Bt Traits Research Articles

Applying a computer model to evaluate the evolution of resistance by western corn rootworm to multiple Bt traits in transgenic maize.

Western corn rootworm, Diabrotica virgifera virgifera (LeConte) (Coleoptera: Chrysomelidae), is a major pest of maize in the United States. Transgenic maize producing insecticidal toxins from the bacterium Bacillus thuringiensis (Bt) have been used to manage this pest since 2003. Refuges of non-Bt maize have been used to delay resistance to Bt maize by western corn rootworm, and are planted in conjunction with maize producing single or multiple (i.e., pyramids) Bt toxins. Two Bt toxins, Cry3Bb1 and Gpp34/Tpp35Ab1, were used individually before being combined as a pyramid, at which point resistance had already evolved to Cry3Bb1. Pyramids targeting western corn rootworm therefore contained at least one toxin to which resistance had evolved. Western corn rootworm has now evolved resistance to all four commercially available Bt toxins used for rootworm management. We used laboratory and field-generated data to parameterize a deterministic model to simulate the effectiveness of refuges and Bt pyramids to delay resistance to Bt maize in western corn rootworm. Resistance to the pyramid of Cry3Bb1 with Gpp34/Tpp35Ab1 evolved more rapidly when resistance to Cry3Bb1 was already present. This effect arose when model conditions affecting initial resistance allele frequency, inheritance of resistance, and fitness costs were varied. Generally, resistance evolved faster when initial resistance allele frequencies were higher, inheritance of resistance was nonrecessive, and fitness costs were absent, which is consistent with previous models that simulated resistance evolution. We conclude that new transgenic pyramids should pair novel, independently acting toxins with abundant refuges to minimize the risk of rapid resistance evolution.

Differential susceptibility of Spodoptera frugiperda (Lepidoptera: Noctuidae) to single versus pyramided Bt traits in Brazilian soybean: what doesn't kill you makes you stronger?

Lepidopteran pest control in agriculture has become heavily dependent on cultivars that express Bacillus thuringiensis (Bt) toxins as 'plant-incorporated protectants'. However, populations of Spodoptera frugiperda (Smith) in Brazil appear resistant to the Bt traits currently available in commercial soybean cultivars. This study evaluated S. frugiperda life history when feeding on three different Bt soybean cultivars. Cultivars expressing Cry1Ac + Cry1F and Cry1A.105 + Cry2Ab2 + Cry1Ac Bt toxins caused 100% larval mortality in S. frugiperda. Both non-Bt and Cry1Ac-expressing soybean induced transgenerational effects that increased the survival of subsequent generations. A Cry1Ac soybean diet reduced the generation time (T) of S. frugiperda relative to non-Bt soybean, resulting in shorter generation time and more rapid population growth. The implications of these results revealed how diet can alter aspects of insect life history and biology, and have important implications for sustainable management of S. frugiperda on soybean. © 2024 Society of Chemical Industry.

Larval density-dependent mortality of western corn rootworm (Coleoptera: Chrysomelidae) in Bt and non-Bt maize and implications on dose calculations†.

Transgenic crops producing insecticidal toxins from the bacterium Bacillus thuringiensis (Bt) have been used to manage insect pests for nearly 30 years. Dose of a Bt crop is key to assessing the risk of resistance evolution because it affects the heritability of resistance traits. Western corn rootworm (Diabrotica virgifera virgifera, LeConte), a major pest of maize, has evolved resistance to all commercially available Bt traits targeting it, and threatens resistance to future transgenic traits. Past research shows the dose of Bt maize targeting western corn rootworm can be confounded by larval density-dependent mortality. We conducted a 2-year field study at two locations to quantify larval density-dependent mortality in Bt and non-Bt maize. We used these results to calculate dose for our method and compared it to three previously published methods. Additionally, adult emergence and root injury were analyzed for predicting initial egg density. Increased pest density caused greater proportions of larvae to die in Bt maize than in non-Bt maize. All methods for calculating dose produced values less than high-dose, and stochastic variation had the greatest impact on dose at high and low pest densities. Our method for calculating dose did not produce values positively correlated with pest density while the three other methods did. To achieve the most accurate calculation of dose for transgenic maize targeting western corn rootworm, density-dependent mortality should be taken into account for both transgenic and non-transgenic maize and assessed at moderate pest densities. © 2024 Society of Chemical Industry.

