism of a Bt toxin by a toxin-binding fragment of alkaline phosphatase. The results here also provide the first evidence of synergism of a Bt toxin by any toxin-binding peptide against resistant insects.

  12. A toxin-binding alkaline phosphatase fragment synergizes Bt toxin Cry1Ac against susceptible and resistant Helicoverpa armigera.

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    Wenbo Chen

    Full Text Available Evolution of resistance by insects threatens the continued success of pest control using insecticidal crystal (Cry proteins from the bacterium Bacillus thuringiensis (Bt in sprays and transgenic plants. In this study, laboratory selection with Cry1Ac yielded five strains of cotton bollworm, Helicoverpa armigera, with resistance ratios at the median lethal concentration (LC50 of activated Cry1Ac ranging from 22 to 1700. Reduced activity and reduced transcription of an alkaline phosphatase protein that binds Cry1Ac was associated with resistance to Cry1Ac in the four most resistant strains. A Cry1Ac-binding fragment of alkaline phosphatase from H. armigera (HaALP1f was not toxic by itself, but it increased mortality caused by Cry1Ac in a susceptible strain and in all five resistant strains. Although synergism of Bt toxins against susceptible insects by toxin-binding fragments of cadherin and aminopeptidase N has been reported previously, the results here provide the first evidence of synergism of a Bt toxin by a toxin-binding fragment of alkaline phosphatase. The results here also provide the first evidence of synergism of a Bt toxin by any toxin-binding peptide against resistant insects.

  13. Distribution and Metabolism of Bt-Cry1Ac Toxin in Tissues and Organs of the Cotton Bollworm, Helicoverpa armigera

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    Zhuoya Zhao

    2016-07-01

    Full Text Available Crystal (Cry proteins derived from Bacillus thuringiensis (Bt have been widely used in transgenic crops due to their toxicity against insect pests. However, the distribution and metabolism of these toxins in insect tissues and organs have remained obscure because the target insects do not ingest much toxin. In this study, several Cry1Ac-resistant strains of Helicoverpa armigera, fed artificial diets containing high doses of Cry1Ac toxin, were used to investigate the distribution and metabolism of Cry1Ac in their bodies. Cry1Ac was only detected in larvae, not in pupae or adults. Also, Cry1Ac passed through the midgut into other tissues, such as the hemolymph and fat body, but did not reach the larval integument. Metabolic tests revealed that Cry1Ac degraded most rapidly in the fat body, followed by the hemolymph, peritrophic membrane and its contents. The toxin was metabolized slowly in the midgut, but was degraded in all locations within 48 h. These findings will improve understanding of the functional mechanism of Bt toxins in target insects and the biotransfer and the bioaccumulation of Bt toxins in arthropod food webs in the Bt crop ecosystem.

  14. Distribution and Metabolism of Bt-Cry1Ac Toxin in Tissues and Organs of the Cotton Bollworm, Helicoverpa armigera.

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    Zhao, Zhuoya; Li, Yunhe; Xiao, Yutao; Ali, Abid; Dhiloo, Khalid Hussain; Chen, Wenbo; Wu, Kongming

    2016-01-01

    Crystal (Cry) proteins derived from Bacillus thuringiensis (Bt) have been widely used in transgenic crops due to their toxicity against insect pests. However, the distribution and metabolism of these toxins in insect tissues and organs have remained obscure because the target insects do not ingest much toxin. In this study, several Cry1Ac-resistant strains of Helicoverpa armigera, fed artificial diets containing high doses of Cry1Ac toxin, were used to investigate the distribution and metabolism of Cry1Ac in their bodies. Cry1Ac was only detected in larvae, not in pupae or adults. Also, Cry1Ac passed through the midgut into other tissues, such as the hemolymph and fat body, but did not reach the larval integument. Metabolic tests revealed that Cry1Ac degraded most rapidly in the fat body, followed by the hemolymph, peritrophic membrane and its contents. The toxin was metabolized slowly in the midgut, but was degraded in all locations within 48 h. These findings will improve understanding of the functional mechanism of Bt toxins in target insects and the biotransfer and the bioaccumulation of Bt toxins in arthropod food webs in the Bt crop ecosystem.

  15. Baseline susceptibility to Cry1Ac insecticidal protein in Heliothis virescens (Lepidoptera: Noctuidae) populations in Brazil.

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    Albernaz, K C; Merlin, B L; Martinelli, S; Head, G P; Omoto, C

    2013-08-01

    The tobacco budworm, Heliothis virescens (F.), is one of the target pests of genetically modified cotton expressing Cry1Ac insecticidal protein (Bt cotton) derived from Bacillus thuringiensis Berliner. This study was conducted to evaluate the susceptibility of field-collected populations of H. virescens to Cry1Ac to establish a baseline for use in an insect resistance management program for Bt cotton in Brazil. Insects were sampled from the main Brazilian cotton-growing regions (Bahia, Mato Grosso, Mato Grosso do Sul, and Goiás) during the cropping seasons of 2007/08 and 2008/09. Cry1Ac susceptibility was estimated by using diet incorporation bioassays. H. virescens was highly susceptible to Cry1Ac protein. The estimated LC50 values varied from 0.18 to 0.66 microg of Cry1Ac/ml of diet among the 2007-2008 populations (approximately 3.7-fold variation). Similarly, the EC50 values based on growth inhibition ranged from 0.0053 to 0.0161 microg of Cry1Ac/ml of diet for the 2007-2008 populations (approximately 3.0-fold variation). A joint analysis of the mortality data across all tested populations was used to develop and validate the diagnostic concentrations of 3.1 and 5.6 microg of Cry1Ac/ml of diet, the upper bound of the confidence interval and twice the LC99 were selected, for resistance monitoring programs of H. virescens to Cry1Ac protein in Brazil.

  16. Similar genetic basis of resistance to Bt toxin Cry1Ac in Boll-selected and diet-selected strains of pink bollworm.

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    Fabrick, Jeffrey A; Tabashnik, Bruce E

    2012-01-01

    Genetically engineered cotton and corn plants producing insecticidal Bacillus thuringiensis (Bt) toxins kill some key insect pests. Yet, evolution of resistance by pests threatens long-term insect control by these transgenic Bt crops. We compared the genetic basis of resistance to Bt toxin Cry1Ac in two independently derived, laboratory-selected strains of a major cotton pest, the pink bollworm (Pectinophora gossypiella [Saunders]). The Arizona pooled resistant strain (AZP-R) was started with pink bollworm from 10 field populations and selected with Cry1Ac in diet. The Bt4R resistant strain was started with a long-term susceptible laboratory strain and selected first with Bt cotton bolls and later with Cry1Ac in diet. Previous work showed that AZP-R had three recessive mutations (r1, r2, and r3) in the pink bollworm cadherin gene (PgCad1) linked with resistance to Cry1Ac and Bt cotton producing Cry1Ac. Here we report that inheritance of resistance to a diagnostic concentration of Cry1Ac was recessive in Bt4R. In interstrain complementation tests for allelism, F(1) progeny from crosses between AZP-R and Bt4R were resistant to Cry1Ac, indicating a shared resistance locus in the two strains. Molecular analysis of the Bt4R cadherin gene identified a novel 15-bp deletion (r4) predicted to cause the loss of five amino acids upstream of the Cry1Ac-binding region of the cadherin protein. Four recessive mutations in PgCad1 are now implicated in resistance in five different strains, showing that mutations in cadherin are the primary mechanism of resistance to Cry1Ac in laboratory-selected strains of pink bollworm from Arizona.

  17. Similar genetic basis of resistance to Bt toxin Cry1Ac in Boll-selected and diet-selected strains of pink bollworm.

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    Jeffrey A Fabrick

    Full Text Available Genetically engineered cotton and corn plants producing insecticidal Bacillus thuringiensis (Bt toxins kill some key insect pests. Yet, evolution of resistance by pests threatens long-term insect control by these transgenic Bt crops. We compared the genetic basis of resistance to Bt toxin Cry1Ac in two independently derived, laboratory-selected strains of a major cotton pest, the pink bollworm (Pectinophora gossypiella [Saunders]. The Arizona pooled resistant strain (AZP-R was started with pink bollworm from 10 field populations and selected with Cry1Ac in diet. The Bt4R resistant strain was started with a long-term susceptible laboratory strain and selected first with Bt cotton bolls and later with Cry1Ac in diet. Previous work showed that AZP-R had three recessive mutations (r1, r2, and r3 in the pink bollworm cadherin gene (PgCad1 linked with resistance to Cry1Ac and Bt cotton producing Cry1Ac. Here we report that inheritance of resistance to a diagnostic concentration of Cry1Ac was recessive in Bt4R. In interstrain complementation tests for allelism, F(1 progeny from crosses between AZP-R and Bt4R were resistant to Cry1Ac, indicating a shared resistance locus in the two strains. Molecular analysis of the Bt4R cadherin gene identified a novel 15-bp deletion (r4 predicted to cause the loss of five amino acids upstream of the Cry1Ac-binding region of the cadherin protein. Four recessive mutations in PgCad1 are now implicated in resistance in five different strains, showing that mutations in cadherin are the primary mechanism of resistance to Cry1Ac in laboratory-selected strains of pink bollworm from Arizona.

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

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

    2014-01-01

    . A customized microarray was used to study gene expression in F. candida specimens that were exposed to Cry1Ab and Cry1Ac. All selected transcripts were subsequently confirmed by qPCR. Eleven transcripts were finally verified, and three of them were annotated. The responses of all eleven transcripts were...

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

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

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

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    Bruce E Tabashnik

    Full Text Available Evolution of resistance in pests threatens the long-term efficacy of insecticidal proteins from Bacillus thuringiensis (Bt used in sprays and transgenic crops. Previous work showed that genetically modified Bt toxins Cry1AbMod and Cry1AcMod effectively countered resistance to native Bt toxins Cry1Ab and Cry1Ac in some pests, including pink bollworm (Pectinophora gossypiella. Here we report that Cry1AbMod and Cry1AcMod were also effective against a laboratory-selected strain of pink bollworm resistant to Cry2Ab as well as to Cry1Ab and Cry1Ac. Resistance ratios based on the concentration of toxin killing 50% of larvae for the resistant strain relative to a susceptible strain were 210 for Cry2Ab, 270 for Cry1Ab, and 310 for Cry1Ac, but only 1.6 for Cry1AbMod and 2.1 for Cry1AcMod. To evaluate the interactions among toxins, we tested combinations of Cry1AbMod, Cry1Ac, and Cry2Ab. For both the resistant and susceptible strains, the net results across all concentrations tested showed slight but significant synergism between Cry1AbMod and Cry2Ab, whereas the other combinations of toxins did not show consistent synergism or antagonism. The results suggest that the modified toxins might be useful for controlling populations of pink bollworm resistant to Cry1Ac, Cry2Ab, or both.

  1. Frequency of alleles conferring resistance to the Bt toxins Cry1Ac and Cry2Ab in Australian populations of Helicoverpa armigera (Lepidoptera: Noctuidae).

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    Mahon, R J; Olsen, K M; Downes, S; Addison, S

    2007-12-01

    Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) is an important lepidopteran pest of cotton (Gossypium spp.) in Australia and the Old World. From 2002, F2 screens were used to examine the frequency of resistance alleles in Australian populations of H. armigera to Bacillus thuringiensis (Bt) CrylAc and Cry2Ab, the two insecticidal proteins present in the transgenic cotton Bollgard II. At that time, Ingard (expressing Cry1Ac) cotton had been grown in Australia for seven seasons, and Bollgard II was about to be commercially released. The principal objective of our study was to determine whether sustained exposure caused an elevated frequency of alleles conferring resistance to Cry1Ac in a species with a track record of evolving resistance to conventional insecticides. No major alleles conferring resistance to Cry1Ac were found. The frequency of resistance alleles for Cry1Ac was <0.0003, with a 95% credibility interval between 0 and 0.0009. In contrast, alleles conferring resistance to Cry2Ab were found at a frequency of 0.0033 (0.0017, 0.0055). The first isolation of this allele was found before the widespread deployment of Bollgard II. For both toxins the experiment-wise detection probability was 94.4%. Our results suggest that alleles conferring resistance to Cry1Ac are rare and that a relatively high baseline frequency of alleles conferring resistance to Cry2Ab existed before the introduction of Bt cotton containing this toxin.

  2. Functional validation of cadherin as a receptor of Bt toxin Cry1Ac in Helicoverpa armigera utilizing the CRISPR/Cas9 system.

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    Wang, Jing; Zhang, Haonan; Wang, Huidong; Zhao, Shan; Zuo, Yayun; Yang, Yihua; Wu, Yidong

    2016-09-01

    Cadherins have been identified as receptors of Bacillus thuringiensis (Bt) Cry1A toxins in several lepidopteran insects including the cotton bollworm, Helicoverpa armigera. Disruption of the cadherin gene HaCad has been genetically linked to resistance to Bt toxin Cry1Ac in H. armigera. By using the CRISPR/Cas9 genome editing system (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated protein 9), HaCad from the Cry1Ac-susceptible SCD strain of H. armigera was successfully knocked out. A single positive CRISPR event with a frame shift deletion of 4 nucleotides was identified and made homozygous to create a knockout line named SCD-Cad. Western blotting confirmed that HaCad was no longer expressed in the SCD-Cad line while an intact HaCad of 210 kDa was present in the parental SCD strain. Insecticide bioassays were used to show that SCD-Cad exhibited 549-fold resistance to Cry1Ac compared with SCD, but no significant change in susceptibility to Cry2Ab. Our results not only provide strong reverse genetics evidence for HaCad as a functional receptor of Cry1Ac, but also demonstrate that the CRISPR/Cas9 technique can act as a powerful and efficient genome editing tool to study gene function in a global agricultural pest, H. armigera.

  3. The cultivation of Bt corn producing Cry1Ac toxins does not adversely affect non-target arthropods.

    Science.gov (United States)

    Guo, Yanyan; Feng, Yanjie; Ge, Yang; Tetreau, Guillaume; Chen, Xiaowen; Dong, Xuehui; Shi, Wangpeng

    2014-01-01

    Transgenic corn producing Cry1Ac toxins from Bacillus thuringiensis (Bt) provides effective control of Asian corn borer, Ostrinia furnacalis (Guenée), and thus reduces insecticide applications. However, whether Bt corn exerts undesirable effects on non-target arthropods (NTAs) is still controversial. We conducted a 2-yr study in Shangzhuang Agricultural Experiment Station to assess the potential impact of Bt corn on field population density, biodiversity, community composition and structure of NTAs. On each sampling date, the total abundance, Shannon's diversity index, Pielou's evenness index and Simpson's diversity index were not significantly affected by Bt corn as compared to non-Bt corn. The "sampling dates" had a significant effect on these indices, but no clear tendencies related to "Bt corn" or "sampling dates X corn variety" interaction were recorded. Principal response curve analysis of variance indicated that Bt corn did not alter the distribution of NTAs communities. Bray-Curtis dissimilarity and distance analysis showed that Cry1Ac toxin exposure did not increase community dissimilarities between Bt and non-Bt corn plots and that the evolution of non-target arthropod community was similar on the two corn varieties. The cultivation of Bt corn failed to show any detrimental evidence on the density of non-target herbivores, predators and parasitoids. The composition of herbivores, predators and parasitoids was identical in Bt and non-Bt corn plots. Taken together, results from the present work support that Bt corn producing Cry1Ac toxins does not adversely affect NTAs.

  4. High Susceptibility to Cry1Ac and Low Resistance Allele Frequency Reduce the Risk of Resistance of Helicoverpa armigera to Bt Soybean in Brazil

    Science.gov (United States)

    Bacalhau, Fabiana B.; Amado, Douglas; Carvalho, Renato A.; Martinelli, Samuel; Head, Graham P.; Omoto, Celso

    2016-01-01

    The Old World bollworm, Helicoverpa armigera (Hübner), was recently introduced into Brazil, where it has caused extensive damage to cotton and soybean crops. MON 87701 × MON 89788 soybean, which expresses the Bt protein Cry1Ac, was recently deployed in Brazil, providing high levels of control against H. armigera. To assess the risk of resistance to the Cry1Ac protein expressed by MON 87701 × MON 89788 soybean in Brazil, we conducted studies to evaluate the baseline susceptibility of H. armigera to Cry1Ac, in planta efficacy including the assessment of the high-dose criterion, and the initial resistance allele frequency based on an F2 screen. The mean Cry1Ac lethal concentration (LC50) ranged from 0.11 to 1.82 μg·mL−1 of diet among all H. armigera field populations collected from crop seasons 2013/14 to 2014/15, which indicated about 16.5-fold variation. MON 87701 × MON 89788 soybean exhibited a high level of efficacy against H. armigera and most likely met the high dose criterion against this target species in leaf tissue dilution bioassays up to 50 times. A total of 212 F2 family lines of H. armigera were established from field collections sampled from seven locations across Brazil and were screened for the presence of MON 87701 × MON 89788 soybean resistance alleles. None of the 212 families survived on MON 87701 × MON 89788 soybean leaf tissue (estimated allele frequency = 0.0011). The responses of H. armigera to Cry1Ac protein, high susceptibility to MON 87701 × MON 89788 soybean, and low frequency of resistance alleles across the main soybean-producing regions support the assumptions of a high-dose/refuge strategy. However, maintenance of reasonable compliance with the refuge recommendation will be essential to delay the evolution of resistance in H. armigera to MON 87701 × MON 89788 soybean in Brazil. PMID:27532632

  5. Bt crops producing Cry1Ac, Cry2Ab and Cry1F do not harm the green lacewing, Chrysoperla rufilabris.

    Directory of Open Access Journals (Sweden)

    Jun-Ce Tian

    Full Text Available The biological control function provided by natural enemies is regarded as a protection goal that should not be harmed by the application of any new pest management tool. Plants producing Cry proteins from the bacterium, Bacillus thuringiensis (Bt, have become a major tactic for controlling pest Lepidoptera on cotton and maize and risk assessment studies are needed to ensure they do not harm important natural enemies. However, using Cry protein susceptible hosts as prey often compromises such studies. To avoid this problem we utilized pest Lepidoptera, cabbage looper (Trichoplusia ni and fall armyworm (Spodoptera frugiperda, that were resistant to Cry1Ac produced in Bt broccoli (T. ni, Cry1Ac/Cry2Ab produced in Bt cotton (T. ni, and Cry1F produced in Bt maize (S. frugiperda. Larvae of these species were fed Bt plants or non-Bt plants and then exposed to predaceous larvae of the green lacewing Chrysoperla rufilabris. Fitness parameters (larval survival, development time, fecundity and egg hatch of C. rufilabris were assessed over two generations. There were no differences in any of the fitness parameters regardless if C. rufilabris consumed prey (T. ni or S. frugiperda that had consumed Bt or non-Bt plants. Additional studies confirmed that the prey contained bioactive Cry proteins when they were consumed by the predator. These studies confirm that Cry1Ac, Cry2Ab and Cry1F do not pose a hazard to the important predator C. rufilabris. This study also demonstrates the power of using resistant hosts when assessing the risk of genetically modified plants on non-target organisms.

  6. The cultivation of Bt corn producing Cry1Ac toxins does not adversely affect non-target arthropods.

    Directory of Open Access Journals (Sweden)

    Yanyan Guo

    Full Text Available Transgenic corn producing Cry1Ac toxins from Bacillus thuringiensis (Bt provides effective control of Asian corn borer, Ostrinia furnacalis (Guenée, and thus reduces insecticide applications. However, whether Bt corn exerts undesirable effects on non-target arthropods (NTAs is still controversial. We conducted a 2-yr study in Shangzhuang Agricultural Experiment Station to assess the potential impact of Bt corn on field population density, biodiversity, community composition and structure of NTAs. On each sampling date, the total abundance, Shannon's diversity index, Pielou's evenness index and Simpson's diversity index were not significantly affected by Bt corn as compared to non-Bt corn. The "sampling dates" had a significant effect on these indices, but no clear tendencies related to "Bt corn" or "sampling dates X corn variety" interaction were recorded. Principal response curve analysis of variance indicated that Bt corn did not alter the distribution of NTAs communities. Bray-Curtis dissimilarity and distance analysis showed that Cry1Ac toxin exposure did not increase community dissimilarities between Bt and non-Bt corn plots and that the evolution of non-target arthropod community was similar on the two corn varieties. The cultivation of Bt corn failed to show any detrimental evidence on the density of non-target herbivores, predators and parasitoids. The composition of herbivores, predators and parasitoids was identical in Bt and non-Bt corn plots. Taken together, results from the present work support that Bt corn producing Cry1Ac toxins does not adversely affect NTAs.

  7. Quantification of Cry1Ac protein at different stages of plant growth in cotton (Gossypium hirsutum L.

    Directory of Open Access Journals (Sweden)

    Muhammad Idrees Khan

    2012-12-01

    Full Text Available The present study was conducted at Central Cotton Research Institute, Multan, Pakistan during cotton growing season 2009-10. Nine cotton cultivars with Cry 1 Ac gene (Mon 531 event selected for current experiment to characterize the toxin level of Cry1Ac protein in different Bt cotton cultivars and to record the variation in Cry1Ac protein at various plant growth stages. It was found that age of plant was having an influence on the expression of gene. Maximum level of endotoxin (0.373g/g was observed in genotype CIM-595 at 100 days of planting. While minimum value (0.166g/g was observed in genotype V-1 at 160 days of planting. Similarly different cotton genotypes showed different boll worm damage % at different growth stages. CEMB-2 was found to be most susceptible genotype showing 93.33 % boll worm damage at 160 days after planting. While V-5 proved to be most resistant showing no boll worm damage at 70 and 100 days after planting.

  8. A comprehensive assessment of the effects of Bt cotton on Coleomegilla maculata demonstrates no detrimental effects by Cry1Ac and Cry2Ab.

    Directory of Open Access Journals (Sweden)

    Yunhe Li

    Full Text Available The ladybird beetle, Coleomegilla maculata (DeGeer, is a common and abundant predator in many cropping systems. Its larvae and adults are predaceous, feeding on aphids, thrips, lepidopteran larvae and plant tissues, such as pollen. Therefore, this species is exposed to insecticidal proteins expressed in insect-resistant, genetically engineered cotton expressing Cry proteins derived from Bacillus thuringiensis (Bt. A tritrophic bioassay was conduced to evaluate the potential impact of Cry2Ab- and Cry1Ac-expressing cotton on fitness parameters of C. maculata using Bt-susceptible and -resistant larvae of Trichoplusia ni as prey. Coleomegilla maculata survival, development time, adult weight and fecundity were not different when they were fed with resistant T. ni larvae reared on either Bt or control cotton. To ensure that C. maculata were not sensitive to the tested Cry toxins independent from the plant background and to add certainty to the hazard assessment, C. maculata larvae were fed artificial diet incorporated with Cry2Ab, Cry1Ac or both at >10 times higher concentrations than in cotton tissue. Artificial diet containing E-64 was included as a positive control. No differences were detected in any life-table parameters between Cry protein-containing diet treatments and the control diet. In contrast, larvae of C. maculata fed the E-64 could not develop to the pupal stage and the 7-d larval weight was significantly negatively affected. In both feeding assays, the stability and bioactivity of Cry proteins in the food sources were confirmed by ELISA and sensitive-insect bioassays. Our results show that C. maculata is not affected by Bt cotton and is not sensitive to Cry2Ab and Cry1Ac at concentrations exceeding the levels in Bt cotton, thus demonstrating that Bt cotton will pose a negligible risk to C. maculata. More importantly, this study demonstrates a comprehensive system for assessing the risk of genetically modified plants on non

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

    Science.gov (United States)

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

    2016-03-23

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

  10. In-silico determination of insecticidal potential of Vip3Aa-Cry1Ac fusion protein against Lepidopteran targets using molecular docking

    Directory of Open Access Journals (Sweden)

    Aftab eAhmad

    2015-12-01

    Full Text Available Study and research of Bt (Bacillus thuringiensis transgenic plants have opened new ways to combat insect pests. Over the decades, however, insect pests, especially the Lepidopteran, have developed tolerance against Bt delta-endotoxins. Such issues can be addressed through the development of novel toxins with greater toxicity and affinity against a broad range of insect receptors. In this computational study, functional domains of Bacillus thuringiensis crystal delta-endotoxin (Cry1Ac insecticidal protein and vegetative insecticidal protein (Vip3Aa have been fused to develop a broad-range Vip3Aa-Cry1Ac fusion protein. Cry1Ac and Vip3Aa are non-homologous insecticidal proteins possessing receptors against different targets within the midgut of insects. The insecticidal proteins were fused to broaden the insecticidal activity. Molecular docking analysis of the fusion protein against aminopeptidase-N (APN and cadherin receptors of five Lepidopteran insects (Agrotis ipsilon, Helicoverpa armigera, Pectinophora gossypiella, Spodoptera exigua and Spodoptera litura revealed that the Ser290, Ser293, Leu337, Thr340 and Arg437 residues of the fusion protein are involved in the interaction with insect receptors. The Helicoverpa armigera cadherin receptor, however, showed no interaction, which might be due to either loss or burial of interactive residues inside the fusion protein. These findings revealed that the Vip3Aa-Cry1Ac fusion protein has a strong affinity against Lepidopteran insect receptors and hence has a potential to be an efficient broad-range insecticidal protein.

  11. In-Silico Determination of Insecticidal Potential of Vip3Aa-Cry1Ac Fusion Protein Against Lepidopteran Targets Using Molecular Docking.

    Science.gov (United States)

    Ahmad, Aftab; Javed, Muhammad R; Rao, Abdul Q; Khan, Muhammad A U; Ahad, Ammara; Din, Salah Ud; Shahid, Ahmad A; Husnain, Tayyab

    2015-01-01

    Study and research of Bt (Bacillus thuringiensis) transgenic plants have opened new ways to combat insect pests. Over the decades, however, insect pests, especially the Lepidopteran, have developed tolerance against Bt delta-endotoxins. Such issues can be addressed through the development of novel toxins with greater toxicity and affinity against a broad range of insect receptors. In this computational study, functional domains of Bacillus thuringiensis crystal delta-endotoxin (Cry1Ac) insecticidal protein and vegetative insecticidal protein (Vip3Aa) have been fused to develop a broad-range Vip3Aa-Cry1Ac fusion protein. Cry1Ac and Vip3Aa are non-homologous insecticidal proteins possessing receptors against different targets within the midgut of insects. The insecticidal proteins were fused to broaden the insecticidal activity. Molecular docking analysis of the fusion protein against aminopeptidase-N (APN) and cadherin receptors of five Lepidopteran insects (Agrotis ipsilon, Helicoverpa armigera, Pectinophora gossypiella, Spodoptera exigua, and Spodoptera litura) revealed that the Ser290, Ser293, Leu337, Thr340, and Arg437 residues of the fusion protein are involved in the interaction with insect receptors. The Helicoverpa armigera cadherin receptor, however, showed no interaction, which might be due to either loss or burial of interactive residues inside the fusion protein. These findings revealed that the Vip3Aa-Cry1Ac fusion protein has a strong affinity against Lepidopteran insect receptors and hence has a potential to be an efficient broad-range insecticidal protein.

  12. Recombinant Gluconacetobacter diazotrophicus containing Cry1Ac gene codes for 130-kDa toxin protein.

    Science.gov (United States)

    Subashini, M; Moushumi Priya, A; Sundarakrishnan, B; Jayachandran, S

    2011-01-01

    Recombinant Gluconacetobacter diazotrophicus containing Cry1Ac gene from Bacillus thuringiensis var. kurstaki borne on pKT230, shuttle vector, was generated. PCR amplification of Cry1Ac gene present in recombinant G. diazotrophicus yielded a 278-bp DNA product. The nitrogenase assay has revealed that the recombinant G. diazotrophicus in sugarcane stem produced similar levels of nitrogenase compared to wild-type G. diazotrophicus. The presence of 130-kDa protein in apoplastic fluid from sugarcane stem harvested from pots inoculated with recombinant G. diazotrophicus shows that the translocated G. diazotrophicus produces 130-kDa protein which is recognized by the hyperimmune antiserum raised against 130-kDa protein. The first instar Eldana saccharina neonate larvae that fed on artificial medium containing recombinant G. diazotrophicus died within 72 h after incubation.

  13. Effects of Soil Salinity on the Expression of Bt Toxin (Cry1Ac) and the Control Efficiency of Helicoverpa armigera in Field-Grown Transgenic Bt Cotton

    Science.gov (United States)

    Luo, Jun-Yu; Zhang, Shuai; Peng, Jun; Zhu, Xiang-Zhen; Lv, Li-Min; Wang, Chun-Yi; Li, Chun-Hua; Zhou, Zhi-Guo; Cui, Jin-Jie

    2017-01-01

    An increasing area of transgenic Bacillus thuringiensis (Bt) cotton is being planted in saline-alkaline soil in China. The Bt protein level in transgenic cotton plants and its control efficiency can be affected by abiotic stress, including high temperature, water deficiency and other factors. However, how soil salinity affects the expression of Bt protein, thus influencing the control efficiency of Bt cotton against the cotton bollworm (CBW) Helicoverpa armigera (Hübner) in the field, is poorly understood. Our objective in the present study was to investigate the effects of soil salinity on the expression of Bt toxin (Cry1Ac) and the control efficiency of Helicoverpa armigera in field-grown transgenic Bt cotton using three natural saline levels (1.15 dS m-1 [low soil-salinity], 6.00 dS m-1 [medium soil-salinity] and 11.46 dS m-1 [high soil-salinity]). We found that the Bt protein content in the transgenic Bt cotton leaves and the insecticidal activity of Bt cotton against CBW decreased with the increasing soil salinity in laboratory experiments during the growing season. The Bt protein content of Bt cotton leaves in the laboratory were negatively correlated with the salinity level. The CBW populations were highest on the Bt cotton grown in medium-salinity soil instead of the high-salinity soil in field conditions. A possible mechanism may be that the relatively high-salinity soil changed the plant nutritional quality or other plant defensive traits. The results from this study may help to identify more appropriate practices to control CBW in Bt cotton fields with different soil salinity levels. PMID:28099508

  14. Detrimental effect of expression ofBt endotoxin Cry1Ac on in vitro regeneration, in vivo growth and development of tobacco and cotton transgenics

    Indian Academy of Sciences (India)

    Preeti Rawat; Amarjeet Kumar Singh; Krishna Ray; Bhupendra Chaudhary; Sanjeev Kumar; Taru Gautam; Shaveta Kanoria; Gurpreet Kaur; Paritosh Kumar; Deepak Pental; Pradeep Kumar Burma

    2011-06-01

    High levels of expression of the cry1Ac gene from Bacillus thuringiensis cannot be routinely achieved in transgenic plants despite modifications made in the gene to improve its expression. This has been attributed to the instability of the transcript in a few reports. In the present study, based on the genetic transformation of cotton and tobacco, we show that the expression of the Cry1Ac endotoxin has detrimental effects on both the in vitro and in vivo growth and development of transgenic plants. A number of experiments on developing transgenics in cotton with different versions of cry1Ac gene showed that the majority of the plants did not express any Cry1Ac protein. Based on Southern blot analysis, it was also observed that a substantial number of lines did not contain the cry1Ac gene cassette although they contained the marker gene nptII. More significantly, all the lines that showed appreciable levels of expression were found to be phenotypically abnormal. Experiments on transformation of tobacco with different constructs expressing the cry1Ac gene showed that in vitro regeneration was inhibited by the encoded protein. Further, out of a total of 145 independent events generated with the different cry1Ac gene constructs in tobacco, only 21 showed expression of the Cry1Ac protein, confirming observations made in cotton that regenerants that express high levels of the Cry1Ac protein are selected against during regeneration of transformed events. This problem was circumvented by targeting the Cry1Ac protein to the chloroplast, which also significantly improved the expression of the protein.

  15. Detrimental effect of expression of Bt endotoxin Cry1Ac on in vitro regeneration, in vivo growth and development of tobacco and cotton transgenics.

    Science.gov (United States)

    Rawat, Preeti; Singh, Amarjeet Kumar; Ray, Krishna; Chaudhary, Bhupendra; Kumar, Sanjeev; Gautam, Taru; Kanoria, Shaveta; Kaur, Gurpreet; Kumar, Paritosh; Pental, Deepak; Burma, Pradeep Kumar

    2011-06-01

    High levels of expression of the cry1Ac gene from Bacillus thuringiensis cannot be routinely achieved in transgenic plants despite modifications made in the gene to improve its expression. This has been attributed to the instability of the transcript in a few reports. In the present study, based on the genetic transformation of cotton and tobacco, we show that the expression of the Cry1Ac endotoxin has detrimental effects on both the in vitro and in vivo growth and development of transgenic plants. A number of experiments on developing transgenics in cotton with different versions of cry1Ac gene showed that the majority of the plants did not express any Cry1Ac protein. Based on Southern blot analysis, it was also observed that a substantial number of lines did not contain the cry1Ac gene cassette although they contained the marker gene nptII. More significantly, all the lines that showed appreciable levels of expression were found to be phenotypically abnormal. Experiments on transformation of tobacco with different constructs expressing the cry1Ac gene showed that in vitro regeneration was inhibited by the encoded protein. Further, out of a total of 145 independent events generated with the different cry1Ac gene constructs in tobacco, only 21 showed expression of the Cry1Ac protein, confirming observations made in cotton that regenerants that express high levels of the Cry1Ac protein are selected against during regeneration of transformed events. This problem was circumvented by targeting the Cry1Ac protein to the chloroplast, which also significantly improved the expression of the protein.

  16. Using resistant prey demonstrates that Bt plants producing Cry1Ac, Cry2Ab, and Cry1F have no negative effects on Geocoris punctipes and Orius insidiosus.

    Science.gov (United States)

    Tian, Jun-Ce; Long, Li-Ping; Wang, Xiang-Ping; Naranjo, Steven E; Romeis, Jörg; Hellmich, Richard L; Wang, Ping; Shelton, Anthony M

    2014-02-01

    Geocoris punctipes (Say) and Orius insidiosus (Say) are generalist predators found in a wide range of crops, including cotton (Gossypium hirsutum L.) and maize (Zea mays L.), where they provide important biological control services by feeding on an array of pests, including eggs and small larvae of caterpillars. A high percentage of cotton and maize in the United States and several other countries are transgenic cultivars that produce one or more of the insecticidal Cry proteins of Bacillus thuringiensis Berliner (Bt). Here we quantify effects of three Cry proteins on the life history of these predators over two generations when they are exposed to these Cry proteins indirectly through their prey. To eliminate the confounding prey quality effects that can be introduced by Bt-susceptible prey, we used Cry1Ac/Cry2Ab-resistant Trichoplusia ni (Hübner) and Cry1 F-resistant Spodoptera frugiperda (J.E. Smith) in a series of tri-trophic studies. Survival, development, adult mass, fecundity, and fertility were similar when predators consumed larvae feeding on Cry1Ac/Cry2Ab cotton or Cry1 F maize compared with prey feeding on isogenic or near-isogenic cotton or maize. Repeated exposure of the same initial cohort over a second generation also resulted in no differences in life-history traits when feeding on non-Bt- or Bt-fed prey. Enzyme-linked immunosorbent assay showed that predators were exposed to Bt Cry proteins from their prey and that these proteins became increasingly diluted as they moved up the food chain. Results show a clear lack of effect of three common and widespread Cry proteins on these two important predator species. The use of resistant insects to eliminate prey quality effects provides a robust and meaningful assessment of exposure and hazard.

  17. High Sensitivity Bacillus thuringiensis Cry1Ac Protein Detections Using Fluorescein Diacetate Nanoparticles.

    Science.gov (United States)

    Liu, Cui; Zhou, Zhen; Zou, Linling; Cao, Yuan-Cheng; Liu, Jun'An; Lin, Yongjun

    2016-03-01

    A highly sensitive transgenic protein analysis method was proposed here based on fluorescein diacetate (FDA). First, FDA was prepared by the ball mill to harvest the nano-sized organic particles. Further examines showed that the FDA size can be controlled by the speed of centrifugation which can obtain FDA in well-distributed size. Cy3 antibody immobilization tests showed that the proteins can attach onto the FDA particles while keep bioactivities. FDA and Cry1Ac antibody immunoassay tests showed that when the FDA particle was in 150 nm, the linear range was 0.01 ng/L-30 μg/mL. And it has the lower detection limitation of 0.01 ng/L, which is 100 times more sensitive than the ELISA methods. These results indicate that the FDA related immunoassays are the promising approach in the transgenic analysis.

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

    Science.gov (United States)

    2010-07-01

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

  19. Degradation of Cry1Ac protein from Bacillus thuringiensis by soil bacteria from transgenic and conventional cotton (Gossypium hirsutum) culture.

    OpenAIRE

    João Paulo Leite Tozzi

    2009-01-01

    Bt é uma bactéria formadora da proteína Cry1Ac, tóxica a lepidópteros. Plantas geneticamente modificadas expressam essa toxina. O objetivo deste trabalho foi isolamento e identificação de bactérias do solo de algodão transgênico e convencional potencialmente biodegradadoras dessa proteína. Estudou-se a dinâmica de crescimento das bactérias em meios com a proteína Cry1Ac ou glucose, a biodegradação, os genes apr, npr e sub. Em solo de algodão convencional a contagem foi menor; para algodão tra...

  20. Preliminary comparing the toxicities of the hybrid cry1Acs fused with different heterogenous genes provided guidance for the fusion expression of Cry proteins.

    Science.gov (United States)

    Tang, Ying; Tong, Jinying; Zhang, Yunlei; Wang, Lei; Hu, Shengbiao; Li, Wenping; Lv, Yuan

    2012-01-01

    In order to provide guidance for selecting suitable heterogenous gene that can efficiently enhance toxicity or broaden insecticidal spectrum of Cry1Ac through fusion expression, two hybrid cry1Acs fused with chitinase-encoding gene tchiB and neurotoxin gene hwtx-1 respectively were constructed and their toxicities were compared. A Bacillus thuringiensis strain harboring the cry1Ac gene in vector pHT315 was used as control. Bioassay revealed that LC(50) (after 72 h) of Cry1Ac protoxin was 41.01 μg mL(-1), while the hybrid cry1Acs fused with tchiB and hwtx-1 were 4.89 and 23.14 μg mL(-1), which were 8.23- and 1.77-fold higher than Cry1Ac protoxin in terms of relative toxicity respectively. Both fusion crystals had a higher toxicity than the original Cry1Ac protein and the toxicity of hybrid cry1Acs fused with hwtx-1 experienced a more significant increase than that fused with tchiB.

  1. Uniform Orientation of Biotinylated Nanobody as an Affinity Binder for Detection of Bacillus thuringiensis (Bt) Cry1Ac Toxin

    Science.gov (United States)

    Li, Min; Zhu, Min; Zhang, Cunzheng; Liu, Xianjin; Wan, Yakun

    2014-01-01

    Nanobodies are the smallest natural fragments with useful properties such as high affinity, distinct paratope and high stability, which make them an ideal tool for detecting target antigens. In this study, we generated and characterized nanobodies against the Cry1Ac toxin and applied them in a biotin-streptavidin based double antibodies (nanobodies) sandwich-ELISA (DAS-ELISA) assay. After immunizing a camel with soluble Cry1Ac toxin, a phage displayed library was constructed to generate Nbs against the Cry1Ac toxin. Through successive rounds of affinity bio-panning, four nanobodies with greatest diversity in CDR3 sequences were obtained. After affinity determination and conjugating to HRP, two nanobodies with high affinity which can recognize different epitopes of the same antigen (Cry1Ac) were selected as capture antibody (Nb61) and detection antibody (Nb44). The capture antibody (Nb61) was biotinylated in vivo for directional immobilization on wells coated with streptavidin matrix. Both results of specificity analysis and thermal stability determination add support for reliability of the following DAS-ELISA with a minimum detection limit of 0.005 μg·mL−1 and a working range 0.010–1.0 μg·mL−1. The linear curve displayed an acceptable correlation coefficient of 0.9976. These results indicated promising applications of nanobodies for detection of Cry1Ac toxin with biotin-streptavidin based DAS-ELISA system. PMID:25474492

  2. Uniform Orientation of Biotinylated Nanobody as an Affinity Binder for Detection of Bacillus thuringiensis (Bt Cry1Ac Toxin

    Directory of Open Access Journals (Sweden)

    Min Li

    2014-12-01

    Full Text Available Nanobodies are the smallest natural fragments with useful properties such as high affinity, distinct paratope and high stability, which make them an ideal tool for detecting target antigens. In this study, we generated and characterized nanobodies against the Cry1Ac toxin and applied them in a biotin-streptavidin based double antibodies (nanobodies sandwich-ELISA (DAS-ELISA assay. After immunizing a camel with soluble Cry1Ac toxin, a phage displayed library was constructed to generate Nbs against the Cry1Ac toxin. Through successive rounds of affinity bio-panning, four nanobodies with greatest diversity in CDR3 sequences were obtained. After affinity determination and conjugating to HRP, two nanobodies with high affinity which can recognize different epitopes of the same antigen (Cry1Ac were selected as capture antibody (Nb61 and detection antibody (Nb44. The capture antibody (Nb61 was biotinylated in vivo for directional immobilization on wells coated with streptavidin matrix. Both results of specificity analysis and thermal stability determination add support for reliability of the following DAS-ELISA with a minimum detection limit of 0.005 μg·mL−1 and a working range 0.010–1.0 μg·mL−1. The linear curve displayed an acceptable correlation coefficient of 0.9976. These results indicated promising applications of nanobodies for detection of Cry1Ac toxin with biotin-streptavidin based DAS-ELISA system.

  3. Feed intake, milk production and composition of crossbred cows fed with insect-protected Bollgard II® cottonseed containing Cry1Ac and Cry2Ab proteins.

    Science.gov (United States)

    Singhal, K K; Tyagi, A K; Rajput, Y S; Singh, M; Kaur, H; Perez, T; Hartnell, G F

    2011-09-01

    Twenty crossbred lactating multiparous cows were used in a 28-day study to compare dry matter intake (DMI), milk yield, milk composition and Bacillus thuringiensis (Bt) protein concentrations in plasma when fed diets containing Bollgard II(®) cottonseed (BGII) or a control non-genetically modified isogenic cottonseed (CON). Bollgard II cottonseed contains the Cry1Ac and Cry2Ab insecticidal proteins that protect cotton plants from feeding damage caused by certain lepidopteran insects. Cows were assigned randomly to the BGII or CON treatments after a 2-week adjustment period. Cows consumed a concentrate containing 40% crushed cottonseed according to milk yield and green maize forage ad libitum. All cows received the same diet but with different crushed cottonseed sources. Cottonseed was included to provide approximately 2.9 kg per cow daily (dry matter basis). The ingredient composition of the concentrate was 40% crushed cottonseed, 15% groundnut cake, 20% corn, 22% wheat bran, 1% salt and 2% mineral mixture. Milk and blood plasma were analyzed for Cry1Ac and Cry2Ab proteins. DMI, BW, milk yield and milk components did not differ between cows on the BGII and CON treatments. Although milk yield and milk fat percentage were not affected by treatment, 4% fat-corrected milk (FCM) production and FCM/kg DMI for cows on the BGII treatment (14.0 kg/cow per day, 1.12 kg/kg) were significantly improved compared with cows on the CON treatment (12.1 kg/cow per day, 0.97 kg/kg). Gossypol contents in BGII cottonseed and conventional cottonseed were similar. Cry1Ac and Cry2Ab2 proteins in Bollgard II cottonseed were 5.53 and 150.8 μg/g, respectively, and were not detected in the milk or plasma samples. The findings suggested that Bollgard II cottonseed can replace conventional cottonseed in dairy cattle diets with no adverse effects on performance and milk composition.

  4. Characterization of the mucosal and systemic immune response induced by Cry1Ac protein from Bacillus thuringiensis HD 73 in mice

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    Vázquez-Padrón R.I.

    2000-01-01

    Full Text Available The present paper describes important features of the immune response induced by the Cry1Ac protein from Bacillus thuringiensis in mice. The kinetics of induction of serum and mucosal antibodies showed an immediate production of anti-Cry1Ac IgM and IgG antibodies in serum after the first immunization with the protoxin by either the intraperitoneal or intragastric route. The antibody fraction in serum and intestinal fluids consisted mainly of IgG1. In addition, plasma cells producing anti-Cry1Ac IgG antibodies in Peyer's patches were observed using the solid-phase enzyme-linked immunospot (ELISPOT. Cry1Ac toxin administration induced a strong immune response in serum but in the small intestinal fluids only anti-Cry1Ac IgA antibodies were detected. The data obtained in the present study confirm that the Cry1Ac protoxin is a potent immunogen able to induce a specific immune response in the mucosal tissue, which has not been observed in response to most other proteins.

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

    Science.gov (United States)

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

    2008-01-01

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

  6. Analysis of resistance to Cry1Ac in field-collected pink bollworm, Pectinophora gossypiella (Lepidoptera:Gelechiidae), populations.

    Science.gov (United States)

    Ojha, Abhishek; Sree, K Sowjanya; Sachdev, Bindiya; Rashmi, M A; Ravi, K C; Suresh, P J; Mohan, Komarlingam S; Bhatnagar, Raj K

    2014-01-01

    High survivorship of pink bollworrm, Pectinophora gossypiella in bolls of Bollgard® cotton hybrids and resistance to Cry1Ac protein, expressed in Bollgard cotton were reported in field-populations collected from the state of Gujarat (western India) in 2010. We have found Cry1Ac-resistance in pink bollworm populations sourced from Bollgard and non-Bt cotton fields in the adjoining states of Maharashtra and Madhya Pradesh in Central India. Further, we observed reduced binding of labeled Cry1Ac protein to receptors localized on the brush-border membrane of pink bollworm larval strains with high tolerance to Cry1Ac. These strains were sourced from Bollgard and conventional cotton fields. A pooled Cry1Ac-resistant strain, further selected on Cry1Ac diet also showed significantly reduced binding to Cry1Ac protein. The reduced binding of Cry1Ac to receptors could be an underlying mechanism for the observed resistance in pink bollworm populations feeding on Bollgard hybrids.

  7. Identification of Cry1Ac and Cry2Ab proteins in transgenic cotton seeds available in Gujarat (India by ELISA method

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    Alka Dohare

    2014-02-01

    Full Text Available Along with the increase market of the transgenic crops, the demand for testing GMOs and for certifying non-GMO foodstuffs has increased dramatically. Within the arena of expanding techniques for identification and quantification of transgenic crops, two major approaches for detecting GMOs are still applicable on large scale, namely ELISA based protein detection and PCR based gene identification. In present study, ELISA techniques was adopted to identify the specific Cry1Ac and Cry2Ab proteins in some transgenic cotton plants seed samples viz., Gujarat cotton hybrid – 6 (BG II, Gujarat cotton hybrid – 8 (BG II and Gujarat cotton hybrid – 10 (BG II from the Gujarat state of India. The study reveals the presence of both Cry1Ac and Cry2Ab proteins in the transgenic seed samples and also demonstrated that the technique of ELISA for identification of Cry1Ac and Cry2Ab proteins is quite handy and easily adoptable.

  8. The interaction of two-spotted spider mites, Tetranychus urticae Koch, with Cry protein production and predation by Amblyseius andersoni (Chant) in Cry1Ac/Cry2Ab cotton and Cry1F maize.

    Science.gov (United States)

    Guo, Yan-Yan; Tian, Jun-Ce; Shi, Wang-Peng; Dong, Xue-Hui; Romeis, Jörg; Naranjo, Steven E; Hellmich, Richard L; Shelton, Anthony M

    2016-02-01

    Crops producing insecticidal crystal (Cry) proteins from the bacterium, Bacillus thuringiensis (Bt), are an important tool for managing lepidopteran pests on cotton and maize. However, the effects of these Bt crops on non-target organisms, especially natural enemies that provide biological control services, are required to be addressed in an environmental risk assessment. Amblyseius andersoni (Acari: Phytoseiidae) is a cosmopolitan predator of the two-spotted spider mite, Tetranychus urticae (Acari: Tetranychidae), a significant pest of cotton and maize. Tri-trophic studies were conducted to assess the potential effects of Cry1Ac/Cry2Ab cotton and Cry1F maize on life history parameters (survival rate, development time, fecundity and egg hatching rate) of A. andersoni. We confirmed that these Bt crops have no effects on the biology of T. urticae and, in turn, that there were no differences in any of the life history parameters of A. andersoni when it fed on T. urticae feeding on Cry1Ac/Cry2Ab or non-Bt cotton and Cry1F or non-Bt maize. Use of a susceptible insect assay demonstrated that T. urticae contained biologically active Cry proteins. Cry proteins concentrations declined greatly as they moved from plants to herbivores to predators and protein concentration did not appear to be related to mite density. Free-choice experiments revealed that A. andersoni had no preference for Cry1Ac/Cry2Ab cotton or Cry1F maize-reared T. urticae compared with those reared on non-Bt cotton or maize. Collectively these results provide strong evidence that these crops can complement other integrated pest management tactics including biological control.

  9. Toxicity and binding analyses of Bacillus thuringiensis toxin Vip3A in Cry1Ac-resistant and-susceptible strains of Helicoverpa armigera (Hübner)

    Institute of Scientific and Technical Information of China (English)

    ZHANG Qian; CHEN Li-zhen; LU Qiong; ZHANG Yan; LIANG Ge-mei

    2015-01-01

    The Bacil us thuringiensis vegetative insecticidal protein, Vip3A, represents a new family of Bt toxin and is currently ap-plied to commercial transgenic cotton. To determine whether the Cry1Ac-resistant Helicoverpa armigera is cross-resistant to Vip3Aa protein, insecticidal activities, proteolytic activations and binding properties of Vip3Aa toxin were investigated using Cry1Ac-susceptible (96S) and Cry1Ac-resistant H. armigera strain (Cry1Ac-R). The toxicity of Vip3Aa in Cry1Ac-R slightly reduced compared with 96S, the resistance ratio was only 1.7-fold. The digestion rate of ful-length Vip3Aa by gut juice extracts from 96S was little faster than that from Cry1Ac-R. Surface plasmon resonance (SPR) showed there was no signiifcant difference between the binding afifnity of Vip3Aa and BBMVs between 96S and Cry1Ac-R strains, and there was no signiifcant competitive binding between Vip3Aa and Cry1Ac in susceptible or resistant strains. So there had little cross-resistance between Vip3Aa and Cry1Ac,Vip3A+Cry proteins maybe the suitable pyramid strategy to control H. armigera in China in the future.

  10. Cry1Ac and Vip3Aa proteins from Bacillus thuringiensis targeting Cry toxin resistance in Diatraea flavipennella and Elasmopalpus lignosellus from sugarcane

    Science.gov (United States)

    2017-01-01

    The biological potential of Vip and Cry proteins from Bacillus is well known and widely established. Thus, it is important to look for new genes showing different modes of action, selecting those with differentiated entomotoxic activity against Diatraea flavipennella and Elasmopalpus lignosellus, which are secondary pests of sugarcane. Therefore, Cry1 and Vip3 proteins were expressed in Escherichia coli, and their toxicities were evaluated based on bioassays using neonate larvae. Of those, the most toxic were Cry1Ac and Vip3Aa considering the LC50 values. Toxins from E. coli were purified, solubilized, trypsinized, and biotinylated. Brush Border Membrane Vesicles (BBMVs) were prepared from intestines of the two species to perform homologous and heterologous competition assays. The binding assays demonstrated interactions between Cry1Aa, Cry1Ac, and Vip3Aa toxins and proteins from the BBMV of D. flavipennella and E. lignosellus. Homologous competition assays demonstrated that binding to one of the BBMV proteins was specific for each toxin. Heterologous competition assays indicated that Vip3Aa was unable to compete for Cry1Ac toxin binding. Our results suggest that Cry1Ac and Vip3Aa may have potential in future production of transgenic sugarcane for control of D. flavipennella and E. lignosellus, but more research is needed on the potential antagonism or synergism of the toxins in these pests. PMID:28123906

  11. Cry1Ac and Vip3Aa proteins from Bacillus thuringiensis targeting Cry toxin resistance in Diatraea flavipennella and Elasmopalpus lignosellus from sugarcane

    Directory of Open Access Journals (Sweden)

    Ana Rita Nunes Lemes

    2017-01-01

    Full Text Available The biological potential of Vip and Cry proteins from Bacillus is well known and widely established. Thus, it is important to look for new genes showing different modes of action, selecting those with differentiated entomotoxic activity against Diatraea flavipennella and Elasmopalpus lignosellus, which are secondary pests of sugarcane. Therefore, Cry1 and Vip3 proteins were expressed in Escherichia coli, and their toxicities were evaluated based on bioassays using neonate larvae. Of those, the most toxic were Cry1Ac and Vip3Aa considering the LC50 values. Toxins from E. coli were purified, solubilized, trypsinized, and biotinylated. Brush Border Membrane Vesicles (BBMVs were prepared from intestines of the two species to perform homologous and heterologous competition assays. The binding assays demonstrated interactions between Cry1Aa, Cry1Ac, and Vip3Aa toxins and proteins from the BBMV of D. flavipennella and E. lignosellus. Homologous competition assays demonstrated that binding to one of the BBMV proteins was specific for each toxin. Heterologous competition assays indicated that Vip3Aa was unable to compete for Cry1Ac toxin binding. Our results suggest that Cry1Ac and Vip3Aa may have potential in future production of transgenic sugarcane for control of D. flavipennella and E. lignosellus, but more research is needed on the potential antagonism or synergism of the toxins in these pests.

  12. Cry1Ac and Vip3Aa proteins from Bacillus thuringiensis targeting Cry toxin resistance in Diatraea flavipennella and Elasmopalpus lignosellus from sugarcane.

    Science.gov (United States)

    Lemes, Ana Rita Nunes; Figueiredo, Camila Soares; Sebastião, Isis; Marques da Silva, Liliane; da Costa Alves, Rebeka; de Siqueira, Herbert Álvaro Abreu; Lemos, Manoel Victor Franco; Fernandes, Odair Aparecido; Desidério, Janete Apparecida

    2017-01-01

    The biological potential of Vip and Cry proteins from Bacillus is well known and widely established. Thus, it is important to look for new genes showing different modes of action, selecting those with differentiated entomotoxic activity against Diatraea flavipennella and Elasmopalpus lignosellus, which are secondary pests of sugarcane. Therefore, Cry1 and Vip3 proteins were expressed in Escherichia coli, and their toxicities were evaluated based on bioassays using neonate larvae. Of those, the most toxic were Cry1Ac and Vip3Aa considering the LC50 values. Toxins from E. coli were purified, solubilized, trypsinized, and biotinylated. Brush Border Membrane Vesicles (BBMVs) were prepared from intestines of the two species to perform homologous and heterologous competition assays. The binding assays demonstrated interactions between Cry1Aa, Cry1Ac, and Vip3Aa toxins and proteins from the BBMV of D. flavipennella and E. lignosellus. Homologous competition assays demonstrated that binding to one of the BBMV proteins was specific for each toxin. Heterologous competition assays indicated that Vip3Aa was unable to compete for Cry1Ac toxin binding. Our results suggest that Cry1Ac and Vip3Aa may have potential in future production of transgenic sugarcane for control of D. flavipennella and E. lignosellus, but more research is needed on the potential antagonism or synergism of the toxins in these pests.

  13. Development of dipsticks for simultaneous detection of vip3A and cry1Ab/cry1Ac transgenic proteins.

    Science.gov (United States)

    Kumar, Rajesh

    2012-01-01

    The number of genetically modified (GM) crops being cultivated and its produce reaching market is increasing every year. The transgenes (vip3A, cry1Ab, and cry1Ac) from Bacillus thuringiensis are being used by plant breeders, apart from other transgenes for developing insect pest-resistant GM crops. It is therefore necessary to develop an easy, rapid, and reliable detection assay to discriminate GM crops and non-GM crops. Dipstick strips using colloidal gold-labeled polyclonal antibodies were developed for simultaneous detection of Vip3A and Cry1Ab/CrylAc proteins. The assay was essentially based on the sandwich format of immunoassay, which was completed within 10 min, and the results were evaluated visually. The detection limits were 50 ng/mL (50 ppb) for both CrylAc and CrylAb proteins, and 100 ng/mL (100 ppb) for Vip3A protein. The developed dipsticks are suitable for on-site simultaneous screening of GM crops bearing two proteins, which, in turn, reduce cost and time of the assay.

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

    Science.gov (United States)

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

    2013-01-01

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

  15. High Expression of Cry1Ac Protein in Cotton (Gossypium hirsutum by Combining Independent Transgenic Events that Target the Protein to Cytoplasm and Plastids.

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    Amarjeet Kumar Singh

    Full Text Available Transgenic cotton was developed using two constructs containing a truncated and codon-modified cry1Ac gene (1,848 bp, which was originally characterized from Bacillus thuringiensis subspecies kurstaki strain HD73 that encodes a toxin highly effective against many lepidopteran pests. In Construct I, the cry1Ac gene was cloned under FMVde, a strong constitutively expressing promoter, to express the encoded protein in the cytoplasm. In Construct II, the encoded protein was directed to the plastids using a transit peptide taken from the cotton rbcSIb gene. Genetic transformation experiments with Construct I resulted in a single copy insertion event in which the Cry1Ac protein expression level was 2-2.5 times greater than in the Bacillus thuringiensis cotton event Mon 531, which is currently used in varieties and hybrids grown extensively in India and elsewhere. Another high expression event was selected from transgenics developed with Construct II. The Cry protein expression resulting from this event was observed only in the green plant parts. No transgenic protein expression was observed in the non-green parts, including roots, seeds and non-green floral tissues. Thus, leucoplasts may lack the mechanism to allow entry of a protein tagged with the transit peptide from a protein that is only synthesized in tissues containing mature plastids. Combining the two events through sexual crossing led to near additive levels of the toxin at 4-5 times the level currently used in the field. The two high expression events and their combination will allow for effective resistance management against lepidopteran insect pests, particularly Helicoverpa armigera, using a high dosage strategy.

  16. Quantitative analysis of differential proteins between the susceptible and resistant Trichoplusia ni cells to Bt Cry1Ac toxin%抗Bt Cry1Ac毒素粉纹夜蛾离体细胞与敏感细胞的差异蛋白定量分析

    Institute of Scientific and Technical Information of China (English)

    刘凯于; 吴彦艳; 彭建新; 洪华珠

    2011-01-01

    苏云金芽胞杆菌(Bacillus thuringiensis,Bt)产生的毒素对许多昆虫具有特异性高效毒杀作用[1-2],但是随着Bt杀虫毒素的广泛使用,昆虫对Bt的抗性风险越来越高.这种抗性机制非常复杂[3],而蛋白质组学技术可较全面揭示昆虫细胞抗Bt毒素的分子机制.作者等[4]通过人工选择获得了高水平抗Bt Cry1Ac毒素的粉纹夜蛾细胞系,在此基础上本试验采用双向凝胶电泳技术对BtCry1Ac毒素敏感粉纹夜蛾细胞和抗性选择细胞的差异蛋白质进行了定量分析.

  17. 烟夜蛾幼虫中肠Bt Cry1Ac毒素受体蛋白cDNA片段的克隆和序列测定%Molecular Cloning and Sequencing of cDNA Fragment Encoding Bt Cry1Ac Toxin Binding Protein from the Midgut of Helicoverpa assulta Guenée Larva

    Institute of Scientific and Technical Information of China (English)

    安世恒; 郭线茹; 罗梅浩; 蒋金炜; 马继盛

    2005-01-01

    利用RT-PCR技术扩增烟夜蛾(Helicoverpa assulta Guenée)幼虫中肠Bt毒素Cry1Ac受体蛋白APN(N-氨基肽酶,aminopeptidase N,APN)基因片段,克隆和测序结果表明,测序得到的812 bp的片段编码270个氨基酸残基,且该片段在阅读框内.通过同源性分析发现,其核苷酸序列与棉铃虫(H. armigera)、澳洲棉铃虫(H. punctigera)、烟芽夜蛾(H. virescens)、舞毒蛾(Lymantria dispar)、小菜蛾(Plutella xylostella)、印度谷螟(Plodia interpunctella)RC688品系和HD198品系、烟草天蛾(Manduca sexta)和家蚕(Bombyx mori)的Cry1Ac受体蛋白基因的同源性分别为97.0%,90.0%,78.0%,63.5%,55.0%,60.3%,61.2%,55.0%和59.0%.推导的烟夜蛾Cry1Ac受体蛋白基因的氨基酸序列与棉铃虫、烟芽夜蛾、斑实夜蛾、舞毒蛾的氨基酸序列同源性分别为95.6%,81.0%,82.7%和55.7%.该片段编码的氨基酸属于氨肽酶家族,与烟夜蛾对Bt Cry1Ac毒素的抗性有关.

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

    Directory of Open Access Journals (Sweden)

    Carmen Sara Hernández-Rodríguez

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

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

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    Linda J Gahan

    2010-12-01

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

  20. Comparison and optimization of the method for Cry1Ac protoxin preparation in HD73 strain

    Institute of Scientific and Technical Information of China (English)

    ZHOU Zi-shan; YANG Su-juan; SHU Chang-long; SONG Fu-ping; ZHOU Xue-ping; ZHANG Jie

    2015-01-01

    Bacil us thuringiensis is one of the most widely used bioinsecticides, and cry gene is the major insecticidal gene. Because Cry1Ac protein shows strong toxicity against many lepidopteran species, it has been applied widely in spraying products and transgenic Bt-crops. The preparation of Cry protoxin is the ifrst step in the very important processes of understanding the insecticidal mechanism, resistance screening, and biosafety assessments. The media for crystal production and the method for Cry protoxin preparation were varied, however, it was not clear which was better for preparing a larger amount of Cry protoxin. In this paper, three media for crystal production and the method for Cry1Ac protoxin preparation from HD73 strain were compared to ifnd an efifcacious way to prepare a large number of Cry1Ac protoxin. The results showed that the 1/2 LB (Luria-Bertani) medium was the ideal medium for crystal production, because the total yield of Cry1Ac protoxin in 300 mL 1/2 LB medium was (112.38±5.64) mg, the highest one among three media;the repeated crystal solubilization method was better for the preparation of the Cry protoxin comparing with the continuous crystal solubilization method. It wil be a reference for other Cry protoxin preparation, especial y for larger number.

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

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    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.

  2. MAPK signaling pathway alters expression of midgut ALP and ABCC genes and causes resistance to Bacillus thuringiensis Cry1Ac toxin in diamondback moth.

    Directory of Open Access Journals (Sweden)

    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.

  3. Transgenic tobacco plants expressing synthetic Cry1Ac and Cry1le genes are more toxic to cotton bollworm than those containing one gene

    Institute of Scientific and Technical Information of China (English)

    LIAN Yun; JIA ZhiWei; HE KangLai; LIU YunJun; SONG FuPing; WANG BaoMin; WANG GuoYing

    2008-01-01

    Transgenic tobacco plants carrying Cry1Ac, Crylle or both genes were obtained. In the leaves of transgenic plants carrying both genes, the contents of CrylAc and Cry1le proteins were 0.173% and 0.131% of the total proteins, respectively, Cry1Ac protein content was 0.182 % and Cry1le protein con-tent was 0.124% of the total proteins in the leaves of transgenic plants containing only one Bt gone. Fresh leaves of transgenic tobacco and wild-type plants were used for the insect bioassay against wild-type and Cry1Ac-resistant cotton bollworm (Helicoverpa armigera). The bioassay results showed that transgenic plants carrying both genes were significantly more toxic to wild-type and Cry1Ac-resistant cotton bollworm than those carrying Cry1Ac or Cry1le alone. This study indicates that the higher toxicity of transgenic tobacco plants carrying both genes is caused by the cooperative function of both Bt proteins, thus providing a potential way to delay the development of insect resis-tance to transgenic crops.

  4. Cloning and Expression Analysis of the cry1Ac Gene from Bacillus thuringiensis Strain 4.0718%Bt4.0718 cry1Ac基因的克隆与表达分析

    Institute of Scientific and Technical Information of China (English)

    丁学知; 张何; 孙运军; 黄潢; 张春艳; 夏立秋

    2006-01-01

    在基因库中比对14种cry1Ac基因序列,发现了同源性很高的上游启动子区域和下游终止子区域.根据这一同源序列设计引物,从Bt4.0718中扩增出包含双启动子和终止子的4.2 kb片段,用PCR-RFLP检测确定其中含有cry1Ac基因.然后将此片段克隆到穿梭载体pHT304中,转化大肠杆菌DH5α和Bt无晶体突变株XZM-101.同时,利用原子力显微镜观察发现重组菌株BXZM34能够产生菱形晶体.%14 cry1Ac genes from GenBank were aligned and the consensus regions in the upstream of promoter and in the downstream of terminator were found. Based on the consensus sequences, a pair of primers was designed and a 4.2 kb element was amplified that includes the dual overlapping promoter and the whole termination-associated sequence from Bacillus thuringiensis strain 4. 0718, and the amplified 4.2 kb element was confirmed to contain the crylAc gene by cryI sub-genetype PCR-RFLP cry gene typing system. The 4.2 kb element was cloned into Bt-E. coli shuttle vector pHT30.4 and the cry1Ac gene was also expressed in E. coli DH5α and acrystalliferous mutant XZM-101. Meanwhile,rhombic crystal was observed from recombinant strain BXZM34 by atomic force microscope.

  5. Cadherin is involved in the action of Bacillus thuringiensis toxins Cry1Ac and Cry2Aa in the beet armyworm, Spodoptera exigua.

    Science.gov (United States)

    Qiu, Lin; Hou, Leilei; Zhang, Boyao; Liu, Lang; Li, Bo; Deng, Pan; Ma, Weihua; Wang, Xiaoping; Fabrick, Jeffrey A; Chen, Lizhen; Lei, Chaoliang

    2015-05-01

    Bacillus thuringiensis (Bt) insecticidal crystal (Cry) proteins are effective against some insect pests in sprays and transgenic crops, although the evolution of resistance could threaten the long-term efficacy of such Bt use. One strategy to delay resistance to Bt crops is to "pyramid" two or more Bt proteins that bind to distinct receptor proteins within the insect midgut. The most common Bt pyramid in cotton (Gossypium hirsutum L.) employs Cry1Ac with Cry2Ab to target several key lepidopteran pests, including the beet armyworm, Spodoptera exigua (Hübner), which is a serious migratory pest of many vegetable crops and is increasingly important in cotton in China. While cadherin and aminopeptidase-N are key receptors of Cry1 toxins in many lepidopterans including S. exigua, the receptor for Cry2A toxins remains poorly characterized. Here, we show that a heterologous expressed peptide corresponding to cadherin repeat 7 to the membrane proximal extracellular domain (CR7-MPED) in the S. exigua cadherin 1b (SeCad1b) binds Cry1Ac and Cry2Aa. Moreover, SeCad1b transcription was suppressed in S. exigua larvae by oral RNA interference and susceptibility to Cry1Ac and Cry2Aa was significantly reduced. These results indicate that SeCad1b plays important functional roles of both Cry1Ac and Cry2Aa, having major implications for resistance management for S. exigua in Bt crops.

