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61.
Génissel A Augustin S Courtin C Pilate G Lorme P Bourguet D 《Proceedings. Biological sciences / The Royal Society》2003,270(1517):791-797
Globally, the estimated total area planted with transgenic plants producing Bacillus thuringiensis (Bt) toxins was 12 million hectares in 2001. The risk of target pests becoming resistant to these toxins has led to the implementation of resistance-management strategies. The efficiency and sustainability of these strategies, including the high-dose plus refuge strategy currently recommended for North American maize, depend on the initial frequency of resistance alleles. In this study, we estimated the initial frequencies of alleles conferring resistance to transgenic Bt poplars producing Cry3A in a natural population of the poplar pest Chrysomela tremulae (Coleoptera: Chrysomelidae). We used the F(2) screen method developed for detecting resistance alleles in natural pest populations. At least three parents of the 270 lines tested were heterozygous for a major Bt resistance allele. We estimated mean resistance-allele frequency for the period 1999-2001 at 0.0037 (95% confidence interval = 0.00045-0.0080) with a detection probability of 90%. These results demonstrate that (i) the F(2) screen method can be used to detect major alleles conferring resistance to Bt-producing plants in insects and (ii) the initial frequency of alleles conferring resistance to Bt toxin can be close to the highest theoretical values that are expected prior to the use of Bt plants if considering fitness costs and typical mutation rates. 相似文献
62.
Application of a Specific and Sensitive Radiometric
Assay for Microbial Lipase Activities in Marine Water Samples
from the Lagoon of
Nouméa 下载免费PDF全文
Nicolas Bourguet Jean-Pascal Torrton Olivier Galy Vincent Arondel Madeleine Goutx 《Applied microbiology》2003,69(12):7395-7400
Marine microbiologists commonly assay lipase activities by using a synthetic fluorescent analog, 4-methylumbelliferyl (MUF)-oleate. The technique is convenient, but it is considered to be unspecific because of the structure of this analog. This study reports the design of a new specific and sensitive lipase assay based on the use of a radiolabeled triglyceride, [3H]triolein. Free fatty acids (FFA) resulting from its hydrolysis are isolated as a function of time in a one-step liquid-liquid extraction and then radioassayed. MUF-oleate and [3H]triolein techniques were compared by measuring lipase activities at similar substrate concentrations along a trophic gradient in the Southwest Lagoon of New Caledonia, near Nouméa. Hydrolysis rates decreased from the nearshore station to the offshore station and showed similar trends regardless of the technique used. Rates decreased from 5.83 to 0.88 nmol of FFA·liter−1·h−1 and from 0.76 to 0.23 nmol of 3H-FFA·liter−1·h−1, respectively. These results appeared to be consistent with bacterial production results, which also decreased similarly (from 0.59 to 0.26 μg of C·liter−1·h−1). However, the ratio of MUF-oleate activities to [3H]triolein activities, which was constant at the offshore stations (3.8 ± 0.1), gradually increased at the nearshore stations (from 4.1 to 7.6). This result shows that the two assays respond in different ways to changes in environmental conditions and validates the need to set up more specific enzymatic assays. 相似文献
63.
Johnson KA Angelucci F Bellelli A Hervé M Fontaine J Tsernoglou D Capron A Trottein F Brunori M 《Biochemistry》2003,42(34):10084-10094
Schistomiasis is a debilitating parasitic disease which affects 200 million people, causing life-threatening complications in 10% of the patients. This paper reports the crystal structure of the Schistosoma haematobium 28 kDa glutathione S-transferase, a multifunctional enzyme involved in host-parasite interactions and presently considered as a promising vaccine candidate against schistosomiasis. The structures of the GSH-free enzyme, as well as the partially (approximately 40%) and almost fully (approximately 80%) GSH-saturated enzyme, exhibit a unique feature, absent in previous GST structures, concerning the crucial and invariant Tyr10 side chain which occupies two alternative positions. The canonical conformer, which allows an H-bond to be formed between the side chain hydroxyl group and the activated thiolate of GSH, is somewhat less than 50% occupied. The new conformer, with the phenoxyl ring on the opposite side of the mobile loop connecting strand 1 and helix 1, is stabilized by a polar interaction with the guanidinium group of the conserved Arg21 side chain. The presence of two conformers of Tyr10 may provide a clue about clarifying the multiple catalytic functions of Sh28GST and might prove to be relevant for the design of specific antischistosomal drugs. The K(d) for GSH binding was determined by equilibrium fluorescence titrations to be approximately 3 microM and by stopped-flow rapid mixing experiments to be approximately 9 microM. The relatively tight binding of GSH by Sh28GST explains the residually bound GSH in the crystal and supports a possible role of GSH as a tightly bound cofactor involved in the catalytic mechanism for prostaglandin D(2) synthase activity. 相似文献
64.