More than just an insect killer: The non-insecticidal activities of Bacillus thuringiensis with biotechnological potential

Bacillus thuringiensis (Bt) is known for the biological control of important insect pests, but scientific advances have revealed several interesting characteristics, in addition to this classical function as a bioinsecticide. To investigate the current knowledge about these non-insecticidal activities, a systematic research on primary data in the scientific literature was conducted on alternative functions of Bt with biotechnological potential. Out of a total of 140 articles selected, 15 non-insecticidal Bt activities were found. Publications related to this topic are available since 1971, and different metadata were reported, such as biomolecules and genes involved in Bt performances in non-insecticidal bioactivities. A total of 11 Bt activities with different effect measures (response variables) were identified, with an average of 48 distinct Bt strains evaluated per activity. Approximately 81.2% of all identified experiments/tests deal with the direct effects of Bt on target cells/organisms, with 36.3% of the strains within these studies tested for antibacterial action; of all microbial targets tested, 92.8% are bacteria, which led to 75.2% of the experimental conditions for all direct activities being performed in vitro. Regarding indirect Bt activities, 67.6% of these studies reported tritrophic Bt-plant-pathogen interactions. Bioremediation also appears as a relevant Bt activity being investigated in-depth. Alternative Bt activities offer innovative ways of developing biotechnology for different areas of anthropic interest; hence, we also focus on the possibility of finding multifunctional strains of Bt, as this may be advantageous from a bioeconomic point of view. Our findings are discussed in terms of research trends, aspects, details and depth of the current knowledge on alternative non-insecticidal Bt traits. We also discuss the potential application of this science for useful technological developments, aiming at solving issues related to human health, sustainable agriculture and environmental preservation/restoration.

Bt corn hybrids expressing mCry3A and eCry3.1Ab Proteins protect corn roots against western corn rootworm injury.

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is one of the most serious pests of corn (Zea mays L.) In 2017 and 2018, studies were conducted in fields with and without known unexpected root injury to Cry3Bb1, to determine root protection by Bt corn hybrids expressing both mCry3A and eCry3.1Ab insecticidal crystal proteins, and hybrids expressing either mCry3A or eCry3.1Ab only against the WCR root injury. Node injury was evaluated using the Iowa State University 0-3 node-injury scale (NIS), and the consistency of root protection was also determined. In 2017, with medium to high larval feeding pressure, the Bt corn hybrids expressing both mCry3A and eCry3.1Ab in the breeding stack, molecular stack, and Bt corn hybrid expressing eCry3.1Ab only, sustained low node injury compared with Bt corn hybrid expressing mCry3A only, and the non-Bt corn. In 2018, with low larval feeding pressure in most of the locations, node injury was not different for the Bt and Non-Bt corn hybrids. Across all locations in both years, the Bt corn hybrids expressing both mCry3A and eCry3.1Ab provided better and consistent node injury protection. Bt corn hybrids expressing both mCry3A and eCry3.1Ab proteins provided better root protection and consistency than the Bt corn hybrid expressing mCry3A only, and non-Bt. Therefore, stacking of Bt traits will be the best option for managing insect resistance. © 2023 Society of Chemical Industry.

Movement of Striacosta albicosta (Smith) (Lepidoptera: Noctuidae) Larvae on Transgenic Bt and Non-Bt Maize.

