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1 The per os infectivity of ascovirus was improved using Bacillus thuringiensis var. kurstaki. 2 Generally, three types of competition were found between ascovirus and Btk. 3 Btk/ascovirus combinations had obvious oral infectivity of S. litura and M. separata larvae. 4 The results provided possible strategies on biocontrol of noctuid pests. This article is protected by copyright. All rights reserved
Huan Yu; Chang‐Jin Yang; Ni Li; Ying Zhao; Zhuang‐Mei Chen; Si‐Jia Yi; Zi‐Qi Li; Michael J. Adang; Guo‐Hua Huang. Novel strategies for the biocontrol of noctuid pests (Lepidoptera) based on improving ascovirus infectivity using Bacillus thuringiensis. Insect Science 2020, 1 .
AMA StyleHuan Yu, Chang‐Jin Yang, Ni Li, Ying Zhao, Zhuang‐Mei Chen, Si‐Jia Yi, Zi‐Qi Li, Michael J. Adang, Guo‐Hua Huang. Novel strategies for the biocontrol of noctuid pests (Lepidoptera) based on improving ascovirus infectivity using Bacillus thuringiensis. Insect Science. 2020; ():1.
Chicago/Turabian StyleHuan Yu; Chang‐Jin Yang; Ni Li; Ying Zhao; Zhuang‐Mei Chen; Si‐Jia Yi; Zi‐Qi Li; Michael J. Adang; Guo‐Hua Huang. 2020. "Novel strategies for the biocontrol of noctuid pests (Lepidoptera) based on improving ascovirus infectivity using Bacillus thuringiensis." Insect Science , no. : 1.
Bacterial insecticidal proteins, such as the Bin toxin from Lysinibacillus sphaericus, could be used more extensively to control insecticide resistant mosquitoes. This study was aimed at identification of mosquito cell proteins binding Bin toxin. Results showed that purified toxin was toxic to Anopheles gambiae larvae and Ag55 cultured cells. Clathrin heavy chain (an endocytosis protein) and glycolytic enzymes such as pyruvate kinase, enolase and dihydrolipoamide dehydrogenase were identified as binders of Bin toxin. The viability of Ag55 cells in the presence of endocytosis inhibitor, pitstop2, was significantly decreased upon Bin treatment, while the inhibitor chlorpromazine did not affect Bin toxicity. Bin toxin treatment decreased ATP production and mitochondrial respiration in Ag55 cells, whereas non-mitochondrial oxygen consumption significantly increased after Bin toxin treatment. These findings are steps towards understanding how Bin toxin kills mosquitoes. Mosquitoes are vectors of pathogens causing human diseases such as dengue fever, yellow fever, zika virus and malaria. An insecticidal toxin from Lysinibacillus sphaericus called Binary, or Bin, toxin could be used more extensively to control insecticide resistant mosquitoes. Bin toxin enter cells in susceptible mosquitoes and induces apoptosis or autophagy. In the current research, we used the malaria mosquito Anopheles gambiae Ag55 cell line as a model. A proteomic-based approach identified proteins that interact with Bin toxin. Interacting proteins include clathrin heavy chain (endocytosis protein) and glycolysis enzymes such as pyruvate kinase, enolase and dihydrolipoamide dehydrogenase. In Ag55 cell toxicity assays, an endocytosis inhibitor, pitstop2, increased Bin toxicity. Real time assays with a Seahorse™ flux analyzer showed that Bin significantly affects mitochondrial respiration, a result consistent with cell death via apoptosis or autophagy. These research findings add insights into how an unusual binary protein exploits cellular machinery to kill mosquitoes.
Muhammad Asam Riaz; Michael J. Adang; Gang Hua; Tatiana Maria Teodoro Rezende; Antonio Mauro Rezende; Guang-Mao Shen. Identification of Lysinibacillus sphaericus Binary toxin binding proteins in a malarial mosquito cell line by proteomics: A novel approach towards improving mosquito control. Journal of Proteomics 2020, 227, 103918 .
AMA StyleMuhammad Asam Riaz, Michael J. Adang, Gang Hua, Tatiana Maria Teodoro Rezende, Antonio Mauro Rezende, Guang-Mao Shen. Identification of Lysinibacillus sphaericus Binary toxin binding proteins in a malarial mosquito cell line by proteomics: A novel approach towards improving mosquito control. Journal of Proteomics. 2020; 227 ():103918.
Chicago/Turabian StyleMuhammad Asam Riaz; Michael J. Adang; Gang Hua; Tatiana Maria Teodoro Rezende; Antonio Mauro Rezende; Guang-Mao Shen. 2020. "Identification of Lysinibacillus sphaericus Binary toxin binding proteins in a malarial mosquito cell line by proteomics: A novel approach towards improving mosquito control." Journal of Proteomics 227, no. : 103918.
Bacillus thuringiensis (Bt) Cry1Fa and Cry1Ab proteins are important Cry toxins due to their high, selective toxicity against a number of lepidopteran species, including important pests of corn and cotton. Competition binding assays are a classical tool for investigating Cry toxin interactions with target pest insects. We developed a fluorescence-based binding assay and assessed Cry1Fa and Cry1Ab toxin binding to brush border membrane preparations from lepidopteran corn pests including Ostrinia nubilalis (European corn borer, ECB), Diatraea grandiosella (south western corn borer, SWCB), and Helicoverpa zea (corn earworm, CEW). Homologous and heterologous competition binding assays with fluorophore-(Alexa488)-labeled Cry1Fa toxin showed that Cry1Fa shares binding site(s) with Cry1Ab toxin in ECB, and SWCB for which Cry1Ab has higher affinity than Cry1Fa. Apart from the shared binding sites, Cry1Ab and Cry1Fa bind an additional site(s) in ECB and SWCB. In CEW, Cry1Fa and Cry1Ab each, has a high affinity binding site(s), which binds the heterologous toxin with low affinity. The Cry1Ab-Cry1Fa toxin binding models for ECB, SWCB and CEW based on our results are considered in the context of what is known about acquired cross-resistance against Cry1Ab and Cry1Fa toxins.
