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Larvicidal efficacy of Catharanthus roseus Linn.(Family:Apocynaceae)leaf extract and bacterial insecticide Bacillus thuringiensis against Anopheles stephensi Liston.

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Asian Pacific Journal of Tropical Medicine 2013年第11期6卷 847-853页

作者： Chellasamy Panneerselvam Kadarkarai Murugan Kalimuthu Kovendan Palanisamy Mahesh Kumar Sekar Ponarulselvam Duraisamy Amerasan Jayapal Subramaniam Jiang-Shiou Hwang DRDO-BU Center for Life Sciences Bharathiar UniversityCoimhatore 641046Tamil NaduIndia Division of Entomology Department of ZoologySchool of Life SciencesBharathiar UniversityCoimbatore 641046Tamil NaduIndia Institute of Marine Biology National Taiwan Ocean UniversityKeelung 202-24Taiwan

Objective:To explore the larvicidal activity of Catharanthus roseus(C.roseus) leaf extract and Raeillus thuringiensis(B.thuringiensis) against the malarial vector Anopheles stephensi(An. stephensi),when being used alone or together.Methods:The larvicidal activity was assayed at various concentrations under the laboratory and field conditions.The LC_(50) and LC_(90) values of the C.roseus leaf extract were determined by probit analysis.Results:The plant extract showed larvicidal effects after 24 h of exposure;however,the highest larval mortality was found in the petroleum ether extract of C.roseus against the first to fourth instars larvae with LC_(50)=3.34,4.48, 5.90 and 8.17 g/L,respectively;B.thuringiensis against the first to fourth instars larvae with LC_(50)=1.***.1.93.2.17 and 2.42 g/L.respectively:and the combined treatment with LC_(50)=2.***.2.41. 2.76 and 3.22 g/L,respectively.No mortality was observed in the control.Conclusions:The petroleum ether extract of C.roseus extract and B.thuringiensis have potential to be used as ideal eco-friendly agents for the control of An.stephensi in vector control programs.The combined treatment with this plant crude extract and bacterial toxin has better larvicidal efficacy against An.stephensi.

关键词： Catharanthus roseus Bacillus thuringiensis Anopheles stephensi Larvicidal activity

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Does cotton bollworm show cross-resistance to the Bacillus thuringiensis toxins Cry1Ac and Cry2Ab? A mini review

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Journal of Arid Land 2020年第2期12卷 349-356页

作者： MA Jihong TIAN Changyan LYU Guanghui MAI Wenxuan State Key Laboratory of Desert and Oasis Ecology Xinjiang Institute of Ecology and GeographyChinese Academy of SciencesUrumqi 830011China University of Chinese Academy of Sciences Beijing 100049China College of Resources and Environment Science Xinjiang UniversityUrumqi 830046China

Since 1996, transgenic Bacillus thuringiensis(Bt) cotton has been commercially grown in numerous countries in an effort to stem the losses caused by key lepidopteran pests. However, the development of pest resistance to Bt toxins has jeopardized the continued utilization of Bt cotton. As a strategy designed to circumvent the development of resistance, Bt cotton varieties expressing two or more toxins targeting the same pest have been introduced. Nevertheless, from the perspective of long-term planting of Bt cotton, the potential risk of cross-resistance to these Bt toxins is a threat that cannot be ignored. In this paper, we review current research(including that based on the analysis of protein binding sites and resistance genes) on the resistance of cotton bollworm(Helicoverpa armigera) to the Bt toxins Cry1 Ac and Cry2 Ab and the interrelationship between these toxins. On the basis of existing evidence, we assume that the actions of Cry1 Ac and Cry2 Ab against cotton bollworm are not completely independent, and then propose the "resistance-associated gene mutation potential hypothesis". Although the mechanisms underlying the resistance of pests to Bt toxins are yet to be comprehensively elucidated, this hypothesis could undoubtedly have important implications for adopting "pyramid" strategy in the future. Further research is recommended to devise strategies to retard the development of H. armigera resistance to Bt cotton, either using different Bt toxins or their various combinations.

