Bt Maize Research Articles

Incipient resistance of the African maize stemborer, Busseola fusca (Lepidoptera: Noctuidae) to Cry2Ab2 maize in South Africa.

Resistance of Busseola fusca (Lepidoptera: Noctuidae) to Cry1Ab was documented in 2006, 7 years after the first cultivation of MON 810 in South Africa. This was mitigated by introducing a second-generation Bacillus thuringiensis (Bt) maize (MON 89034), which contains the Cry1A.105 and Cry2Ab2 proteins. The first reports of B. fusca infestations of MON 89034 maize came in the KwaZulu-Natal province (2017-2018 cropping season), followed by reports in the Mpumalanga province (2022-2023 season). Here we report results of artificial diet- and plant-based laboratory assays to assess the susceptibility of B. fusca populations to the Bt proteins in MON 89034. Larvae were sampled from nine locations which included three where greater than expected injury due to B. fusca had been reported to MON 89034. Larval mortality in assays with diet-incorporated Cry2Ab2 protein was 100% for all except the three problem populations, showing that the Cry2Ab2 protein in MON 89034 is highly efficacious against B. fusca. In contrast, assays with Cry1A.105 did not cause significant mortality in any of the B. fusca populations including larvae from a susceptible reference. Larval survival on leaf tissue of MON 89034 maize after 7 days ranged between 75% and 91% for the three problematic B. fusca populations, compared to 0.4% to 9.6% for the five other populations. Therefore, MON 89034 is effectively a single-mode-of-action technology against B. fusca and carries an inherent high risk for the evolution of resistance. This study shows that the three B. fusca populations collected from locations with greater than expected damage to MON 89034 have resistance to the Cry2Ab2 protein and therefore to MON 89034 maize. This research emphasizes the importance of resistance monitoring and implementation of effective insect resistance management tactics. © 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

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.

Impact of dual Bt-transgenic maize (2A7) on soil microbial communities and enzyme activities: A comparative study with control variety Z58

The impacts of transgenic crops on soil microbiology and fertility are critical in determining their biosafety. While transgenic crops can alter soil microbes, their effects are often context-dependent; therefore, the ecological importance of these changes remains a topic of ongoing research. Using high-throughput sequencing, we investigated the effects of Bacillus thuringiensis (Bt) maize expressing the mcry1Ab and mcry2Ab genes (2A7) on soil nutrient dynamics, as well as the diversity and function of soil microbial communities, including bacteria and fungi, within different soil compartments. Our findings revealed a plant-shaped rhizosphere (RS) microbial community as a result of the selective recruitment of microorganisms from the surrounding environment. The transgene insertion had a significant impact on the RS niche, and several species eventually became associated with Z58 and 2A7 plants. For example, Neocosmospora rubicola fungal and Pantoea dispersa bacterial microorganisms were significantly decreased in the dual Bt-transgenic 2A7 rhizosphere but enriched in the Z58 rhizospheres. The activity of soil enzymes such as urease, invertase, and alkaline phosphatase was boosted by Bt-transgenic 2A7. LefSe analysis identified significant bacterial and fungal biomarker species that were responsible for the differential effects of Bt-transgenic 2A7 and control Z58 within rhizosphere soils. Mantel analysis further demonstrated that the root exudates of 2A7 altered nutrient-acquisition enzymes by influencing biomarker taxa. PICRUSt2 functional characterization revealed a significantly higher abundance of the phosphate-starvation-inducible protein in control Z58 than in Bt-transgenic 2A7. Furthermore, taxonomy, alpha (Shannon diversity), and beta diversity analyses all revealed niche-driven microbial profile differentiation. Niche partitioning also had a significant impact on N- and P-related COGs as well. Our findings suggests that Bt-transgenic 2A7 modulates rhizosphere microbial communities by affecting biomarker taxa and soil enzyme activity. These findings will promote sustainable agriculture practices by advancing our knowledge of the ecological effects of Bt crops on soil microbial communities.

