scholarly journals Engineering of Bacillus thuringiensis Cry Proteins to Enhance the Activity against Western Corn Rootworm

Toxins ◽  
2019 ◽  
Vol 11 (3) ◽  
pp. 162 ◽  
Author(s):  
Jingtong Hou ◽  
Ruth Cong ◽  
Michi Izumi-Willcoxon ◽  
Hana Ali ◽  
Yi Zheng ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
3D Structure ◽  
Diabrotica Virgifera Virgifera ◽  
Western Corn Rootworm ◽  
Corn Rootworm ◽  
Diabrotica Virgifera ◽  
Cry Protein ◽  
Neutral Ph ◽  
Highly Active ◽  
Cry Toxin

A novel Bacillus thuringiensis Cry protein, Cry8Hb, active against Diabrotica virgifera virgifera (Western corn rootworm, WCRW) was discovered. Unexpectedly, the anti-rootworm activity of the Cry8Hb toxin was enhanced significantly by fusing Escherichia coli maltose binding protein (MBP) to this Cry toxin. While the exact mechanism of the activity enhancement remains indefinite, it is probable that the enhancement is a result of increased solubility of the MBP-Cry8Hb fusion in the rootworm midgut. This hypothesis was examined using a synthetic Cry3 protein called IP3-1, which was not soluble at a neutral pH like Cry8Hb and marginally active to WCRW. When IP3-1 was fused to MBP, its anti-WCRW activity was enhanced 13-fold. To further test the hypothesis, DNA shuffling was performed on IP3-1 to increase the solubility without MBP. Screening of shuffled libraries found six new IP3 variants showing very high anti-WCRW activity without MBP. Sequence and 3D structure analysis of those highly active, shuffled IP3 variants revealed several charge-altering mutations such as Lys to Glu on the putative MBP-attaching side of the IP3 molecule. It is likely that those mutations make the protein acidic to substitute the functions of MBP including enhancing the solubility of IP3 at a neutral pH.

scholarly journals Cry75Aa (Mpp75Aa) Insecticidal Proteins for Controlling the Western Corn Rootworm, Diabrotica virgifera virgifera, (Coleoptera: Chrysomelidae), Isolated from the Insect Pathogenic Bacteria Brevibacillus laterosporus.

2020 ◽  
Author(s):  
David Bowen ◽  
Yong Yin ◽  
Stanislaw Flasinski ◽  
Catherine Chay ◽  
Gregory Bean ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Pathogenic Bacteria ◽  
Insect Pests ◽  
Diabrotica Virgifera Virgifera ◽  
Western Corn Rootworm ◽  
Cross Resistance ◽  
Corn Rootworm ◽  
Diabrotica Virgifera ◽  
Insecticidal Proteins ◽  
Brevibacillus Laterosporus

This study describes three closely related proteins, cloned from Brevibacillus laterosporus strains, that are lethal upon feeding to Diabrotica virgifera virgifera LeConte, the western corn rootworm (WCR). Mpp75Aa1, Mpp75Aa2 and Mpp75Aa3 were toxic to WCR larvae when fed purified protein. Transgenic plants expressing each mMpp75Aa protein were protected from feeding damage and showed significant reduction in adult emergence from infested plants by both susceptible and Cry3Bb1 and Cry34Ab1/Cry35Ab1-resistant WCR. These results demonstrate that proteins from B. laterosporus are as efficacious as the well-known Bacillus thuringiensis (Bt) insecticidal proteins in controlling major insect pests such as WCR. The deployment of transgenic maize expressing mMpp75Aa along with other active molecules lacking cross-resistance have the potential to be a useful tool for control of WCR populations resistant to current Bt traits. IMPORTANCE Insects feeding on roots of crops can damage the plant roots resulting in yield loss due to poor water and nutrient uptake and plant lodging. In maize the western corn rootworm (WCR) can cause severe damage to the roots resulting in significant economic loss for farmers. Genetically modified (GM) expressing Bacillus thuringiensis (Bt) insect control proteins, has provided a solution for control of these pests. In recent years populations of WCR resistant to the Bt proteins in commercial GM maize have emerged. There is a need to develop new insecticidal traits for the control of WCR populations resistant to current commercial traits. New proteins with commercial level efficacy on WCR from sources other than Bt are becoming more critical. The Mpp75Aa proteins, from B. laterosporus, when expressed in maize, are efficacious against the resistant populations of WCR and have the potential to provide solutions for control of resistant WCR.

scholarly journals Binary Toxins from Bacillus thuringiensis Active against the Western Corn Rootworm, Diabrotica virgifera virgifera LeConte

2004 ◽  
Vol 70 (8) ◽  
pp. 4889-4898 ◽  
Author(s):  
James A. Baum ◽  
Chi-Rei Chu ◽  
Mark Rupar ◽  
Gregory R. Brown ◽  
William P. Donovan ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Insect Pest ◽  
Diabrotica Virgifera Virgifera ◽  
The United States ◽  
Western Corn Rootworm ◽  
Binary Toxin ◽  
Corn Rootworm ◽  
Diabrotica Virgifera ◽  
Crystal Proteins ◽  
Genes Encoding