关键词： cross-resistance genetically modified cotton cotton bollworm Bacillus thuringiensis Cry1Ac Cry2Ab

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MOLECULAR CLONING AND EXPRESSION OF Bacillus thuringiensis subsp. galleriae INSECTICIDAL CRYSTAL PROTEIN GENES IN Escherichia coli

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Science China Chemistry 1989年第7期32卷 830-836页

作者：陈骐范云六 Laboratory of Molecular Biology Chinese Academy of Agricultural Sciences Beijing

The location of the toxin gene of B. thuringiensis subsp. galleriae (H5ab) on the Mr-130Mdplasmid is determined by molecular cloning. Double digestion fragments (BamHⅠ and SalⅠ)and PstⅠ restriction fragments as well, from the 130 Md plasmid of B. thuringiensis subsp.galleriae, are ligated with the cloning vector pAT 153 respectively and transformed into E.coli strain HB 101. Out of 208 transformants, three colonies (FG2, FG9, FG19) give posi-tive hybridization reaction using the HD-1 delta-endotoxin gene as a probe. They are presum-ed to contain the delta-endotoxin gene of B. thuringiensis subsp. galleriae. Western bolt assaysindicate that Mr-130 kDal and 68 kDal, crystal proteins produced by clone FG 2 react withanticrystal protein antibody. The protein extracts of clone FG2 are lethal to Ostrinia furna-calis (Guenee). This is the first report with regard to the cloning and expression of the B. thuringiensissubsp. galleriae (H5ab) delta-endotoxin gene.

关键词： Bacillus thuringiensis subsp galleriae (H5ab) delta-endotoxin gene cloning and expression gene location

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

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Asian Pacific Journal of Tropical Biomedicine 2013年第2期3卷 111-117页

作者： Zulfaidah Penata Gama Nobukazu Nakagoshi Suharjono Faridah Setyowati Graduate School for International Development(IDEC) Hiroshima University Department of Biology Faculty of ScienceBrawijaya University

Objective:To investigate the toxicity of indigenous Bacillus thuringiensis（B.thuringiensis）isolates from Malang City for controlling Aedes aegypti（Ae.aegypti）larvae.Methods:Soil samples were taken from Purwantoro and Sawojajar sub-districts.Bacterial isolation was performed using B.thuringiensis selective media.Phenotypic characteristics of the isolates were obtained with the simple matching method.The growth and prevalence of spores were determined by the Total Plate Count method,and toxicity tests were also performed on the third instar larval stage of Ae.aegypti.The percentage of larval mortality was analysed using probit regression.The LC50was analysed by ANOVA,and the Tukey HSD interval was 95%.Results:Among the 33 selected bacterial isolates,six were obtained（PWR4-31,PWR4-32,SWJ4-2b,SWJ4-4b,SWJ-4k and SWJ5-1）that had a similar phenotype to reference B.thuringiensis.Based on the dendrogram,all of the bacterial isolates were 71%similar.Three isolates that had a higher prevalence of reference B.thuringiensis were PWR4-32,SWJ4-4b and SW5-1,of which the spore prevalence was 52.44%,23.59%,34.46%,respectively.These three indigenous isolates from Malang City successfully killed Ae.aegypti larvae.The PWR4-32 isolates were the most effective at killing the larvae.Conclusions:Six indigenous B.thuringiensis isolates among the 33 bacterial isolates found in the Sawojajar and Purwantoro sub-districts were toxic to the third instar larvae of Ae.aegypti.The PWR4-32 isolates were identical to tbe reference B.thuringiensis and had 88%phenotype similarity.The PWR4-32 isolates had the highest spore prevalence（52.44%）,and the early stationary phase occurred at 36 h.The PWR4-32 isolates were the most effective at killing Ae.aegypti larvae(LC50-72 h=2.3×108 cells/mL).