Reben Raeman; Gang Hua; Qi Zhang; Michael J. Adang. Fluorescent analyses of Bacillus thuringiensis Cry1Fa and Cry1Ab toxin binding sites on brush border membrane vesicles of Ostrinia nubilalis (Hübner), Diatraea grandiosella (Dyar), and Helicoverpa zea (Boddie) larvae. Pesticide Biochemistry and Physiology 2020, 167, 104592 .
AMA StyleReben Raeman, Gang Hua, Qi Zhang, Michael J. Adang. Fluorescent analyses of Bacillus thuringiensis Cry1Fa and Cry1Ab toxin binding sites on brush border membrane vesicles of Ostrinia nubilalis (Hübner), Diatraea grandiosella (Dyar), and Helicoverpa zea (Boddie) larvae. Pesticide Biochemistry and Physiology. 2020; 167 ():104592.
Chicago/Turabian StyleReben Raeman; Gang Hua; Qi Zhang; Michael J. Adang. 2020. "Fluorescent analyses of Bacillus thuringiensis Cry1Fa and Cry1Ab toxin binding sites on brush border membrane vesicles of Ostrinia nubilalis (Hübner), Diatraea grandiosella (Dyar), and Helicoverpa zea (Boddie) larvae." Pesticide Biochemistry and Physiology 167, no. : 104592.
Cry3Bb toxin from Bacillus thuringiensis is an important insecticidal protein due to its potency against coleopteran pests, especially rootworms. Cadherin, a protein in the insect midgut epithelium, is a receptor of Cry toxins; in some insect species toxin-binding domains of cadherins-synergized Cry toxicity. Previously, we reported that the DvCad1-CR8-10 fragment of Diabrotica virgifera virgifera cadherin-like protein (GenBank Accession #EF531715) enhanced Cry3Bb toxicity to the Colorado Potato Beetle (CPB), Leptinotarsa decimlineata (L. decimlineata). We report that individual CR domains of the DvCad1-CR8-10 fragment were found to have strong binding affinities to α-chymotrypsin-treated Cry3Bb. The dissociation constant (Kd) of Cry3Bb binding to the CR8, CR9, and CR10 domain was 4.9 nM, 28.2 nM, and 4.6 nM, respectively. CR8 and CR10, but not CR9, enhanced Cry3Bb toxicity against L. decimlineata and the lesser mealworm Alphitobius diaperinus neonates. In-frame deletions of the DvCad1-CR10 open reading frame defined a high-affinity binding and synergistic site to a motif in residues I1226–D1278. A 26 amino acid peptide from the high affinity Cry3Bb-binding region of CR10 functioned as a Cry3Bb synergist against coleopteran larvae.
Youngjin Park; Gang Hua; Suresh Ambati; Milton Taylor; Michael J. Adang. Binding and Synergizing Motif within Coleopteran Cadherin Enhances Cry3Bb Toxicity on the Colorado Potato Beetle and the Lesser Mealworm. Toxins 2019, 11, 386 .
AMA StyleYoungjin Park, Gang Hua, Suresh Ambati, Milton Taylor, Michael J. Adang. Binding and Synergizing Motif within Coleopteran Cadherin Enhances Cry3Bb Toxicity on the Colorado Potato Beetle and the Lesser Mealworm. Toxins. 2019; 11 (7):386.
Chicago/Turabian StyleYoungjin Park; Gang Hua; Suresh Ambati; Milton Taylor; Michael J. Adang. 2019. "Binding and Synergizing Motif within Coleopteran Cadherin Enhances Cry3Bb Toxicity on the Colorado Potato Beetle and the Lesser Mealworm." Toxins 11, no. 7: 386.
Global climate change and acquired resistance to insecticides are threats to world food security. Drosophila suzukii, a devastating invasive pest in many parts of the world, causes substantial economic losses to fruit production industries, forcing farmers to apply broad-spectrum insecticides frequently. This could lead to the development of insecticide resistance. We determined the Lethal Concentration 50 (median lethal concentration, LC50) values of zeta-cypermethrin, spinosad, and malathion insecticides against D. suzukii colonies established from Clarke and Pierce county Georgia, United States. The LC50 values were 3 fold higher in the Pierce county population for all insecticide treatments. We then used RNA sequencing to analyze the responses of Pierce and Clarke population flies surviving a LC50 treatment of the 3 insecticides. We identified a high number of differentially expressed genes that are likely involved in detoxification and reduced cuticular penetration, especially in the Pierce population, with extensive overlap in differentially expressed genes between the 3 insecticide treatments. Finally, we predicted fewer nonsynonymous single nucleotide variants having deleterious effects on protein function among detoxification, insecticide target, and cuticular protein encoding genes in Pierce flies. Thus a combination of increased gene expression and fewer deleterious single nucleotide variants highlights molecular mechanisms underlying the higher LC50 values for Pierce population flies.
Ruchir Mishra; Joanna C. Chiu; Gang Hua; Nilesh Tawari; Michael J. Adang; Ashfaq A. Sial. High throughput sequencing reveals Drosophila suzukii responses to insecticides. Insect Science 2017, 25, 928 -945.
