Highly Effective Broad Spectrum Chimeric Larvicide That Targets Vector Mosquitoes Using a Lipophilic Protein

Bideshi, Dennis K.; Park, Hyun-Woo; Hice, Robert H.; Wirth, Margaret C.; Federici, Brian A.

doi:10.1038/s41598-017-11717-9

Cited by 20 publications

(15 citation statements)

References 33 publications

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“…We also show that the mixture of BinA subunit with Cyt1Aa was enough to induce toxicity in resistant larvae and for the internalization of BinA in the midgut cells. These data are consistent with the larvicidal activity previously recorded for a recombinant chimeric protein formed by BinA and Cyt1Aa against Bin-resistant C. quinquefasciatus and A. aegypti larvae (66). Our results indicate that BinA entry into the cytoplasm, which could be likely due to the pore formation of Cyt1Aa, is sufficient to provoke mortality and therefore to overcome the failures related to the BinB receptor-specific binding step (10,13,65,67).…”

Section: Discussionsupporting

confidence: 92%

Functional Bacillus thuringiensis Cyt1Aa Is Necessary To Synergize Lysinibacillus sphaericus Binary Toxin (Bin) against Bin-Resistant and -Refractory Mosquito Species

Nascimento

Torres-Quintero

Molina

et al. 2020

Appl Environ Microbiol

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The binary (Bin) toxin from Lysinibacillus sphaericus is effective to mosquito larvae, but its utilization is threatened by the development of insect resistance. Bin toxin is composed of the BinB subunit required for binding to midgut receptors and the BinA subunit that causes toxicity after cell internalization, mediated by BinB. Culex quinquefasciatus resistance to this toxin is caused by mutations that prevent expression of Bin toxin receptors in the midgut. Previously, it was shown that the Cyt1Aa toxin from Bacillus thuringiensis subsp. israelensis restores Bin toxicity to Bin-resistant C. quinquefasciatus and to Aedes aegypti larvae, which are naturally devoid of functional Bin receptors. Our goal was to elucidate the mechanism involved in Cyt1Aa synergism with Bin in such larvae. In vivo assays showed that the mixture of Bin toxin, or its BinA subunit, with Cyt1Aa was effective to kill resistant larvae. However, no specific binding interaction between Cyt1Aa and the Bin toxin, or its subunits, was observed. The synergy between Cyt1Aa and Bin toxins is dependent on functional Cyt1Aa, as demonstrated by using the nontoxic Cyt1AaV122E mutant toxin affected in oligomerization and membrane insertion, which was unable to synergize Bin toxicity in resistant larvae. The synergism correlated with the internalization of Bin or BinA into anterior and medium midgut epithelial cells, which occurred only in larvae treated with wild-type Cyt1Aa toxin. This toxin is able to overcome failures in the binding step involving BinB receptor by allowing the internalization of Bin toxin, or its BinA subunit, into the midgut cells. IMPORTANCE One promising management strategy for mosquito control is the utilization of a mixture of L. sphaericus and B. thuringiensis subsp. israelensis insecticidal toxins. From this set, Bin and Cyt1Aa toxins synergize and display toxicity to resistant C. quinquefasciatus and to A. aegypti larvae, whose midgut cells lack Bin toxin receptors. Our data set provides evidence that functional Cyt1Aa is essential for internalization of Bin or its BinA subunit into such cells, but binding interaction between Bin and Cyt1Aa is not observed. Thus, this mechanism contrasts with that for the synergy between Cyt1Aa and the B. thuringiensis subsp. israelensis Cry toxins, where active Cyt1Aa is not necessary but a specific binding between Cry and Cyt1Aa is required. Our study established the initial molecular basis of the synergy between Bin and Cyt1Aa, and these findings enlarge our knowledge of their mode of action, which could help to develop improved strategies to cope with insect resistance.

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Section: Discussionsupporting

confidence: 92%

Functional Bacillus thuringiensis Cyt1Aa Is Necessary To Synergize Lysinibacillus sphaericus Binary Toxin (Bin) against Bin-Resistant and -Refractory Mosquito Species

Nascimento

Torres-Quintero

Molina

et al. 2020

Appl Environ Microbiol

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“…Here we show, for the first time, that exchange of loops involved in insect specificity among toxins that are not related structurally is feasible and could provide tools for changing insect specificity and potentially for providing tools for countering resistance. A recent publication showed that Cyt1Aa-BinA hybrid toxins can be constructed resulting in hybrid-toxins with a broad-spectrum against mosquitoes including Anopheles gambiae, C. quinquefasciatus and A. aegypti 32 . These data indicate that Cyt1Aa toxin may accept larger domains and produce active proteins, suggesting that in the future other Cyt1Aa-hybrids could be constructed containing entire domains that could effectively enhance a lethal effect by Cyt1Aa in other targets.…”

Section: Discussionmentioning

confidence: 99%

“…Cyt1Aa, Cyt1Aa-hybrid, Cry11Aa, Cry1Ab or Cry1Ab-G439D protoxins were produced in B. thuringiensis 407 − acrystalliferous strain transformed with wild type or mutated plasmids. Plasmid pWF45 has cyt1Aa gene 32 or the new constructions of cyt1Aa-hybrid genes. Plasmid pCG6 contains cry11Aa gene 35 .…”