Toxic Effects of Bt-(Cry1Ab+Vip3Aa) Maize (“DBN3601T’’ Event) on the Asian Corn Borer Ostrinia furnacalis (Guenée) in Southwestern China

Asian corn borer (ACB), Ostrinia furnacalis, is an important agricultural pest affecting maize production in southwestern China, but knowledge of the toxic effect of Bt maize on the pest has been insufficient until now. In this study, we determined the susceptibility of ACB to Cry1Ab, Vip3Aa, and their complex proteins and evaluated the efficacy of Chinese domestic Bt-(Cry1Ab+Vip3Aa) maize (“DBN3601T” event) against the pest in Yunnan Province of southwestern China. The susceptible bioassay indicated that the LC50 values of the Cry1Ab and Cry1Ab+Vip3Aa proteins expressed by the Bt maize varieties against ACB larvae were 51.42 and 46.85 ng/g, respectively; however, the ACB larva was insensitive to the Vip3Aa protein. The Cry1Ab+Vip3Aa protein contents in V6–V8 leaves, VT tassels, R1 silks, R2 kernels, R3 stalks and R3 cobs of the Bt-(Cry1Ab+Vip3Aa) maize were 114.20, 30.69, 3.77, 8.92, 11.09 and 10.99 μg/g, respectively. The larval feeding test indicated that the Bt-(Cry1Ab+Vip3Aa) maize was more toxic to the early instar larvae, and the survival time of larvae fed on the leaves was the shortest, while it survived the longest on stalks. The identification of maize resistance levels in the field showed that both larval density and plant damage score of Bt-(Cry1Ab+Vip3Aa) maize were significantly lower than those in conventional maize. It is concluded that the Bt-(Cry1Ab+Vip3Aa) maize can be used for control of the ACB in southwestern China.

Control Efficacy of the Bt Maize Event DBN3601T Expressing Cry1Ab and Vip3Aa Proteins against Beet Armyworm, Spodoptera exigua (Hübner), in China.

The beet armyworm, Spodoptera exigua (Hübner), is a major pest of maize, cotton, soybean, and many other crops globally. Despite the widespread deployment of Bt transgenic maize for pest control worldwide, the efficacy of Bt lepidopteran-resistant transgenic maize in managing S. exigua remains rarely studied. In this study, we quantified the expression level of pyramided Cry1Ab and Vip3Aa toxins in Bt maize (event DBN3601T) and evaluated their control efficiency against S. exigua under both laboratory and field conditions. The enzyme-linked immunosorbent assay (ELISA) results showed that the expression levels of Cry1Ab and Vip3Aa proteins in DBN3601T maize tissues followed a decreasing order as follows: V5-leaf > V8-leaf > VT-tassel > R2-kernel > R1-silk. Diet-overlay assay results showed that the LC50 values of Cry1Ab and Vip3Aa proteins against S. exigua larvae were 11.66 ng/cm2 and 27.74 ng/cm2, respectively, with corresponding GIC50 values at 1.59 ng/cm2 and 7.93 ng/cm2. Bioassay using various tissues of the DBN3601T maize indicated that after 7 days of infestation, mortality rates of neonates and third-instar larvae ranged from 86% to 100% and 58% to 100%, respectively. Mortality was highest on V5 and V8 leaves, followed by R2-kernel, VT-tassel, and R1-silk. Field trials demonstrated that DBN3601T maize exhibited significantly lower larval density, damage rate, and leaf damage score compared to non-Bt maize. Field cage trial showed that the control efficacy of DBN3601T maize at the vegetative stage could reach 98%. These findings provide a theoretical basis for utilizing Bt transgenic maize to enhance the sustainable management of S. exigua in Asia.