  6. Transmission of Cry1Ac from transgenic Bt cotton to the predatory ladybug Propylaea japonica via the whitefly Bemisia tabaci%转Bt基因棉花中Cry1Ac蛋白经烟粉虱途径向龟纹瓢虫的传递

    Institute of Scientific and Technical Information of China (English)

    周福才; 顾爱祥; 杨益众; 苏宏华; 胡其靖; 杨爱民

    2015-01-01

    【目的】烟粉虱Bemisia tabaci是转基因棉的非靶标害虫,对棉花生产造成严重影响。本文探讨转Bt基因棉花中Cry1Ac蛋白在棉花-烟粉虱-龟纹瓢食物链中的传递规律,以期为转基因棉的环境安全评价提供科学依据。【方法】在实验室条件下,以常规棉SM3号、33、SY321为对照,分析转Bt基因棉花GK12、XM33B、SGK321叶片、在这些棉花上取食的烟粉虱、以及捕食烟粉虱的瓢虫体内Cry1Ac蛋白含量。同时,将取食转 Bt 基因棉花上的烟粉虱的瓢虫转接到对应的受体亲本棉花上,分析瓢虫体内Cry1Ac蛋白含量变化规律。【结果】在转 Bt基因棉花上取食的烟粉虱成虫和若虫以及它们的蜜露中均能检测到Cry1Ac蛋白,以转Bt基因棉花上的烟粉虱若虫为食料的龟纹瓢虫体Propylaea japonica内也能检测到Cry1Ac蛋白。龟纹瓢虫取食转Bt基因棉花上的烟粉虱若虫1 d后体内即能检测到Cry1Ac蛋白,并且随着取食时间的延长,体内Cry1Ac蛋白的含量逐渐增加,但到第6~8天后Cry1Ac蛋白的含量相对稳定。取食3个不同品种棉花上烟粉虱若虫的龟纹瓢虫体内Cry1Ac蛋白的含量存在明显差异,这种差异与棉花叶片上表达的 Cry1Ac 蛋白量呈正相关。但取食后6 d,在3个品种棉花上取食的龟纹瓢虫体内的Cry1Ac蛋白含量之间没有明显的差异。以转Bt基因棉花上的烟粉虱若虫为食料的龟纹瓢虫转移到对应的常规棉亲本上以后,体内的 Cry1Ac 蛋白的含量迅速下降,但10 d 后仍能检测到微量的 Cry1Ac 蛋白。【结论】转Bt基因棉花中的Cry1Ac蛋白可以通过烟粉虱途径传递到龟纹瓢虫体内,龟纹瓢虫对食料中的 Cry1Ac 蛋白具有富集作用,并且 Cry1Ac 蛋白的富集存在饱和现象,富集饱和量与食料中的 Cry1Ac含量无关;龟纹瓢虫脱离含有Cry1Ac蛋白的食料环境后,体内的Cry1Ac

  7. Resistance of Trichoplusia ni to Bacillus thuringiensis toxin Cry1Ac is independent of alteration of the cadherin-like receptor for Cry toxins.

    Directory of Open Access Journals (Sweden)

    Xin Zhang

    Full Text Available Alteration of binding sites for Bacillus thuringiensis (Bt toxins in insect midgut is the major mechanism of high-level resistance to Bt toxins in insects. The midgut cadherin is known to be a major binding protein for Bt Cry1A toxins and linkage of Bt-resistance to cadherin gene mutations has been identified in lepidopterans. The resistance to Bt toxin Cry1Ac evolved in greenhouse populations of Trichoplusia ni has been identified to be associated with the down-regulation of an aminopeptidase N (APN1 gene by a trans-regulatory mechanism and the resistance gene has been mapped to the locus of an ABC transporter (ABCC2 gene. However, whether cadherin is also involved with Cry1Ac-resistance in T. ni requires to be understood. Here we report that the Cry1Ac-resistance in T. ni is independent of alteration of the cadherin. The T. ni cadherin cDNA was cloned and the cadherin sequence showed characteristic features known to cadherins from Lepidoptera. Various T. ni cadherin gene alleles were identified and genetic linkage analysis of the cadherin alleles with Cry1Ac-resistance showed no association of the cadherin gene with the Cry1Ac-resistance in T. ni. Analysis of cadherin transcripts showed no quantitative difference between the susceptible and Cry1Ac-resistant T. ni larvae. Quantitative proteomic analysis of midgut BBMV proteins by iTRAQ-2D-LC-MS/MS determined that there was no quantitative difference in cadherin content between the susceptible and the resistant larvae and the cadherin only accounted for 0.0014% (mol% of the midgut BBMV proteins, which is 1/300 of APN1 in molar ratio. The cadherin from both the susceptible and resistant larvae showed as a 200-kDa Cry1Ac-binding protein by toxin overlay binding analysis, and nano-LC-MS/MS analysis of the 200-kDa cadherin determined that there is no quantitative difference between the susceptible and resistant larvae. Results from this study indicate that the Cry1Ac-resistance in T. ni is

  8. Resistance of Trichoplusia ni to Bacillus thuringiensis toxin Cry1Ac is independent of alteration of the cadherin-like receptor for Cry toxins.

    Science.gov (United States)

    Zhang, Xin; Tiewsiri, Kasorn; Kain, Wendy; Huang, Lihua; Wang, Ping

    2012-01-01

    Alteration of binding sites for Bacillus thuringiensis (Bt) toxins in insect midgut is the major mechanism of high-level resistance to Bt toxins in insects. The midgut cadherin is known to be a major binding protein for Bt Cry1A toxins and linkage of Bt-resistance to cadherin gene mutations has been identified in lepidopterans. The resistance to Bt toxin Cry1Ac evolved in greenhouse populations of Trichoplusia ni has been identified to be associated with the down-regulation of an aminopeptidase N (APN1) gene by a trans-regulatory mechanism and the resistance gene has been mapped to the locus of an ABC transporter (ABCC2) gene. However, whether cadherin is also involved with Cry1Ac-resistance in T. ni requires to be understood. Here we report that the Cry1Ac-resistance in T. ni is independent of alteration of the cadherin. The T. ni cadherin cDNA was cloned and the cadherin sequence showed characteristic features known to cadherins from Lepidoptera. Various T. ni cadherin gene alleles were identified and genetic linkage analysis of the cadherin alleles with Cry1Ac-resistance showed no association of the cadherin gene with the Cry1Ac-resistance in T. ni. Analysis of cadherin transcripts showed no quantitative difference between the susceptible and Cry1Ac-resistant T. ni larvae. Quantitative proteomic analysis of midgut BBMV proteins by iTRAQ-2D-LC-MS/MS determined that there was no quantitative difference in cadherin content between the susceptible and the resistant larvae and the cadherin only accounted for 0.0014% (mol%) of the midgut BBMV proteins, which is 1/300 of APN1 in molar ratio. The cadherin from both the susceptible and resistant larvae showed as a 200-kDa Cry1Ac-binding protein by toxin overlay binding analysis, and nano-LC-MS/MS analysis of the 200-kDa cadherin determined that there is no quantitative difference between the susceptible and resistant larvae. Results from this study indicate that the Cry1Ac-resistance in T. ni is independent of cadherin

  9. Climate change and genetically modified insecticidal plants. Plant-herbivore interactions and secondary chemistry of Bt Cry1Ac-toxin producing oilseed rape (Brassica napus L.) under elevated CO{sub 2} or O{sub 3}

    Energy Technology Data Exchange (ETDEWEB)

    Himanen, S.

    2008-07-01

    Transgenic insect-resistant plants producing Bacillus thuringiensis (Bt) crystalline endotoxins are the first commercial applications of genetically modified crops and their use has steadily expanded over the last ten years. Together with the expanding agricultural use of transgenic crops, climate change is predicted to be among the major factors affecting agriculture in the coming years. Plants, herbivores and insects of higher trophic levels are all predicted to be affected by the current atmospheric climate change. However, only very few studies to date have addressed the sustained use and herbivore interactions of Bt-producing plants under the influence of these abiotic factors. The main objective of this study was to comparatively assess the performance of a Bt Cry1Ac toxin-producing oilseed rape line and its non-transgenic parent line in terms of vegetative growth and allocation to secondary defence compounds (glucosinolates and volatile terpenoids), and the performance of Bt-target and nontarget insect herbivores as well as tritrophic interaction functioning on these lines. For this, several growth chamber experiments with vegetative stage non-Bt and Bt plants facing exposures to doubled atmospheric CO{sub 2} level alone or together with increased temperature and different regimes of elevated O{sub 3} were conducted. The main hypothesis of this work was that Bt-transgenic plants have reduced performance or allocation to secondary compounds due to the cost of producing Bt toxin under changed abiotic environments. The Bt-transgenic oilseed rape line exhibited slightly delayed vegetative growth and had increased nitrogen and reduced carbon content compared to the non-transgenic parent line, but the physiological responses (i.e. biomass gain and photosynthesis) of the plant lines to CO{sub 2} and O{sub 3} enhancements were equal. Two aphid species, non-susceptible to Bt Cry1Ac, showed equal performance and reproduction on both plant lines under elevated CO{sub 2

  10. Altered binding of the Cry1Ac toxin to larval membranes but not to the toxin-binding protein in Plodia interpunctella selected for resistance to different Bacillus thuringiensis isolates.

    Science.gov (United States)

    Mohammed, S I; Johnson, D E; Aronson, A I

    1996-11-01

    Immunoblotting and cytochemical procedures were used to determine whether toxin binding was altered in strains of the Indianmeal moth, Plodia interpunctella, selected for resistance to various strains of Bacillus thuringiensis. Each of these B. thuringiensis subspecies produces a mixture of protoxins, primarily Cry1 types, and the greatest insect resistance is to the Cry1A protoxins. In several cases, however, there was also resistance to toxins not present in the B. thuringiensis strains used for selection. The Cry1Ab and Cry1Ac toxins bound equally well over a range of toxin concentrations and times of incubation to a single protein of ca. 80-kDa in immunoblots of larval membrane extracts from all of the colonies. This binding protein is essential for toxicity since a mutant Cry1Ac toxin known to be defective in binding and thus less toxic bound poorly to the 80-kDa protein. This binding protein differed in size from the major aminopeptidase N antigens implicated in toxin binding in other insects. Binding of fluorescently labeled Cry1Ac or Cry1Ab toxin to larval sections was found at the tips of the brush border membrane prepared from the susceptible but not from any of the resistant P. interpunctella. Accessibility of a major Cry1A-binding protein appears to be altered in resistant larvae and could account for their broad resistance to several B. thuringiensis toxins.

  11. Cry1Ac蛋白降解菌株FJSB3的分离鉴定及降解特性%ISOLATION AND IDENTIFICATION OF Cry1Ac PROTEIN-DEGRADING BACTERIA AND ITS DEGRADATION CHARACTERISTICS

    Institute of Scientific and Technical Information of China (English)

    赵辉欣; 刘阳; 邢福国

    2011-01-01

    A degradation of Cry1Ac protein in rice straw,bacterium FJSB3 capable of highly degrading Cry1Ac protein was isolated from the agricultural soil where the transgenic Cry1Acrice was grown.The FJSB3 was identified preliminarily as Stenotrophomonas sp.by phenotype observation,16S rDNA analysis and electron microscope.SDS-PAGE showed that FJSB3 could degrade Btprotein.Based on the single factor experiments,the optimal conditions of degrading Cry1Ac protein in rice straw were: 30℃,initial concentration of bacterium 2.0×106 CFU/ml,pH 7.0,C/N 25/1.Under these conditions,the degradation rate of Cry1Ac protein reached 92.86%.%从转Cry1Ac基因水稻种植田土壤中,分离纯化得到1株能高效降解Cry1Ac蛋白的细菌FJSB3。通过表型特征、16S rDNA扩增和电镜观察,初步鉴定FJSB3为寡养单胞菌(Stenotrophomonas sp.)。FJSB3发酵液能降解粗Bt蛋白。通过单因素试验确定FJSB3降解水稻秸秆中Cry1Ac蛋白的最适条件为:发酵温度30℃,初始菌液浓度2.0×106CFU/ml,pH 7.0,碳氮比(C/N)25/1。在此条件下,水稻秸秆中Cry1Ac蛋白降解率达到92.86%。

  12. Signal Peptide of Potato PinⅡ Enhances the Expression of Cry1Ac in Transgenic Tobacco

    Institute of Scientific and Technical Information of China (English)

    Yun-Jun LIU; Yuan YUAN; Jun ZHENG; Ya-Zhong TAO; Zhi-Gang DONG; Jian-Hua WANG; Guo-Ying WANG

    2004-01-01

    The modified Cry1Ac was expressed in transgenic tobacco plants.To allow secretion of the Cry1Ac protein into the intercellular space,the signal peptide sequence of potato proteinase inhibitor Ⅱ(pinⅡ)was N-terminally fused to the CrylAc encoding region.Expression of Cry1Ac in transgenic tobacco plants was assayed with ELISA.The results showed that pinⅡ signal peptide sequence enhanced the expression of Cry 1 Ac protein and led to the secretion of the Cry 1Ac protein in transgenic tobacco plants.GFP gene was also fused to the signal peptide sequence and transformed to tobacco.The results of fluorescent detection showed that GFP had localized in the apoplast of transgenic plants.

  13. EXPRESIÓN DE LA PROTEÍNA Cry1Ac EN TEJIDOS DE LÍNEASTRANSGÉNICAS DE PAPA (Solanum tuberosum spp. ANDÍGENAVAR. DIACOL CAPIRO. Cry1Ac Protein Expression in Tissues of Potato (Solanumtuberosum spp. Andígena Transgenic Lines Var. Diacol Capiro

    Directory of Open Access Journals (Sweden)

    PABLO ANDRÉS VANEGAS ARAUJO

    genetic plant breeding related to the introduction of Cry genes which codify insecticidal crystal proteins is an alternative for reducing the insect attack in commercial crops. In this work, the insertion, transcription and expression of Cry1Ac gen was characterized in different tissues and three development stages of two transgenic lines of Solanum tuberosum variety Diacol Capiro that were previously transformed by Agrobacterium tumefaciens method. The characterization was realized by PCR, RT-PCR and ELISA techniques. The gen insertion and transcription was confirmed using primers for Cry1Ac gen that amplified a specific band of 766 bp. The protein expression levels were higher than 45 µg/g and were not significantly different between the analyzed lines nor the three development stages. Furthermore, taking into account some relevant phenotypic features, no significant differences were found between transgenic lines and controls. The results suggest that monitoring and biosecurity assays are necessary with this vegetal material because their high level expression inside all the tissues analyzed that could affect non-targeted insects.

  14. Agrotis segetum midgut putative receptor of Bacillus thuringiensis vegetative insecticidal protein Vip3Aa16 differs from that of Cry1Ac toxin

    OpenAIRE

    Ben Hamadou-Charfi, Dorra; Boukedi, Hanen; Abdelkefi-Mesrati, Lobna; Tounsi, Slim; Jaoua, Samir

    2013-01-01

    Considering the fact that Agrotis segetum is one of the most pathogenic insects to vegetables and cereals in the world, particularly in Africa, the mode of action of Vip3Aa16 of Bacillus thuringiensis BUPM95 and Cry1Ac of the recombinant strain BNS3Cry-(pHTcry1Ac) has been examined in this crop pest. A. segetum proteases activated the Vip3Aa16 protoxin (90kDa) yielding three bands of about 62, 45, 22kDa and the activated form of the toxin was active against this pest with an LC50 of about 86n...

  15. 棉铃虫中肠氨肽酶 APN4与 Cry1Ac、Cry2Aa 结合能力的比较%Comparison of the binding affinity of the APN4 receptor in Helicoverpa armigera (Hübner) to the Cry1Ac and Cry2Aa insecticidal proteins

    Institute of Scientific and Technical Information of China (English)

    袁向东; 葛朝虹; 肖玉涛; 梁革梅

    2015-01-01

    Objectives] Bt (Bacillus thuringiensis) insecticidal proteins have been widely adopted to control agricultural pests because of their high target specificity. The binding of a Bt insecticidal protein to its specific receptor in the insect midgut plays a key role in the insecticidal action process. Aminopeptidase N (APN) is one of the major Bt protein receptors. To better characterize the molecular mechanism underlying the insecticidal activity of different Bt insecitcidal proteins, and lay the foundation for Bt resistance management and novel Bt insecticidal protein development, we analyzed the binding affinity of aminopeptidase N4 (APN4) to the Cry1Ac and Cry2Aa insecitcidal proteins in Helicoverpa armigera. [Methods] The binding affinity of aminopeptidase N4 (APN4) was assessed using ligand blot analysis and an ELISA binding assay, respectively. [Results] The results show that recombinant APN4 could bind to both Cry1Ac and Cry2A; their respective dissociation constants were 46.7 nmol/L and 26.5 nmol/L. [Conclusion] The results suggest that there was no significant difference in the binding affinity of APN4 to Cry1Ac and Cry2Aa in H. armigera.%【目的】 Bt 杀虫蛋白(Bacillus thuringiensis)具有高度的靶标特异性,已经被广泛用于农业害虫防治。Bt 杀虫蛋白要发挥杀虫活性,必须首先与其受体蛋白结合,氨肽酶 N(Aminopeptidase N)是一类重要的 Bt 受体蛋白。因此,分析该受体与 Bt 杀虫蛋白的结合能力,可为进一步明确不同 Bt 的分子作用机制、Bt 的抗性治理以及新 Bt 的开发应用等提供借鉴。【方法】本文利用 Ligand blot 和 Elisa 方法比较了棉铃虫 Helicoverpa armigera 中肠 APN4(Aminopeptidase N4,APN4)与 Cry1Ac、Cry2Aa 的结合能力。【结果】原核表达的 APN4片段与活化的 Cry1Ac、Cry2Aa 都可以结合,解离常数(Kd)分别是48.59 nmol/L和21.73 nmol/L。【结论】 APN4片段与 Cry1Ac、Cry2Aa 的结合能力

  16. Vip3Aa tolerance response of Helicoverpa armigera populations from a Cry1Ac cotton planting region.

    Science.gov (United States)

    An, Jingjie; Gao, Yulin; Wu, Kongming; Gould, Fred; Gao, Jianhua; Shen, Zhicheng; Lei, Chaoliang

    2010-12-01

    Transgenic cotton, Gossypium hirsutum L., that expresses the Bacillus thuringiensis (Bt) Cry1Ac toxin, holds great promise in controlling target insect pests. Evolution of resistance by target pests is the primary threat to the continued efficacy of Bt cotton. To thwart pest resistance evolution, a transgenic cotton culitvar that produces two different Bt toxins, cry1Ac and vip3A genes, was proposed as a successor of cry1Ac cotton. This article reports on levels of Vip3Aa tolerance in Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) populations from the Cry1Ac cotton planting region in China based on bioassays of the F1 generation of isofemale lines. In total, 80 isofemale families of H. armigera from Xiajin county of Shandong Province (an intensive Bt cotton planting area) and 93 families from Anci county of Hebei Province (a multiple-crop system including corn [Zea mays L.] , soybean [Glycine max (L.) Merr.], peanut (Arachis hypogaea L.), and Bt cotton) were screened with a discriminating concentration of both Cry1Ac- and Vip3A-containing diets in 2009. From data on the relative average development rates and percentage of larval weight inhibition of F1 full-sib families tested simultaneously on Cry1Ac and Vip3Aa, results indicate that responses to Cry1Ac and Vip3Aa were not genetically correlated in field population ofH. armigera. This indicates that the threat of cross-resistance between Cry1Ac and Vip3A is low in field populations of H. armigera. Thus, the introduction of Vip3Aa/Cry1Ac-producing lines could delay resistance evolution in H. armigera in Bt cotton planting area of China.

  17. Produção da toxina Cry1Ac e preferência para alimentação e oviposição de Alabama argillacea em algodão Bt sob estresse hídrico

    OpenAIRE

    Mauricio Silva de Lima; Jorge Braz Torres

    2011-01-01

    O objetivo deste trabalho foi avaliar a produção da toxina Cry1Ac de Bacillus thuringiensis (Bt) em plantas de algodão geneticamente modificado (algodão Bt), e a preferência para alimentação e oviposição de Alabama argillacea confinada em algodão Bt e não Bt sob estresse hídrico. As variedades de algodão Bt - Acala 90B e NuOpal - e não Bt - Acala 90 e DeltaOpal - foram cultivadas em microparcelas, com e sem estresse hídrico. A produção de toxina foi avaliada em folhas e partes reprodutivas da...

  18. 基因枪介导法获得转BtCry1Ac基因抗虫玉米植株的研究%Genetic Transformation of BtCry1Ac into Maize by Particle Bombardment

    Institute of Scientific and Technical Information of China (English)

    铁双贵; 柏松; 岳润清; 齐建双; 王延召; 孙静; 陈小洁; 田保明

    2012-01-01

    CrylA c gene was introduced into immature embryo callus using particle bombardment. After selection, transgenic plants were regenerated. Some of them were positive by PCR and immune test paper analysis, it was demonstrated that the Cry1 Ac gene had been intergraded into plants genome and expressed in transgenic plants. Bioassay results showed that T) generation transgenic plants had high resistance to the Asian corn borer.%利用基因枪介导法将含有Cry1Ac基因的质粒转入玉米愈伤组织,获得转Bt基因玉米植株,对所获得的转基因植株进行PCR分析和金标免疫试纸鉴定.结果表明,部分转化植株呈阳性,说明目的基因已经整合到玉米基因组中.将转基因植株自交获得T1代种子,对T1代植株进行田间抗虫鉴定,结果表明,与对照相比转基因玉米植株的抗虫活性明显强于非转基因植株.

  19. Produção da toxina Cry1Ac e preferência para alimentação e oviposição de Alabama argillacea em algodão Bt sob estresse hídrico

    Directory of Open Access Journals (Sweden)

    Mauricio Silva de Lima

    2011-05-01

    Full Text Available O objetivo deste trabalho foi avaliar a produção da toxina Cry1Ac de Bacillus thuringiensis (Bt em plantas de algodão geneticamente modificado (algodão Bt, e a preferência para alimentação e oviposição de Alabama argillacea confinada em algodão Bt e não Bt sob estresse hídrico. As variedades de algodão Bt - Acala 90B e NuOpal - e não Bt - Acala 90 e DeltaOpal - foram cultivadas em microparcelas, com e sem estresse hídrico. A produção de toxina foi avaliada em folhas e partes reprodutivas das variedades de algodão Bt. Para o teste de escolha para oviposição, mariposas de A. argillacea foram liberadas em telado que continha as quatro variedades do algodão. Após 72 horas, o número de ovos foi quantificado. Para a preferência de alimentação, foram avaliadas lagartas com três e 10 dias de idade. A produção da toxina Cry1Ac foi superior nas folhas da parte apical das plantas, das duas variedades sob estresse hídrico, e similar nas brácteas e cascas de maçãs. Lagartas e mariposas de A. argillacea não diferenciam algodão Bt e não Bt na escolha quanto à alimentação e à oviposição, respectivamente. Entretanto, as mariposas preferem ovipositar em plantas sem estresse hídrico.

  20. Increased long-flight activity triggered in beet armyworm by larval feeding on diet containing Cry1Ac protoxin.

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    Xing Fu Jiang

    Full Text Available Evaluating ecological safety and conducting pest risk analysis for transgenic crops are vitally important before their commercial planting. The beet armyworm, Spodoptera exigua, a long-distance migratory insect pest, is not a direct target of transgenic Cry1Ac-expressing cotton in China, but nevertheless it has recently become an important pest. Migrants leaving their natal field arrive in other appropriate habitat far away in a short time, often followed by larval outbreaks. S. exigua has low susceptibility to Cry1Ac. However, our results from laboratory experiments identified (i sublethal effects of Cry1Ac protoxin on larval development rate, larval and pupal weight, and adult lifetime fecundity, and (ii increased long-flight behavior triggered by Cry1Ac which may contribute to larval outbreaks elsewhere. No significant differences in larval mortality, pupation rate, adult emergence rate, longevity, pre-oviposition period, or oviposition period were observed between controls and larvae fed on artificial diet incorporating a low concentration of Cry1Ac protoxin. The negative sublethal effects on some developmental and reproductive traits and lack of effect on others suggest they do not contribute to the observed severity of S. exigua outbreaks after feeding on Cry1Ac cotton. Interestingly, the percentage of long fliers increased significantly when larvae were reared on diet containing either of two low-dose treatments of Cry1Ac, suggesting a possible increased propensity to disperse long distances triggered by Cry1Ac. We hypothesize that negative effects on development and reproduction caused by Cry1Ac in the diet are offset by increased flight propensity triggered by the poor food conditions, thereby improving the chances of escaping adverse local conditions before oviposition. Increased long-flight propensity in turn may amplify the area damaged by outbreak populations. This phenomenon might be common in other migratory insect pests receiving

  1. Evaluation of cytotoxic and antimicrobial effects of two Bt Cry proteins on a GMO safety perspective.

    Science.gov (United States)

    Farias, Davi Felipe; Viana, Martônio Ponte; de Oliveira, Gustavo Ramos; Beneventi, Magda Aparecida; Soares, Bruno Marques; Pessoa, Claudia; Pessoa, Igor Parra; Silva, Luciano Paulino; Vasconcelos, Ilka Maria; de Sá, Maria Fátima Grossi; Carvalho, Ana Fontenele Urano

    2014-01-01

    Studies have contested the innocuousness of Bacillus thuringiensis (Bt) Cry proteins to mammalian cells as well as to mammals microbiota. Thus, this study aimed to evaluate the cytotoxic and antimicrobial effects of two Cry proteins, Cry8Ka5 (a novel mutant protein) and Cry1Ac (a widely distributed protein in GM crops). Evaluation of cyto- and genotoxicity in human lymphocytes was performed as well as hemolytic activity coupled with cellular membrane topography analysis in mammal erythrocytes. Effects of Cry8Ka5 and Cry1Ac upon Artemia sp. nauplii and upon bacteria and yeast growth were assessed. The toxins caused no significant effects on the viability (IC50 > 1,000 µg/mL) or to the cellular DNA integrity of lymphocytes (no effects at 1,000 µg/mL). The Cry8Ka5 and Cry1Ac proteins did not cause severe damage to erythrocytes, neither with hemolysis (IC50 > 1,000 µg/mL) nor with alterations in the membrane. Likewise, the Cry8Ka5 and Cry1Ac proteins presented high LC50 (755.11 and >1,000 µg/mL, resp.) on the brine shrimp lethality assay and showed no growth inhibition of the microorganisms tested (MIC > 1,000 µg/mL). This study contributed with valuable information on the effects of Cry8Ka5 and Cry1Ac proteins on nontarget organisms, which reinforce their potential for safe biotechnological applications.

  2. 分泌抗Bt Cry1Ac蛋白单克隆抗体杂交瘤细胞株的建立%The Establishment of Hybridoma Cell Line Secreting Specific Monoclonal Antibodies Against Bt Cry1Ac

    Institute of Scientific and Technical Information of China (English)

    乔艳红; 张维; 林敏; 张杰; 潘家荣

    2005-01-01

    从苏云金芽孢杆菌HD-73中提取Bt Cry1Ac蛋白,电镜观察Bt Cry1Ac蛋白为标准的菱形.用SP2/0-Ag14骨髓瘤细胞与经该Bt Cry1Ac蛋白免疫的BALB/c小鼠的脾细胞融合,经3次克隆化,筛出两株稳定分泌Bt Cry1Ac单克隆的杂交瘤细胞株1C3、2F3.两株细胞均具有抗Bt Cry1Ac特异性,与Bt Cry1Ab和Bt Cry2A无明显的交叉反应,经亚型鉴定均为IgG1.腹水滴度均为1∶1024000.

  3. Downregulation and Mutation of a Cadherin Gene Associated with Cry1Ac Resistance in the Asian Corn Borer, Ostrinia furnacalis (Guenée

    Directory of Open Access Journals (Sweden)

    Tingting Jin

    2014-09-01

    Full Text Available Development of resistance in target pests is a major threat to long-term use of transgenic crops expressing Bacillus thuringiensis (Bt Cry toxins. To manage and/or delay the evolution of resistance in target insects through the implementation of effective strategies, it is essential to understand the basis of resistance. One of the most important mechanisms of insect resistance to Bt crops is the alteration of the interactions between Cry toxins and their receptors in the midgut. A Cry1Ac-selected strain of Asian corn borer (ACB, Ostrinia furnacalis, a key pest of maize in China, evolved three mutant alleles of a cadherin-like protein (OfCAD (MPR-r1, MPR-r2 and MPR-r3, which mapped within the toxin-binding region (TBR. Each of the three mutant alleles possessed two or three amino acid substitutions in this region, especially Thr1457→Ser. In highly resistant larvae (ACB-Ac200, MPR-r2 had a 26-amino acid residue deletion in the TBR, which resulted in reduced binding of Cry1Ac compared to the MPR from the susceptible strain, suggesting that the number of amino acid deletions influences the level of resistance. Furthermore, downregulation of OfCAD gene (ofcad transcription was observed in the Cry1Ac resistant strain, ACB-Ac24, suggesting that Cry1Ac resistance in ACB is associated with the downregulation of the transcript levels of the cadherin-like protein gene. The OfCAD identified from ACB exhibited a high degree of similarity to other members of the cadherin super-family in lepidopteran species.

  4. Effects of Bacillus thuringiensis toxin Cry1Ac and Beauveria bassiana on Asiatic corn borer (Lepidoptera: Crambidae).

    Science.gov (United States)

    Ma, Xiao-Mu; Liu, Xiao-Xia; Ning, Xia; Zhang, Bo; Han, Fei; Guan, Xiu-Min; Tan, Yun-Feng; Zhang, Qing-Wen

    2008-10-01

    In this study, interactions between Cry1Ac, a toxic crystal protein produced by Bacillus thuringiensis (Berliner), and Beauveria bassiana on the mortality and survival of Ostrinia furnacalis was evaluated in the laboratory. The results showed that Cry1Ac is toxic to O. furnacalis. Not only were larval growth and development delayed, but pupation, pupal weight and adult emergency also decreased when larvae were fed on artificial diet containing purified Cry1Ac toxin. When third instars O. furnacalis were exposed to combination of B. bassiana (1.8x10(5), 1.8x10(6) or 1.8x10(7) conidia ml(-1)) and Cry1Ac, (0.2 or 0.8 microg g(-1)), the effect on mortality was additive, however, the combinations of sublethal concentrations showed antagonism between Cry1Ac (3.2 or 13 microg g(-1)) and B. bassiana (1.8x10(5) or 1.8x10(6) conidia ml(-1)). When neonates were reared on sublethal concentrations of Cry1AC until the third instar, and survivors exposed B. bassiana conidial suspension, such treatments showed additive effect on mortality of O. furnacalis except for the combination of Cry1Ac (0.2 microg g(-1)) and B. bassiana (1.8x10(6) conidia ml(-1)) that showed antagonism.

  5. Diversity in gut microflora of Helicoverpa armigera populations from different regions in relation to biological activity of Bacillus thuringiensis δ-endotoxin Cry1Ac.

    Science.gov (United States)

    Paramasiva, Inakarla; Shouche, Yogesh; Kulkarni, Girish Jayant; Krishnayya, Pulipaka Venkata; Akbar, Shaik Mohammed; Sharma, Hari Chand

    2014-12-01

    Transgenic crops expressing toxin proteins from Bacillus thuringiensis (Bt) have been deployed on a large scale for management of Helicoverpa armigera. Resistance to Bt toxins has been documented in several papers, and therefore, we examined the role of midgut microflora of H. armigera in its susceptibility to Bt toxins. The susceptibility of H. armigera to Bt toxin Cry1Ac was assessed using Log-dose-Probit analysis, and the microbial communities were identified by 16S rRNA sequencing. The H. armigera populations from nine locations harbored diverse microbial communities, and had some unique bacteria, suggesting a wide geographical variation in microbial community in the midgut of the pod borer larvae. Phylotypes belonging to 32 genera were identified in the H. armigera midgut in field populations from nine locations. Bacteria belonging to Enterobacteriaceae (Order Bacillales) were present in all the populations, and these may be the common members of the H. armigera larval midgut microflora. Presence and/or absence of certain species were linked to H. armigera susceptibility to Bt toxins, but there were no clear trends across locations. Variation in susceptibility of F1 neonates of H. armigera from different locations to the Bt toxin Cry1Ac was found to be 3.4-fold. These findings support the idea that insect migut microflora may influence the biological activity of Bt toxins.

  6. Binding and Oligomerization of Modified and Native Bt Toxins in Resistant and Susceptible Pink Bollworm.

    Directory of Open Access Journals (Sweden)

    Josue Ocelotl

    Full Text Available Insecticidal proteins from Bacillus thuringiensis (Bt are used extensively in sprays and transgenic crops for pest control, but their efficacy is reduced when pests evolve resistance. Better understanding of the mode of action of Bt toxins and the mechanisms of insect resistance is needed to enhance the durability of these important alternatives to conventional insecticides. Mode of action models agree that binding of Bt toxins to midgut proteins such as cadherin is essential for toxicity, but some details remain unresolved, such as the role of toxin oligomers. In this study, we evaluated how Bt toxin Cry1Ac and its genetically engineered counterpart Cry1AcMod interact with brush border membrane vesicles (BBMV from resistant and susceptible larvae of Pectinophora gossypiella (pink bollworm, a global pest of cotton. Compared with Cry1Ac, Cry1AcMod lacks 56 amino acids at the amino-terminus including helix α-1; previous work showed that Cry1AcMod formed oligomers in vitro without cadherin and killed P. gossypiella larvae harboring cadherin mutations linked with >1000-fold resistance to Cry1Ac. Here we found that resistance to Cry1Ac was associated with reduced oligomer formation and insertion. In contrast, Cry1AcMod formed oligomers in BBMV from resistant larvae. These results confirm the role of cadherin in oligomerization of Cry1Ac in susceptible larvae and imply that forming oligomers without cadherin promotes toxicity of Cry1AcMod against resistant P. gossypiella larvae that have cadherin mutations.

  7. Asymmetrical cross-resistance between Bacillus thuringiensis toxins Cry1Ac and Cry2Ab in pink bollworm.

    Science.gov (United States)

    Tabashnik, Bruce E; Unnithan, Gopalan C; Masson, Luke; Crowder, David W; Li, Xianchun; Carrière, Yves

    2009-07-21

    Transgenic crops producing Bacillus thuringiensis (Bt) toxins kill some key insect pests and can reduce reliance on insecticide sprays. Sustainable use of such crops requires methods for delaying evolution of resistance by pests. To thwart pest resistance, some transgenic crops produce 2 different Bt toxins targeting the same pest. This "pyramid" strategy is expected to work best when selection for resistance to 1 toxin does not cause cross-resistance to the other toxin. The most widely used pyramid is transgenic cotton producing Bt toxins Cry1Ac and Cry2Ab. Cross-resistance between these toxins was presumed unlikely because they bind to different larval midgut target sites. Previous results showed that laboratory selection with Cry1Ac caused little or no cross-resistance to Cry2A toxins in pink bollworm (Pectinophora gossypiella), a major cotton pest. We show here, however, that laboratory selection of pink bollworm with Cry2Ab caused up to 420-fold cross-resistance to Cry1Ac as well as 240-fold resistance to Cry2Ab. Inheritance of resistance to high concentrations of Cry2Ab was recessive. Larvae from a laboratory strain resistant to Cry1Ac and Cry2Ab in diet bioassays survived on cotton bolls producing only Cry1Ac, but not on cotton bolls producing both toxins. Thus, the asymmetrical cross-resistance seen here does not threaten the efficacy of pyramided Bt cotton against pink bollworm. Nonetheless, the results here and previous evidence indicate that cross-resistance occurs between Cry1Ac and Cry2Ab in some key cotton pests. Incorporating the potential effects of such cross-resistance in resistance management plans may help to sustain the efficacy of pyramided Bt crops.

  8. Effects of transgenic Bt (Cry1Ab/Cry1Ac) rice on the population development of Plodia interpunctella%转Bt基因稻谷对印度谷螟生长发育的影响

    Institute of Scientific and Technical Information of China (English)

    蒋海燕; 王振华; 华红霞; 高艮亮; 蔡万伦; 杨长举

    2011-01-01

    The effect of the transgenic Bt rice CrylAb/ CrylAc gene on the feeding of Plodia interpunctella ( Hlibner)on stored rice was investigated. Population parameters of P. Interpunctella were investigated after adding different ratios (70% , 50% , 30% , 10% ) of transgenic Bt rice flour into a conventional artificial diet at 30°C. And RH75%. The content of Bt protein in 3rd instar larva and the transgenic Bt rice grain was measured using the ELISA method. The results show that significant mortality of 1" instar larva was observed 72 h after feeding on Bt rice flour. Larval mortality rates and development time increased significantly as the content of transgenic Bt rice flour in the artificial diet increased. For research purposes, 2. 35 u.g/g was the optimal sub-lethal dose of Cryl Ab/Cryl Ac protein for P. Interpunctella.%为建立仓储阶段转Bt水稻安全性评价中靶标害虫抗性汰选研究体系,配制了含不同比例(70%,50%,30%,10%)转Bt基因(Cry1Ab/Cry IAbc)明辉63水稻谷粉(简称Bt谷粉)的人工饲料饲喂印度谷螟Plodiainterpunctella (Hübner),测定其对1~3龄幼虫在72 h内的急性毒力,及对印度谷螟种群生长发育的影响,并采用ELISA法检测转基因稻谷和末龄幼虫体内Bt蛋白含量.结果发现:4种比例人工饲料对幼虫的毒力作用均发生在取食48 h后,72 h后剂量效应明显.含Bt水稻较高比例的饲料对印度谷螟发育的负面效应明显:幼虫死亡率高,发育历期延长.Bt蛋白在幼虫体内含量与对应饲料中的含量基本成正比.综合考虑,将Bt杀虫蛋白含量2.35 μg/g作为转Bt基因稻谷对印度谷螟的亚致死剂量最为合适.

  9. Transgenic Cabbage Expressing Cry1Ac1 Does Not Affect the Survival and Growth of the Wolf Spider, Pardosa astrigera L. Koch (Araneae: Lycosidae)

    Science.gov (United States)

    Kim, Young-Joong; Lee, Joon-Ho; Harn, Chee Hark; Kim, Chang-Gi

    2016-01-01

    Both herbivores that consume transgenic crops and their predators can be exposed to insecticidal proteins expressed in those crops. We conducted a tritrophic bioassay to evaluate the ecotoxicological impacts that Bt cabbage (Brassica oleracea var. capitata) expressing Cry1Ac1 protein might have on the wolf spider (Pardosa astrigera), a non-target generalist predator. Enzyme-Linked Immunosorbent Assays indicated that protein levels were 4.61 ng g-1 dry weight in fruit flies (Drosophila melanogaster) fed with the transgenic cabbage and 1.86 ng g-1 dry weight in the wolf spiders that preyed upon them. We also compared the life history traits of spiders collected from Bt versus non-Bt cabbage and found no significant differences in their growth, survival, and developmental rates. Because Bt cabbage did not affect the growth of fruit flies, we conclude that any indirect effects that this crop had on the wolf spider were probably not mediated by prey quality. Therefore, exposure to Cry1Ac1 protein when feeding upon prey containing that substance from transgenic cabbage has only a negligible influence on those non-target predatory spiders. PMID:27055120

  10. Transgenic Cabbage Expressing Cry1Ac1 Does Not Affect the Survival and Growth of the Wolf Spider, Pardosa astrigera L. Koch (Araneae: Lycosidae).

    Science.gov (United States)

    Kim, Young-Joong; Lee, Joon-Ho; Harn, Chee Hark; Kim, Chang-Gi

    2016-01-01

    Both herbivores that consume transgenic crops and their predators can be exposed to insecticidal proteins expressed in those crops. We conducted a tritrophic bioassay to evaluate the ecotoxicological impacts that Bt cabbage (Brassica oleracea var. capitata) expressing Cry1Ac1 protein might have on the wolf spider (Pardosa astrigera), a non-target generalist predator. Enzyme-Linked Immunosorbent Assays indicated that protein levels were 4.61 ng g(-1) dry weight in fruit flies (Drosophila melanogaster) fed with the transgenic cabbage and 1.86 ng g(-1) dry weight in the wolf spiders that preyed upon them. We also compared the life history traits of spiders collected from Bt versus non-Bt cabbage and found no significant differences in their growth, survival, and developmental rates. Because Bt cabbage did not affect the growth of fruit flies, we conclude that any indirect effects that this crop had on the wolf spider were probably not mediated by prey quality. Therefore, exposure to Cry1Ac1 protein when feeding upon prey containing that substance from transgenic cabbage has only a negligible influence on those non-target predatory spiders.

  11. Characterization and functional analysis ofβ-1,3-galactosyltransferase involved in Cry1Ac resistance from Helicoverpa armigera (Hübner)

    Institute of Scientific and Technical Information of China (English)

    ZHANG Li-li; LIANG Ge-mei; GAO Xi-wu; CAO Guang-chun; GUO Yu-yuan

    2015-01-01

    Carbohydrate chains are the principal antigens by which Bacil us thuringiensis (Bt) identify receptor proteins. The interaction between the antigen and Bt causes a pore in the membrane of midgut epithelial cel s of insects. Receptor proteins, such as aminopeptidase N and alkaline phosphatase, are glycoproteins. Cadherin is another cel surface receptor protein which has potential glycosylation sites. Glycosyltransferase is very important for the synthesis and modiifcation of receptor proteins. It can indirectly inlfuence the function of Bt. The 1 950 bp ful-length cDNA encodingβ-1,3-galactosyltransferase was cloned from the the midgut of Helicoverpa armigera by degenerative PCR combined with RACE techniques (GAL-Harm, GenBank accession no.:GQ904195.1) with two potential N-glycosylation sites (157NNTI160 and 272NKTL275). Protein sequence alignments revealed that H. armigeraβ-1,3-galactosyltransferase shared high identity withβ-1,3-galactosyltransferase in other insect species. The expression level of theβ-1,3-galactosyltransferase gene in Cry1Ac-resistant H. armigera larvae was 9.2-fold higher than that in susceptible strain. The function ofβ-1,3-galactosyltransferase was investigated using RNAi technique. The result showed Cry1Ac enhanced the toxicity against the siRNA-treated larvae compared with non-siRNA-treated ones, which indicatedβ-1,3-galactosyltransferase played an important role for the insecticidal toxicity of Cry1Ac in H. armigera.

  12. Interactions of Bacillus thuringiensis Cry1Ac toxin in genetically engineered cotton with predatory heteropterans.

    Science.gov (United States)

    Torres, Jorge B; Ruberson, John R

    2008-06-01

    A number of cotton varieties have been genetically transformed with genes from Bacillus thuringiensis (Bt) to continuously produce Bt endotoxins, offering whole plant and season-long protection against many lepidopteran larvae. Constant whole-plant toxin expression creates a significant opportunity for non-target herbivores to acquire and bio-accumulate the toxin for higher trophic levels. In the present study we investigated movement of Cry1Ac toxin from the transgenic cotton plant through specific predator-prey pairings, using omnivorous predators with common cotton pests as prey: (1) the beet armyworm, Spodoptera exigua (Lepidoptera: Noctuidae), with the predator Podisus maculiventris (Heteroptera: Pentatomidae); (2) the two-spotted spider mite, Tetranychus urticae (Acarina: Tetranychidae), with the predatory big-eyed bug Geocoris punctipes (Heteroptera: Geocoridae) and (3) with the predatory damsel bug Nabis roseipennis (Heteropera: Nabidae); and (4) the thrips Frankliniella occidentalis (Thysanoptera: Thripidae) with the predatory pirate bug Orius insidiosus (Heteroptera: Anthocoridae). We quantified Cry1Ac toxin in the cotton plants, and in the pests and predators, and the effects of continuous feeding on S. exigua larvae fed either Bt or non-Bt cotton on life history traits of P. maculiventris. All three herbivores were able to convey Cry1Ac toxin to their respective predators. Among the herbivores, T. urticae exhibited 16.8 times more toxin in their bodies than that expressed in Bt-cotton plant, followed by S. exigua (1.05 times), and F. occidentalis immatures and adults (0.63 and 0.73 times, respectively). Of the toxin in the respective herbivorous prey, 4, 40, 17 and 14% of that amount was measured in the predators G. punctipes, P. maculiventris, O. insidiosus, and N. roseipennis, respectively. The predator P. maculiventris exhibited similar life history characteristics (developmental time, survival, longevity, and fecundity) regardless of the prey's food

  13. Dual mode of action of Bt proteins: protoxin efficacy against resistant insects

    OpenAIRE

    2015-01-01

    Transgenic crops that produce Bacillus thuringiensis (Bt) proteins for pest control are grown extensively, but insect adaptation can reduce their effectiveness. Established mode of action models assert that Bt proteins Cry1Ab and Cry1Ac are produced as inactive protoxins that require conversion to a smaller activated form to exert toxicity. However, contrary to this widely accepted paradigm, we report evidence from seven resistant strains of three major crop pests showing that Cry1Ab and Cry1...

  14. Effects of different brush border membrane vesicle isolation protocols on proteomic analysis of Cry1 Ac binding proteins from the midgut of Helicoverpa armigera

    Institute of Scientific and Technical Information of China (English)

    Li-Zhen Chen; Ge-Mei Liang; Brian G.Rector; Jie Zhang; Kong-Ming Wu; Yu-Yuan Guo

    2008-01-01

    Brush border membrane vesicles(BBMV)isolated from insect midguts have been widely used to study CrylA binding proteins.Sample preparation is important in two-dimensional electrophoresis(2-DE),so to determine a suitable BBMV preparation method in Helicoverpa armigera for 2-DE,we compared three published BBMV preparation methods mostly used in sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE).All mctllods yielded similar types and numbers of binding proteins,but in different quantifies.The Abdul.Rauf and Ellar protocol was the best of the three,but had limitations.Sufficient protein qu antity iS important for research involving limited numbers of insects,such as studies of insect resistance to Bacillus thuringiensis in the field.Consequently,we integrated the three BBMV isolation methods into a single protocol that yielded high quantities of BBMV proteins from H.armigera larval midguts.which proved suitable for 2-DE analysis.

  15. Potential factors impacting season-long expression of Cry1Ac in 13 commercial varieties of Bollgard® cotton

    Directory of Open Access Journals (Sweden)

    John J. Adamczyk, Jr.

    2001-11-01

    Full Text Available Thirteen commercial varieties of transgenic Cry1Ac Bacillus thuringiensis Berliner (Bt cotton were examined across two sites in 2000 for potential factors that impact endotoxin expression. In all cases, two varieties (NuCOTN 33B and DP 458B/RR, Delta and Pineland Co., Scott, MS expressed more Cry1Ac than the other 11 varieties in various plant structures. These two varieties share the same parental background (DP 5415. Furthermore, when the next generation of plants were tested in the greenhouse, the same varietal patterns were exhibited. These data strongly suggest that factors such as parental background had a stronger impact on the expression of Cry1Ac than the environment.

  16. Dual mode of action of Bt proteins: protoxin efficacy against resistant insects.

    Science.gov (United States)

    Tabashnik, Bruce E; Zhang, Min; Fabrick, Jeffrey A; Wu, Yidong; Gao, Meijing; Huang, Fangneng; Wei, Jizhen; Zhang, Jie; Yelich, Alexander; Unnithan, Gopalan C; Bravo, Alejandra; Soberón, Mario; Carrière, Yves; Li, Xianchun

    2015-10-12

    Transgenic crops that produce Bacillus thuringiensis (Bt) proteins for pest control are grown extensively, but insect adaptation can reduce their effectiveness. Established mode of action models assert that Bt proteins Cry1Ab and Cry1Ac are produced as inactive protoxins that require conversion to a smaller activated form to exert toxicity. However, contrary to this widely accepted paradigm, we report evidence from seven resistant strains of three major crop pests showing that Cry1Ab and Cry1Ac protoxins were generally more potent than the corresponding activated toxins. Moreover, resistance was higher to activated toxins than protoxins in eight of nine cases evaluated in this study. These data and previously reported results support a new model in which protoxins and activated toxins kill insects via different pathways. Recognizing that protoxins can be more potent than activated toxins against resistant insects may help to enhance and sustain the efficacy of transgenic Bt crops.

  17. Evaluation of genetically modified sugarcane lines carrying Cry 1AC gene using molecular marker techniques.

    Science.gov (United States)

    Ismail, Roba M

    2013-01-01

    Five genetically modified insect resistant sugarcane lines harboring the Bt Cry 1AC gene to produce insecticidal proteins were compared with non-transgenic control by using three types of molecular marker techniques namely, RAPD, ISSR and AFLP. These techniques were applied on transgenic and non-transgenic plants to investigate the genetic variations, which may appear in sugarcane clones. This variation might demonstrate the genomic changes associated with the transformation process, which could change important molecular basis of various biological phenomena. Genetic variations were screened using 22 different RAPD primers, 10 ISSR primers and 13 AFLP primer combinations. Analysis of RAPD and ISSR banding patterns gave no exclusive evidence for genetic variations. Meanwhile, the percentage of polymorphic bands was 0.45% in each of RAPD and ISSR, while the polymorphism generated by AFLP analysis was 1.8%. The maximum percentage of polymorphic bands was 1.4%, 1.1% and 5.5% in RAPD, ISSR and AFLP, respectively. These results demonstrate that most transgenic lines showed genomic homogeneity and verified minor genomic changes. Dendrograms revealing the relationships among the transgenic and control plants were developed from the data of each of the three marker types.

  18. Effects of transgenic Cry1Ac + CpTI cotton on non-target mealybug pest Ferrisia virgata and its predator Cryptolaemus montrouzieri.

    Science.gov (United States)

    Wu, Hongsheng; Zhang, Yuhong; Liu, Ping; Xie, Jiaqin; He, Yunyu; Deng, Congshuang; De Clercq, Patrick; Pang, Hong

    2014-01-01

    Recently, several invasive mealybugs (Hemiptera: Pseudococcidae) have rapidly spread to Asia and have become a serious threat to the production of cotton including transgenic cotton. Thus far, studies have mainly focused on the effects of mealybugs on non-transgenic cotton, without fully considering their effects on transgenic cotton and trophic interactions. Therefore, investigating the potential effects of mealybugs on transgenic cotton and their key natural enemies is vitally important. A first study on the effects of transgenic cotton on a non-target mealybug, Ferrisia virgata (Cockerell) (Hemiptera: Pseudococcidae) was performed by comparing its development, survival and body weight on transgenic cotton leaves expressing Cry1Ac (Bt toxin) + CpTI (Cowpea Trypsin Inhibitor) with those on its near-isogenic non-transgenic line. Furthermore, the development, survival, body weight, fecundity, adult longevity and feeding preference of the mealybug predator Cryptolaemus montrouzieri Mulsant (Coleoptera: Coccinellidae) was assessed when fed F. virgata maintained on transgenic cotton. In order to investigate potential transfer of Cry1Ac and CpTI proteins via the food chain, protein levels in cotton leaves, mealybugs and ladybirds were quantified. Experimental results showed that F. virgata could infest this bivalent transgenic cotton. No significant differences were observed in the physiological parameters of the predator C. montrouzieri offered F. virgata reared on transgenic cotton or its near-isogenic line. Cry1Ac and CpTI proteins were detected in transgenic cotton leaves, but no detectable levels of both proteins were present in the mealybug or its predator when reared on transgenic cotton leaves. Our bioassays indicated that transgenic cotton poses a negligible risk to the predatory coccinellid C. montrouzieri via its prey, the mealybug F. virgata.

  19. Effects of Transgenic Cry1Ac + CpTI Cotton on Non-Target Mealybug Pest Ferrisia virgata and Its Predator Cryptolaemus montrouzieri

    Science.gov (United States)

    Wu, Hongsheng; Zhang, Yuhong; Liu, Ping; Xie, Jiaqin; He, Yunyu; Deng, Congshuang; De Clercq, Patrick; Pang, Hong

    2014-01-01

    Recently, several invasive mealybugs (Hemiptera: Pseudococcidae) have rapidly spread to Asia and have become a serious threat to the production of cotton including transgenic cotton. Thus far, studies have mainly focused on the effects of mealybugs on non-transgenic cotton, without fully considering their effects on transgenic cotton and trophic interactions. Therefore, investigating the potential effects of mealybugs on transgenic cotton and their key natural enemies is vitally important. A first study on the effects of transgenic cotton on a non-target mealybug, Ferrisia virgata (Cockerell) (Hemiptera: Pseudococcidae) was performed by comparing its development, survival and body weight on transgenic cotton leaves expressing Cry1Ac (Bt toxin) + CpTI (Cowpea Trypsin Inhibitor) with those on its near-isogenic non-transgenic line. Furthermore, the development, survival, body weight, fecundity, adult longevity and feeding preference of the mealybug predator Cryptolaemus montrouzieri Mulsant (Coleoptera: Coccinellidae) was assessed when fed F. virgata maintained on transgenic cotton. In order to investigate potential transfer of Cry1Ac and CpTI proteins via the food chain, protein levels in cotton leaves, mealybugs and ladybirds were quantified. Experimental results showed that F. virgata could infest this bivalent transgenic cotton. No significant differences were observed in the physiological parameters of the predator C. montrouzieri offered F. virgata reared on transgenic cotton or its near-isogenic line. Cry1Ac and CpTI proteins were detected in transgenic cotton leaves, but no detectable levels of both proteins were present in the mealybug or its predator when reared on transgenic cotton leaves. Our bioassays indicated that transgenic cotton poses a negligible risk to the predatory coccinellid C. montrouzieri via its prey, the mealybug F. virgata. PMID:24751821

  20. Effects of transgenic Cry1Ac + CpTI cotton on non-target mealybug pest Ferrisia virgata and its predator Cryptolaemus montrouzieri.

    Directory of Open Access Journals (Sweden)

    Hongsheng Wu

    Full Text Available Recently, several invasive mealybugs (Hemiptera: Pseudococcidae have rapidly spread to Asia and have become a serious threat to the production of cotton including transgenic cotton. Thus far, studies have mainly focused on the effects of mealybugs on non-transgenic cotton, without fully considering their effects on transgenic cotton and trophic interactions. Therefore, investigating the potential effects of mealybugs on transgenic cotton and their key natural enemies is vitally important. A first study on the effects of transgenic cotton on a non-target mealybug, Ferrisia virgata (Cockerell (Hemiptera: Pseudococcidae was performed by comparing its development, survival and body weight on transgenic cotton leaves expressing Cry1Ac (Bt toxin + CpTI (Cowpea Trypsin Inhibitor with those on its near-isogenic non-transgenic line. Furthermore, the development, survival, body weight, fecundity, adult longevity and feeding preference of the mealybug predator Cryptolaemus montrouzieri Mulsant (Coleoptera: Coccinellidae was assessed when fed F. virgata maintained on transgenic cotton. In order to investigate potential transfer of Cry1Ac and CpTI proteins via the food chain, protein levels in cotton leaves, mealybugs and ladybirds were quantified. Experimental results showed that F. virgata could infest this bivalent transgenic cotton. No significant differences were observed in the physiological parameters of the predator C. montrouzieri offered F. virgata reared on transgenic cotton or its near-isogenic line. Cry1Ac and CpTI proteins were detected in transgenic cotton leaves, but no detectable levels of both proteins were present in the mealybug or its predator when reared on transgenic cotton leaves. Our bioassays indicated that transgenic cotton poses a negligible risk to the predatory coccinellid C. montrouzieri via its prey, the mealybug F. virgata.

  1. Evaluation of Cytotoxic and Antimicrobial Effects of Two Bt Cry Proteins on a GMO Safety Perspective

    Directory of Open Access Journals (Sweden)

    Davi Felipe Farias

    2014-01-01

    Full Text Available Studies have contested the innocuousness of Bacillus thuringiensis (Bt Cry proteins to mammalian cells as well as to mammals microbiota. Thus, this study aimed to evaluate the cytotoxic and antimicrobial effects of two Cry proteins, Cry8Ka5 (a novel mutant protein and Cry1Ac (a widely distributed protein in GM crops. Evaluation of cyto- and genotoxicity in human lymphocytes was performed as well as hemolytic activity coupled with cellular membrane topography analysis in mammal erythrocytes. Effects of Cry8Ka5 and Cry1Ac upon Artemia sp. nauplii and upon bacteria and yeast growth were assessed. The toxins caused no significant effects on the viability (IC50>1,000 µg/mL or to the cellular DNA integrity of lymphocytes (no effects at 1,000 µg/mL. The Cry8Ka5 and Cry1Ac proteins did not cause severe damage to erythrocytes, neither with hemolysis (IC50>1,000 µg/mL nor with alterations in the membrane. Likewise, the Cry8Ka5 and Cry1Ac proteins presented high LC50 (755.11 and >1,000 µg/mL, resp. on the brine shrimp lethality assay and showed no growth inhibition of the microorganisms tested (MIC>1,000 µg/mL. This study contributed with valuable information on the effects of Cry8Ka5 and Cry1Ac proteins on nontarget organisms, which reinforce their potential for safe biotechnological applications.

  2. Enhancement of Bacillus thuringiensis insecticidal activity by combining Cry1Ac and bi-functional toxin HWTX-XI from spider.

    Science.gov (United States)

    Sun, Yunjun; Fu, Zujiao; He, Xiaohong; Yuan, Chunhua; Ding, Xuezhi; Xia, Liqiu

    2016-03-01

    In order to assess the potency of bi-functional HWTX-XI toxin from spider Ornithoctonus huwena in improving the insecticidal activity of Bacillus thuringiensis, a fusion gene of cry1Ac and hwtx-XI was constructed and expressed in an acrystalliferous B. thuringiensis strain Cry(-)B. Western blot analysis and microscopic observation revealed that the recombinant strain could express 140-kDa Cry1Ac-HWTX-XI fusion protein and produce parasporal inclusions during sporulation. Bioassay using the larvae of Helicoverpa armigera and Spodoptera exigua showed that the Cry1Ac-HWTX-XI fusion was more toxic than the control Cry1Ac protoxin, as revealed by 95% lethal concentration. Our study indicated that the HWTX-XI from spider might be a candidate for enhancing the toxicity of B. thuringiensis products.

  3. Antagonism between Cry1Ac1 and Cyt1A1 toxins of bacillus thuringiensis

    Science.gov (United States)

    del Rincon-Castro MC; Barajas-Huerta; Ibarra

    1999-05-01

    Most strains of the insecticidal bacterium Bacillus thuringiensis have a combination of different protoxins in their parasporal crystals. Some of the combinations clearly interact synergistically, like the toxins present in B. thuringiensis subsp. israelensis. In this paper we describe a novel joint activity of toxins from different strains of B. thuringiensis. In vitro bioassays in which we used pure, trypsin-activated Cry1Ac1 proteins from B. thuringiensis subsp. kurstaki, Cyt1A1 from B. thuringiensis subsp. israelensis, and Trichoplusia ni BTI-Tn5B1-4 cells revealed contrasting susceptibility characteristics. The 50% lethal concentrations (LC50s) were estimated to be 4,967 of Cry1Ac1 per ml of medium and 11.69 ng of Cyt1A1 per ml of medium. When mixtures of these toxins in different proportions were assayed, eight different LC50s were obtained. All of these LC50s were significantly higher than the expected LC50s of the mixtures. In addition, a series of bioassays were performed with late first-instar larvae of the cabbage looper and pure Cry1Ac1 and Cyt1A1 crystals, as well as two different combinations of the two toxins. The estimated mean LC50 of Cry1Ac1 was 2.46 ng/cm2 of diet, while Cyt1A1 crystals exhibited no toxicity, even at very high concentrations. The estimated mean LC50s of Cry1Ac1 crystals were 15.69 and 19.05 ng per cm2 of diet when these crystals were mixed with 100 and 1,000 ng of Cyt1A1 crystals per cm2 of diet, respectively. These results indicate that there is clear antagonism between the two toxins both in vitro and in vivo. Other joint-action analyses corroborated these results. Although this is the second report of antagonism between B. thuringiensis toxins, our evidence is the first evidence of antagonism between toxins from different subspecies of B. thuringiensis (B. thuringiensis subsp. kurstaki and B. thuringiensis subsp. israelensis) detected both in vivo and in vitro. Some possible explanations for this relationship are discussed.

  4. Cytotoxicity and binding profiles of activated Cry1Ac and Cry2Ab to three insect cell lines

    Science.gov (United States)

    While Cry1Ac has been known to bind with larval midgut proteins cadherin, APN (amino peptidase N), ALP (alkaline phosphatase) and ABCC2 (ATP-binding cassette transporter subfamily C2), little is known about the receptors of Cry2Ab. To provide a clue to the receptors of Cry2Ab, we tested the baselin...

  5. Cis-mediated down-regulation of a trypsin gene associated with Bt resistance in cotton bollworm.

    Science.gov (United States)

    Liu, Chenxi; Xiao, Yutao; Li, Xianchun; Oppert, Brenda; Tabashnik, Bruce E; Wu, Kongming

    2014-11-27

    Transgenic plants producing insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) are useful for pest control, but their efficacy is reduced when pests evolve resistance. Here we examined the mechanism of resistance to Bt toxin Cry1Ac in the laboratory-selected LF5 strain of the cotton bollworm, Helicoverpa armigera. This strain had 110-fold resistance to Cry1Ac protoxin and 39-fold resistance to Cry1Ac activated toxin. Evaluation of five trypsin genes revealed 99% reduced transcription of one trypsin gene (HaTryR) was associated with resistance. Silencing of this gene with RNA interference in susceptible larvae increased their survival on diets containing Cry1Ac. Bioassays of progeny from crosses revealed that resistance to Cry1Ac was genetically linked with HaTryR. We identified mutations in the promoter region of HaTryR in the resistant strain. In transfected insect cell lines, transcription was lower when driven by the resistant promoter compared with the susceptible promoter, implicating cis-mediated down-regulation of HaTryR transcription as a mechanism of resistance. The results suggest that H. armigera can adapt to Bt toxin Cry1Ac by decreased expression of trypsin. Because trypsin activation of protoxin is a critical step in toxicity, transgenic plants with activated toxins rather than protoxins might increase the durability of Bt crops.

  6. The role of β18-β19 loop structure in insecticidal activity of Cry1Ac toxin from Bacillus thuringiensis

    Institute of Scientific and Technical Information of China (English)

    XIA LiQiu; WANG FaXiang; DING XueZhi; ZHAO XinMin; FU ZuJiao; QUAN MeiFang; YU ZiNiu

    2008-01-01

    The β18-β19 loop in domain Ⅲ of Cry1Ac toxin is unique among Bacillus thuringiensis Cry proteins. In this study, the role of the loop structure in insecticidal activity of Cry1Ac toxin was investigated. Alanine scanning mutations within the loop were initially generated and most mutants were over-expressed and reduced toxicity at different degrees, except mutant N546A that showed almost 2 times enhanced toxicity against Helicoverpa armigera larvae. Further mutagenic analysis of N546 re-vealed that a charged amino acid in this position would cause very unfavorable influence on insecti-cidal activity. In addition, the deletion of N546 led to protein instability because of destruction of the loop integrity. Besides, mutant W544F was much more toxic than W544Y, indicating that hydrophobic nature of the position was important for maintaining the stability and activity of Cry1Ac protein. These findings are the first biological evidence for a structural function of β18-β19 loop in insecticidal activity of the Cry1Ac toxin.

  7. Evaluation of Bt Corn with Pyramided Genes on Efficacy and Insect Resistance Management for the Asian Corn Borer in China

    Science.gov (United States)

    Bai, Shuxiong; Wang, Zhenying; He, Kanglai

    2016-01-01

    A Bt corn hybrid (AcIe) with two Bt genes (cry1Ie and cry1Ac) was derived by breeding stack from line expressing Cry1Ie and a line expressing Cry1Ac. Efficacy of this pyramided Bt corn hybrid against the Asian corn borer (ACB), Ostrinia furnacalis, was evaluated. We conducted laboratory bioassays using susceptible and resistant ACB strains fed on artificial diet or fresh plant tissues. We also conducted field trials with artificial infestations of ACB neonates at the V6 and silk stages. The toxin-diet bioassay data indicated that mixtures of Cry1Ac and Cry1Ie proteins had synergistic insecticidal efficacy. The plant tissue bioassay data indicated that Bt corn hybrids expressing either a single toxin (Cry1Ac or Cry1Ie) or two toxins had high efficacy against susceptible ACB. Damage ratings in the field trials indicated that the Bt corn hybrids could effectively protect against 1st and the 2nd generation ACB in China. The hybrid line with two Bt genes showed a higher efficacy against ACB larvae resistant to Cry1Ac or CryIe than the hybrid containing one Bt gene, and the two gene hybrid would have increased potential for managing or delaying the evolution of ACB resistance to Bt corn plants. PMID:28006032

  8. 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,

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

    OpenAIRE

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

    2013-01-01

    First generation of insect-protected transgenic corn (Bt-corn) was based on the expression of Cry1Ab or Cry1Fa proteins. Currently, the trend is the combination of two or more genes expressing proteins that bind to different targets. In addition to broadening the spectrum of action, this strategy helps to delay the evolution of resistance in exposed insect populations. One of such examples is the combination of Cry1A.105 with Cry1Fa and Cry2Ab to control O. nubilalis and S. frugiperda. Cry1A....

  10. Non-recessive Bt toxin resistance conferred by an intracellular cadherin mutation in field-selected populations of cotton bollworm.