AMA StyleRuchir Mishra, Joanna C. Chiu, Gang Hua, Nilesh Tawari, Michael J. Adang, Ashfaq A. Sial. High throughput sequencing reveals Drosophila suzukii responses to insecticides. Insect Science. 2017; 25 (6):928-945.
Chicago/Turabian StyleRuchir Mishra; Joanna C. Chiu; Gang Hua; Nilesh Tawari; Michael J. Adang; Ashfaq A. Sial. 2017. "High throughput sequencing reveals Drosophila suzukii responses to insecticides." Insect Science 25, no. 6: 928-945.
Bacillus thuringiensis is a Gram-positive aerobic bacterium that produces insecticidal crystalline inclusions during sporulation phases of the mother cell. The virulence factor, known as parasporal crystals, is composed of Cry and Cyt toxins. Most Cry toxins display a common 3-domain topology. Cry toxins exert intoxication through toxin activation, receptor binding and pore formation in a suitable larval gut environment. The mosquitocidal toxins of Bt subsp. israelensis (Bti) were found to be highly active against mosquito larvae and are widely used for vector control. Bt subsp. jegathesan is another strain which possesses high potency against broad range of mosquito larvae. The present review summarizes characterized receptors for Cry toxins in mosquito larvae, and will also discuss the diversity and effects of 3-D mosquitocidal Cry toxin and the ongoing research for Cry toxin mechanisms generated from investigations of lepidopteran and dipteran larvae.
Qi Zhang; Gang Hua; Michael J. Adang. Effects and mechanisms of Bacillus thuringiensis crystal toxins for mosquito larvae. Insect Science 2016, 24, 714 -729.
AMA StyleQi Zhang, Gang Hua, Michael J. Adang. Effects and mechanisms of Bacillus thuringiensis crystal toxins for mosquito larvae. Insect Science. 2016; 24 (5):714-729.
Chicago/Turabian StyleQi Zhang; Gang Hua; Michael J. Adang. 2016. "Effects and mechanisms of Bacillus thuringiensis crystal toxins for mosquito larvae." Insect Science 24, no. 5: 714-729.
Binary toxin (Bin) produced by Lysinibacillus sphaericus is toxic to Culex and Anopheles mosquito larvae. It has been used world-wide for control of mosquitoes that vector disease. The Bin toxin interacts with the glucosidase receptor, Cpm1, in Culex and its orthologue, Agm3, in Anopheles mosquitoes. However, the exact mechanism of its mode of action is not clearly understood. It is essential to understand mode of action of Bin toxin to circumvent the resistance that develops over generations of exposure. A suitable model cell line will facilitate investigations of the molecular action of Bin toxin. Here we report Bin toxin activity on Ag55 cell line that has been derived from an actual target, Anopheles gambiae larvae. The Bin toxin, both in pro and active forms, kills the Ag55 cells within 24 h. Bin toxin internalizes in Ag55 cells and also induces vacuolation as tracked by Lysotracker dye. The dose response studies showed that 1.5 nM of Bin toxin is sufficient to induce vacuolation and Ag55 cell death. Presence of α-glucosidase gene (Agm3) expression in the Ag55 cells was also confirmed. Thus, Ag55 cells constitute an appropriate model system to decipher the mode of Bin action in mosquito larvae.
Ramesh S. Hire; Gang Hua; Qi Zhang; Ruchir Mishra; Michael J. Adang. Anopheles gambiae Ag55 cell line as a model for Lysinibacillus sphaericus Bin toxin action. Journal of Invertebrate Pathology 2015, 132, 105 -110.
AMA StyleRamesh S. Hire, Gang Hua, Qi Zhang, Ruchir Mishra, Michael J. Adang. Anopheles gambiae Ag55 cell line as a model for Lysinibacillus sphaericus Bin toxin action. Journal of Invertebrate Pathology. 2015; 132 ():105-110.
Chicago/Turabian StyleRamesh S. Hire; Gang Hua; Qi Zhang; Ruchir Mishra; Michael J. Adang. 2015. "Anopheles gambiae Ag55 cell line as a model for Lysinibacillus sphaericus Bin toxin action." Journal of Invertebrate Pathology 132, no. : 105-110.
The Cry11Ba protein of Bacillus thuringiensis subsp. jegathesan crystals has uniquely high toxicity against a spectrum of mosquito species. The high potency of Cry11Ba against Anopheles gambiae is caused by recognition of multiple midgut proteins including glycosyl phosphatidylinositol-anchored alkaline phosphatase AgALP1, aminopeptidase AgAPN2, α-amylase AgAmy1 and α-glucosidase Agm3 that bind Cry11Ba with high affinity and function as putative receptors. The cadherin AgCad2 in An. gambiae larvae also binds Cry11Ba with high affinity (Kd = 12 nM) and is considered a putative receptor, while cadherin AgCad1 bound Cry11Ba with low affinity (Kd = 766 nM), a property not supportive for a Cry11Ba receptor role. Here, we show the in vivo involvement of AgCad1 in Cry11Ba toxicity in An. gambiae larvae using chitosan/DsiRNA nanoparticles to inhibit AgCad expression in larvae. Cry11Ba was significantly less toxic to AgCad1-silenced larvae than to control larvae. Because AgCad1 was co-suppressed by AgCad2 DsRNAi, the involvement of AgCad2 in Cry11Ba toxicity could not be ascertained. The ratio of AgCad1:AgCad2 transcript level is 36:1 for gut tissue in 4th instar larvae. Silencing AgCad expression had no effect on transcript levels of other binding receptors of Cry11Ba. We conclude that AgCad1 and possibly AgCad2 in An. gambiae larvae are functional receptors of Cry11Ba toxin in vivo.