An extended investigation on allele frequency of resistance to Bt maize in the southwestern corn borer and sugarcane borer (Lepidoptera: Crambidae) in the mid-south region of the United States

The southwestern corn borer, Diatraea grandiosella Dyar, and sugarcane borer, Diatraea saccharalis (F.) (Lepidoptera: Crambidae), are two important stalk borers targeted by Bacillus thuringiensis (Bt) maize in the Americas. Practical resistance to Bt maize in D. grandiosella is emerging in the southwest region of the United States. In this study, Bt maize leaf tissue was used to screen F2 family-lines of these two stalk borers for resistance alleles (RA) to Bt maize expressing one or two of the four common Bt proteins: Cry1A.105, Cry2Ab2, Cry1F, and Cry1Ab. The family-lines were developed from single-pairing or group-mating of field-collected D. grandiosella from non-Bt maize in Tennessee and D. saccharalis from sugarcane in Louisiana during 2019–2022. The F2 screen identified one possible major RA to Cry1A.105 maize in 155 D. grandiosella and 198 D. saccharalis individuals collected from fields, while for the maize expressing one or two of the other three Bt proteins examined, no major RAs were detected in 94–208 field-collected individuals of the two species. Bayesian analysis showed that an upper-level resistance allele frequency (RAF) for major alleles to Cry1A.105 maize was 0.0063 in D. grandiosella and 0.0050 in D. saccharalis. With 95% probability, the RAF of major alleles in D. grandiosella was <0.0093 for Cry2Ab2 maize, <0.0155 for Cry1F maize, and <0.0150 for Cry1A/Cry2A maize; and for D. saccharalis, it was <0.0071 for Cry2Ab2 maize and <0.0090 for Cry1Ab maize. These findings suggest that RAFs for major alleles to the common Bt maize are still low in the two stalk borers. However, minor RAs in both species, especially to Cry2Ab2 and Cry1A.105 maize in D. saccharalis, were not rare. The results of this study, together with other data, suggest that, after 25 years of Bt maize use, the technology is still effective and field resistance to maize expressing the Cry proteins has not evolved in these two stalk borers in the mid-south region of the United States.

Maize Kernel-Associated Metagenomes Reveal Potential Microbe–Microbe Interactions that Underlie Fusarium Ear Rot Disease

As an alternative to host resistance, transgenic expression of entomocidal and antimicrobial proteins from Bacillus thuringiensis ( Bt) in maize can mitigate Fusarium ear rot (FER). This study evaluated FER in Bt and conventional maize and the role of microbes in the kernel-associated metagenome using quantitative reduced representation sequencing. Our results revealed significant differences in FER severity across environments and varieties and between inoculation treatments. The lower FER scores of conventional maize Spectrum 6416 relative to other varieties highlighted resistant alleles in maize germplasm, whereas strong correlations indicated FER-induced yield loss. Mitigation of FER by transgenes was validated by about threefold enrichment of Fusarium verticillioides ( Fv) postinoculation compared with nonsignificant Fv enrichment in conventional maize. Although the major causal pathogen of FER, Fv, was the most abundant species in the metagenomes ( r = 0.41 to 0.49), a similar degree of correlation was observed between FER and several Fusarium spp. ( r = 0.2 to 0.56). The potential FER-suppressing properties of Talaromyces stipitatus and Ustilago maydis were indicated by negative correlations with FER ( r = –0.22 to –0.42), Fv, and some Fusarium spp. The more FER-resistant varieties consistently enriched for potential FER-suppressing Burkholderia cenocepacia (negatively correlated with Fv). This suggests host genetic background-dependent recruitment of beneficial microbes that suppress pathogens and that microbe–microbe interactions play a role in FER severity. Quantitative and species/strain-level metagenomic profiles hold promise for identifying robust disease-suppressing microbes and microbes that act in synergy with pathogens and for developing holobiont-aware breeding strategies that account for host–microbiome coevolution and host genotype, microbiome/metagenome, and environment interactions. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Fall armyworm (Lepidoptera: Noctuidae): practical resistance of 2 Brazilian populations to Cry1A.105 + Cry2Ab and Cry1F Bt maize.