关键词： Aedes aegypti Bacillus thuringiensis LC50 Toxicological studies

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Construction of polyhedrin-positive recombinant viruswith expression of truncated δ-endotoxin fromBacillus thuringiensis in insect cell

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Chinese Science Bulletin 1996年第7期41卷 597-603页

作者：王福山黄永秀齐义鹏刘子夜杨远征 Institute of Virology Wuhan University Wuhan 430072 China

Baculoviruses have been widely used as biological agents because of their specificpathogenicity for target insect and harmlessness to mammals, birds and plants as well asadvantages with persistence and epidemics as pestcides. A major drawback for morewide-spread use of these viruses is their low virulenee and slow speed of action, which re-stricted their use. No enhancement in pathogenicity of recombinant viruses was observedwith insertion of the Bacillus thuringiensis full-length endotoxin cryIA(c) and cryIA(b) geneinto the AcNOV (Autographa californica nuclear polyhedrosis virus) genome under the con-trol of polyhedrin gene promoter. The reason may be that protoxin, full-length genesproduct of recombinant virus in infected insect cells, which was short of insect gut alkalienvironment, cannot be degraded into active toxic polypeptide. Expression level of 3’

关键词： Bacillus thuringiensis δ-endotoxin polyhedrin gene expression recombinant virus.

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Expression of Bacillus thuringiensis δ-Endotoxin Gene With Recombinant Baculovirus in Insect Cell

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Chinese Science Bulletin 1994年第5期39卷 424-429页

作者：齐义鹏黄永秀沈英谭业平王福山 Virology Institute of Wuhan University Wuhan 430072 PRC

Insect baculovirus(IBV) is a biological insecticide in agriculture and forestry, butits slow speed of killing pests and narrow range of host make its application quitelimited. Hence, polyhedrin (ocu) gene of IBV was c... 详细信息

关键词： bacillus thuringiensis cryIA(c) gene gene expression baculovirus transfer vector recombinant virus cotransfection.

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EXPRESSION OF THE Bacillus thuringiensis SUBSP, KURSTAKI HD-1 δ-ENDOTOXIN GENE IN Escherichia coli

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Chinese Science Bulletin 1992年第2期37卷 146-149页

作者：陈骐范云六 Biotechnology Research Center Chinese Academy of Agricultural Sciences Beijing 100081 PRC

In 1987, Vaeck and Fischhoff reported the transformation of the B. thuringiensis (abbreviated to B. t. ) δ-endotoxin gene into tobacco and tomato respectively, and obtained the transgenic plants. Since then, more att... 详细信息

关键词： Bacillus thuringiensis δ-endotoxin gene expression.

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Insilico structural analysis of parasporin 2 protein sequences of non-toxic bacillus thuringiensis

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Journal of Biomedical Science and Engineering 2010年第4期3卷 415-421页

作者： Ayyasamy Mahalakshmi Rajaiah Shenbagarathai 不详

The unusual and remarkable property of parasporin 2 of non-insecticidal Bacillus thuringiensis is specifically recognizing and selectively targeting human leukemic cell lines. The 37-kDa inactive nascent protein is proteolytically cleaved to the 30-kDa active form that loses both the N-terminal and the C-terminal segments. Accumulated cytological and biochemical observations on parasporin-2 imply that the protein is a pore-forming toxin. To confirm the hypothesis, insilico analysis was performed using homology modeling. The resulting model of parasporin 2 protein is unusually elongated and mainly comprises long β-strands aligned with its long axis. It is similar to aerolysin-type β-pore-forming toxins, which strongly reinforce the pore-forming hypothesis. The molecule can be divided into three domains. Domain 1, comprising a small β-sheet sandwiched by short α-helices, is probably the target-binding module. Two other domains are both β-sandwiches and thought to be involved in oligomerization and pore formation. Domain 2 has a putative channel-forming β-hairpin characteristic of aerolysin-type toxins. The surface of the protein has an extensive track of exposed side chains of serine and threonine residues. The track might orient the molecule on the cell membrane when domain 1 binds to the target until oligomerization and pore formation are initiated. The β-hairpin has such a tight structure that it seems unlikely to reform as postulated in a recent model of pore formation developed for aerolysin-type toxins. Parasporin 2 (Accession no: BAD35170) protein sequence analysis indicated two different domains namely, aerolysin toxin and clostridium toxin domain based on different database searches (CDD and Pfam). It showed a close similarity with the available PDB template (PDB id: 2ZTB) of parasporin which has cytocidal activity against MOLT-4, HL60 and Jurkat cell lines. Based on the PSI Blast analysis, 3D structures of the domains were predicted by using Swiss model server. Accuracy of the prediction of 3D structure o