Qi Zhang; Gang Hua; Michael J. Adang. Chitosan/DsiRNA nanoparticle targeting identifies AgCad1 cadherin in Anopheles gambiae larvae as an in vivo receptor of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan. Insect Biochemistry and Molecular Biology 2015, 60, 33 -38.
AMA StyleQi Zhang, Gang Hua, Michael J. Adang. Chitosan/DsiRNA nanoparticle targeting identifies AgCad1 cadherin in Anopheles gambiae larvae as an in vivo receptor of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan. Insect Biochemistry and Molecular Biology. 2015; 60 ():33-38.
Chicago/Turabian StyleQi Zhang; Gang Hua; Michael J. Adang. 2015. "Chitosan/DsiRNA nanoparticle targeting identifies AgCad1 cadherin in Anopheles gambiae larvae as an in vivo receptor of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan." Insect Biochemistry and Molecular Biology 60, no. : 33-38.
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.
Benjamin R. Deist; Michael A. Rausch; Maria Teresa Fernandez-Luna; Michael J. Adang; Bryony C. Bonning. Bt Toxin Modification for Enhanced Efficacy. Toxins 2014, 6, 3005 -3027.
AMA StyleBenjamin R. Deist, Michael A. Rausch, Maria Teresa Fernandez-Luna, Michael J. Adang, Bryony C. Bonning. Bt Toxin Modification for Enhanced Efficacy. Toxins. 2014; 6 (10):3005-3027.
Chicago/Turabian StyleBenjamin R. Deist; Michael A. Rausch; Maria Teresa Fernandez-Luna; Michael J. Adang; Bryony C. Bonning. 2014. "Bt Toxin Modification for Enhanced Efficacy." Toxins 6, no. 10: 3005-3027.
The lesser mealworm, Alphitobius diaperinus, is a serious cosmopolitan pest of commercial poultry facilities because of its involvement in structural damage to poultry houses, reduction in feed conversion efficiency, and transfer of avian and human pathogens. Cry3Aa, Cry3Bb, and Cry8Ca insecticidal proteins of Bacillus thuringiensis are used to control coleopteran larvae. Cadherins localized in the midgut epithelium function as receptors for Cry toxins in lepidopteran, coleopteran, and dipteran insects. Previously, we demonstrated that the truncated cadherin (DvCad1) from Diabrotica virgifera virgifera, which consists of the C-terminal cadherin repeats (CR) 8–10 and expressed in Escherichia coli, enhanced Cry3Aa and Cry3Bb toxicity against several coleopteran species. Here we report that the DvCad1-CR8–10 enhances Cry3Aa, Cry3Bb, and Cry8Ca toxicity to lesser mealworm. Previously, by an enzyme linked immunosorbent microplate assay, we demonstrated that the DvCad1-CR8–10 binds activated-Cry3Aa (11.8 nM), -Cry3Bb (1.4 nM), and now report that CR8–10 binds activated-Cry8Ca (5.7 nM) toxin. The extent of Cry toxins enhancement by DvCad1-CR8–10, which ranged from 3.30- to 5.93-fold, may have practical application for lesser mealworm control in preventing avian and human pathogen transfer in poultry facilities.
Youngjin Park; Gang Hua; Milton D. Taylor; Michael J. Adang. A coleopteran cadherin fragment synergizes toxicity of Bacillus thuringiensis toxins Cry3Aa, Cry3Bb, and Cry8Ca against lesser mealworm, Alphitobius diaperinus (Coleoptera: Tenebrionidae). Journal of Invertebrate Pathology 2014, 123, 1 -5.
AMA StyleYoungjin Park, Gang Hua, Milton D. Taylor, Michael J. Adang. A coleopteran cadherin fragment synergizes toxicity of Bacillus thuringiensis toxins Cry3Aa, Cry3Bb, and Cry8Ca against lesser mealworm, Alphitobius diaperinus (Coleoptera: Tenebrionidae). Journal of Invertebrate Pathology. 2014; 123 ():1-5.
Chicago/Turabian StyleYoungjin Park; Gang Hua; Milton D. Taylor; Michael J. Adang. 2014. "A coleopteran cadherin fragment synergizes toxicity of Bacillus thuringiensis toxins Cry3Aa, Cry3Bb, and Cry8Ca against lesser mealworm, Alphitobius diaperinus (Coleoptera: Tenebrionidae)." Journal of Invertebrate Pathology 123, no. : 1-5.
Bacillus thuringiensis (Bt) Cry proteins are used as components of biopesticides or expressed in transgenic crops to control diverse insect pests worldwide. These Cry toxins bind to receptors on the midgut brush border membrane and kill enterocytes culminating in larval mortality. Cadherin proteins have been identified as Cry toxin receptors in diverse lepidopteran, coleopteran, and dipteran species. In the present work we report a 185 kDa cadherin (AdCad1) from larvae of the lesser mealworm (Alphitobius diaperinus) larvae as the first identified receptor for Cry3Bb toxin. The AdCad1 protein contains typical structural components for Cry toxin receptor cadherins, including nine cadherin repeats (CR9), a membrane-proximal extracellular domain (MPED) and a cytosolic region. Peptides corresponding to the CR9 and MPED regions bound Cry3Bb toxin with high affinities (23 nM and 40 nM) and significantly synergized Cry3Bb toxicity against A. diperinus larvae. Silencing of AdCad1 expression through RNA interference resulted in highly reduced susceptibility to Cry3Bb in A. diperinus larvae. The CR9 peptide fed with toxin to RNAi-treated larvae restored Cry3Bb toxicity. These results are evidences that AdCad1 is a functional receptor of Cry3Bb toxin and that exogenously fed CR9 peptide can overcome the effect of reduced AdCad1expression on Cry3Bb toxicity to larvae.