In the Americas, transgenic cropsproducing insecticidal proteins fromBacillus thuringiensis Berliner (Bt,Bacillales: Bacillaceae) have been used widely to manage fall armyworm (FAW, Spodoptera frugiperda [J.E. Smith]). As resistance to Cry1 single-gene Bt maize (Zea mays L.)rapidly evolved in some FAW populations, pyramided Bt maizehybrids producing Cry1, Cry2, or Vip3Aa proteins were introduced in the 2010s. We examined field-evolved resistance to single- and dual-protein Bt maize hybrids in 2 locations in southeastern Brazil, where plant damage by FAW larvae far exceeded the economic threshold in 2017. We collected late-instar larvae in Cry1A.105 + Cry2Ab and Cry1F maize fields and established 2 FAW populations in the laboratory. The F1 offspring reared on the foliage of Bt and non-Bt maize plants (Cry1A.105 + Cry2Ab and Cry1F) showed neonate-to-adult survival rates as high as 70% for both populations. There was no significant difference in the life-table parameters of armyworms reared on non-Bt and Bt maize foliage, indicating complete resistance to Cry1A.105 + Cry2Ab maize. Larval survival rates of reciprocal crosses of a susceptible laboratory strain and the field-collected populations indicated nonrecessive resistance to Cry1F and a recessive resistance to Cry1A.105 + Cry2Ab maize. When relaxing the selection pressure, the armyworm fitness varied on Cry1A.105 + Cry2Ab and non-Bt maize; the resistance was somewhat stable across 12 generations, without strong fitness costs, although one of the lines died confounded by a depleted-quality, artificial rearing diet. To our knowledge, this is the first report documenting the practical resistance of FAW to a pyramided Bt crop. We discuss the implications for resistance management.

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.

Field evaluation the effect of two transgenic Bt maize events on predatory arthropods in the Huang-Huai-Hai summer maize-growing region of China.

To illustrate the impact of genetically modified (GM) Bt maize on the natural enemy communities in the Huang-Huai-Hai summer maize-growing region in China, the abundance of 7 common predator taxa (Geocoris pallidipennis Costa, Harmonia axyridis (Pallas), lacewings, Orius sauteri (Poppius), Propylea japonica (Thunberg), spiders, and Staphylinidae) was quantitatively evaluated by comparing Bt-Cry1Ab DBN9936 and Bt-Cry1Ab/Cry2Aj Ruifeng 125 events to their near non-Bt isolines during the growing season from 2016 to 2019. A total of 10,302, 19,793, 13,536, and 5,672 individuals were observed during 4 years, and the abundance of each taxa on Bt maize varied between sample dates among those arthropod taxa. Shannon-Wiener diversity index of predator communities from 7 taxa showed very similar temporal dynamics and principal response curve analyses to examine community-level effects showed no significant differences in predator abundance in Bt maize compared with non-Bt maize. We conclude that the 2 Bt maize hybrids did not adversely affect the predator community in the Huang-Huai-Hai summer maize-growing region of China.

Insecticidal Effects of Transgenic Maize Bt-Cry1Ab, Bt-Vip3Aa, and Bt-Cry1Ab+Vip3Aa against the Oriental Armyworm, Mythimna separata (Walker) in Southwest China.

The oriental armyworm, Mythimna separata (Walker), an important migratory pest of maize and wheat, is posing a severe threat to maize production in Asian countries. As source areas of spring-summer emigratory populations, the control of M. separata in southwestern China is of great significance for East Asian maize production. To assess the toxicity of Bt maize against the pest, bioassays of Bt-(Cry1Ab+Vip3Aa) maize (event DBN3601T), Bt-Cry1Ab maize (event DBN9936), and Bt-Vip3Aa maize (event DBN9501) were conducted in Yunnan province of southwest China. There were significant differences in insecticidal activity between the three Bt maize events, and DBN3601T presented the highest insecticidal role. The results also indicated that the insecticidal effect of various Bt maize tissues took an order in leaf > kernel > silk, which is highly consistent with the expression amounts of Bt insecticidal protein in leaf (69.69 ± 1.18 μg/g), kernel (11.69 ± 0.75 μg/g), and silk (7.32 ± 0.31 μg/g). In field trials, all larval population densities, plant damage rates, and leaf damage levels of DBN3601T maize were significantly lower than the conventional maize. This research indicated that the DBN3601T event had a high control efficiency against M. separata and could be deployed in southwest China for the management of M. separata.