    Science.gov (United States)

    Zhang, Haonan; Wu, Shuwen; Yang, Yihua; Tabashnik, Bruce E; Wu, Yidong

    2012-01-01

    Transgenic crops producing Bacillus thuringiensis (Bt) toxins have been planted widely to control insect pests, yet evolution of resistance by the pests can reduce the benefits of this approach. Recessive mutations in the extracellular domain of toxin-binding cadherin proteins that confer resistance to Bt toxin Cry1Ac by disrupting toxin binding have been reported previously in three major lepidopteran pests, including the cotton bollworm, Helicoverpa armigera. Here we report a novel allele from cotton bollworm with a deletion in the intracellular domain of cadherin that is genetically linked with non-recessive resistance to Cry1Ac. We discovered this allele in each of three field-selected populations we screened from northern China where Bt cotton producing Cry1Ac has been grown intensively. We expressed four types of cadherin alleles in heterologous cell cultures: susceptible, resistant with the intracellular domain mutation, and two complementary chimeric alleles with and without the mutation. Cells transfected with each of the four cadherin alleles bound Cry1Ac and were killed by Cry1Ac. However, relative to cells transfected with either the susceptible allele or the chimeric allele lacking the intracellular domain mutation, cells transfected with the resistant allele or the chimeric allele containing the intracellular domain mutation were less susceptible to Cry1Ac. These results suggest that the intracellular domain of cadherin is involved in post-binding events that affect toxicity of Cry1Ac. This evidence is consistent with the vital role of the intracellular region of cadherin proposed by the cell signaling model of the mode of action of Bt toxins. Considered together with previously reported data, the results suggest that both pore formation and cell signaling pathways contribute to the efficacy of Bt toxins.

  11. Non-recessive Bt toxin resistance conferred by an intracellular cadherin mutation in field-selected populations of cotton bollworm.

    Directory of Open Access Journals (Sweden)

    Haonan Zhang

    Full Text Available Transgenic crops producing Bacillus thuringiensis (Bt toxins have been planted widely to control insect pests, yet evolution of resistance by the pests can reduce the benefits of this approach. Recessive mutations in the extracellular domain of toxin-binding cadherin proteins that confer resistance to Bt toxin Cry1Ac by disrupting toxin binding have been reported previously in three major lepidopteran pests, including the cotton bollworm, Helicoverpa armigera. Here we report a novel allele from cotton bollworm with a deletion in the intracellular domain of cadherin that is genetically linked with non-recessive resistance to Cry1Ac. We discovered this allele in each of three field-selected populations we screened from northern China where Bt cotton producing Cry1Ac has been grown intensively. We expressed four types of cadherin alleles in heterologous cell cultures: susceptible, resistant with the intracellular domain mutation, and two complementary chimeric alleles with and without the mutation. Cells transfected with each of the four cadherin alleles bound Cry1Ac and were killed by Cry1Ac. However, relative to cells transfected with either the susceptible allele or the chimeric allele lacking the intracellular domain mutation, cells transfected with the resistant allele or the chimeric allele containing the intracellular domain mutation were less susceptible to Cry1Ac. These results suggest that the intracellular domain of cadherin is involved in post-binding events that affect toxicity of Cry1Ac. This evidence is consistent with the vital role of the intracellular region of cadherin proposed by the cell signaling model of the mode of action of Bt toxins. Considered together with previously reported data, the results suggest that both pore formation and cell signaling pathways contribute to the efficacy of Bt toxins.

  12. New resistance mechanism in Helicoverpa armigera threatens transgenic crops expressing Bacillus thuringiensis Cry1Ac toxin.

    Science.gov (United States)

    Gunning, Robin V; Dang, Ho T; Kemp, Fred C; Nicholson, Ian C; Moores, Graham D

    2005-05-01

    In Australia, the cotton bollworm, Helicoverpa armigera, has a long history of resistance to conventional insecticides. Transgenic cotton (expressing the Bacillus thuringiensis toxin Cry1Ac) has been grown for H. armigera control since 1996. It is demonstrated here that a population of Australian H. armigera has developed resistance to Cry1Ac toxin (275-fold). Some 70% of resistant H. armigera larvae were able to survive on Cry1Ac transgenic cotton (Ingard) The resistance phenotype is inherited as an autosomal semidominant trait. Resistance was associated with elevated esterase levels, which cosegregated with resistance. In vitro studies employing surface plasmon resonance technology and other biochemical techniques demonstrated that resistant strain esterase could bind to Cry1Ac protoxin and activated toxin. In vivo studies showed that Cry1Ac-resistant larvae fed Cy1Ac transgenic cotton or Cry1Ac-treated artificial diet had lower esterase activity than non-Cry1Ac-fed larvae. A resistance mechanism in which esterase sequesters Cry1Ac is proposed.

  13. Multi-Toxin Resistance Enables Pink Bollworm Survival on Pyramided Bt Cotton.

    Science.gov (United States)

    Fabrick, Jeffrey A; Unnithan, Gopalan C; Yelich, Alex J; DeGain, Ben; Masson, Luke; Zhang, Jie; Carrière, Yves; Tabashnik, Bruce E

    2015-11-12

    Transgenic crops producing Bacillus thuringiensis (Bt) proteins kill key insect pests, providing economic and environmental benefits. However, the evolution of pest resistance threatens the continued success of such Bt crops. To delay or counter resistance, transgenic plant "pyramids" producing two or more Bt proteins that kill the same pest have been adopted extensively. Field populations of the pink bollworm (Pectinophora gossypiella) in the United States have remained susceptible to Bt toxins Cry1Ac and Cry2Ab, but field-evolved practical resistance to Bt cotton producing Cry1Ac has occurred widely in India. Here we used two rounds of laboratory selection to achieve 18,000- to 150,000-fold resistance to Cry2Ab in pink bollworm. Inheritance of resistance to Cry2Ab was recessive, autosomal, conferred primarily by one locus, and independent of Cry1Ac resistance. We created a strain with high resistance to both toxins by crossing the Cry2Ab-resistant strain with a Cry1Ac-resistant strain, followed by one selection with Cry2Ab. This multi-toxin resistant strain survived on field-collected Bt cotton bolls producing both toxins. The results here demonstrate the risk of evolution of resistance to pyramided Bt plants, particularly when toxins are deployed sequentially and refuges are scarce, as seen with Bt cotton and pink bollworm in India.

  14. The Spodoptera exigua (Lepidoptera: Noctuidae) ABCC2 Mediates Cry1Ac Cytotoxicity and, in Conjunction with Cadherin, Contributes to Enhance Cry1Ca Toxicity in Sf9 Cells.

    Science.gov (United States)

    Ren, Xiang-Liang; Jiang, Wei-Li; Ma, Ya-Jie; Hu, Hong-Yan; Ma, Xiao-Yan; Ma, Yan; Li, Guo-Qing

    2016-12-01

    In insects, the mode of Cry1A toxins action has been studied in detail and many receptors that participate in the process are known. Recent evidence has revealed that an ABC transporter (ABCC2) is involved in conferring resistance to Cry1A toxins and that ABCC2 could be a receptor of Cry1A. However, it is not known whether Cry1Ca interacts with the same receptor proteins as Cry1A. In this study, we report the cloning of an ABC transporter gene, SeABCC2b, from the midgut of Spodoptera exigua (Hübner) larvae, and its expression in Sf9 cells for a functional analysis. The addition of Cry1Ca and Cry1Ac to Sf9 cell culture caused swelling in 28.5% and 93.9% of the SeABCC2-expressing cells, respectively. In contrast, only 7.4% and 1.3% of the controls cells swelled in the presence of Cry1Ca and Cry1Ac. Thus, SeABCC2b-expressing Sf9 cells had increased susceptibility to Cry1Ca and Cry1Ac. Similarly, S. exigua cadherin (SeCad1b) expressed in Sf9 cells caused 47.1% and 1.8% of the SeCad1b-expressing cells to swell to Cry1Ca and Cry1Ac exposure. Therefore, Sf9 cells expressing SeCad1b were more sensitive to Cry1Ca than Cry1Ac. Together, our data suggest that SeABCC2b from S. exigua mediates Cry1Ac cytotoxicity and, in conjunction with SeCad1b, contributes to enhance Cry1Ca toxicity in Sf9 cells.

  15. Alternative splicing and highly variable cadherin transcripts associated with field-evolved resistance of pink bollworm to bt cotton in India.

    Directory of Open Access Journals (Sweden)

    Jeffrey A Fabrick

    Full Text Available Evolution of resistance by insect pests can reduce the benefits of insecticidal proteins from Bacillus thuringiensis (Bt that are used extensively in sprays and transgenic crops. Despite considerable knowledge of the genes conferring insect resistance to Bt toxins in laboratory-selected strains and in field populations exposed to Bt sprays, understanding of the genetic basis of field-evolved resistance to Bt crops remains limited. In particular, previous work has not identified the genes conferring resistance in any cases where field-evolved resistance has reduced the efficacy of a Bt crop. Here we report that mutations in a gene encoding a cadherin protein that binds Bt toxin Cry1Ac are associated with field-evolved resistance of pink bollworm (Pectinophora gossypiella in India to Cry1Ac produced by transgenic cotton. We conducted laboratory bioassays that confirmed previously reported resistance to Cry1Ac in pink bollworm from the state of Gujarat, where Bt cotton producing Cry1Ac has been grown extensively. Analysis of DNA from 436 pink bollworm from seven populations in India detected none of the four cadherin resistance alleles previously reported to be linked with resistance to Cry1Ac in laboratory-selected strains of pink bollworm from Arizona. However, DNA sequencing of pink bollworm derived from resistant and susceptible field populations in India revealed eight novel, severely disrupted cadherin alleles associated with resistance to Cry1Ac. For these eight alleles, analysis of complementary DNA (cDNA revealed a total of 19 transcript isoforms, each containing a premature stop codon, a deletion of at least 99 base pairs, or both. Seven of the eight disrupted alleles each produced two or more different transcript isoforms, which implicates alternative splicing of messenger RNA (mRNA. This represents the first example of alternative splicing associated with field-evolved resistance that reduced the efficacy of a Bt crop.

  16. Alternative splicing and highly variable cadherin transcripts associated with field-evolved resistance of pink bollworm to bt cotton in India.

    Science.gov (United States)

    Fabrick, Jeffrey A; Ponnuraj, Jeyakumar; Singh, Amar; Tanwar, Raj K; Unnithan, Gopalan C; Yelich, Alex J; Li, Xianchun; Carrière, Yves; Tabashnik, Bruce E

    2014-01-01

    Evolution of resistance by insect pests can reduce the benefits of insecticidal proteins from Bacillus thuringiensis (Bt) that are used extensively in sprays and transgenic crops. Despite considerable knowledge of the genes conferring insect resistance to Bt toxins in laboratory-selected strains and in field populations exposed to Bt sprays, understanding of the genetic basis of field-evolved resistance to Bt crops remains limited. In particular, previous work has not identified the genes conferring resistance in any cases where field-evolved resistance has reduced the efficacy of a Bt crop. Here we report that mutations in a gene encoding a cadherin protein that binds Bt toxin Cry1Ac are associated with field-evolved resistance of pink bollworm (Pectinophora gossypiella) in India to Cry1Ac produced by transgenic cotton. We conducted laboratory bioassays that confirmed previously reported resistance to Cry1Ac in pink bollworm from the state of Gujarat, where Bt cotton producing Cry1Ac has been grown extensively. Analysis of DNA from 436 pink bollworm from seven populations in India detected none of the four cadherin resistance alleles previously reported to be linked with resistance to Cry1Ac in laboratory-selected strains of pink bollworm from Arizona. However, DNA sequencing of pink bollworm derived from resistant and susceptible field populations in India revealed eight novel, severely disrupted cadherin alleles associated with resistance to Cry1Ac. For these eight alleles, analysis of complementary DNA (cDNA) revealed a total of 19 transcript isoforms, each containing a premature stop codon, a deletion of at least 99 base pairs, or both. Seven of the eight disrupted alleles each produced two or more different transcript isoforms, which implicates alternative splicing of messenger RNA (mRNA). This represents the first example of alternative splicing associated with field-evolved resistance that reduced the efficacy of a Bt crop.

  17. Elite Indica transgenic rice plants expressing modified Cry1Ac endotoxin of Bacillus thuringiensis show enhanced resistance to yellow stem borer (Scirpophaga incertulas).

    Science.gov (United States)

    Khanna, H K; Raina, S K

    2002-08-01

    Bt-transgenics of elite indica rice breeding lines (IR-64, Pusa Basmati-1 and Karnal Local) were generated through biolistic or Agrobacterium-mediated approaches. A synthetic cry1Ac gene, codon optimised for rice and driven by the maize ubiquitin-1 promoter, was used. Over 200 putative transformants of IR-64 and Pusa Basmati-1 and 26 of the Karnal Local were regenerated following use of the hpt (hygromycin phosphotransferase) selection system. Initial transformation frequency was in the range of 1 to 2% for particle bombardment while it was comparatively higher (approximately 9%) for Agrobacterium. An improved selection procedure, involving longer selection on the antibiotic-supplemented medium, enhanced the frequency of Bt-transformants and reduced the number of escapes. Molecular evaluation revealed multiple transgene insertions in transformants, whether generated through biolistic or Agrobacterium. In the latter case, it was also observed that all genes on the T-DNA do not necessarily get transferred as an intact insert. Selected Bt-lines of IR-64 and Pusa Basmati-1, having Bt-titers of 0.1% (of total soluble protein) and above were evaluated for resistance against manual infestation of freshly hatched neonate larvae of yellow stem borers collected from a hot spot stem borer infested area in northern India. Several Bt-lines were identified showing 100% mortality of larvae, within 4-days of infestation, in cut-stem as well as vegetative stage whole plant assays. However, there was an occasional white head even among such plants when assayed at the reproductive stage. Results are discussed in the light of resistance management strategies for deployment of Bt-rice.

  18. The protoxin Cry1Ac of Bacillus thuringiensis improves the protection conferred by intranasal immunization with Brucella abortus RB51 in a mouse model.

    Science.gov (United States)

    González-González, Edith; García-Hernández, Ana Lilia; Flores-Mejía, Raúl; López-Santiago, Rubén; Moreno-Fierros, Leticia

    2015-02-25

    Brucellosis is a zoonotic disease affecting many people and animals worldwide. Preventing this infection requires improving vaccination strategies. The protoxin Cry1Ac of Bacillus thuringiensis is an adjuvant that, in addition to increasing the immunogenicity of different antigens, has shown to be protective in different models of parasitic infections. The objective of the present study was to test whether the intranasal co-administration of pCry1Ac with the RB51 vaccine strain of Brucella abortus confers protection against an intranasal challenge with the virulent strain B. abortus 2308 in BALB/c mice. The results showed that co-administration of pCry1Ac and RB51, increased the immunoprotection conferred by the vaccine as evidenced by the following: (1) decrease of the splenic bacterial load when challenged intranasally with the virulent strain; (2) greater in vivo cytotoxic activity in response to the transference of previously infected cells; (3) further proliferation of cytotoxic TCD8+ cells in response to stimulation with heat-inactivated bacteria; (4) increased production of TNF-α and IFN-γ; and (5) significant IgG2a response. These results indicate that the use of the Cry1Ac protein as a mucosal adjuvant via the intranasal route can be a promising alternative for improving current RB51 vaccine against brucellosis.

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

    Science.gov (United States)

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

    2014-01-01

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

  20. Effects of antiserums of cadherin, aminopeptidase N and alkaline phosphatase on the toxicities of Cry1Ac and Cry2Aa in Helicoverpa armigera (Lepidoptera: Noctuidae)%棉铃虫中肠钙粘蛋白、氨肽酶N及碱性磷酸酯酶的抗血清对Cry1Ac和Cry2Aa毒力的影响

    Institute of Scientific and Technical Information of China (English)

    王冰洁; 袁向东; 赵曼; 刘臣; 陈琳; 梁革梅

    2016-01-01

    [Aim] Cry1A and Cry2A toxins play insecticidal roles by specifically binding with receptor proteins on insect midgut,and are widely used in Bt transgenic crops now.This study aims to further understand the action mechanisms of Cry2A and the roles of the functional receptor proteins of Cry1 A in the toxicity of Cry2A.[Methods] We firstly extracted brush border membrane vesicles (BBMV) of midgut,prepared antibody and antiserums of cadherin (CAD),aminopeptidase N (APN),and alkaline phosphatase (ALP) in Helicoverpa armigera.Then,after detecting the antiserums of these three receptor proteins in BBMV by Western blot,we compared the effects of antiserums of these three receptor proteins on the toxicities of Cry1Ac and Cry2Aa in the susceptible and Cry1Ac-resistant H.armigera (BtR) by the antibody blocking technology.[Results] For the susceptible H.armigera,the antiserums of these three receptor proteins not only significantly reduced the toxicity of Cry1 Ac,but also remarkably reduced the toxicity of Cry2Aa.Among them,anti-APN antiserum had the biggest impact on the toxicity of Cry1Ac,causing the mortality of H.armigera larvae to be reduced by 84.44%,and anti-ALP antiserum had the greatest effect on the toxicity of Cry2Aa,causing the larval mortality to be reduced by 71.04% compared with the control (without treatment by antiserum).The toxicity of Cry1Ac to Cry1Ac-resistant H.armigera (BtR) was obviously reduced and the toxicity of Cry2Aa also became less to these resistant larvae.The influences of the antiserums of these three receptors on the toxicity of Cry1Ac to the Cry1Acresistant strain of H.armigera (BtR) were smaller than those to the susceptible strain.Especially,the inhibition percentage of anti-CAD and-APN antiserums to the toxicity of Cry1 Ac decreased significantly.The effects of anti-CAD and-ALP antiserums on the toxicity of Cry2Aa to the Cry1Ac-resistant strain (BtR) and the susceptible strain were not significantly different,but the anti-APN antiserum

  1. An Improved Method for Purifying Cry1Ac Protein from Bacillus thuringiensis%一种纯化苏云金杆菌Cry1Ac蛋白的方法

    Institute of Scientific and Technical Information of China (English)

    王发祥; 刘永乐; 丁学知; 夏立秋

    2011-01-01

    The research of structure and function of Cry proteins was greatly restricted by their purification method. Therefore, a method that purifying Bacillus thuringiensis CrylAc protoxin by using isoelectric precipitation and purifying Bacillus thuringiensis CrylAc activated toxin by using molecular sieve chromatography was proposed. The results showed that the CrylAc protoxin had good effects of precipitation and purification when Ph value was 5.05; activating the purified protoxin to conduct molecular sieve chromatography, high concentration of CrylAc activated toxin could be obtained from the maximum peak of chromatography. This improved method not only has good purification effect, but also contribute a renovation to the purification method of Cry protein, which is significant for promoting researches on structure and function of Cry protein.%Cry蛋白的纯化技术在很大程度上制约了其结构与功能的研究.为此,提出了一种以等电点沉淀方法纯化苏云金杆菌Cry1Ac的原毒素,用分子筛层析纯化其活性毒素的方法.结果表明:Cry1Ac的原毒素在pH值5.05条件下沉淀纯化效果好;以纯化的原毒素激活进行分子筛层析,收集其最大峰便可获得较高浓度的活性毒素;该方法不仅纯化效果好,而且省去层析后的浓缩步骤,是Cry蛋白的纯化方法研究中的一次革新,对推动Cry蛋白的结构和功能研究具有重要的意义.

  2. Effect of silencing midgut cadherin gene on susceptibility of Plutella xylostella larvae to Cry1Ac toxin%沉默cadherin基因对小菜蛾敏感性的影响

    Institute of Scientific and Technical Information of China (English)

    杨中侠; 杨柳; 谭济才; 吴青君; 王少丽; 徐宝云; 张友军

    2013-01-01

    钙粘蛋白(cadherin)是许多鳞翅目昆虫中苏云金芽孢杆菌(Bt)毒素的受体.利用RNAi技术对小菜蛾中肠cadherin基因进行沉默处理,明确了cadherin基因沉默对小菜蛾对Cry1 Ac毒素敏感性的影响.结果表明,注射目标dsRNA后的第1天至第5天,Cry1 Ac敏感及抗性小菜蛾品系cadherin的基因表达量均显著减少,尤以第2天的减少量最为明显;Cry1 Ac毒力测定结果表明,沉默cadherin基因能在一定程度上提高幼虫的成活率,即可使小菜蛾对Cry1 Ac毒素的敏感性降低.研究初步表明,cadherin基因的功能与小菜蛾对Cry1Ac毒素的抗性有关.%Cadherin was confirmed to be the receptor of Bacillus thuringiensis (Bt) insecticidal protein in several Lepidoptera insects.Cadherin gene in Plutella xylostella was silenced by RNAi technique and effect of cadherin gene silencing on susceptibility of P.xylostella larvae to Cry1Ac toxin was investigated.The results showed that the expression level of cadherin gene reduced from the first day to the fifth day after the dsRNA corresponding to cadherin segment was injected,and the most significant reduction was observed sharply at the second day.The susceptibility of resistant and susceptible strain of P.xylostella larvae to Cry1Ac toxin decreased after cadherin gene being silenced.The results suggested that cadherin gene was related to resistance of P.xylostella to Cry1Ac toxin.

  3. Transformation and evaluation of Cry1Ac+Cry2A and GTGene in Gossypium hirsutum L.

    Directory of Open Access Journals (Sweden)

    Agung Nugroho Puspito

    2015-11-01

    Full Text Available More than 50 countries around the globe cultivate cotton on a large scale. It is a major cash crop of Pakistan and is considered white gold because it is highly important to the economy of Pakistan. In addition to its importance, cotton cultivation faces several problems, such as insect pests, weeds, and viruses. In the past, insects have been controlled by insecticides, but this method caused a severe loss to the economy. However, conventional breeding methods have provided considerable breakthroughs in the improvement of cotton, but it also has several limitations. In comparison with conventional methods, biotechnology has the potential to create genetically modified plants that are environmentally safe and economically viable. In this study, a local cotton variety VH 289 was transformed with two Bt genes (Cry1Ac and Cry2A and a herbicide resistant gene (cp4 EPSPS using the Agrobacterium mediated transformation method. The constitutive CaMV 35S promoter was attached to the genes taken from Bacillus thuringiensis (Bt and to an herbicide resistant gene during cloning, and this promoter was used for the expression of the genes in cotton plants. This construct was used to develop the Glyphosate Tolerance Gene (GTGene for herbicide tolerance and insecticidal gene (Cry1Ac and Cry2A for insect tolerance in the cotton variety VH 289. The transgenic cotton variety performed 85% better compared with the non-transgenic variety. The study results suggest that farmers should use the transgenic cotton variety for general cultivation to improve the production of cotton.

  4. Differential proteomic analysis of Bt Cry1Ac toxin-resistant and susceptible BTI-Tn-5B1-4 cells%Bt Cry1Ac毒素筛选的粉纹夜蛾离体抗性细胞与敏感细胞蛋白质组的差异分析

    Institute of Scientific and Technical Information of China (English)

    蒋才富; 刘凯于; 彭蓉; 郑进; 彭建新; 洪华珠

    2004-01-01

    采用双向电泳技术和肽质量指纹图谱技术分析了Bt Cry1Ac毒素筛选的粉纹夜蛾Trichoplusia ni抗性BTI-Tn-5B1-4细胞与同源敏感细胞的蛋白质组的差异.用Melanie ViewerⅡ软件在抗性细胞的双向电泳图谱上检测到的平均蛋白质点数为707±25个(n=3),在敏感细胞的双向电泳图谱上检测到的平均蛋白质点数为637±19个(n=3),其中分辨率高、重复性良好的显著差异点有10余个.对其中的一个敏感细胞特有的显著差异斑点进行了肽质量指纹图谱分析,经数据库查询表明该蛋白质与胞质外周蛋白具有同源性.

  5. 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

    Directory of Open Access Journals (Sweden)

    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.

  6. Effect of crop plants on fitness costs associated with resistance to Bacillus thuringiensis toxins Cry1Ac and Cry2Ab in cabbage loopers

    Science.gov (United States)

    Wang, Ran; Tetreau, Guillaume; Wang, Ping

    2016-01-01

    Fitness costs associated with resistance to Bacillus thuringiensis (Bt) toxins critically impact the development of resistance in insect populations. In this study, the fitness costs in Trichoplusia ni strains associated with two genetically independent resistance mechanisms to Bt toxins Cry1Ac and Cry2Ab, individually and in combination, on four crop plants (cabbage, cotton, tobacco and tomato) were analyzed, in comparison with their near-isogenic susceptible strain. The net reproductive rate (R0) and intrinsic rate of increase (r) of the T. ni strains, regardless of their resistance traits, were strongly affected by the host plants. The ABCC2 gene-linked mechanism of Cry1Ac resistance was associated with relatively low fitness costs, while the Cry2Ab resistance mechanism was associated with higher fitness costs. The fitness costs in the presence of both resistance mechanisms in T. ni appeared to be non-additive. The relative fitness of Bt-resistant T. ni depended on the specific resistance mechanisms as well as host plants. In addition to difference in survivorship and fecundity, an asynchrony of adult emergence was observed among T. ni with different resistance mechanisms and on different host plants. Therefore, mechanisms of resistance and host plants available in the field are both important factors affecting development of Bt resistance in insects. PMID:26868936

  7. Effect of crop plants on fitness costs associated with resistance to Bacillus thuringiensis toxins Cry1Ac and Cry2Ab in cabbage loopers.

    Science.gov (United States)

    Wang, Ran; Tetreau, Guillaume; Wang, Ping

    2016-02-12

    Fitness costs associated with resistance to Bacillus thuringiensis (Bt) toxins critically impact the development of resistance in insect populations. In this study, the fitness costs in Trichoplusia ni strains associated with two genetically independent resistance mechanisms to Bt toxins Cry1Ac and Cry2Ab, individually and in combination, on four crop plants (cabbage, cotton, tobacco and tomato) were analyzed, in comparison with their near-isogenic susceptible strain. The net reproductive rate (R0) and intrinsic rate of increase (r) of the T. ni strains, regardless of their resistance traits, were strongly affected by the host plants. The ABCC2 gene-linked mechanism of Cry1Ac resistance was associated with relatively low fitness costs, while the Cry2Ab resistance mechanism was associated with higher fitness costs. The fitness costs in the presence of both resistance mechanisms in T. ni appeared to be non-additive. The relative fitness of Bt-resistant T. ni depended on the specific resistance mechanisms as well as host plants. In addition to difference in survivorship and fecundity, an asynchrony of adult emergence was observed among T. ni with different resistance mechanisms and on different host plants. Therefore, mechanisms of resistance and host plants available in the field are both important factors affecting development of Bt resistance in insects.

  8. Cry1Ac+Cry2Ab基因棉与转Cry1Ac+EPSPS基因棉在黄萎病菌压力环境下检测技术和生存竞争能力研究%The Study on Detection Technology and Survival Competitiveness of Transgenic Cry1Ac+Cry2Ab Cotton and Transgenic Cry1Ac+EPSPS Cotton Under the Pressure of Verticillium dahliae

    Institute of Scientific and Technical Information of China (English)

    乔艳艳; 张兴华; 马艳; 李捷; 杨兆光

    2013-01-01

    旨在研究2种转双价基因棉在棉黄萎病菌压力环境下的生存竞争能力.以‘赣棉杂1号’和‘赣棉11号’为对照,在黄萎病圃检测转Cry1Ac+Cry2Ab基因棉、转Cry1Ac+EPSPS基因棉发病情况、生长和产量竞争能力.2个转双价基因棉抗黄萎病性不显著的强于2个对照;转Cry1Ac+EPSPS基因棉株高、覆盖度增长竞争能力相对较强,转Cry1Ac+ Cry2Ab基因棉相对较弱;转Cry1Ac+EPSPS基因棉的单株成铃和果枝层数较高,增长竞争能力也较强,转Cry1Ac+Cry2Ab基因棉较弱;2个转双价基因棉最后的脱落率与2个对照差异不显著;转Cry1Ac+ EPSPS基因棉籽棉、皮棉产量介于2个对照之间,而转Cry1Ac+Cry2Ab基因棉的产量低于2个对照.转Cry1Ac+Cry2Ab基因棉推广种植可行性相对较差,转Cry1Ac+EPSPS基因棉推广种植可行性一般.

  9. Pretreatment with Cry1Ac Protoxin Modulates the Immune Response, and Increases the Survival of Plasmodium-Infected CBA/Ca Mice

    Directory of Open Access Journals (Sweden)

    Martha Legorreta-Herrera

    2010-01-01

    Full Text Available Malaria is a major global health problem that kills 1-2 million people each year. Despite exhaustive research, naturally acquired immunity is poorly understood. Cry1A proteins are potent immunogens with adjuvant properties and are able to induce strong cellular and humoral responses. In fact, it has been shown that administration of Cry1Ac protoxin alone or with amoebic lysates induces protection against the lethal infection caused by the protozoa Naegleria fowleri. In this work, we studied whether Cry1Ac is able to activate the innate immune response to induce protection against Plasmodium berghei ANKA (lethal and P. chabaudi AS (nonlethal parasites in CBA/Ca mice. Treatment with Cry1Ac induced protection against both Plasmodium species in terms of reduced parasitaemia, longer survival time, modulation of pro- and anti-inflammatory cytokines, and increased levels of specific antibodies against Plasmodium. Understanding how to boost innate immunity to Plasmodium infection should lead to immunologically based intervention strategies.

  10. Biosafety assessment of transgenic Bt cotton on model animals

    Directory of Open Access Journals (Sweden)

    Sadia Bano

    2016-05-01

    Full Text Available Abstract Background: To know the effects of transgenic crops on soil microorganisms, animals and other expected hazards due to the introduction of GM crops into the environment is critical both scientifically and environmentally. The work was conducted to study the effect of insecticidal Bt protein on Rats and Earthworms. Methods: For this purpose, animals like rat and soil organisms like Earthworm were selected. Rats were selected on the basis of its 95% homology on genomic, cellular and enzymatic level with human while earthworm were preferred on the basis of their direct contact with soil to evaluate the impact of Bt (Cry1AC crop field soil on earthworm, secreted by root exudates of Bt cotton. Several physical, molecular, biochemical and histological analyses were performed on both Rats/Earthworms fed on standard diet (control group as well containing Bt protein (experimental group. Results: Molecular analyses such as immune Dot blot, SDS-PAGE, ELISA and PCR, confirmed the absence of Cry1Ac protein in blood and urine samples of rats, which were fed with Bt protein in their diet. Furthermore, histological studies showed that there was no difference in cellular architecture in liver, heart, kidney and intestine of Bt and non-Bt diet fed rats. To see the effect of Bt on earthworm two different groups were studied, one with transgenic plant field soil supplemented with grinded leaves of cotton and second group with non-Bt field soil. Conclusions: No lethal effects of transgenic Bt protein on the survival of earthworm and rats were observed. Bradford assay, Dipstick assay ELISA demonstrated the absence of Cry1Ac protein in the mid-gut epithelial tissue of earthworm. The results of present study will be helpful in successful deployment and commercial release of genetically modified crop in Pakistan.

  11. The development of resistance to Cry1Ac and cross-resistance in an isolated cell strain BTI-TN-5B1 of Trichoplusia ni (Hübner)%粉纹夜蛾离体细胞对Bt Cry1Ac毒素的抗性发展及其交互抗性

    Institute of Scientific and Technical Information of China (English)

    刘凯于; 郑丙莲; 蒋才富; 洪华珠; 彭建新

    2003-01-01

    以Cry1Ac活性毒素逐代筛选抗性粉纹夜蛾离体细胞BTI-TN-5B1,抗性发展速度呈S形曲线,经56代选择后,抗性比上升至1 280倍.在去除选择压力后,抗性比逐渐下降.低水平抗性细胞抗性衰退较快,而高水平抗性细胞抗性衰退较缓慢.抗性比衰退至1.5倍的细胞在复筛后抗性比恢复较快.抗性细胞对苏云金杆菌工程菌GC-91(Bt GC-91)、苏云金杆菌鲇泽亚种(B.t.awazai)和苏云金杆菌库斯塔克亚种(B.t.kurstaki)的复合毒素晶体的活性毒素都具有较高的交互抗性,但对农药无交互抗性,对杆状病毒的敏感性也没发生变化.

  12. 粉纹夜蛾细胞中Bt Cry1Ac选择抗性的研究及离体品系与毒素结合的比较%Comparison of studies on Cry1Ac-binding in resistant and susceptible cells in Trichoplusia ni BTI-Tn-5B1

    Institute of Scientific and Technical Information of China (English)

    刘凯于; 蒋才富; 洪华珠; 彭建新; 余泽华

    2003-01-01

    采用Bt Cry1Ac活性毒素对粉纹夜蛾BTI-Tn-5B1细胞进行56代筛选后获得了抗性比为1280倍的抗性细胞.ELISA检测表明抗性细胞总蛋白和膜蛋白结合的Cry1Ac数量都少于敏感细胞.配体结合Western杂交实验显示:抗性细胞和敏感细胞的膜蛋白与总蛋白都有5条电泳迁移率相同的毒素结合多肽带,其分子量分别为207,158.5,118.8,72,38.5 kD;抗性细胞的118.8和72 kD的阳性带比敏感细胞的略弱,这可能与抗性的形成相关.

  13. Common, but complex, mode of resistance of Plutella xylostella to Bacillus thuringiensis toxins Cry1Ab and Cry1Ac.

    Science.gov (United States)

    Sayyed, Ali H; Gatsi, Roxani; Ibiza-Palacios, M Sales; Escriche, Baltasar; Wright, Denis J; Crickmore, Neil

    2005-11-01

    A field collected population of Plutella xylostella (SERD4) was selected in the laboratory with Bacillus thuringiensis endotoxins Cry1Ac (Cry1Ac-SEL) and Cry1Ab (Cry1Ab-SEL). Both subpopulations showed similar phenotypes: high resistance to the Cry1A toxins and little cross-resistance to Cry1Ca or Cry1D. A previous analysis of the Cry1Ac-SEL showed incompletely dominant resistance to Cry1Ac with more than one factor, at least one of which was sex influenced. In the present study reciprocal mass crosses between Cry1Ab-SEL and a laboratory susceptible population (ROTH) provided evidence that Cry1Ab resistance was also inherited as incompletely dominant trait with more than one factor, and at least one of the factors was sex influenced. Analysis of single pair mating indicated that Cry1Ab-SEL was still heterogeneous for Cry1Ab resistance genes, showing genes with different degrees of dominance. Binding studies showed a large reduction of specific binding of Cry1Ab and Cry1Ac to midgut membrane vesicles of the Cry1Ab-SEL subpopulation. Cry1Ab-SEL was found to be more susceptible to trypsin-activated Cry1Ab toxin than protoxin, although no defect in toxin activation was found. Present and previous results indicate a common basis of resistance to both Cry1Ab and Cry1Ac in selected subpopulations and suggest that a similar set of resistance genes are responsible for resistance to Cry1Ab and Cry1Ac and are selected whichever toxin was used. The possibility of an incompletely dominant trait of resistant to these toxins should be taken into account when considering refuge resistance management strategies.

  14. Effects of transgenic cotton with cry1Ac plus cry2Ab on arthropod communities%转cry1Ac+cry2Ab基因棉对棉田节肢动物群落的影响

    Institute of Scientific and Technical Information of China (English)

    雒珺瑜; 崔金杰; 张帅; 陆雪君

    2011-01-01

    The effects of transgenic cotton with cry1Ac + cry2Ab on arthropod communities were studied, with transgenic cotton with crylAc and conventional cotton as the controls. The results showed that there were no significant differences in the structure and composition of arthropod community, pest sub-community and natural enemy communities among the three cotton fields. The numbers of piercing-sucking pests, trunk boring pests, defoliating insects and spiders, ladybirds and predatory natural enemies were decreased in transgenic cotton with cry1Ac+ cry2Ab fields than in transgenic cotton with cry1Ac and conventional cotton fields. The diversity indexes of arthropod community, pest sub-community and natural enemy communities showed no significant difference a-mong the three cotton fields, indicating that there was no significant difference in the community stability of arthropod community, pest sub-community and natural enemy communities among different cotton fields.%以Bt棉和常规棉为对照,研究了转cry1Ac+ cry2Ab棉对棉田节肢动物群落的影响.研究结果表明,转cry1Ac+ cry2Ab棉田、转crylAc棉田和常规棉田节肢动物群落、害虫亚群落和天敌亚群落的结构与组成无明显差异;转cry1Ac+ cry2Ab棉田刺吸类害虫、蛀食类害虫和食叶类害的数量均低于转cry1Ac棉田和常规棉田;转cry1Ac +cry2Ab棉田蜘蛛类、瓢虫类和捕食类天敌的数量均低于转cry1Ac棉田和常规棉田;转cry1Ac+ cry2Ab棉田节肢动物群落、害虫亚群落和天敌亚群落的多样性指数与转cry1Ac棉田和常规棉田相比均无明显差异,说明转cry1Ac+ cry2Ab棉田节肢动物群落、害虫亚群落和天敌亚群落的稳定性和转cry1Ac棉和常规棉相比均无明显差异.

  15. Frequency of alleles conferring resistance to the Bacillus thuringiensis toxins Cry1Ac and Cry2Ab in Australian populations of Helicoverpa punctigera (Lepidoptera: Noctuidae) from 2002 to 2006.

    Science.gov (United States)

    Downes, S; Parker, T L; Mahon, R J

    2009-04-01

    Helicoverpa punctigera and Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) are important pests of field and horticultural crops in Australia. The former is endemic to the continent, whereas the latter is also distributed in Africa and Asia. Although H. armigera rapidly developed resistance to virtually every group of insecticide used against it, there is only one report of resistance to an insecticide in H. punctigera. In 1996 the Australian cotton industry adopted Ingard, which expresses the Bacillus thuringiensis (Bt) toxin gene cry1Ac. In 2004/2005, Bollgard II (which expresses Cry1Ac and Cry2Ab) replaced Ingard and has subsequently been grown on 80% of the area planted to cotton, Gossypium hirsutum L. From 2002/2003 to 2006/2007, F2 screens were used to detect resistance to Cry1Ac or Cry2Ab. We detected no alleles conferring resistance to Cry1Ac; the frequency was < 0.0005 (n = 2,180 alleles), with a 95% credibility interval between 0 and 0.0014. However, during the same period, we detected alleles that confer resistance to Cry2Ab at a frequency of 0.0018 (n = 2,192 alleles), with a 95% credibility interval between 0.0005 and 0.0040. For both toxins, the experiment-wise detection probability was 94%, i.e., if there actually was a resistance allele in any tested lines, we would have detected it 94% of the time. The first isolation of Cry2Ab resistance in H. punctigera was before the widespread deployment of Bollgard II. This finding supports our published notion for H. armigera that alleles conferring resistance to Cry2Ab may be present at detectable frequencies in populations before selection by transgenic crops.

  16. Mis-splicing of the ABCC2 gene linked with Bt toxin resistance in Helicoverpa armigera.

    Science.gov (United States)

    Xiao, Yutao; Zhang, Tao; Liu, Chenxi; Heckel, David G; Li, Xianchun; Tabashnik, Bruce E; Wu, Kongming

    2014-08-26

    Toxins from the bacterium Bacillus thuringiensis (Bt) are used widely for insect control in sprays and transgenic plants, but their efficacy is reduced when pests evolve resistance. Previous work showed that mutations in a gene encoding the transporter protein ABCC2 are linked with resistance to Bt toxins Cry1Ab, Cry1Ac or both in four species of Lepidoptera. Here we compared the ABCC2 gene of Helicoverpa armigera (HaABCC2) between susceptible strains and a laboratory-selected strain with >1,000-fold resistance to Cry1Ac relative its susceptible parent strain. We discovered a 73-base pair (bp) insertion in the cDNA of the resistant strain that generates a premature stop codon expected to yield a truncated ABCC2 protein. Sequencing of genomic DNA revealed that this insertion is an intron that is not spliced out because of a 6-bp deletion at its splicing site. Analysis of progeny from crosses revealed tight genetic linkage between HaABCC2 and resistance to Cry1Ac. These results provide the first evidence that mis-splicing of a gene encoding an ABCC2 protein confers resistance to a Bt toxin.

  17. Nutrition affects insect susceptibility to Bt toxins

    Science.gov (United States)

    Deans, Carrie A.; Behmer, Spencer T.; Tessnow, Ashley E.; Tamez-Guerra, Patricia; Pusztai-Carey, Marianne; Sword, Gregory A.

    2017-01-01

    Pesticide resistance represents a major challenge to global food production. The spread of resistance alleles is the primary explanation for observations of reduced pesticide efficacy over time, but the potential for gene-by-environment interactions (plasticity) to mediate susceptibility has largely been overlooked. Here we show that nutrition is an environmental factor that affects susceptibility to Bt toxins. Protein and carbohydrates are two key macronutrients for insect herbivores, and the polyphagous pest Helicoverpa zea self-selects and performs best on diets that are protein-biased relative to carbohydrates. Despite this, most Bt bioassays employ carbohydrate-biased rearing diets. This study explored the effect of diet protein-carbohydrate content on H. zea susceptibility to Cry1Ac, a common Bt endotoxin. We detected a 100-fold increase in LC50 for larvae on optimal versus carbohydrate-biased diets, and significant diet-mediated variation in survival and performance when challenged with Cry1Ac. Our results suggest that Bt resistance bioassays that use ecologically- and physiologically-mismatched diets over-estimate susceptibility and under-estimate resistance. PMID:28045087

  18. Nutrition affects insect susceptibility to Bt toxins

    Science.gov (United States)

    Deans, Carrie A.; Behmer, Spencer T.; Tessnow, Ashley E.; Tamez-Guerra, Patricia; Pusztai-Carey, Marianne; Sword, Gregory A.

    2017-01-01

    Pesticide resistance represents a major challenge to global food production. The spread of resistance alleles is the primary explanation for observations of reduced pesticide efficacy over time, but the potential for gene-by-environment interactions (plasticity) to mediate susceptibility has largely been overlooked. Here we show that nutrition is an environmental factor that affects susceptibility to Bt toxins. Protein and carbohydrates are two key macronutrients for insect herbivores, and the polyphagous pest Helicoverpa zea self-selects and performs best on diets that are protein-biased relative to carbohydrates. Despite this, most Bt bioassays employ carbohydrate-biased rearing diets. This study explored the effect of diet protein-carbohydrate content on H. zea susceptibility to Cry1Ac, a common Bt endotoxin. We detected a 100-fold increase in LC50 for larvae on optimal versus carbohydrate-biased diets, and significant diet-mediated variation in survival and performance when challenged with Cry1Ac. Our results suggest that Bt resistance bioassays that use ecologically- and physiologically-mismatched diets over-estimate susceptibility and under-estimate resistance.

  19. Susceptibility of legume pod borer (LPB), Maruca vitrata to delta-endotoxins of Bacillus thuringiensis (Bt) in Taiwan.

    Science.gov (United States)

    Srinivasan, R

    2008-01-01

    Baseline susceptibility of legume pod borer (LPB) to the insecticidal crystal proteins (ICPs) from Bacillus thuringiensis, viz, Cry1Aa, Cry1Ab, Cry1Ac, Cry1Ca and Cry2Aa was assessed in Taiwan. Insect bioassays were performed by incorporating the Bt delta-endotoxins into the LPB artificial diet. The efficacy of different Bt delta-endotoxins against second instar larvae of LPB showed that the toxin Cry1Ab was the most potent toxin (LC(50) 0.207ppm), followed by Cry1Ca, Cry1Aa, Cry2Aa and Cry1Ac in descending order, with LC(50)s 0.477ppm, 0.812ppm, 1.058ppm and 1.666ppm, respectively. Hence, Cry1Ab and/or Cry1Ca toxins would provide effective control of early larval stages of LPB.

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

    Science.gov (United States)

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

    2016-04-01

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

  1. Cry1Ac Transgenic Sugarcane Does Not Affect the Diversity of Microbial Communities and Has No Significant Effect on Enzyme Activities in Rhizosphere Soil within One Crop Season

    Directory of Open Access Journals (Sweden)

    Dinggang eZhou

    2016-03-01

    Full Text Available Cry1Ac transgenic sugarcane provides a promising way to control stem-borer pests. Biosafety assessment of soil ecosystem for cry1Ac transgenic sugarcane is urgently needed because of the important role of soil microorganisms in nutrient transformations and element cycling, however little is known. This study aimed to explore the potential impact of cry1Ac transgenic sugarcane on rhizosphere soil enzyme activities and microbial community diversity, and also to investigate whether the gene flow occurs through horizontal gene transfer. We found no horizontal gene flow from cry1Ac sugarcane to soil. No significant difference in the population of culturable microorganisms between the non-GM and cry1Ac transgenic sugarcane was observed, and there were no significant interactions between the sugarcane lines and the growth stages. A relatively consistent trend at community-level, represented by the functional diversity index, was found between the cry1Ac sugarcane and the non-transgenic lines. Most soil samples showed no significant difference in the activities of four soil enzymes: urease, protease, sucrose, and acid phosphate monoester between the non-transgenic and cry1Ac sugarcane lines. We conclude, based on one crop season, that the cry1Ac sugarcane lines may not affect the microbial community structure and functional diversity of the rhizosphere soil and have few negative effects on soil enzymes.

  2. Susceptibility of field populations of sugarcane borer from non-Bt and Bt maize plants to five individual Cry toxins

    Institute of Scientific and Technical Information of China (English)

    Fangneng Huang; Mukti N.Ghimire; B.Rogers Leonard; Yu-Cheng Zhu; Graham P.Head

    2012-01-01

    Sugarcane borer,Diatraea saccharalis (F.),is a major target of transgenic maize expressing Bacillus thuringiensis (Bt) proteins in South America and the US midsouth region.Resistance development in targct pest populations is a major threat to the sustainable use of Bt crops.In our field trials in 2009,a significant number of live borers and plant injury from D.saccharalis were observed in an experimental SmartStaxTM maize line.The objective of this study was to assess the relative susceptibility of two field populations ofD.saccharalis collected from non-Bt and Bt maize plants containing SmartStaxTM traits to five individual Cry proteins.The five Bt proteins included two proteins (Cry1A.105 and Cry2Ab2) that were expressed in SmartStaxTM maize plants and three other common Bt proteins (Cry1Aa,Cry1Ab and Cry1Ac) that were not produced in SmartStaxTM.Larval mortality and growth inhibition on Bt diet of the fourth gcneration after field collections were evaluated 7 days after release of neonates on the diet surface.The laboratory bioassays showed that 50% lethal concentration (LC50) values for Cry1 A.105 and Cry2Ab2 for the population originated from Bt plants were 3.55-and 1.34-fold greater,respectively,than those of the population collected from non-Bt plants.In contrast,relative to the population from non-Bt plants,the LC50 of the population sampled from Bt plants were 3.85-,2.5-and 1.64-fold more sensitive to Cry1Aa,Cry1Ab and Cry1Ac,respectively.The resuits did not provide clear evidence to conclude that the observed field survival of D.saccharalis on Bt plants was associated with increased levels of resistance.

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

    Directory of Open Access Journals (Sweden)

    B.M. Ribeiro

    1998-06-01

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

  4. Bacillus thuringiensis insecticidal crystal proteins affect lifespan and reproductive performance of Helicoverpa armigera and Spodoptera exigua adults.

    Science.gov (United States)

    Zhang, Ying; Ma, Yan; Wan, Pin-Jun; Mu, Li-Li; Li, Guo-Qing

    2013-04-01

    Being delivered as sprays or expressed in plant, Bacillus thuringiensis (Bt) crystalline proteins (Cry toxins) display insecticidal activities against numerous Lepidopteran, Dipteran, and Coleopteran larvae. Comparative study of toxicities of Bt Cry toxins between larvae and adults may afford important new insights into the interactions of the toxins with receptor proteins in host insect, and represent intriguing targets for the control of insect pests. However, the effectiveness of Bt Cry toxins in insect adults has paid less attention. In the present article, the effectiveness of Cry1Ac and Cry1Ca on lifespans and reproductive performance of Helicoverpa armigera (Hübner) and Spodoptera exigua (Hübner) adults were evaluated by in vivo experiments. Considering transgenic plants express modified, truncated versions of cry genes yielding active toxin fragment, we used activated Bt toxins at the concentration of 500, 100, and 20 microg/ml in a 10% sucrose aquous solution. At the highest concentration, Cry1Ac and Cry1Ca shortened 48.1 and 48.9% of H. armigera female lifespan, and 43.5 and 38.5% of S. exigua female lifespan, and they reduced 37.8 and 40.3%, and 50.5 and 47.4% of H. armigera and S. exigua male lifespans respectively. Bt toxins negatively affected copulation. Exposure to 500 microg/ml of Cry1Ac and Cry1Ca greatly reduced 50.0 and 46.8%, and 58.7 and 57.3% spermatophore acceptance by H. armigera and S. exigua females, respectively. Similarly, Cry1Ac and Cry1Ca exposure decreased 40.0 and 50.3%, and 61.3 and 60.0% of spermatophore transfer by H. armigera and S. exigua males, respectively. Moreover, exposure females rather than males to 500 microg/ml of Cry1Ac and Cry1Ca significantly dropped 57.5 and 57.5% of the number of eggs laid by H. armigera, and 35.4 and 45.8% of the number of egg masses deposited by S. exigua. In contrast, both Cry1Ac and Cry1Ca did not negatively influence the egg hatchability. At the middle and the lowest concentrations, however

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

    Science.gov (United States)

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

    2013-06-01

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

  6. Evaluation of two cotton varieties CRSP1 and CRSP2 for genetic transformation efficiency, expression of transgenes Cry1Ac + Cry2A, GT gene and insect mortality

    Directory of Open Access Journals (Sweden)

    Arfan Ali

    2016-03-01

    Full Text Available Expression of the transgene with a desirable character in crop plant is the ultimate goal of transgenic research. Transformation of two Bt genes namely Cry1Ac and Cry2A cloned as separate cassette under 35S promoter in pKHG4 plant expression vector was done by using shoot apex cut method of Agrobacterium. Molecular confirmation of putative transgenic cotton plants for Cry1Ac, Cry2A and GT gene was done through PCR and ELISA. Transformation efficiency of CRSP-1 and CRSP-2 was calculated to be 1.2 and 0.8% for Cry1Ac while 0.9 and 0.6% for Cry2A and 1.5 and 0.7% for GTG respectively. CRSP-1 was found to adopt natural environment (acclimatized earlier than CRSP-2 when exposed to sunlight for one month. Expression of Cry1Ac, Cry2A and GTG was found to be 1.2, 1 and 1.3 ng/μl respectively for CRSP-1 as compared to CRSP-2 where expression was recorded to be 0.9, 0.5 and 0.9 ng/μl respectively. FISH analysis of the transgenic CRSP-1 and CRSP-2 demonstrated the presence of one and two copy numbers respectively. Similarly, the response of CRSP-1 against Glyphosate @1900 ml/acre was far better with almost negligible necrotic spot and efficient growth after spray as compared to CRSP-2 where some plants were found to have necrosis and negative control where the complete decay of plant was observed after seven days of spray assay. Similarly, almost 100% mortality of 2nd instar larvae of Heliothis armigera was recorded after three days in CRSP-1 as compared CRSP-2 where insect mortality was found to be less than 90%. Quantitatively speaking non transgenic plants were found with 23–90% leaf damage by insect, while CRSP-1 was with less than 5% and CRSP-2 with 17%. Taken together CRSP1 was found to have better insect control and weedicide resistance along with its natural ability of genetic modification and can be employed by the valuable farmers for better insect control and simultaneously for better production.

  7. Impact of Bt-cotton on soil microbiological and biochemical attributes

    Directory of Open Access Journals (Sweden)

    Sanaullah Yasin

    2016-10-01

    Full Text Available Transgenic Bt-cotton produces Bt-toxins (Cry proteins which may accumulate and persist in soil due to their binding ability on soil components. In the present study, the potential impacts of Bt- and non-Bt genotypes of cotton on soil microbial activity, substrate use efficiency, viable microbial population counts, and nutrient dynamics were studied. Two transgenic Bt-cotton genotypes (CIM-602 CIM-599 expressing cry1 Ac gene and two non-Bt cotton genotypes (CIM-573 and CIM-591 were used to evaluate their impact on biological and chemical properties of soil across the four locations in Punjab. Field trials were conducted at four locations (Central Cotton Research Institute-Multan, Naseer Pur, Kot Lal Shah, and Cotton Research Station-Bahawalpur of different agro-ecological zones of Punjab. Rhizosphere soil samples were collected by following standard procedure from these selected locations. Results reveled that Bt-cotton had no adverse effect on microbial population (viable counts and enzymatic activity of rhizosphere soil. Bacterial population was more in Bt-cotton rhizosphere than that of non-Bt cotton rhizosphere at all locations. Phosphatase, dehydrogenase, and oxidative metabolism of rhizosphere soil were more in Bt-cotton genotypes compared with non-Bt cotton genotypes. Cation exchange capacity, total nitrogen, extractable phosphorous, extractable potassium, active carbon, Fe and Zn contents were higher in rhizosphere of Bt-cotton genotypes compared with non-Bt cotton genotypes. It can be concluded from present study that the cultivation of Bt-cotton expressing cry1 Ac had apparently no negative effect on metabolic, microbiological activities, and nutrient dynamics of soils. Further work is needed to investigate the potential impacts of Bt-cotton on ecology of soil-dwelling insects and invertebrates before its recommendation for extensive cultivation.

  8. Tradeoff between reproduction and resistance evolution to Bt-toxin in Helicoverpa armigera: regulated by vitellogenin gene expression.

    Science.gov (United States)

    Zhang, W N; Xiao, H J; Liang, G M; Guo, Y Y; Wu, K M

    2014-08-01

    Evolution of resistance to insecticides usually has fitness tradeoffs associated with adaptation to the stress. The basic regulation mechanism of tradeoff between reproduction and resistance evolution to Bacillus thuringiensis (Bt) toxin in the cotton bollworm, Helicoverpa armigera (Ha), based on the vitellogenin (Vg) gene expression was analyzed here. The full-length cDNA of the Vg gene HaVg (JX504706) was cloned and identified. HaVg has 5704 base pairs (bp) with an open reading frame (ORF) of 5265 bp, which encoded 1756 amino acid protein with a predicted molecular mass of 197.28 kDa and a proposed isoelectric point of 8.74. Sequence alignment analysis indicated that the amino acid sequence of HaVg contained all of the conserved domains detected in the Vgs of the other insects and had a high similarity with the Vgs of the Lepidoptera insects, especially Noctuidae. The resistance level to Cry1Ac Bt toxin and relative HaVg mRNA expression levels among the following four groups: Cry1Ac-susceptible strain (96S), Cry1Ac-resistant strain fed on artificial diet with Bt toxin for 135 generations (BtR stands for the Cry1Ac Bt resistance), progeny of the Cry1Ac-resistant strain with a non-Bt-toxin artificial diet for 38 generations (CK1) and the direct descendants of the 135th-generation resistant larvae which were fed on an artificial diet without the Cry1Ac protein (CK2) were analyzed. Compared with the 96S strain, the resistance ratios of the BtR strain, the CK1 strain and the CK2 strain were 2917.15-, 2.15- and 2037.67-fold, respectively. The maximum relative HaVg mRNA expression levels of the BtR strain were approximately 50% less than that of the 96S strain, and the coming of maximum expression was delayed for approximately 4 days. The overall trend of the HaVg mRNA expression levels in the CK1 strain was similar to that in the 96S strain, and the overall trend of the HaVg mRNA expression levels in the CK2 strain was similar to that in the BtR strain. Our results

  9. Performance and cross-crop resistance of Cry1F-maize selected Spodoptera frugiperda on transgenic Bt cotton: implications for resistance management.

    Science.gov (United States)

    Yang, Fei; Kerns, David L; Brown, Sebe; Kurtz, Ryan; Dennehy, Tim; Braxton, Bo; Head, Graham; Huang, Fangneng

    2016-06-15

    Transgenic crops producing Bacillus thuringiensis (Bt) proteins have become a primary tool in pest management. Due to the intensive use of Bt crops, resistance of the fall armyworm, Spodoptera frugiperda, to Cry1F maize has occurred in Puerto Rico, Brazil, and some areas of the southeastern U.S. The sustainability of Bt crops faces a great challenge because the Cry1F-maize resistant S. frugiperda may also infest other Bt crops in multiple cropping ecosystems. Here we examined the survival and plant injury of a S. frugiperda population selected with Cry1F maize on three single-gene and five pyramided Bt cotton products. Larvae of Cry1F-susceptible (SS), -heterozygous (RS), and -resistant (RR) genotypes of S. frugiperda were all susceptible to the pyramided cotton containing Cry1Ac/Cry2Ab, Cry1Ac/Cry1F/Vip3A, Cry1Ab/Cry2Ae, or Cry1Ab/Cry2Ae/Vip3A, and the single-gene Cry2Ae cotton. Pyramided cotton containing Cry1Ac/Cry1F was effective against SS and RS, but not for RR. These findings show that the Cry1F-maize selected S. frugiperda can cause cross-crop resistance to other Bt crops expressing similar insecticidal proteins. Resistance management and pest management programs that utilize diversify mortality factors must be implemented to ensure the sustainability of Bt crops. This is especially important in areas where resistance to single-gene Bt crops is already widespread.

  10. Influence of Dual-Bt Protein Corn on Bollworm, Helicoverpa zea (Boddie), Survivorship on Bollgard II Cotton.

    Science.gov (United States)

    Von Kanel, M B; Gore, J; Catchot, A; Cook, D; Musser, F; Caprio, M

    2016-04-01

    Similar Cry proteins are expressed in both Bt corn, Zea mays L., and cotton, Gossypium hirsutum (L.), commercial production systems. At least one generation of corn earworm, Helicoverpa zea (Boddie), completes development on field corn in the Mid-South before dispersing across the landscape into other crop hosts like cotton. A concern is that Bt corn hybrids may result in selection for H. zea populations with a higher probability of causing damage to Bt cotton. The objective of this study was to determine the susceptibility of H. zea offspring from moths that developed on non-Bt and VT Triple Pro (VT3 PRO) field corn to lyophilized Bollgard II cotton tissue expressing Cry1Ac and Cry2Ab. Offspring of individuals reared on VT3 PRO expressing Cry1A.105 and Cry2Ab had a significantly higher LC50 two out of the three years this study was conducted. Excess larvae were placed on artificial diet and allowed to pupate to determine if there were any inheritable fitness costs associated with parental development on VT3 PRO corn. Offspring resulting from males collected from VT3 PRO had significantly lower pupal weight and longer pupal duration compared with offspring of individuals collected from non-Bt corn. However, offspring from females collected from VT3 PRO were not different from non-Bt offspring. Paternal influence on offspring in insects is not commonly observed, but illustrates the side effects of development on a transgenic plant expressing less than a high dose, 25 times the concentration needed to kill susceptible larvae.

  11. Development of a Triple Gene Cry1Ac-Cry2Ab-EPSPS Construct and Its Expression in Nicotiana benthamiana for Insect Resistance and Herbicide Tolerance in Plants

    Science.gov (United States)

    Naqvi, Rubab Z.; Asif, Muhammad; Saeed, Muhammad; Asad, Shaheen; Khatoon, Asia; Amin, Imran; Mukhtar, Zahid; Bashir, Aftab; Mansoor, Shahid

    2017-01-01

    Insect pest complex, cotton leaf curl disease and weeds pose major threat to crop production worldwide, including Pakistan. To address these problems, in the present study a triple gene construct harboring Cry1Ac, Cry2Ab, and EPSPS cassettes has been developed for plant specifically in cotton transformation against lepidopteron insect-pests and weeds. Nicotiana benthamiana (tobacco) was used as a model system for characterization of this triple gene construct. The construct has been assembled in plant expression vector and transformed in N. benthamiana. In six transgenic tobacco lines the integration of Cry1Ac-Cry2Ab-EPSPS in tobacco genome was checked by PCR, while successful protein expression of all the three genes was confirmed through immunostrip assay. Efficacy of Cry1Ac and Cry2Ab was evaluated through insect bioassay using armyworm (Spodoptera littoralis). These transgenic tobacco plants showed significant insect mortality as compared to control plants during insect bioassay. Three out of six tested transgenic lines L3, L5, and L9 exhibited 100% mortality of armyworm, while three other lines L1, L10, and L7 showed 86, 80, and 40% mortality, respectively. This construct can readily be used with confidence to transform cotton and other crops for the development of insect resistant and herbicide tolerant transgenic plants. The transgenic crop plants developed using this triple gene construct will provide an excellent germplasm resource for the breeders to improve their efficiency in developing stable homozygous lines as all the three genes being in a single T-DNA border will inherit together. PMID:28174591

  12. Monitoring and adaptive resistance management in Australia for Bt-cotton: current status and future challenges.

    Science.gov (United States)

    Downes, Sharon; Mahon, Rod; Olsen, Karen

    2007-07-01

    In the mid-1990 s the Australian Cotton industry adopted an insect-resistant variety of cotton (Ingard) which expresses the Bt toxin Cry1Ac that is specific to a group of insects including the target Helicoverpa armigera. A conservative resistance management plan (RMP), that restricted the area planted to Ingard, was implemented to preserve the efficacy of Cry1Ac until two-gene transgenic cotton was available. In 2004/05 Bollgard II replaced Ingard as the transgenic cotton available in Australia. It improves on Ingard by incorporating an additional insecticidal protein (Cry2Ab). If an appropriate refuge is grown, there is no restriction on the area planted to Bollgard II. In 2004/05 and 2005/06 the Bollgard II acreage represented approximately 80 of the total area planted to cotton in Australia. The sensitivity of field-collected populations of H. armigera to Bt products was assayed before and subsequent to the widespread deployment of Ingard cotton. In 2002 screens against Cry2Ab were developed in preparation for replacement of Ingard with Bollgard II. There have been no reported field failures of Bollgard II due to resistance. However, while alleles that confer resistance to H. armigera in the field are rare for Cry1Ac, they are surprisingly common for Cry2Ab. We present an overview of the current approach adopted in Australia to monitor and adaptively manage resistance to Bt-cotton in field populations of H. armigera and discuss the implications of our findings to date. We also highlight future challenges for resistance management in Australia, many of which extend to other Bt-crop and pest systems.

  13. Qualitative and event-specific real-time PCR detection methods for Bt brinjal event EE-1.

    Science.gov (United States)

    Randhawa, Gurinder Jit; Sharma, Ruchi; Singh, Monika

    2012-01-01

    Bt brinjal event EE-1 with cry1Ac gene, expressing insecticidal protein against fruit and shoot borer, is the first genetically modified food crop in the pipeline for commercialization in India. Qualitative polymerase chain reaction (PCR) along with event-specific conventional as well as real-time PCR methods to characterize the event EE-1 is reported. A multiplex (pentaplex) PCR system simultaneously amplifying cry1Ac transgene, Cauliflower Mosaic Virus (CaMV) 35S promoter, nopaline synthase (nos) terminator, aminoglycoside adenyltransferase (aadA) marker gene, and a taxon-specific beta-fructosidase gene in event EE-1 has been developed. Furthermore, construct-specific PCR, targeting the approximate 1.8 kb region of inserted gene construct comprising the region of CaMV 35S promoter and cry1Ac gene has also been developed. The LOD of developed EE-1 specific conventional PCR assay is 0.01%. The method performance of the reported real-time PCR assay was consistent with the acceptance criteria of Codex Alimentarius Commission ALINORM 10/33/23, with the LOD and LOQ values of 0.05%. The developed detection methods would not only facilitate effective regulatory compliance for identification of genetic traits, risk assessment, management, and postrelease monitoring, but also address consumer concerns and resolution of legal disputes.

  14. Target and nontarget effects of novel "triple-stacked" Bt-transgenic cotton 1: canopy arthropod communities.

    Science.gov (United States)

    Whitehouse, M E A; Wilson, L J; Davies, A P; Cross, D; Goldsmith, P; Thompson, A; Harden, S; Baker, G

    2014-02-01

    Transgenic cotton varieties (Bollgard II) expressing two proteins (Cry1Ac and Cry2Ab) from Bacillus thuringiensis (Bt) have been widely adopted in Australia to control larvae of Helicoverpa. A triple-stacked Bt-transgenic cotton producing Cry1Ac, Cry2Ab, and Vip3A proteins (Genuity Bollgard III) is being developed to reduce the chance that Helicoverpa will develop resistance to the Bt proteins. Before its introduction, nontarget effects on the agro-ecosystem need to be evaluated under field conditions. By using beatsheet and suction sampling methods, we compared the invertebrate communities of unsprayed non-Bt-cotton, Bollgard II, and Bollgard III in five experiments across three sites in Australia. We found significant differences between invertebrate communities of non-Bt and Bt (Bollgard II and Bollgard III) cotton only in experiments where lepidopteran larval abundance was high. In beatsheet samples where lepidopterans were absent (Bt crops), organisms associated with flowers and bolls in Bt-cotton were more abundant. In suction samples, where Lepidoptera were present (i.e., in non-Bt-cotton), organisms associated with damaged plant tissue and frass were more common. Hence in our study, Bt- and non-Bt-cotton communities only differed when sufficient lepidopteran larvae were present to exert both direct and indirect effects on species assemblages. There was no overall significant difference between Bollgard II and III communities, despite the addition of the Vip gene in Bollgard III. Consequently, the use of Bollgard III in Australian cotton provides additional protection against the development of resistance by Helicoverpa to Bt toxins, while having no additional effect on cotton invertebrate communities.

  15. New insight to structure-function relationship of GalNAc mediated primary interaction between insecticidal Cry1Ac toxin and HaALP receptor of Helicoverpa armigera.

    Science.gov (United States)

    Sengupta, Anindita; Sarkar, Anindya; Priya, Prerna; Ghosh Dastidar, Shubhra; Das, Sampa

    2013-01-01

    Over the last few decades Cry1Ac toxin has been widely used in controlling the insect attack due to its high specificity towards target insects. The pore-forming toxin undergoes a complex mechanism in the insect midgut involving sequential interaction with specific glycosylated receptors in which terminal GalNAc molecule plays a vital role. Recent studies on Cry toxins interactions with specific receptors revealed the importance of several amino acid residues in domain III of Cry1Ac, namely Q509, N510, R511, Y513 and W545, serve as potential binding sites that surround the putative GalNAc binding pocket and mediate the toxin-receptor interaction. In the present study, alanine substitution mutations were generated in the Cry1Ac domain III region and functional significance of those key residues was monitored by insect bioassay on Helicoverpa armigera larvae. In addition, ligand blot analysis and SPR binding assay was performed to monitor the binding characteristics of Cry1Ac wild type and mutant toxins towards HaALP receptor isolated from Helicoverpa armigera. Mutagenesis data revealed that, alanine substitutions in R511, Y513 and W545 substantially impacted the relative affinity towards HaALP receptor and toxicity toward target insect. Furthermore, in silico study of GalNAc-mediated interaction also confirmed the important roles of these residues. This structural analysis will provide a detail insight for evaluating and engineering new generation Cry toxins to address the problem of change in insect behavioral patterns.

