Similar Genetic Basis of Resistance to Bt Toxin Cry1Ac in Boll-Selected and Diet-Selected Strains of Pink Bollworm

Genetically engineered cotton and corn plants producing insecticidal Bacillus thuringiensis (Bt) toxins kill some key insect pests. Yet, evolution of resistance by pests threatens long-term insect control by these transgenic Bt crops. We compared the genetic basis of resistance to Bt toxin Cry1Ac in two independently derived, laboratory-selected strains of a major cotton pest, the pink bollworm (Pectinophora gossypiella [Saunders]). The Arizona pooled resistant strain (AZP-R) was started with pink bollworm from 10 field populations and selected with Cry1Ac in diet. The Bt4R resistant strain was started with a long-term susceptible laboratory strain and selected first with Bt cotton bolls and later with Cry1Ac in diet. Previous work showed that AZP-R had three recessive mutations (r1, r2, and r3) in the pink bollworm cadherin gene (PgCad1) linked with resistance to Cry1Ac and Bt cotton producing Cry1Ac. Here we report that inheritance of resistance to a diagnostic concentration of Cry1Ac was recessive in Bt4R. In interstrain complementation tests for allelism, F1 progeny from crosses between AZP-R and Bt4R were resistant to Cry1Ac, indicating a shared resistance locus in the two strains. Molecular analysis of the Bt4R cadherin gene identified a novel 15-bp deletion (r4) predicted to cause the loss of five amino acids upstream of the Cry1Ac-binding region of the cadherin protein. Four recessive mutations in PgCad1 are now implicated in resistance in five different strains, showing that mutations in cadherin are the primary mechanism of resistance to Cry1Ac in laboratory-selected strains of pink bollworm from Arizona

Lessons from agriculture for the sustainable management of malaria vectors.

Willem Takken and colleagues argue for the expansion of insecticide monotherapy in malaria control by taking lessons from agriculture and including more sustainable integrated vector management strategies

Shared midgut binding sites for Cry1A.105, Cry1Aa, Cry1Ab, Cry1Ac and Cry1Fa proteins from Bacillus thuringiensis in two important corn pests, Ostrinia nubilalis and Spodoptera frugiperda

First generation of insect-protected transgenic corn (Bt-corn) was based on the expression of Cry1Ab or Cry1Fa proteins. Currently, the trend is the combination of two or more genes expressing proteins that bind to different targets. In addition to broadening the spectrum of action, this strategy helps to delay the evolution of resistance in exposed insect populations. One of such examples is the combination of Cry1A.105 with Cry1Fa and Cry2Ab to control O. nubilalis and S. frugiperda. Cry1A.105 is a chimeric protein with domains I and II and the C-terminal half of the protein from Cry1Ac, and domain III almost identical to Cry1Fa. The aim of the present study was to determine whether the chimeric Cry1A.105 has shared binding sites either with Cry1A proteins, with Cry1Fa, or with both, in O. nubilalis and in S. frugiperda. Brush-border membrane vesicles (BBMV) from last instar larval midguts were used in competition binding assays with 125I-labeled Cry1A.105, Cry1Ab, and Cry1Fa, and unlabeled Cry1A.105, Cry1Aa, Cry1Ab, Cry1Ac, Cry1Fa, Cry2Ab and Cry2Ae. The results showed that Cry1A.105, Cry1Ab, Cry1Ac and Cry1Fa competed with high affinity for the same binding sites in both insect species. However, Cry2Ab and Cry2Ae did not compete for the binding sites of Cry1 proteins. Therefore, according to our results, the development of cross-resistance among Cry1Ab/Ac, Cry1A.105, and Cry1Fa proteins is possible in these two insect species if the alteration of shared binding sites occurs. Conversely, cross-resistance between these proteins and Cry2A proteins is very unlikely in such case

Genetic hitchhiking and resistance evolution to transgenic Bt toxins: insights from the African stalk borer Busseola fusca (Noctuidae)

Since transgenic crops expressing Bacillus thuringiensis (Bt) toxins were first released, resistance evolution leading to failure in control of pests populations has been observed in a number of species. Field resistance of the moth Busseola fusca was acknowledged 8 years after Bt maize was introduced in South Africa. Since then, field resistance of this corn borer has been observed at several locations, raising questions about the nature, distribution and dynamics of the resistance trait. Using genetic markers, our study identified four outlier loci clearly associated with resistance. In addition, genetic structure at neutral loci reflected extensive gene flow among populations. A realistically parameterised model suggests that resistance could travel in space at speed of several kilometres a year. Markers at outlier loci delineated a geographic region associated with resistance spread. This was an area of approximately 100 km radius, including the location where resistance was first reported. Controlled crosses corroborated these findings and showed significant differences of progeny survival on Bt plants depending on the origin of the resistant parent. Last, our study suggests diverse resistance mutations, which would explain the widespread occurrence of resistant larvae in Bt fields across the main area of maize production in South Africa