Exposure of lepidopteran pests to Bacillus thuringiensis (Bt) proteins has been shown to affect the behavior of larvae, including increased movement and avoidance of Bt-expressing plants or diet. Therefore, we hypothesized that the behavior of western bean cutworm, Striacosta albicosta (Smith) (Lepidoptera: Noctuidae), an important pest of maize, could be affected when exposed to Bt plants. To test this hypothesis, we conducted a series of artificial arena and on-plant experiments to determine S. albicosta neonate behavior when exposed to Bt and non-Bt plant tissue. Video tracking experiments presented neonate larvae with the choice of Bt or non-Bt pollen in a Petri dish for 15 min while being video recorded for analysis with EthoVision software. This study showed an increase in mean velocity and total time spent moving for larvae in the presence of Cry1F vs. non-Bt when compared with Vip3A vs. non-Bt or Cry1F vs. Vip3A. However, there was no difference in total distance moved or time spent in the food zone for all scenarios. Maize tissue choice experiments allowed neonatal larvae the choice of feeding on Bt or non-Bt tassel or leaves for 9 h in Petri dish arenas. This experiment showed that larvae preferred tassel tissue over leaves but did not indicate that larvae could distinguish between Bt and non-Bt tissue. In contrast, on-plant experiments (including a whole plant neonate dispersal study under controlled conditions and an in-field silking behavior experiment) indicated that the presence of Cry1F and Vip3A Bt toxins increased plant abandonment, suggesting that larvae are able to detect and avoid Bt toxins. The discrepancy of these results is likely due to the on-plant studies providing more field-realistic environmental conditions and a longer duration of exposure to Bt toxins for the behavioral experiments. Our results represent the first steps in understanding the complex behavior of S. albicosta when exposed to Bt plants. A better understanding of the response of larvae when exposed to Bt traits can aid in the management of this pest, particularly for the design of resistance management strategies and refuge design.

Removing neonicotinoid seed treatments has negligible effects on refuge function and crop protection in transgenic maize targeting western corn rootworm (Coleoptera: Chrysomelidae).

Nearly all maize seed sold in the United States includes a neonicotinoid seed treatment (NST), meant to protect seedlings against early-season insect pests. For key pests, including western corn rootworm (Diabrotica virgifera virgifera LeConte) (D.v.v), insecticidal proteins derived from Bacillus thuringiensis (Bt) are expressed in plant tissues as alternatives to soil-applied insecticides. Insect resistance management (IRM) plans use non-Bt "refuges" to encourage survival of Bt-susceptible D.v.v., which maintains susceptible alleles in the population. In non-cotton producing regions, IRM guidelines require a minimum 5% blended refuge for maize expressing more than 1 trait targeting D.v.v. Prior work has shown that 5% blends yield insufficient proportions of refuge beetles to contribute reliably to IRM. Whether NSTs interfere with survivorship of refuge beetles is unknown. Our objective was to determine whether NSTs affect proportions of refuge beetles, and secondarily, to determine whether NSTs provide agronomic advantages over Bt seed alone. To reveal host plant type (i.e., Bt or refuge), we used a stable isotope (15N) to mark refuge plants in plots with 5% seed blends. To assess refuge performance between treatments, we compared proportions of beetles from respective natal hosts. In all site-years, NSTs showed inconsistent effects on proportions of refuge beetles. Treatment comparisons showed inconsistent agronomic benefits of NSTs when combined with Bt traits. Our results demonstrate that NSTs have a negligible impact on refuge performance and reinforces the assertion that 5% blends are serving little benefit for IRM. Plant stand and yield were not improved by NSTs.

Bt cotton and modern insecticide adoption decreases Helicoverpa spp. population recruitment in a subtropical cropping system

AbstractExtensive crop sampling for Helicoverpa spp. pupae was undertaken (2017–2021) to provide insights as to how pest population dynamics have altered in a subtropical cropping system following the introduction of transgenic Bt cotton and newer generation insecticides for pulse production. Previously (1996–1999), a pattern of year‐round population cycling and build‐up was identified to occur between summer cotton (non‐Bt) and winter chickpea, enabled by widespread resistance to broad‐spectrum insecticides used at the time. Current pupae sampling was unable to recover pupae from Bt cotton, suggesting this crop had become a population sink rather than a source for Helicoverpa spp. Pupae were less abundant in pulses, with monthly counts varying from 0 to 3192/ha in chickpea being a fraction of previous densities that ranged from 10,000 to 100,000/ha whilst pupae were not detected in mungbean crops. Poor survival (0%–39.5%) of pupae collected from chickpea, likely due to sub‐lethal insecticide exposure during the larval stage, further limited population recruitment. The highest densities of pupae (up to 18,666/ha) were routinely recovered from unsprayed pigeon pea grown as refuges for Bt cotton resistance management, although Tachinid spp. parasitoids caused increasing pupal mortality with refuge age. This study suggests that the high control efficacy afforded by transgenic Bt traits incorporated into cotton and newer insecticides used on pulse crops has provided a form of area‐wide management that may explain suppression of Helicoverpa spp. pupae densities compared to levels previously recorded across the cropping system.