Section: Methodsmentioning

confidence: 99%

Engineering Bacillus thuringiensis Cyt1Aa toxin specificity from dipteran to lepidopteran toxicity

Torres-Quintero

Gómez

Pacheco

et al. 2018

Sci Rep

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The Cyt and Cry toxins are different pore-forming proteins produced by Bacillus thuringiensis bacteria, and used in insect-pests control. Cry-toxins have a complex mechanism involving interaction with several proteins in the insect gut such as aminopeptidase N (APN), alkaline phosphatase (ALP) and cadherin (CAD). It was shown that the loop regions of domain II of Cry toxins participate in receptor binding. Cyt-toxins are dipteran specific and interact with membrane lipids. We show that Cry1Ab domain II loop3 is involved in binding to APN, ALP and CAD receptors since point mutation Cry1Ab-G439D affected binding to these proteins. We hypothesized that construction of Cyt1A-hybrid proteins providing a binding site that recognizes gut proteins in lepidopteran larvae could result in improved Cyt1Aa toxin toward lepidopteran larvae. We constructed hybrid Cyt1Aa-loop3 proteins with increased binding interaction to Manduca sexta receptors and increased toxicity against two Lepidopteran pests, M. sexta and Plutella xylostella. The hybrid Cyt1Aa-loop3 proteins were severely affected in mosquitocidal activity and showed partial hemolytic activity but retained their capacity to synergize Cry11Aa toxicity against mosquitos. Our data show that insect specificity of Cyt1Aa toxin can be modified by introduction of loop regions from another non-related toxin with different insect specificity.

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“…Of note, several of these recombinant/modified toxins have been introduced in genetically modified (GM) plants directly producing the toxins, thereby circumventing several limitations of topical insecticide applications (e.g., limited persistence, difficult timing of applications, and higher production costs) [ 53 ]. Cyt1Aa toxin from Bti has also been used to generate chimeras, expanding its host spectrum to other mosquito species when fused to the binary Tpp1Aa/2Aa (formerly BinAB) from Ls [ 74 ] and to lepidopteran when the recognition loop 3 of domain II from Cry1Ab was inserted between its loops 1 and 2 [ 75 ]. Interestingly, interdomains exchanges conducted between Cry11Aa and Cry11Ba from Bti and Btj , respectively, highlighted that while some domain combinations improved the mosquitocidal activity, others completely abolished the ability of the chimeras to form inclusions [ 76 , 77 ].…”

Section: Producing New Crystalline Toxins For the Development Of Innovative Bioinsecticidesmentioning

confidence: 99%

“…Moreover, the checkerboard-like crystal packing of Cry3Aa offers large solvent channels that could be able to accommodate a cargo protein up to ~50 Å in diameter ( Figure 3 ) [ 31 , 45 ]. This contrasts with other toxins, such as Cyt1Aa, which pack into crystals that might be too dense to allow a cargo protein to cocrystallize [ 88 ], although recent attempts suggest that it might not be an insuperable barrier [ 74 ].…”

Section: Functionalizing Toxin Crystals For the Development Of New Biotechnological Toolsmentioning

confidence: 99%

Can (We Make) Bacillus thuringiensis Crystallize More Than Its Toxins?

Tetreau

Андреева

Banneville

et al. 2021

Toxins

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The development of finely tuned and reliable crystallization processes to obtain crystalline formulations of proteins has received growing interest from different scientific fields, including toxinology and structural biology, as well as from industry, notably for biotechnological and medical applications. As a natural crystal-making bacterium, Bacillus thuringiensis (Bt) has evolved through millions of years to produce hundreds of highly structurally diverse pesticidal proteins as micrometer-sized crystals. The long-term stability of Bt protein crystals in aqueous environments and their specific and controlled dissolution are characteristics that are particularly sought after. In this article, I explore whether the crystallization machinery of Bt can be hijacked as a means to produce (micro)crystalline formulations of proteins for three different applications: (i) to develop new bioinsecticidal formulations based on rationally improved crystalline toxins, (ii) to functionalize crystals with specific characteristics for biotechnological and medical applications, and (iii) to produce microcrystals of custom proteins for structural biology. By developing the needs of these different fields to figure out if and how Bt could meet each specific requirement, I discuss the already published and/or patented attempts and provide guidelines for future investigations in some underexplored yet promising domains.

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Highly Effective Broad Spectrum Chimeric Larvicide That Targets Vector Mosquitoes Using a Lipophilic Protein

Cited by 20 publications

References 33 publications

Functional Bacillus thuringiensis Cyt1Aa Is Necessary To Synergize Lysinibacillus sphaericus Binary Toxin (Bin) against Bin-Resistant and -Refractory Mosquito Species

Functional Bacillus thuringiensis Cyt1Aa Is Necessary To Synergize Lysinibacillus sphaericus Binary Toxin (Bin) against Bin-Resistant and -Refractory Mosquito Species

Engineering Bacillus thuringiensis Cyt1Aa toxin specificity from dipteran to lepidopteran toxicity

Can (We Make) Bacillus thuringiensis Crystallize More Than Its Toxins?

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