  16. Effects of 90-Day Feeding of Transgenic Maize BT799 on the Reproductive System in Male Wistar Rats

    Directory of Open Access Journals (Sweden)

    Qian-ying Guo

    2015-12-01

    Full Text Available BT799 is a genetically modified (GM maize plant that expresses the Cry1Ac gene from Bacillus thuringiensis (Bt. The Cry1Ac gene was introduced into maize line Zhen58 to encode the Bt crystal protein and thus produce insect-resistant maize BT799. Expression of Bt protein in planta confers resistance to Lepidopteran pests and corn rootworms. The present study was designed to investigate any potential effects of BT799 on the reproductive system of male rats and evaluate the nutritional value of diets containing BT799 maize grain in a 90-day subchronic rodent feeding study. Male Wistar rats were fed with diets containing BT799 maize flours or made from its near isogenic control (Zhen58 at a concentration of 84.7%, nutritionally equal to the standard AIN-93G diet. Another blank control group of male rats were treated with commercial AIN-93G diet. No significant differences in body weight, hematology and serum chemistry results were observed between rats fed with the diets containing transgenic BT799, Zhen58 and the control in this 13-week feeding study. Results of serum hormone levels, sperm parameters and relative organ/body weights indicated no treatment-related side effects on the reproductive system of male rats. In addition, no diet-related changes were found in necropsy and histopathology examinations. Based on results of the current study, we did not find any differences in the parameters tested in our study of the reproductive system of male rats between BT799 and Zhen58 or the control.

  17. Cross-resistance to toxins used in pyramided Bt crops and resistance to Bt sprays in Helicoverpa zea.

    Science.gov (United States)

    Welch, Kara L; Unnithan, Gopalan C; Degain, Ben A; Wei, Jizhen; Zhang, Jie; Li, Xianchun; Tabashnik, Bruce E; Carrière, Yves

    2015-11-01

    To delay evolution of resistance by insect pests, farmers are rapidly increasing their use of transgenic crops producing two or more Bacillus thuringiensis (Bt) toxins that kill the same pest. A key condition favoring durability of these "pyramided" crops is the absence of cross-resistance between toxins. Here we evaluated cross-resistance in the major lepidopteran pest Helicoverpa zea (Boddie) to Bt toxins used in pyramids. In the laboratory, we selected a strain of this pest with Bt toxin Cry1Ac followed by selection with MVP II, a formulation containing a hybrid protoxin that is identical to Cry1Ac in the active portion of the toxin and 98.5% identical overall. We calculated the resistance ratio as the EC50 (concentration causing mortality or failure to develop beyond the first instar of 50% of larvae) for the laboratory-selected strain divided by the EC50 for its field-derived parent strain that was not selected in the laboratory. The resistance ratio was 20.0-33.9 (mean=27.0) for MVP II, 57.0 for Cry1Ac, 51.3 for Cry1A.105, 22.4 for Cry1Ab, 3.3 for Cry2Ab, 1.8 for Cry1Fa, and 1.6 for Vip3Aa. Resistance ratios were 2.9 for DiPel ES and 2.0 for Agree VG, which are commercial Bt spray formulations containing Cry1Ac, other Bt toxins, and Bt spores. By the conservative criterion of non-overlap of 95% fiducial limits, the EC50 was significantly higher for the selected strain than its parent strain for MVP II, Cry1Ac, Cry1A.105, Cry1Ab, Cry2Ab and DiPel ES. For Cry1Fa, Vip3Aa, and Agree VG, significantly lower susceptibility to a high concentration indicated low cross-resistance. The resistance ratio for toxins other than Cry1Ac was associated with their amino acid sequence similarity to Cry1Ac in domain II. Resistance to Cry1Ac and the observed cross-resistance to other Bt toxins could accelerate evolution of H. zea resistance to currently registered Bt sprays and pyramided Bt crops.

  18. Efficacy of genetically modified Bt toxins against insects with different genetic mechanisms of resistance.

    Science.gov (United States)

    Tabashnik, Bruce E; Huang, Fangneng; Ghimire, Mukti N; Leonard, B Rogers; Siegfried, Blair D; Rangasamy, Murugesan; Yang, Yajun; Wu, Yidong; Gahan, Linda J; Heckel, David G; Bravo, Alejandra; Soberón, Mario

    2011-10-09

    Transgenic crops that produce Bacillus thuringiensis (Bt) toxins are grown widely for pest control, but insect adaptation can reduce their efficacy. The genetically modified Bt toxins Cry1AbMod and Cry1AcMod were designed to counter insect resistance to native Bt toxins Cry1Ab and Cry1Ac. Previous results suggested that the modified toxins would be effective only if resistance was linked with mutations in genes encoding toxin-binding cadherin proteins. Here we report evidence from five major crop pests refuting this hypothesis. Relative to native toxins, the potency of modified toxins was >350-fold higher against resistant strains of Plutella xylostella and Ostrinia nubilalis in which resistance was not linked with cadherin mutations. Conversely, the modified toxins provided little or no advantage against some resistant strains of three other pests with altered cadherin. Independent of the presence of cadherin mutations, the relative potency of the modified toxins was generally higher against the most resistant strains.

  19. 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.

  20. Expression of Cry1Ac gene in entomogenous fungus Beaveria bassiana%苏云金芽孢杆菌晶体杀虫蛋白CrylAc基因在虫生真菌球孢白僵菌中的表达

    Institute of Scientific and Technical Information of China (English)

    刘振邦; 任小芳; 宋文婧; 朱虹; 黄勃; 李增智

    2011-01-01

    Bt delta-endotoxin Cry1Ac gene was cloned by PCR method and ligated with plasmid pbarGPE1 to construct a recombinant plasmid pbarGPEl -Cry1Ac. This recombinant plasmid pbarGPE1-Cry1Ac was then transferred into a wild-type strain of Beauveria bassiana by blastospore transform, resulting in a multicopy trans-formant. The results of bioassay against the Masson's pine caterpillar, Dendrolimus punctatus, showed that LC25 of the transformant was reduced by 5.41 times, LD25 was decreased by 7.1 times, and LT25 shortened by 2.2 days,indicating a substantial virulence enhancement. Compared to spray treatment, the cumulative mortality of feeding treatment and feeding with spray treatment were increased by 55.6% and 63.0%, and LT50 shortened by 4.5 days and 5.3 days. These data indicated that the stomach toxicity of Bt delta-endotoxin gene enhanced the virulence of B. Bassiana greatly. And this is the first expression of Cry 1 Ac gene in the insect fungi.%依据苏云金芽孢杆菌晶体杀虫蛋白CrylAc基因序列设计引物进行PCR扩增,纯化产物与栽体pbarGPEl连接,构建真菌表达载体pBARGPE1-CrylAc,通过芽生孢子转化法转入野生型球孢白僵菌中,获得多拷贝转化子;毒力测定结果表明转化子对马尾松毛虫的LC25比出发株降低5.4倍,LD25减少7.1倍,LT25缩短2.2 d,毒力明显提高;喂食处理和虫体喷雾喂食结合处理与单纯虫体喷雾处理相比,累计致死率分别提高了55.6%和63.0%,致死中时分别缩短了4.5d和5.3 d.结果表明,苏云金芽孢杆菌晶体杀虫蛋白Cry1Ac基因的表达显著提高了球孢白僵菌的毒力.这是CrylAc基因首次在虫生真菌中表达.

  1. New insight to structure-function relationship of GalNAc mediated primary interaction between insecticidal Cry1Ac toxin and HaALP receptor of Helicoverpa armigera.

    Directory of Open Access Journals (Sweden)

    Anindita Sengupta

    Full Text Available Over the last few decades Cry1Ac toxin has been widely used in controlling the insect attack due to its high specificity towards target insects. The pore-forming toxin undergoes a complex mechanism in the insect midgut involving sequential interaction with specific glycosylated receptors in which terminal GalNAc molecule plays a vital role. Recent studies on Cry toxins interactions with specific receptors revealed the importance of several amino acid residues in domain III of Cry1Ac, namely Q509, N510, R511, Y513 and W545, serve as potential binding sites that surround the putative GalNAc binding pocket and mediate the toxin-receptor interaction. In the present study, alanine substitution mutations were generated in the Cry1Ac domain III region and functional significance of those key residues was monitored by insect bioassay on Helicoverpa armigera larvae. In addition, ligand blot analysis and SPR binding assay was performed to monitor the binding characteristics of Cry1Ac wild type and mutant toxins towards HaALP receptor isolated from Helicoverpa armigera. Mutagenesis data revealed that, alanine substitutions in R511, Y513 and W545 substantially impacted the relative affinity towards HaALP receptor and toxicity toward target insect. Furthermore, in silico study of GalNAc-mediated interaction also confirmed the important roles of these residues. This structural analysis will provide a detail insight for evaluating and engineering new generation Cry toxins to address the problem of change in insect behavioral patterns.

  2. Protection against Naegleria fowleri infection in mice immunized with Cry1Ac plus amoebic lysates is dependent on the STAT6 Th2 response.

    Science.gov (United States)

    Carrasco-Yepez, M; Rojas-Hernandez, S; Rodriguez-Monroy, M A; Terrazas, L I; Moreno-Fierros, L

    2010-01-01

    We previously reported that intranasal administration of Cry1Ac protoxin alone or in combination with amoebic lysates increases protection against Naegleria fowleri meningoencephalitis in mice. Those results suggested that both antibody responses and innate immune mechanisms may be participating in the protective effects observed. The present study was aimed to investigate whether the STAT6-induced Th2 immune response is essential for the resistance to N. fowleri infection, conferred by immunization with amoebic lysates plus Cry1Ac. STAT6-deficient (STAT6-/-) and wild-type (STAT6+/+) BALB/c mice were immunized by the intranasal route with a combination of N. fowleri lysates plus Cry1Ac, and subsequently challenged with lethal doses of N. fowleri trophozoites. STAT6+/+ mice displayed 100% protection, while no protection was observed in STAT6-/- mice. Significantly higher titres of Th2-associated IgG1 as well as interleukin-4 (IL-4) were found in STAT6+/+ mice, whereas in STAT6-/- mice significantly more IL-12 and IFN-gamma as well as significantly higher titres of Th1-associated IgG2a were detected. Thus, whereas protected STAT6+/+-immunized mice elicited a Th-2 type inclined immune response that produced predominantly humoral immunity, unprotected STAT6-/- mice exhibited a polarized Th1 type cellular response. These findings suggest that the STAT6-signalling pathway is critical for defence against N. fowleri infection.

  3. Agronomic performance, chromosomal stability and resistance to velvetbean caterpillar of transgenic soybean expressing cry1Ac gene Performance agronômica, estabilidade cromossômica e resistência à lagarta-da-soja em soja transgênica que expressa o gene cry1Ac

    Directory of Open Access Journals (Sweden)

    Milena Schenkel Homrich

    2008-07-01

    Full Text Available The objective of this work was to analyze the agronomic performance and chromosomal stability of transgenic homozygous progenies of soybean [Glycine max (L. Merrill.], and to confirm the resistance of these plants against Anticarsia gemmatalis. Eleven progenies expressing cry1Ac, hpt and gusA genes were evaluated for agronomic characteristics in relation to the nontransformed parent IAS 5 cultivar. Cytogenetical analysis was carried out on transgenic and nontransgenic plants. Two out of the 11 transgenic progenies were also evaluated, in vitro and in vivo, for resistance to A. gemmatalis. Two negative controls were used in resistance bioassays: a transgenic homozygous line, containing only the gusA reporter gene, and nontransgenic 'IAS 5' plants. The presence of cry1Ac transgene affected neither the development nor the yield of plants. Cytogenetical analysis showed that transgenic plants presented normal karyotype. In detached-leaf bioassay, cry1Ac plants exhibited complete efficacy against A. gemmatalis, whereas negative controls were significantly damaged. Whole-plant feeding assay confirmed a very high protection of cry1Ac against velvetbean caterpillar, while nontransgenic 'IAS 5' plants and homozygous gusA line exhibited 56.5 and 71.5% defoliation, respectively. The presence of cry1Ac transgene doesn't affect the majority of agronomic traits (including yield of soybean and grants high protection against A. gemmatalis.O objetivo deste trabalho foi analisar a performance agronômica e a estabilidade cromossômica de progênies transgênicas homozigotas de soja [Glycine max (L. Merrill.], e confirmar a resistência dessas plantas a Anticarsia gemmatalis. Onze progênies com expressão dos genes cry1Ac, hpt e gusA foram avaliadas quanto às características agronômicas, em relação à cultivar parental IAS 5 não transformada. Análises citogenéticas foram realizadas em plantas transgênicas e não transgênicas. Duas das 11 prog

  4. 基于cry1Ac表达调控元件的苏云金芽孢杆菌表达载体构建%Construction of Bacillus thuringiensis Expression Vector by Using Regulatory Elements from cry1Ac Gene

    Institute of Scientific and Technical Information of China (English)

    郑文; 叶伟星; 彭东海; 孙明

    2012-01-01

    通过克隆cry1Ac基因BtⅠ-BtⅡ启动子、SD序列以及终止子,并引入pET28a的His标签和多克隆位点,在穿梭载体pHT304的基础上构建了一个苏云金芽孢杆菌表达载体pBMB1A.将cry1Ac以及具有非典型BtⅠ-BtⅡ启动子的cry2Ab、cry5Ba、cry6Aa、cry7Ba、cry55Aa6个基因装载到该表达载体上,转入BMB171构建了重组菌株,通过复红简单染色后的显微镜镜检结果表明,重组菌株BMB1A-1Ac、BMB1A-5Ba、BMB1A-7Ba和BMB1A-55Aa均能形成正常晶体,SDS-PAGE结果也证实这4个重组菌株的重组质粒均能表达出目的蛋白.同时,选取重组菌株BMB1A-1Ac来考察His标签对Cry1Ac杀虫活性的影响,生物活性测定结果显示重组菌株的LC50值与对照菌株相比无明显差异.该载体可用于快速克隆表达不同类别的Cry蛋白.%A Bacillus thuringiensis expression vector named Pbmbia was constructed by cloning BtI-BtII promoter, SD-sequence, and transcription terminator of cry 1 Ac gene, and adding His-tag and the following multiple cloning sites (MCS) from pET28a into the shuttle vector Pht304. Cry 1 Ac and other five cry genes with atypical Btl-Btll promoter including cry2Ab, cry5Ba, cry6Aa, cry7Ba, cry55Aa were cloned into Pbmbia. Microscopic observation showed that the recombinant strains containing CrylAc, Cry5Ba, Cry7Ba and Cry55Aa could form parasporal inclusion bodies; And SDS-PAGE proved that all of them could produce the corresponding major insecticidal crystal proteins bands. Meanwhile, CrylAc was selected to determine the effect of His-tag on its insecticidal activity, and the bioassay result showed that there was no significant difference between the LC50 of the recombinant strain and the control strain. The expression vector constructed was suitable for rapid cloning and expression of different cry genes.

  5. A seed mixture increases dominance of resistance to Bt cotton in Helicoverpa zea.

    Science.gov (United States)

    Brévault, Thierry; Tabashnik, Bruce E; Carrière, Yves

    2015-05-07

    Widely grown transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) can benefit agriculture, but adaptation by pests threatens their continued success. Refuges of host plants that do not make Bt toxins can promote survival of susceptible insects and delay evolution of resistance, particularly if resistance is inherited as a recessive trait. However, data have been lacking to compare the dominance of resistance when Bt and non-Bt seeds are planted in random mixtures versus separate blocks. Here we report results from greenhouse experiments with transgenic cotton producing Bt toxin Cry1Ac and the bollworm, Helicoverpa zea, showing that the dominance of resistance was significantly higher in a seed mixture relative to a block of Bt cotton. The proportion of larvae on non-Bt cotton plants in the seed mixture was also significantly higher than expected under the null hypothesis of random distribution. In simulations based on observed survival, resistance evolved 2- to 4.5-fold faster in the seed mixture relative to separate blocks of Bt and non-Bt cotton. These findings support previous modelling results indicating that block refuges may be more effective than seed mixtures for delaying resistance in pests with mobile larvae and inherently low susceptibility to the toxins in Bt crops.

  6. Natural enemies delay insect resistance to Bt crops.

    Science.gov (United States)

    Liu, Xiaoxia; Chen, Mao; Collins, Hilda L; Onstad, David W; Roush, Richard T; Zhang, Qingwen; Earle, Elizabeth D; Shelton, Anthony M

    2014-01-01

    We investigated whether development of resistance to a Bt crop in the presence of a natural enemy would be slower than without the natural enemy and whether biological control, in conjunction with a Bt crop, could effectively suppress the pest population. Additionally, we investigated whether insecticide-sprayed refuges of non-Bt crops would delay or accelerate resistance to the Bt crop. We used a system of Bt broccoli expressing Cry1Ac, a population of the pest Plutella xylostella with a low frequency of individuals resistant to Cry1Ac and the insecticide spinosad, and a natural enemy, Coleomegilla maculata, to conduct experiments over multiple generations. The results demonstrated that after 6 generations P. xylostella populations were very low in the treatment containing C. maculata and unsprayed non-Bt refuge plants. Furthermore, resistance to Bt plants evolved significantly slower in this treatment. In contrast, Bt plants with no refuge were completely defoliated in treatments without C. maculata after 4-5 generations. In the treatment containing sprayed non-Bt refuge plants and C. maculata, the P. xylostella population was low, although the speed of resistance selection to Cry1Ac was significantly increased. These data demonstrate that natural enemies can delay resistance to Bt plants and have significant implications for integrated pest management (IPM) with Bt crops.

  7. Natural enemies delay insect resistance to Bt crops.

    Directory of Open Access Journals (Sweden)

    Xiaoxia Liu

    Full Text Available We investigated whether development of resistance to a Bt crop in the presence of a natural enemy would be slower than without the natural enemy and whether biological control, in conjunction with a Bt crop, could effectively suppress the pest population. Additionally, we investigated whether insecticide-sprayed refuges of non-Bt crops would delay or accelerate resistance to the Bt crop. We used a system of Bt broccoli expressing Cry1Ac, a population of the pest Plutella xylostella with a low frequency of individuals resistant to Cry1Ac and the insecticide spinosad, and a natural enemy, Coleomegilla maculata, to conduct experiments over multiple generations. The results demonstrated that after 6 generations P. xylostella populations were very low in the treatment containing C. maculata and unsprayed non-Bt refuge plants. Furthermore, resistance to Bt plants evolved significantly slower in this treatment. In contrast, Bt plants with no refuge were completely defoliated in treatments without C. maculata after 4-5 generations. In the treatment containing sprayed non-Bt refuge plants and C. maculata, the P. xylostella population was low, although the speed of resistance selection to Cry1Ac was significantly increased. These data demonstrate that natural enemies can delay resistance to Bt plants and have significant implications for integrated pest management (IPM with Bt crops.

  8. 苏云金芽孢杆菌Ly30株cry1Ac基因的克隆及表达%Cloning and Expression of cry1Ac Gene from A Novel Bacillus thuringiensis Ly30 Strain

    Institute of Scientific and Technical Information of China (English)

    姚江; 张杰; 宋福平; 李长友; 刘旭光; 黄大昉

    2003-01-01

    Bt菌Ly30株是中国自行分离的对多种害虫具有高毒力的苏云金芽孢杆菌(Bacillusthunngiensis,Bt),经CAPS(cleaved amplified polymorphic sequences)系统鉴定,它含有cry1Ac基因.以全长基因PCR产物的粘端定向克隆的方法,设计1对特异引物,分别引入Sa1Ⅰ和BamHⅠ酶切位点.以Ly30质粒DNA为模板扩增cry1Ac全长基因,与表达载体pET-21b相应的酶切产物连接,转化大肠杆菌(Eschrichia coli),获得含有cry1Ac基因重组质粒pEKLy1Ac.该基因的亚克隆和序列测定结果表明,其编码区为3 534bp,编码蛋白分子量为133.5kD,含1 177个氨基酸,等电点为4.8,与Cry1Ac3同源性最高,存在4个氨基酸的差异,与Cry1Ac10之间则有6个氨基酸的不同.该基因序列已在GenBank中登记注册为AF482767,并被国际Bt杀虫晶体蛋白基因命名委员会正式命名为cry1Ac14.该基因经诱导获得高效表达,SDS-PAGE电泳检测到明显的133.5kD蛋白带.室内生物测定结果表明,诱导表达的Cry1Ac蛋白对棉铃虫、甜菜夜蛾等鳞翅目害虫幼虫均有较高的杀虫活性,其LC50值分别为19.236和3.276μg/g饲料.

  9. Introgression of Bt Genes in Novel Germplasm and Contribution to Indian Cotton Economy

    Institute of Scientific and Technical Information of China (English)

    VIDYASAGAR; Parchuri

    2008-01-01

    Emergence of transgenic Bt-cotton technology has opened up a new chapter in Indian cotton production in 21st century.The cry1Ac gene of Monsanto derived from American Upland Coker-312 background was not directly suitable for varied cotton growing situations in India.Delivery of Bt-gene

  10. Comparison of susceptibility of Chilo suppressalis and Bombyx mori to five Bacillus thuringiensis proteins.

    Science.gov (United States)

    Jiao, Yaoyu; Yang, Yan; Meissle, Michael; Peng, Yufa; Li, Yunhe

    2016-05-01

    Transformation of rice with genes encoding insecticidal Cry proteins from Bacillus thuringiensis (Bt) should confer high resistance to target lepidopteran pests, such as Chilo suppressalis, and low toxicity to non-target organisms, such as silkworm Bombyx mori. Five purified Cry proteins that have been used for plant transformation were tested using dietary exposure assays. The susceptibility of C. suppressalis larvae to the five insecticidal proteins in the decreasing order was: Cry1Ca>Cry1Ab>Cry1Ac>Cry2Aa>Cry1Fa. However, the toxicities of the Cry proteins to B. mori were in the order: Cry1Fa>Cry1Ca>Cry2Aa>Cry1Ab>Cry1Ac. The Cry1Ca, Cry1Ab and Cry1Ac proteins exhibited relatively high toxicity to C. suppressalis larvae, with EC50 values of 16.4, 45.8 and 89.6ng/g, respectively. The toxicities of the three Cry proteins to B. mori larvae were 8, 14, and 22times lower, with EC50 values of 138.3, 628.4 and 1939.2ng/g, respectively. The Cry1Fa and Cry2Aa proteins showed high toxicity to B. mori larvae, with EC50 values of 135.7 and 373.9ng/g, respectively, but low toxicity to C. suppressalis larvae, with EC50 values of 6092.1 and 1208.5ng/g, respectively. We thus conclude that Cry1Ab, Cry1Ac and Cry1Ca are appropriate for transforming rice to control lepidopteran rice pests. In contrast, Cry1Fa and Cry2Aa are not appropriate due to their high toxicity to silkworm larvae and low activity against the target pest.

  11. Early warning of cotton bollworm resistance associated with intensive planting of Bt cotton in China.

    Science.gov (United States)

    Zhang, Haonan; Yin, Wei; Zhao, Jing; Jin, Lin; Yang, Yihua; Wu, Shuwen; Tabashnik, Bruce E; Wu, Yidong

    2011-01-01

    Transgenic crops producing Bacillus thuringiensis (Bt) toxins kill some key insect pests, but evolution of resistance by pests can reduce their efficacy. The predominant strategy for delaying pest resistance to Bt crops requires refuges of non-Bt host plants to promote survival of susceptible pests. To delay pest resistance to transgenic cotton producing Bt toxin Cry1Ac, farmers in the United States and Australia planted refuges of non-Bt cotton, while farmers in China have relied on "natural" refuges of non-Bt host plants other than cotton. Here we report data from a 2010 survey showing field-evolved resistance to Cry1Ac of the major target pest, cotton bollworm (Helicoverpa armigera), in northern China. Laboratory bioassay results show that susceptibility to Cry1Ac was significantly lower in 13 field populations from northern China, where Bt cotton has been planted intensively, than in two populations from sites in northwestern China where exposure to Bt cotton has been limited. Susceptibility to Bt toxin Cry2Ab did not differ between northern and northwestern China, demonstrating that resistance to Cry1Ac did not cause cross-resistance to Cry2Ab, and implying that resistance to Cry1Ac in northern China is a specific adaptation caused by exposure to this toxin in Bt cotton. Despite the resistance detected in laboratory bioassays, control failures of Bt cotton have not been reported in China. This early warning may spur proactive countermeasures, including a switch to transgenic cotton producing two or more toxins distinct from Cry1A toxins.

  12. Early warning of cotton bollworm resistance associated with intensive planting of Bt cotton in China.

    Directory of Open Access Journals (Sweden)

    Haonan Zhang

    Full Text Available Transgenic crops producing Bacillus thuringiensis (Bt toxins kill some key insect pests, but evolution of resistance by pests can reduce their efficacy. The predominant strategy for delaying pest resistance to Bt crops requires refuges of non-Bt host plants to promote survival of susceptible pests. To delay pest resistance to transgenic cotton producing Bt toxin Cry1Ac, farmers in the United States and Australia planted refuges of non-Bt cotton, while farmers in China have relied on "natural" refuges of non-Bt host plants other than cotton. Here we report data from a 2010 survey showing field-evolved resistance to Cry1Ac of the major target pest, cotton bollworm (Helicoverpa armigera, in northern China. Laboratory bioassay results show that susceptibility to Cry1Ac was significantly lower in 13 field populations from northern China, where Bt cotton has been planted intensively, than in two populations from sites in northwestern China where exposure to Bt cotton has been limited. Susceptibility to Bt toxin Cry2Ab did not differ between northern and northwestern China, demonstrating that resistance to Cry1Ac did not cause cross-resistance to Cry2Ab, and implying that resistance to Cry1Ac in northern China is a specific adaptation caused by exposure to this toxin in Bt cotton. Despite the resistance detected in laboratory bioassays, control failures of Bt cotton have not been reported in China. This early warning may spur proactive countermeasures, including a switch to transgenic cotton producing two or more toxins distinct from Cry1A toxins.

  13. Bacillus thuringiensis plants expressing Cry1Ac, Cry2Ab and Cry1F are not toxic to the assassin bug, Zelus renardii

    Science.gov (United States)

    Cotton and maize delivering insecticidal crystal (Cry) proteins from the bacterium, Bacillus thuringiensis (Bt), have been commercialized since 1996. Bt plants are subjected to environmental risk assessments for non-target organisms, especially natural enemies that suppress pest populations. In th...

  14. Sustained susceptibility of pink bollworm to Bt cotton in the United States.

    Science.gov (United States)

    Tabashnik, Bruce E; Morin, Shai; Unnithan, Gopalan C; Yelich, Alex J; Ellers-Kirk, Christa; Harpold, Virginia S; Sisterson, Mark S; Ellsworth, Peter C; Dennehy, Timothy J; Antilla, Larry; Liesner, Leighton; Whitlow, Mike; Staten, Robert T; Fabrick, Jeffrey A; Li, Xianchun; Carrière, Yves

    2012-01-01

    Evolution of resistance by pests can reduce the benefits of transgenic crops that produce toxins from Bacillus thuringiensis (Bt) for insect control. One of the world's most important cotton pests, pink bollworm (Pectinophora gossypiella), has been targeted for control by transgenic cotton producing Bt toxin Cry1Ac in several countries for more than a decade. In China, the frequency of resistance to Cry1Ac has increased, but control failures have not been reported. In western India, pink bollworm resistance to Cry1Ac has caused widespread control failures of Bt cotton. By contrast, in the state of Arizona in the southwestern United States, monitoring data from bioassays and DNA screening demonstrate sustained susceptibility to Cry1Ac for 16 y. From 1996-2005, the main factors that delayed resistance in Arizona appear to be abundant refuges of non-Bt cotton, recessive inheritance of resistance, fitness costs associated with resistance and incomplete resistance. From 2006-2011, refuge abundance was greatly reduced in Arizona, while mass releases of sterile pink bollworm moths were made to delay resistance as part of a multi-tactic eradication program. Sustained susceptibility of pink bollworm to Bt cotton in Arizona has provided a cornerstone for the pink bollworm eradication program and for integrated pest management in cotton. Reduced insecticide use against pink bollworm and other cotton pests has yielded economic benefits for growers, as well as broad environmental and health benefits. We encourage increased efforts to combine Bt crops with other tactics in integrated pest management programs.

  15. Effects of transgenic Cry1A+CpTI cotton and Cry1Ac toxin on the parasitoid, Campoketis chlorideae (Hymenoptera:Ichneumonidae)

    Institute of Scientific and Technical Information of China (English)

    XIAO-XIALIU; CHANG-GUISUN; QING-WENZHANG

    2005-01-01

    The effect oftransgenic double genes, Cry1A + CpTlcotton and Cry1Ac toxin on the parasitoid, Carnpoketis chlorideae Uchida of cotton bollworm, Helicoverpa armigera (Hübner), was investigated in the laboratory. Helicoverpa armigera larvae when in the first,second and third instar could not survive if fed on transgenic cotton leaves. Consequently,C. chlorideae larvae could not complete their development if parasitizing on such hosts.After H. armigera larvae were reared on transgenic or traditional cotton leaves for 12-48 hours, they were parasitized by C. chlorideae females. Parasitized larvae continued to feed on transgenic or traditional cotton for 12-48 h. The present results showed that the body weight of larvae of the parasitoids were significantly reduced when parasitized hosts fed on transgenic cotton leaves compared to those fed on traditional cotton. Duration of egg and larvae stage were significantly prolonged, pupal and adult weight of C. chloridae was decreased when the host larvae fed on transgenic cotton leaves longer than 48 h. The development duration of C. chlorideae pupae on the hosts fed on transgenic cotton leaves in each treatment was not significantly different from those of controls. The longevity of parasitoid females and males fed with a solution containing Cry1Ac toxin was not significantly different with that of the control.

  16. Cross-resistance of Cry1Ac-resistant Heliocoverpa armigera Hübner%抗Bt棉铃虫种群的交互抗性研究

    Institute of Scientific and Technical Information of China (English)

    卢美光; 芮昌辉; 赵建周; 孟香清

    2000-01-01

    分别用对Cry1Ac抗性达11.6倍的Bt棉叶和Bt粉剂(Btk)汰选棉铃虫(Heliocoverpa armigera)种群(对Btk的抗性指数为5.2倍),测定了其对不同Bt制剂、Bt毒素、化学农药的交互抗性.结果表明:停止汰选后,Bt棉叶汰选种群对Cry1Ac的抗性指数为5.9倍,对Cry1Ac+1C的交互抗性为3.7倍,对Btk的敏感性降低,对Bta、Cry2A无交互抗性;Btk汰选种群对Btk的抗性指数为4.1倍,对Bta、 Cry1Ac、Cry1Ac+1C的交互抗性为2.3、3.1、2.3倍,对Cry2A无交互抗性;两汰选种群对化学农药灭多威、辛硫磷、氯氰菊酯的敏感性均有所增加.

  17. 表面等离子共振传感器检测转基因植物中苏云金芽孢杆菌cry1Ac蛋白的方法研究%Using a Surface Plasmon Resonance Sensor for the Detection of Bt cry1Ac Protein

    Institute of Scientific and Technical Information of China (English)

    杜方; 纪淑娟; 黄新

    2014-01-01

    为建立表面等离子共振传感器检测转基因植物中苏云金芽孢杆菌cry1Ac蛋白的方法,根据生物分子间的相互作用,在金片表面修饰cry1Ac蛋白的特异性单克隆抗体,对不同稀释梯度的cry1Ac蛋白进行检测研究.结果表明:该方法可有效的检测到cry1Ac蛋白,检测灵敏度可达到10ng· mL-1,与cry2A蛋白及CP4-EPSPS蛋白无交叉反应.说明利用表面等离子共振方法检测cry1Ac蛋白操作简单,省时且灵敏度高,特异性强,可用于对cry1Ac蛋白的定量检测.

  18. Biological characteristics of black armyworm Spodoptera cosmioides on genetically modified soybean and corn crops that express insecticide Cry proteins

    Directory of Open Access Journals (Sweden)

    Gabriela Vieira Silva

    2016-09-01

    Full Text Available ABSTRACT This study aimed to evaluate the development and reproduction of the black armyworm, Spodoptera cosmioides when larvae fed on leaves of Bt-corn hybrids, expressing a single Cry1F and also Cry1F, Cry1A.105 and Cry2Ab2 in pyramided corn and their non-Bt-isoline (hybrid 2B688, as well as on leaves of two soybean isolines expressing the Cry1Ac protein and its non-Bt isoline (A5547-227. We also assessed the effect of these Bt and non-Bt plants on the leaf consumption rate of S. cosmioides larvae. This pest was unable to develop when fed on any of the corn isolines (Bt and non-Bt. When both 1st and 3rd instar larvae were fed on corn leaf, mortality was 100% in both Bt and non-Bt corn. In contrast, when corn leaves were offered to 5th instar larvae, there were survivors. Defoliation and leaf consumption was higher with non-Bt corn than with both of the Bt corn isolines. There was no negative effect of Bt soybean leaves on the development and reproduction of S. cosmioides with respect to all evaluated parameters. Our study indicates that both Bt and non-Bt corn adversely affect the development of S. cosmioides while Bt soybean did not affect its biology, suggesting that this lepidopteran has major potential to become an important pest in Bt soybean crops.

  19. Early detection of field-evolved resistance to Bt cotton in China: cotton bollworm and pink bollworm.

    Science.gov (United States)

    Tabashnik, Bruce E; Wu, Kongming; Wu, Yidong

    2012-07-01

    Transgenic crops producing Bacillus thuringiensis (Bt) toxins kill some major insect pests, but pests can evolve resistance and thereby reduce the effectiveness of such Bt crops. The main approach for slowing pest adaptation to Bt crops uses non-Bt host plants as "refuges" to increase survival of susceptible pests. To delay evolution of pest resistance to cotton producing Bt toxin Cry1Ac, several countries have required refuges of non-Bt cotton, while farmers in China have relied on "natural" refuges of non-Bt host plants other than cotton. This strategy is designed for cotton bollworm (Helicoverpa armigera), which attacks many crops and is the primary target of Bt cotton in China, but it does not apply to pink bollworm (Pectinophora gossypiella), which feeds almost entirely on cotton in China. Here we review evidence of field-evolved resistance to Cry1Ac by cotton bollworm in northern China and by pink bollworm in the Yangtze River Valley of China. For both pests, results of laboratory diet bioassays reveal significantly decreased susceptibility of field populations to Cry1Ac, yet field control failures of Bt cotton have not been reported. The early detection of resistance summarized here may spur countermeasures such as planting Bt cotton that produces two or more distinct toxins, increased planting of non-Bt cotton, and integration of other management tactics together with Bt cotton.

  20. Expresión de la proteína Cry1Ac en tejidos de líneas transgénicas de papa (Solanum tuberosum spp. Andígena Var. Diacol capiro.

    Directory of Open Access Journals (Sweden)

    Blanco Martinez Jennifer Teresa

    2010-08-01

    ="ES-TRAD">y no mostraron diferencias significativas entre las líneas analizadas, ni entre las tres etapas del desarrollo. No se evidenciaron diferencias significativas entre las líneas transgénicas con respecto al control al hacer un análisis de algunas características fenotípicas relevantes. Los resultados encontrados sugieren la realización de seguimientos y ensayos de bioseguridad sobre este material, ya que los altos niveles de expresión en todos los tejidos analizados, pueden afectar a organismos no blanco.

     

    ABSTRACT

    Potato is the fourth of the most important crop and foodstuff in the world, in Colombia around 2.8 millions of tons are produced annually supporting economically 90000 families. In the country, the major economic impact in the crop is caused by Tecia solanivora that originates loses up to 100% in the tuber production. The genetic plant breeding related to the introduction of Cry genes which codify insecticidal crystal proteins is an alternative for reducing the insect attack in commercial crops. In this work, the insertion, transcription and expression of Cry1Ac gen was characterized in different tissues and three development stages of two

  1. FROM Qutn TO Bt COTTON: DEVELOPMENT, ADOPTION AND PROSPECTS. A REVIEW.

    Science.gov (United States)

    Maik, W; Abid, M A; Cheema, H M N; Khan, A A; Iqbal, M Z; Qayyum, A; Hanif, M; Bibi, N; Yuan, S N; Yasmeen, A; Mahmood, A; Ashraf, J

    2015-01-01

    Cotton has unique history of domestication, diversification, and utilization. Globally it is an important cash crop that provides raw material for textile industry. The story of cotton started from human civilization and the climax arrived with the efforts of developing transgenic cotton for various traits. Though conventional breeding brought steady improvement in developing resistance against biotic stresses but recent success story of gene transferfrom Bacillus thuringiensis into cotton showed game changing effects on cotton cultivation. Amongst various families of insecticidal proteins Bt Cry-toxins received more attention because of specificity against receptors on the cell membranes of insect midgut epithelial cells. Rapid Bt cotton adoption by farmers due to its economic and environmental benefits has changed the landscape of cotton cultivation in many countries. But the variable expression of Bt transgene in the newly developed Bt cotton genotypes in tropical environment is questionable. Variability of toxin level in different plant parts at various life stage of plant is an outcome of genotypic interaction with environmental factors. Temporal gene expression of Cry1Ac is also blamed for the epigenetic background in which transgene has been inserted. The presence of genotypes with sub-lethal level of Bt toxin might create resistance in Lepidopteron insects, limiting the use of Bt cotton in future, with the opportunityfor other resistance development strategies to get more attention like gene stacking. Until the farmers get access to more recent technology, best option is to delay the development of resistance by applying Insect Resistance Management (IRM) strategies.

  2. Interactions Between Exogenous Bt Insecticidal Protein and Cotton Terpenoid Aldehydes

    Institute of Scientific and Technical Information of China (English)

    ZHANG Yong-jun; GUO Yu-yuan; WU Kong-ming; WANG Wu-gang

    2002-01-01

    The contents of terpenoid aldehydes in Bt transgenic cotton and their non-Bt parental varieties were analyzed by the HPLC method. Statistical analysis of variance showed that Bt insecticidal protein Bt-ICP expression has no negative effect on the synthesis of gossypol, total heliocides and total resistant terpenoids.The results of the combined dosage test of Bt-ICP and gossypoi in vitro showed that there is no interaction between gossypol and Bt-ICP on the mortality of cotton boilworm larvae Helicoverpa armigera (Hubnner). It is indicated that the actions of Bt-ICP and gossypol on cotton bollworm are additive. Therefore, it is advantageous to combine Bt-ICP with cotton terpenoid aldehydes in controlling cotton bollworm.

  3. Helicoverpa armigera baseline susceptibility to Bacillus thuringiensis Cry toxins and resistance management for Bt cotton in India.

    Science.gov (United States)

    Gujar, G T; Kalia, V; Kumari, A; Singh, B P; Mittal, A; Nair, R; Mohan, M

    2007-07-01

    Transgenic cotton that produces insecticidal proteins from Bacillus thuringiensis (Bt), often referred to as Bt cotton, is widely grown in many countries. Bt cotton with a single cry1A gene and stacked also with cry2A gene has provided satisfactory protection against the damage by the lepidopteran bollworms, especially the cotton bollworm, Helicoverpa armigera (Hübner) which is considered as a key pest. The baseline susceptibility of the larvae of H. armigera to Cry1Ac and other toxins carried out in many countries has provided a basis for monitoring resistance. There is no evidence of development of field-level resistance in H. armigera leading to the failure of Bt cotton crop anywhere in the world, despite the fact that Bt cotton was grown on the largest ever area of 12.1 million hectares in 2006 and its cumulative cultivation over the last 11 years has surpassed the annual cotton area in the world. Nevertheless, the Bt resistance management has become a necessity to sustain Bt cotton and other transgenic crops in view of potential of the target insects to evolve Cry toxin resistance.

  4. A comparison of spider communities in Bt and non-Bt rice fields.

    Science.gov (United States)

    Lee, Sue Yeon; Kim, Seung Tae; Jung, Jong Kook; Lee, Joon-Ho

    2014-06-01

    To assess the potential adverse effects of a Bt rice line (Japonica rice cultivar, Nakdong) expressing a synthetic cry1Ac1 gene, C7-1-9-1-B, which was highly active against all larval stages of Cnaphalocrocis medinalis (Guenée) (Lepidoptera: Crambidae), we investigated the community structure of spiders in Bt and non-Bt rice fields during the rice-growing season in 2007 and 2008 in Chungcheongnam-do, Korea. Spiders were surveyed with a sweep net and suction device. Suction sampling captured more spiders, measured in terms of species level and abundance, than sweeping. Araneidae and Thomisidae were captured more by sweeping, and certain species were captured only by sweeping. These findings show that both suction and sweep sampling methods should be used because these methods are most likely complementary. In total, 29 species in 23 genera and nine families were identified from the 4,937 spiders collected, and both Bt and non-Bt rice fields showed a typical Korean spider assemblage. The temporal patterns of spider species richness and spider abundance were very similar between Bt and non-Bt rice, although significant differences in species richness were observed on a few occasions. Overall, spider community structure, including diversity, the dominant species, and abundance did not differ between Bt and non-Bt rice. The results of the study indicated that the transgenic Cry1Ac rice lines tested in this study had no adverse effects on the spider community structure of the rice fields.

  5. BT799玉米对亚洲玉米螟抗性研究%Resistance of transgenic Bt corn variety BT799 to the Asian corn borer

    Institute of Scientific and Technical Information of China (English)

    王月琴; 何康来; 江帆; 王依冬; 张天涛; 王振营; 白树雄

    2014-01-01

    Objectives] To review the resistance of transgenic Bt corn to target pests which is one of the primarily steps for research and development of insect-resistant, transgenic corn. This review mainly evaluates the resistance of transgenic Cry1Ac gene corn variety BT799 to the Asian corn borer (ACB), Ostrinia furnacalis (Guenée), and measured the quantity of Cry1Ac protein expressed in corn plant tissues. [Methods] Enzyme-linked immunosorbent assays, laboratory bioassay and field studies with artificial infestation were employed in this study. [Results] The expression levels of Cry1Ac protein were 768.0 ng/g (protein/fresh leaves), 1452.8-2978.5 ng/g (protein/dry mass of silk, husk and young kernels). Leaf-feeding rates in field trials indicated that BT799 and CC-2XBT799 were highly resistant to ACB. The survival of ACB larvae feeding on Zhengdan 958K, a single cross hybrid containing BT799, was significantly lower (0-37.5%) than that of larvae that fed on non-Bt Zhengdan 958 (89.9%-100.0%). In addition, larval survival of Cry1Ac-, Cry1Ie-, and Cry1F-selected ACB strains (ACB-AcR, ACB-IeR and ACB-FR) were significantly different when fed on Zhengdan 958K. ACB-IeR had the lowest survival followed by ACB-FR, both of them had significantly lower survival than those that fed on the control Zhengdan 958. However, the survival of ACB-AcR was not significantly different from larvae feeding on the control Zhengdan 958. [Conclusion] These results suggests that the transgenic Bt corn variety BT799 is highly toxic to the ACB and can provide effective control for ACB in the field.%【目的】抗螟性鉴定是转基因抗虫玉米研发的重要一环。本文主要就转基因玉米BT799对亚洲玉米螟的抗性展开评价,同时测定了BT799植株组织中Cry1Ac蛋白的表达量。【方法】采用了酶联免疫吸附测定法(ELISA)、室内生测和田间人工接虫鉴定3种方法。【结果】转基因抗虫玉米BT799组织中Cry1Ac蛋白含量分别为768

  6. Large-scale test of the natural refuge strategy for delaying insect resistance to transgenic Bt crops.

    Science.gov (United States)

    Jin, Lin; Zhang, Haonan; Lu, Yanhui; Yang, Yihua; Wu, Kongming; Tabashnik, Bruce E; Wu, Yidong

    2015-02-01

    The 'natural refuge strategy" for delaying insect resistance to transgenic cotton that produces insecticidal proteins from Bacillus thuringiensis (Bt) relies on refuges of host plants other than cotton that do not make Bt toxins. We tested this widely adopted strategy by comparing predictions from modeling with data from a four-year field study of cotton bollworm (Helicoverpa armigera) resistance to transgenic cotton producing Bt toxin Cry1Ac in six provinces of northern China. Bioassay data revealed that the percentage of resistant insects increased from 0.93% in 2010 to 5.5% in 2013. Modeling predicted that the percentage of resistant insects would exceed 98% in 2013 without natural refuges, but would increase to only 1.1% if natural refuges were as effective as non-Bt cotton refuges. Therefore, the results imply that natural refuges delayed resistance, but were not as effective as an equivalent area of non-Bt cotton refuges. The percentage of resistant insects with nonrecessive inheritance of resistance increased from 37% in 2010 to 84% in 2013. Switching to Bt cotton producing two or more toxins and integrating other control tactics could slow further increases in resistance.

  7. Effects of Rice Genetically Modified With Cry1Ac/sck on Intestinal Microflor of Broiler Chicken%转Cry1Ac/sck基因糙米作为肉仔鸡日粮原料对肠道群的影响研究

    Institute of Scientific and Technical Information of China (English)

    秦海峰; 刘阳; 邢福国; 王会娟

    2012-01-01

    To assess whether the transgenic brown rice used in broiler's diet had any deleterious effects on intestinal micoflor of broiler chicken, a 42- day feeding trial was employed. Three hunder sixty one- day old broilers were divided into three groups. One group was fed a diet containing transgenic brown rice and the other group was fed a diet containing corresponding nontransgenic brown rice. Three groups were fed a diet containing corn. Entericmicroorganisms at the 21 and 42 day of age were determined. No histopathological damages were detected. From this study, it was concluded that there were no adverse effects of the transgenic brown rice on the broiler chicken's entericmicroorganisms. The transgenic brown rice used in broiler's diet was safe on entericmicroorganisms of brolier chicken.%为评价转Cry1Ac/sck基因糙米作为肉仔鸡日粮原料对肠道微生态的影响.将360只1日龄肉仔鸡随机分成3组,第一组饲喂含转基因糙米日粮;第二组饲喂含非转基因糙米日粮;第三组饲喂非转基因玉米日粮,试验期42天.在21、42天时测定肉仔鸡肠道菌群情况.结果表明:在实验期内,转基因糙米对肉仔鸡肠道菌群未产生明显影响.转Cry1Ac/sck基因糙米作为肉仔鸡日粮原料对肉仔鸡肠道微生态是安全的.

  8. Effect of pyramiding Bt and CpTI genes on resistance of cotton to Helicoverpa armigera (Lepidoptera: Noctuidae) under laboratory and field conditions

    NARCIS (Netherlands)

    Cui, J.J.; Luo, J.Y.; Werf, van der W.; Ma, Y.; Xia, J.Y.

    2011-01-01

    Transgenic cotton (Gossypium hirsutum L.) varieties, adapted to China, have been bred that express two genes for resistance to insects. the Cry1Ac gene from Bacillus thuringiensis (Berliner) (Bt), and a trypsin inhibitor gene from cowpea (CpTI). Effectiveness of the double gene modification in confe

  9. Interaction of proteinase inhibitors with Cry1Ac toxicity and presence of fifteen chymotrypsin cDNAs in midgut of the tobacco budworm Heliothis virescens (F.) (Lepidoptera: noctuidae)

    Science.gov (United States)

    Extensive adoption of Bt cotton places heavy selection pressure on lepidopteran insects. The potential development of Bt resistance is a major risk in the durability of Bt plant technology. Another concern is the surging of non-lepidopteran pests not controlled by Bt toxins. Intensive cloning and se...

  10. Study on Detection Technology and Survival Competition Ability of Transgenic Cry1Ac + EPSPS Cotton: Based on Glyphosate Pressure and No Control on Target Pests%转抗虫抗草甘膦除草剂基因(Cry1Ac+EPSPS)棉的检测技术与生存竞争能力研究——基于草甘膦压力和对靶标害虫不防治环境下

    Institute of Scientific and Technical Information of China (English)

    李捷; 张兴华; 马艳; 杨兆光; 乔艳艳

    2013-01-01

    此研究为国家转抗虫抗草甘膦除草剂基因(Cry1Ac+ EPSPS)棉环境安全评价技术建立而设.在草甘膦压力和对靶标害虫不防治环境下的检测技术,以杂交抗虫棉赣棉杂1号和非转基因棉赣棉11号为2个对照,来检测评价转抗虫抗草甘膦除草剂基因(Cry1Ac+ EPSPS)棉的生存竞争能力.结果:转Cry1Ac+ EPSPS棉与两个对照相比不显示抗虫优势;株高增长无竞争力;覆盖度有不显著的弱竞争力;单株成铃、单铃籽棉重、单铃皮棉重、衣分率、脱落率、籽棉产量、皮棉产量与赣棉杂1号比呈现显著或板显著的负竞争优势,与赣棉11号比无竞争优势;果枝层数前期与两对照相比有极显著的负竞争优势,后期与赣棉杂1号比无竞争优势,与赣棉11号比有显著的负竞争优势.结论:转Cry1Ac+ EPSPS棉推广种植可行性较差.本项检测技术和评价方法有较好的适用性.

  11. Selection and genotype analysis of Bt isolates with high virulence against Athetis lepigone (Möschler)%对二点委夜蛾高毒力苏云金芽胞杆菌的筛选及其基因型分析

    Institute of Scientific and Technical Information of China (English)

    杨云鹤; 宋萍; 王振营; 董志平; 王勤英

    2014-01-01

    【目的】获得对二点委夜蛾Athetis lepigone(Möschler)高毒力的苏云金芽胞杆菌(Bt)菌株,寻找对该虫具有特异杀虫活性的蛋白毒素,探索 Bt 菌株或其杀虫基因应用于二点委夜蛾防治的可行性。【方法】通过生物测定方法比较了36株苏云金芽胞杆菌和一株恶臭假单胞工程菌PHB-cry1Ab对二点委夜蛾幼虫的杀虫活性,同时利用PCR-RFLP方法对这些菌株的基因型进行了分析。【结果】不同Bt菌株对二点委夜蛾幼虫的杀虫活性差别很大,杀虫活性高的菌株都含有 cry1Ac 基因。饲毒72 h 后含单基因的Bt HD-73菌株(cry1Ac)对二点委夜蛾2龄幼虫的毒力(LC50值为188.51μg/g)明显高于含多基因的Bt SC-40菌株(cry1Ac,cry2Ac,cry1I,vip3A)的毒力(LC50值为418.13μg/g)。含有vip3A基因的Bt SC-40和Bt HD-1营养期上清液对二点委夜蛾2龄幼虫表现出一定的杀虫活性(72 h死亡率分别达到42.5%和57.4%),而无vip3A基因的Bt HD-73营养期上清液未表现出明显的杀虫活性。【结论】由cry1Ac基因编码的Cry1Ac蛋白对二点委夜蛾幼虫具有特异杀虫活性,Vip3A蛋白对二点委夜蛾幼虫可能也有一定的杀虫活性。%[Objectives] To obtain Bt strains and insecticidal proteins with high insecticidal activity against Athetis lepigone (Möschler). [Methods] The insecticidal activities of 36 Bt strains and an engineered Pseudomonas putida with a cry1Ab gene were evaluated on second instar larvae of A. lepigone by bioassay. PCR-RFLP analysis was performed to verify the genotypes of all Bt isolates. [Results] The bioassay results indicated obvious differences in the insecticidal activities of the different Bt isolates tested. The results also revealed that the Bt strains with high insecticidal activity all contained the cry1Ac gene. Furthermore, the results of a comparison of the toxicities of Bt SC-40 (cry1Ac, cry2Ac, cry1I, vip3A) and Bt HD-73

  12. Single amino acid mutation in an ATP-binding cassette transporter gene causes resistance to Bt toxin Cry1Ab in the silkworm, Bombyx mori.

    Science.gov (United States)

    Atsumi, Shogo; Miyamoto, Kazuhisa; Yamamoto, Kimiko; Narukawa, Junko; Kawai, Sawako; Sezutsu, Hideki; Kobayashi, Isao; Uchino, Keiro; Tamura, Toshiki; Mita, Kazuei; Kadono-Okuda, Keiko; Wada, Sanae; Kanda, Kohzo; Goldsmith, Marian R; Noda, Hiroaki

    2012-06-19

    Bt toxins derived from the arthropod bacterial pathogen Bacillus thuringiensis are widely used for insect control as insecticides or in transgenic crops. Bt resistance has been found in field populations of several lepidopteran pests and in laboratory strains selected with Bt toxin. Widespread planting of crops expressing Bt toxins has raised concerns about the potential increase of resistance mutations in targeted insects. By using Bombyx mori as a model, we identified a candidate gene for a recessive form of resistance to Cry1Ab toxin on chromosome 15 by positional cloning. BGIBMGA007792-93, which encodes an ATP-binding cassette transporter similar to human multidrug resistance protein 4 and orthologous to genes associated with recessive resistance to Cry1Ac in Heliothis virescens and two other lepidopteran species, was expressed in the midgut. Sequences of 10 susceptible and seven resistant silkworm strains revealed a common tyrosine insertion in an outer loop of the predicted transmembrane structure of resistant alleles. We confirmed the role of this ATP-binding cassette transporter gene in Bt resistance by converting a resistant silkworm strain into a susceptible one by using germline transformation. This study represents a direct demonstration of Bt resistance gene function in insects with the use of transgenesis.

  13. The effects of high temperature level on square Bt protein concentration of Bt cotton

    Institute of Scientific and Technical Information of China (English)

    WANG Jun; CHEN Yuan; YAO Meng-hao; LI Yuan; WEN Yu-jin; ZHANG Xiang; CHEN De-hua

    2015-01-01

    Higher bol worm survival rates were detected after high temperature presented during square period in Bt cotton. The objective of this study was to investigate the effects of high temperature level on the Bt efifcacy of two different types of Bt cotton cultivars at squaring stage. During the 2011 to 2013 cotton growth seasons, high temperature treatments ranged from 34 to 44°C in climate chambers, and ifeld experiments under high temperature weather with various temperature levels were conducted to investigate the effects of the high temperature level on square Bt protein concentration and nitrogen metabolism. The climate chamber experiments showed that the square insecticidal protein contents reduced after 24 h elevated temperature treatments for both cultivars, whereas signiifcant declines of the square insecticidal protein contents were detected at temperature >38°C, and only slightly numerical reductions were observed when temperature below 38°C. Similar high temperature responses were also observed at the two ifeld experimental sites in 2013. Correspondingly, high temperature below 38°C seems have little effect on the square amino acid concentrations, soluble protein contents, glutam-ic-pyruvic transaminase (GPT) and glutamic-oxalacetic transaminase (GOT) activities as wel as protease and peptidase activities; however, when the temperature was above 38°C, reduced soluble protein contents, enhanced amino acid con-centrations, decreased GPT and GOT activities, bolstered protease and peptidase activities in square were detected. In general, the higher the temperature is (>38°C), the larger the changes for the above compound contents and key enzymes activities of the square protein cycle. The ifndings indicated that the unstable insect resistance of the square was related to high temperature level during square stage.

  14. [Effects of high temperature on Bt protein content and nitrogen metabolic physiology in boll wall of Bt cotton].

    Science.gov (United States)

    Wang, Jun; Abidallah, Eltayib H M A; Hua, Ming-ming; Heng, Li; Lyu, Chun-hua; Chen, De-hua

    2015-10-01

    Bt cotton cultivar Sikang 1 (a conventional cultivar) and Sikang 3 (a hybrid cultivar) from China, and 99B (a conventional cultivar) and Daiza 1 (a hybrid cultivar) from USA were selected as experimental materials, the ball wall Bt protein content and nitrogen metabolic physiology were investigated under different high temperature levels at peak boll stage. The results showed that the Bt protein content of boll wall decreased with the increasing temperature. Compared with the control (32 °C, the boll wall Bt protein content decreased significantly when the temperature was above 38 °C for the conventional cultivars and above 40 °C for the hybrid cultivars. The Bt protein contents of cultivar Sikang 1 and 99B decreased by 53.0% and 69.5% respectively with the temperature at 38 °C, and that of cultivar Sikang 3 and Daiza 1 decreased by 64.8% and 54.1% respectively with the temperature at 40 °C. Greater reductions in the boll wall soluble protein contents and GPT activities, larger increments for the boll wall free amino acid contents and proteinsase activities were also observed when the boll wall Bt protein content was significantly reduced. Therefore, high temperature resulted in the reduction of Bt protein synthesis and increase of the insecticidal protein degradation in the boll wall significantly, which caused the reductions in boll wall Bt protein content and insect resistance.

  15. Cloning eleven midgut trypsin cDNAs and evaluating the interaction of proteinase inhibitors with Cry1Ac against the tobacco budworm Heliothis virescens (F.) (Lepidoptera: Noctuidae)

    Science.gov (United States)

    Midgut trypsins are associated with Bt protoxin activation and toxin degradation. Proteinase inhibitors have potential insecticidal toxicity against a wide range of insect species. Proactive action to examine trypsin gene profiles and proteinase inhibitors for interaction with Bt toxin is necessary ...

  16. Effect of pyramiding Bt and CpTI genes on resistance of cotton to Helicoverpa armigera (Lepidoptera: Noctuidae) under laboratory and field conditions

    OpenAIRE

    2011-01-01

    Transgenic cotton (Gossypium hirsutum L.) varieties, adapted to China, have been bred that express two genes for resistance to insects. the Cry1Ac gene from Bacillus thuringiensis (Berliner) (Bt), and a trypsin inhibitor gene from cowpea (CpTI). Effectiveness of the double gene modification in conferring resistance to cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae), was studied in laboratory and field experiments. In each experiment, performance of Bt+CpTI cotton was c...

  17. Interação do algodoeiro BT submetido ao estresse hídrico e Alabama argillacea (Hübner) (Lepidoptera: Noctuidae)

    OpenAIRE

    de Lima, Mauricio Silva

    2010-01-01

    The genetically modified cotton to produce toxins from the bacterium Bacillus thuringiensis (Bt) has the objective of offering plant resistance against lepidopteran larvae. The cotton crop cultivated in the Semiarid region can be subjected to the condition of water stress. Thus, this work investigated the expression of the Cry1Ac toxin in two varieties of Bt-cotton Acala 90B and NuOpal cultivated under the conditions of irrigated and water stress. In addition, the oviposition and feeding pref...

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

    Science.gov (United States)

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

    2016-07-01

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

  19. Feeding and dispersal behavior of the cotton leafworm, Alabama argillacea (Hübner) (Lepidoptera: Noctuidae), on Bt and non-Bt cotton: implications for evolution and resistance management.

    Science.gov (United States)

    Ramalho, Francisco S; Pachú, Jéssica K S; Lira, Aline C S; Malaquias, José B; Zanuncio, José C; Fernandes, Francisco S

    2014-01-01

    The host acceptance of neonate Alabama argillacea (Hübner) (Lepidoptera: Noctuidae) larvae to Bt cotton plants exerts a strong influence on the potential risk that this pest will develop resistance to Bt cotton. This will also determine the efficiency of management strategies to prevent its resistance such as the "refuge-in-the-bag" strategy. In this study, we assessed the acceptance of neonate A. argillacea larvae to Bt and non-Bt cotton plants at different temperatures during the first 24 h after hatching. Two cotton cultivars were used in the study, one a Bt DP 404 BG (Bollgard) cultivar, and the other, an untransformed isoline, DP 4049 cultivar. There was a greater acceptance by live neonate A. argillacea larvae for the non-Bt cotton plants compared with the Bt cotton plants, especially in the time interval between 18 and 24 h. The percentages of neonate A. argillacea larvae found on Bt or non-Bt plants were lower when exposed to temperatures of 31 and 34 °C. The low acceptance of A. argillacea larvae for Bt cotton plants at high temperatures stimulated the dispersion of A. argillacea larvae. Our results support the hypothesis that the dispersion and/or feeding behavior of neonate A. argillacea larvae is different between Bt and non-Bt cotton. The presence of the Cry1Ac toxin in Bt cotton plants, and its probable detection by the A. argillacea larvae tasting or eating it, increases the probability of dispersion from the plant where the larvae began. These findings may help to understand how the A. argillacea larvae detect the Cry1Ac toxin in Bt cotton and how the toxin affects the dispersion behavior of the larvae over time. Therefore, our results are extremely important for the management of resistance in populations of A. argillacea on Bt cotton.

  20. Insect Resistance to Bacillus thuringiensis Toxin Cry2Ab Is Conferred by Mutations in an ABC Transporter Subfamily A Protein.

    Directory of Open Access Journals (Sweden)

    Wee Tek Tay

    2015-11-01

    Full Text Available The use of conventional chemical insecticides and bacterial toxins to control lepidopteran pests of global agriculture has imposed significant selection pressure leading to the rapid evolution of insecticide resistance. Transgenic crops (e.g., cotton expressing the Bt Cry toxins are now used world wide to control these pests, including the highly polyphagous and invasive cotton bollworm Helicoverpa armigera. Since 2004, the Cry2Ab toxin has become widely used for controlling H. armigera, often used in combination with Cry1Ac to delay resistance evolution. Isolation of H. armigera and H. punctigera individuals heterozygous for Cry2Ab resistance in 2002 and 2004, respectively, allowed aspects of Cry2Ab resistance (level, fitness costs, genetic dominance, complementation tests to be characterised in both species. However, the gene identity and genetic changes conferring this resistance were unknown, as was the detailed Cry2Ab mode of action. No cross-resistance to Cry1Ac was observed in mutant lines. Biphasic linkage analysis of a Cry2Ab-resistant H. armigera family followed by exon-primed intron-crossing (EPIC marker mapping and candidate gene sequencing identified three independent resistance-associated INDEL mutations in an ATP-Binding Cassette (ABC transporter gene we named HaABCA2. A deletion mutation was also identified in the H. punctigera homolog from the resistant line. All mutations truncate the ABCA2 protein. Isolation of further Cry2Ab resistance alleles in the same gene from field H. armigera populations indicates unequal resistance allele frequencies and the potential for Bt resistance evolution. Identification of the gene involved in resistance as an ABC transporter of the A subfamily adds to the body of evidence on the crucial role this gene family plays in the mode of action of the Bt Cry toxins. The structural differences between the ABCA2, and that of the C subfamily required for Cry1Ac toxicity, indicate differences in the

  1. Insect Resistance to Bacillus thuringiensis Toxin Cry2Ab Is Conferred by Mutations in an ABC Transporter Subfamily A Protein

    Science.gov (United States)

    Tay, Wee Tek; Mahon, Rod J.; Heckel, David G.; Walsh, Thomas K.; Downes, Sharon; James, William J.; Lee, Sui-Fai; Reineke, Annette; Williams, Adam K.; Gordon, Karl H. J.

    2015-01-01

    The use of conventional chemical insecticides and bacterial toxins to control lepidopteran pests of global agriculture has imposed significant selection pressure leading to the rapid evolution of insecticide resistance. Transgenic crops (e.g., cotton) expressing the Bt Cry toxins are now used world wide to control these pests, including the highly polyphagous and invasive cotton bollworm Helicoverpa armigera. Since 2004, the Cry2Ab toxin has become widely used for controlling H. armigera, often used in combination with Cry1Ac to delay resistance evolution. Isolation of H. armigera and H. punctigera individuals heterozygous for Cry2Ab resistance in 2002 and 2004, respectively, allowed aspects of Cry2Ab resistance (level, fitness costs, genetic dominance, complementation tests) to be characterised in both species. However, the gene identity and genetic changes conferring this resistance were unknown, as was the detailed Cry2Ab mode of action. No cross-resistance to Cry1Ac was observed in mutant lines. Biphasic linkage analysis of a Cry2Ab-resistant H. armigera family followed by exon-primed intron-crossing (EPIC) marker mapping and candidate gene sequencing identified three independent resistance-associated INDEL mutations in an ATP-Binding Cassette (ABC) transporter gene we named HaABCA2. A deletion mutation was also identified in the H. punctigera homolog from the resistant line. All mutations truncate the ABCA2 protein. Isolation of further Cry2Ab resistance alleles in the same gene from field H. armigera populations indicates unequal resistance allele frequencies and the potential for Bt resistance evolution. Identification of the gene involved in resistance as an ABC transporter of the A subfamily adds to the body of evidence on the crucial role this gene family plays in the mode of action of the Bt Cry toxins. The structural differences between the ABCA2, and that of the C subfamily required for Cry1Ac toxicity, indicate differences in the detailed mode

  2. [Advances in effects of insecticidal crystal proteins released from transgenic Bt crops on soil ecology].

    Science.gov (United States)

    Zhou, Xue-Yong; Liu, Ning; Zhao, Man; Li, He; Zhou, Lang; Tang, Zong-Wen; Cao, Fei; Li, Wei

    2011-05-01

    With the large scale cultivation of transgenic crops expressing Bacillus thuringiensis (Bt) insecticidal crystal proteins in the world, the problem of environmental safety caused by these Bt crops has received extensive attention. These insecticidal crystal proteins can be released into the soil continuously in the growing period of Bt plants. If their accumulation of the insecticidal crystal proteins exceeds consumption by insect larvae and degradation by the environmental factors, these insecticidal crystal proteins could constitute a hazard to non-target insects and soil microbiota. There are three main ways to release insecticidal crystal proteins into soil for Bt plants: root exudates, pollen falling, and crop reside returning. The Bt insecticidal crystal proteins released into soil can be adsorbed rapidly by active soil particles and the absorption equilibrium attained within 1-3 h. The adsorption protects Bt insecticidal crystal proteins against soil microbial degradation or enzyme degradation, which leads to remarkable prolong of the persistence of insecticidal activity. The change of soil microorganism species is an important index for evaluating the effect of Bt plants on soil ecology. The research showed that these insecticidal crystal proteins released by the Bt plant root exudates or Bt organism had no toxicity to the soil earthworms, nematodes, protozoa, bacteria and fungi; however, it could reduce the mycelium length of the arbuscular mycorrhizal fungi (AMF) and restrain AMF to form invasion unit. The influencing degree of Bt protein on soil enzyme activity varied with the releasing modes or growth period of Bt crops. Bt Cry1Ab protein can be taken up from soil by parts of following crops; however, different results were obtained with different commercial kits. To better understand the soil ecological evaluation about the insecticidal crystal proteins released from transgenic Bt crops, this review provides a comprehensive overview about the release

  3. Cross-resistance responses of CrylAc-selected Heliothis virescens (Lepidoptera: Noctuidae) to the Bacillus thuringiensis protein vip3A.