Effect of Bt traits on transgenic rice’s growth and weed competitiveness

Transgene escape could lead to genetically modified rice establishing wild populations in the natural environment and competing for survival space with weeds. However, whether the expression of the Bacillus thuringiensis (Bt) gene in rice will alter the relationship between transgene plants and weeds and induce undesirable environmental consequences are poorly understood. Thus, field experiments were conducted to investigate the weed competitiveness and assess the ecological risk of transgenic Bt rice under herbicide-free and lepidopterous pest-controlled environments. Results showed that weed–rice competition in the direct-sowing (DS) field was earlier and more severe than that in the transplanting (TP) field, which resulted in a significant decrease in biomass and yield in DS. However, conventional Bt and non-Bt rice yield was not significantly different. The weed number, weed coverage ratio, and weed diversity of conventional Bt rice were significantly higher than those of non-Bt rice at the early growth and mature stages, especially in DS plots, suggesting that Bt traits did not increase the weed competitiveness of transgenic rice and had no negative effect on weed diversity. Grain yield and weed number varied between different hybrid rice lines, but those differences were insignificant between Bt and non-Bt rice. The number of insects increased with the increase of weeds in hybrid rice plots, whereas the insect number and diversity did not display a significant difference between Bt and non-Bt rice. Therefore, the ecological risk of transgenic Bt rice is comparable to non-Bt rice.

Management of Insect Pests with Bt Crops in the United States.

Genetically engineered corn and cotton that produce insecticidal toxins derived from the bacterium Bacillus thuringiensis (Bt) have been used to manage insect pests in the United States and elsewhere. In some cases, this has led to regional suppression of pest populations and pest eradication within the United States, and these outcomes were associated with reductions in conventional insecticides and increased profits for farmers. In other instances, pests evolved resistance to multiple Bt traits, compromising the capacity of Bt crops to manage pests and leading to increased feeding injury to crops in the field. Several aspects of pest biology and pest-crop interactions were associated with cases where pests remained susceptible versus instances where pests evolved resistance. The viability of future transgenic traits can be improved by learning from these past outcomes. In particular, efforts should be made to delay resistance by increasing the prevalence of refuges and using integrated pest management.

Characterizing the sublethal effects of SmartStax PRO dietary exposure on life history traits of the western corn rootworm, Diabrotica virgifera virgifera LeConte.

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, is an economically important pest of field corn (Zea mays L.) across the United States (U.S.) Corn Belt. Repeated use of transgenic hybrids expressing Bacillus thuringiensis (Bt) proteins has selected for field-evolved resistance to all current rootworm-active Bt proteins. The newest product available for WCR management is SmartStax® PRO, a rootworm-active pyramid containing Cry3Bb1, Cry34/35Ab1 [now reclassified as Gpp34Ab1/Tpp35Ab1] and a new mode of action, DvSnf7 dsRNA. Understanding the fitness of adult WCR after dietary exposure to SmartStax® PRO will identify potential impacts on WCR population dynamics and inform efforts to optimize resistance management strategies. Therefore, the objective of the present study was to characterize the effect of SmartStax® PRO dietary exposure on WCR life history traits. Adult WCR were collected during 2018 and 2019 from emergence tents placed over replicated field plots of SmartStax® PRO or non-rootworm Bt corn at a site with a history of rootworm-Bt trait use and suspected resistance to Cry3Bb1 and Cry34/35Ab1. Adult survival was reduced by 97.1–99.7% in SmartStax® PRO plots relative to the non-rootworm Bt corn plots during the study. Individual male/female pairs were fed different diets of ear tissue to simulate lifetime or adult exposure. Life history parameters measured included adult longevity, adult head capsule width, lifetime female egg production, and egg viability. Results indicate that lifetime or adult exposure to SmartStax® PRO significantly reduced adult longevity and lifetime egg production. Larval exposure to SmartStax® PRO significantly reduced WCR adult size. Results from this study collectively suggest that SmartStax® PRO may negatively impact WCR life history traits, which may lead to reduced population growth when deployed in an area with WCR resistance to Bt traits.

Structural and functional insights into the first Bacillus thuringiensis vegetative insecticidal protein of the Vpb4 fold, active against western corn rootworm.