65.
Insecticide resistance and dominance levels 总被引:11,自引:0,他引:11
Dominance has been assessed in different ways in insecticide resistance studies, based on three phenotypic traits: the insecticide concentration required to give a particular mortality (DLC), mortality at a particular insecticide dose (DML), and fitness in treated areas (DWT). We propose a general formula for estimating dominance on a scale of 0 to 1 (0 = complete recessivity and 1 = complete dominance). DLC, DML, and DWT are not directly related and their values depend on genetic background and environmental conditions. We also show that pest management strategies can have the consequence to increase DWT via the selection of dominance modifiers. Studies on resistance to Bacillus thuringiensis toxins provide the ultimate example of the complexity of the definition of the concept of dominance. Almost all studies have focused on calculation of DLC, which provides little information about the efficiency of pest management programs. For instance, one assumption of the high dose/refuge strategy is that Bacillus thuringiensis resistance must be effectively recessive (i.e., DML must be close to zero). However, DWT, rather than DML, is relevant to the resistance management strategy. Therefore, we strongly suggest that the time has come to focus on fitness dominance levels in the presence and absence of insecticide. 相似文献
66.
67.
Integrating Phosphoproteome and Transcriptome Reveals New Determinants of Macrophage Multinucleation
68.
69.
Maxime Bourgognon Rebecca Klippstein Khuloud T. Al-Jamal 《Journal of visualized experiments : JoVE》2015,(102)
The large majority of in vitro nanotoxicological studies have used immortalized cell lines for their practicality. However, results from nanoparticle toxicity testing in immortalized cell lines or primary cells have shown discrepancies, highlighting the need to extend the use of primary cells for in vitro assays. This protocol describes the isolation of mouse liver macrophages, named Kupffer cells, and their use to study nanoparticle toxicity. Kupffer cells are the most abundant macrophage population in the body and constitute part of the reticulo-endothelial system (RES), responsible for the capture of circulating nanoparticles. The Kupffer cell isolation method reported here is based on a 2-step perfusion method followed by purification on density gradient. The method, based on collagenase digestion and density centrifugation, is adapted from the original protocol developed by Smedsrød et al. designed for rat liver cell isolation and provides high yield (up to 14 x 106 cells per mouse) and high purity (>95%) of Kupffer cells. This isolation method does not require sophisticated or expensive equipment and therefore represents an ideal compromise between complexity and cell yield. The use of heavier mice (35-45 g) improves the yield of the isolation method but also facilitates remarkably the procedure of portal vein cannulation. The toxicity of functionalized carbon nanotubes f-CNTs was measured in this model by the modified LDH assay. This method assesses cell viability by measuring the lack of structural integrity of Kupffer cell membrane after incubation with f-CNTs. Toxicity induced by f-CNTs can be measured consistently using this assay, highlighting that isolated Kupffer cells are useful for nanoparticle toxicity testing. The overall understanding of nanotoxicology could benefit from such models, making the nanoparticle selection for clinical translation more efficient. 相似文献
70.
Gaetane Nocturne Stephan Pavy Saida Boudaoud Raphaèle Seror Philippe Goupille Philippe Chanson Désirée van der Heijde Floris van Gaalen Francis Berenbaum Xavier Mariette Karine Briot Antoine Feydy Pascal Claudepierre Philippe Dieudé Joanne Nithitham Kimberly E. Taylor Lindsey A. Criswell Maxime Dougados Christian Roux Corinne Miceli-Richard 《PloS one》2015,10(8)