    Science.gov (United States)

    Jackson, R E; Marcus, M A; Gould, F; Bradley, J R; Van Duyn, J W

    2007-02-01

    One susceptible and three Cry1Ac-resistant strains of tobacco budworm, Heliothis virescens (F.) (Lepidoptera: Noctuidae), were used in laboratory studies to determine the level of cross-resistance between the Bacillus thuringiensis (Berliner) toxins Cry1Ac and Vip3A by using concentration-mortality and leaf tissue experiments. Concentration-mortality data demonstrated that the three Cry1Ac-resistant H. virescens strains, YHD2, KCBhyb, and CxC, were at least 215- to 316-fold resistant to Cry1Ac compared with the susceptible strain, YDK. Results from Vip3A concentration-mortality tests indicated that mortality was similar among all four H. virescens strains. Relative larval growth on Cry1Ac reflected concentration-mortality test results, because YHD2 larval growth was mostly unaffected by the Cry1Ac concentrations tested. Growth ratios for KCBhyb and CXC indicated that they had a more moderate level of resistance to Cry1Ac than did YHD2. Relative larval growth on Vip3A was highly variable at lower concentrations, but it was more consistent on concentrations of Vip3A above 25 microg/ml. Differences in larval growth among strains on Vip3A were not as pronounced as seen in Cry1Ac experiments. Mortality and larval growth also was assessed in leaf tissue bioassays in which YDK, CxC, and KCBhyb neonates were placed onto leaf disks from non-Bt and Bt cotton, Gossypium hirsutum L., for 5 d. Three Bt lines were used in an initial bioassay and consisted of two Vip3A-containing lines, COT203 and COT102, and a Cry1Ac-producing line. Mortality of KCBhyb and CXC was lower than that of YDK larvae in the presence of leaf tissue from the Cry1Ac-producing line. Additionally, increased larval growth and leaf tissue consumption on Cry1Ac-containing leaf disks was observed for KCBhyb and CXC. Mortality and larval weights were similar among strains when larvae were fed leaf tissue of either non-Bt, COT203, or COT102. A subsequent leaf tissue bioassay was conducted that evaluated four

  4. Spodoptera frugiperda (J.E. Smith) with field-evolved resistance to Bt maize are susceptible to Bt pesticides.

    Science.gov (United States)

    Jakka, S R K; Knight, V R; Jurat-Fuentes, J L

    2014-10-01

    Field-evolved resistance to maize event TC1507 expressing the Cry1Fa toxin from Bacillus thuringiensis (Bt) was detected in populations of Spodoptera frugiperda from Puerto Rico. We tested for cross-resistance to purified Cry1A toxins and commercial Bt pesticides in susceptible (Benzon) and TC1507-resistant (456) strains of S. frugiperda. Larvae from the 456 strain exhibited cross-resistance to Cry1Ab and Cry1Ac toxins, while no differences in susceptibility to XenTari WG and DiPel ES pesticides were detected. These data support cross-resistance to toxins that share binding sites with Cry1Fa and no cross-resistance to Bt pesticides in S. frugiperda with field-evolved resistance to Bt maize.

  5. Hematotoxicity and genotoxicity evaluations in Swiss mice intraperitoneally exposed to Bacillus thuringiensis (var kurstaki) spore crystals genetically modified to express individually Cry1Aa, Cry1Ab, Cry1Ac, or Cry2Aa.

    Science.gov (United States)

    Mezzomo, Bélin Poletto; Miranda-Vilela, Ana Luisa; Barbosa, Lilian Carla Pereira; Albernaz, Vanessa Lima; Grisolia, Cesar Koppe

    2016-08-01

    Bacillus thuringiensis (Bt) has been widely used in foliar sprays as part of integrated pest management strategies against insect pests of agricultural crops. Since the advent of genetically modified plants expressing Bt δ-endotoxins, the bioavailability of Cry proteins has increased, and therefore for biosafety reasons their adverse effects should be studied, mainly for nontarget organisms. We evaluated, in Swiss mice, the hematotoxicity and genotoxicity of the genetically modified strains of Bt spore crystals Cry1Aa, 1Ab, 1Ac, or 2Aa at 27 mg/kg, and Cry1Aa, 1Ab and 2Aa also at 136 and 270 mg/kg, administered with a single intraperitoneal injection 24 h before euthanasia. Controls received filtered water or cyclophosphamide. Blood samples collected by cardiac puncture were used to perform hemogram, and bone marrow was extracted for the micronucleus test. Bt spore crystals presented toxicity for lymphocytes when in higher doses, which varied according to the type of spore crystal studied, besides promoting cytotoxic and genotoxic effects for the erythroid lineage of bone marrow, mainly at highest doses. 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 indicated that these Bt spore crystals were not harmless to mice. This suggests that a more specific approach should be taken to increase knowledge about their toxicological properties and to establish the toxicological risks to nontarget organisms. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 970-978, 2016.

  6. 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

  7. [Advances in safety studies of soil Bt toxin proteins released from transgenic Bt crops].

    Science.gov (United States)

    Bai, Yaoyu; Jiang, Mingxing; Cheng, Jia; Jiang, Yonghou

    2003-11-01

    Commercialized transgenic Bt (Bacillus thuringiensis) crops are permitted for field growth in a large scale, which leads to significant issues of ecological risk assessment in soil ecosystem. In this paper, some general safety problems involving in the soil Bt active toxins released from insect-resistant transgenic Bt crops in the forms of plant residues, root exudates and pollens were reviewed, including their adsorption by soil active-particles, their insecticidal activity, persistence, and biodegradation by soil microbes, and their effects on soil organisms.

  8. EXPRESIÓN DE LA PROTEÍNA Cry1Ac EN TEJIDOS DE LÍNEAS TRANSGÉNICAS DE PAPA (Solanum tuberosum spp. ANDÍGENA VAR. DIACOL CAPIRO.

    Directory of Open Access Journals (Sweden)

    PABLO ANDRÉS VANEGAS ARAUJO

    2010-01-01

    Full Text Available La papa (Solanum sp. es el cuarto producto alimenticio más importante en el mundo. En Colombia anualmente se producen alrededor de 2,8 millones de toneladas, sirviendo como sustento económico a 90.000 familias. En el país, Tecia solanivora genera el mayor impacto económico en el cultivo con pérdidas de hasta el 100% en la producción de tubérculos. El fitomejoramiento vía introducción de genes Cry, que codifican para cristales proteicos insecticidas, constituye una alternativa para reducir el ataque de insectos en cultivos de interés comercial. En este trabajo se caracterizó la inserción, transcripción y expresión del gen Cry1Ac en diferentes tejidos y en tres etapas del desarrollo para dos líneas transgénicas de Solanum tuberosum spp. andígena variedad Diacol Capiro generadas previamente por transformación con Agrobacterium tumefaciens. La caracterización se realizó a través de técnicas de PCR, RT-PCR y ELISA. Se corroboró la inserción y transcripción del gen utilizando primers que amplificaron una banda específica de 766pb para Cry1Ac. Los niveles de expresión de la proteína llegaron a ser mayores a 45μg/g y no mostraron diferencias significativas entre las líneas analizadas, ni entre las tres etapas del desarrollo. No se evidenciaron diferencias significativas entre las líneas transgénicas con respecto al control al hacer un análisis de algunas características fenotípicas relevantes. Los resultados encontrados sugieren la realización de seguimientos y ensayos de bioseguridad sobre este material, ya que los altos niveles de expresión en todos los tejidos analizados, pueden afectar a organismos no blanco.

  9. 转苏云金杆菌毒蛋白基因玉米植株的获得及其抗虫性分析%Maize Transformation of cry1Ac3 Gene and Insect Resistance of Their Transgenic Plants

    Institute of Scientific and Technical Information of China (English)

    李慧芬; 李旭刚; 刘翔; 常团结; 肖桂芳; 朱祯

    2002-01-01

    The success for genetic transformation of maize (Zea mays L.) is highly related to genotype of target material.A few model varieties can be induced into type Ⅱ callus,which can be easily transformed with high regeneration frequency.However,most of cultivars could be only induced into type Ⅰ callus,which is difficult to be transformed with low regeneration.Thus,studying on the conditions of induction and transformation for type Ⅰ callus will show great importance for improving elite of maize directly with genetic engineering.Bacillus thuringiensis toxin protein (cry1Ac3) gene was successfully delivered into type Ⅰ calli of two elite inbred lines of maize,340 and E28,via particle bombardment in this work.Fertile transgenic corn plants were obtained through phosphinothricin (PPT) or hygromycin B (HygB) selection,and the results of PCR,Southern blot assay and ELISA showed that foreign genes had been integrated into maize genome and expressed.In the meantime,strong resistance of some transgenic plants to corn borer was showed through bioassay.In addition,the comparison of selective effect between PPT and HygB showed that PPT,as a selective agent,was better than HygB for the growth and regeneration of resistant calli.%玉米( Zea mays L.)转化成功与否与基因型密切相关.在转化过程中,除少数模式品种能够形成再生频率较高且易转化的Ⅱ型愈伤组织外,大多数栽培品种往往只能够形成再生频率较低且不易转化的Ⅰ型愈伤组织.因此探索Ⅰ型愈伤组织的诱导及其转化条件,提高转化效率,对直接改良玉米优良自交系具有重要意义.应用基因枪转化技术将苏云金杆菌( Bacillus thuringiensis ) cry1Ac3基因导入玉米优良自交系E28及340的Ⅰ型胚性愈伤组织中,经过膦丝菌素(PPT)或潮霉素(HygB)筛选,获得了再生植株.经PCR检测、Southern blot分析及Bt毒蛋白ELISA检测证实,外源基因已整合到玉米基因组中,并已获得表达.抗虫性

  10. Vip3Aa及Cry1Ac对棉铃虫幼虫多种酶活力的影响%Effects of Vip3Aa and Cry1Ac on enzyme activity in cotton bollworm Helicoverpa armigera larvae

    Institute of Scientific and Technical Information of China (English)

    张彦; 梁革梅; 高珍

    2012-01-01

    .室内生测结果表明,Vip3Aa对棉铃虫的杀虫效果低于Cry1Ac,但Vip3Aa对棉铃虫幼虫生长有明显的抑制作用.取食含Cry1Ac、Vip3Aa或Cry1Ac+Vip3Aa饲料的棉铃虫,总蛋白酶和类胰凝乳蛋白酶活性很快升高;但经Cry1Ac处理12h后这2种酶活性与对照差异不显著或低于对照,而取食含Vip3Aa饲料的棉铃虫酶活力显著高于对照的时间明显延长,而且类胰蛋白酶活性也显著高于对照;表明Cry1Ac降解速度比Vip3Aa快,可能是由于降解2种蛋白参与的酶系存在差异,同时Cry1Ac+Vip3Aa混用可以延长蛋白被酶解的时间.谷胱甘肽S-转移酶和α-乙酸萘酯酶活性在棉铃虫取食含Vip3Aa、Cry1Ac或Cry1Ac+Vip3Aa蛋白的饲料后活性升高,说明这2种酶可能参与了对Cry1Ac、Vip3Aa的解毒作用.但Cry1Ac、Vip3Aa对氨肽酶活性影响不大,可能在毒蛋白发挥毒性的过程中与氨肽酶活力变化无关.

  11. The natural refuge policy for Bt cotton (Gossypium L. in Pakistan – a situation analysis

    Directory of Open Access Journals (Sweden)

    Muhammad Sajjad Ali

    2013-07-01

    Full Text Available Bt cotton (event Cry1Ac was formally commercialized in Pakistan in 2010. However, there has been an increasing trend of planting unauthorized Bt cotton germplasm in farmers' fields since 2003 with a high rate of adoption in the core cotton areas especially in the province Punjab. The transgenic cotton technology has provided the growers with substantial economic benefits and has reduced their dependence on pesticides for pest control, especially against Helicoverpa armigera (Hubner. However, keeping in view the capacity of this insect to develop resistance against novel chemical formulations, it is easily speculated that Bt toxin, too, is no exception. Refuge crop policy for mono transgenic crop events has helped in delaying the rate of resistance evolution in the target pests. Thus, in Pakistan, where planting of structured refuge crops along Bt cotton fields is not mandatory, the effectiveness and durability of Bt cotton technology may decrease due to a number of factors which are discussed in this review.

  12. Cry1Ac抗、感棉铃虫碱性磷酸酯酶(ALP1)的表达量比较%The Expression Level of Alkaline Phosphatase (ALP1) in Cry1Ac-Resistant and Susceptible Cotton Bollworm Helicoverpa armigera Hübner

    Institute of Scientific and Technical Information of China (English)

    张涛; 张丽丽; 魏纪珍; 肖玉涛; 梁革梅; 周瑞阳

    2013-01-01

    [目的]通过比较Cry1Ac抗、感棉铃虫昆虫中肠碱性磷酸酶(ALP1)表达量的差异及抗性棉铃虫取食Cry1Ac蛋白对ALP1表达量的影响,分析ALP1表达量变化与抗性的关系,为进一步明确Bt抗性机制、制定抗性治理策略提供理论依据.[方法]利用实时荧光定量PCR技术,比较敏感棉铃虫、Cry1Ac抗性棉铃虫取食含Cry1Ac蛋白的饲料和正常饲料后ALP1表达量的差异.[结果]ALP1在棉铃虫整个发育期都表达,幼虫的表达量最高,蛹期表达量最低,在成虫体内也有较高的表达;ALP1在幼虫中肠表达量最高,其次是后肠、前肠、马氏管,表皮中的表达量最低;与敏感品系相比,对Cry1Ac具有中等水平抗性的棉铃虫ALP1表达量明显增加,尤其是取食含Cry1Ac蛋白饲料的抗性棉铃虫幼虫的ALP1的表达量显著升高,但抗性棉铃虫取食正常饲料后,2、3龄幼虫的ALP1的表达量与敏感棉铃虫差异不显著;所有的抗性品系4龄棉铃虫幼虫中肠ALP1的表达量都显著高于敏感品系,而且随着棉铃虫抗性倍数的升高,ALP1的表达量呈逐渐降低的趋势.[结论]抗性棉铃虫中肠ALP1表达量的改变可能与Cry1Ac抗性、Cry1Ac代谢有一定的关系.%[Objective] The objective of this study is to provide a theoretical basis for further clarifying Cry1Ac resistance mechanism and constituting reasonable resistance management strategy,the difference of ALP1 expression level between Cry1Ac-resistant and susceptible Helicoverpa armigera,the effect of Cry1Ac toxin on ALP1 expression level in Cry1Ac-resistant larvae and the relationship between ALP 1 and Cry 1 Ac-resistant strains were analyzed.[Method] The expression levels of ALP1 in susceptible and Cry1Ac-resistant H.armigera larvae feeding on artificial diet containing Cry1Ac and no-Cry1Ac were compared using real-time quantitative PCR analysis.[Result] The ALP1 could express in whole development period ofH.armigera,the highest expression level

  13. 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

  14. Synergism and antagonism between Bacillus thuringiensis Vip3A and Cry1 proteins in Heliothis virescens, Diatraea saccharalis and Spodoptera frugiperda.

    Directory of Open Access Journals (Sweden)

    Ana Rita Nunes Lemes

    Full Text Available 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.

  15. 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.

  16. Effects of Transgenic Brown Rice on Nutritional of Broiler Chicken%转Cry1Ac/sck基因糙米作为肉仔鸡日粮原料的营养安全性评价

    Institute of Scientific and Technical Information of China (English)

    秦海峰; 刘阳; 邢福国

    2011-01-01

    A 42-day feeding trial was employed to evaluate whether the transgenic brown rice used in broiler' s diet had any deleterious effects on gowth performance, on organ index, on carcass trait, on composition of muscle of broiler chicken. Three hunder sixty one-day old broilers were divided into three groups. One group was fed a diet containing transgenic brown rice and the other group was fed a diet containing corresponding nontransgenic brown rice. Three group was fed a diet containing corn. Growth Performance and organ index at the 21 and 42 day of age were determined, carcass trait and composition of muscle were measured at the end of the experiment. The results showed that no significant difference were observed(P>0.05) in the growth performance, in organ index and carcass trait and composition of muscle in two phases between the two groups. No histopathological damages were detected. From this study, it was concluded that there were no adverse effects of the transgenic brown rice on the broiler chicken' s health.%为了研究转Cry1Ac/sck基因糙米作为肉仔鸡日粮原料的营养安全性,将360只1日龄肉仔鸡随机分成3组.一组饲喂含转基因糙米日粮,另一组饲喂含非转基因糙米日粮,第三组饲喂非转基因玉米日粮.试验期42 d.测定肉仔鸡的生产性能,器官指数,屠体性状,肌肉成分,21,42 d测定生产性能,器官指数.42 d测定屠体性状和肌肉成分.结果显示,转基因糙米对肉仔鸡的生产性能、器官指数、屠体性状、肌肉成分和健康状况在试验期内未产生明显的有害影响(P>0.05).

  17. Insecticidal activity of residual Bt protein at the second trophic level

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Measurements were taken of Bt protein expressed in the leaves of transgenic cotton (Gossypium hirsutum) transformed with a synthesized Bt (Bacillus thuringiensis) cry1A gene and its persistent level in larval bodies and faeces of a non-targeted insect pest, beet armyworm (Spodoptera exigua). We performed enzyme linked immunosorbent assays (ELISA) and bioassays using neonate larvae of cotton bollworm (Helicoverpa armigera) to detect the insecticidal activity of residual Bt protein at the second trophic level. The results showed that Bt protein content in functional leaves was different at various developmental stages and was different among plants at the same stage. Even though Bt protein concentration in the larval bodies and faeces decreased 97.5%-99% compared to that found in cotton leaves subsequently fed to beet armyworm larvae, it still had a lethal effect on neonate cotton bollworm larvae. Therefore, Bt protein present at the second trophic level had insecticidal activity. This result is important in understanding and predicting the effect of transgenic plants on nontarget organisms.

  18. Evidence of field-evolved resistance of Spodoptera frugiperda to Bt corn expressing Cry1F in Brazil that is still sensitive to modified Bt toxins.

    Science.gov (United States)

    Monnerat, Rose; Martins, Erica; Macedo, Cristina; Queiroz, Paulo; Praça, Lilian; Soares, Carlos Marcelo; Moreira, Helio; Grisi, Isabella; Silva, Joseane; Soberon, Mario; Bravo, Alejandra

    2015-01-01

    Brazil ranked second only to the United States in hectares planted to genetically modified crops in 2013. Recently corn producers in the Cerrado region reported that the control of Spodoptera frugiperda with Bt corn expressing Cry1Fa has decreased, forcing them to use chemicals to reduce the damage caused by this insect pest. A colony of S. frugiperda was established from individuals collected in 2013 from Cry1Fa corn plants (SfBt) in Brazil and shown to have at least more than ten-fold higher resistance levels compared with a susceptible colony (Sflab). Laboratory assays on corn leaves showed that in contrast to SfLab population, the SfBt larvae were able to survive by feeding on Cry1Fa corn leaves. The SfBt population was maintained without selection for eight generations and shown to maintain high levels of resistance to Cry1Fa toxin. SfBt showed higher cross-resistance to Cry1Aa than to Cry1Ab or Cry1Ac toxins. As previously reported, Cry1A toxins competed the binding of Cry1Fa to brush border membrane vesicles (BBMV) from SfLab insects, explaining cross-resistance to Cry1A toxins. In contrast Cry2A toxins did not compete Cry1Fa binding to SfLab-BBMV and no cross-resistance to Cry2A was observed, although Cry2A toxins show low toxicity to S. frugiperda. Bioassays with Cry1AbMod and Cry1AcMod show that they are highly active against both the SfLab and the SfBt populations. The bioassay data reported here show that insects collected from Cry1Fa corn in the Cerrado region were resistant to Cry1Fa suggesting that resistance contributed to field failures of Cry1Fa corn to control S. frugiperda.

  19. Evidence of field-evolved resistance of Spodoptera frugiperda to Bt corn expressing Cry1F in Brazil that is still sensitive to modified Bt toxins.

    Directory of Open Access Journals (Sweden)

    Rose Monnerat

    Full Text Available Brazil ranked second only to the United States in hectares planted to genetically modified crops in 2013. Recently corn producers in the Cerrado region reported that the control of Spodoptera frugiperda with Bt corn expressing Cry1Fa has decreased, forcing them to use chemicals to reduce the damage caused by this insect pest. A colony of S. frugiperda was established from individuals collected in 2013 from Cry1Fa corn plants (SfBt in Brazil and shown to have at least more than ten-fold higher resistance levels compared with a susceptible colony (Sflab. Laboratory assays on corn leaves showed that in contrast to SfLab population, the SfBt larvae were able to survive by feeding on Cry1Fa corn leaves. The SfBt population was maintained without selection for eight generations and shown to maintain high levels of resistance to Cry1Fa toxin. SfBt showed higher cross-resistance to Cry1Aa than to Cry1Ab or Cry1Ac toxins. As previously reported, Cry1A toxins competed the binding of Cry1Fa to brush border membrane vesicles (BBMV from SfLab insects, explaining cross-resistance to Cry1A toxins. In contrast Cry2A toxins did not compete Cry1Fa binding to SfLab-BBMV and no cross-resistance to Cry2A was observed, although Cry2A toxins show low toxicity to S. frugiperda. Bioassays with Cry1AbMod and Cry1AcMod show that they are highly active against both the SfLab and the SfBt populations. The bioassay data reported here show that insects collected from Cry1Fa corn in the Cerrado region were resistant to Cry1Fa suggesting that resistance contributed to field failures of Cry1Fa corn to control S. frugiperda.

  20. Can interactions between Bt proteins be predicted and how should effects on non-target organisms of GM crops with multiple Bt Proteins be assessed?

    NARCIS (Netherlands)

    Schrijver, De A.; Clercq, de P.; Booij, K.; Maagd, de R.A.; Frankenhuyzen, van K.

    2014-01-01

    Genes expressing Bacillus thuringiensis (Bt) toxins have been incorporated into genetically modified (GM) plants to render these resistant to certain insect pests. Of particular interest have been the genes encoding Cry (Crystal) proteins, but also the gene encoding the vegetative insecticidal prote

  1. Bt resistance in Australian insect pest species.

    Science.gov (United States)

    Downes, Sharon; Walsh, Tom; Tay, Wee Tek

    2016-06-01

    Bt cotton was initially deployed in Australia in the mid-1990s to control the polyphagous pest Helicoverpa armigera (Hübner) which was intractably resistant to synthetic chemistries. A conservative strategy was enforced and resistance to first generation single toxin technology was managed. A decade later, shortly after the release of dual toxin cotton, high baseline frequencies of alleles conferring resistance to one of its components prompted a reassessment of the thinking behind the potential risks to this technology. Several reviews detail the characteristics of this resistance and the nuances of deploying first and second generation Bt cotton in Australia. Here we explore recent advances and future possibilities to estimate Bt resistance in Australian pest species and define what we see as the critical data for enabling effective pre-emptive strategies. We also foreshadow the imminent deployment of three toxin (Cry1Ac, Cry2Ab, Vip3A) Bollgard 3 cotton, and examine aspects of resistance to its novel component, Vip3A, that we believe may impact on its stewardship.

  2. Proteomic analysis of Cry2Aa-binding proteins and their receptor function in Spodoptera exigua

    Science.gov (United States)

    Qiu, Lin; Zhang, Boyao; Liu, Lang; Ma, Weihua; Wang, Xiaoping; Lei, Chaoliang; Chen, Lizhen

    2017-01-01

    The bacterium Bacillus thuringiensis produces Crystal (Cry) proteins that are toxic to a diverse range of insects. Transgenic crops that produce Bt Cry proteins are grown worldwide because of their improved resistance to insect pests. Although Bt “pyramid” cotton that produces both Cry1A and Cry2A is predicted to be more resistant to several lepidopteran pests, including Spodoptera exigua, than plants that produce Cry1Ac alone, the mechanisms responsible for the toxicity of Cry2Aa in S. exigua are not well understood. We identified several proteins that bind Cry2Aa (polycalin, V-ATPase subunits A and B, actin, 4-hydroxybutyrate CoA-transferase [4-HB-CoAT]), and a receptor for activated protein kinase C (Rack), in S. exigua. Recombinant, expressed versions of these proteins were able to bind the Cry2Aa toxin in vitro assays. RNA interference gene knockdown of the Se-V-ATPase subunit B significantly decreased the susceptibility of S. exigua larvae to Cry2Aa, whereas knockdown of the other putative binding proteins did not. Moreover, an in vitro homologous competition assay demonstrated that the Se-V-ATPase subunit B binds specifically to the Cry2Aa toxin, suggesting that this protein acts as a functional receptor of Cry2Aa in S. exigua. This the first Cry2Aa toxin receptor identified in S. exigua brush-border membrane vesicles. PMID:28067269

  3. 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.

  4. A high-throughput liquid bead array-based screening technology for Bt presence in GMO manipulation.

    Science.gov (United States)

    Fu, Wei; Wang, Huiyu; Wang, Chenguang; Mei, Lin; Lin, Xiangmei; Han, Xueqing; Zhu, Shuifang

    2016-03-15

    The number of species and planting areas of genetically modified organisms (GMOs) has been rapidly developed during the past ten years. For the purpose of GMO inspection, quarantine and manipulation, we have now devised a high-throughput Bt-based GMOs screening method based on the liquid bead array. This novel method is based on the direct competitive recognition between biotinylated antibodies and beads-coupled antigens, searching for Bt presence in samples if it contains Bt Cry1 Aa, Bt Cry1 Ab, Bt Cry1 Ac, Bt Cry1 Ah, Bt Cry1 B, Bt Cry1 C, Bt Cry1 F, Bt Cry2 A, Bt Cry3 or Bt Cry9 C. Our method has a wide GMO species coverage so that more than 90% of the whole commercialized GMO species can be identified throughout the world. Under our optimization, specificity, sensitivity, repeatability and availability validation, the method shows a high specificity and 10-50 ng/mL sensitivity of quantification. We then assessed more than 1800 samples in the field and food market to prove capacity of our method in performing a high throughput screening work for GMO manipulation. Our method offers an applicant platform for further inspection and research on GMO plants.

  5. Biological activity of Bt proteins expressed in different structures of transgenic corn against Spodoptera frugiperda

    Directory of Open Access Journals (Sweden)

    Daniel Bernardi

    2016-06-01

    Full Text Available ABSTRACT: Spodoptera frugiperda (J. E. Smith is the main target pest of Bt corn technologies, such as YieldGard VT PRO(tm (Cry1A.105/Cry2Ab2 and PowerCore(tm (Cry1A.105/Cry2Ab2/Cry1F. In this study, it was evaluated the biological activity of Bt proteins expressed in different plant structures of YieldGard VT PRO(tm and PowerCore(tm corn against S. frugiperda . Complete mortality of S. frugiperda neonates was observed on leaf-disc of both Bt corn technologies. However, the mortality in silks and grains was lower than 50 and 6%, respectively. In addition, more than 49% of the surviving larvae in silks and grains completed the biological cycle. However, all life table parameters were negatively affected in insects that developed in silks and grains of both Bt corn events. In summary, the low biological activity of Bt proteins expressed on silks and grains of YieldGard VT PRO(tm and PowerCore(tm corn can contribute to the resistance evolution in S. frugiperda populations.

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

    Science.gov (United States)

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

    2016-05-05

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

  7. Identification and chromosomal mapping of insect-resistant gene Bt in up-land cotton%陆地棉Bt抗虫基因类型鉴定与染色体定位

    Institute of Scientific and Technical Information of China (English)

    安百伟; 赵亮; 狄佳春; 陈旭升

    2016-01-01

    In order to define the inheritance of insect⁃resistant traits, insect⁃resistant gene types and the position of the insect⁃resistant gene on chromosomes in a Bt transgenic cotton germplasm N099, the insect resistance and susceptibility was evaluated in a F2 population of non⁃resistant N099 and resistant Hai7124 ( Gossypium barbadense) . Results showed that the resistance was controlled by a single pair of dominant gene. N099 was identified by Bt protein strip and DNA⁃PCR to contain bivalent Bt insect⁃resistant gene, Cry1Ac+Cry2A. A pair of polymorphic primer NAU2912 two hundred and thirty⁃four pairs of primers covering 26 cotton chromosomes was found in a near⁃isogenic pool consisting of resistant&susceptible F2 cotton plants to be linked with the Bt gene Cry1Ac+Cry2A. Known the primer on cotton chromosome 26, then other prim⁃ers on the chromosome flanking 50 cM from the primer were screened, which received 16 pairs of primers linked with the Bt gene. It was located between SSR marker dc40260 and cgr6702, with genetic distance 1�8 cM and 2�9 cM respectively. Thus, the bivalent Bt gene is mapped on the cotton chromosome 26.%为明确Bt抗虫棉种质系N099的抗虫性状遗传方式、抗虫基因类型以及抗虫基因插入染色体的位置,用N099为父本与非抗虫海岛棉海7124杂交,分析了F2分离世代遗传方式,结果表明该抗虫棉的抗虫性状呈简单的1对显性基因控制的质量遗传方式。进一步在Bt毒蛋白和DNA 2个层面对该抗虫基因类型进行鉴定,结果显示N099所含抗虫基因为双价Bt基因Cry1Ac+Cry2A。而后利用234对核心引物对F2代抗、感株组成的近等基因混池进行多态性筛选,发现其中1对多态性引物NAU2912与该Bt基因连锁。已知引物NAU2912位于棉花第26染色体上,通过对该引物上、下50�0 cM之间的其他引物进行多态性筛选,共获得16对引物与双价Bt基因相连锁,目

  8. Degradation of Cry1Ab Protein Within Transgenic Bt Maize Tissue by Composite Microbial System of MC1

    Institute of Scientific and Technical Information of China (English)

    Meng Yao; Gu Wan-rong; Ye Le-fu; Chen Dong-sheng; Li Jing; Wei Shi

    2014-01-01

    Environmental safety issues involved in transgenic plants have become the concern of researchers, practitioners and policy makers in recent years. Potential differences between Bt maize (ND1324 and ND2353 expressing the insecticidal Cry1Ab protein) and near-isogenic non-Bt varieties (ND1392 and ND223) in their influence on the composite microbial system of MC1 during the fermentation process were studied during 2011-2012. Cry1Ab protein in Bt maize residues didn't affect characteristics of lignocellulose degradation by MC1, pH of fermentation broth decreasing at initial stage and increasing at later stage of degradation. The quality of various volatile products in fermentation broth showed that no significant difference of residues fermentation existed between Bt maize and non-Bt maize. During the fermentation MC1 efficiently degraded maize residues by 83%-88%, and cellulose, hemicelluloses and lignin content decreased by 70%-72%, 72%-75% and 30%-37%, respectively. Besides that, no consistent difference was found between Bt and non-Bt maize residues lignocellulose degradation by MC1 during the fermentation process. MC1 degraded 88%-89% Cry1Ab protein in Bt maize residues, and in the fermentation broth of MC1 and bacteria of MC1 Cry1Ab protein was not detected. DGGE profile analyses revealed that the microbial community drastically changed during 1-3 days and became stable until the 9th day. Though the dominant strains at different fermentation stages had significantly changed, no difference on the dominant strains was observed between Bt and non-Bt maize at different stages. Our study indicated that Cry1Ab protein did not influence the growth characteristic of MC1.

  9. Bt基因和Bt蛋白检测技术研究进展%Research on Progress of the Detection of BT Protein and BT Gene

    Institute of Scientific and Technical Information of China (English)

    汪倩; 向晓玲; 许耀心; 倪晓强; 姜佳燕; 龚云飞

    2012-01-01

    BT protein is a kind of specific insecticidal activity of protein crystallization produced by bacillus. Through the transgenic technology, we switch Bt gene into the crops such as rice and soybean to cultivate a good resistance to insects breed, then without using pesticides and some other harmful substances, the plant diseases and insect pests can be prevented and cured efficiently. However, animal experiment and clinical study found that Bt protein may damage mammalian immune organs and immune cells, and may also influence the genetic structure of the gene pool and genetic diversity of the popula- tion. It also has serious influence on the soil specific organisms function, soil biodiversity, and the soil enzyme activity and so on. What' s more, Bt protein in transgenic rice may enrich in human bodies along the food chain and do potential harm to humans. Therefore, the research on detecting of Bt genc or protein is necessary. So, the research status of Bt gene and pro- tein are briefly reviewed in this paper.%通过转基因技术将可表达杀虫特性蛋白的Bt基因转入大豆、水稻等农作物中,培育出优良的抗虫品种,从而在不使用农药等有害物质的前提下,对病虫害起到高效防治作用。然而动物试验及临床研究发现,Bt蛋白可能导致哺乳动物的免疫器官和免疫细胞损伤,也可能影响基因库的遗传结构及群体遗传多样性,对土壤特异生物类群功能、土壤生物多样性以及土壤酶活性等也有不同程度的影响,且Bt蛋白有可能沿着食物链在人体内富集,对人体形成潜在的危害。因此,开展对Bt基因和Bt蛋白的监控研究是必要的,该文对Bt的相关研究现状作一简要综述。

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

    Science.gov (United States)

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

    2009-07-01

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

  11. Bioinformatic analysis for allergenicity assessment of Bacillus thuringiensis Cry proteins expressed in insect-resistant food crops.

    Science.gov (United States)

    Randhawa, Gurinder Jit; Singh, Monika; Grover, Monendra

    2011-02-01

    The novel proteins introduced into the genetically modified (GM) crops need to be evaluated for the potential allergenicity before their introduction into the food chain to address the safety concerns of consumers. At present, there is no single definitive test that can be relied upon to predict allergic response in humans to a new protein; hence a composite approach to allergic response prediction is described in this study. The present study reports on the evaluation of the Cry proteins, encoded by cry1Ac, cry1Ab, cry2Ab, cry1Ca, cry1Fa/cry1Ca hybrid, being expressed in Bt food crops that are under field trials in India, for potential allergenic cross-reactivity using bioinformatics search tools. The sequence identity of amino acids was analyzed using FASTA3 of AllergenOnline version 10.0 and BLASTX of NCBI Entrez to identify any potential sequence matches to allergen proteins. As a step further in the detection of allergens, an independent database of domains in the allergens available in the AllergenOnline database was also developed. The results indicated no significant alignment and similarity of Cry proteins at domain level with any of the known allergens revealing that there is no potential risk of allergenic cross-reactivity.

  12. Influence of the Ingestion of Cry1Ac Toxin on Trypsin-Like Protease in Helicoverpa Armigera Midgut%喂食Cry1Ac毒素对棉铃虫中肠类胰蛋白酶的影响

    Institute of Scientific and Technical Information of China (English)

    袁灿; 夏立秋; 丁学知; 田野; 尹佳

    2009-01-01

    苏云金芽胞杆菌(Bacillus thuringiensis,Bt)在芽胞形成过程中产生的原毒素被敏感昆虫吞食进入消化道后,在中肠蛋白酶的作用下经历一系列酶的剪切活化过程,形成具有杀虫活性的毒性肽.类胰蛋白酶因其在昆虫中肠内蛋白质水解和Bt毒素活化中起着重要作用而受到广泛研究.本研究提取了棉铃虫4龄幼虫的中肠BBMV蛋白,利用SDS-PAGE分析鉴定到喂食Cry1Ac毒素24 h后,棉铃虫幼虫中肠类胰蛋白酶的表达量呈明显下降趋势.该下降趋势很有可能是作为一种昆虫对Bt毒素作用的早期反应,通过影响Bt毒素在中肠中的酶解活化过程,从而为昆虫提供即时保护.

  13. Survival and Development of Spodoptera frugiperda and Chrysodeixis includens (Lepidoptera: Noctuidae) on Bt Cotton and Implications for Resistance Management Strategies in Brazil.

    Science.gov (United States)

    Sorgatto, Rodrigo J; Bernardi, Oderlei; Omoto, Celso

    2015-02-01

    In Brazil, Spodoptera frugiperda (J. E. Smith) and Chrysodeixis includens (Walker) are important cotton pests and target of control of Bollgard II (Cry1Ac/Cry2Ab2) and WideStrike (Cry1Ac/Cry1F) cotton technologies. To subsidize an insect resistance management program, we conducted laboratory studies to evaluate the toxicity of these Bt cotton plants throughout larval development of S. frugiperda and C. includens. In bioassays with leaf disc, the efficacy of both Bt cotton plants against neonates was >80% for S. frugiperda and 100% for C. includens. However, S. frugiperda larvae that survived on Bt cotton had >76% of growth inhibition and stunting. In bioassays with S. frugiperda and C. includens larvae fed on non-Bt near-isoline during different time period (from 3 to 18 d) and then transferred to Bollgard II or WideStrike leaves showed that larval susceptibility decreased as larval age increased. For Bollgard II cotton, in all S. frugiperda instars, there were larvae that reached the pupal and adult stages. In contrast, on WideStrike cotton, a few larvae in fifth and sixth instar completed the biological cycle. For C. includens, some larvae in sixth instar originated adults in both Bt cotton plants. In conclusion, Bollgard II and WideStrike cotton technologies showed high efficacy against neonates of S. frugiperda and C. includens. However, the mortality of these species decreases as larval age increase, allowing insect survival in a possible seed mixture environment and favoring the resistance evolution.

  14. 转双Bt基因巨霸杨外源基因表达及抗虫性检测%Exogenous Gene Expression and Insect Resistance Detection of Transgenic Populus deltocdes ‘55/56’ ×P.deltoc-descv‘2KEN8’ with Double Bt Gene

    Institute of Scientific and Technical Information of China (English)

    赵洁; 杜沙沙; 邱彤; 杨敏生; 王进茂

    2016-01-01

    To improve the efficiency of genetically modified ( gm) polar Bt gene expression and expand the insect-resistant spec-trum, we chose the Populu s deltocdes‘55/56’ ×P.deltocdescv‘2KEN8’ with doubleB t gene of Cr y1Ac and Cry3A of one-year seedlings as materials , and tested the expression of exogenous genes and insect resistance .The purpose gene was inte-grated into the genome in the P.deltocdes‘ 55/56’ ×P.deltocdecsv ‘2KEN8’ by PCR identification .We used the fluores-cent quantitative PCR technique and ELISA to detect the four gm strains transcription abundance Bt toxic protein gene ex-pression content in the leaves .The difference existed between the two Bt gene transcription abundance in different strains:Cry3A gene transcription abundance was significantly higher than Cry1Ac.Two kinds of Bt toxic protein expression quantity were significantly different between each strain .Cry3A toxic protein levels in each strain were significantly higher than Cry1Ac.Each strains’ insect-resistant effect to lepidoptera pests , larvae (Hyphantria cunea) was not obvious with no sig-nificant difference between different strains .The strains showed very high insect-resistant effect to coleoptera pests ( Pla-giodera versicolora), and were significantly higher than that of single gene Cry3A poplars high strain CC84, and there was significant difference between different strains .%为提高转基因杨树中Bt基因的表达效率并扩大抗虫谱,以同时转入Cry1Ac和Cry3A基因的转双Bt基因巨霸杨( Populus deltocdes‘55/56’×P.deltocdes‘2KEN8’)株系1年生苗为材料,对外源基因的表达和抗虫性进行检测。经PCR检测,证明目的基因已被整合到巨霸杨基因组中。利用荧光定量PCR和ELISA技术检测了4个转基因株系叶片中Bt基因的转录丰度和毒蛋白表达量。结果表明:不同株系间Bt基因的转录丰度存在显著差异,Cry3A基因的转录丰度显著高于Cry1Ac

  15. Impacts of Bt rice expressing Cry1C or Cry2A protein on the performance of nontarget leafhopper, Nephotettix cincticeps (Hemiptera: Cicadellidae), under laboratory and field conditions.

    Science.gov (United States)

    Lu, Z B; Tian, J C; Wang, W; Xu, H X; Hu, C; Guo, Y Y; Peng, Y F; Ye, G Y

    2014-02-01

    Transgenic rice expressing Bacillus thuringiensis Berliner (Bt) protein can effectively control target insects including stem borers and leaf folders. However, the potential effects of Bt rice on nontarget organisms including nontarget herbivores have not been fully evaluated. In the current study, ecological fitness parameters of the nontarget herbivore, Nephotettix cincticeps (Uhler) (Hemiptera: Cicadellidae), fed on T1C-19 (Cry1C) or T2A-1 (Cry2A) rice were compared with non-Bt rice (MH63) under laboratory conditions. A 2-yr field trial was also conducted to monitor the population dynamics of N. cincticeps in the Bt and control rice plots using the vacuum-suction machine and yellow sticky card traps. Laboratory results showed that there were no significant differences in some of biological parameters including egg developmental duration, adult fresh weight, adult longevity, and oviposition period when N. cincticeps fed on Bt or non-Bt rice was compared. However, the survival rate of N. cincticeps nymphs fed on T2A-1 Bt rice plants was significantly higher than that on the control. When N. cincticeps fed on T1C-19 Bt rice plants, its nymphal duration was significantly longer and fecundity significantly lower compared with those fed on both T2A-1 Bt and non-Bt rice plants; the preoviposition period of N. cincticeps fed on T1C-19 and T2A-1 Bt rice was also significantly shorter than those on non-Bt rice. Nonetheless, both seasonal density and population dynamics of N. cincticeps adults and nymphs were similar between Bt (T1C-19 and T2A-1) and non-Bt rice plots under field conditions. In conclusion, our results indicate that our two tested Bt rice lines would not lead to higher population of N. cincticeps. Long-term experiments to monitor the population dynamics of N. cincticeps at large scale need to be carried out to confirm the current results.

  16. Prediction on Antigenic Epitope Characteristics of Bt Cry2Ab Protein in Transgenic Crops

    Institute of Scientific and Technical Information of China (English)

    Jierong GAO; Ying HE; Zehong ZOU; Ailin TAO; Yuncan AI

    2012-01-01

    Abstract [Objective] This study aimed to predict the structural characteristics of Bt Cry2Ab protein in transgenic crops with bioinformatic analysis to provide the theoreti- cal clues for design of antibody Cry2Ab. [Method] The amino acid sequence of Cry2Ab protein was searched from NCBI database. The B cell epitopes were pre- dicted with DNAStar. The binding affinity between Cry2Ab protein and MHC-II molecules was analyzed with NetMHCII 2.2 Server to predict the T cell epitopes. [Result] Prediction result suggested the potential B cell epitope of Cry2Ab locating in the region of 208-215. Analysis of the binding affinity between Cry2Ab and MHC-II molecules suggested the regions of 177-185, 299-307 and 255-263 were the po- tential T cell epitopes. Human with HLA-DRB10101 alleles and HLA-DRB10701 al- leles were more sensitive to Cry2Ab protein. [Conclusion] This study facilitates to un- derstand the structural characteristics of Cry2Ab protein and provides a new clue to improve the assessment method for potential allergenicity of genetically modified food.

  17. [Bt transgenic crops for insect-resistance and modification of Bt protein and utilization of stacking strategy].

    Science.gov (United States)

    Li, Chen; Liu, Bolin

    2015-01-01

    Insecticidal protein genes from Bacillus thuringiensis are currently the most widely used insect-resistant genes. They have been transferred to many crops for breeding and production. Among them, cotton, maize, potato and other insect-resistant crops are commercialized, creating considerable economic benefit. In this review, we summarized advances in identifying functional genes and transgenic crops for insect resistance, compared different strategies for enhancing vigor of insecticidal protein and utilizing gene stacking as well as listing valuable groups of stacked genes. In addition, the methods for multiple gene transformation was discussed.

  18. 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.

  19. Studies on Rapid Quantitative Analysis of Bt Toxin by Using Envirologix Kits in Transgenic Rice

    Institute of Scientific and Technical Information of China (English)

    XIE Xiao-bo; SHU Qing-yao

    2002-01-01

    Investigations were done on the usefulness of Envirologix Cry1Ab/Cry1Ac Plate kits for quantitative analysis of Bt toxin content in transgenic rice grains. Two transgenic rice lines: Kemingdao 1 (KMD1) and Kemingdao 2 (KMD2), transformed with a cry1Ab gene, and their parental variety, cv.Xiushui 11, were used as positive and negative samples. Results showed that the correlation coefficients as high as 0. 985 - 0. 998, significant at probability level of 0.05 or 0.01, were obtained for linear regression equations by using the appended calibrators of the kits. No significant differences were detected for values of same rice sample obtained from different trials or by using different lots of the product (kit). The detectable Bt toxin content by this method could be as low as 0.5 ng/g. The Envirologix Kit could be useful for rapid quantitative detection of Bt toxin in rice grains because of its preciseness, simplicity and time saving.

  20. [Occurrence dynamics of migratory pest insects Cnaphalocrocis medinalis and Sogatella furcifera in transgenic Bt rice field in Xing'an County of Guangxi Province].

    Science.gov (United States)

    Sui, He; Li, Zhi-Yi; Xu, Yan-Bo; Han, Chao; Han, Lan-Zhi; Chen, Fa-Jun

    2011-11-01

    An investigation was conducted in Xing' an County of Guangxi Province in 2010 to study the occurrence and damage characteristics of two unique migratory pest insects, rice leaffolder (Cnaphalocrocis medinalis) and white-backed planthopper (Sogatella furcifera), in a transgenic Bt rice (cv. HH1 with dual Cry1Ab+Cry1Ac genes) field, taking the corresponding non-transgenic parent (cv. MH63) field as the control. No significant differences were observed in the abundance of C. medinalis eggs and larvae in the two fields, but the percentage of fold-leaf plants and the fold-leaf rate per plant were significantly lower in transgenic Bt rice field than in the control, suggesting that transgenic Bt rice had higher resistance against the target pest insect C. medinalis. As for S. furcifera, its occurrence dynamics of nymphs, adults (including macro- and brachypterous forms), and whole population had no significant differences between the two fields, but the abundance of the nymphs and brachypterous adults at the peak stage of S. furcifera occurrence was obviously higher in transgenic Bt rice field than in the control, while the macropterous abundance was in adverse. The sex ratio of female of the macropterous adults at the late growth stage of rice was generally lower in transgenic Bt rice field than in the control. These results suggested that under the background of large area commercial production of transgenic Bt rice, the occurrence and harm of the non-target pest insect S. furcifera could become more complicated.

  1. Toxicity of Bacillus thuringiensis (L.) Cry proteins against summer fruit tortrix (Adoxophyes orana - Fischer von Rösslerstamm).

    Science.gov (United States)

    Radosavljevic, Jelena; Naimov, Samir

    2016-07-01

    The activity of seven Cry1, one Cry9 and one hybrid Cry1 protoxins against neonate larvae of summer fruit tortrix (Adoxophyes orana - Fischer von Rösslerstamm) has been investigated. Cry1Ia is identified as the most toxic protein, followed by Cry1Aa and Cry1Ac. Cry1Ca, Cry1Cb, Cry1Da and Cry1Fa were less active, while SN19 (Cry1 hybrid protein with domain composition 1Ba/1Ia/1Ba) and Cry9Aa exhibited negligible toxicity against A. orana. In vitro trypsin-activated Cry1Ac is still less active than Cry1Ia protoxin, suggesting that toxicity of Cry1Ia is most probably due to more complex differences in further downstream processing, toxin-receptor interactions and pore formation in A. orana's midgut epithelium.

  2. Generation of insect-resistant and glyphosate-tolerant rice by introduction of a T-DNA containing two Bt insecticidal genes and an EPSPS gene.

    Science.gov (United States)

    Zhao, Qi-chao; Liu, Ming-hong; Zhang, Xian-wen; Lin, Chao-yang; Zhang, Qing; Shen, Zhi-cheng

    2015-10-01

    Insect resistance and glyphosate tolerance have been two of the most important traits in the genetic improvement of various crops. In this study, two Bacillus thuringiensis (Bt) insecticidal genes, Cry1Ac and Cry1Ig, and a modified glyphosate-tolerant 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene (G10) were combined into a single transferred DNA (T-DNA) fragment and introduced into rice by Agrobacterium-mediated transformation. A transgenic line with single-copy T-DNA insertion named GAI-14 was found to be highly resistant to striped stem borer and rice leaf roller, and tolerant to glyphosate. Analysis of T-DNA border sequence suggested that the transgenes were inserted at the chromosome 3 and appeared to have not interrupted any known or putative genes. A field trial observed no significant difference in the basic agronomic traits between GAI-14 and the recipient rice.

  3. Bacillus thuringiensis (Bt)

    Science.gov (United States)

    2004-01-01

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

  4. Resistance to Bt maize in Mythimna unipuncta (Lepidoptera: Noctuidae) is mediated by alteration in Cry1Ab protein activation.

    Science.gov (United States)

    González-Cabrera, Joel; García, Matías; Hernández-Crespo, Pedro; Farinós, Gema P; Ortego, Félix; Castañera, Pedro

    2013-08-01

    Bt maize cultivars based on the event MON810 (expressing Cry1Ab) have shown high efficacy for controlling corn borers. However, their efficiency for controlling some secondary lepidopteran pests such as Mythimna unipuncta has been questioned, raising concerns about potential outbreaks and its economic consequences. We have selected a resistant strain (MR) of M. unipuncta, which is capable of completing its life cycle on Bt maize and displays a similar performance when feeding on both Bt and non-Bt maize. The proteolytic activation of the protoxin and the binding of active toxin to brush border membrane vesicles were investigated in the resistant and a control strain. A reduction in the activity of proteolytic enzymes, which correlates with impaired capacity of midgut extracts to activate the Cry1Ab protoxin has been observed in the resistant strain. Moreover, resistance in larvae of the MR strain was reverted when treated with Cry1Ab toxin activated with midgut juice from the control strain. All these data indicate that resistance in the MR strain is mediated by alteration of toxin activation rather than to an increase in the proteolytic degradation of the protein. By contrast, binding assays performed with biotin labelled Cry1Ab suggest that binding to midgut receptors does not play a major role in the resistance to Bt maize. Our results emphasize the risk of development of resistance in field populations of M. unipuncta and the need to consider this secondary pest in ongoing resistance management programs to avoid the likely negative agronomic and environmental consequences.

  5. Study on Spatial-temporal Dynamics of Bt Toxic Protein Expression in Insect-resistant Transgenic Cotton and Its Degradation in Soil

    Institute of Scientific and Technical Information of China (English)

    Yiwen ZHANG; Lianrong WANG; Liancheng ZHANG; Jun ZHANG; Xinbo JI; Jinmao WANG

    2012-01-01

    [Ob.jcctive] This study aimed to investigate the spatial-temporal dynamics of Bt toxic protein expression in insect-resistant transgenic cotton and its degradation in soil. [Meth~d] BtcrylAc toxic protein expression in roots, stems and leaves of trans- genic cotton Guoshen GK45 at different developmental stages and the annual aver- age content of BtCrylAc toxin protein in the topsoil, rhizosphere soil and following cotton-growing area were explored and analyzed by using enzyme linked immuno sorbed assay (ELISA). [Result] The content of exogenous BtCrylAc toxin protein de- creased during the growth process of insect-resistant transgenic cotton; to be specif- ic, the content of BtCrylAc toxin protein in cotton stems and leaves decreased more slowly and always maintained a high level, while that in roots decreased rapidly and reached a minimum level to the following plant growth and development stage. BtCrylAc toxin protein was detected in topsoil of both non-transgenic and transgenic cotton-growing areas, and the content of BtCrylAc toxin protein increased in topsoil of following cotton-growing area, which was very low in rhizosphere soil. [Conclusion] Determination of BtcrylAc toxic protein provides scientific basis for the risk assess- ment of the cultivation of genetically modified crops and the safety evaluation of soil ecosystem.

  6. Isolation and Identification of Bacillus thuringensis (Bt) from Coalfields%煤矿中苏云金芽胞杆菌(Bt)菌株的分离及其鉴定

    Institute of Scientific and Technical Information of China (English)

    林白容; 章秋萍; 王毅斌; 关雄

    2014-01-01

    (2-DE/MS) showed that isolated Bt strain encoded approximately 130 kD crystal protein, which was then identified as insecticidal protein Cry1Ac by mass spectrometry (MALDI-TOF/TOF-MS). Molecular chaperone GroEL, ATP F0F1 synthase subunit beta, elongation factor Tu and pyruvate dehydrogenase were identified by MALDI-TOF/TOF-MS, respectively. Further understanding of protein functions were conducted through bioinformatics analysis, leading to correct folding of nascent peptides, energy production and conversion, as well as improving the ability of cells to withstand adverse external stimuli. This work provides basic data for discovering new Bt resources, new insecticidal protein and using microbial to control heavy metal pollution.

  7. Relationship Between Leaf C/N Ratio and Insecticidal Protein Expression in Bt Cotton as Affected by High Temperature and N Rate

    Institute of Scientific and Technical Information of China (English)

    ZHANG Xiang; LÜ Chun-hua; CHEN Yuan; WANG Gui-xia; CHEN Yuan; CHEN De-hua

    2014-01-01

    Expression of insecticidal protein for transgenic Bacillus thuringiensis (Bt) cotton is unstable and related to nitrogen metabolism. The objective of this study was to investigate the relationship between leaf carbon nitrogen ratio (C/N) and insecticidal efifcacy of two Bt cotton cultivars. C/N ratio and Bt protein content were both measured at peak square period and peak boll period respectively under 5-7 d high temperature and different nitrogen fertilizer rates on the Yangzhou University Farm and the Ludong Cotton Farm, China. All plants were grown in ifeld. The results showed that the C/N ratio enhanced slightly and the Bt protein content remained stable at peak square period, but significant increases for the C/N ratio and decreases markedly for the leaf Bt protein concentration were detected at the peak boll period. The similar patterns at the two growth periods were found for the leaf C/N ratio and Bt protein content by different N fertilizer treatments. When nitrogen rate was from 0 to 600 kg ha-1, the C/N ratio was reduced by 0.017 and 0.006 for Sikang 1 and Sikang 3 at peak square period, compared to the 1.350 to 1.143 reduction for Sikang 1 and Sikang 3 at peak boll period, respectively. Correspondingly, the leaf Bt protein contents were bolstered by 2.6-11.8 and 26.9-36.9% at the two different growth periods, respectively. The results suggested that enhanced C/N ratio by high temperature and nitrogen application may result in the reduction of insectiocidal efifcacy in Bt cotton, especially in peak boll period.

  8. Comparison of exogenous gene expression and insect-resistance ability of transgenic 741 poplars with single and double Bt genes%转单、双Bt基因741杨外源基因表达和抗虫性比较

    Institute of Scientific and Technical Information of China (English)

    王桂英; 杨敏生; 霍雪梅; 刘晓杰

    2012-01-01

    [目的]研究联合使用两种或两种以上的抗虫基因的抗虫效果,同时鉴定并筛选出转双Bt基因741杨对鳞翅目和鞘翅目害虫有较强抗性的株系.[方法]选取转三基因(Cry3Aa+ Cry1Ac+ API)741杨8个株系、转双基因(Cry1Ac+ API)741杨1个株系和转单基因(Cry3Aa)741杨3个株系为试材,从外源基因PCR检测、毒蛋白表达和抗虫性三方面对转基因株系进行对比分析.[结果]经PCR扩增后各转基因株系出现了预期的电泳条带.ELISA蛋白检测显示转基因株系中都有与所含基因相应的Bt杀虫蛋白表达.用转基因株系新鲜叶片进行柳蓝叶甲Plagiodera versicolora和美国白蛾Hyphantria cunea室内饲虫实验表明:转入不同抗虫基因的杨树对昆虫的抗性具有选择性,对非靶标昆虫没有毒杀作用.转双Bt基因741杨具有双抗性,不同转基因株系表现出高中低的抗性水平:在对柳蓝叶甲的抗性上,筛选出的其中5个高抗株系(pCCA1,pCCA2,pCCA5,pCCA6和pCCA9)的抗性水平明显比含Cry3Aa单Bt基因的3个高抗株系(pCC11,pCC53和pCC84)高;在对美国白蛾的抗性上,有7个株系(pCCA2 ~pCCA7和pCCA9)的抗性水平与含Cry1Ac单Bt基因株系(pB29)表现一致,只有1个株系(pCCA1)对美国白蛾表现出了极低的抗性.[结论]多个抗虫基因在741杨上的联合使用,不仅扩大了抗虫谱,其中的高抗株系还具有了更高的抗虫能力,有效地发挥了基因的叠加效应.%[ Aim ] To study the combining effects of two or more insect-resistance genes in plants, as well as to screen and confirm double Bt transgenic 741 poplar lines with high resistance against coleopteran and lepidopteran insects. [Methods] Eight lines with three transgenes ( CryiAa + Cry1Ac + API) , one line with double transgenes ( CryXAc +API) and three lines with single transgene ( CryiAa) of transgenic 741 poplar were used as experimental materials. Comparative studies were conducted from three aspects; PCR

  9. Transfer of Bt-toxin protein gene into maize by high-velocity microprojectile bombardments and regeneration of transgenic plants

    Institute of Scientific and Technical Information of China (English)

    王国英; 杜天兵; 张宏; 谢友菊; 戴景瑞; 米景九; 李太源; 田颖川; 乔利亚; 莽克强

    1995-01-01

    Bt-toxin protein gene was successfully transferred into maize by the microprojectile bombard-ments of cell suspension,embryogenic calli and immature embryos with a Chinese-made particle gun(JQ-700).Although the bombarded embryogenic calli and immature embryos produced less mean transformants per dishthan the cell suspensions,they were the suitable materials for maize transformation because their culture andregeneration have been achieved in most maize cultivars.The evaluation on the resistance of transgenic plantsto corn borer shows the significant difference between them,from highly resistant to susceptible.

  10. Continuous evolution of B. thuringiensis toxins overcomes insect resistance

    OpenAIRE

    2016-01-01

    The Bacillus thuringiensis δ-endotoxins (Bt toxins) are widely used insecticidal proteins in engineered crops that provide agricultural, economic, and environmental benefits. The development of insect resistance to Bt toxins endangers their long-term effectiveness. We developed a phage-assisted continuous evolution (PACE) selection that rapidly evolves high-affinity protein-protein interactions, and applied this system to evolve variants of the Bt toxin Cry1Ac that bind a cadherin-like recept...

  11. Resistance to Bacillus thuringiensis Mediated by an ABC Transporter Mutation Increases Susceptibility to Toxins from Other Bacteria in an Invasive Insect.

    Directory of Open Access Journals (Sweden)

    Yutao Xiao

    2016-02-01

    Full Text Available Evolution of pest resistance reduces the efficacy of insecticidal proteins from the gram-positive bacterium Bacillus thuringiensis (Bt used widely in sprays and transgenic crops. Recent efforts to delay pest adaptation to Bt crops focus primarily on combinations of two or more Bt toxins that kill the same pest, but this approach is often compromised because resistance to one Bt toxin causes cross-resistance to others. Thus, integration of Bt toxins with alternative controls that do not exhibit such cross-resistance is urgently needed. The ideal scenario of negative cross-resistance, where selection for resistance to a Bt toxin increases susceptibility to alternative controls, has been elusive. Here we discovered that selection of the global crop pest, Helicoverpa armigera, for >1000-fold resistance to Bt toxin Cry1Ac increased susceptibility to abamectin and spineotram, insecticides derived from the soil bacteria Streptomyces avermitilis and Saccharopolyspora spinosa, respectively. Resistance to Cry1Ac did not affect susceptibility to the cyclodiene, organophospate, or pyrethroid insecticides tested. Whereas previous work demonstrated that the resistance to Cry1Ac in the strain analyzed here is conferred by a mutation disrupting an ATP-binding cassette protein named ABCC2, the new results show that increased susceptibility to abamectin is genetically linked with the same mutation. Moreover, RNAi silencing of HaABCC2 not only decreased susceptibility to Cry1Ac, it also increased susceptibility to abamectin. The mutation disrupting ABCC2 reduced removal of abamectin in live larvae and in transfected Hi5 cells. The results imply that negative cross-resistance occurs because the wild type ABCC2 protein plays a key role in conferring susceptibility to Cry1Ac and in decreasing susceptibility to abamectin. The negative cross-resistance between a Bt toxin and other bacterial insecticides reported here may facilitate more sustainable pest control.

  12. Resistance to Bacillus thuringiensis Mediated by an ABC Transporter Mutation Increases Susceptibility to Toxins from Other Bacteria in an Invasive Insect

    Science.gov (United States)

    Zhang, Dandan; Gong, Lingling; He, Fei; Soberón, Mario; Bravo, Alejandra; Tabashnik, Bruce E.; Wu, Kongming

    2016-01-01

    Evolution of pest resistance reduces the efficacy of insecticidal proteins from the gram-positive bacterium Bacillus thuringiensis (Bt) used widely in sprays and transgenic crops. Recent efforts to delay pest adaptation to Bt crops focus primarily on combinations of two or more Bt toxins that kill the same pest, but this approach is often compromised because resistance to one Bt toxin causes cross-resistance to others. Thus, integration of Bt toxins with alternative controls that do not exhibit such cross-resistance is urgently needed. The ideal scenario of negative cross-resistance, where selection for resistance to a Bt toxin increases susceptibility to alternative controls, has been elusive. Here we discovered that selection of the global crop pest, Helicoverpa armigera, for >1000-fold resistance to Bt toxin Cry1Ac increased susceptibility to abamectin and spineotram, insecticides derived from the soil bacteria Streptomyces avermitilis and Saccharopolyspora spinosa, respectively. Resistance to Cry1Ac did not affect susceptibility to the cyclodiene, organophospate, or pyrethroid insecticides tested. Whereas previous work demonstrated that the resistance to Cry1Ac in the strain analyzed here is conferred by a mutation disrupting an ATP-binding cassette protein named ABCC2, the new results show that increased susceptibility to abamectin is genetically linked with the same mutation. Moreover, RNAi silencing of HaABCC2 not only decreased susceptibility to Cry1Ac, it also increased susceptibility to abamectin. The mutation disrupting ABCC2 reduced removal of abamectin in live larvae and in transfected Hi5 cells. The results imply that negative cross-resistance occurs because the wild type ABCC2 protein plays a key role in conferring susceptibility to Cry1Ac and in decreasing susceptibility to abamectin. The negative cross-resistance between a Bt toxin and other bacterial insecticides reported here may facilitate more sustainable pest control. PMID:26872031

  13. Construction of a new plant expression vector containing two insect resistant genes and its expression in transgenic tobacco plants

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    A new plant expression vector (pBS29K-BA) containing two insect resistant genes, a synthetic chimeric gene BtS29K encoding the activated insecticidal protein Cry1Ac and a gene API-BA encoding the arrowhead (Sagittaria sagittifolia L.) proteinase inhibitor (API) A and B, is constructed. Transgenic tobacco plants expressing these two genes are obtained through Agrobacterium-mediated transformation of tobacco leaf discs. The average expression levels of Cry1Ac and API-BA proteins in transgenic plants are of 3.2 μg and 4.9 μg per gram fresh leaf respectively. The results of insecticidal assay of transgenic plants indicate that the pBS29K-BA transformed plants are more resistant to insect damage than the plants expressing the Cry1Ac gene or API-BA gene alone.

  14. Isolation and characterization of gut bacterial proteases involved in inducing pathogenicity of Bacillus thuringiensis toxin in cotton bollworm, Helicoverpa armigera

    Directory of Open Access Journals (Sweden)

    Visweshwar Regode

    2016-10-01

    Full Text Available Bacillus thuringiensis (Bt toxin proteins are deployed in transgenic plants for pest management. The present studies were aimed at characterization of gut bacterial proteases involved in activation of inactive Cry1Ac protoxin (pro-Cry1Ac to active toxin in Helicoverpa armigera. Bacterial strains were isolated from H. armigera midgut and screened for their proteolytic activation towards pro-Cry1Ac. Among twelve gut bacterial isolates seven isolates showed proteolytic activity, and proteases from three isolates (IVS1, IVS2 and IVS3 were found to be involved in the proteolytic conversion of pro-Cry1Ac into active toxin. The proteases from IVS1, IVS2 and IVS3 isolates were purified to 11.90-, 15.50- and 17.20-fold, respectively. The optimum pH and temperature for gut bacterial protease activity was 8.0 and 40 oC. Maximum inhibition of total proteolytic activity was exerted by PMSF followed by EDTA. Fluorescence zymography revealed that proteases from IVS1, IVS2, and IVS3 were chymotrypsin-like and showing protease band at ~15, 65 and 15 kDa, respectively. Active Cry1Ac formed from processing pro-Cry1Ac by gut bacterial proteases exhibited toxicity towards H. armigera. The gut bacterial isolates IVS1, IVS2 and IVS3 showed homology with Bacillus thuringiensis (CP003763.1, Vibrio fischeri (CP000020.2 and Escherichia coli (CP011342.1, respectively. Proteases produced by midgut bacteria are involved in proteolytic processing of Bt protoxin and play a major role in inducing pathogenicity of Bt toxins in H. armigera.

  15. Genetic transformation and expression of transgenic lines of Populus x euramericana with insect-resistance and salt-tolerance genes.

    Science.gov (United States)

    Yang, R L; Wang, A X; Zhang, J; Dong, Y; Yang, M S; Wang, J M

    2016-04-29

    We characterized new transgenic varieties of poplar with multiple insect-resistant and salt stress tolerant genes. Two insect-resistant Bacillus thuringiensis (Bt) genes, Cry1Ac and Cry3A, and a salt-tolerant gene, Betaine aldehyde dehydrogenase (BADH) were inserted into a vector, p209-Cry1Ac-Cry3A-BADH. The clone of Populus x euramericana was transformed by the vector using the Agrobacterium-mediated method. Three transgenic lines were assessed using genetic detection and resistance expression analysis. PCR revealed that exogenous genes Cry1Ac, Cry3A, BADH and selective marker gene NPTII were present in three transgenic lines. Quantitative real-time PCR (qPCR) showed significant differences in the transcriptional abundance of three exogenous genes in different lines. Results of assays for Bt toxic proteins showed that the Cry1Ac and Cry3A toxic protein content of each line was 12.83-26.32 and 2108.91-2724.79 ng/g, respectively. The Cry1Ac toxic protein content of different lines was significantly different; the Cry3A toxic protein content was about 100 times higher than that of the Cry1Ac toxic protein. The insect-resistance test revealed the mortality rate of transgenic lines to Hyphantria cunea L1 larvae varied by 42.2-66.7%, which was significantly higher than non-transgenic lines. The mortality rate of L1 and L2 Plagiodera versicolora larvae was 100%. The insecticidal effect of transgenic lines to P. versicolora larvae was higher than that to H. cunea larvae. NaCl stress tolerance of three transgenic lines under 3-6% NaCl concentration was significantly higher than that of non-transgenic lines.