The western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte, is a major maize pest in the United States causing significant economic loss. The emergence of field-evolved resistant WCR to Bacillus thuringiensis (Bt) traits has prompted the need to discover and deploy new insecticidal proteins in transgenic maize. In the current study we determined the crystal structure and mode of action (MOA) of the Vpb4Da2 protein (formerly known as Vip4Da2) from Bt, the first identified insecticidal Vpb4 protein with commercial level control against WCR. The Vpb4Da2 structure exhibits a six-domain architecture mainly comprised of antiparallel β-sheets organized into β-sandwich layers. The amino-terminal domains 1–3 of the protein share structural homology with the protective antigen (PA) PA14 domain and encompass a long β-pore forming loop as in the clostridial binary-toxB module. Domains 5 and 6 at the carboxyl-terminal half of Vpb4Da2 are unique as this extension is not observed in PA or any other structurally-related protein other than Vpb4 homologs. These unique Vpb4 domains adopt the topologies of carbohydrate-binding modules known to participate in receptor-recognition. Functional assessment of Vpb4Da2 suggests that domains 4–6 comprise the WCR receptor binding region and are key in conferring the observed insecticidal activity against WCR. The current structural analysis was complemented by in vitro and in vivo characterizations, including immuno-histochemistry, demonstrating that Vpb4Da2 follows a MOA that is consistent with well-characterized 3-domain Bt insecticidal proteins despite significant structural differences.

Fused: a promising molecular target for an RNAi-based strategy to manage Bt resistance in Plutella xylostella (L.)

Bacillus thuringiensis (Bt), a naturally occurring entomopathogenic soil bacterium, has been the active ingredient of sprayable Bt biopesticides for over a century. Insecticidal Bt Cry proteins are particularly well suited for use as plant-incorporated protectants in transgenic crops (Bt crops) due to their specificity against insect pests and safety for non-target organisms. The sustainability of these Bt products, however, has been challenged by the development of resistance in the field. RNA interference (RNAi), a species-specific control alternative that has been deregulated and commercialized in the USA and Canada, provides a new mode of action to complement the existing Bt products. Based on our preliminary research, we hypothesized that pyramiding/integrating Bt with RNAi can address/manage resistance issues related to Bt traits. To examine this overarching hypothesis, we (1) cloned and characterized a serine/threonine kinase gene (fused) of the Hedgehog (Hh) signaling pathway in the diamondback moth, Plutella xylostella (L.), a global superpest; (2) profiled Pxfused expression in Bt-resistant P. xylostella strains; and (3) investigated the involvement of Pxfused in Bt Cry1Ac resistance in P. xylostella. Pxfused expression was elevated ubiquitously in all Bt-resistant strains, and silencing of Pxfused led to larval and pupal mortality in both Cry1Ac-susceptible and -resistant strains, suggesting that Pxfused is a potential target for RNAi-based resistance management. Taken together, our results not only identify a molecular target to control a devastating lepidopteran pest, but also shed light on a novel resistance management strategy through the integration of two biotechnological techniques with distinct modes of action.

Resistance to Bt Maize by Western Corn Rootworm: Effects of Pest Biology, the Pest-Crop Interaction and the Agricultural Landscape on Resistance.

Simple SummarySince the 1990s, an important innovation in the management of agricultural pest insects has been the commercial cultivation of genetically engineered crops that produce insecticidal toxins, which in turn act to protect plants from feeding injury by insects. To date, these transgenic crops, which include cotton, maize and soybean, have produced insecticidal proteins derived from the bacterium Bacillus thuringiensis (Bt). Benefits associated with planting of Bt crops include reduced feeding injury from pest insects, decreased yield losses from pests and less harm to the environment. However, the evolution of Bt resistance by insect pests can diminish these benefits. One serious insect pest currently managed with Bt maize is the western corn rootworm. The larval stage of this insect feeds on maize roots and can substantially reduce yield. In some parts of the US Corn Belt, western corn rootworm rapidly adapted to Bt maize, and currently, some populations show resistance to all commercially available Bt traits. This review summarizes the time course of resistance development in the field, key factors contributing to resistance evolution, and steps that biotechnology companies, farmers and regulatory agencies can take to delay additional cases of pest resistance to current and future transgenic technologies.The western corn rootworm, Diabrotica virgifera virgifera LeConte, is among the most serious pests of maize in the United States. Since 2003, transgenic maize that produces insecticidal toxins from the bacterium Bacillus thuringiensis (Bt) has been used to manage western corn rootworm by killing rootworm larvae, which feed on maize roots. In 2009, the first cases of field-evolved resistance to Bt maize were documented. These cases occurred in Iowa and involved maize that produced Bt toxin Cry3Bb1. Since then, resistance has expanded to include other geographies and additional Bt toxins, with some rootworm populations displaying resistance to all commercially available Bt traits. Factors that contributed to field-evolved resistance likely included non-recessive inheritance of resistance, minimal fitness costs of resistance and limited adult dispersal. Additionally, because maize is the primary agricultural crop on which rootworm larvae can survive, continuous maize cultivation, in particular continuous cultivation of Bt maize, appears to be another key factor facilitating resistance evolution. More diversified management of rootworm larvae, including rotating fields out of maize production and using soil-applied insecticide with non-Bt maize, in addition to planting refuges of non-Bt maize, should help to delay the evolution of resistance to current and future transgenic traits.