Gang Hua; Youngjin Park; Michael J. Adang. Cadherin AdCad1 in Alphitobius diaperinus larvae is a receptor of Cry3Bb toxin from Bacillus thuringiensis. Insect Biochemistry and Molecular Biology 2013, 45, 11 -17.
AMA StyleGang Hua, Youngjin Park, Michael J. Adang. Cadherin AdCad1 in Alphitobius diaperinus larvae is a receptor of Cry3Bb toxin from Bacillus thuringiensis. Insect Biochemistry and Molecular Biology. 2013; 45 ():11-17.
Chicago/Turabian StyleGang Hua; Youngjin Park; Michael J. Adang. 2013. "Cadherin AdCad1 in Alphitobius diaperinus larvae is a receptor of Cry3Bb toxin from Bacillus thuringiensis." Insect Biochemistry and Molecular Biology 45, no. : 11-17.
Bacillus thuringiensis subsp. jegathesan produces Cry11Ba crystal protein with high toxicity to mosquito larvae. The Cry11Ba toxicity is dependent on its receptors on mosquito larval midgut epithelial cells. Previously, a cadherin-like protein (AgCad2), aminopeptidase (AgAPN2) and alkaline phosphatase (AgALP1) were reported to be involved in regulation of Cry11Ba toxicity on Anopheles gambiae larvae. Here, the cDNAs encoding α-amylase (AgAmy1) and α-glucosidase (Agm3) were cloned from A. gambiae larva midgut. Both are glycophosphatidylinositol (GPI) anchored proteins on brush border membranes (BBMV). Immunohistochemistry revealed their localization on different regions of the larval midgut. AgAmy1 and Agm3 bound Cry11Ba with high affinity, 37.6 nM and 21.1 nM respectively. Cry11Ba toxicity against A. gambiae larvae was neutralized by both AgAmy1 and Agm3. The results provide evidence that both AgAmy1 and Agm3 function as receptors of Cry11Ba in A. gambiae.
Qi Zhang; Gang Hua; Krishnareddy Bayyareddy; Michael J. Adang. Analyses of α-amylase and α-glucosidase in the malaria vector mosquito, Anopheles gambiae, as receptors of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan. Insect Biochemistry and Molecular Biology 2013, 43, 907 -915.
AMA StyleQi Zhang, Gang Hua, Krishnareddy Bayyareddy, Michael J. Adang. Analyses of α-amylase and α-glucosidase in the malaria vector mosquito, Anopheles gambiae, as receptors of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan. Insect Biochemistry and Molecular Biology. 2013; 43 (10):907-915.
Chicago/Turabian StyleQi Zhang; Gang Hua; Krishnareddy Bayyareddy; Michael J. Adang. 2013. "Analyses of α-amylase and α-glucosidase in the malaria vector mosquito, Anopheles gambiae, as receptors of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan." Insect Biochemistry and Molecular Biology 43, no. 10: 907-915.
In an effort to study the mode of action of Cry11Ba, we identified toxin binding proteins in Anopheles gambiae larval midgut and investigated their receptor roles. Previously, an aminopeptidase (AgAPN2) and an alkaline phosphatase (AgALP1) were identified as receptors for Cry11Ba toxin in A. gambiae. However, an A. gambiae cadherin (AgCad1) that bound Cry11Ba with low affinity (Kd = 766 nM) did not support a receptor role of AgCad1 for Cry11Ba. Here, we studied a second A. gambiae cadherin (AgCad2) that shares 14% identity to AgCad1. Immunohistochemical study showed that the protein is localized on A. gambiae larval midgut apical membranes. Its cDNA was cloned and the protein was analyzed as a transmembrane protein containing 14 cadherin repeats. An Escherichia coli expressed CR14MPED fragment of AgCad2 bound Cry11Ba with high affinity (Kd = 11.8 nM), blocked Cry11Ba binding to A. gambiae brush border vesicles and reduced Cry11Ba toxicity in bioassays. Its binding to Cry11Ba could be completely competed off by AgCad1, but only partially competed by AgALP1. The results are evidence that AgCad2 may function as a receptor for Cry11Ba in A. gambiae larvae.
Gang Hua; Qi Zhang; Rui Zhang; Amir M. Abdullah; Paul J. Linser; Michael J. Adang. AgCad2 cadherin in Anopheles gambiae larvae is a putative receptor of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan. Insect Biochemistry and Molecular Biology 2013, 43, 153 -161.
AMA StyleGang Hua, Qi Zhang, Rui Zhang, Amir M. Abdullah, Paul J. Linser, Michael J. Adang. AgCad2 cadherin in Anopheles gambiae larvae is a putative receptor of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan. Insect Biochemistry and Molecular Biology. 2013; 43 (2):153-161.
Chicago/Turabian StyleGang Hua; Qi Zhang; Rui Zhang; Amir M. Abdullah; Paul J. Linser; Michael J. Adang. 2013. "AgCad2 cadherin in Anopheles gambiae larvae is a putative receptor of Cry11Ba toxin of Bacillus thuringiensis subsp. jegathesan." Insect Biochemistry and Molecular Biology 43, no. 2: 153-161.