Video-tracking and On-plant Tests Show Cry1Ab Resistance Influences Behavior and Survival of Neonate Ostrinia nubilalis Following Exposure to Bt Maize

To examine how resistance to Bacillus thuringiensis (Bt) toxins influences movement and survival of European corn borer (Ostrinia nubilalis [Hübner]) neonates, the responses of Cry1Ab-resistant , -susceptible, and hybrid (F1) larvae were examined using two different techniques. First, using an automated video-tracking system, aspects of O. nubilalis movement were quantified in the presence of artificial diet incorporating 50% non-Bt or insect-resistant Cry1Ab maize tissue. Second, O. nubilalis dispersal and survival were measured 48–72 h after hatching on a Cry1Ab maize plant surrounded by two non-Bt maize plants. Video tracking indicated the presence of Cry1Ab tissue increased the total distance moved (m), time moving (%), and time away from the diet (%) for O. nubilalis while decreasing meander (degrees/cm). However, resistant larvae showed reduced movement and increased meander (≈localized searching) relative to susceptible or hybrid larvae on diet incorporating Cry1Ab tissue. Conversely, when placed onto Cry1Ab maize plants, resistant larvae were more likely than susceptible O. nubilalis to disperse onto adjacent non-Bt plants. The difference in on-plant dispersal seems to reflect greater survival after toxin exposure for resistant larvae rather than increased activity. These results suggest that simplified ‘Petri dish’ tests may not be predictive of larval movement among non-Bt and insect-resistant Bt maize plants. Because models of O. nubilalis resistance evolution incorporate various movement and survival parameters, improved data for on-plant behavior and survival of Bt- resistant , -susceptible, and hybrid larvae should help preserve the efficacy of transgenic insect-resistant maize

An individual-based model of the evolution of pesticide resistance in heterogeneous environments : Control of meligethes aeneus population in oilseed rape crops

Copyright: © 2014 Stratonovitch et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Preventing a pest population from damaging an agricultural crop and, at the same time, preventing the development of pesticide resistance is a major challenge in crop protection. Understanding how farming practices and environmental factors interact with pest characteristics to influence the spread of resistance is a difficult and complex task. It is extremely challenging to investigate such interactions experimentally at realistic spatial and temporal scales. Mathematical modelling and computer simulation have, therefore, been used to analyse resistance evolution and to evaluate potential resistance management tactics. Of the many modelling approaches available, individual-based modelling of a pest population offers most flexibility to include and analyse numerous factors and their interactions. Here, a pollen beetle (Meligethes aeneus) population was modelled as an aggregate of individual insects inhabiting a spatially heterogeneous landscape. The development of the pest and host crop (oilseed rape) was driven by climatic variables. The agricultural land of the landscape was managed by farmers applying a specific rotation and crop protection strategy. The evolution of a single resistance allele to the pyrethroid lambda cyhalothrin was analysed for different combinations of crop management practices and for a recessive, intermediate and dominant resistance allele. While the spread of a recessive resistance allele was severely constrained, intermediate or dominant resistance alleles showed a similar response to the management regime imposed. Calendar treatments applied irrespective of pest density accelerated the development of resistance compared to ones applied in response to prescribed pest density thresholds. A greater proportion of springs own oilseed rape was also found to increase the speed of resistance as it increased the period of insecticide exposure. Our study demonstrates the flexibility and power of an individual-based model to simulate how farming practices affect pest population dynamics, and the consequent impact of different control strategies on the risk and speed of resistance development.Peer reviewe

Fitness Cost of Resistance to Bt Cotton Linked with Increased Gossypol Content in Pink Bollworm Larvae