  16. Food safety knowledge on the Bt mutant protein Cry8Ka5 employed in the development of coleopteran-resistant transgenic cotton plants

    NARCIS (Netherlands)

    Felipe Farias, Davi; Peijnenburg, A.A.C.M.; Grossi-de-Sá, Maria F.; Carvalho, A.F.U.

    2015-01-01

    Insecticidal Cry proteins from Bacillus thuringiensis (Bt) have been exploited in the development of genetically modified (GM) crops for pest control. However, several pests are still difficult to control such as the coleopteran boll weevil Anthonomus grandis. By applying in vitro molecular evolutio

  17. Use of Bt-resistant caterpillars to assess the effect of Cry proteins on beneficial natural enemies

    Science.gov (United States)

    A concern related to the use of insect-resistant Bt-transgenic plants is their potential to harm non-target organisms, especially natural enemies of important crop pests. A few studies purporting to show negative effects of Bt plants on non-target organisms had tremendous negative effects on the per...

  18. Susceptibility of field populations of the fall armyworm (Lepidoptera: Noctuidae) from Florida and Puerto Rico to purified Cry1F protein and corn leaf tissue containing single and pyramided Bt genes

    Science.gov (United States)

    Larval survival of Cry1F-susceptible (FL), -resistant (PR and Cry1F-RR), and -heterozygous (FL x PR and Cry1F-RS) populations of the fall armyworm, Spodoptera frugiperda (J.E. Smith) to purified Cry1F protein and corn leaf tissue of seven Bacillus thuringiensis (Bt) corn hybrids and five non-Bt corn...

  19. Effect of High Temperature Stress on Bt Insecticidal Protein Content and Nitrogen Metabolism of Square in Bt Cotton%高温胁迫对Bt棉蕾中杀虫蛋白含量及氮代谢的影响

    Institute of Scientific and Technical Information of China (English)

    衡丽; 李亚兵; 胡大鹏; 王桂霞; 吕春花; 张祥; 陈源; 陈德华

    2016-01-01

    为明确高温气候下Bt棉抗虫性下降的原因,以2个不同类型Bt棉品种为材料,2011年于盛蕾期设计38℃高温胁迫3、5和7 d;2012年设计38℃/25℃的昼夜变温胁迫4、7和10 d,研究其对转Bt棉蕾的杀虫蛋白表达量影响及其生理机理。结果表明,持续高温胁迫7 d内,棉蕾中Bt杀虫蛋白含量呈下降趋势,持续胁迫3 d内下降幅度最大,与未胁迫相比,泗抗3号和泗抗1号分别下降18.71%和26.54%;昼夜变温下高温胁迫4 d时蕾中Bt杀虫蛋白表达量无显著变化,当胁迫7 d以上时, Bt杀虫蛋白含量显著下降,泗抗3号和泗抗1号分别比对照下降11.32%和14.18%。持续高温或昼夜变温高温持续胁迫下,棉蕾中游离氨基酸含量和蛋白酶活性都显著增加,可溶性蛋白含量和 GPT 活性都显著下降。相关分析表明,游离氨基酸含量、蛋白酶活性与Bt杀虫蛋白含量呈极显著负相关;可溶性蛋白含量和GPT活性与Bt杀虫蛋白含量呈极显著正相关。分析表明,高温胁迫下蛋白质合成功能下降,分解能力增强导致可溶蛋白含量,包括Bt杀虫蛋白含量下降。常规种泗抗1号的抗虫性和氮代谢受高温的影响较大。%To explore the reason for low insect resistance under high temperature climate in Bt cotton production, we selected two Bt cotton cultivars as experimental material with treatments of 38℃ for three, five and seven days in 2011, and 38℃/25℃ at day /night for four, seven and ten days, respectively at the artificial climate chamber in 2012. The results showed that the Bt insecti-cidal protein contents declined within seven days under 38℃, with the largest reduction occurred within three days. Compared with control, the square Bt insecticidal protein contents decreased by 18.71% and 26.54% for cultivar Sikang 3 (SK-3) and Sikang 1 (SK-1), respectively. Under 38℃/25℃ at day/night, the square Bt insecticidal protein contents had no obvious reduction within

  20. 一些化学物质对苏云金芽孢杆菌Cry1Ac毒素与昆虫离体细胞相互作用的影响%Influence of some chemicals on the action of Bacillus thuringiensis Cry1Ac toxin with cultured insect cells

    Institute of Scientific and Technical Information of China (English)

    刘凯于; 阎江洪; 康薇; 郑进; 洪华珠

    2005-01-01

    为探讨苏云金芽孢杆菌Bacillus thuringiensis(Bt)杀虫晶体蛋白与昆虫细胞的相互作用,以Bt Cry1Ac毒素和对该毒素敏感的粉纹夜蛾Trichoplusia ni离体细胞BTI-TN-5B1-4为材料,研究了一些化学物质对Cry1Ac毒素与昆虫离体细胞相互作用的影响.结果表明:N-糖基化抑制剂衣霉素、蛋白质合成抑制剂放线菌酮、胞吞作用抑制剂莫能菌素和胰蛋白酶预处理,都能不同程度地提高BTI-TN-5B1-4细胞对Cry1Ac毒素的敏感性,其中胰蛋白酶预处理的作用最明显;而N-乙酰半乳糖胺不能抑制Cry1Ac毒素对这种离体细胞的毒力.

  1. Comparison of three transgenic Bt rice lines for insecticidal protein expression and resistance against a target pest, Chilo suppressalis (Lepidoptera: Crambidae).

    Science.gov (United States)

    Wang, Ya-Nan; Ke, Kai-Qie; Li, Yun-He; Han, Lan-Zhi; Liu, Yan-Min; Hua, Hong-Xia; Peng, Yu-Fa

    2016-02-01

    Two transgenic rice lines (T2A-1 and T1C-19b) expressing cry2A and cry1C genes, respectively, were developed in China, targeting lepidopteran pests including Chilo suppressalis (Walker) (Lepidoptera: Crambidae). The seasonal expression of Cry proteins in different tissues of the rice lines and their resistance to C. suppressalis were assessed in comparison to a Bt rice line expressing a cry1Ab/Ac fusion gene, Huahui 1, which has been granted a biosafety certificate. In general, levels of Cry proteins were T2A-1 > Huahui 1 > T1C-19b among rice lines, and leaf > stem > root among rice tissues. The expression patterns of Cry protein in the rice line plants were similar: higher level at early stages than at later stages with an exception that high Cry1C level in T1C-19b stems at the maturing stage. The bioassay results revealed that the three transgenic rice lines exhibited significantly high resistance against C. suppressalis larvae throughout the rice growing season. According to Cry protein levels in rice tissues, the raw and corrected mortalities of C. suppressalis caused by each Bt rice line were the highest in the seedling and declined through the jointing stage with an exception for T1C-19b providing an excellent performance at the maturing stage. By comparison, T1C-19b exhibited more stable and greater resistance to C. suppressalis larvae than T2A-1, being close to Huahui 1. The results suggest cry1C is an ideal Bt gene for plant transformation for lepidopteran pest control, and T1C-19b is a promising Bt rice line for commercial use for tolerating lepidopteran rice pests.

  2. Cross-Resistance between Cry1 Proteins in Fall Armyworm (Spodoptera frugiperda May Affect the Durability of Current Pyramided Bt Maize Hybrids in Brazil.

    Directory of Open Access Journals (Sweden)

    Daniel Bernardi

    Full Text Available 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 (isofamilies 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

  3. 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

  4. Expression of Bt Protein in Transgenic Insect-resistant Rice%转基因抗虫水稻中Bt蛋白表达量的研究

    Institute of Scientific and Technical Information of China (English)

    于志晶; 蔡勤安; 林秀峰; 马瑞

    2012-01-01

    [Objective] The aim was to study the expression of Bt protein in transgenic pest-resistant rice. [ Method] Enzyme-linked immu-nosorbent assay (ELISA) was used to measure Bt protein expression in different parts of transgenic pest-resistant rice at same growth stage. [Result] Absolute content of Bt protein expression from high to low was as follows: leaves>immature seeds and glumes>roots>stems in different tissues of transgenic rice in grain-filling stage; Bt protein contents of transgenic rice changed a little in different growth stages (tillering stage, booting stage, and grain-filling stage) ; in general, concentration of Bt protein declined a little in later growth stage, for which, the resistibility would not be influenced too much. [ Conclusion ] The experiment is of great importance for pest prevention and transgenic rice breeding.%[目的]对转基因抗虫水稻中Bt蛋白表达量进行研究.[方法]应用酶联免疫吸附测定法(ELISA)定量检测转基因抗虫水稻生相同生长时期不同部位的Bt蛋自表达量.[结果]转基因水稻灌浆期不同组织中Bt蛋白表达绝对含量的高低顺序为:叶片>未成熟种子及颖壳>根>茎杆;在水稻不同的生长发育(分蘖期、抽穗期和灌浆期)阶段,转基因Bt水稻中Bt蛋白的含量有一些变化;一般在水稻生长后期Bt蛋白的浓度有所下降,但幅度不大,对其抗性不会造成太大影响[结论]该试验对田间害虫的防治以及转基因水稻的育种都具有重要意义.

  5. Impacts of Environmental Factors on Degradation of Cry1Ab Insecticidal Protein in Leaf-Blade Powders of Transgenic Bt Rice

    Institute of Scientific and Technical Information of China (English)

    BAI Yao-yu; JIANG Ming-xing; CHENG Jia-an

    2007-01-01

    The determination of the environmental fate of Bt insecticidal protein released by Bt rice plants in paddy soils is a key issue in its ecological risk assessment. In this study, the impacts of soil water content, pH, and temperature on the degradation of Cry1Ab protein expressed in the leaves of Bt rice KMD2 were studied in the laboratory. Three types of paddy soils were used, i.e., blue clayey paddy soil, pale paddy soil on quaternary red soil, and marine-fluvigenic yellow loamy paddy soil. Ground powders of KMD2 leaf blades were mixed with each type of soil, and degradation dynamics of Cry1Ab were measured using enzyme-linked immunosorbent assay (ELISA). The degradation rate of Cry1Ab was high at the early experimental stage, but slowed down steadily at middle and later stages, which could be described by exponential equations, with the half-life period of degradation determined as 1.8-4.0 d. The soil water content, pH, and temperature could affect the degradation of Cry1Ab, but the effects of soil pH and temperature were relatively greater. In general,Cry 1 Ab degradations were slower under lower soil pH and temperature conditions, especially for marine-fluvigenic yellow loamy paddy soil.

  6. Use of a pollen-based diet to expose the ladybird beetle Propylea japonica to insecticidal proteins.

    Directory of Open Access Journals (Sweden)

    Xiaojie Zhang

    Full Text Available A rape seed pollen-based diet was developed and found to be suitable for use in a dietary exposure assay for Propylea japonica. Using the diet, we established and validated a dietary exposure assay by using the protease inhibitor E-64 as positive control. Dose-dependent responses were documented for all observed life-table parameters of P. japonica including survival, pupation and eclosion rates, development time and adult weight. Results suggested that the dietary assay can detect the effects of insecticidal compounds on the survival and development of P. japonica. Using the established dietary assay, we subsequently tested the toxicity of Cry1Ab, Cry1Ac and Cry1F proteins that are expressed by transgenic maize, cotton or rice plants to P. japonica larvae. The diet containing E-64 was included as a positive control. Survival and development of P. japonica larvae were not adversely affected when the diet contained purified Cry1Ab, Cry1Ac, or Cry1F at 500 µg/g diet representing a worst-case exposure scenario. In contrast, P. japonica larvae were adversely affected when the diet contained E-64. The bioactivity and stability of the Cry proteins in the diet and Cry protein uptake by the ladybird larvae were confirmed by bioassay with a Cry-sensitive insect species and by ELISA. The current study describes a suitable experimental system for assessing the potential effects of gut-active insecticidal compounds on ladybird beetle larvae. The experiments with the Cry proteins demonstrate that P. japonica larvae are not sensitive to Cry1Ab, Cry1Ac and Cry1F.

  7. Use of a pollen-based diet to expose the ladybird beetle Propylea japonica to insecticidal proteins.

    Science.gov (United States)

    Zhang, Xiaojie; Li, Yunhe; Romeis, Jörg; Yin, Xinming; Wu, Kongming; Peng, Yufa

    2014-01-01

    A rape seed pollen-based diet was developed and found to be suitable for use in a dietary exposure assay for Propylea japonica. Using the diet, we established and validated a dietary exposure assay by using the protease inhibitor E-64 as positive control. Dose-dependent responses were documented for all observed life-table parameters of P. japonica including survival, pupation and eclosion rates, development time and adult weight. Results suggested that the dietary assay can detect the effects of insecticidal compounds on the survival and development of P. japonica. Using the established dietary assay, we subsequently tested the toxicity of Cry1Ab, Cry1Ac and Cry1F proteins that are expressed by transgenic maize, cotton or rice plants to P. japonica larvae. The diet containing E-64 was included as a positive control. Survival and development of P. japonica larvae were not adversely affected when the diet contained purified Cry1Ab, Cry1Ac, or Cry1F at 500 µg/g diet representing a worst-case exposure scenario. In contrast, P. japonica larvae were adversely affected when the diet contained E-64. The bioactivity and stability of the Cry proteins in the diet and Cry protein uptake by the ladybird larvae were confirmed by bioassay with a Cry-sensitive insect species and by ELISA. The current study describes a suitable experimental system for assessing the potential effects of gut-active insecticidal compounds on ladybird beetle larvae. The experiments with the Cry proteins demonstrate that P. japonica larvae are not sensitive to Cry1Ab, Cry1Ac and Cry1F.

  8. Food safety knowledge on the Bt mutant protein Cry8Ka5 employed in the development of coleopteran-resistant transgenic cotton plants

    OpenAIRE

    2015-01-01

    Insecticidal Cry proteins from Bacillus thuringiensis (Bt) have been exploited in the development of genetically modified (GM) crops for pest control. However, several pests are still difficult to control such as the coleopteran boll weevil Anthonomus grandis. By applying in vitro molecular evolution to the cry8Ka1 gene sequence, variants were generated with improved activity against A. grandis. Among them, Cry8Ka5 mutant protein showed coleoptericidal activity 3-fold higher (LC50 2.83 μg/mL)...

  9. Bt toxin modification for enhanced efficacy.

    Science.gov (United States)

    Deist, Benjamin R; Rausch, Michael A; Fernandez-Luna, Maria Teresa; Adang, Michael J; Bonning, Bryony C

    2014-10-22

    Insect-specific toxins derived from Bacillus thuringiensis (Bt) provide a valuable resource for pest suppression. Here we review the different strategies that have been employed to enhance toxicity against specific target species including those that have evolved resistance to Bt, or to modify the host range of Bt crystal (Cry) and cytolytic (Cyt) toxins. These strategies include toxin truncation, modification of protease cleavage sites, domain swapping, site-directed mutagenesis, peptide addition, and phage display screens for mutated toxins with enhanced activity. Toxin optimization provides a useful approach to extend the utility of these proteins for suppression of pests that exhibit low susceptibility to native Bt toxins, and to overcome field resistance.

  10. Interaction between Functional Domains of Bacillus thuringiensis Insecticidal Crystal Proteins

    Science.gov (United States)

    Rang, Cécile; Vachon, Vincent; de Maagd, Ruud A.; Villalon, Mario; Schwartz, Jean-Louis; Bosch, Dirk; Frutos, Roger; Laprade, Raynald

    1999-01-01

    Interactions among the three structural domains of Bacillus thuringiensis Cry1 toxins were investigated by functional analysis of chimeric proteins. Hybrid genes were prepared by exchanging the regions coding for either domain I or domain III among Cry1Ab, Cry1Ac, Cry1C, and Cry1E. The activity of the purified trypsin-activated chimeric toxins was evaluated by testing their effects on the viability and plasma membrane permeability of Sf9 cells. Among the parental toxins, only Cry1C was active against these cells and only chimeras possessing domain II from Cry1C were functional. Combination of domain I from Cry1E with domains II and III from Cry1C, however, resulted in an inactive toxin, indicating that domain II from an active toxin is necessary, but not sufficient, for activity. Pores formed by chimeric toxins in which domain I was from Cry1Ab or Cry1Ac were slightly smaller than those formed by toxins in which domain I was from Cry1C. The properties of the pores formed by the chimeras are therefore likely to result from an interaction between domain I and domain II or III. Domain III appears to modulate the activity of the chimeric toxins: combination of domain III from Cry1Ab with domains I and II of Cry1C gave a protein which was more strongly active than Cry1C. PMID:10388684

  11. Identification of top-down forces regulating cotton aphid population growth in transgenic Bt cotton in central China.

    Directory of Open Access Journals (Sweden)

    Peng Han

    Full Text Available The cotton aphid Aphis gossypii Glover is the main aphid pest in cotton fields in the Yangtze River Valley Cotton-planting Zone (YRZ in central China. Various natural enemies may attack the cotton aphid in Bt cotton fields but no studies have identified potential specific top-down forces that could help manage this pest in the YRZ in China. In order to identify possibilities for managing the cotton aphid, we monitored cotton aphid population dynamics and identified the effect of natural enemies on cotton aphid population growth using various exclusion cages in transgenic Cry1Ac (Bt+CpTI (Cowpea trypsin inhibitor cotton field in 2011. The aphid population growth in the open field (control was significantly lower than those protected or restricted from exposure to natural enemies in the various exclusion cage types tested. The ladybird predator Propylaea japonica Thunberg represented 65% of Coccinellidae predators, and other predators consisted mainly of syrphids (2.1% and spiders (1.5%. The aphid parasitoids Aphidiines represented 76.7% of the total count of the natural enemy guild (mainly Lysiphlebia japonica Ashmead and Binodoxys indicus Subba Rao & Sharma. Our results showed that P. japonica can effectively delay the establishment and subsequent population growth of aphids during the cotton growing season. Aphidiines could also reduce aphid density although their impact may be shadowed by the presence of coccinellids in the open field (likely both owing to resource competition and intraguild predation. The implications of these results are discussed in a framework of the compatibility of transgenic crops and top-down forces exerted by natural enemy guild.

  12. Identification of top-down forces regulating cotton aphid population growth in transgenic Bt cotton in central China.

    Science.gov (United States)

    Han, Peng; Niu, Chang-ying; Desneux, Nicolas

    2014-01-01

    The cotton aphid Aphis gossypii Glover is the main aphid pest in cotton fields in the Yangtze River Valley Cotton-planting Zone (YRZ) in central China. Various natural enemies may attack the cotton aphid in Bt cotton fields but no studies have identified potential specific top-down forces that could help manage this pest in the YRZ in China. In order to identify possibilities for managing the cotton aphid, we monitored cotton aphid population dynamics and identified the effect of natural enemies on cotton aphid population growth using various exclusion cages in transgenic Cry1Ac (Bt)+CpTI (Cowpea trypsin inhibitor) cotton field in 2011. The aphid population growth in the open field (control) was significantly lower than those protected or restricted from exposure to natural enemies in the various exclusion cage types tested. The ladybird predator Propylaea japonica Thunberg represented 65% of Coccinellidae predators, and other predators consisted mainly of syrphids (2.1%) and spiders (1.5%). The aphid parasitoids Aphidiines represented 76.7% of the total count of the natural enemy guild (mainly Lysiphlebia japonica Ashmead and Binodoxys indicus Subba Rao & Sharma). Our results showed that P. japonica can effectively delay the establishment and subsequent population growth of aphids during the cotton growing season. Aphidiines could also reduce aphid density although their impact may be shadowed by the presence of coccinellids in the open field (likely both owing to resource competition and intraguild predation). The implications of these results are discussed in a framework of the compatibility of transgenic crops and top-down forces exerted by natural enemy guild.

  13. An Evaluation of Methods for Assessing the Impacts of Bt-maize on Non-target Organisms with Leafhoppers%利用叶蝉评价转基因玉米对非靶标生物的风险

    Institute of Scientific and Technical Information of China (English)

    冯艳杰; 葛阳; 谭树乾; 陈红星; 郭艳艳; 张恺祺; 石旺鹏

    2014-01-01

    The application of transgenic insect-resistant crops has been an important way to replace insecticides. But transgenic insect-resistant crops have potential negative effects, especially the effect on the non-target organisms and resistance to GM. Visual assessment and yellow traps were used for investigating the dynamic of leafhoppers and main arthropods in Cry1Ac transgenic insect-resistant corn field for two years. The study showed that planthoppers and leafhoppers were highly abundant and diverse in maize fields, their mid-term occurrence was 28.9%-43.1% of sucking pest insects, and the population dynamic had no significant difference between Bt-maize and the untreated near-isogenic maize. Leafhoppers were directly exposed to these entomotoxic proteins and could also serve as a source of Cry protein exposure to predatory arthropods. Therefore, it was reasonable to use Auchenorrhyncha in the risk assessment of GM corn. So the risk of Bt corn on non-target organisms could be evaluated using leafhoppers as an indicator species. The results in field tests showed that this method was feasible.%转基因抗虫作物应用是替代杀虫剂的重要途径。但转基因作物可能存负面效应,特别是对非靶标生物的影响及害虫产生抗性。采用直接和陷阱调查结合方法,连续多年对转Cry1Ac抗虫基因玉米实验田叶蝉和主要节肢动物数量动态跟踪调查,结果发现叶蝉是玉米田优势非靶标生物种群之一,中期发生比例占刺吸式害虫的28.9%~43.1%,且种群动态与主要非靶标节肢动物群落动态基本一致。又鉴于叶蝉直接刺吸接触到Bt杀虫蛋白,可直接反映转基因作物对非靶标生物的安全性,因此可利用叶蝉作为转基因抗虫玉米安全性评价指示生物来评价转基因玉米对非靶标生物群落风险,田间测试结果证明此方法是可行的。

  14. 转Bt毒蛋白基因玉米及其抗虫性研究进展%Transgenic Corn with Bt Toxin Protein Gene and Insect-resistance

    Institute of Scientific and Technical Information of China (English)

    杨春英; 宋建成

    2001-01-01

    本文从转Bt毒蛋白基因玉米的培育及商品化,Bt毒蛋白基因在转基因玉米中的遗传分离与整合、对玉米螟及其它害虫的杀虫效果、对天敌种群数量和玉米病害发生程度的影响、玉米螟对转Bt毒蛋白基因玉米产生抗性及解决措施、应用转Bt毒蛋白基因玉米潜在的生态风险性等方面对国内外最新研究进展进行了综述。%This review briefly focused on the progress at home and overseas in the study of transgenic corn transformed with Bt toxic protein gene,including the breeding and commercial production of transgenic corn transformed with Bt toxic protein gene,genetic segregation and combination of Bt toxin protein in transgenic corn,insecticidal effect on corn borer and other pests,influence on natural enemy population amount and corn disease,corn borer tolerance to Bt toxic protein gene and countermeasures,and the potential ecological risk of transgenic corn transformed with Bt toxic protein gene.

  15. Bt pollen dispersal and Bt kernel mosaics: integrity of non-Bt refugia for lepidopteran resistance management in maize.

    Science.gov (United States)

    Burkness, Eric C; Hutchison, W D

    2012-10-01

    Field trials were conducted at Rosemount, MN in 2009 and 2010, to measure pollen movement from Bt corn to adjacent blocks of non-Bt refuge corn. As the use of Bt corn hybrids continues to increase in the United States, and new insect resistance management (IRM) plans are implemented, it is necessary to measure the efficacy of these IRM plans. In Minnesota, the primary lepidopteran pests of corn include the European corn borer, Ostrinia nubilalis (Hübner) and corn earworm, Helicoverpa zea (Boddie). The primary IRM plan in transgenic corn is the use of hybrids expressing a high dose of insecticidal proteins and an insect refuge containing hybrids not expressing insecticidal proteins that produce susceptible insects. Wind-assisted pollen movement in corn occurs readily, and is the primary method of pollination for corn. The combination of pollen movement and viability determines the potential for cross pollination of refuge corn. In 2009 and 2010, cross pollination occurred with the highest frequency on the north and east sides of Bt corn fields, but was found at some level in all directions. Highest levels of cross pollination (75%) were found within the first four rows (3 m) of non-Bt corn adjacent to Bt corn, and in general decreasing levels of cross pollination were found the further the non-Bt corn was planted from the Bt corn. A mosaic of Bt cross-pollinated kernels was found throughout the ear, but in both years the ear tip had the highest percentage of cross-pollinated kernels; this pattern may be linked to the synchrony of pollen shed and silking between Bt and non-Bt corn hybrids. The dominant wind direction in both years was from WNW. However, in both years, there were also prevailing winds from SSW and WSW. Further studies are needed to quantify Bt levels in cross-pollinated kernels, measure the Bt dose of such kernels and associated lepidopteran pest survival, and measure the impact of Bt pollen on lepidopteran pests, particularly when considering the

  16. Engineering modified Bt toxins to counter insect resistance.

    Science.gov (United States)

    Soberón, Mario; Pardo-López, Liliana; López, Idalia; Gómez, Isabel; Tabashnik, Bruce E; Bravo, Alejandra

    2007-12-07

    The evolution of insect resistance threatens the effectiveness of Bacillus thuringiensis (Bt) toxins that are widely used in sprays and transgenic crops. Resistance to Bt toxins in some insects is linked with mutations that disrupt a toxin-binding cadherin protein. We show that susceptibility to the Bt toxin Cry1Ab was reduced by cadherin gene silencing with RNA interference in Manduca sexta, confirming cadherin's role in Bt toxicity. Native Cry1A toxins required cadherin to form oligomers, but modified Cry1A toxins lacking one alpha-helix did not. The modified toxins killed cadherin-silenced M. sexta and Bt-resistant Pectinophora gossypiella that had cadherin deletion mutations. Our findings suggest that cadherin promotes Bt toxicity by facilitating toxin oligomerization and demonstrate that the modified Bt toxins may be useful against pests resistant to standard Bt toxins.

  17. 转Bt基因水稻种子中Bt蛋白含量测定方法的研究%Protein Content in Bt- Gene Rice Seed

    Institute of Scientific and Technical Information of China (English)

    陈笑芸; 汪小福; 周育; 缪青梅; 方敬; 徐俊锋

    2012-01-01

    采用ELISA技术检测转Bt基因水稻中Bt蛋白的含量,判断水稻样品中是否含有转基因成分.利用研磨、酶标、孵育等技术对样品进行前处理.采用阳性质控物浓度等倍稀释方法,建立标准曲线,相关系数为0.997 4.用一系列不同转基因含量的标准基体材料,分析方法的最低检测限,灵敏度达0.1%.通过对我国进入生产性试验的转Bt基因水稻品系TT51 -1和科丰6号的测试,表明该方法与普通PCR方法真实性和灵敏度一致,可以广泛应用于转Bt基因水稻及其粗加工产品的转基因成分检测.为转基因生物安全监管和安全性评价提供技术支撑.%In this study, the enzyme - linked immunosorbent assay (ELISA) has been develpoed for the detection of Bt protein content in transgenic Bt rice to evaluate the presence of genetically modified ingredients in rice samples. The sample pre -treatment step involved in grinding,enzymatic labeling reaction and incubation process. The standard curve was constructed by dilution of the the positive control material with the same dilution factor, and the correlation coefficient is 0. 997 4. A range of different standards containing of genetically modified ingredients were used to analyze the limit of detection method. The results showed the detection sensitivity is up to 0.1% . Through testing the transgenic Bt rice strain TT51-1 and Kefeng 6 which are putting into the production test in China, the ELISA method developed demonstrated consistency with ordinary PCR methods in both authenticity and sensitivity aspects. Therefore, this ELISA method could be widely applied to detect genetically modified ingredients in transgenic Bt rice or raw commodities, providng technical support for the safety regulation and evalution of genetically modified organisms.

  18. Expression of Cry1Ab protein in a marker-free transgenic Bt rice line and its efficacy in controlling a target pest, Chilo suppressalis (Lepidoptera: Crambidae).

    Science.gov (United States)

    Wang, Yanan; Zhang, Lei; Li, Yunhe; Liu, Yanmin; Han, Lanzhi; Zhu, Zhen; Wang, Feng; Peng, Yufa

    2014-04-01

    A marker-free Bt transgenic rice line, mfb-MH86, was recently developed in China, which contains a cry1Ab gene driven by a ubiquitin promoter. This Bt gene confers resistance to a range of lepidopteran species, including the striped stem borer, Chilo suppressalis (Walker). The expression of Cry1Ab protein in mfb-MH86 leaves, stems and leaf sheaths (hereinafter referred to as stems), and roots was evaluated throughout the rice-growing season using an enzyme-linked immunosorbent assay. In addition, mfb-MH86 resistance to C. suppressalis, a major pest of rice, was evaluated in a laboratory bioassay with field-collected rice stems. Cry1Ab protein levels of mfb-MH86 were highest in leaves (9.71-34.09 μg/g dry weight [DW]), intermediate in stems (7.66-18.51 μg/g DW), and lowest in roots (1.95-13.40 μg/g DW). In all tissues, Cry1Ab levels in mfb-MH86 were higher in seedling and tillering stages than in subsequent growth stages. In the laboratory bioassay, mortality of C. suppressalis after 6 d of feeding on mfb-MH86 stems was 100% throughout the rice-growing season; mortality of C. suppressalis when feeding on stems of the nontransformed isoline, MH86, ranged from 15.0 to 38.3%. The results indicate that Cry1Ab protein levels in mfb-MH86 stems are sufficient to protect plants against C. suppressalis throughout the rice-growing season. Although our results are promising, further comprehensive evaluations of mfb-MH86, including field surveys, will be needed before commercial use.

  19. Purification of the insecticidal Cry2Ad protein from a Bt-isolated BRC-HZP10 strain and toxin assay to the diamondback moth, Plutella xylostella (L.).

    Science.gov (United States)

    Liao, J Y; Gao, Y Q; Wu, Q Y; Zhu, Y C; You, M S

    2015-07-13

    The present study aims to characterize the Cry2Ad toxin protein isolated from a Bacillus thuringiensis strain, BRC-HZP10, which have a potential insecticidal activity against larvae of the diamondback moth, Plutella xylostella (L.). The crude Bt toxin proteins were isolated and purified by cation exchange chromatography, then equilibrated with 0.2 M NaOH buffer, pH 4.0, followed by ultraviolet detection at 280 nm and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A refined Cry2Ad toxin protein with 88.34% purity was eventually obtained and used for a bioassay by feeding it to P. xylostella. The results showed conspicuous insecticidal activity towards P. xylostella with 50% lethal concentration of 6.84 μg/mL and 95% confidence interval of 5.77-7.91 mg/mL. At a concentration of 16.38 μg/mL, the intake of Cry2Ad protein significantly shortened the oviposition period and larval developmental duration, but significantly reduced the fecundity and egg hatchability of the population compared to those of control (without treatment with Cry2Ad protein) (P protein plays an effective role in controlling the population of P. xylostella.

  20. Improvement and efficient display of Bacillus thuringiensis toxins on M13 phages and ribosomes.

    Science.gov (United States)

    Pacheco, Sabino; Cantón, Emiliano; Zuñiga-Navarrete, Fernando; Pecorari, Frédéric; Bravo, Alejandra; Soberón, Mario

    2015-12-01

    Bacillus thuringiensis (Bt) produces insecticidal proteins that have been used worldwide in the control of insect-pests in crops and vectors of human diseases. However, different insect species are poorly controlled by the available Bt toxins or have evolved resistance to these toxins. Evolution of Bt toxicity could provide novel toxins to control insect pests. To this aim, efficient display systems to select toxins with increased binding to insect membranes or midgut proteins involved in toxicity are likely to be helpful. Here we describe two display systems, phage display and ribosome display, that allow the efficient display of two non-structurally related Bt toxins, Cry1Ac and Cyt1Aa. Improved display of Cry1Ac and Cyt1Aa on M13 phages was achieved by changing the commonly used peptide leader sequence of the coat pIII-fusion protein, that relies on the Sec translocation pathway, for a peptide leader sequence that relies on the signal recognition particle pathway (SRP) and by using a modified M13 helper phage (Phaberge) that has an amber mutation in its pIII genomic sequence and preferentially assembles using the pIII-fusion protein. Also, both Cry1Ac and Cyt1Aa were efficiently displayed on ribosomes, which could allow the construction of large libraries of variants. Furthermore, Cry1Ac or Cyt1Aa displayed on M13 phages or ribosomes were specifically selected from a mixture of both toxins depending on which antigen was immobilized for binding selection. These improved systems may allow the selection of Cry toxin variants with improved insecticidal activities that could counter insect resistances.

  1. Toxicity of Cry1A toxins from Bacillus thuringiensis to CF1 cells does not involve activation of adenylate cyclase/PKA signaling pathway.

    Science.gov (United States)

    Portugal, Leivi; Muñóz-Garay, Carlos; Martínez de Castro, Diana L; Soberón, Mario; Bravo, Alejandra

    2017-01-01

    Bacillus thuringiensis (Bt) bacteria produce Cry toxins that are able to kill insect pests. Different models explaining the mode of action of these toxins have been proposed. The pore formation model proposes that the toxin creates pores in the membrane of the larval midgut cells after interaction with different receptors such as cadherin, aminopeptidase N and alkaline phosphatase and that this pore formation activity is responsible for the toxicity of these proteins. The alternative model proposes that interaction with cadherin receptor triggers an intracellular cascade response involving protein G, adenylate cyclase (AC) and protein kinase A (PKA). In addition, it was shown that Cry toxins induce a defense response in the larvae involving the activation of mitogen-activated kinases such as MAPK p38 in different insect orders. Here we analyzed the mechanism of action of Cry1Ab and Cry1Ac toxins and a collection of mutants from these toxins in the insect cell line CF1 from Choristoneura fumiferana, that is naturally sensitive to these toxins. Our results show that both toxins induced permeability of K(+) ions into the cells. The initial response after intoxication with Cry1Ab and Cry1Ac toxins involves the activation of a defense response that involves the phosphorylation of MAPK p38. Analysis of activation of PKA and AC activities indicated that the signal transduction involving PKA, AC and cAMP was not activated during Cry1Ab or Cry1Ac intoxication. In contrast we show that Cry1Ab and Cry1Ac activate apoptosis. These data indicate that Cry toxins can induce an apoptotic death response not related with AC/PKA activation. Since Cry1Ab and Cry1Ac toxins affected K(+) ion permeability into the cells, and that mutant toxins affected in pore formation are not toxic to CF1, we propose that pore formation activity of the toxins is responsible of triggering cell death response in CF1cells.

  2. Research Progress of Maize Transformation with Bt Toxin Protein Gene%转Bt毒蛋白基因玉米的研究进展

    Institute of Scientific and Technical Information of China (English)

    谢树章; 雷开荣; 林清

    2011-01-01

    In this paper, firstly, we review research progress of transgenic maize with insect resistant, classify and summarize main approaches of maize transformation with Bt toxin protein gene and screening of transgenic lines, then the bioassay for insect resistance and evaluation and utilization of genetic stability are introduced,too. Finally, we discuss the technical difficulties and prospect of transgenic maize with insect resistant. We hold that in order to promote better development of maize transformation with Bt toxin protein gene, the key technologies must be further study positively.%此文首先回顾了国内外抗虫转基因玉米研究发展历程,归纳总结了转Bt毒蛋白基因玉米的常用转化方法和转基因玉米转化体的鉴定方法,对转Bt毒蛋白基因玉米后续实验的抗虫性分析鉴定及遗传稳定性评价与利用也进行了介绍.进而探讨了Bt毒蛋白基因在玉米遗传转化上善待解决的问题以及未来的发展方向.认为应加大力度对瓶颈技术进行深入研究,以期转Bt毒蛋白基因玉米能够获得更好的发展.

  3. IT-BT convergence technology

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2012-12-15

    This book explains IT-BT convergence technology as the future technology, which includes a prolog, easy IT-BT convergence technology that has infinite potentials for new value, policy of IT-BT convergence technology showing the potential of smart Korea, IT-BT convergence opening happy future, for the new future of IT powerful nation Korea with IT-BT convergence technology and an epilogue. This book reveals the conception, policy, performance and future of IT-BT convergence technology.

  4. Screening of Bt isolates with insecticidal activity against Apolygus lucorum (Heteroptera:Miridae)and bioassay of Cry15Aa polypeptides%对绿盲蝽具有杀虫活性Bt菌株的筛选及Cry15Aa蛋白活性的研究

    Institute of Scientific and Technical Information of China (English)

    张学雯; 束长龙; 陆宴辉; 刘春颖; 张杰; 高继国

    2016-01-01

    Transgenic cotton with cry genes has controlled the main cotton pests (Helicoverpa armigera )effective-ly,however another non target mirid bugs (Heteroptera:Miridae)with sucking mouthparts have increased popu-lation sizes year by year.It has not been reported to the effective insecticidal genes of Bacillus thuringiensis a-gainst Apolygus lucorum (Meyer-Dür),in this study,Bt insecticidal genes and isolates stored in our laboratory were prepared to assay the insecticidal activity against A.lucorum .The cry 15Aa gene which belongs to Mtx in our lab was used to perform the bioassay to A.lucorum ,the result showed that Cry15Aa polypeptides was certain insecticidal activity against A.lucorum .After bioassay against A.lucorum ,twenty A.lucorum toxic Bt isolates were obtained.The LC-MS/MS identification showed these isolates might contains Cry1Aa,Cry1Ab,Cry1Ac, Cry1Ae,Cry1Af,Cry1Ag,Cry1Ah,Cry1Ba,Cry1Be,Cry8Ha and other proteins which are insecticidal to lepi-doptera and coleopteran pests,and four of them contains peptide fragment similar to Cry15Aa which belong to Mtx protein.But subsequence gene identification results indicated that these isolates not contains full length cry 15Aa gene,it suggested that these isolates contains new cry 15 type genes.The results indicated that these Bt i-solates may contain new insecticidal proteins toxic to mirid bugs.The obtaining of novel Bt isolates and the discovery of Cry15Aa with insecticidal activity against A.lucorum are significant to the biological control for A.lucorum .%Bt 棉有效控制了棉田主要害虫棉铃虫(Helicoverpa armigera ),然而原来处于次要地位的刺吸式口器害虫盲蝽(Heteroptera:Miridae)为害逐年加重,目前对绿盲蝽(Apolygus lucorum Meyer-Dür)有效的抗虫基因未见报道。本研究以本实验室保存的 Bt 杀虫基因和菌株为材料,对绿盲蝽进行杀虫活性筛选。利用本实验室先前克隆的Mtx 类杀虫基因 cry15Aa 表达产物进行绿

  5. General Research Situation of Resistance of Target Insect to Bt Insecticidal Crystal Protein%目标昆虫对Bt杀虫晶体蛋白抗性的研究概况

    Institute of Scientific and Technical Information of China (English)

    杨宙; 康美花; 陈红萍; 刘建华; 曹丰生

    2012-01-01

    This article summarized the resistance of many target insect species to Bt insecticidal crystal protein, and introduced the mechanism of generating resistance and its hereditary characters.%综述了多种目标昆虫对Bt杀虫晶体蛋白的抗性机制和遗传特性.

  6. Food safety knowledge on the Bt mutant protein Cry8Ka5 employed in the development of coleopteran-resistant transgenic cotton plants.

    Science.gov (United States)

    Farias, Davi F; Peijnenburg, Ad A C M; Grossi-de-Sá, Maria F; Carvalho, Ana F U

    2015-01-01

    Insecticidal Cry proteins from Bacillus thuringiensis (Bt) have been exploited in the development of genetically modified (GM) crops for pest control. However, several pests are still difficult to control such as the coleopteran boll weevil Anthonomus grandis. By applying in vitro molecular evolution to the cry8Ka1 gene sequence, variants were generated with improved activity against A. grandis. Among them, Cry8Ka5 mutant protein showed coleoptericidal activity 3-fold higher (LC50 2.83 μg/mL) than that of the original protein (Cry8Ka1). Cry8Ka5 has been used in breeding programs in order to obtain coleopteran-resistant cotton plants. Nevertheless, there is some concern in relation to the food safety of transgenic crops, especially to the heterologously expressed proteins. In this context, our research group has performed risk assessment studies on Cry8Ka5, using the tests recommended by Codex as well as tests that we proposed as alternative and/or complementary approaches. Our results on the risk analysis of Cry8Ka5 taken together with those of other Cry proteins, point out that there is a high degree of certainty on their food safety. It is reasonable to emphasize that most safety studies on Cry proteins have essentially used the Codex approach. However, other methodologies would potentially provide additional information such as studies on the effects of Cry proteins and derived peptides on the indigenous gastrointestinal microbiota and on intestinal epithelial cells of humans. Additionally, emerging technologies such as toxicogenomics potentially will offer sensitive alternatives for some current approaches or methods.

  7. 转基因抗虫耐除草剂(Cry1Ac+EPSPS)玉米对草甘膦耐受性研究%Tolerance of transgenic and herbicide-and insect-resistant corn to glyphosate

    Institute of Scientific and Technical Information of China (English)

    江帆; 刘凯强; 张天涛; 白树雄; 王振营; 何康来

    2016-01-01

    Insect-resistant cry 1Ac gene and glyphosate-tolerant gene were used to construct an insect-resistant/glyphosate-tolerant bivalent plant which had been patented.It effectively improved complex traits in corn.This study mainly evaluated the tolerance of insect-resistant/glyphosate-tolerant bivalent corn and glyphosate-tolerant corn to glyphosate,and measured the quantity of Cry1Ac and CP4 EPSPS protein expressed in corn tissues,in or-der to provide the basis for the improvement and application of the maize varieties.Insect-resistant/glyphosate-tolerant bivalent corn ‘CC-2×BT799’,insect-resistant corn ‘BT799’,herbicide-resistant corn ‘CC-2’,and con-ventional corn ‘Zheng 58’were compared by investigating the survival rates and heights of plants after spraying herbicides.Cry1Ac and CP4 EPSPS protein expression was measured by ELISA.‘CC-2’and ‘CC-2×BT799’per-formed excellent tolerance to glyphosate after spraying glyphosate with recommended and double recommended rates at the whorl stage.ELISA detection showed that CP4 EPSPS protein expression in the tassel was slightly higher than in leaves.CP4 EPSPS protein expression in ‘CC-2 ×BT799’was slightly higher than in ‘CC-2’. Cry1Ac protein expression in BT799 and ‘CC-2×BT799’was nearly the same in the tassel.However,Cry1Ac protein expression in BT799 leaves was about 2 times of that in ‘CC-2×BT799’.Insect-resistant/glyphosate-tolerant corn showed strong resistance to glyphosate.Cry1Ac protein expression varied among the corn varieties and tissues.%将具有我国自主知识产权的抗虫、耐除草剂基因转化到玉米中获得兼具抗虫和耐除草剂的复合性状转基因玉米,实现了玉米复杂性状的有效改良,研究了转耐除草剂 CC-2基因和可表达 Bt 毒素的 cry1Ac 基因的双抗玉米对除草剂的耐受性及目标蛋白表达量。通过调查喷施除草剂后玉米植株存活率及株高和生长发育情况,研究了转抗虫

  8. The halo effect: suppression of pink bollworm on non-Bt cotton by Bt cotton in China.

    Directory of Open Access Journals (Sweden)

    Peng Wan

    Full Text Available In some previously reported cases, transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt have suppressed insect pests not only in fields planted with such crops, but also regionally on host plants that do not produce Bt toxins. Here we used 16 years of field data to determine if Bt cotton caused this "halo effect" against pink bollworm (Pectinophora gossypiella in six provinces of the Yangtze River Valley of China. In this region, the percentage of cotton hectares planted with Bt cotton increased from 9% in 2000 to 94% in 2009 and 2010. We found that Bt cotton significantly decreased the population density of pink bollworm on non-Bt cotton, with net decreases of 91% for eggs and 95% for larvae on non-Bt cotton after 11 years of Bt cotton use. Insecticide sprays targeting pink bollworm and cotton bollworm (Helicoverpa armigera decreased by 69%. Previously reported evidence of the early stages of evolution of pink bollworm resistance to Bt cotton in China has raised concerns that if unchecked, such resistance could eventually diminish or eliminate the benefits of Bt cotton. The results reported here suggest that it might be possible to find a percentage of Bt cotton lower than the current level that causes sufficient regional pest suppression and reduces the risk of resistance.

  9. The halo effect: suppression of pink bollworm on non-Bt cotton by Bt cotton in China.

    Science.gov (United States)

    Wan, Peng; Huang, Yunxin; Tabashnik, Bruce E; Huang, Minsong; Wu, Kongming

    2012-01-01

    In some previously reported cases, transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) have suppressed insect pests not only in fields planted with such crops, but also regionally on host plants that do not produce Bt toxins. Here we used 16 years of field data to determine if Bt cotton caused this "halo effect" against pink bollworm (Pectinophora gossypiella) in six provinces of the Yangtze River Valley of China. In this region, the percentage of cotton hectares planted with Bt cotton increased from 9% in 2000 to 94% in 2009 and 2010. We found that Bt cotton significantly decreased the population density of pink bollworm on non-Bt cotton, with net decreases of 91% for eggs and 95% for larvae on non-Bt cotton after 11 years of Bt cotton use. Insecticide sprays targeting pink bollworm and cotton bollworm (Helicoverpa armigera) decreased by 69%. Previously reported evidence of the early stages of evolution of pink bollworm resistance to Bt cotton in China has raised concerns that if unchecked, such resistance could eventually diminish or eliminate the benefits of Bt cotton. The results reported here suggest that it might be possible to find a percentage of Bt cotton lower than the current level that causes sufficient regional pest suppression and reduces the risk of resistance.

  10. Advances in effects of insecticidal crystal proteins released from transgenic Bt crops on soil ecology%转Bt基因作物释放杀虫晶体蛋白对土壤生态安全的影响

    Institute of Scientific and Technical Information of China (English)

    周学永; 刘宁; 赵曼; 李河; 周浪; 唐宗文; 曹斐; 李巍

    2011-01-01

    With the large scale cultivation of transgenic crops expressing Bacillus thuringiensi.s (Bt) insecticidal crystal proteins in the world, the problem of environmental safety caused by these Bt crops has received extensive attention.These insecticidal crystal proteins can be released into the soil continuously in the growing period of Bt plants.If their accumulation of the insecticidal crystal proteins exceeds consumption by insect larvae and degradation by the environmental factors,these insecticidal crystal proteins could constitute a hazard to non-target insects and soil microbiota.There are three main ways to release insecticidal crystal proteins into soil for Bt plants: root exudates, pollen falling, and crop reside retuming.The Bt insecticidal crystal proteins released into soil can be adsorbed rapidly by active soil particles and the absorption equilibrium attained within l-3 h.The adsorption protects Bt insecticidal crystal proteins against soil microbial degradation or enzyme degradation, which leads to remarkable prolong of the persistence of insecticidal activity.The change of soil microorganism species is an important index for evaluating the effect of Bt plants on soil ecology.The research showed that these insecticidal crystal proteins released by the Bt plant root exudates or Bt organism had no toxicity to the soil earthworms, nematodes, protozoa, bacteria and fungi; however, it could reduce the mycelium length of the arbuscular mycorrhizal fungi (AMF) and restrain AMF to form mvasion unit.The influencing degree of Bt protein on soil enzyme activity varied with the releasing modes or growth period of Bt crops.Bt CryIAb protein can be taken up from soil by parts of following crops; however, different results were obtained with different commercial kits.To better understand the soil ecological evaluation about the insecticidal crystal proteins released from transgenic Bt crops, this review provides a comprehensive overview about the release, adsorption

  11. Development of transgenic sorghum for insect resistance against the spotted stem borer (Chilo partellus).

    Science.gov (United States)

    Girijashankar, V; Sharma, H C; Sharma, Kiran K; Swathisree, V; Prasad, L Sivarama; Bhat, B V; Royer, Monique; Secundo, Blanca San; Narasu, M Lakshmi; Altosaar, I; Seetharama, N

    2005-11-01

    Transgenic sorghum plants expressing a synthetic cry1Ac gene from Bacillus thuringiensis (Bt) under the control of a wound-inducible promoter from the maize protease inhibitor gene (mpiC1) were produced via particle bombardment of shoot apices. Plants were regenerated from the transformed shoot apices via direct somatic embryogenesis with an intermittent three-step selection strategy using the herbicide Basta. Molecular characterisation based on polymerase chain reaction and Southern blot analysis revealed multiple insertions of the cry1Ac gene in five plants from three independent transformation events. Inheritance and expression of the Bt gene was confirmed in T(1) plants. Enzyme-linked immunosorbant assay indicated that Cry1Ac protein accumulated at levels of 1-8 ng per gram of fresh tissue in leaves that were mechanically wounded. Transgenic sorghum plants were evaluated for resistance against the spotted stem borer (Chilo partellus Swinhoe) in insect bioassays, which indicated partial resistance to damage by the neonate larvae of the spotted stem borer. Reduction in leaf damage 5 days after infestation was up to 60%; larval mortality was 40%, with the surviving larvae showing a 36% reduction in weight over those fed on control plants. Despite the low levels of expression of Bt delta-endotoxin under the control of the wound-inducible promoter, the transgenic plants showed partial tolerance against first instar larvae of the spotted stem borer.

  12. 转Bt基因玉米叶片中Bt蛋白的表达和分泌研究%Study on Expression and Exudation of Bt Protein in Leaves of Transgenic Insect- resistant Corn

    Institute of Scientific and Technical Information of China (English)

    田晓燕; 赵蕾; 孙红炜; 李凡; 赵辉; 颜世磊; 路兴波

    2012-01-01

    With the transgenic insect - resistant corn Mon810 and its non - transgenic parent as materials , the enzyme - linked immunosorbent assay (ELISA ) method was used to detect the expression and quanti-tate the exudation of Bt protein in corn leaves. The results showed that Bt protein could be detected in tissue and on surface of leaves. The Bt content in tissues showed a rising trend from seedling stage to silking stage with the advances of vegetable growth, and reached the peak at the silking stage and then declined gradually. The variation of Bt exudation on leaf surface had no obvious regularity.%以转Bt玉米Mon810及其亲本非转基因玉米为研究对象,通过酶联免疫吸附测定法(ELISA)定量检测Mon810玉米叶中Bt蛋白表达量和叶表Bt蛋白分泌量,结果表明叶组织和叶表均可检测到Bt蛋白,组织内Bt蛋白量从苗期到抽丝期随着植株营养生长的加快呈上升趋势,至抽丝期达到顶峰,之后逐渐降低;叶表Bt蛋白分泌量变化无明显规律性.

  13. Effect on Stresses of 18°C and Different Relative Humidities on Bt Protein Ex-pression at Squaring Stage in Bt Cotton%蕾期低温及湿度胁迫对Bt棉杀虫蛋白表达量的影响

    Institute of Scientific and Technical Information of China (English)

    陈源; 顾超; 王桂霞; 吕春花; 刘晓飞; 张祥; 陈德华

    2013-01-01

      以Bt基因来源于美国和我国的常规棉花品种DP410B和泗抗1号、杂交种岱杂1号和泗抗3号为材料,应用盆栽试验,探讨蕾期18℃低温及不同相对湿度不同时间胁迫对叶片 Bt 杀虫蛋白表达量的影响.结果表明,蕾期18℃下不同湿度6 h胁迫对叶片Bt蛋白的含量都没有显著影响,但胁迫48 h导致2个不同Bt来源常规品种的叶片Bt杀虫蛋白表达量显著下降,与对照相比,下降6.8%~7.2%,杂交种泗抗3号也明显下降,岱杂1号则未受影响;18℃下高湿度与低湿度胁迫间没有显著影响.因此蕾期长时低温会对Bt棉抗虫性有一定影响,影响程度与品种及类型密切相关.%The objective of this study was to investigate the effect of stresses of 18°C and different relative humidities on the leaf resistance to the insect at squaring stage in Bt cotton. The two conventional cultivars (DP410B and Sikang 1) and two hybrids (Daiza 1 and Sikang 3) from the US and China, respectively, were used. The potted cotton plants were stressed at 18°C and dif-ferent relative humidities (90%, 70%, and 50%) for 6 hours and 48 hours, respectively, then we determined leaf Bt protein con-tents. The results showed that the short period (6 hours) stress of 18°C and different relative humidities obviously had no effect on the leaf Bt protein contents. but the leaf Bt protein contents sharply reduced in the two Bt conventional cultivars after the longer period (48 hours) stress of 18°C and different relative humidities compared with the control, with the reduction from 6.8%to 7.2%. The leaf Bt protein content of hybrid Sikang 3 markedly decreased while that of hybrid Daiza 1 remained unchanged. There was no difference in the effect between the high relative humidity and low relative humidity at 18°C, so the low temperature duration may affect the Bt cotton resistance to insect at squaring stage, which is closely related to cultivars.

  14. The Binding Characterization of Cry Insecticidal Proteins to the Brush Border Membrane Vesicles of Helicoverpa armigera, Spodoptera exigua, Spodoptera litura and Agrotis ipsilon

    Institute of Scientific and Technical Information of China (English)

    LU Qiong; CAO Guang-chun; ZHANG Li-li; LIANG Ge-mei; GAO Xi-wu; ZHANG Yong-jun; GUO Yu-yuan

    2013-01-01

    Cry toxins produced by Bacillus thuringiensis (Bt) are effective biological insecticides against certain insect species. However, there are potential risks of the evolved resistance of insects to Cry toxin owing to decreased binding of toxins to target sites in the brush border membranes of the larva midgut. The Cry toxins with different binding sites in the larval midgut have been considered to be a good combination to deploy in delaying resistance evolution. Bioassay results demonstrated that the toxicity of different Cry toxins ranked differently for each species. The toxicity ranking was Cry1Ac>Cry1Ab>Cry2Ab for Helicoverpa armigera, Cry1B>Cry1C>Cry2Ab for Spodoptera exigua, and Cry2Ab>Cry1B>Cry1C for S. litura. Only Cry2Ab was toxic to Agrotis ipsilon. Binding experiments were performed with 125I-Cry1Ab, 125I-Cry1Ac, 125I-Cry1B, 125I-Cry1C, 125I-Cry2Ab and the brush border membranes vesicles (BBMV) from H. armigera, S. exigua, S. litura and A. ipsilon. The binding of Cry1Ab and Cry1Ac was shown to be saturable by incubating with increasing concentrations of H. armigera BBMV (Kd=(45.00±2.01) nmol L-1 and (12.80±0.18) nmol L-1, respectively;Bmax=(54.95±1.79) ng and (55.44±0.91) ng, separately). The binding of Cry1B was shown to be saturable by incubating with increasing concentrations of S. exigua BBMV (Kd=(23.26±1.66) nmol L-1;Bmax=(65.37±1.87) ng). The binding of 125I-Cry toxins was shown to be non-saturable by incubating with increasing concentrations of S. litura and A. ipsilon BBMV. In contrast, Cry1B and Cry1C showed some combination with the BBMV of S. litura, and a certain amount of Cry2Ab could bind to the BBMV of A. ipsilon. These observations suggest that a future strategy could be devised for the focused combination of specific cry genes in transgenic crops to control target pests, widen the spectrum of insecticide effectiveness and postpone insect resistance evolution.

  15. Can pyramids and seed mixtures delay resistance to Bt crops?

    Science.gov (United States)

    The primary strategy for delaying evolution of pest resistance to transgenic crops that produce insecticidal proteins from Bacillus thuringiensis (Bt) entails refuges of plants that do not produce Bt toxins and thus allow survival of susceptible pests. Recent advances include using refuges together...

  16. 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.

  17. DECOMPOSITION OF BT COTTON AND NON BT COTTON RESIDUES UNDER VARIED SOIL TYPES

    Directory of Open Access Journals (Sweden)

    Sujata Kumari

    2014-04-01

    Full Text Available Use of the insecticidal cry proteins from the bacterium, Bacillus thuringiensis (Bt in cotton has raised a number of concerns, including the ecological impact on soil ecosystems.Greenhouse study was conducted during the 2011 wet season (March to August at the Institute of Agricultural Sciences of Banaras Hindu University. It was carried out on three different soil orders that includedentisol, inceptisol and alfisol. Bt cotton (var.NCS-138 and its non-transgenic isoline (var.NCS-138 were grown until maturity. A no crop pot was maintained for all the three soil orders. The highest rate of decomposition was found in alluvial soil compared to black and red soils in 50 days after incorporation (DAI. Thereafter the rate of decomposition was slowed downby100 DAI and the constant rate of decomposition was found in 150 DAI. The rate of decomposition was higher in non Bt than Bt crop residues.

  18. 利用分子标记辅助选择培育转 Bt基因抗虫水稻%Breeding of Trans-Bt Gene Rice Varieties with Pest Resistance by Means of Marker-Assisted Selection (MAS)

    Institute of Scientific and Technical Information of China (English)

    张琳; 魏志刚; 裴婷婷; 胡颂平; 余霞; 张静; 罗利军; 叶水峰; 刘国兰

    2014-01-01

    为培育转Bt基因抗虫水稻品种材料,以传统杂交的方法将3个Bt明恢63中的Bt基因cry1C*、cry2A*和cry1Ac/Ab,转育到水稻品种秀水123、湘晴、旱恢3号和沪旱1B中;以回交二代自交三代(BC2F3)水稻叶片为材料,利用分子标记辅助选择、田间除草剂筛选和试纸条检测方法,以获得新型转Bt基因株系。并在全生育期实行除不喷施农药外的正常田间管理的转基因植株的田间抗虫鉴定方法筛选出抗虫性良好、农艺性状优良的株系,为后续抗虫育种提供材料。 BC2 F3代PCR检测及试纸条检测的结果显示分别获得cry1C*和cry2A*基因的沪旱1B、秀水123和湘晴的纯合株系。同时,在田间自然诱发虫害条件下,转基因株系对稻纵卷叶螟虫表现出强抗性。%In order to breed trans-Bt gene rice materials with pest resistance,three Bt genes,cry1Ac/Ab, cry1C* and cry2A*,from the donor parents developed by Huazhong Agricultural University were introgressed into Xiushui123,Xiangqing,Hanhui No.3 and Huhan1B,namely Minghui63( Cry1C*) ,Minghui63( Cry2A*) and Minghui63 ( Cry1Ab/Ac) respectively using the method of traditional hybrid;then the new type lines of trans-Bt gene rice were by obtained means of maker-assisted selection ( MAS) ,screening of basta test in the field and test paper strip detection.In addition trans-gene rice lines of good pest resistance and fine agricultural traits were scteened by using the assays of insect-resistance in the field under normal field management but without spraying pesticide in the whole bearing stage of these trans-gene lines providing the materials for fur-ther anti-pest breeding.The homozygous lines with cry1C* and cry2A were obtained in the different back-grounds ( Xiushui123,Xiangqing and Huhan1B) respectively by the detection of PCR and test paper strip.At the same time,Bt lines showed excellent resistance to rice leaf roller in the field under the condition of its natu

  19. Imidacloprid affects the functional response of predator Podisus nigrispinus (Dallas) (Heteroptera: Pentatomidae) to strains of Spodoptera frugiperda (J.E. Smith) on Bt cotton.

    Science.gov (United States)

    Malaquias, J B; Ramalho, F S; Omoto, C; Godoy, W A C; Silveira, R F

    2014-03-01

    Podisus nigrispinus (Dallas) (Heteroptera: Pentatomidae) is one of the most common asopine species in the neotropical region and its occurrence was reported in several countries of South and Central America, as an important biological control agent for many crops. This study was carried out to identify the imidacloprid impacts on the functional response of predator P. nigrispinus fed on Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) strain resistant to lambda-cyhalothrin, on Bt cotton expressing Cry1Ac (Bollgard(®)). Spodoptera frugiperda larvae were used in the following conditions: resistant (1) and susceptible (2) strains to lambda-cyhalothrin fed on Bollgard(®) cotton leaves (DP 404 BG); and resistant (3) and susceptible (4) strains to lambda-cyhalothrin fed on non-genetically modified cotton leaves (cultivar DP4049). The predatory behavior of P. nigrispinus was affected by imidacloprid and the type II asymptotic curve was the one that best described the functional response data. Handling time (T h ) of predator females did not differ among treatments in the presence of imidacloprid. The attack rate did decrease, however, due to an increase in the density of larvae offered. Regardless of the treatment (S. frugiperda strain or cotton cultivar), the predation of P. nigrispinus females on S. frugiperda larvae was significantly lower when exposed to imidacloprid, especially at a density of 16 larvae/predator. The predation behavior of P. nigrispinus on S. frugiperda larvae is affected by the insecticide imidacloprid showing that its applications should be used in cotton crop with caution.

  20. E.QUALITY@BT...

    Science.gov (United States)

    Macmillan, Roderick H.

    1996-01-01

    Describes a management system developed by BT Laboratories (United Kingdom) that is based on ISO 9001 using the World Wide Web, a hypermedia system, and part of the Internet. Subject matter is presented as an alphabetical list of linked entries, numerous navigational techniques are available, and searching options function within an index file.…

  1. [Effects of transgenic Bt crops on non-target soil animals].

    Science.gov (United States)

    Yuan, Yi-gang; Ge, Feng

    2010-05-01

    Transgenic Bt crops are widely planted around the world. With the quick development and extension of genetically modified crops, it is needed to make a deep study on the effects of Bt crops on soil ecosystem. This paper reviewed the research progress on the effects of transgenic Bt crops on the population dynamics and community structure of soil animals, e.g., earthworm, nematode, springtail, mite, and beetle, etc. The development history of Bt crops was introduced, the passway the Bt protein comes into soil as well as the residual and degradation of Bt protein in soil were analyzed, and the critical research fields about the ecological risk analysis of transgenic Bt crops on non-target soil animals in the future were approached, which would provide a reference for the research of the effects of transgenic Bt crops on non-target soil animals.

  2. Green algae (Chlorella pyrenoidosa) adsorbs Bacillus thurigiensis (Bt) toxin, Cry1Ca insecticidal protein, without an effect on growth.

    Science.gov (United States)

    Wang, Jiamei; Chen, Xiuping; Li, Yunhe; Su, Changqing; Ding, Jiatong; Peng, Yufa

    2014-08-01

    The effect of purified Cry1Ca insecticidal protein on the growth of Chlorella pyrenoidosa was studied in a three-generation toxicity test. The C. pyrenoidosa medium with a density of 5.4 × 10(5) cells/mL was subcultured for three generations with added Cry1Ca at 0, 10, 100, and 1000 µg/L, and cell numbers were determined daily. To explore the distribution of Cry1Ca in C. pyrenoidosa and the culture medium, Cry1Ca was added at 1000 µg/L to algae with a high density of 4.8 × 10(6) cells/mL, and Cry1Ca content was determined daily in C. pyrenoidosa and the culture medium by enzyme-linked immunosorbent assays. Our results showed that the growth curves of C. pyrenoidosa exposed to 10, 100, and 1000 µg/L of Cry1Ca almost overlapped with that of the blank control, and there were no statistically significant differences among the four treatments from day 0 to day 7, regardless of generation. Moreover, the Cry1Ca content in the culture medium and in C. pyrenoidosa sharply decreased under exposure of 1000 µg/L Cry1Ca with high initial C. pyrenoidosa cell density. The above results demonstrate that Cry1Ca in water can be rapidly adsorbed and degraded by C. pyrenoidosa, but it has no suppressive or stimulative effect on algae growth.

  3. Quality of laboratory studies assessing effects of Bt-proteins on non-target organisms: minimal criteria for acceptability.

    Science.gov (United States)

    De Schrijver, Adinda; Devos, Yann; De Clercq, Patrick; Gathmann, Achim; Romeis, Jörg

    2016-08-01

    The potential risks that genetically modified plants may pose to non-target organisms and the ecosystem services they contribute to are assessed as part of pre-market risk assessments. This paper reviews the early tier studies testing the hypothesis whether exposure to plant-produced Cry34/35Ab1 proteins as a result of cultivation of maize 59122 is harmful to valued non-target organisms, in particular Arthropoda and Annelida. The available studies were assessed for their scientific quality by considering a set of criteria determining their relevance and reliability. As a case-study, this exercise revealed that when not all quality criteria are met, weighing the robustness of the study and its relevance for risk assessment is not obvious. Applying a worst-case expected environmental concentration of bioactive toxins equivalent to that present in the transgenic crop, confirming exposure of the test species to the test substance, and the use of a negative control were identified as minimum criteria to be met to guarantee sufficiently reliable data. This exercise stresses the importance of conducting studies meeting certain quality standards as this minimises the probability of erroneous or inconclusive results and increases confidence in the results and adds certainty to the conclusions drawn.

  4. Development of Bt Rice and Bt Maize in China and Their Efficacy in Target Pest Control

    Science.gov (United States)

    Liu, Qingsong; Hallerman, Eric; Peng, Yufa; Li, Yunhe

    2016-01-01

    Rice and maize are important cereal crops that serve as staple foods, feed, and industrial material in China. Multiple factors constrain the production of both crops, among which insect pests are an important one. Lepidopteran pests cause enormous yield losses for the crops annually. In order to control these pests, China plays an active role in development and application of genetic engineering (GE) to crops, and dozens of GE rice and GE maize lines expressing insecticidal proteins from the soil bacterium Bacillus thuringiensis (Bt) have been developed. Many lines have entered environmental release, field testing, and preproduction testing, and laboratory and field experiments have shown that most of the Bt rice and Bt maize lines developed in China exhibited effective control of major target lepidopteran pests on rice (Chilo suppressalis, Scirpophaga incertulas, and Cnaphalocrocis medinalis) and maize (Ostrinia furnacalis), demonstrating bright prospects for application. However, none of these Bt lines has yet been commercially planted through this writing in 2016. Challenges and perspectives for development and application of Bt rice and maize in China are discussed. This article provides a general context for colleagues to learn about research and development of Bt crops in China, and may shed light on future work in this field. PMID:27763554

  5. Development of Bt Rice and Bt Maize in China and Their Efficacy in Target Pest Control

    Directory of Open Access Journals (Sweden)

    Qingsong Liu

    2016-10-01

    Full Text Available Rice and maize are important cereal crops that serve as staple foods, feed, and industrial material in China. Multiple factors constrain the production of both crops, among which insect pests are an important one. Lepidopteran pests cause enormous yield losses for the crops annually. In order to control these pests, China plays an active role in development and application of genetic engineering (GE to crops, and dozens of GE rice and GE maize lines expressing insecticidal proteins from the soil bacterium Bacillus thuringiensis (Bt have been developed. Many lines have entered environmental release, field testing, and preproduction testing, and laboratory and field experiments have shown that most of the Bt rice and Bt maize lines developed in China exhibited effective control of major target lepidopteran pests on rice (Chilo suppressalis, Scirpophaga incertulas, and Cnaphalocrocis medinalis and maize (Ostrinia furnacalis, demonstrating bright prospects for application. However, none of these Bt lines has yet been commercially planted through this writing in 2016. Challenges and perspectives for development and application of Bt rice and maize in China are discussed. This article provides a general context for colleagues to learn about research and development of Bt crops in China, and may shed light on future work in this field.