Lipid rafts are microdomains in the plasma membrane of eukaryotic cells. Among their many functions, lipid rafts are involved in cell toxicity caused by pore forming bacterial toxins including Bacillus thuringiensis (Bt) Cry toxins. We isolated lipid rafts from brush border membrane vesicles (BBMV) of Aedes aegypti larvae as a detergent resistant membrane (DRM) fraction on density gradients. Cholesterol, aminopeptidase (APN), alkaline phosphatase (ALP) and the raft marker flotillin were preferentially partitioned into the lipid raft fraction. When mosquitocidal Cry4Ba toxin was preincubated with BBMV, Cry4Ba localized to lipid rafts. A proteomic approach based on one-dimensional gel electrophoresis, in-gel trypsin digestion, followed by liquid chromatography–mass spectrometry (geLC–MS/MS) identified a total of 386 proteins. Of which many are typical lipid raft marker proteins including flotillins and glycosylphosphatidylinositol (GPI)-anchored proteins. Identified raft proteins were annotated in silico for functional and physicochemical characteristics. Parameters such as distribution of isoelectric point, molecular mass, and predicted post-translational modifications relevant to lipid raft proteins (GPI anchorage and myristoylation or palmitoylation) were analyzed for identified proteins in the DRM fraction. From a functional point of view, this study identified proteins implicated in Cry toxin interactions as well as membrane-associated proteins expressed in the mosquito midgut that have potential relevance to mosquito biology and vector management.
Krishnareddy Bayyareddy; Xiang Zhu; Ron Orlando; Michael J. Adang. Proteome Analysis of Cry4Ba Toxin-interacting Aedes aegypti Lipid Rafts using geLC–MS/MS. Journal of Proteome Research 2012, 11, 5843 -5855.
AMA StyleKrishnareddy Bayyareddy, Xiang Zhu, Ron Orlando, Michael J. Adang. Proteome Analysis of Cry4Ba Toxin-interacting Aedes aegypti Lipid Rafts using geLC–MS/MS. Journal of Proteome Research. 2012; 11 (12):5843-5855.
Chicago/Turabian StyleKrishnareddy Bayyareddy; Xiang Zhu; Ron Orlando; Michael J. Adang. 2012. "Proteome Analysis of Cry4Ba Toxin-interacting Aedes aegypti Lipid Rafts using geLC–MS/MS." Journal of Proteome Research 11, no. 12: 5843-5855.
Bacillus thuringiensis var. israelensis (Bti) is a natural larval mosquito pathogen producing pore-forming toxins targeting the midgut of Diptera larvae. It is used worldwide for mosquito control. Resistance mechanisms of an Aedes aegypti laboratory strain selected for 30 generations with field-collected leaf litter containing Bti toxins were investigated in larval midguts at two levels: 1. gene transcription using DNA microarray and RT-qPCR and 2. differential expression of brush border membrane proteins using DIGE (Differential In Gel Electrophoresis).
Guillaume Tetreau; Krishnareddy Bayyareddy; Christopher Jones; Renaud Stalinski; Muhammad A Riaz; Margot Paris; Jean-Philippe David; Michael J Adang; Laurence Després. Larval midgut modifications associated with Bti resistance in the yellow fever mosquito using proteomic and transcriptomic approaches. BMC Genomics 2012, 13, 248 -248.
AMA StyleGuillaume Tetreau, Krishnareddy Bayyareddy, Christopher Jones, Renaud Stalinski, Muhammad A Riaz, Margot Paris, Jean-Philippe David, Michael J Adang, Laurence Després. Larval midgut modifications associated with Bti resistance in the yellow fever mosquito using proteomic and transcriptomic approaches. BMC Genomics. 2012; 13 (1):248-248.
Chicago/Turabian StyleGuillaume Tetreau; Krishnareddy Bayyareddy; Christopher Jones; Renaud Stalinski; Muhammad A Riaz; Margot Paris; Jean-Philippe David; Michael J Adang; Laurence Després. 2012. "Larval midgut modifications associated with Bti resistance in the yellow fever mosquito using proteomic and transcriptomic approaches." BMC Genomics 13, no. 1: 248-248.
The Cry proteins produced byBacillus thuringiensis(Bt) are the most widely used biopesticides effective against a range of crop pests and disease vectors. Like chemical pesticides, development of resistance is the primary threat to the long-term efficacy of Bt toxins. Recently discovered cadherin-based Bt Cry synergists showed the potential to augment resistance management by improving efficacy of Cry toxins. However, the mode of action of Bt Cry synergists is thus far unclear. Here we elucidate the mechanism of cadherin-based Cry toxin synergism utilizing two cadherin peptides,Spodoptera frugiperdaCad (SfCad) andManduca sextaCad (MsCad), which differentially enhance Cry1Fa toxicity toSpodoptera frugiperdaneonates. We show that differential SfCad- and MsCad-mediated protection of Cry1Fa toxin in theSpodoptera frugiperdamidgut correlates with differential Cry1Fa toxicity enhancement. Both peptides exhibited high affinity for Cry1Fa toxin and an increased rate of Cry1Fa-induced pore formation inS. frugiperda. However, only SfCad bound theS. frugiperdabrush border membrane vesicle and more effectively prolonged the stability of Cry1Fa toxin in the gut, explaining higher Cry1Fa enhancement by this peptide. This study shows that cadherin fragments may enhanceB. thuringiensistoxicity by at least two different mechanisms or a combination thereof: (i) protection of Cry toxin from protease degradation in the insect midgut and (ii) enhancement of pore-forming ability of Cry toxin.
Khalidur Rahman; Mohd Amir F. Abdullah; Suresh Ambati; Milton D. Taylor; Michael J. Adang. Differential Protection of Cry1Fa Toxin against Spodoptera frugiperda Larval Gut Proteases by Cadherin Orthologs Correlates with Increased Synergism. Applied and Environmental Microbiology 2011, 78, 354 -362.