Fitness costs of resistance to Bacillus thuringiensis (Bt) crops occur in the absence of Bt toxins, when individuals with resistance alleles are less fit than individuals without resistance alleles. As costs of Bt resistance are common, refuges of non-Bt host plants can delay resistance not only by providing susceptible individuals to mate with resistant individuals, but also by selecting against resistance. Because costs typically vary across host plants, refuges with host plants that magnify costs or make them less recessive could enhance resistance management. Limited understanding of the physiological mechanisms causing fitness costs, however, hampers attempts to increase costs. In several major cotton pests including pink bollworm (Pectinophora gossypiella), resistance to Cry1Ac cotton is associated with mutations altering cadherin proteins that bind this toxin in susceptible larvae. Here we report that the concentration of gossypol, a cotton defensive chemical, was higher in pink bollworm larvae with cadherin resistance alleles than in larvae lacking such alleles. Adding gossypol to the larval diet decreased larval weight and survival, and increased the fitness cost affecting larval growth, but not survival. Across cadherin genotypes, the cost affecting larval growth increased as the gossypol concentration of larvae increased. These results suggest that increased accumulation of plant defensive chemicals may contribute to fitness costs associated with resistance to Bt toxins

Vip3A Resistance Alleles Exist at High Levels in Australian Targets before Release of Cotton Expressing This Toxin

Crops engineered to produce insecticidal crystal (Cry) proteins from the soil bacterium Bacillus thuringiensis (Bt) have revolutionised pest control in agriculture. However field-level resistance to Bt has developed in some targets. Utilising novel vegetative insecticidal proteins (Vips), also derived from Bt but genetically distinct from Cry toxins, is a possible solution that biotechnical companies intend to employ. Using data collected over two seasons we determined that, before deployment of Vip-expressing plants in Australia, resistance alleles exist in key targets as polymorphisms at frequencies of 0.027 (n = 273 lines, 95% CI = 0.019–0.038) in H. armigera and 0.008 (n = 248 lines, 0.004–0.015) in H. punctigera. These frequencies are above mutation rates normally encountered. Homozygous resistant neonates survived doses of Vip3A higher than those estimated in field-grown plants. Fortunately the resistance is largely, if not completely, recessive and does not confer resistance to the Bt toxins Cry1Ac or Cry2Ab already deployed in cotton crops. These later characteristics are favourable for resistance management; however the robustness of Vip3A inclusive varieties will depend on resistance frequencies to the Cry toxins when it is released (anticipated 2016) and the efficacy of Vip3A throughout the season. It is appropriate to pre-emptively screen key targets of Bt crops elsewhere, especially those such as H. zea in the USA, which is not only closely related to H. armigera but also will be exposed to Vip in several varieties of cotton and corn

Resistance of Trichoplusia ni to Bacillus thuringiensis Toxin Cry1Ac Is Independent of Alteration of the Cadherin-Like Receptor for Cry Toxins

Alteration of binding sites for Bacillus thuringiensis (Bt) toxins in insect midgut is the major mechanism of high-level resistance to Bt toxins in insects. The midgut cadherin is known to be a major binding protein for Bt Cry1A toxins and linkage of Bt-resistance to cadherin gene mutations has been identified in lepidopterans. The resistance to Bt toxin Cry1Ac evolved in greenhouse populations of Trichoplusia ni has been identified to be associated with the down-regulation of an aminopeptidase N (APN1) gene by a trans-regulatory mechanism and the resistance gene has been mapped to the locus of an ABC transporter (ABCC2) gene. However, whether cadherin is also involved with Cry1Ac-resistance in T. ni requires to be understood. Here we report that the Cry1Ac-resistance in T. ni is independent of alteration of the cadherin. The T. ni cadherin cDNA was cloned and the cadherin sequence showed characteristic features known to cadherins from Lepidoptera. Various T. ni cadherin gene alleles were identified and genetic linkage analysis of the cadherin alleles with Cry1Ac-resistance showed no association of the cadherin gene with the Cry1Ac-resistance in T. ni. Analysis of cadherin transcripts showed no quantitative difference between the susceptible and Cry1Ac-resistant T. ni larvae. Quantitative proteomic analysis of midgut BBMV proteins by iTRAQ-2D-LC-MS/MS determined that there was no quantitative difference in cadherin content between the susceptible and the resistant larvae and the cadherin only accounted for 0.0014% (mol%) of the midgut BBMV proteins, which is 1/300 of APN1 in molar ratio. The cadherin from both the susceptible and resistant larvae showed as a 200-kDa Cry1Ac-binding protein by toxin overlay binding analysis, and nano-LC-MS/MS analysis of the 200-kDa cadherin determined that there is no quantitative difference between the susceptible and resistant larvae. Results from this study indicate that the Cry1Ac-resistance in T. ni is independent of cadherin alteration