Susceptibility of Fall Armyworm Field Populations to Vip3Aa/Cry Bt Maize in a Tropical Agricultural Region

Fall armyworm (FAW, Spodoptera frugiperda) is a polyphagous and migratory lepidopteran pest insect in field crops and is notoriously invasive worldwide. In large portions of the Americas, its populations are managed using transgenic maize or cotton varieties producing insecticidal proteins from Bacillus thuringiensis (Bt), primarily Vip3Aa pyramided with Cry Bt proteins. We determined the susceptibility of FAW field populations from locations pressured with such maize hybrids for over five years. We used time–mortality bioassays with F1 third-instar larvae of six geographically distinct populations collected in maize fields of a tropical agricultural region encompassing four Brazilian states. We maintained the neonate progeny from the field populations on an artificial diet until the third instar, and then determined their survival curves on the foliage of three Vip3Aa/Cry-producing Bt maize hybrids. Death of the mid-size, third-instar FAWs occurred relatively rapidly, with larval mortality rates reaching 98–100% in less than five days regardless of Bt maize hybrid. However, median survival time (ST50) for the larvae differed among the populations, with the lowest and highest ST50 values occurring for PI-Cr (42 h, 1.75 d) and PI-Ur populations (66–90 h, 2.75–3.75 h), respectively. Therefore, the F1 third-instar larvae of FAW populations were largely susceptible to Vip3Aa/Cry-producing maize foliage, and the most contrasting susceptibility occurred in the insects from Piauí state, Brazil. These results indicate that progeny of FAWs from areas highly pressured with Vip3Aa/Cry Bt maize hybrids are killed on maize foliage producing Vip3Aa and Cry Bt proteins despite field reports of increased leaf damage by the larvae in some locations. This research informs decision making for Bt-crop resistance management by producers, technicians, and researchers in local, regional, and world agriculture.

Pressure from insect-resistant maize on protected butterflies and moths.

Intensification in agriculture affects many insect species, including butterflies. Insect-resistant crops, such as Bt (Bacillus thuringiensis) maize, which produces a toxin active against Lepidoptera, are an alternative to insecticide sprays. Genetically modified crops are regulated in most countries and require an environmental risk assessment. In the European Union, such assessments include the use of simulation models to predict the effects on nontarget Lepidoptera (NTL). To support the assessment of protected NTL, we extended an individual-based, stochastic, spatially explicit mathematical model (LepiX) to include a wider range of exposure scenarios, a species-sensitivity distribution, and an option for repeated exposure of individuals. We applied the model to transgenic maize DAS-1507, which expresses a high concentration of Bt toxin in pollen that may be consumed by NTL larvae on their host plants nearby. Even in the most conservative scenario without repeated exposure, mortality estimates for highly sensitive species ranged from 41% to 6% at distances of 10-1000 m from the nearest maize field. Repeated exposure can cause additional mortality and thus is relevant for the overall risk assessment. Uncertainties in both exposure and ecotoxicity estimates strongly influenced the predicted mortalities. Care should be taken to include these uncertainties in the model scenarios used for decision-making. In accordance with other modeling results, our simulations demonstrated that mean mortality may not be safe for protected species. With its high pollen expression, DAS-1507 maize may pose risks to sensitive and protected butterfly and moth species that may be difficult to manage. High expression of Bt toxin in pollen is unnecessary for controlling target pests. Consequently, we suggest that Bt maize with high pollen expression not be cultivated in regions where protected butterflies are to be conserved.

