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

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

doi:10.1038/s41598-017-11717-9

Dennis K Bideshi, Robert H Hice + Show 3 more

Open Access

https://doi.org/10.1038/s41598-017-11717-9

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Abstract

Two mosquitocidal bacteria, Bacillus thuringiensis subsp. israelensis (Bti) and Lysinibacillus sphaericus (Ls) are the active ingredients of commercial larvicides used widely to control vector mosquitoes. Bti’s efficacy is due to synergistic interactions among four proteins, Cry4Aa, Cry4Ba, Cry11Aa, and Cyt1Aa, whereas Ls’s activity is caused by Bin, a heterodimer consisting of BinA, the toxin, and BinB, a midgut-binding protein. Cyt1Aa is lipophilic and synergizes Bti Cry proteins by increasing midgut binding. We fused Bti’s Cyt1Aa to Ls’s BinA yielding a broad-spectrum chimeric protein highly mosquitocidal to important vector species including Anopheles gambiae, Culex quinquefasciatus, and Aedes aegypti, the latter an important Zika and Dengue virus vector insensitive to Ls Bin. Aside from its vector control potential, our bioassay data, in contrast to numerous other reports, provide strong evidence that BinA does not require conformational interactions with BinB or microvillar membrane lipids to bind to its intracellular target and kill mosquitoes.

Highlights

The Lysinibacillus sphaericus (Ls) binary toxin (Bin) is a heterodimer of two related propeptides, BinA, a toxin (42 kDa), and BinB, a midgut microvillar binding protein (51 kDa), which co-crystallize during synthesis[5, 6, 12,13,14]
In a proof-of-concept study, we fused the Cyt1Aa protoxin, which has high affinity for mosquito microvilli lipids, to the BinA protoxin, yielding the chimeric protein, Cyt1Aa-BinA (69.6 kDa). We evaluated this construct for stable synthesis in 4Q7, an acrystalliferous strain of Bacillus thuringiensis subsp. israelensis (Bti), and for the efficacy of this recombinant chimeric strain against larvae of mosquito species belonging to the three most important genera of disease vectors, Anopheles, Aedes, and Culex
The high toxicity we obtained against Ae. aegypti is potentially important, as our chimera expanded the target spectrum of BinA to include this species, which lacks a BinB receptor and is poorly sensitive to Ls Bin[1]

Summary

Introduction

The Ls binary toxin (Bin) is a heterodimer of two related propeptides, BinA, a toxin (42 kDa), and BinB, a midgut microvillar binding protein (51 kDa), which co-crystallize during synthesis[5, 6, 12,13,14]. BinB binds to a glycoprotein receptor, the first identified being a glycosylphosphatidylinositol (GPI)-anchored α-glucosidase[15]. That act at the microvillar surface, BinA and BinB are internalized and act intracellularly killing cells by autophagy and/or apoptosis[5, 16, 17], during which large cytoplasmic vacuoles are formed followed by midgut exfoliation that results in larval death. At LC90 levels, Ls mortality peaks at 48 hours post-treatment due to Bin’s internalization process, whereas with Bti maximum mortality occurs at 24 hours post-treatment

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