  6. A seed mixture increases dominance of resistance to Bt cotton in Helicoverpa zea

    OpenAIRE

    2015-01-01

    Widely grown transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) can benefit agriculture, but adaptation by pests threatens their continued success. Refuges of host plants that do not make Bt toxins can promote survival of susceptible insects and delay evolution of resistance, particularly if resistance is inherited as a recessive trait. However, data have been lacking to compare the dominance of resistance when Bt and non-Bt seeds are planted in random mixtures ...

  7. Comparative Analysis of Genomics and Proteomics in the New Isolated Bacillus thuringiensis X022 Revealed the Metabolic Regulation Mechanism of Carbon Flux Following Cu(2+) Treatment.

    Science.gov (United States)

    Quan, Meifang; Xie, Junyan; Liu, Xuemei; Li, Yang; Rang, Jie; Zhang, Tong; Zhou, Fengjuan; Xia, Liqiu; Hu, Shengbiao; Sun, Yunjun; Ding, Xuezhi

    2016-01-01

    Bacillus thuringiensis (Bt) X022 is a novel strain isolated from soil in China, and showed strong insecticidal activity against several Lepidopteran pests. In this work, we performed whole genome sequencing of this Bt strain using the next-generation sequencing technology, and further conducted a comparative analysis with the proteomics data of the specific spore-release period based on LC-MS/MS approach. The Bt X022 genome consisted of one circular chromosomal DNA and seven plasmids, which were further functionally annotated using the RAST server. Comparative analysis of insecticidal substances showed that X022 contained genes coding for three Cry proteins (Cry1Ac, Cry1Ia and Cry2Ab) and a vegetative insecticidal protein (Vip3A). However, three insecticidal crystal proteins (ICPs) (Cry1Ca, Cry1Ac and Cry1Da) were detected by proteomics in the spore-release period. Moreover, a putative biosynthetic gene cluster and the metabolic pathway for poly-β-hydroxybutyrate in Bt X022 were deduced based on the comparative analysis of genomic and proteomic data, which revealed the metabolic regulation mechanism of carbon flux correlated with increased production of ICPs caused by Cu(2+.) Hence, these results provided a deeper understanding of the genetic background and protein expression profile of Bt X022. This study established a foundation for directed genetic modification and further application of this new isolated Bt strain.

  8. Comparative analysis of genomics and proteomics in the new isolated Bacillus thuringiensis X022 revealed the metabolic regulation mechanism of carbon flux following Cu2+ treatment

    Directory of Open Access Journals (Sweden)

    Meifang eQuan

    2016-05-01

    Full Text Available Bacillus thuringiensis (Bt X022, which showed strong insecticidal activity against several Lepidopteran pests, is a novel strain isolated from soil in China. In this work, we performed whole genome sequencing of this Bt strain using the next-generation sequencing technology, and further conducted a comparative analysis with the proteomics data of the specific spore-release period based on LC-MS/MS approach. The Bt X022 genome consisted of one circular chromosomal DNA and seven plasmids, which were further functionally annotated using the RAST server. Comparative analysis of insecticidal substances showed that X022 contained genes coding for three Cry proteins (Cry1Ac, Cry1Ia and Cry2Ab and a vegetative insecticidal protein (Vip3A. However, three insecticidal crystal proteins (ICPs (Cry1Ca, Cry1Ac and Cry1Da were detected by proteomics in the spore-release period. Moreover, a putative biosynthetic gene cluster and the metabolic pathway for poly-β-hydroxybutyrate in Bt X022 were deduced based on the comparative analysis of genomic and proteomic data, which revealed the metabolic regulation mechanism of carbon flux correlated with increased production of ICPs caused by Cu2+ . Hence, these results provided a deeper understanding of the genetic background and protein expression profile of Bt X022. This study established a foundation for directed genetic modification and further application of this new isolated Bt strain.

  9. Comparative Analysis of Genomics and Proteomics in the New Isolated Bacillus thuringiensis X022 Revealed the Metabolic Regulation Mechanism of Carbon Flux Following Cu2+ Treatment

    Science.gov (United States)

    Quan, Meifang; Xie, Junyan; Liu, Xuemei; Li, Yang; Rang, Jie; Zhang, Tong; Zhou, Fengjuan; Xia, Liqiu; Hu, Shengbiao; Sun, Yunjun; Ding, Xuezhi

    2016-01-01

    Bacillus thuringiensis (Bt) X022 is a novel strain isolated from soil in China, and showed strong insecticidal activity against several Lepidopteran pests. In this work, we performed whole genome sequencing of this Bt strain using the next-generation sequencing technology, and further conducted a comparative analysis with the proteomics data of the specific spore-release period based on LC-MS/MS approach. The Bt X022 genome consisted of one circular chromosomal DNA and seven plasmids, which were further functionally annotated using the RAST server. Comparative analysis of insecticidal substances showed that X022 contained genes coding for three Cry proteins (Cry1Ac, Cry1Ia and Cry2Ab) and a vegetative insecticidal protein (Vip3A). However, three insecticidal crystal proteins (ICPs) (Cry1Ca, Cry1Ac and Cry1Da) were detected by proteomics in the spore-release period. Moreover, a putative biosynthetic gene cluster and the metabolic pathway for poly-β-hydroxybutyrate in Bt X022 were deduced based on the comparative analysis of genomic and proteomic data, which revealed the metabolic regulation mechanism of carbon flux correlated with increased production of ICPs caused by Cu2+. Hence, these results provided a deeper understanding of the genetic background and protein expression profile of Bt X022. This study established a foundation for directed genetic modification and further application of this new isolated Bt strain. PMID:27303381

  10. In vivo fluorescence observation of parasporal inclusion formation in Bacillus thuringiensis

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    A recombinant gene expressing a Cry1Ac-GFP fusion protein with a molecular mass of approximately 160 kD was constructed to investigate the expression of cry1Ac,the localization of its gene product Cry1Ac,and its role in crystal development in Bacillus thuringiensis.The cry1Ac-gfp fusion gene under the control of the cry1Ac promoter was cloned into the plasmid pHT304,and this construct was designated pHTcry1Ac-gfp.pHTcry1Ac-gfp was transformed into the crystal-negative strain,HD-73 cry-,and the resulting strain was named HD-73-(pHTcry1Ac-gfp).The gfp gene was then inserted into the large HD-73 endogenous plasmid pHT73 and fused with the 3’ terminal of the cry1Ac gene by homologous recombination,yielding HD-73Φ(cry1Ac-gfp)3534.Laser confocal microscopy and Western blot analyses showed for the first time that the Cry1Ac-GFP fusion proteins in both HD-73-(pHTcry1Ac-gfp) and HD-73Φ(cry1Ac-gfp)3534 were produced during asymmetric septum formation.Surprisingly,the Cry1Ac-GFP fusion protein showed polarity and was located near the septa in both strains.There was no significant difference between Cry1Ac-GFP and Cry1Ac in their toxicity to Plutella xylostella larvae.

  11. Molecular and insecticidal characterization of a Cry1I protein toxic to insects of the families Noctuidae, Tortricidae, Plutellidae, and Chrysomelidae.

    Science.gov (United States)

    Ruiz de Escudero, Iñigo; Estela, Anna; Porcar, Manuel; Martínez, Clara; Oguiza, José A; Escriche, Baltasar; Ferré, Juan; Caballero, Primitivo

    2006-07-01

    The most notable characteristic of Bacillus thuringiensis is its ability to produce insecticidal proteins. More than 300 different proteins have been described with specific activity against insect species. We report the molecular and insecticidal characterization of a novel cry gene encoding a protein of the Cry1I group with toxic activity towards insects of the families Noctuidae, Tortricidae, Plutellidae, and Chrysomelidae. PCR analysis detected a DNA sequence with an open reading frame of 2.2 kb which encodes a protein with a molecular mass of 80.9 kDa. Trypsin digestion of this protein resulted in a fragment of ca. 60 kDa, typical of activated Cry1 proteins. The deduced sequence of the protein has homologies of 96.1% with Cry1Ia1, 92.8% with Cry1Ib1, and 89.6% with Cry1Ic1. According to the Cry protein classification criteria, this protein was named Cry1Ia7. The expression of the gene in Escherichia coli resulted in a protein that was water soluble and toxic to several insect species. The 50% lethal concentrations for larvae of Earias insulana, Lobesia botrana, Plutella xylostella, and Leptinotarsa decemlineata were 21.1, 8.6, 12.3, and 10.0 microg/ml, respectively. Binding assays with biotinylated toxins to E. insulana and L. botrana midgut membrane vesicles revealed that Cry1Ia7 does not share binding sites with Cry1Ab or Cry1Ac proteins, which are commonly present in B. thuringiensis-treated crops and commercial B. thuringiensis-based bioinsecticides. We discuss the potential of Cry1Ia7 as an active ingredient which can be used in combination with Cry1Ab or Cry1Ac in pest control and the management of resistance to B. thuringiensis toxins.

  12. Gene cloning and expression of cadherin in midgut of Helicoverpa armigera and its Cry1A binding region

    Institute of Scientific and Technical Information of China (English)

    WANG Guirong; WU Kongming; LIANG Gemei; GUO Yuyuan

    2005-01-01

    Cadherins belong to one of the families of animal glycoproteins responsible for calcium-dependent cell-cell adhesion. Recent literatures showed that the cadherin-like in midgut of several insects served as the receptor of Bt toxin Cry1A and the variation of cadherin-like is related to insect's resistance to Cry1A. The full-length cDNA encoding cadherin-like of Helicoverpa armigera is cloned by degenerate PCR and RACE techniques and the gene was designated as BtR-harm, which is 5581 bp in full-length, encoding 1730 amino acid residues (BtR-harm was deposited in GenBank and the accession number is AF519180). Its predicted molecular weight and isoelectric point were 195.39 kDa and 4.23, respectively. The inferred amino acid sequence includes a signal sequence, 11 cadherin repeats, a membrane-proximal region, a transmembrane region and a cytoplasmic region. Sequence analysis indicated that the deduced protein sequence was most similar to the cadherin-like from Heliothis virescens with 84.2% identity and highly similar to three other lepidopteran cadherin from Bombyx mori, Manduca sexta and Pectinophora gossypiella, with the sequence identities of 60.3.6%, 57.5% and 51.0%, respectively. The cDNA encoding cadherin gene was expressed successfully in E. coli and the recombinant proteins can bind with Cry1Ac. Truncation analysis and binding experiment of BtR-harm revealed that the Cry1A binding region was a contiguous 244-amino acid sequence, which located between amino acid 1217 and 1461. Semi-quantitative RT-PCR analysis showed that BtR-harm was highly expressed in midgut of H. armigera, very low expressed in foregut and hindgut and was not expressed in other tissues. After H. armigera producing resistance to Cry1Ac, the expression quantity of BtR-harm significantly decreased in midgut of H. armigera. It is the first confirmation that BtR-harm can function as receptor of Cry1Ac in H. armigera and the binding region was located on a contiguous 244 amino acid sequence

  13. The Expression of Bt Toxin Protein in Transgenic Plants%Bt毒蛋白在转基因植物中的表达

    Institute of Scientific and Technical Information of China (English)

    魏俊杰

    2012-01-01

    With the promotion of insect-resistant genetically modified crops and large-scale commercial cultivation,Bt toxin in transgenic plants expression is increasingly attracted attention.In this paper,corn,cotton,rice and other insect-resistant transgenic crops expressing the Bt toxin were discussed.%随着抗虫转基因植物的推广和大面积商业化种植,Bt毒蛋白在转基因植物中的表达引起了人们的重视。综述了玉米、棉花、水稻等抗虫转基因植物的Bt毒蛋白表达及其时空变化规律。

  14. Three cadherin alleles associated with resistance to Bacillus thuringiensis in pink bollworm

    Science.gov (United States)

    Morin, Shai; Biggs, Robert W.; Sisterson, Mark S.; Shriver, Laura; Ellers-Kirk, Christa; Higginson, Dawn; Holley, Daniel; Gahan, Linda J.; Heckel, David G.; Carrière, Yves; Dennehy, Timothy J.; Brown, Judith K.; Tabashnik, Bruce E.

    2003-01-01

    Evolution of resistance by pests is the main threat to long-term insect control by transgenic crops that produce Bacillus thuringiensis (Bt) toxins. Because inheritance of resistance to the Bt toxins in transgenic crops is typically recessive, DNA-based screening for resistance alleles in heterozygotes is potentially much more efficient than detection of resistant homozygotes with bioassays. Such screening, however, requires knowledge of the resistance alleles in field populations of pests that are associated with survival on Bt crops. Here we report that field populations of pink bollworm (Pectinophora gossypiella), a major cotton pest, harbored three mutant alleles of a cadherin-encoding gene linked with resistance to Bt toxin Cry1Ac and survival on transgenic Bt cotton. Each of the three resistance alleles has a deletion expected to eliminate at least eight amino acids upstream of the putative toxin-binding region of the cadherin protein. Larvae with two resistance alleles in any combination were resistant, whereas those with one or none were susceptible to Cry1Ac. Together with previous evidence, the results reported here identify the cadherin gene as a leading target for DNA-based screening of resistance to Bt crops in lepidopteran pests. PMID:12695565

  15. Bt crops benefit natural enemies to control non-target pests.

    Science.gov (United States)

    Tian, Jun-Ce; Yao, Ju; Long, Li-Ping; Romeis, Jörg; Shelton, Anthony M

    2015-11-12

    Crops producing insecticidal crystal (Cry) proteins from Bacillus thuringiensis (Bt) control important lepidopteran pests. However, pests such as aphids not susceptible to Cry proteins may require other integrated pest management (IPM) tactics, including biological control. We fed aphids on Bt and non-Bt plants and analyzed the Bt protein residue in aphids and compared the effects of Bt plants and a pyrethroid, lambda-cyhalothrin, on the performance of three natural enemies (predators: Coleomegilla maculata and Eupeodes americanus; parasitoid Aphidius colemani) of the green peach aphid, Myzus persicae. No Bt protein residues in aphids were detected and no significant differences were recorded in the performance of pyrethroid-resistant aphids that fed on Bt broccoli expressing Cry1Ab or Cry1C, or on non-Bt broccoli plants treated or not treated with the pyrethroid. This indicated the aphids were not affected by the Cry proteins or the pyrethroid, thus removing any effect of prey quality. Tri-trophic experiments demonstrated that no C. maculata and E. americanus survived consumption of pyrethroid-treated aphids and that ovipositional behavior of A. colemani was impaired when provided with pyrethroid-treated aphids. In contrast, natural enemies were not affected when fed aphids reared on Bt broccoli, thus demonstrating the safety of these Bt plants for IPM.

  16. Response of the gypsy moth, Lymantria dispar to transgenic poplar, Populus simonii x P. nigra, expressing fusion protein gene of the spider insecticidal peptide and Bt-toxin C-peptide.

    Science.gov (United States)

    Cao, Chuan-Wang; Liu, Gui-Feng; Wang, Zhi-Ying; Yan, Shan-Chun; Ma, Ling; Yang, Chuan-Ping

    2010-01-01

    The response of the Asian gypsy moth Lymantria dispar (L.) (Lepidoptera: Lymantriidae) to a fusion gene consisting of the spider, Atrax robustus Simon (Araneae: Hexanthelidae) ω-ACTX-Ar1 sequence coding for an ω-atracotoxin and a sequence coding for the Bt-toxin C-peptide, expressed in transgenic poplar Populus simonii x P. nigra L. (Malphigiales: Salicaceae) was investigated. Individual performance, feeding selection, midgut proteinase activity and nutrition utilization were monitored. The growth and development of L. dispar were significantly affected by continually feeding on the transgenic poplar, with the larval instars displaying significantly shorter developmental times than those fed on nontransgenic poplar, but pupation was delayed. Mortality was higher in populations fed transgenic poplar leaves, than for larvae fed nontransgenic poplar leaves. The cumulative mortality during all stages of larvae fed transgenic leaves was 92% compared to 16.7% of larvae on nontransgenic leaves. The highest mortality observed was 71.7% in the last larval instar stage. A two-choice test showed that fifth-instar larvae preferred to feed on nontransgenic leaves at a ratio of 1:1.4. Feeding on transgenic leaves had highly significant negative effects on relative growth of larvae, and the efficiency of conversion of ingested and digested food. Activity of major midgut proteinases was measured using substrates TAME and BTEE showed significant increases in tryptase and chymotrypsinlike activity (9.2- and 9.0-fold, respectively) in fifth-instar larvae fed on transgenic leaves over control. These results suggest transgenic poplar is resistant to L. dispar, and the mature L. dispar may be weakened by the transgenic plants due to Bt protoxins activated by elevated major midgut proteinase activity. The new transgenic poplar expressing fusion protein genes of Bt and a new spider insecticidal peptide are good candidates for managing gypsy moth.

  17. Advances in genetic engineering of insect-resistant Populus spp. with Bt toxic protein genes%杨树Bt抗虫基因工程研究进展

    Institute of Scientific and Technical Information of China (English)

    张蕾; 崔建国; 王洪魁

    2005-01-01

    本文介绍了苏云金芽孢杆菌(Bt)毒蛋白的杀虫机理和Bt毒蛋白基因的分类;概述了Bt毒蛋白基因在杨树基因工程中的应用现状,探讨了当前杨树Bt抗虫基因工程中存在的主要问题,并展望了杨树抗虫基因工程在杨树遗传改良中的应用前景.

  18. Insect resistance to Bt crops: lessons from the first billion acres.

    Science.gov (United States)

    Tabashnik, Bruce E; Brévault, Thierry; Carrière, Yves

    2013-06-01

    Evolution of resistance in pests can reduce the effectiveness of insecticidal proteins from Bacillus thuringiensis (Bt) produced by transgenic crops. We analyzed results of 77 studies from five continents reporting field monitoring data for resistance to Bt crops, empirical evaluation of factors affecting resistance or both. Although most pest populations remained susceptible, reduced efficacy of Bt crops caused by field-evolved resistance has been reported now for some populations of 5 of 13 major pest species examined, compared with resistant populations of only one pest species in 2005. Field outcomes support theoretical predictions that factors delaying resistance include recessive inheritance of resistance, low initial frequency of resistance alleles, abundant refuges of non-Bt host plants and two-toxin Bt crops deployed separately from one-toxin Bt crops. The results imply that proactive evaluation of the inheritance and initial frequency of resistance are useful for predicting the risk of resistance and improving strategies to sustain the effectiveness of Bt crops.

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

    Science.gov (United States)

    Knaak, N; Franz, A R; Santos, G F; Fiuza, L M

    2010-08-01

    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 microgxcm-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.

  1. THE EFFECT OF ARTIFICIAL INOCULATION WITH SELECTED FUSARIUM STRAINS ON NUTRITIONAL QUALITY AN ENSILING PROCESS OF BT MAIZE

    Directory of Open Access Journals (Sweden)

    Ludmila KŘÍŽOVÁ

    2011-01-01

    Full Text Available The objective of the study was to compare the nutritive value and mycotoxin content of maize forage and silage of near isogenic control MONUMENTAL (C and Bt maize (MONSANTO, MON 810 that was either untreated (Bt or artificially inoculated with Fusarium strains (I-Bt. The inoculation was made in the growing crop in milk stage of maturity. Plants were harvested at the soft dough stage of maturity and ensiled in microsilage tubes. The content of forage dry matter (DM was 307.6 g/kg in C, 306.9 g/kg in Bt and 298.0 g/kg in I-Bt. All forages were positive for deoxynivalenol, aflatoxin, fumonisins and zearalenone (P>0.05. Content of DM was the lowest in I-Bt silage (285.5 g/kg and differed significantly from C (296.7 g/kg or Bt (303.7 g/kg, P<0.05. Content of crude protein (CP was the lowest in I-Bt silage (79.0 g/kg and differed significantly from C or Bt (85.7 or 81.9 g/kg, respectively, P<0.05. Silages Bt and I-Bt had lower pH (3.93 and 3.96, respectively than silage C (4.02, P<0.05. Silage I-Bt tended to have a higher degree of proteolysis 9.18 % measured as N-NH3 (% of total N than silages C or Bt (8.64 or 8.9 %, respectively, P>0.05. Lactic acid was predominant product of fermentation in all silages, however silage I-Bt tended to have lower content of lactic acid (20.96 g/kg than C or Bt (24.76 or 23.82 g/kg, P>0.05. I-Bt silage contained lower levels of eoxynivalenol (602 ppb than C or Bt silage (748 and 690 ppb, respectively, P<0.05. Content of fumonisins and zearalenone in C did not differ from I-Bt (P<0.05 but both were lower than in Bt (P<0.05. In conclusion, nutritional value a fermentation parameters of Bt silage were similar to C except of CP content and pH that was lower in Bt (P<0.05. I-Bt silage had lower content of DM, CP and fat than Bt silage (P<0.05. Controversially, concentrations of mycotoxins in I-Bt silage were lower than in Bt.

  2. How to cope with insect resistance to Bt toxins?

    Science.gov (United States)

    Bravo, Alejandra; Soberón, Mario

    2008-10-01

    Transgenic Bt crops producing insecticidal crystalline proteins from Bacillus thuringiensis, so-called Cry toxins, have proved useful in controlling insect pests. However, the future of Bt crops is threatened by the evolution of insect resistance. Understanding how Bt toxins work and how insects become resistant will provide the basis for taking measures to counter resistance. Here we review possible mechanisms of resistance and different strategies to cope with resistance, such as expression of several toxins with different modes of action in the same plant, modified Cry toxins active against resistant insects, and the potential use of Cyt toxins or a fragment of cadherin receptor. These approaches should provide the means to assure the successful use of Bt crops for an extended period of time.

  3. Bt crop effects on functional guilds of non-target arthropods: a meta-analysis.

    Directory of Open Access Journals (Sweden)

    L LaReesa Wolfenbarger

    Full Text Available BACKGROUND: Uncertainty persists over the environmental effects of genetically-engineered crops that produce the insecticidal Cry proteins of Bacillus thuringiensis (Bt. We performed meta-analyses on a modified public database to synthesize current knowledge about the effects of Bt cotton, maize and potato on the abundance and interactions of arthropod non-target functional guilds. METHODOLOGY/PRINCIPAL FINDINGS: We compared the abundance of predators, parasitoids, omnivores, detritivores and herbivores under scenarios in which neither, only the non-Bt crops, or both Bt and non-Bt crops received insecticide treatments. Predators were less abundant in Bt cotton compared to unsprayed non-Bt controls. As expected, fewer specialist parasitoids of the target pest occurred in Bt maize fields compared to unsprayed non-Bt controls, but no significant reduction was detected for other parasitoids. Numbers of predators and herbivores were higher in Bt crops compared to sprayed non-Bt controls, and type of insecticide influenced the magnitude of the difference. Omnivores and detritivores were more abundant in insecticide-treated controls and for the latter guild this was associated with reductions of their predators in sprayed non-Bt maize. No differences in abundance were found when both Bt and non-Bt crops were sprayed. Predator-to-prey ratios were unchanged by either Bt crops or the use of insecticides; ratios were higher in Bt maize relative to the sprayed non-Bt control. CONCLUSIONS/SIGNIFICANCE: Overall, we find no uniform effects of Bt cotton, maize and potato on the functional guilds of non-target arthropods. Use of and type of insecticides influenced the magnitude and direction of effects; insecticde effects were much larger than those of Bt crops. These meta-analyses underscore the importance of using controls not only to isolate the effects of a Bt crop per se but also to reflect the replacement of existing agricultural practices. Results will

  4. Potential Use of Proteinase Inhibitors, Avidin, and Other Bio-reagents for Synergizing Bt Performance and Delaying Resistance Development to Bt

    Science.gov (United States)

    After being ingested by target insects, the insecticidal proteins from Bacillus thuringiensis (Bt) need to go through a proteolytic process by insect midgut proteinases to become activated. At the same time, Bt can be hydrolyzed and degraded by midgut proteinases to become non-toxic to target insect...

  5. Harmonia axyridis (Coleoptera: Coccinellidae) exhibits no preference between Bt and non-Bt maize fed Spodoptera frugiperda (Lepidoptera: Noctuidae).

    Science.gov (United States)

    Dutra, Carla C; Koch, Robert L; Burkness, Eric C; Meissle, Michael; Romeis, Joerg; Hutchison, William D; Fernandes, Marcos G

    2012-01-01

    A recent shift in managing insect resistance to genetically engineered (GE) maize consists of mixing non-GE seed with GE seed known as "refuge in a bag", which increases the likelihood of predators encountering both prey fed Bt and prey fed non-Bt maize. We therefore conducted laboratory choice-test feeding studies to determine if a predator, Harmonia axyridis, shows any preference between prey fed Bt and non-Bt maize leaves. The prey species was Spodoptera frugiperda, which were fed Bt maize (MON-810), expressing the single Cry1Ab protein, or non-Bt maize. The predators were third instar larvae and female adults of H. axyridis. Individual predators were offered Bt and non-Bt fed prey larvae that had fed for 24, 48 or 72 h. Ten and 15 larvae of each prey type were offered to third instar and adult predators, respectively. Observations of arenas were conducted at 1, 2, 3, 6, 15 and 24 h after the start of the experiment to determine the number and type of prey eaten by each individual predator. Prey larvae that fed on non-Bt leaves were significantly larger than larvae fed Bt leaves. Both predator stages had eaten nearly all the prey by the end of the experiment. However, in all combinations of predator stage and prey age, the number of each prey type consumed did not differ significantly. ELISA measurements confirmed the presence of Cry1Ab in leaf tissue (23-33 µg/g dry weight) and S. frugiperda (2.1-2.2 µg/g), while mean concentrations in H. axyridis were very low (0.01-0.2 µg/g). These results confirm the predatory status of H. axyridis on S. frugiperda and that both H. axyridis adults and larvae show no preference between prey types. The lack of preference between Bt-fed and non-Bt-fed prey should act in favor of insect resistance management strategies using mixtures of GE and non-GE maize seed.

  6. Responses of the cutworm Spodoptera litura (Lepidoptera: Noctuidae) to two Bt corn hybrids expressing Cry1Ab

    Science.gov (United States)

    Yinghua, Shu; Yan, Du; Jin, Chen; Jiaxi, Wei; Jianwu, Wang

    2017-01-01

    To examine the responses of the secondary lepidopteran pest Spodoptera litura to two Bacillus thuringiensis (Bt) corn hybrids [5422Bt1 (Event Bt11), 5422CBCL (MON810)] expressing Cry1Ab, larval bioassays with Cry1Ab toxin, corn leaves or kernels and bagging on corn plants were conducted. The results showed that larvae displayed a similar performance when fed kernels, but not leaves of 5422Bt1, 5422CBCL and their near-isogenic non-Bt corn (5422). Significantly higher Cry1Ab amounts were detected in larvae fed leaves than kernels of both Bt hybrids, with different molecular weights of protein band in plants (72 and 90 kDa for 5422Bt1 and 5422CBCL, respectively), gut contents (65 kDa), feces (50 kDa), which indicated that larvae had lower ingestion, higher degradation and excretion of Cry1Ab when fed kernels not leaves of both Bt hybrids. Significantly higher levels of cadherin-like receptors and alkaline phosphatase transcripts were detected in larvae fed leaves than kernels of two Bt hybrids. Catalase, superoxide dismutase and glutathione-S-transferase activities in larvae fed 5422Bt1 leaves were significantly higher than that of 5422 treatments. Therefore, S. litura had low susceptibility to 5422Bt1 and 5422CBCL when larvae fed kernels not leaves of Bt corn. Additionally, S. litura presented a much stronger tolerance to 5422CBCL than 5422Bt1. PMID:28186125

  7. Food Safety of Bt Cry Proteins from Biotechnology-derived Crops%转基因抗虫作物中苏云金芽孢杆菌Cry蛋白的食品安全问题

    Institute of Scientific and Technical Information of China (English)

    王春玲; Bruce Hammond

    2011-01-01

    苏云金芽孢杆菌(Bt)微生物制剂是农业、林业和饮用水等领域用来控制靶标害虫幼虫的有效工具,至今已经有50余年的使用历史。同时其在美国、欧洲和其他一些国家被广泛用于经过认证的有机农业生产之中。目前已获审批的转基因Bt作物中最常使用的是Cry蛋白。Cry蛋白的作用机制、食品安全性以及致敏性已经经过啮齿类动物、农场动物和人体内试验和生物信息学研究的严格检验。Cry蛋白的杀虫作用只在靶标害虫的碱性消化道内,与中肠上皮细胞的特异蛋白受体结合才能起到杀虫作用,而其他非靶标生物体内(人类、猕猴、小鼠、大鼠和牛等)都被证明没有这种特异蛋白质受体。美国、欧洲和其他国家的管理机构都已经证实了转基因Bt作物和Cry蛋白在农作物和饮用水中残留的安全性。食物加工过程能够最大化地减少转基因作物中功能性Cry蛋白的摄入。转基因抗虫作物有利于降低农药杀虫剂的使用的同时,也能够有效防止玉米中伏马菌毒素的污染。%Bt microbial insecticides have been used to control larval insect pests that feed on agricultural crops, forest and drinking water for over 50 years. They are also widely used in certified organic agricultural food production in the United States, Europe and other countries. Cry proteins are most commonly introduced in registered Bt crops. Mode of action, food safety and allergenicity of Cry protein have been rigorously tested using rodent, farm animals, and human experiment and bioinformatics research tools. The insecticidal function of Cry protein only works in the alkaline environment of the insect gut where specific receptors on the membranes of midgut epithelial cells present. Other non-target organisms (humans, rhesus monkeys, mice, rats and cows etc.) lack these receptors. Regulatory agencies in US, EU and other countries have confirmed the history

  8. Effects of Transgenic Cry 1 Ac plus CpTI Cotton on the Bioecology of Main Parasitoids in Laboratory Studies

    Institute of Scientific and Technical Information of China (English)

    CUI Jin-jie; VANDER Weft Wopke; MA Yah; LUO Jun-yu

    2008-01-01

    @@ Effects of transgenic CrylAc plus CpTI cotton (double genes cotton) on development of main parasitoids were studied in the laboratory.Compared with the traditional cotton,the differences of the moulting rate and adult weight of cotton aphid-Aphydius from double gene cotton field were not obvious.

  9. Next-generation transgenic cotton: pyramiding RNAi and Bt counters insect resistance.

    Science.gov (United States)

    Ni, Mi; Ma, Wei; Wang, Xiaofang; Gao, Meijing; Dai, Yan; Wei, Xiaoli; Zhang, Lei; Peng, Yonggang; Chen, Shuyuan; Ding, Lingyun; Tian, Yue; Li, Jie; Wang, Haiping; Wang, Xiaolin; Xu, Guowang; Guo, Wangzhen; Yang, Yihua; Wu, Yidong; Heuberger, Shannon; Tabashnik, Bruce E; Zhang, Tianzhen; Zhu, Zhen

    2017-02-15

    Transgenic crops producing insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) are extensively cultivated worldwide. To counter rapidly increasing pest resistance to crops that produce single Bt toxins, transgenic plant 'pyramids' producing two or more Bt toxins that kill the same pest have been widely adopted. However, cross-resistance and antagonism between Bt toxins limit the sustainability of this approach. Here we describe development and testing of the first pyramids of cotton combining protection from a Bt toxin and RNA interference (RNAi). We developed two types of transgenic cotton plants producing double-stranded RNA (dsRNA) from the global lepidopteran pest Helicoverpa armigera designed to interfere with its metabolism of juvenile hormone (JH). We focused on suppression of JH acid methyltransferase (JHAMT), which is crucial for JH synthesis, and JH-binding protein (JHBP), which transports JH to organs. In 2015 and 2016, we tested larvae from a Bt-resistant strain and a related susceptible strain of H. armigera on seven types of cotton: two controls, Bt cotton, two types of RNAi cotton (targeting JHAMT or JHBP) and two pyramids (Bt cotton plus each type of RNAi). Both types of RNAi cotton were effective against Bt-resistant insects. Bt cotton and RNAi acted independently against the susceptible strain. In computer simulations of conditions in northern China, where millions of farmers grow Bt cotton as well as abundant non-transgenic host plants of H. armigera, pyramided cotton combining a Bt toxin and RNAi substantially delayed resistance relative to using Bt cotton alone.

  10. Exploring the midgut transcriptome and brush border membrane vesicle proteome of the rice stem borer, Chilo suppressalis (Walker.

    Directory of Open Access Journals (Sweden)

    Weihua Ma

    Full Text Available The rice stem borer, Chilo suppressalis (Walker (Lepidoptera: Pyralidae, is one of the most detrimental pests affecting rice crops. The use of Bacillus thuringiensis (Bt toxins has been explored as a means to control this pest, but the potential for C. suppressalis to develop resistance to Bt toxins makes this approach problematic. Few C. suppressalis gene sequences are known, which makes in-depth study of gene function difficult. Herein, we sequenced the midgut transcriptome of the rice stem borer. In total, 37,040 contigs were obtained, with a mean size of 497 bp. As expected, the transcripts of C. suppressalis shared high similarity with arthropod genes. Gene ontology and KEGG analysis were used to classify the gene functions in C. suppressalis. Using the midgut transcriptome data, we conducted a proteome analysis to identify proteins expressed abundantly in the brush border membrane vesicles (BBMV. Of the 100 top abundant proteins that were excised and subjected to mass spectrometry analysis, 74 share high similarity with known proteins. Among these proteins, Western blot analysis showed that Aminopeptidase N and EH domain-containing protein have the binding activities with Bt-toxin Cry1Ac. These data provide invaluable information about the gene sequences of C. suppressalis and the proteins that bind with Cry1Ac.

  11. Exploring the midgut transcriptome and brush border membrane vesicle proteome of the rice stem borer, Chilo suppressalis (Walker).

    Science.gov (United States)

    Ma, Weihua; Zhang, Zan; Peng, Chuanhua; Wang, Xiaoping; Li, Fei; Lin, Yongjun

    2012-01-01

    The rice stem borer, Chilo suppressalis (Walker) (Lepidoptera: Pyralidae), is one of the most detrimental pests affecting rice crops. The use of Bacillus thuringiensis (Bt) toxins has been explored as a means to control this pest, but the potential for C. suppressalis to develop resistance to Bt toxins makes this approach problematic. Few C. suppressalis gene sequences are known, which makes in-depth study of gene function difficult. Herein, we sequenced the midgut transcriptome of the rice stem borer. In total, 37,040 contigs were obtained, with a mean size of 497 bp. As expected, the transcripts of C. suppressalis shared high similarity with arthropod genes. Gene ontology and KEGG analysis were used to classify the gene functions in C. suppressalis. Using the midgut transcriptome data, we conducted a proteome analysis to identify proteins expressed abundantly in the brush border membrane vesicles (BBMV). Of the 100 top abundant proteins that were excised and subjected to mass spectrometry analysis, 74 share high similarity with known proteins. Among these proteins, Western blot analysis showed that Aminopeptidase N and EH domain-containing protein have the binding activities with Bt-toxin Cry1Ac. These data provide invaluable information about the gene sequences of C. suppressalis and the proteins that bind with Cry1Ac.

  12. Effects of Cry1Ab Bt maize straw return on bacterial community of earthworm Eisenia fetida.

    Science.gov (United States)

    Shu, Yinghua; Zhang, Yanyan; Zeng, Huilan; Zhang, Yahui; Wang, Jianwu

    2017-04-01

    The eco-toxicological effects of Bacillus thuringiensis (Bt) maize on earthworm life-history traits were widely studied and the results were controversial, while their effects on earthworm bacterial community have been rarely studied. Here, effects of two hybrids of Bt maize [5422Bt1 (event Bt11) and 5422CBCL (MON810)] straw return on Eisenia fetida bacterial community were investigated by the terminal restriction fragment length polymorphism (T-RFLP) and polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) combing with DNA sequencing, compared to near-isogenic non-Bt maize (5422). Bt maize straw return had significant effects on soil nutrients, especially for available nitrogen (N). The significant differences were shown in soil bacterial community between Bt and non-Bt maize treatments on the 75(th) and 90(th) d, which was closely correlated with soil available N, P and K rather than Cry1Ab protein. There was no statistically significant difference in the bacterial community of earthworm gut contents between Bt and non-Bt maize treatments. The significant differences in the bacterial community of earthworm casts were found among three maize varieties treatments, which were closely correlated with Cry1Ab protein and N levels. The differentiated bacterial species in earthworm casts mainly belonged to Proteobacteria, including Brevundimonas, Caulobacter, Pseudomonas, Stenotrophomonas, Methylobacterium, Asticcacaulis and Achromobacter etc., which were associated with the mineralization, metabolic process and degradation of plants residues. Therefore, Bt maize straw return caused changes in the bacterial community of E. fetida casts, which was possibly caused by the direct (Cry1Ab protein) and non-expected effects (N levels) of Bt maize straw.

  13. Characterization of NCAM expression and function in BT4C and BT4Cn glioma cells

    DEFF Research Database (Denmark)

    1991-01-01

    -substratum binding assay in which the binding of BT4C and BT4Cn cells to NCAM immobilized to glass was assessed. We found that BT4C cells adhere specifically to NCAM, and that adhesion is inhibited by anti-NCAM Fab'-fragments, while no specific binding of BT4Cn cells to NCAM was observed. The BT4C and BT4Cn cell...

  14. 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.

  15. Complete genome sequence of Bacillus thuringiensis serovar galleriae strain HD-29, a typical strain of commercial biopesticide.

    Science.gov (United States)

    Zhu, Lei; Tian, Long-Jun; Zheng, Jinshui; Gao, Qiu-Ling; Wang, Yue-Ying; Peng, Dong-Hai; Ruan, Li-Fang; Sun, Ming

    2015-02-10

    Bacillus thuringiensis serovar galleriae is highly toxic to Lepidoptera insect pests, and has been widely used as Bt biopesticide in many countries. Here we reported the complete genome of strain HD-29, a standard serotype strain in galleriae serovariety. More than previous work reported, it harbors ten plasmids, and three large ones carry eight insecticidal protein genes (cry1Aa, cry1Ac, cry1Ca, cry1Da, cry1Ia, cry2Ab, cry9Ea and vip3Aa) and an intact zwittermicin A biosynthetic gene cluster.

  16. Diminishing returns from increased percent Bt cotton: the case of pink bollworm.

    Science.gov (United States)

    Huang, Yunxin; Wan, Peng; Zhang, Huannan; Huang, Minsong; Li, Zhaohua; Gould, Fred

    2013-01-01

    Regional suppression of pests by transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) has been reported in several cropping systems, but little is known about the functional relationship between the ultimate pest population density and the pervasiveness of Bt crops. Here we address this issue by analyzing 16 years of field data on pink bollworm (Pectinophora gossypiella) population density and percentage of Bt cotton in the Yangtze River Valley of China. In this region, the percentage of cotton hectares planted with Bt cotton increased from 9% in 2000 to 94% in 2009 and 2010. We find that as the percent Bt cotton increased over the years, the cross-year growth rate of pink bollworm from the last generation of one year to the first generation of the next year decreased. However, as the percent Bt cotton increased, the within-year growth rate of pink bollworm from the first to last generation of the same year increased, with a slope approximately opposite to that of the cross-year rates. As a result, we did not find a statistically significant decline in the annual growth rate of pink bollworm as the percent Bt cotton increased over time. Consistent with the data, our modeling analyses predict that the regional average density of pink bollworm declines as the percent Bt cotton increases, but the higher the percent Bt cotton, the slower the decline in pest density. Specifically, we find that 95% Bt cotton is predicted to cause only 3% more reduction in larval density than 80% Bt cotton. The results here suggest that density dependence can act against the decline in pest density and diminish the net effects of Bt cotton on suppression of pink bollworm in the study region. The findings call for more studies of the interactions between pest density-dependence and Bt crops.

  17. Breeding Strategies for Delaying Insect Resistance to Bt Toxic Protein%延缓害虫对Bt毒蛋白耐受性的育种策略

    Institute of Scientific and Technical Information of China (English)

    陈军; 刘友全; 卢孟柱

    2003-01-01

    在分析害虫对转Bt 毒蛋白基因植株产生耐受性的特点、发生机制的基础上,重点探讨了为延缓害虫对Bt毒蛋白耐受性的育种策略:(1)培育双价或多价Bt毒蛋白抗虫品种;(2)寻找能使Bt毒蛋白基因在植物中高效表达的启动子;(3)使用组织特异型或诱导型启动子以减轻对害虫持续的选择压;(4)将Bt毒蛋白基因转入叶绿体基因组以解决原核与真核细胞对遗传密码子偏爱的不同;(5)通过突变等技术改变Bt毒蛋白一级结构,从分子水平延缓害虫产生耐受性.

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

    Science.gov (United States)

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

    2011-06-01

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

  19. Modeling evolution of resistance of sugarcane borer (Lepidoptera: Crambidae) to transgenic Bt corn.

    Science.gov (United States)

    Kang, J; Huang, F; Onstad, D W

    2014-08-01

    Diatraea saccharalis (F.) (Lepidoptera: Crambidae) is a target pest of transgenic corn expressing Bacillus thuringiensis (Bt) protein, and the first evidence of resistance by D. saccharalis to Cry1Ab corn was detected in a field population in northeast Louisiana in 2004. We used a model of population dynamics and genetics of D. saccharalis to 1) study the effect of interfield dispersal, the first date that larvae enter diapause for overwintering, toxin mortality, the proportion of non-Bt corn in the corn patch, and the area of a crop patch on Bt resistance evolution; and 2) to identify gaps in empirical knowledge for managing D. saccharalis resistance to Bt corn. Increasing, the proportion of corn refuge did not always improve the durability of Bt corn if the landscape also contained sugarcane, sorghum, or rice. In the landscape, which consisted of 90% corn area, 5% sorghum area, and 5% rice area, the durability of single-protein Bt corn was 40 yr when the proportion of corn refuge was 0.2 but 16 yr when the proportion of corn refuge was 0.5. The Bt resistance evolution was sensitive to a change (from Julian date 260 to 272) in the first date larvae enter diapause for overwintering and moth movement. In the landscapes with Bt corn, non-Bt corn, sugarcane, sorghum, and rice, the evolution of Bt resistance accelerated when larvae entered diapause for overwintering early. Intermediate rates of moth movement delayed evolution of resistance more than either extremely low or high rates. This study suggested that heterogeneity in the agrolandscapes may complicate the strategy for managing Bt resistance in D. saccharalis, and designing a Bt resistance management strategy for D. saccharalis is challenging because of a lack of empirical data about overwintering and moth movement.

  20. Comparing gene expression profiles between Bt and non-Bt rice in response to brown planthopper infestation

    Directory of Open Access Journals (Sweden)

    Fang eWang

    2015-12-01

    Full Text Available Bt proteins are the most widely used insecticidal proteins in transgenic crops for improving insect resistance. We previously observed longer nymphal developmental duration and lower fecundity in brown planthopper (BPH fed on Bt rice line KMD2, although Bt insecticidal protein Cry1Ab could rarely concentrate in this non-target rice pest. In the present study, we performed microarray analysis in an effort to detect Bt-independent variation, which might render Bt rice more defensive and/or less nutritious to BPH. We detected 3,834 and 3,273 differentially expressed probe-sets in response to BPH infestation in non-Bt parent Xiushui 11 and Bt rice KMD2, respectively, only 439 of which showed significant differences in expression between rice lines. Our analysis revealed a shift from growth to defense responses in response to BPH infestation, which was also detected in many other studies of plants suffering biotic and abiotic stresses. Chlorophyll biosynthesis and basic metabolism pathways were inhibited in response to infestation. IAA and GA levels decreased as a result of the repression of biosynthesis-related genes or the induction of inactivation-related genes. In accordance with these observations, a number of IAA-, GA-, BR-signaling genes were downregulated in response to BPH. Thus, the growth of rice plants under BPH attack was reduced and defense related hormone signaling like JA, SA and ET were activated. In addition, growth-related hormone signaling pathways, such as GA, BR and auxin signaling pathways, as well as ABA, were also found to be involved in BPH-induced defense. On the other side, 51 probe-sets (represented 50 genes that most likely contribute to the impact of Bt rice on BPH were identified, including three early nodulin genes, four lipid metabolic genes, 14 stress response genes, three TF genes and genes with other functions. Two transcription factor genes, bHLH and MYB, together with lipid transfer protein genes LTPL65 and

  1. Nye oplysninger om bl.a. miljømæssig risikovurdering af PAT-protein og glufosinatammonium herbicider. Zea mays (Bt11). Supplerende materiale om Bt11. Modtaget 21-02-2005, deadline 29-03-2005, svar 07-03-2005

    DEFF Research Database (Denmark)

    Strandberg, Morten Tune

    2005-01-01

    Mail: Det tilsendte supplerende materiale om Bt-11 majsen (mail pr. 21-02-2005) omhandler bl.a. eventuelle uønskede konsekvenser ved ulovlig anvendelse af glufosinat ammonium herbicider i forbindelse med dyrkningen. Vi har allerede indraget dette aspekt i forbindelse med den økologiske risikovurd...

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

    Science.gov (United States)

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

    2014-01-01

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

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

    Science.gov (United States)

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

    2014-01-01

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

  4. Pest trade-offs in technology: reduced damage by caterpillars in Bt cotton benefits aphids.

    Science.gov (United States)

    Hagenbucher, Steffen; Wäckers, Felix L; Wettstein, Felix E; Olson, Dawn M; Ruberson, John R; Romeis, Jörg

    2013-05-07

    The rapid adoption of genetically engineered (GE) plants that express insecticidal Cry proteins derived from Bacillus thuringiensis (Bt) has raised concerns about their potential impact on non-target organisms. This includes the possibility that non-target herbivores develop into pests. Although studies have now reported increased populations of non-target herbivores in Bt cotton, the underlying mechanisms are not fully understood. We propose that lack of herbivore-induced secondary metabolites in Bt cotton represents a mechanism that benefits non-target herbivores. We show that, because of effective suppression of Bt-sensitive lepidopteran herbivores, Bt cotton contains reduced levels of induced terpenoids. We also show that changes in the overall level of these defensive secondary metabolites are associated with improved performance of a Bt-insensitive herbivore, the cotton aphid, under glasshouse conditions. These effects, however, were not as clearly evident under field conditions as aphid populations were not correlated with the amount of terpenoids measured in the plants. Nevertheless, increased aphid numbers were visible in Bt cotton compared with non-Bt cotton on some sampling dates. Identification of this mechanism increases our understanding of how insect-resistant crops impact herbivore communities and helps underpin the sustainable use of GE varieties.

  5. Insecticidal Mechanism of Bt Vegetative Insecticidal Proteins and their Application in GM Crops%新型Bt杀虫蛋白:VIP杀虫的机理与植物转基因应用

    Institute of Scientific and Technical Information of China (English)

    刘辰; 谢柳; 张文飞

    2008-01-01

    苏云金芽孢杆菌(Bacillus thuringiensis,简称 Bt)是目前应用最多的生物杀虫剂.它能够产生多种杀虫因子,其中最主要是杀虫晶体蛋白(insecticidal crystal proteins.ICP)和营养期杀虫蛋(vegetative insecticidal protein,VIP).VIP在Bt营养期生长阶段开始分泌,它们不形成蛋白晶体,和ICP在进化上没有同源性.VIP的杀虫活性很高,达到了纳克级水平,为一类新型杀虫蛋白,被认为是第二代生物杀虫剂.VIP根据蛋白质序列同源性主要分为三类:VIP1、VIP2和VIP3.VIP1和VIP2共同构成二元毒素对鞘翅目萤叶甲科昆虫具有特异性毒杀作用,VIP3对鳞翅目害虫具有很高的杀虫活性.由于VIP对一些可能对ICP产生抗性的害虫具有高效的毒杀作用,而且害虫对ICP和VIP产生交互抗性的几率也很小,因此转VIP植物研究受到了科学家的青睐,并已有不少成功的报道和专利.目前为止,已经利用V1P3构建了转基因水稻、转基因玉米、转基因棉花等多种转基冈作物.

  6. Regeneration of sugarcane elite breeding lines and engineering of stem borer resistance.

    Science.gov (United States)

    Weng, Li-Xing; Deng, Haihua; Xu, Jin-Ling; Li, Qi; Wang, Lian-Hui; Jiang, Zide; Zhang, Hai Bao; Li, Qiwei; Zhang, Lian-Hui

    2006-02-01

    Five elite sugarcane breeding lines were tested for efficiency in embryogenesis and plant regeneration. All of them produced regenerative embryogenic calli but with varied efficiencies. To engineer strongly insect-resistant sugarcanes, the GC content of a truncated cry1Ac gene, which encodes the active region of Cry1Ac insecticidal delta-endotoxin, was increased from the original 37.4 to 47.5% following the sugarcane codon usage pattern. The synthetic cry1Ac gene (s-cry1Ac) was placed under the control of maize ubiquitin promoter and introduced by microprojectile bombardment into the embryogenic calli of sugarcane lines YT79-177 and ROC16. Southern blotting analysis showed that multicopies of s-cry1Ac were integrated into the genomes of transgenic sugarcane lines. Immunoblotting analysis identified 18 transgenic lines expressing detectable levels of s-Cry1Ac, which were estimated in the range of 1.8-10.0 ng mg(-1) total soluble proteins. Four transgenic and two parental lines were assayed for sugarcane stem borer resistance in leaf tissue feeding trials and greenhouse plant assays. The results showed that, while the untransformed control lines were severely damaged in both leaves and stems, the transgenic sugarcane lines expressing high levels of s-Cry1Ac proteins were highly resistant to sugarcane stem borer attack, resulting in complete mortality of the inoculated insects within 1 week after inoculation.

  7. The food and environmental safety of Bt crops.

    Science.gov (United States)

    Koch, Michael S; Ward, Jason M; Levine, Steven L; Baum, James A; Vicini, John L; Hammond, Bruce G

    2015-01-01

    Bacillus thuringiensis (Bt) microbial pesticides have a 50-year history of safety in agriculture. Cry proteins are among the active insecticidal ingredients in these pesticides, and genes coding for Cry proteins have been introduced into agricultural crops using modern biotechnology. The Cry gene sequences are often modified to enable effective expression in planta and several Cry proteins have been modified to increase biological activity against the target pest(s). Additionally, the domains of different but structurally conserved Cry proteins can be combined to produce chimeric proteins with enhanced insecticidal properties. Environmental studies are performed and include invertebrates, mammals, and avian species. Mammalian studies used to support the food and feed safety assessment are also used to support the wild mammal assessment. In addition to the NTO assessment, the environmental assessment includes a comparative assessment between the Bt crop and the appropriate conventional control that is genetically similar but lacks the introduced trait to address unintended effects. Specific phenotypic, agronomic, and ecological characteristics are measured in the Bt crop and the conventional control to evaluate whether the introduction of the insect resistance has resulted in any changes that might cause ecological harm in terms of altered weed characteristics, susceptibility to pests, or adverse environmental impact. Additionally, environmental interaction data are collected in field experiments for Bt crop to evaluate potential adverse effects. Further to the agronomic and phenotypic evaluation, potential movement of transgenes from a genetically modified crop plants into wild relatives is assessed for a new pest resistance gene in a new crop. This review summarizes the evidence for safety of crops containing Cry proteins for humans, livestock, and other non-target organisms.

  8. The Food and Environmental Safety of Bt Crops

    Directory of Open Access Journals (Sweden)

    Michael Stephen Koch

    2015-04-01

    Full Text Available Bt (Bacillus thuringiensis microbial pesticides have a 50-year history of safe use in agriculture. Cry proteins are among the active insecticidal ingredients in these pesticides, and genes coding for Cry proteins have been introduced into agricultural crops using modern biotechnology. The Cry gene sequences are often modified to enable effective expression in planta and several Cry proteins have been modified to increase biological activity against the target pest(s. Additionally, the domains of different but structurally conserved Cry proteins can be combined to produce chimeric proteins with enhanced insecticidal properties. Environmental studies are performed and include invertebrates, mammals and avian species. Mammalian studies used to support the food and feed safety assessment are also used to support the wild mammal assessment. In addition to the NTO assessment, the environmental assessment includes a comparative assessment between the Bt crop and the appropriate conventional control that is genetically similar but lacks the introduced trait to address unintended effects. Specific phenotypic, agronomic, and ecological characteristics are measured in the Bt crop and the conventional control to evaluate whether the introduction of the insect resistance has resulted in any changes that might cause ecological harm in terms of altered weed characteristics, susceptibility to pests, or adverse environmental impact. Additionally, environmental interaction data are collected in field experiments for Bt crop to evaluate potential adverse effects. Further to the agronomic and phenotypic evaluation, potential movement of transgenes from a genetically modified crop plants into wild relatives is assessed for a new pest resistance gene in a new crop. This review summarizes the evidence for safety of crops containing Cry proteins for humans, livestock, and other non-target organisms.

  9. Molecular and biochemical analysis of the plastidic ADP-glucose transporter (ZmBT1) from Zea mays.

    NARCIS (Netherlands)

    Kirchberger, S.; Leroch, M.; Huynen, M.A.; Wahl, M.; Neuhaus, H.E.; Tjaden, J.

    2007-01-01

    Physiological studies on the Brittle1 maize mutant have provided circumstantial evidence that ZmBT1 (Zea mays Brittle1 protein) is involved in the ADP-Glc transport into maize endosperm plastids, but up to now, no direct ADP-Glc transport mediated by ZmBT1 has ever been shown. The heterologous synth

  10. Molecular and biochemical analysis of the plastidic ADP-glucose transporter (ZmBT1) from Zea mays

    NARCIS (Netherlands)

    Kirchberger, S.; Leroch, M.; Huynen, M.A.; Wahl, M.; Neuhaus, H.E.; Tjaden, J.

    2007-01-01

    Physiological studies on the Brittle1 maize mutant have provided circumstantial evidence that ZmBT1 (Zea mays Brittle1 protein) is involved in the ADP-Glc transport into maize endosperm plastids, but up to now, no direct ADP-Glc transport mediated by ZmBT1 has ever been shown. The heterologous synth

  11. Preparative Purification and Bioassay of Bt Toxin from Cry1Ab Transgenic Rice

    Institute of Scientific and Technical Information of China (English)

    WU Jian-min; YE Qin-fu

    2004-01-01

    A method of extracting and purifying Cry1Ab protein(Bt toxin) from Cry1Ab transgenic rice was established. Most of the Bt toxin present in the tissue of Cry1Ab transgenic rice was extracted effectively with a solution of 50 mmol/LNa2CO3 and NaHCO3. The crude protein containing Bt toxin was obtained after the pretreatment of Cry1Ab transgenic rice with ultra-filtration, ammonium sulfate precipitation and centrifugation. The dialysed crude protein was futher separated on DEAE Sephadex A-50 columns and Sephadex G-150 columns. The protein bound on DEAE Sephadex A-50 gel was eluted with buffer solution B(10 mmol/L trisHCl buffer+1. 0 mmol/L EDTA, pH=8.0) mixed with 0. 1, 0. 3, 0. 5 and 0. 8 mol/L NaCl in a discontinuous gradient elution mode. The peak of the Bt toxin eluted from the columns was identified by ELISA and bioassayed with larvae of tobacco hornworms and silkworms. The purity and the bioactivity of the Bt toxin were determined by means of SDS-PAGE and larvicidal assay, respectively. The purity of the Bt toxin obtained by this method is high, and its insecticidal activity is retained after the toxin is purified.

  12. Characterization of NCAM expression and function in BT4C and BT4Cn glioma cells

    DEFF Research Database (Denmark)

    1991-01-01

    The neural cell adhesion molecule, NCAM, plays an important role in cell-cell adhesion. Therefore, we have studied NCAM expression in the glioma cell lines BT4C and BT4Cn. We demonstrate that the 2 cell lines differ in their metastatic ability; while BT4C cells have a very low capacity...... for producing experimental metastases, that of BT4Cn cells is high. In BT4C cells NCAM is synthesized as 4 polypeptides with Mr's of 190,000, 140,000, 115,000 and 97,000. The 140,000, 115,000 and 97,000 polypeptides are glycosylated and for the 140,000 and 115,000 polypeptides sulfatation is observed......-substratum binding assay in which the binding of BT4C and BT4Cn cells to NCAM immobilized to glass was assessed. We found that BT4C cells adhere specifically to NCAM, and that adhesion is inhibited by anti-NCAM Fab'-fragments, while no specific binding of BT4Cn cells to NCAM was observed. The BT4C and BT4Cn cell...

  13. Bacillus thuringiensis (Bt) toxin susceptibility and isolation of resistance mutants in the nematode Caenorhabditis elegans.

    OpenAIRE

    2000-01-01

    The protein toxins produced by Bacillus thuringiensis (Bt) are the most widely used natural insecticides in agriculture. Despite successful and extensive use of these toxins in transgenic crops, little is known about toxicity and resistance pathways in target insects since these organisms are not ideal for molecular genetic studies. To address this limitation and to investigate the potential use of these toxins to control parasitic nematodes, we are studying Bt toxin action and resistance in ...

  14. Pest trade-offs in technology: reduced damage by caterpillars in Bt cotton benefits aphids

    OpenAIRE

    2013-01-01

    The rapid adoption of genetically engineered (GE) plants that express insecticidal Cry proteins derived from Bacillus thuringiensis (Bt) has raised concerns about their potential impact on non-target organisms. This includes the possibility that non-target herbivores develop into pests. Although studies have now reported increased populations of non-target herbivores in Bt cotton, the underlying mechanisms are not fully understood. We propose that lack of herbivore-induced secondary metabolit...

  15. Dependence of dielectric properties on BT particle size in EP/BT composites

    Institute of Scientific and Technical Information of China (English)

    YANG Xiaojun; YANG Zhimin; MAO Changhui; DU Jun

    2006-01-01

    The polymer-ceramic composites of epoxy resin (EP) and barium titanate (BT) were prepared.BT powders of different BT particle sizes from 100 nm to 1 μm were used in the preparation.The dielectric properties, such as dielectric constant, dielectric loss and electrical breakdown strength, of the EP/BT composites were studied.The morphology of the composites was characterized by the means of scanning electron microscopy (SEM).The results show that the dielectric constant of the composites is much higher than the epoxy matrix at frequency range from 1 kHz to 10 MHz, and it is also obviously dependent on the size of BT particles.The electrical breakdown strength of the composites decreases with the increase of the BT content.The dependence of electrical breakdown strength on BT particle sizes was also discussed.

  16. Overexpression of a Foreign Bt Gene in Cotton Affects the Low-Molecular-Weight Components in Root Exudates

    Institute of Scientific and Technical Information of China (English)

    YAN Wei-Dong; SHI Wei-Ming; LI Bao-Hai; ZHANG Min

    2007-01-01

    Most research in the past using genetically modified crops (GM crops) has focused on the ecological safety of foreign gene (i.e., the gene flow), gene products (for example, Bt (Bacillus thuringiensis) protein), and the safety of transgenic food for humans. In this study, changes in both the species and amounts of low-molecular-weight components in cotton (Gossypium hirsutum L.) root exudates after foreign Bt gene overexpression were investigated under different nutritional conditions. Transgenic cotton containing Bt (Bt-cotton), supplemented with all the mineral nutrients, secreted more organic acids than the wild-type cotton (WT). When nitrogen was removed from the full-nutrient solution, the amount of organic acids secretion of Bt-cotton was lesser than that of WT. The roots of the transgenic cotton secreted lesser amounts of amino acids and soluble sugars than the WT roots in the full-nutrient solution. Deficiencies of P and K caused a large increase in the total amino acid and soluble sugar secretions of both Bt-cotton and WT, with larger increases observed in Bt-cotton. Because transferring the foreign Bt gene into cotton can result in alterations in the components of the root exudates, with the effect varying depending on the nutritional status, the cultivation of genetically modified crops, such as Bt-cotton, in soil environments should be more carefully assessed, and the possible effects as a result of the alterations in the root exudate components should be considered.

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

    Science.gov (United States)

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

  18. Effects of Bt-transgenic rice cultivation on planktonic communities in paddy fields and adjacent ditches

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yongbo, E-mail: liuyb@craes.org.cn [State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012 (China); Liu, Fang [State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012 (China); Wang, Chao [Pearl River Fisheries Research Institute, Chinese Academy of Fishery Science, Guangzhou 510380 (China); Quan, Zhanjun [State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012 (China); Li, Junsheng, E-mail: lijsh@creas.org.cn [State Key Laboratory of Environmental Criteria and Risk Assessment, Chinese Research Academy of Environmental Sciences, Beijing 100012 (China)

    2016-09-15

    The non-target effects of transgenic plants are issues of concern; however, their impacts in cultivated agricultural fields and adjacent natural aquatic ecosystems are poorly understood. We conducted field experiments during two growing seasons to determine the effects of cultivating Bacillus thuringiensis (Bt)-transgenic rice on the phytoplankton and zooplankton communities in a paddy field and an adjacent ditch. Bt toxin was detected in soil but not in water. Water quality was not significantly different between non-Bt and Bt rice fields, but varied among up-, mid- and downstream locations in the ditch. Cultivation of Bt-transgenic rice had no effects on zooplankton communities. Phytoplankton abundance and biodiversity were not significantly different between transgenic and non-transgenic rice fields in 2013; however, phytoplankton were more abundant in the transgenic rice field than in the non-transgenic rice field in 2014. Water quality and rice type explained 65.9% and 12.8% of this difference in 2014, respectively. Phytoplankton and zooplankton were more abundant in mid- and downstream, than upstream, locations in the ditch, an effect that we attribute to water quality differences. Thus, the release of Bt toxins into field water during the cultivation of transgenic crops had no direct negative effects on plankton community composition, but indirect effects that alter environmental conditions should be taken into account during the processes of management planning and policymaking. - Highlights: • We detect fusion Cry1Ab/1Ac proteins from Bt rice entering into aquatic ecosystems. • Bt-transgenic rice cultivation have no significant effect on zooplankton community. • Bt-transgenic rice cultivation have indirect effect on phytoplankton community. • Water quality explains the difference of plankton communities in adjacent ditches.

  19. Pest management through Bacillus thuringiensis (Bt) in a tea-silkworm ecosystem: status and potential prospects.

    Science.gov (United States)

    Dashora, Kavya; Roy, Somnath; Nagpal, Akanksha; Roy, Sudipta Mukhopadhyay; Flood, Julie; Prasad, Anjali Km; Khetarpal, Ravinder; Neave, Suzanne; Muraleedharan, N

    2017-03-01

    Bacillus thuringiensis (Bt) is a soil bacterium that forms spores containing crystals comprising one or more Cry or Cyt proteins having potential and specific insecticidal activity. Different strains of Bt produce different types of toxins, affecting a narrow taxonomic group of insects. Therefore, it is used in non-chemical pest management, including inherent pest resistance through GM crops. The specificity of action of Bt toxins reduces the concern of adverse effects on non-target species, a concern which remains with chemical insecticides as well. To make use of Bt more sustainable, new strains expressing novel toxins are actively being sought globally. Since Bt is successfully used against many pests including the lepidopteran pests in different crop groups, the insecticidal activity against Samia cynthia (Drury) (Eri silkworm) and Antheraea assamensis Helfer (Muga silkworm) becomes a concern in the state of Assam in India which is a predominantly tea- and silk-producing zone. Though Bt can be used as an effective non-chemical approach for pest management for tea pests in the same geographical region, yet, it may potentially affect the silk industry which depends on silkworm. There is a need to identify the potentially lethal impact (through evaluating their mortality potential) of local Bt strains on key silkworm species in North Eastern India. This will allow the use of existing Bt for which the silkworms have natural resistance. Through this review, the authors aim to highlight recent progress in the use of Bt and its insecticidal toxins in tea pest control and the potential sensitivity for tea- and silk-producing zone of Assam in India.

  20. 75 FR 19388 - Pesticide Product; Registration Application

    Science.gov (United States)

    2010-04-14

    ... Classification System (NAICS) codes have been provided to assist you and others in determining whether this... protectant, Bacillus thuringiensis Cry1Ac Protein and the Genetic Material ] (Vector PV-GMIR9) Necessary...

  1. Development and Validation of a Fluorescent Multiplexed Immunoassay for Measurement of Transgenic Proteins in Cotton (Gossypium hirsutum).

    Science.gov (United States)

    Yeaman, Grant R; Paul, Sudakshina; Nahirna, Iryna; Wang, Yongcheng; Deffenbaugh, Andrew E; Liu, Zi Lucy; Glenn, Kevin C

    2016-06-22

    In order to provide farmers with better and more customized alternatives to improve yields, combining multiple genetically modified (GM) traits into a single product (called stacked trait crops) is becoming prevalent. Trait protein expression levels are used to characterize new GM products and establish exposure limits, two important components of safety assessment. Developing a multiplexed immunoassay capable of measuring all trait proteins in the same sample allows for higher sample throughput and savings in both time and expense. Fluorescent (bead-based) multiplexed immunoassays (FMI) have gained wide acceptance in mammalian research and in clinical applications. In order to facilitate the measurement of stacked GM traits, we have developed and validated an FMI assay that can measure five different proteins (β-glucuronidase, neomycin phosphotransferase II, Cry1Ac, Cry2Ab2, and CP4 5-enolpyruvyl-shikimate-3-phosphate synthase) present in cotton leaf from a stacked trait product. Expression levels of the five proteins determined by FMI in cotton leaf tissues have been evaluated relative to expression levels determined by enzyme-linked immunosorbent assays (ELISAs) of the individual proteins and shown to be comparable. The FMI met characterization requirements similar to those used for ELISA. Therefore, it is reasonable to conclude that FMI results are equivalent to those determined by conventional individual ELISAs to measure GM protein expression levels in stacked trait products but with significantly higher throughput, reduced time, and more efficient use of resources.