AMA StyleKhalidur Rahman, Mohd Amir F. Abdullah, Suresh Ambati, Milton D. Taylor, Michael J. Adang. Differential Protection of Cry1Fa Toxin against Spodoptera frugiperda Larval Gut Proteases by Cadherin Orthologs Correlates with Increased Synergism. Applied and Environmental Microbiology. 2011; 78 (2):354-362.
Chicago/Turabian StyleKhalidur Rahman; Mohd Amir F. Abdullah; Suresh Ambati; Milton D. Taylor; Michael J. Adang. 2011. "Differential Protection of Cry1Fa Toxin against Spodoptera frugiperda Larval Gut Proteases by Cadherin Orthologs Correlates with Increased Synergism." Applied and Environmental Microbiology 78, no. 2: 354-362.
Cry11Ba produced by Bacillus thuringiensis subsp. jegathesan is an active toxin for larvae of the mosquito Anopheles gambiae. A 106-kDa aminopeptidase N (APN), called AgAPN2, was previously identified as a Cry11Ba receptor in A. gambiae. A 70-kDa fragment of AgAPN2 expressed in Escherichia coli binds Cry11Ba with high affinity (K(d) = 6.4 nM) and inhibits Cry11Ba activity by 98% in bioassays [Zhang et al. (2008) Biochemistry 47, 11263-11272]. To identify regions involved in toxicity, we truncated the 70-kDa APN fragment into peptides of 28- and 30-kDa ta and tb, respectively, and tested their abilities to mediate toxicity and bind Cry11Ba. While AgAPN2ta reduced Cry11Ba toxicity by 85%, AgAPN2tb showed a significant enhancement effect on Cry11Ba toxicity. The purified peptides showed evidence of structural folding and bound the same site(s) on Cry11Ba with high affinity. The inhibitory AgAPN2ta blocked Cry11Ba binding to brush border membrane vesicles (BBMV) of A. gambiae whereas the toxicity enhancing AgAPN2tb increased Cry11Ba binding on BBMV. A deletion at the N-terminus ((336)S-P(420)) of AgAPN2ta significantly reduced AgAPN2ta binding to Cry11Ba and its inhibitory effect. Deletion of the central region ((676)I-W(760)) of AgAPN2tb eliminated its increased toxin binding and toxicity enhancement effect without affecting Cry11Ba binding. A "bridge" model is proposed for AgAPN2tb action whereby the peptide binds Cry11Ba and vectors it to sites on the larval midgut.
Rui Zhang; Gang Hua; Jeffrey L. Urbauer; Michael J. Adang. Synergistic and Inhibitory Effects of Aminopeptidase Peptides onBacillus thuringiensisCry11Ba Toxicity in the MosquitoAnopheles gambiae. Biochemistry 2010, 49, 8512 -8519.
AMA StyleRui Zhang, Gang Hua, Jeffrey L. Urbauer, Michael J. Adang. Synergistic and Inhibitory Effects of Aminopeptidase Peptides onBacillus thuringiensisCry11Ba Toxicity in the MosquitoAnopheles gambiae. Biochemistry. 2010; 49 (39):8512-8519.
Chicago/Turabian StyleRui Zhang; Gang Hua; Jeffrey L. Urbauer; Michael J. Adang. 2010. "Synergistic and Inhibitory Effects of Aminopeptidase Peptides onBacillus thuringiensisCry11Ba Toxicity in the MosquitoAnopheles gambiae." Biochemistry 49, no. 39: 8512-8519.
A peptide from cadherin AgCad1 of Anopheles gambiae larvae was reported as a synergist of Bacillus thuringiensis subsp. israelensis Cry4Ba's toxicity to the Anopheles mosquito (G. Hua, R. Zhang, M. A. Abdullah, and M. J. Adang, Biochemistry 47:5101-5110, 2008). We report that CR11 to the membrane proximal extracellular domain (MPED) (CR11-MPED) and a longer peptide, CR9 to CR11 (CR9-11), from AgCad1 act as synergists of Cry4Ba's toxicity to Aedes aegypti larvae, but a Diabrotica virgifera virgifera cadherin-based synergist of Cry3 (Y. Park, M. A. F. Abdullah, M. D. Taylor, K. Rahman, and M. J. Adang, Appl. Environ. Microbiol. 75:3086-3092, 2009) did not affect Cry4Ba's toxicity. Peptides CR9-11 and CR11-MPED bound Cry4Ba with high affinity (13 nM and 23 nM, respectively) and inhibited Cry4Ba binding to the larval A. aegypti brush border membrane. The longer CR9-11 fragment was more potent than CR11-MPED in enhancing Cry4Ba against A. aegypti .
Youngjin Park; Gang Hua; Mohd Amir F. Abdullah; Khalidur Rahman; Michael J. Adang. Cadherin Fragments from Anopheles gambiae Synergize Bacillus thuringiensis Cry4Ba's Toxicity against Aedes aegypti Larvae. Applied and Environmental Microbiology 2009, 75, 7280 -7282.
AMA StyleYoungjin Park, Gang Hua, Mohd Amir F. Abdullah, Khalidur Rahman, Michael J. Adang. Cadherin Fragments from Anopheles gambiae Synergize Bacillus thuringiensis Cry4Ba's Toxicity against Aedes aegypti Larvae. Applied and Environmental Microbiology. 2009; 75 (22):7280-7282.
Chicago/Turabian StyleYoungjin Park; Gang Hua; Mohd Amir F. Abdullah; Khalidur Rahman; Michael J. Adang. 2009. "Cadherin Fragments from Anopheles gambiae Synergize Bacillus thuringiensis Cry4Ba's Toxicity against Aedes aegypti Larvae." Applied and Environmental Microbiology 75, no. 22: 7280-7282.