Populations of Helicoverpa zea (Boddie) in the Southeastern United States are Commonly Resistant to Cry1Ab, but Still Susceptible to Vip3Aa20 Expressed in MIR 162 Corn.

sThe corn earworm, Helicoverpa zea (Boddie), is a major pest targeted by pyramided Bacillus thuringiensis (Bt) corn and cotton in the U.S. Cry1Ab is one of the first insecticidal toxins used in Bt crops, while Vip3A is a relatively new toxin that has recently been incorporated into Cry corn with event MIR 162 and Cry cotton varieties to generate pyramided Bt traits targeting lepidopteran pests including H. zea. The objectives of this study were to determine the current status and distribution of the Cry1Ab resistance, and evaluate the susceptibility to Vip3Aa20 expressed in MIR 162 corn in H. zea in the southeastern U.S. During 2018 and 2019, 32 H. zea populations were collected from non-Bt corn (19 populations), Cry corn (12), and Cry/Vip3A cotton (1) across major corn areas in seven southeastern states of the U.S. Susceptibility of these populations to Cry1Ab and Vip3Aa20 was determined using diet-overlay bioassays. Compared to a known susceptible insect strain, 80% of the field populations were 13- to >150-fold resistant to Cry1Ab, while their response to Vip3Aa20 ranged from >11-fold more susceptible to 9-fold more tolerant. Mean susceptibility to each Bt toxin was not significantly different between the two groups of the populations collected from non-Bt and Bt crops, as well as between the two groups of the populations collected during 2018 and 2019. The results show that resistance to Cry1Ab in H. zea is widely distributed across the region. However, the Cry1Ab-resistant populations are not cross-resistant to Vip3Aa20, and H. zea in the region is still susceptible to the Vip3Aa20 toxin. Vip3Aa20 concentrations between 5 and 10 µg/cm2 may be used as diagnostic concentrations for susceptibility monitoring in future. Additional studies are necessary to elucidate the impact of the selection with Bt corn on resistance evolution in H. zea to Vip3A cotton in the U.S.

Needs Assessment for Corn Insect Pest Management in Iowa

AbstractCorn, Zea mays L. (Cyperales: Poaceae), production is essential to Iowa’s economy and provides food, feed, and fuel beyond the state. Farmers face a number of challenges in corn production, including insect pests, and understanding farmers’ perceptions and concerns is critical for extension personnel to provide adequate and relevant programming. The Iowa State University Agriculture and Natural Resources Crops Team conducted a survey of Iowa farmers and agribusiness professionals during the 2019–2020 extension season. The goal was to understand farmer perceptions of corn pests and whether farmers experienced challenges with resistance to Bt traits or insecticides. Additionally, we aimed to realize farmers’ confidence in developing resistance management plans and discussing those needs with their input suppliers and advisors. Our survey revealed that northern corn rootworm, Diabrotica barberi Smith & Lawrence (Coleoptera: Chrysomelidae), was the pest of greatest concern across the state. People were generally more concerned about belowground Bt trait performance than aboveground traits, though only 12% of respondents reported Bt trait failure for corn rootworms. In general, people were not concerned about the performance of seed treatments or soil- or foliar-applied insecticides. Farmers generally felt very confident in developing a resistance management plan and discussing their needs with their advisors. The results of this survey will be used to target extension programming throughout the state and increase confidence for farmers and agribusiness employees to discuss resistance management plans together.