  2. 广西兴安转Bt水稻大田两迁害虫发生动态%Occurrence dynamics of migratory pest insects Cnaphalocrocis medinalis and Sogatella furcifera in transgenic Bt rice field in Xing' an County of Guangxi Province

    Institute of Scientific and Technical Information of China (English)

    隋贺; 李志毅; 徐艳博; 韩超; 韩兰芝; 陈法军

    2011-01-01

    An investigation was conducted in Xing' an County of Guangxi Province in 2010 to study the occurrence and damage characteristics of two unique migratory pest insects, rice leaffolder ( Cnaphalocrocis medinalis) and white-backed planthopper (Sogatella furcifera) , in a transgenic Bt rice (cv. HH1 with dual CrylAb + CrylAc genes) field, taking the corresponding non-transgenic parent (cv. MH63) field as the control. No significant differences were observed in the abundance of C. Medinalis eggs and larvae in the two fields, but the percentage of fold-leaf plants and the fold-leaf rate per plant were significantly lower in transgenic Bt rice field than in the control, suggesting that transgenic Bt rice had higher resistance against the target pest insect C. Medinalis. As for 5. Furcifera, its occurrence dynamics of nymphs, adults (including macro- and brachypterous forms) , and whole population had no significant differences between the two fields, but the abundance of the nymphs and brachypterous adults at the peak stage of S. Furcifera occurrence was obviously higher in transgenic Bt rice field than in the control, while the macropterous abundance was in adverse. The sex ratio of female of the macropterous adults at the late growth stage of rice was generally lower in transgenic Bt rice field than in the control. These results suggested that under the background of large area commercial production of transgenic Bt rice, the occurrence and harm of the non-target pest insect S. Furcifera could become more complicated.%以转Bt水稻华恢1号(Cry1Ac/ CryAb融合基因型,简称HH1)及其对照亲本明恢63(简称MH63)稻田两迁害虫稻纵卷叶螟和白背飞虱为研究对象,系统研究转基因抗虫水稻种植下两迁害虫的发生规律及其致害力差异.结果表明:转Bt水稻及其对照亲本上稻纵卷叶螟的落卵量和幼虫发生量无显著差异,但转Bt水稻的卷叶株率和卷叶率都显著低于对照亲本.表明转Bt水稻对靶标

  3. Effects of transgenic Bt rice on growth, reproduction, and superoxide dismutase activity of Folsomia candida (Collembola: Isotomidae) in laboratory studies.

    Science.gov (United States)

    Bai, Yaoyu; Yan, Ruihong; Ke, Xin; Ye, Gongyin; Huang, Fangneng; Luo, Yongming; Cheng, Jiaan

    2011-12-01

    Transgenic rice expressing Bacillus thuringiensis (Bt) CrylAb protein is expected to be commercialized in China in the near future. The use of Bt rice for controlling insect pests sparks intensive debates regarding its biosafety. Folsomia candida is an euedaphic species and is often used as a "standard" test organism in assessing effects of environmental pollutants on soil organisms. In this study, growth, development, reproduction, and superoxide dismutase activity (SOD) of F. candida were investigated in the laboratory for populations reared on leaf tissue or leaf-soil mixtures of two CrylAb rice lines and a non-Bt rice isoline. Two independent tests were performed: 1) a 35-d test using petri dishes containing yeast diet (positive control) or fresh rice leaf tissue, and 2) a 28-d test in soil-litter microcosms containing yeast or a mixture of soil and rice leaf tissue. Biological parameters measured in both tests were number of progeny production, population growth rate, and SOD activity. For the petri dish test, data measured also included insect body length and number of exuviation. There were no significant differences between the populations reared on Bt and non-Bt rice leaf tissue in all measured parameters in both tests and for both Bt rice lines, suggesting no significant effects of the CrylAb protein in Bt rice on F. candida in the laboratory studies. Results of this study should add additional biosafety proofs for use of Bt rice to manage rice pests in China.

  4. Response of the Gypsy Moth, Lymantria dispar to Transgenic Poplar, Populus simonii x P. nigra, Expressing Fusion Protein Gene of the Spider Insecticidal Peptide and Bt-toxin C-peptide

    OpenAIRE

    Cao, Chuan-Wang; Liu, Gui-Feng; Wang, Zhi-Ying; Yan, Shan-Chun; Ma, Ling; Yang, Chuan-Ping

    2010-01-01

    The response of the Asian gypsy moth Lymantria dispar (L.) (Lepidoptera: Lymantriidae) to a fusion gene consisting of the spider, Atrax robustus Simon (Araneae: Hexanthelidae) ω?-ACTX-Ar1 sequence coding for an ω?-atracotoxin and a sequence coding for the Bt-toxin C-peptide, expressed in transgenic poplar Populus simonii x P. nigra L. (Malphigiales: Salicaceae) was investigated. Individual performance, feeding selection, midgut proteinase activity and nutrition utilization were monitored. The...

  5. Development of stem borer resistant transgenic parental lines involved in the production of hybrid rice.

    Science.gov (United States)

    Ramesh, S; Nagadhara, D; Pasalu, I C; Kumari, A Padma; Sarma, N P; Reddy, V D; Rao, K V

    2004-07-15

    Stem borer resistant transgenic parental lines, involved in hybrid rice, were produced by Agrobacterium-mediated gene transfer method. Two pSB111 super-binary vectors containing modified cry1Ab/cry1Ac genes driven by maize ubiquitin promoter, and herbicide resistance gene bar driven by cauliflower mosaic virus 35S promoter were, used in this study. Embryogenic calli after co-cultivation with Agrobacterium were selected on the medium containing phosphinothricin. Southern blot analyses of primary transformants revealed the stable integration of bar, cry1Ab and cry1Ac coding sequences into the genomes of three parental lines with a predominant single copy integration and without any rearrangement of T-DNA. T1 progeny plants disclosed a monogenic pattern (3:1) of transgene segregation as confirmed by molecular analyses. Furthermore, the co-segregation of bar and cry genes in T1 progenies suggested that the transgenes are integrated at a single site in the rice genome. In different primary transformants with alien inbuilt resistance, the levels of cry proteins varied between 0.03 and 0.13% of total soluble proteins. These transgenic lines expressing insecticidal proteins afforded substantial resistance against stem borers. This is the first report of its kind dealing with the introduction of Bacillus thuringiensis (Bt) cry genes into the elite parental lines involved in the development of hybrid rice.

  6. Delaying corn rootworm resistance to Bt corn.

    Science.gov (United States)

    Tabashnik, Bruce E; Gould, Fred

    2012-06-01

    Transgenic crops producing Bacillus thuringiensis (Bt) toxins for insect control have been successful, but their efficacy is reduced when pests evolve resistance. To delay pest resistance to Bt crops, the U.S. Environmental Protection Agency (EPA) has required refuges of host plants that do not produce Bt toxins to promote survival of susceptible pests. Such refuges are expected to be most effective if the Bt plants deliver a dose of toxin high enough to kill nearly all hybrid progeny produced by matings between resistant and susceptible pests. In 2003, the EPA first registered corn, Zea mays L., producing a Bt toxin (Cry3Bb1) that kills western corn rootworm, Diabrotica virgifera virgifera LeConte, one of the most economically important crop pests in the United States. The EPA requires minimum refuges of 20% for Cry3Bb1 corn and 5% for corn producing two Bt toxins active against corn rootworms. We conclude that the current refuge requirements are not adequate, because Bt corn hybrids active against corn rootworms do not meet the high-dose standard, and western corn rootworm has rapidly evolved resistance to Cry3Bb1 corn in the laboratory, greenhouse, and field. Accordingly, we recommend increasing the minimum refuge for Bt corn targeting corn rootworms to 50% for plants producing one toxin active against these pests and to 20% for plants producing two toxins active against these pests. Increasing the minimum refuge percentage can help to delay pest resistance, encourage integrated pest management, and promote more sustainable crop protection.

  7. 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.

  8. Binding of Bt Cry toxins to lepidopteran midgut aminopeptidase N and the relationship between their interactions with Bt resistance%鳞翅目昆虫氨肽酶N与Bt毒素的结合及其与Bt抗性的关系

    Institute of Scientific and Technical Information of China (English)

    马文静; 韩兰芝; 尹新明; 曹广春; 苏丽娟

    2011-01-01

    随着Bt Cry作物在我国的广泛应用和推广,靶标害虫对其抗性风险已成为Bt Cry作物生态安全研究的重要内容.氨肽酶N(Aminopeptidase N,APN)是位于昆虫中肠刷状缘膜囊泡(Brush Border Membrane Vesicles,BBMV)上Bt Cry毒素重要的受体蛋白之一,它与Bt Cry毒素的结合能力决定了Bt Cry毒素的杀虫活性及昆虫对Bt抗性的产生.本文从APN的结构特征与分类、APN与Bt Cry毒素的结合特异性、结合位点、结合过程中的分子互作机制及APN变异导致昆虫抗性产生几方面系统综述了鳞翅目昆虫中肠Bt Cry受体蛋白-氨肽酶N与Bt Cry 毒素的结合及其与Bt抗性关系的研究进展.%With the wide application and popularization of Bt Cry crop, the resistance of target pests a-gainst Bt Cry crop has become the focus of studies on its ecological safety. Aminopeptidase N (APN) is one of Bt-toxin receptor proteins located in brush border membrane vesicles ( BBMV) of insect midgut. Its binding capability with Bt Cry toxin determines the toxin's insecticidal spectrum and insect Bt resistance. This paper systemically reviews the progress of researches on APN structural characteristics and classfication, the binding feature, binding sites and interactions mechanism of Bt Cry toxin and APN as well as Bt resistance resulting from APN mutations.

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

    Science.gov (United States)

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

    2015-06-01

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

  10. Selection of Homozygous Cotton Lines Transformed with Two Insect-Resistant Genes

    Institute of Scientific and Technical Information of China (English)

    WU Jia-he; TIAN Ying-chuan; LUO Xiao-li; GUO Hong-nian; SHI Yue-jin; CHEN Xiao-ying; JIA Yan-tao; XIAO Juan-li; ZHANG Xian-long

    2003-01-01

    A plant expression vector containing a chimeric Bt29K gene coding for the activated Cry1Ac protein and the arrowhead proteinase inhibitior gene API-B were introduced into the cotton cultivar Jihe321 mediated by Agrobactertium tumefaciens. Based on the results of kanamycin resistant testing, PCR detection for both foreign genes and insect bioassay using Heliethis armigera, nine transgenic homozygous cotton lines with insect-resistance of more than 90% and better agronomic traits were bred through six generations from the original transgenic plants. Results from insect bioassay and sequence analysis of the PCR products of plants from some homozygous lines indicated that the chimeric Bt29K gene was stably inherited in these transgenic cotton lines. The main agronomic characters of these homozygous cotton lines, such as boll productivity and fibre strength, were better than that of the original cotton cv. Jihe321.

  11. Transgenic Bt Cotton Does Not Disrupt the Top-Down Forces Regulating the Cotton Aphid in Central China

    Science.gov (United States)

    Yao, Yong-Sheng; Han, Peng; Niu, Chang-Ying; Dong, Yong-Cheng; Gao, Xi-Wu; Cui, Jin-Jie; Desneux, Nicolas

    2016-01-01

    Top-down force is referred to arthropod pest management delivered by the organisms from higher trophic levels. In the context of prevalent adoption of transgenic Bt crops that produce insecticidal Cry proteins derived from Bacillus thuringiensis (Bt), it still remains elusive whether the top-down forces are affected by the insect-resistant traits that introduced into the Bt crops. We explored how Bt cotton affect the strength of top-down forces via arthropod natural enemies in regulating a non-target pest species, the cotton aphid Aphis gossypii Glover, using a comparative approach (i.e. Bt cotton vs. conventional cotton) under field conditions. To determine top-down forces, we manipulated predation/parasitism exposure of the aphid to their natural enemies using exclusion cages. We found that the aphid population growth was strongly suppressed by the dominant natural enemies including Coccinellids, spiders and Aphidiines parasitoids. Coccinellids, spiders and the assemblage of other arthropod natural enemies (mainly lacewings and Hemipteran bugs) are similarly abundant in both plots, but with the parasitoid mummies less abundant in Bt cotton plots compared to the conventional cotton plots. However, the lower abundance of parasitoids in Bt cotton plots alone did not translate into differential top-down control on A. gossypii populations compared to conventional ones. Overall, the top-down forces were equally strong in both plots. We conclude that transgenic Bt cotton does not disrupt the top-down forces regulating the cotton aphid in central China. PMID:27870914

  12. High susceptibility of Bt maize to aphids enhances the performance of parasitoids of lepidopteran pests.

    Directory of Open Access Journals (Sweden)

    Cristina A Faria

    Full Text Available Concerns about possible undesired environmental effects of transgenic crops have prompted numerous evaluations of such crops. So-called Bt crops receive particular attention because they carry bacteria-derived genes coding for insecticidal proteins that might negatively affect non-target arthropods. Here we show a remarkable positive effect of Bt maize on the performance of the corn leaf aphid Rhopalosiphum maidis, which in turn enhanced the performance of parasitic wasps that feed on aphid honeydew. Within five out of six pairs that were evaluated, transgenic maize lines were significantly more susceptible to aphids than their near-isogenic equivalents, with the remaining pair being equally susceptible. The aphids feed from the phloem sieve element content and analyses of this sap in selected maize lines revealed marginally, but significantly higher amino acid levels in Bt maize, which might partially explain the observed increased aphid performance. Larger colony densities of aphids on Bt plants resulted in an increased production of honeydew that can be used as food by beneficial insects. Indeed, Cotesia marginiventris, a parasitoid of lepidopteran pests, lived longer and parasitized more pest caterpillars in the presence of aphid-infested Bt maize than in the presence of aphid-infested isogenic maize. Hence, depending on aphid pest thresholds, the observed increased susceptibility of Bt maize to aphids may be either a welcome or an undesirable side effect.

  13. Impacts of Bt crops on non-target organisms and insecticide use patterns

    Science.gov (United States)

    Bacillus thuringiensis (Bt), a bacterium capable of producing insecticidal proteins is ubiquitous in the environment, and the genes coding for these proteins are now becoming ubiquitous in major crop plants via recombinant DNA technology where they provide host plant resistance to major lepidopteran...

  14. 苏云金芽胞杆菌6618杀虫菌株的分离和鉴定%Isolation and Characterization of an Insecticidal Bacillus thuringiensis Strain 6618

    Institute of Scientific and Technical Information of China (English)

    张晨; 黄菲; 陆秀青; 谢俊雁; 丁学知; 夏立秋; 孙运军

    2015-01-01

    从我国不同地区采集118份土样,利用温度筛选法分离获得一株苏云金芽胞杆菌( Bacillus thuringien-sis,简称Bt)6618,镜检观察发现该菌株能产生典型的菱形晶体,PCR分析表明其含有cry1类杀虫基因。采用SDS-PAGE和质谱分析发现该菌株主要产生130 kD原毒素,其组分由Cry1Ae和Cry1Ac原毒素组成。基因序列分析表明该菌株的cry1 Ac基因为已知的 cry1 Ac1,而 cry1 Ae为新型杀虫基因。毒力生测表明该菌株对棉铃虫( Helicoverpa armigera)幼虫具有显著的杀虫效果。筛选获得的高毒力Bt菌株6618为丰富我国苏云金芽胞杆菌储备和研发新型高效生物杀虫剂提供了菌株资源。%118 soil samples were collected from different parts of our country and a new Bacillus thuringiensis strain 6618 was isolated by temperature screening method. This strain could produce typical bipyramidal crystal under micro-scope observation. PCR analysis showed that it contained cry1-type gene. SDS-PAGE analysis indicated that this strain expressed a major protein band about 130 kD, which was verified by mass spectrometry analysis to be the products of Cry1Ae and Cry1Ac protoxins. Sequence analysis showed that the cry1Ac gene in this strain was identical to the pub-lished cry1Ac1 gene, while the cry1Ae gene was found to be different from the known gene. The crystal-spore mixture of the strain 6618 exhibited high toxicity towards the larvae of Helicoverpa armigera. The isolated Bacillus thuringiensis strain 6618 with high toxicity will not only enrich the reserve of Bacillus thuringiensis strains of our country, but also pro-vide resource for the development of new efficient biological pesticide.

  15. A challenge for the seed mixture refuge strategy in Bt maize: impact of cross-pollination on an ear-feeding pest, corn earworm.

    Directory of Open Access Journals (Sweden)

    Fei Yang

    Full Text Available To counter the threat of insect resistance, Bacillus thuringiensis (Bt maize growers in the U.S. are required to plant structured non-Bt maize refuges. Concerns with refuge compliance led to the introduction of seed mixtures, also called RIB (refuge-in-the-bag, as an alternative approach for implementing refuge for Bt maize products in the U.S. Maize Belt. A major concern in RIB is cross-pollination of maize hybrids that can cause Bt proteins to be present in refuge maize kernels and negatively affect refuge insects. Here we show that a mixed planting of 5% nonBt and 95% Bt maize containing the SmartStax traits expressing Cry1A.105, Cry2Ab2 and Cry1F did not provide an effective refuge for an important above-ground ear-feeding pest, the corn earworm, Helicoverpa zea (Boddie. Cross-pollination in RIB caused a majority (>90% of refuge kernels to express ≥ one Bt protein. The contamination of Bt proteins in the refuge ears reduced neonate-to-adult survivorship of H. zea to only 4.6%, a reduction of 88.1% relative to larvae feeding on ears of pure non-Bt maize plantings. In addition, the limited survivors on refuge ears had lower pupal mass and took longer to develop to adults.

  16. A challenge for the seed mixture refuge strategy in Bt maize: impact of cross-pollination on an ear-feeding pest, corn earworm.

    Science.gov (United States)

    Yang, Fei; Kerns, David L; Head, Graham P; Leonard, B Rogers; Levy, Ronnie; Niu, Ying; Huang, Fangneng

    2014-01-01

    To counter the threat of insect resistance, Bacillus thuringiensis (Bt) maize growers in the U.S. are required to plant structured non-Bt maize refuges. Concerns with refuge compliance led to the introduction of seed mixtures, also called RIB (refuge-in-the-bag), as an alternative approach for implementing refuge for Bt maize products in the U.S. Maize Belt. A major concern in RIB is cross-pollination of maize hybrids that can cause Bt proteins to be present in refuge maize kernels and negatively affect refuge insects. Here we show that a mixed planting of 5% nonBt and 95% Bt maize containing the SmartStax traits expressing Cry1A.105, Cry2Ab2 and Cry1F did not provide an effective refuge for an important above-ground ear-feeding pest, the corn earworm, Helicoverpa zea (Boddie). Cross-pollination in RIB caused a majority (>90%) of refuge kernels to express ≥ one Bt protein. The contamination of Bt proteins in the refuge ears reduced neonate-to-adult survivorship of H. zea to only 4.6%, a reduction of 88.1% relative to larvae feeding on ears of pure non-Bt maize plantings. In addition, the limited survivors on refuge ears had lower pupal mass and took longer to develop to adults.

  17. Cannibalism of Helicoverpa zea (Lepidoptera: Noctuidae) from Bacillus thuringiensis (Bt) transgenic corn versus non-Bt corn.

    Science.gov (United States)

    Chilcutt, Charles F

    2006-06-01

    Because of the importance of cannibalism in population regulation of Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) in corn, Zea mays L., it is useful to understand the interactions between Bacillus thuringiensis (Bt) transgenic corn and cannibalism. To determine the effects of Bt corn on cannibalism in H. zea, pairs of the same or different instars were taken from Bt or non-Bt corn and placed on artificial diet in proximity. Cannibalism occurred in 91% of pairs and was approximately 7% greater for pairs of larvae reared from Bt transgenic corn (95%) than from non-Bt corn (88%). Also, first instar by first instar pairs had a lower rate of cannibalism than other pairs. Time until cannibalism was not different for larvae from Bt corn versus non-Bt corn. Pupation rate of cannibals and surviving victims was not different for pairs from Bt corn versus non-Bt corn. Finally, cannibalism increased pupation rate of cannibals from both Bt and non-Bt corn by approximately 23 and 12%, respectively, although the increases were not significant. Thus, negative effects of Bt on larvae were compensated by increased cannibalism in comparison with larvae reared on non-Bt corn, which increased larval survival to levels comparable with larvae reared on non-Bt plants.

  18. Tolerance of Bt corn (MON 810) to maize stem borer, Chilo partellus (Lepidoptera: Pyralidae).

    Science.gov (United States)

    Singh, Ramkumar; Channappa, Ravi K; Deeba, Farah; Nagaraj, Nandi J; Sukavaneaswaran, Mohan K; Manjunath, T M

    2005-11-01

    Transgenic corn (MON 810), expressing the Bacillus thuringiensis (Bt) protein, Cry1Ab, was evaluated under greenhouse conditions for its tolerance to the maize stem borer, Chilo partellus. Bt corn (MON 810) provided effective protection against the stem borer even under a high level of larval infestation in the greenhouse. The observed tolerance is examined and discussed in the light of the susceptibility of C. partellus to the Cry1Ab protein in laboratory bioassays. The implications of the tissue concentrations of Cry1Ab in MON 810, and baseline susceptibility recorded in the current study, for insect-resistance management are discussed.

  19. Domains of Bacillus thuringiensis crystal proteins involved in insecticidal activity

    NARCIS (Netherlands)

    Bosch, H.J.; Schipper, B.; Kleij, van der H.; Maagd, de R.A.; Stiekema, W.J.

    1994-01-01

    The expected increase in application of Bacillus thuringiensis (Bt) in crop protection makes it necessary to anticipate the development of Bt-resistant insects. To safeguard the long-term use of Bt-based insecticides, we studied the mode of action of Bt crystal proteins. CryIA(b), CryIC and CryIE ar

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

    OpenAIRE

    2016-01-01

    Transgenic cotton expressing the Cry 1Ab protein of Bacillus thuringiensis developing against Helocoverpa armigera may be affect on secondary pest such as Bemisia tabaci. In this study effects of Bt cotton on demographic parameters of B. tabaci were assessed and the data analyzed using the age specific, two-sex life table parameters. Results showed that getting to the adulthood stage, was faster on non-Bt cotton in comparison with Bt cotton. Also the fecundity was higher on non-Bt cotton than...

  1. Performance and cross-crop resistance of Cry1F-maize selected Spodoptera frugiperda on transgenic Bt cotton: implications for resistance management

    OpenAIRE

    Fei Yang; Kerns, David L.; Sebe Brown; Ryan Kurtz; Tim Dennehy; Bo Braxton; Graham Head; Fangneng Huang

    2016-01-01

    Transgenic crops producing Bacillus thuringiensis (Bt) proteins have become a primary tool in pest management. Due to the intensive use of Bt crops, resistance of the fall armyworm, Spodoptera frugiperda, to Cry1F maize has occurred in Puerto Rico, Brazil, and some areas of the southeastern U.S. The sustainability of Bt crops faces a great challenge because the Cry1F-maize resistant S. frugiperda may also infest other Bt crops in multiple cropping ecosystems. Here we examined the survival and...

  2. Influence of Bt rice plants on nitrogen fixation bacteria and horizontal transfer of the gene for Nitrogenase iron protein%种植转Bt水稻对固氮细菌多样性和固氮酶铁蛋白基因nifH的水平转移的影响

    Institute of Scientific and Technical Information of China (English)

    任少华; 徐斌; 黄晶心; 王婧; 章振亚; 陈艳; 肖明

    2012-01-01

    Enumeration of culturable bacteria colonies on nitrogen fixing medium (NFM) and LB medium conducted by this study revealed statistically significant difference among various soil samples collected from both transgenic Bacillus thuringiensis (Bt) rice fields and non-transgenic Bacillus thuringiensis (Bt) rice field.The transgenic Bt rice had generated impact on the quantities of microorganism communities in soil.Similar results were obtained by measuring shoot lengths after seeds germination.This also suggested that the planting of transgenic Bt rice may affect rice cultivation in the same field.However,such inhibitory effect was often transient in duration.Phylogenetic tree of 16S rDNAs showed a great genetic diversity of our isolated nitrogen fixation bacteria strains,and found that the majority of clones were related to two groups:one group belongs to the classes of Actinobacteria and the other group is Proteobacteria including α-Proteobacteri,more than half of nitrogen fixation bacteria (92%) in the rhizosphere belong to Actinobacteria.The sequences of the 16S rDNAs and the Nitrogenase iron protein coding genes (nifH) of eight nitrogen-fixing bacteria isolated from the transgenic Bt rice soil field were also compared.A lack of phylogenetic congruence of the nifH genes and 16S rDNAs might indicate the occurrence of relatively recent in situ horizontal transfer of the nifH gene,possibly as a direct or indirect consequence of transgenic Bt rice.%通过对具有不同种植年限和不同种植强度的转Bt基因水稻与非转Bt基因水稻土壤中的细菌以及固氮细菌群落结构进行研究,发现转Bt水稻的种植可能会影响土壤中微生物群落的多样性,但是这种影响的可能只是暂时的,通过对测量种植水稻的芽长实验也得出相似的结论.另外,根据16S rDNA基因构建的系统发育进化树揭示了本实验分离的固氮细菌的遗传多样性,发现实验土壤中的固氮细

  3. DYNAMICS OF EXPRESSION OF Bt INSECTICIDAL PROTEIN GENE IN THE TRANSGENIC RAPESEED AND ITS INSECT- RESISTANCE ACTIVITY%Bt杀虫蛋白基因在转基因油菜中的动态表达与其抗虫性研究

    Institute of Scientific and Technical Information of China (English)

    林良斌; 官春云; 杨焕文; 李永忠; 杨志新

    2001-01-01

    采用ELISA方法检测Bt杀虫基因在转基因油菜植株中的表达。试验结果表明:在第3至第9叶期Bt杀虫蛋白基因在转基因油莱植株中稳定、高效地表达,Bt杀虫蛋白的表达量为170ng/25mg~260ng25/mg鲜叶,占植物可溶性蛋白的0.067%~0.105%,其抗虫效果高达42.5%~80.0%。但从第11叶期后Bt杀虫蛋白表达量显著地降低,只有7ng/25mg~50ng/25mg鲜叶,占植物可溶性蛋白的0.003%~0.02%。%ELISA was adopted to analyze the expression of the insecticidal protein gene Bt in the transgenic rapeseed plants. The results showed that the expression of the protein gene was stable and highly efficient at the 3~9-leaf stage of the plants, with an expressing quantity of 170~260ng per25 mg fresh leaf,accounting for 0.067%~0.105% of the total soluble proteins of the plant and with an insecticidal activity of ashigh as 42.5%~80.0% .From the 11-leaf stage onward,the expressing quantity of Bt dropped sharply, being 7~50 ng per 25 mg fiesh leaf, accounting for 0.003%~0.02% of the total soluble proteins of the plants.

  4. 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

  5. Effects of transgenic Cry1Ac + CpTI cotton on non-target mealybug pest Ferrisia virgata and its predator Cryptolaemus montrouzieri

    OpenAIRE

    2014-01-01

    Recently, several invasive mealybugs (Hemiptera: Pseudococcidae) have rapidly spread to Asia and have become a serious threat to the production of cotton including transgenic cotton. Thus far, studies have mainly focused on the effects of mealybugs on non-transgenic cotton, without fully considering their effects on transgenic cotton and trophic interactions. Therefore, investigating the potential effects of mealybugs on transgenic cotton and their key natural enemies is vitally important. A ...

  6. Cry1F resistance in fall armyworm Spodoptera frugiperda: single gene versus pyramided Bt maize.

    Directory of Open Access Journals (Sweden)

    Fangneng Huang

    Full Text Available Evolution of insect resistance to transgenic crops containing Bacillus thuringiensis (Bt genes is a serious threat to the sustainability of this technology. However, field resistance related to the reduced efficacy of Bt maize has not been documented in any lepidopteran pest in the mainland U.S. after 18 years of intensive Bt maize planting. Here we report compelling evidence of field resistance in the fall armyworm, Spodoptera frugiperda (J.E. Smith, to Cry1F maize (TC 3507 in the southeastern region of the U.S. An F2 screen showed a surprisingly high (0.293 Cry1F resistance allele frequency in a population collected in 2011 from non-Bt maize in south Florida. Field populations from non-Bt maize in 2012-2013 exhibited 18.8-fold to >85.4-fold resistance to purified Cry1F protein and those collected from unexpectedly damaged Bt maize plants at several locations in Florida and North Carolina had >85.4-fold resistance. In addition, reduced efficacy and control failure of Cry1F maize against natural populations of S. frugiperda were documented in field trials using Cry1F-based and pyramided Bt maize products in south Florida. The Cry1F-resistant S. frugiperda also showed a low level of cross-resistance to Cry1A.105 and related maize products, but not to Cry2Ab2 or Vip3A. The occurrence of Cry1F resistance in the U.S. mainland populations of S. frugiperda likely represents migration of insects from Puerto Rico, indicating the great challenges faced in achieving effective resistance management for long-distance migratory pests like S. frugiperda.

  7. Cry1F resistance in fall armyworm Spodoptera frugiperda: single gene versus pyramided Bt maize.

    Science.gov (United States)

    Huang, Fangneng; Qureshi, Jawwad A; Meagher, Robert L; Reisig, Dominic D; Head, Graham P; Andow, David A; Ni, Xinzi; Kerns, David; Buntin, G David; Niu, Ying; Yang, Fei; Dangal, Vikash

    2014-01-01

    Evolution of insect resistance to transgenic crops containing Bacillus thuringiensis (Bt) genes is a serious threat to the sustainability of this technology. However, field resistance related to the reduced efficacy of Bt maize has not been documented in any lepidopteran pest in the mainland U.S. after 18 years of intensive Bt maize planting. Here we report compelling evidence of field resistance in the fall armyworm, Spodoptera frugiperda (J.E. Smith), to Cry1F maize (TC 3507) in the southeastern region of the U.S. An F2 screen showed a surprisingly high (0.293) Cry1F resistance allele frequency in a population collected in 2011 from non-Bt maize in south Florida. Field populations from non-Bt maize in 2012-2013 exhibited 18.8-fold to >85.4-fold resistance to purified Cry1F protein and those collected from unexpectedly damaged Bt maize plants at several locations in Florida and North Carolina had >85.4-fold resistance. In addition, reduced efficacy and control failure of Cry1F maize against natural populations of S. frugiperda were documented in field trials using Cry1F-based and pyramided Bt maize products in south Florida. The Cry1F-resistant S. frugiperda also showed a low level of cross-resistance to Cry1A.105 and related maize products, but not to Cry2Ab2 or Vip3A. The occurrence of Cry1F resistance in the U.S. mainland populations of S. frugiperda likely represents migration of insects from Puerto Rico, indicating the great challenges faced in achieving effective resistance management for long-distance migratory pests like S. frugiperda.

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

    Directory of Open Access Journals (Sweden)

    Laura M.N. Pinto

    2012-06-01

    Full Text Available 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.

  9. Daxx Interacts With Phage ΦBT1 Integrase and Inhibits Its Recombination%Daxx与ΦBT1相互作用并抑制其重组活性

    Institute of Scientific and Technical Information of China (English)

    王臻臻; 王然; 李文娟; 薛京伦; 陈金中

    2013-01-01

    The bacterial phage ΦBT1 integrase is a promising tool due to its site-specific transgene character.It enriches the site-specific transgenic tools and provides the possibility for multiple site-specific transgenic manipulations.To improve its safety as a vector of gene therapy,it is necessary to investigate the potential interactions between ΦBT1 and proteins in mammalian host cells.Yeast mating and co-immunoprecipitation assay indicated that a tetrapeptide 433RFAL436 in ΦBT1 integrase was responsible for ΦBT1 and Daxx interaction.It was also demonstrated that over-expression of Daxx could reduce ΦBT1 mediated recombination rate in 293T cells by using ΦBT1 report system.It is the first time to identify a cellular protein interacting with ΦBT1 integrase and inhibiting its recombination efficiency.This result might be useful for improving the ΦBT1 integrase mediated transgene methods and directing the selection of target cells for ΦBT1 integrase.%噬菌体整合酶BT1因其具有可介导转基因位点特异性整合的能力而成为了基因治疗中一种有效的工具,它丰富了转基因载体的选择,并使得多位点特异性整合成为可能.为了能够更加安全有效地利用ΦBT1整合酶作为基因治疗载体,有必要了解ΦBT1整合酶与宿主细胞内蛋白质相互作用的情况.酵母配对实验与免疫共沉淀实验揭示了ΦBT1整合酶中4个氨基酸433RFAL436对整合酶与PML-NBs蛋白Daxx的结合起到了关键作用.通过进一步构建并利用ΦBT1整合酶哺乳动物细胞报告系统,证实过表达Daxx会抑制ΦBT1整合酶在293T细胞中的重组效率.以上结果表明,细胞内的蛋白质可以与ΦBT1整合酶发生相互作用并抑制其重组活性,对于改善ΦBT1整合酶介导的转基因操作以及选择ΦBT1整合酶靶细胞方面具有重要的参考意义.

  10. Bacillus thuringiensis (Bt for the Control of Insect Pests in Stored Tobacco: A Review

    Directory of Open Access Journals (Sweden)

    Blanc M

    2014-12-01

    Full Text Available Among the insect species causing infestations and serious damages to stored commodities, the cigarette beetle, Lasiodermaserricorne (F. and the tobacco moth, Ephestiaelutella (Hübner are the major pests of both raw and manufactured tobacco. Post-harvest tobacco control is achieved through sanitation, insect monitoring, and fumigation with phosphine. However, insect resistance to phosphine and control failures have been reported, and increasing regulatory pressure is being exerted on fumigants. Biological control agents such as Bacillus thuringiensis (Bt appear to be environmentally sound and potentially viable alternatives to chemical control. Bt is a bacterium that produces insecticidal crystal proteins during the sporulation phase and has been, for more than 40 years, the microorganism of choice for the biocontrol of phytophagous insect pests. It produces insecticidal crystal proteins that display specific activity against certain orders of insects and become active upon ingestion by the insect. Our laboratory has conducted extensive research and worldwide surveys to evaluate the presence of Bt in stored tobacco and has confirmed previous findings indicating that Bt may be considered part of the naturally occurring phylloplanemicroflora. Several Bt strains were isolated from tobacco and characterized by DNA and protein profiling. The insecticidal activity of selected strains and of two commercial products against the larvae of L. serricorne was determined by diet incorporation assays. Moreover, the stability of Bt spores and crystal proteins on cured tobacco leaves was assessed over a storage period of time of 30 months. Cigarette prototypes were made with Bt-treated tobacco. Standard cigarette and smoke evaluations did not show any significant difference between the test and control cigarettes. Although the tested Bt strains and products did not yield satisfactory levels of mortality at the required times and doses, the experimental results

  11. Modeling evolution of resistance of sugarcane borer (Lepidoptera: Crambidae) to transgenic Bt corn

    NARCIS (Netherlands)

    Kang, J.; Huang, F.; Onstad, D. W.

    2014-01-01

    Diatraea saccharalis (F.) (Lepidoptera: Crambidae) is a target pest of transgenic corn expressing Bacillus thuringiensis (Bt) protein, and the first evidence of resistance by D. saccharalis to Cry1Ab corn was detected in a field population in northeast Louisiana in 2004. We used a model of populatio

  12. Incipient resistance of Helicoverpa punctigera to the Cry2Ab Bt toxin in Bollgard II cotton.

    Directory of Open Access Journals (Sweden)

    Sharon Downes

    Full Text Available Combinations of dissimilar insecticidal proteins ("pyramids" within transgenic plants are predicted to delay the evolution of pest resistance for significantly longer than crops expressing a single transgene. Field-evolved resistance to Bacillus thuringiensis (Bt transgenic crops has been reported for first generation, single-toxin varieties and the Cry1 class of proteins. Our five year data set shows a significant exponential increase in the frequency of alleles conferring Cry2Ab resistance in Australian field populations of Helicoverpa punctigera since the adoption of a second generation, two-toxin Bt cotton expressing this insecticidal protein. Furthermore, the frequency of cry2Ab resistance alleles in populations from cropping areas is 8-fold higher than that found for populations from non-cropping regions. This report of field evolved resistance to a protein in a dual-toxin Bt-crop has precisely fulfilled the intended function of monitoring for resistance; namely, to provide an early warning of increases in frequencies that may lead to potential failures of the transgenic technology. Furthermore, it demonstrates that pyramids are not 'bullet proof' and that rapid evolution to Bt toxins in the Cry2 class is possible.

  13. Impact of corn earworm injury on yield of transgenic corn producing Bt toxins in the Carolinas.

    Science.gov (United States)

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

    2014-06-01

    Transgenic corn, Zea mays L., hybrids expressing insecticidal Cry proteins from Bacillus thuringiensis (Bt) and insecticide applications to suppress injury from Helicoverpa zea (Boddie) were evaluated in Florence, SC, and in Plymouth, NC, in 2012 and 2013. Based on kernel area injured, insecticide applications (chlorantraniliprole) every 3-4 d from R1 until H. zea had cycled out of corn reduced injury by 80-93% in Florence and 94-95% in Plymouth. Despite intensive applications of insecticide (13-18 per trial), limited injury still occurred in all treated plots in 2012, except in DKC 68-03 (Genuity VT Double PRO), based on kernels injured (both locations) and proportion of injured ears (Florence only). In 2013, ear injury was low in Plymouth, with no kernel injury in any insecticide-treated plots, except P1498R (non-Bt) and P1498YHR (Optimum Intrasect). Injury in Florence in 2013 did not occur in treated plots of DKC 68-04 (non-Bt), DKC 68-03 (Genuity VT Double PRO), and N785-3111 (Agrisure Viptera). Yields were not significantly affected by insecticide treatment and were not statistically different among near-isolines with and without Bt traits. Yields were not significantly associated with kernel injury based on regression analyses. The value of using Bt corn hybrids to manage H. zea is discussed.

  14. Practice Tests for the TOEFL iBT

    CERN Document Server

    Stirling, Bruce

    2012-01-01

    Practice Tests for the TOEFL iBT contains four TOEFL tests, with answer keys. Perfect for self-study and classrooms. Each TOEFL iBT Practice Test...* reflects the design of the official TOEFL internet-based test* tests academic English-language proficiency expected of university students in the United States, Canada, Australia, New Zealand, Ireland, Scotland and England* provides extra practice before you take the official TOEFL iBT* will help you identify those areas of academic English you need to improve for a higher TOEFL iBT score* will give you an unofficial, TOEFL iBT range score within

  15. Commercial production of transgenic Bt insect-resistant cotton varieties and the resistance management for bollworm (Helicoverpa armigera Hubner)

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    There are currently three kinds of transgenic Bt insect-resistant cotton germplasm lines, Shanxi 94-24, Zhongxin 94 and R19, in China. They showed high resistance to the neonate larvae of bollworm (Helicoverpa armigera). Transgenic Bt insect-resistant cotton varieties or hybrids have been bred using the three kinds of germplasm lines as parents. Our researches reveal that there exist different expressions in resistant level at different developmental stages in the three categories of germplasm lines. When neonate larvae are fed with leaves of cotton plant at the seeding stage with less than 10 leaves on the main stem, the mortality of the neonate larvae is 100%, but the resistance level will decline at later season. When Bt gene has been transferred to the cotton genome, it can be steadily transferred to the progeny, the level of resistance to bollworm keeps fundamentally uniform. Such insects as tobacco budworm (Heliothis virencens) in laboratory directive selection are very apt to produce resistance to the Bt insecticidal crystal protein. From the present crop system of cotton region in the Yangtze and Yellow River Valleys, and the expression characteristic of transgenic Bt resistant cotton, we suggest that the resistance to toxin protein in bollworm is not apt to be produced if the transgenic Bt insect-resistant cotton varieties are released and grown in the regions except in the Xinjiang cotton region. The managing strategies to delay or retard the resistance are discussed.

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

    Science.gov (United States)

    Wang, Yueqin; Wang, Yidong; Wang, Zhenying; Bravo, Alejandra; Soberón, Mario; He, Kanglai

    2016-01-01

    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.

  17. BT's adoption of customer centric design.

    Science.gov (United States)

    Chamberlain, Mark; Esquivel, Jacqueline; Miller, Fiona; Patmore, Jeff

    2015-01-01

    Between 2005 and 2010 BT underwent a major transformation from a company with a special section devoted to 'older and disabled consumers' to a company with an inclusive design strategy. The mainstreaming of these issues responded to a demand for better, more user-friendly communications products and growing awareness of the importance of previously marginalised consumer groups. It also took place alongside the development and publication of BS7000-6, a guide to inclusive design management. Based on several product design case studies, this paper reflects on how and why this transformation was seen as necessary for future success, and how the transformation was achieved. The evolution of BT's approach has continued since, but this paper looks back in time, and documents the transformation up to 2010 and reflects the state of the company in 2010 rather than at the time of publication.

  18. 捷波朗BT8010

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    捷波朗最新发布的蓝牙耳机产品BT8010采用了独特的双听筒设计,支持电话簿和均衡器等个性化功能、OLED显示屏对比度良好,“Jog Wheel缓动轮”设计大大提高了产品的操作感受。

  19. A critical assessment of the effects of Bt transgenic plants on parasitoids.

    Directory of Open Access Journals (Sweden)

    Mao Chen

    Full Text Available The ecological safety of transgenic insecticidal plants expressing crystal proteins (Cry toxins from the bacterium Bacillus thuringiensis (Bt continues to be debated. Much of the debate has focused on nontarget organisms, especially predators and parasitoids that help control populations of pest insects in many crops. Although many studies have been conducted on predators, few reports have examined parasitoids but some of them have reported negative impacts. None of the previous reports were able to clearly characterize the cause of the negative impact. In order to provide a critical assessment, we used a novel paradigm consisting of a strain of the insect pest, Plutella xylostella (herbivore, resistant to Cry1C and allowed it to feed on Bt plants and then become parasitized by Diadegma insulare, an important endoparasitoid of P. xylostella. Our results indicated that the parasitoid was exposed to a biologically active form of the Cy1C protein while in the host but was not harmed by such exposure. Parallel studies conducted with several commonly used insecticides indicated they significantly reduced parasitism rates on strains of P. xylostella resistant to these insecticides. These results provide the first clear evidence of the lack of hazard to a parasitoid by a Bt plant, compared to traditional insecticides, and describe a test to rigorously evaluate the risks Bt plants pose to predators and parasitoids.

  20. Mode of inheritance for biochemical traits in genetically engineered cotton under water stress.

    Science.gov (United States)

    Abid, Muhammad Ali; Malik, Waqas; Yasmeen, Azra; Qayyum, Abdul; Zhang, Rui; Liang, Chengzhen; Guo, Sandui; Ashraf, Javaria

    2016-02-02

    Drought is an abiotic environmental stress that can significantly reduce crop productivity. We examined the mode of inheritance for different biochemical traits including total soluble proteins, chlorophyll a, chlorophyll b, total chlorophyll, carotenoids, total phenolic contents and enzymatic antioxidants (superoxide dismutase, peroxidase and catalase), and their relationship with Bacillus thuringiensis (Bt) toxin under control and drought conditions. Eight genetically diverse cotton genotypes were selfed for two generations to ensure homozygosity. Fifteen F1 hybrids were developed by crossing five non-Bt female lines with three Bt male testers. The F1 hybrids and eight parents were finally evaluated under control (100 % field capacity (FC)) and drought (50 % FC) conditions in 2013. The biochemical traits appeared to be controlled by non-additive gene action with low narrow sense heritability estimates. The estimates of general combining ability and specific combining ability for all biochemical traits were significant under control and drought conditions. The genotype-by-trait biplot analysis showed the better performance of Bt cotton hybrids when compared with their parental genotypes for various biochemical traits under control and drought conditions. The biplot and path coefficient analyses revealed the prevalence of different relationships between Cry1Ac toxin and biochemical traits in the control and drought conditions. In conclusion, biochemical traits could serve as potential biochemical markers for breeding Bt cotton genotypes without compromising the optimal level of Bt toxin.

  1. Detection and monitoring of insect resistance to transgenic Bt crops

    Institute of Scientific and Technical Information of China (English)

    FANGNENG HUANG

    2006-01-01

    Transgenic crops expressing Bacillus thuringiensis (Bt) endotoxins have become one of the most important tools for managing corn and cotton insect pests in the US and other countries. The widespread adoption of transgenic Bt crops could place a high degree of selection pressure on the target insect populations and accelerate development of resistance, raising concerns about the long-term durability of Bt plants as an effective pest management tool. Conservation of Bt susceptibility in insects has become one of the most active research areas in modern agriculture. One of the key factors for a successful Bt resistance management plan is to have a cost-effective monitoring system that can provide information on: (i) the initial Bt resistance allele frequencies at low levels in field insect populations; and (ii) early shifts in Bt resistance allele frequencies so that proactive measures for managing resistance can be deployed well before field control failures. Developing such a monitoring program has been difficult because: (i) resistance traits that occur at very low frequencies are hard to detect; (ii) many factors affect the sensitivity and accuracy of a Bt resistance monitoring program; and (iii) monitoring resistance is costly. Several novel methods for detecting Bt resistance alleles developed during the last decade have made a cost-effective monitoring system possible. Future studies should focus on how to improve and standardize the methodologies for insect sampling and Bt resistance detection.

  2. Exposure and nontarget effects of transgenic Bt corn debris in streams.

    Science.gov (United States)

    Jensen, Peter D; Dively, Galen P; Swan, Christopher M; Lamp, William O

    2010-04-01

    Corn (Zea mays L.) transformed with a gene from the bacterium Bacillus thuringiensis (Bt) comprises 49% of all corn in the United States. The input of senesced corn tissue expressing the Bt gene may impact stream-inhabiting invertebrates that process plant debris, especially trichopteran species related to the target group of lepidopteran pests. Our goal was to assess risk associated with transgenic corn debris entering streams. First, we show the input of corn tissue after harvest was extended over months in a stream. Second, using laboratory bioassays based on European corn borer [Ostrinia nubilalis (Hübner)], we found no bioactivity of Cry1Ab protein in senesced corn tissue after 2 wk of exposure to terrestrial or aquatic environments. Third, we show that Bt near-isolines modify growth and survivorship of some species of invertebrates. Of the four nontarget invertebrate species fed Bt near-isolines, growth of two closely related trichopterans was not negatively affected, whereas a tipulid crane fly exhibited reduced growth rates, and an isopod exhibited reduced growth and survivorship on the Cry1Ab near-isoline but not on the stacked Cry1Ab + Cry3Bb1 near-isoline. Because of lack of evidence of bioactivity of Bt after 2 wk and because of lack of nontarget effects on the stacked near-isoline, we suggest that tissue-mediated differences, and not the presence of the Cry1Ab protein, caused the different responses among the species. Overall, our results provide evidence that adverse effects to aquatic nontarget shredders involve complex interactions arising from plant genetics and environment that cannot be ascribed to the presence of Cry1Ab proteins.

  3. Consequences for Protaphorura armata (Collembola: Onychiuridae) following exposure to genetically modified Bacillus thuringiensis (Bt) maize and non-Bt maize

    Energy Technology Data Exchange (ETDEWEB)

    Heckmann, Lars-Henrik [National Environmental Research Institute, Department of Terrestrial Ecology, Vejlsovej 25, PO Box 314, DK-8600 Silkeborg (Denmark); Griffiths, Bryan S. [Scottish Crop Research Institute, Department of Soil Plant Dynamics, Invergowrie, Dundee DD2 5DA (United Kingdom); Caul, Sandra [Scottish Crop Research Institute, Department of Soil Plant Dynamics, Invergowrie, Dundee DD2 5DA (United Kingdom); Thompson, Jacqueline [Scottish Crop Research Institute, Department of Soil Plant Dynamics, Invergowrie, Dundee DD2 5DA (United Kingdom); Pusztai-Carey, Marianne [Case Western Reserve University, Cleveland, OH 44106 (United States); Moar, William J. [Auburn University, Department of Entomology and Plant Pathology, Auburn, AL 36849 (United States); Andersen, Mathias N. [Danish Institute of Agricultural Sciences, Research Centre Foulum, PO Box 50, DK-8830 Tjele (Denmark); Krogh, Paul Henning [National Environmental Research Institute, Department of Terrestrial Ecology, Vejlsovej 25, PO Box 314, DK-8600 Silkeborg (Denmark)]. E-mail: phk@dmu.dk

    2006-07-15

    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 Protaphorura armata. For comparison three non-Bt maize varieties, Rivaldo (isogenic to Cascade), Monumental (isogenic to MEB307) and DK242, and two control diets based on baker's yeast (uncontaminated and contaminated with Bt toxin Cry1Ab) were also tested. Due to a lower C:N ratio, individuals reared on yeast performed significantly better in all of the measured endpoints than those reared on maize. P. armata performed equally well when reared on two Bt and three non-Bt maize varieties. Although there were no negative effects of Bt maize in this experiment, we recommend future studies on Bt crops to focus on species interactions in long-term, multi-species experiments. - Protaphorura armata performed equally well when reared on two Bt and three non-Bt maize varieties.

  4. Dispersal and movement behavior of neonate European corn borer (Lepidoptera: Crambidae) on non-Bt and transgenic Bt corn.

    Science.gov (United States)

    Goldstein, Jessica A; Mason, Charles E; Pesek, John

    2010-04-01

    Neonate movement and dispersal behavior of the European corn borer, Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae), were investigated under controlled conditions on Bacillus thuringiensis (Bt) and non-Bt corn, Zea mays L., to assess plant abandonment, dispersal from their natal plant, and silking behavior after Bt and non-Bt preexposure. With continuous airflow, neonates on a Bt corn plant for 24 h abandoned that plant 1.78 times more frequently than neonates on a non-Bt corn plant. Indirect evidence indicated that at least one third of the neonates were capable of ballooning within 24 h. In the greenhouse, some neonates were recovered after 24 h from plants 76 and 152 cm away that likely ballooned from their natal plant. After 1 h of preexposure on a Bt corn leaf, neonates placed on a new corn leaf and observed for 10 min began silking off of a new Bt leaf significantly sooner than a new non-Bt leaf. Results suggest that neonates are unable to detect Bt in the corn within 10 min but that they can detect it within the first hour.

  5. TOEFL strategies a complete guide to the iBT

    CERN Document Server

    Stirling, Bruce

    2016-01-01

    TOEFL students all ask: How can I get a high TOEFL iBT score? Answer: Learn argument scoring strategies. Why? Because the TOEFL iBT recycles opinion-based and fact-based arguments for testing purposes from start to finish. In other words, the TOEFL iBT is all arguments. That's right, all arguments. If you want a high score, you need essential argument scoring strategies. That is what TOEFL STRATEGIES A COMPLETE GUIDE gives you, and more!

  6. Effects of transgenic Bacillus thuringiensis maize grain on B. thuringiensis-susceptible Plodia interpunctella (Lepidoptera: Pyralidae).

    Science.gov (United States)

    Giles, K L; Hellmich, R L; Iverson, C T; Lewis, L C

    2000-06-01

    Percentage survivorship, developmental time, adult body length, and sex ratio of Plodia interpunctella (Hübner) reared on field-produced grain from sixteen cultivars of maize, Zea mays L., including several transgenic Bacillus thuringiensis (Bt) Berliner hybrids and selected non-Bt isolines, were evaluated under laboratory conditions. Compared with isolines, development was delayed and survivorship reduced for P. interpunctella reared on grain from transgenic hybrids with the CaMV/35s promoter that express Cry1Ab protein. Similarly, compared with non-Bt hybrids, a transgenic hybrid with the CaMV/35s promoter that expresses Cry9C protein delayed development, decreased survivorship, and caused reductions in adult body length of P. interpunctella. In contrast, no significant differences in P. interpunctella developmental times or survivorship were observed between transgenic hybrids with the PEPC promoter expressing Cry1Ab and their isolines. Additionally, developmental time, survivorship, and adult body length were similar between P. interpunctella reared on a transgenic hybrid with the CaMV/35s promoter expressing Cry1Ac and non-Bt hybrids. Our data demonstrate that transgenic Bt maize grain, especially grain from hybrids with the CaMV/35s promoter expressing Cry1Ab or Cry9C, can significantly affect B. thuringiensis-susceptible P. interpunctella populations up to 4 or 5 mo after harvest.

  7. The compatibility of a nucleopolyhedrosis virus control with resistance management for Bacillus thuringiensis: co-infection and cross-resistance studies with the diamondback moth, Plutella xylostella.

    Science.gov (United States)

    Raymond, B; Sayyed, A H; Wright, D J

    2006-10-01

    The use of genetically modified crops expressing Bacillus thuringiensis (Bt) toxins can lead to the reduction in application of broad-spectrum pesticides and an increased opportunity for supplementary biological control. Bt microbial sprays are also used by organic growers or as part of integrated pest management programs that rely on the use of natural enemies. In both applications the evolution of resistance to Bt toxins is a potential problem. Natural enemies (pathogens or insects) acting in combination with toxins can accelerate or decelerate the evolution of resistance to Bt. In the present study we investigated whether the use of a nucleopolyhedrovirus (AcMNPV) could potentially affect the evolution of resistance to the Bt toxin Cry1Ac in Plutella xylostella. At low toxin doses there was evidence for antagonistic interactions between AcMNPV and Cry1Ac resistant and susceptible insects. However, this antagonism was much stronger and more widespread for susceptible larvae; interactions were generally not distinguishable from additive for resistant larvae. Selection for resistance to Cry1Ac in two populations of P. xylostella with differing resistance mechanisms did not produce any correlated changes in resistance to AcMNPV. Stronger antagonistic interactions between Bt and AcMNPV on susceptible rather than resistant larvae can decrease the relative fitness between Bt-resistant and susceptible larvae. These interactions and the lack of cross-resistance between virus and toxin suggest that the use of NPV is compatible with resistance management to Bt products.

  8. The development and status of Bt rice in China.

    Science.gov (United States)

    Li, Yunhe; Hallerman, Eric M; Liu, Qingsong; Wu, Kongming; Peng, Yufa

    2016-03-01

    Multiple lines of transgenic rice expressing insecticidal genes from the bacterium Bacillus thuringiensis (Bt) have been developed in China, posing the prospect of increases in production with decreased application of pesticides. We explore the issues facing adoption of Bt rice for commercial production in China. A body of safety assessment work on Bt rice has shown that Bt rice poses a negligible risk to the environment and that Bt rice products are as safe as non-Bt control rice products as food. China has a relatively well-developed regulatory system for risk assessment and management of genetically modified (GM) plants; however, decision-making regarding approval of commercial production has become politicized, and two Bt rice lines that otherwise were ready have not been allowed to enter the Chinese agricultural system. We predict that Chinese farmers would value the prospect of increased yield with decreased use of pesticide and would readily adopt production of Bt rice. That Bt rice lines may not be commercialized in the near future we attribute to social pressures, largely due to the low level of understanding and acceptance of GM crops by Chinese consumers. Hence, enhancing communication of GM crop science-related issues to the public is an important, unmet need. While the dynamics of each issue are particular to China, they typify those in many countries where adoption of GM crops has been not been rapid; hence, the assessment of these dynamics might inform resolution of these issues in other countries.

  9. TMD factorization and evolution at large $b_T$

    Energy Technology Data Exchange (ETDEWEB)

    Collins, John [Pennsylvania State Univ., University Park, PA (United States); Rogers, Ted [Thomas Jefferson National Accelerator Facility (TJNAF), Newport News, VA (United States); Old Dominion Univ., Norfolk, VA (United States)

    2015-07-20

    In using transverse-momentum-dependent (TMD) parton densities and fragmentation functions, important non-perturbative information is at large transverse position $b_T$. This concerns both the TMD functions and their evolution. Fits to high energy data tend to predict too rapid evolution when extrapolated to low energies where larger values of $b_T$ dominate. I summarize a new analysis of the issues. It results in a proposal for much weaker $b_T$ dependence at large $b_T$ for the evolution kernel, while preserving the accuracy of the existing fits. The results are particularly important for using transverse-spin-dependent functions like the Sivers function.

  10. Two BT-VSA Solvable Models

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Variable separation approach that is based on Backlund transformation (BT-VSA) is extended to solve the (3+1)-dimensional Jimbo-Miwa equation and the (1+1)-dimensional Drinfel'd-Sokolov-Wilson equation. New ex act solutions, which include some low-dimensional functions, are obtained. One of the low-dimensional function is arbitrary and another must satisfy a Riccati equation. Some new localized excitations can be derived from (2+1)-dimensional localized excitations and for simplification, we omit those in this letter.

  11. Testing pollen of single and stacked insect-resistant Bt-maize on in vitro reared honey bee larvae.

    Science.gov (United States)

    Hendriksma, Harmen P; Härtel, Stephan; Steffan-Dewenter, Ingolf

    2011-01-01

    The ecologically and economic important honey bee (Apis mellifera) is a key non-target arthropod species in environmental risk assessment (ERA) of genetically modified (GM) crops. Honey bee larvae are directly exposed to transgenic products by the consumption of GM pollen. But most ERA studies only consider responses of adult bees, although Bt-proteins primarily affect the larval phases of target organisms. We adopted an in vitro larvae rearing system, to assess lethal and sublethal effects of Bt-pollen consumption in a standardized eco-toxicological bioassay. The effects of pollen from two Bt-maize cultivars, one expressing a single and the other a total of three Bt-proteins, on the survival and prepupae weight of honey bee larvae were analyzed. The control treatments included pollen from three non-transgenic maize varieties and of Heliconia rostrata. Three days old larvae were fed the realistic exposure dose of 2 mg pollen within the semi-artificial diet. The larvae were monitored over 120 h, until the prepupal stage, where larvae terminate feeding and growing. Neither single nor stacked Bt-maize pollen showed an adverse effect on larval survival and the prepupal weight. In contrast, feeding of H. rostrata pollen caused significant toxic effects. The results of this study indicate that pollen of the tested Bt-varieties does not harm the development of in vitro reared A. mellifera larvae. To sustain the ecosystem service of pollination, Bt-impact on A. mellifera should always be a crucial part of regulatory biosafety assessments. We suggest that our approach of feeding GM pollen on in vitro reared honey bee larvae is well suited of becoming a standard bioassay in regulatory risk assessments schemes of GM crops.

  12. Testing pollen of single and stacked insect-resistant Bt-maize on in vitro reared honey bee larvae.

    Directory of Open Access Journals (Sweden)

    Harmen P Hendriksma

    Full Text Available The ecologically and economic important honey bee (Apis mellifera is a key non-target arthropod species in environmental risk assessment (ERA of genetically modified (GM crops. Honey bee larvae are directly exposed to transgenic products by the consumption of GM pollen. But most ERA studies only consider responses of adult bees, although Bt-proteins primarily affect the larval phases of target organisms. We adopted an in vitro larvae rearing system, to assess lethal and sublethal effects of Bt-pollen consumption in a standardized eco-toxicological bioassay. The effects of pollen from two Bt-maize cultivars, one expressing a single and the other a total of three Bt-proteins, on the survival and prepupae weight of honey bee larvae were analyzed. The control treatments included pollen from three non-transgenic maize varieties and of Heliconia rostrata. Three days old larvae were fed the realistic exposure dose of 2 mg pollen within the semi-artificial diet. The larvae were monitored over 120 h, until the prepupal stage, where larvae terminate feeding and growing. Neither single nor stacked Bt-maize pollen showed an adverse effect on larval survival and the prepupal weight. In contrast, feeding of H. rostrata pollen caused significant toxic effects. The results of this study indicate that pollen of the tested Bt-varieties does not harm the development of in vitro reared A. mellifera larvae. To sustain the ecosystem service of pollination, Bt-impact on A. mellifera should always be a crucial part of regulatory biosafety assessments. We suggest that our approach of feeding GM pollen on in vitro reared honey bee larvae is well suited of becoming a standard bioassay in regulatory risk assessments schemes of GM crops.

  13. Does Bt maize cultivation affect the non-target insect community in the agro ecosystem?

    Directory of Open Access Journals (Sweden)

    Daniela Chaves Resende

    2016-03-01

    Full Text Available ABSTRACT The cultivation of genetically modified crops in Brazil has led to the need to assess the impacts of this technology on non-target species. Under field conditions, the potential effect on insect biodiversity was evaluated by comparing a homogeneous corn field with conventional and transgenic maize, expressing different Bt proteins in seven counties of Minas Gerais, Brazil. The richness pattern of non-target insect species, secondary pests and natural enemies were observed. The results do not support the hypothesis that Bt protein affects insect biodiversity. The richness and diversity data of insects studied were dependent on the location and other factors, such as the use of insecticides, which may be a major factor where they are used.

  14. Effects of Bt-maize material on the life cycle of the land snail Cantareus aspersus

    DEFF Research Database (Denmark)

    Kramarz, Paulina; de Vaufleury, Annette; Gimbert, Frédéric

    2009-01-01

    Insect resistant Bt-maize (MON 810) expresses active Cry1Ab endotoxin derived from Bacillus thuringiensis (Bt). Snails constitute non-target soil species potentially exposed to Bt-toxin through consumption of plant material and soil in fields where transgenic plants have been grown. We studied...... the effect of the Cry1Ab toxin on survival, growth and egg hatchability of the snail Cantareus aspersus. From the age of 4 to 88 weeks, snails were fed either powdered Bt-maize or non-Bt-maize and exposed to soil samples collected after harvesting either the Bt-maize or non-Bt-maize. We applied four...... treatments: non-Bt soil + non-Bt-maize (MM); Bt soil + Bt-maize (BB), non-Bt soil + Bt-maize (MB), Bt soil + non-Bt-maize (BM). Eggs laid by snails not exposed to Bt-toxin were also exposed to the two types of soils (Bt and non-Bt soil). At the end of growth (47 weeks of exposure), snails exposed to Bt...

  15. Consumption of Bt Maize Pollen Containing Cry1Ie Does Not Negatively Affect Propylea japonica (Thunberg (Coleoptera: Coccinellidae

    Directory of Open Access Journals (Sweden)

    Yonghui Li

    2017-03-01

    Full Text Available Propylea japonica (Thunberg (Coleoptera: Coccinellidae are prevalent predators and pollen feeders in East Asian maize fields. They are therefore indirectly (via prey and directly (via pollen exposed to Cry proteins within Bt-transgenic maize fields. The effects of Cry1Ie-producing transgenic maize pollen on the fitness of P. japonica was assessed using two dietary-exposure experiments in the laboratory. In the first experiment, survival, larval developmental time, adult fresh weight, and fecundity did not differ between ladybirds consuming Bt or non-Bt maize pollen. In the second experiment, none of the tested lethal and sublethal parameters of P. japonica were negatively affected when fed a rapeseed pollen-based diet containing Cry1Ie protein at 200 μg/g dry weight of diet. In contrast, the larval developmental time, adult fresh weight, and fecundity of P. japonica were significantly adversely affected when fed diet containing the positive control compound E-64. In both experiments, the bioactivity of the Cry1Ie protein in the food sources was confirmed by bioassays with a Cry1Ie-sensitive lepidopteran species. These results indicated that P. japonica are not affected by the consumption of Cry1Ie-expressing maize pollen and are not sensitive to the Cry1Ie protein, suggesting that the growing of Bt maize expressing Cry1Ie protein will pose a negligible risk to P. japonica.

  16. Consumption of Bt Maize Pollen Containing Cry1Ie Does Not Negatively Affect Propylea japonica (Thunberg) (Coleoptera: Coccinellidae)

    Science.gov (United States)

    Li, Yonghui; Liu, Yanmin; Yin, Xinming; Romeis, Jörg; Song, Xinyuan; Chen, Xiuping; Geng, Lili; Peng, Yufa; Li, Yunhe

    2017-01-01

    Propylea japonica (Thunberg) (Coleoptera: Coccinellidae) are prevalent predators and pollen feeders in East Asian maize fields. They are therefore indirectly (via prey) and directly (via pollen) exposed to Cry proteins within Bt-transgenic maize fields. The effects of Cry1Ie-producing transgenic maize pollen on the fitness of P. japonica was assessed using two dietary-exposure experiments in the laboratory. In the first experiment, survival, larval developmental time, adult fresh weight, and fecundity did not differ between ladybirds consuming Bt or non-Bt maize pollen. In the second experiment, none of the tested lethal and sublethal parameters of P. japonica were negatively affected when fed a rapeseed pollen-based diet containing Cry1Ie protein at 200 μg/g dry weight of diet. In contrast, the larval developmental time, adult fresh weight, and fecundity of P. japonica were significantly adversely affected when fed diet containing the positive control compound E-64. In both experiments, the bioactivity of the Cry1Ie protein in the food sources was confirmed by bioassays with a Cry1Ie-sensitive lepidopteran species. These results indicated that P. japonica are not affected by the consumption of Cry1Ie-expressing maize pollen and are not sensitive to the Cry1Ie protein, suggesting that the growing of Bt maize expressing Cry1Ie protein will pose a negligible risk to P. japonica. PMID:28300767

  17. Cotton bollworm resistance to Bt transgenic cotton: A case analysis

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Cotton bollworm (Helicoverpa armigera) is one of the most serious insect pests of cotton. Transgenic cotton expressing Cry toxins derived from a soil bacterium, Bacillus thuringiensis (Bt), has been produced to target this pest. Bt cotton has been widely planted around the world, and this has resulted in efficient control of bollworm populations with reduced use of synthetic insecticides. However, evolution of resistance by this pest threatens the continued success of Bt cotton. To date, no field populations of bollworm have evolved significant levels of resistance; however, several laboratory-selected Cry-resistant strains of H. armigera have been obtained, which suggests that bollworm has the capacity to evolve resistance to Bt. The development of resistance to Bt is of great concern, and there is a vast body of research in this area aimed at ensuring the continued success of Bt cotton. Here, we review studies on the evolution of Bt resistance in H. armigera, focusing on the biochemical and molecular basis of Bt resistance. We also discuss resistance management strategies, and monitoring programs implemented in China, Australia, and India.