Specific Bacillus thuringiensis Berliner (Bt) toxins are effective against a narrow spectrum of species. While specificity is an advantage for limiting adverse effects on non-target organisms, it is also the primary drawback of Bt's application for controlling multiple pest species in agriculture, forestry and other areas. Recently, it was reported that a small toxin-binding fragment of Manduca sexta (Joh.) cadherin acts as a synergist of Bt toxins to M. sexta, Heliothis virescens F. and Helicoverpa zea (Boddie). These insects are quite susceptible to the Cry1A toxins. The first aim of the present study was to determine if longer-sized fragments of M. sexta cadherin differed in the level of toxin enhancement. The second aim was to examine enhancement of Bt toxins against relatively Bt-tolerant insects Agrotis ipsilon (Hufn.) and Spodoptera exigua (Hübner). Cadherin fragments longer than previously reported had improved synergistic properties. Significant enhancement of Bt Cry1A toxins against A. ipsilon and S. exigua was found. A cadherin fragment also increased Cry1C toxicity to S. exigua. The commercial development of this synergist has the potential to widen the spectrum of Bt toxicity to other important agricultural lepidopteran insect pests and thus increase its usefulness in agriculture.
Mohd Amir F Abdullah; Saad Moussa; Milton D Taylor; Michael J Adang. Manduca sexta(Lepidoptera: Sphingidae) cadherin fragments function as synergists for Cry1A and Cry1CBacillus thuringiensistoxins against noctuid mothsHelicoverpa zea, Agrotis ipsilonandSpodoptera exigua. Pest Management Science 2009, 65, 1097 -1103.
AMA StyleMohd Amir F Abdullah, Saad Moussa, Milton D Taylor, Michael J Adang. Manduca sexta(Lepidoptera: Sphingidae) cadherin fragments function as synergists for Cry1A and Cry1CBacillus thuringiensistoxins against noctuid mothsHelicoverpa zea, Agrotis ipsilonandSpodoptera exigua. Pest Management Science. 2009; 65 (10):1097-1103.
Chicago/Turabian StyleMohd Amir F Abdullah; Saad Moussa; Milton D Taylor; Michael J Adang. 2009. "Manduca sexta(Lepidoptera: Sphingidae) cadherin fragments function as synergists for Cry1A and Cry1CBacillus thuringiensistoxins against noctuid mothsHelicoverpa zea, Agrotis ipsilonandSpodoptera exigua." Pest Management Science 65, no. 10: 1097-1103.
Alkaline phosphatases (ALPs, EC 3.1.3.1) isolated from lepidopteran and dipteran species are identified as receptors for Cry1Ac and Cry11Aa toxins, respectively [Jurat-Fuentes, J. L., and Adang, M. J. (2004) Eur. J. Biochem. 7, 3127-3135; Fernandez, L. E., et al. (2006) Biochem. J. 396, 77-84]. In our study, an alkaline phosphatase cDNA (AgALP1) was cloned from the midgut of Anopheles gambiae larvae. The encoded 63 kDa protein has a predicted glycosylphosphatidylinositol (GPI) anchor omega-site ((526)Asp), an N-glycosylation site ((239)Asn-Leu-Thr), and an O-glycosylation site ((312)Ser). AgALP1(t) was expressed in Escherichia coli and used to prepare antiserum and to analyze the interaction of AgALP with mosquitocidal Cry11Ba toxin. Anti-AgALP serum localized AgALP to the apical brush border in the anterior and posterior midgut of larvae and detected a 65 kDa species on a blot of brush border membrane vesicles (BBMVs) protein prepared from larvae. ALP activity was released from larval BBMVs prepared by phosphatidylinositol-specific phospholipase C (PIPLC) treatment, and after separation by two-dimensional gel electrophoresis and blotting, a chain of doublet spots at 65 kDa was detected by anti-AgALP. A subset of these doublet spots bound Cry11Ba on a reprobed blot. Heterologously expressed AgALP1(t) bound [(125)I]Cry11Ba on dot blots and reduced the level of binding of [(125)I]Cry11Ba to brush border membrane vesicles by 41%, a percentage comparable to that of unlabeled Cry11Ba and aminopeptidase AgAPN2(t1) peptide. AgALP1(t) binds Cry11Ba toxin with a high affinity (23.9 nM) and shares a binding site on Cry11Ba with AgAPN2(t1). In bioassays against An. gambiae larvae, the presence of AgALP1(t) reduced larval mortality from 78 to 8%. We conclude that AgALP1 is a binding protein and a functional receptor for Cry11Ba toxin.
Gang Hua; Rui Zhang; Krishnareddy Bayyareddy; Michael J. Adang. Anopheles gambiae Alkaline Phosphatase Is a Functional Receptor of Bacillus thuringiensis jegathesan Cry11Ba Toxin. Biochemistry 2009, 48, 9785 -9793.
AMA StyleGang Hua, Rui Zhang, Krishnareddy Bayyareddy, Michael J. Adang. Anopheles gambiae Alkaline Phosphatase Is a Functional Receptor of Bacillus thuringiensis jegathesan Cry11Ba Toxin. Biochemistry. 2009; 48 (41):9785-9793.
Chicago/Turabian StyleGang Hua; Rui Zhang; Krishnareddy Bayyareddy; Michael J. Adang. 2009. "Anopheles gambiae Alkaline Phosphatase Is a Functional Receptor of Bacillus thuringiensis jegathesan Cry11Ba Toxin." Biochemistry 48, no. 41: 9785-9793.