Bt cotton seed purity in Burkina Faso: status and lessons learnt

BackgroundSince the commercial release of Bt cotton in Burkina Faso in 2009, the issue of seed purity in producers’ fields has rarely been addressed in an unbiased and objective manner. The potential for contamination of conventional seed varieties with Bt traits and the consequent threat to the continuation of organic cotton production has been documented. However, studies are rare on the varietal purity of Bt cotton seeds, despite the implications for the effectiveness and sustainability of their use.This paper compensates for the lack of research on the varietal purity of cotton seeds in Burkina Faso by reporting the results of Enzyme linked immunosorbent assay tests collected in 2015 on samples of both conventional and Bt varieties from 646 fields.ResultsAccording to the conservative criteria used to declare the presence of a Bt gene in a given variety (more than 10% of seeds of conventional variety exhibit Bt traits, and at least 90% of seeds of Bt variety exhibit Bt traits), seed purity was very questionable for both types of variety. For the supposedly conventional variety, the Cry1Ac gene was observed in 63.6% of samples, the Cry2Ab gene was observed in 59.3% of samples, and both genes were detected in 52.2% of the seed samples. Only 29.3% of the seeds that were supposed to be of conventional type contained no Bt genes. Conversely, for the labeled Bt variety, the Cry1Ac gene was found in only 59.6% of samples, the Cry2Ab gene was found in 53.6% of the samples, and both genes were found in 40.4% of the samples. Finally, for the seeds that were supposed to contain both genes (Bollguard 2), both Cry1Ac and Cry2Ab genes were found in only 40.4% of the samples, only one of the genes was found in 32.4% of the samples, and 27.2% of the seeds in the samples contained neither.Two factors are responsible for the severe lack of seed purity. First, conventional varieties are being contaminated with Bt traits because of a failure to revise the seed production scheme in Burkina Faso to prevent cross-pollination. Second, the original Bt seeds provided to Burkina Faso lacked varietal purity.The organic sector plays a very minor role in the cotton sector of Burkina Faso (production of organic cotton totaled 453 t in 2018/2019, out of national cotton production of 183 000 t). Nevertheless, the lack of purity in conventional seed varieties is a threat to efforts to expand certified organic cotton production. The poor presence of Bt proteins in supposed Bt varieties undermines their effectiveness in controlling pests and increases the likelihood of the development of resistance among pest populations.ConclusionOur results show the extent of purity loss when inadequate attention is paid to the preservation of seed purity. Pure conventional seeds could vanish in Burkina Faso, while Bt seeds do not carry the combination of the expected Bt traits. Any country wishing to embark on the use of Bt cotton, or to resume its use, as in the case of Burkina Faso, must first adjust its national seed production scheme to ensure that procedures to preserve varietal purity are enforced. The preservation of varietal purity is necessary to enable the launch or the continuation of identity-cotton production. In addition, the preservation of varietal purity is necessary to ensure the sustainable effectiveness of Bt cotton. In order to ensure that procedures to preserve varietal purity are observed, seed purity must be tested regularly, and test results must be published.

Applying a Selection Experiment to Test for Fitness Costs of Bt Resistance in Western Corn Rootworm (Coleoptera: Chrysomelidae) and the Effect of Density on Fitness Costs.

Western corn rootworm, Diabrotica virgifera virgifera LeConte, is a serious pest of corn and is often managed with transgenic corn producing insecticidal toxins from the bacterium Bacillus thuringiensis (Bt). This pest has developed field-evolved resistance to all commercially available Bt traits, beginning with Cry3Bb1 in 2009. Fitness costs may accompany Bt resistance, where individuals with alleles for Bt resistance have reduced fitness on non-Bt corn compared to Bt-susceptible individuals. In conjunction with non-Bt refuges, fitness costs can delay the evolution of Bt resistance. Importantly, ecological factors may affect the presence and magnitude of fitness costs. For western corn rootworm, available data suggest that fitness costs of Bt resistance may be present in some cases. Using two Cry3Bb1-resistant western corn rootworm strains (Hopkinton and Cresco), a fitness-cost experiment was performed by rearing rootworm in the absence of Bt for six generations to test for fitness costs of Cry3Bb1 resistance and the effect of larval rearing density on fitness costs. Fitness costs were detected for both strains; however, strains were still resistant to Cry3Bb1 corn at the end of the experiment. Cresco experienced a greater loss of resistance at low versus high density, but no effect of density was detected in Hopkinton. Our study shows that fitness costs can accompany Bt resistance in western corn rootworm and may be more pronounced under low larval density. Even though fitness costs were present, it appears that rootworm populations may remain resistant to Cry3Bb1 corn for years after resistance has evolved.