关键词： Parasporin Homology Model Non-Toxic Bacillus thuringiensis Cell Lines

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Cloning of a New Truncated cry1AcGene from an Indian Isolate of Bacillus thuringiensis

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Advances in Microbiology 2014年第1期4卷 55-62页

作者： A. Ramalakshmi R. Manikandan V. Balasubramani V. Udayasuriyan Department of Agricultural Microbiology Tamil Nadu Agricultural University Coimbatore India Department of Plant Biotechnology Tamil Nadu Agricultural University Coimbatore India

Transgenic Bt crops producing insecticidal crystal proteins from Bacillus thuringiensis (Bt), so-called Cry toxins, have proved useful in controlling insect pests. Among the cry toxins, Cry1A toxins are important because of high toxicity to lepidopteran pests and their widespread distribution among Bt strains. In Cry1A proteins, toxin fragment is comprised of about 620 amino acids of N-terminal region and C-terminal half is not required for toxicity. Four indigenous isolates of Bt viz., T15, T16, T20 and T31 were screened by PCR-RFLP for 3’-truncated cry1A gene(s) corresponding to toxin fragment. RFLP analysis of cry1A amplicons obtained from the four isolates of Bt showed presence of cry1Ac-type gene alone in three isolates. One of the cry1Ac-postive isolates, T15 which showed 100 percent mortality in Helicoverpa armigera, was selected for cloning of DNA fragment of about 2.1 kb containing 3’-truncated cry1Ac gene. Nucleotide sequence data generated for 3’-truncated cry1Ac gene of T15 showed 98 to 99 percent homology with 1958 bp of already reported sequences of all cry1Ac genes (cry1Ac1 to cry1Ac24). Deduced amino acid sequence of cry1Ac of Bt strain, T15 showed one to four percent variation in comparison to all reported Cry1Ac holotypes (Cry1Ac1 to Cry1Ac24) by differing at 5 to 19 positions. This suggests that the cry1Ac toxin of Bt isolate, T15 is a new kind of its group.

关键词： Bacillus thuringiensis cry1Ac Cloning

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Solid-State Fermentation for the Concomitant Production of δ-Endotoxin and Endospore from Bacillus thuringiensissubsp. kurstaki

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Advances in Bioscience and Biotechnology 2014年第10期5卷 797-804页

作者： Veloorvalappil Narayanan Jisha Sailas Benjamin Enzyme Technology Laboratory Biotechnology Division Department of Botany University of Calicut Kerala India

Water stress and limited aeration imparted by solid-state fermentation (SSF) were reported as crucial factors for the enhancement of endospore production by Bacillus thuringiensis (Bt);and thus, more δ-endotoxin could be produced concomitantly with reduced time. Therefore, Bt subsp. kurstaki (Btk) was employed in the present study to evaluate its efficiency for the concomitant production of endospores and δ-endotoxin in LB medium supplemented with various naturally available agricultural products, i.e., flours of soybean, Bengal gram or jack seed at various concentrations (10%, 20%, 30%, 40%, 50%, 60%, 80% or 100%, all w/v). After 12 h fermentation, the supernatant in it was centrifuged off aseptically to obtain solid substrate for subsequent SSF. Of them, soybean (30%) supplemented medium was the best for the enhanced production of endospore and δ-crystals. The maximum yield of endospores during solid-state fermentation was observed 48 h, i.e., compared to submerged fermentation in LB, it was 24 h less gestation period. In control sample, the endospores achieved the maximum length (1.10 ± 0.13 μm) and diameter (0.63 ± 0.07 μm) at 72 h;while in soybean supplemented medium, the maximum length (2.10 ± 0.16 μm) and diameter (1.63 ± 0.16 μm) were at 48 h and 72 h, respectively. Upon staining, acridine orange specifically stained the endospores;malachite green-saffranin stained both δ-crystals and endospores;and coomassie brilliant blue specifically stained δ-endotoxin. Briefly, normal gestation period or harvest time for Btk is 72 h, which could be reduced to 48 h, if SSF is employed as demonstrated in this study.

关键词： Bacillus thuringiensis subsp. kurstaki Spore Solid-State Fermentation δ-Endotoxin

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