Cry1Ac and Vip3Aa proteins from Bacillus thuringiensis targeting Cry toxin resistance in Diatraea flavipennella and Elasmopalpus lignosellus from sugarcane.

Ana Rita Nunes Lemes,Manoel Victor Franco Lemos,Liliane Marques Da Silva,Odair Aparecido Fernandes,Herbert Álvaro Abreu De Siqueira,Camila Soares Figueiredo,Rebeka Da Costa Alves,Janete Apparecida Desidério,Isis Sebastião

doi:10.7717/peerj.2866

Ana Rita Nunes Lemes, Manoel Victor Franco Lemos + Show 7 more

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https://doi.org/10.7717/peerj.2866

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Abstract

The biological potential of Vip and Cry proteins from Bacillus is well known and widely established. Thus, it is important to look for new genes showing different modes of action, selecting those with differentiated entomotoxic activity against Diatraea flavipennella and Elasmopalpus lignosellus, which are secondary pests of sugarcane. Therefore, Cry1 and Vip3 proteins were expressed in Escherichia coli, and their toxicities were evaluated based on bioassays using neonate larvae. Of those, the most toxic were Cry1Ac and Vip3Aa considering the LC50 values. Toxins from E. coli were purified, solubilized, trypsinized, and biotinylated. Brush Border Membrane Vesicles (BBMVs) were prepared from intestines of the two species to perform homologous and heterologous competition assays. The binding assays demonstrated interactions between Cry1Aa, Cry1Ac, and Vip3Aa toxins and proteins from the BBMV of D. flavipennella and E. lignosellus. Homologous competition assays demonstrated that binding to one of the BBMV proteins was specific for each toxin. Heterologous competition assays indicated that Vip3Aa was unable to compete for Cry1Ac toxin binding. Our results suggest that Cry1Ac and Vip3Aa may have potential in future production of transgenic sugarcane for control of D. flavipennella and E. lignosellus, but more research is needed on the potential antagonism or synergism of the toxins in these pests.

Highlights

Sugarcane is grown in several countries of tropical and sub-tropical regions, playing an important role in the economy, especially sugar and alcohol production (Moore, 2005; Srikanth, Subramonian & Premachandran, 2011)
Cry1Ac was most toxic to D. flavipennella and E. lignosellus (LC50 = 8.6 and 15.6 ng/cm2, respectively) of all protoxins tested
While Vip3Aa was similar to Cry1Aa and Cry1Ca in toxicity to E. lignosellus, it was much less toxic to D. flavipennella (LC50 = 495ng/cm2)

Summary

Introduction

Sugarcane is grown in several countries of tropical and sub-tropical regions, playing an important role in the economy, especially sugar and alcohol production (Moore, 2005; Srikanth, Subramonian & Premachandran, 2011). Direct losses are due to galleries burrowed by larvae in sugarcane stems, which can lead to yield reductions or ‘dead heart’ (apical bud death), in 3-month-old plants (Pinto, Machado & Oliveira, 2009; Garcia, 2013). This species is abundant in sugarcane producing areas of Northeastern Brazil and is considered more damaging than D. saccharalis (Mendonca et al, 1996; Pinto, 2006). E. lignosellus is a polyphagous insect whose larvae destroy the meristematic tissue bellow the soil surface and/ or the phloem vessels of saplings, affecting crop stand (Viana, Cruz & Waquil, 2000; Viana, 2004)

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