Baseline Susceptibility of a Laboratory Strain of Northern Corn Rootworm, Diabrotica barberi (Coleoptera: Chrysomelidae) to Bacillus thuringiensis Traits in Seedling, Single Plant, and Diet-Toxicity Assays

The northern corn rootworm (NCR), Diabrotica barberi Smith & Lawrence, is an economic pest of maize in the U.S. Corn Belt. The objective of this study was to determine the baseline susceptibility of a laboratory NCR strain to Bt proteins eCry3.1Ab, mCry3A, Cry3Bb1, and Cry34/35Ab1 using seedling, single plant, and diet-toxicity assays. Plant assays were performed in greenhouse using corn hybrids expressing one of the Bt proteins and each respective near-isoline. Diet-toxicity assays, consisting of Bt proteins overlaid onto artificial diet were also conducted. In both plant assays, significantly more larvae survived Cry34/35Ab1-expressing corn compared with all other Bt-expressing corn, and larvae that survived eCry3.1Ab-expressing corn had significantly smaller head capsule widths compared with larvae that survived Cry34/35Ab1-expressing corn. In seedling assays, larvae surviving eCry3.1Ab-expressing corn also had significantly smaller head capsule widths compared with larvae that survived mCry3A-expressing corn. Additionally, larvae that survived mCry3A-expressing corn weighed significantly more than larvae surviving eCry3.1Ab- and Cry34/35Ab1-expressing corn. In single plant assays, no significant differences in larval dry weight was observed between any of the Bt-expressing corn. In diet assays, LC50s ranged from 0.14 (eCry3.1Ab) to 10.6 µg/cm2 (Cry34/35Ab1), EC50s ranged from 0.12 (Cry34/35Ab1) to 1.57 µg/cm2 (mCry3A), IC50s ranged from 0.08 (eCry3.1Ab) to 2.41 µg/cm2 (Cry34/35Ab1), and MIC50s ranged from 2.52 (eCry3.1Ab) to 14.2 µg/cm2 (mCry3A). These results establish the toxicity of four Bt proteins to a laboratory diapausing NCR strain established prior to the introduction of Bt traits and are important for monitoring resistance evolution in NCR field populations.

Comparing Populations of Western Corn Rootworm (Coleoptera: Chrysomelidae) in Regions With and Without a History of Injury to Cry3 Corn.

Transgenic corn expressing insecticidal proteins derived from the bacterium Bacillus thuringiensis (Bt) is an important pest management tool. Western corn rootworm, Diabrotica virgifera virgifera LeConte, is a key pest of corn in the midwestern United States that has developed field-evolved resistance to all available Bt traits. The first Bt trait to be commercialized for management of rootworm was Cry3Bb1 in 2003, and field-evolved resistance appeared in 2009. In this study, we examined fields in counties where greater-than-expected injury to Cry3 (Cry3Bb1 or mCry3A) corn roots (>1 node) had previously been reported (problem counties) and counties where injury had not been reported (non-problem counties). Four to eight fields were sampled per county in 2015, 2016, and 2017 to quantify rootworm abundance, root injury, Cry3Bb1resistance, and rootworm management strategies. Rootworm abundance, root injury, and resistance to Cry3Bb1 did not differ between county types. Management tactics differed between county types, with problem counties growing more corn, using more soil insecticide, and growing more Cry34/35Ab1 corn. Additionally, a comparison of root injury to Bt and non-Bt corn within fields indicated that farmers derived an economic benefit from planting Bt corn to manage corn rootworm. Our results suggest that rootworm populations are similar between problem and non-problem counties in Iowa due to similar levels of selection pressure on Cry3 corn, but problem county fields have applied more management tactics due to previous rootworm issues in the area.