ABSTRACT The western corn rootworm, Diabrotica virgifera virgifera LeConte, is a significant pest of corn in the United States. The development of transgenic corn hybrids resistant to rootworm feeding damage depends on the identification of genes encoding insecticidal proteins toxic to rootworm larvae. In this study, a bioassay screen was used to identify several isolates of the bacterium Bacillus thuringiensis active against rootworm. These bacterial isolates each produce distinct crystal proteins with approximate molecular masses of 13 to 15 kDa and 44 kDa. Insect bioassays demonstrated that both protein classes are required for insecticidal activity against this rootworm species. The genes encoding these proteins are organized in apparent operons and are associated with other genes encoding crystal proteins of unknown function. The antirootworm proteins produced by B. thuringiensis strains EG5899 and EG9444 closely resemble previously described crystal proteins of the Cry34A and Cry35A classes. The antirootworm proteins produced by strain EG4851, designated Cry34Ba1 and Cry35Ba1, represent a new binary toxin. Genes encoding these proteins could become an important component of a sustainable resistance management strategy against this insect pest.

scholarly journals Detection of alternative splicing in Diabrotica virgifera virgifera LeConte, in association with Bt resistance using RNA-seq and PacBio Iso-Seq

2020 ◽  
Author(s):  
Zixiao Zhao ◽  
Christine G. Elsik ◽  
Bruce E. Hibbard ◽  
Kent S. Shelby
Keyword(s):  
Alternative Splicing ◽  
Matrix Protein ◽  
Diabrotica Virgifera Virgifera ◽  
Western Corn Rootworm ◽  
Corn Rootworm ◽  
Diabrotica Virgifera ◽  
Rna Seq ◽  
Bt Resistance ◽  
Bt Toxins ◽  
Alternatively Spliced

AbstractBackgroundAlternative splicing is one of the major mechanisms that increases transcriptome diversity in eukaryotes, including insect species that have gained resistance to pesticides and Bt toxins. In western corn rootworm (Diabrotica virgifera virgifera LeConte), neither alternative splicing nor its role in resistance to Bt toxins has been studied.ResultsTo investigate the mechanisms of Bt resistance we carried out single-molecule real-time (SMRT) transcript sequencing and Iso-seq analysis on resistant, eCry3.1Ab-selected and susceptible, unselected, western corn rootworm neonate midguts which fed on seedling maize with and without eCry3.1Ab for 12 and 24 hours. We present transcriptome-wide alternative splicing patterns of western corn rootworm midgut in response to feeding on eCry3.1Ab-expressing corn using a comprehensive approach that combines both RNA-seq and SMRT transcript sequencing techniques. We found that 67.73% of multi-exon genes are alternatively spliced, which is consistent with the high transposable element content of the genome. One of the alternative splicing events we identified was a novel peritrophic matrix protein with two alternative splicing isoforms. Analysis of differential exon usage between resistant and susceptible colonies showed that in eCry3.1Ab-resistant western corn rootworm, expression of one isoform was significantly higher than in the susceptible colony, while no significant differences between colonies were observed with the other isoform.ConclusionOur results provide the first survey of alternative splicing in western corn rootworm and suggest that the observed alternatively spliced isoforms of peritrophic matrix protein may be associated with eCry3.1Ab resistance in western corn rootworm.

Soybean Foliage Consumption Reduces Adult Western Corn Rootworm (Diabrotica virgifera virgifera)(Coleoptera: Chrysomelidae) Survival and Stimulates Flight

2021 ◽  
Author(s):  
Joseph L Spencer ◽  
Timothy R Mabry ◽  
Eli Levine ◽  
Scott A Isard
Keyword(s):  
Wind Tunnel ◽  
Crop Rotation ◽  
Diabrotica Virgifera Virgifera ◽  
Western Corn Rootworm ◽  
Egg Laying ◽  
Corn Rootworm ◽  
Diabrotica Virgifera ◽  
Egg Maturation ◽  
Behavioral Resistance ◽  
Collection Sites

Abstract Western corn rootworm, Diabrotica virgifera virgifera LeConte, biology is tied to the continuous availability of its host (corn, Zea mays L.). Annual rotation of corn with a nonhost, like soybean (Glycine max (L.) Merrill) was a reliable tactic to manage western corn rootworm. Behavioral resistance to annual crop rotation (rotation resistance) allowed some eastern U.S. Corn Belt populations to circumvent rotation by laying eggs in soybean and in cornfields. When active in soybean, rotation-resistant adults commonly consume foliage, in spite of detrimental effects on beetle survival. Rotation-resistant beetle activity in soybean is enabled by the expression of certain proteinases and an adapted gut microbiota that provide limited protection from soybean antiherbivore defenses. We investigated the effects of corn and soybean herbivory on rotation-resistant female survival and initiation of flight using mortality assays and wind tunnel flight tests. Among field-collected females tested with mortality assays, beetles from collection sites in a cornfield survived longer than those from collection sites in a soybean field. However, reduced survival due to soybean herbivory could be restored by consuming corn tissues. Field-collected beetles that fed on a soybean tissue laboratory diet or only water were more likely to fly in a wind tunnel than corn-feeding beetles. Regardless of collection site and laboratory diet, 90.5% of beetles that flew oriented their flights upwind. Diet-related changes in the probability of flight provide a proximate mechanism for interfield movement that facilitates restorative feeding and the survival of females previously engaged in soybean herbivory. Rotation-resistant western corn rootworm females feeding on soybean tissues experience reduced survival in mortality assays and display increased flight probability (which may facilitate flight back to a cornfield where consumption of host tissues improves survival potential and facilitates maturation of eggs). The consequences of soybean herbivory provide a proximal mechanism for behavioral resistance to crop rotation. Increased egg-laying probability while feeding on soybean tissues, facilitation of egg maturation while feeding on corn tissues, and interfield movement are previously documented consequences.

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