Expression of Cry1Ab/2Aj Protein in Genetically Engineered Maize Plants and Its Transfer in the Arthropod Food Web

While transgenic Bacillus thuringiensis (Bt) maize provides pest resistance and a reduced application of chemical pesticides, a comprehensive environmental risk assessment is mandatory before its field release. This research determined the concentrations of Bt protein in plant tissue and in arthropods under field conditions in Gongzhuling City, northeastern China, to provide guidance for the selection of indicator species for non-target risk assessment studies. Bt maize expressing Cry1Ab/2Aj and non-transformed near-isoline were grown under identical environmental and agricultural conditions. Cry1Ab/2Aj was detected in plant tissues and arthropods collected from Bt maize plots during pre-flowering, flowering, and post-flowering. The expression of Cry1Ab/2Aj varied across growth stages and maize tissues, as well as in the collected arthropods at the three growth stages. Therefore, representative species should be chosen to cover the whole growing season and to represent different habitats and ecological functions. Dalbulus maidis (Hemiptera: Cicadellidae), Rhopalosiphum padi (Hemiptera: Aphididae), Heteronychus arator (Coleoptera: Scarabaeidae), and Somaticus angulatus (Coleoptera: Tenebrionidae) are suitable non-target herbivores. Propylea japonica (Coleoptera: Coccinellidae), Paederus fuscipes (Coleoptera: Staphylinidae), Chrysoperla nipponensis (Neuroptera: Chrysopidae), and spiders are suggested predators. Apis cerana and Apis mellifera ligustica (both Hymenoptera: Apidae) represent pollinators and Folsomia candida (Collembola: Isotomidae) decomposers.

Susceptibility of northern corn rootworm (Diabrotica barberi) populations to Cry3Bb1 and Gpp34/Tpp35Ab1 proteins in seedling and diet overlay toxicity assays.

The northern corn rootworm, Diabrotica barberi Smith & Lawrence (Coleoptera: Chrysomelidae) is a major pest of maize in the United States Corn Belt. Recently, resistance to Bacillus thuringiensis (Bt) maize was reported in North Dakota and increased use of Bt maize hybrids could facilitate resistance evolution in other maize-producing states. In this study, susceptibility to Bt proteins was evaluated in wild D. barberi populations from 8 fields collected in 5 different states (Minnesota, Missouri, Nebraska, Iowa, and North Dakota). Field populations were compared to a susceptible D. barberi colony in seedling and diet toxicity assays conducted with 3 concentrations of Cry3Bb1 (0.4, 4.0, and 40.0 µg/cm2) and Gpp34/Tpp35Ab1 (previously called Cry34/35Ab1; 1.4, 14.0, and 140.0 µg/cm2). The 2019 population from Meeker Co., Minnesota (MN-2019), exhibited the lowest mortality to Cry3Bb1 and also had nominally lowest mortality to Gpp34/Tpp35Ab1 at the highest concentrations tested in diet toxicity assays. Percent second instar was also highest for larvae of the Minnesota population surviving Cry3Bb1. In seedling assays, MN and IA-2018 populations exhibited the highest proportion survival and dry weight to both proteins expressed in corn. No significant differences in mortality, percent second instar, and dry weight were observed at the highest concentration for both proteins among the populations collected in in 2020. Most D. barberi populations were still highly susceptible to Cry3Bb1 and Gpp34/Tpp35Ab1 proteins based on diet and seedling assays, but resistance appears to be developing in some D. barberi populations. Now that methods are available, resistance monitoring may also be needed for D. barberi in some regions.

Effects of non-lethal Cry1F toxin exposure on the growth, immune response, and intestinal microbiota of silkworm (Bombyx mori)

Bt (Bacillus thuringiensis) maize is expected to be commercial cultivated widely in China. When Bt maize is planted near mulberry trees, it renders silkworms (Bombyx mori) vulnerable, as they belong to the same class as the Lepidoptera insects targeted by Bt maize. Cry1F has been found to be highly toxic to silkworms, particularly in their early larval stages. In this study, we aimed to assess the effects of non-lethal Cry1F exposure on the growth, immune response, and intestinal microbiota in silkworms. The results showed that feeding silkworms with mulberry leaves soaked in 100 μg/mL Cry1F for 96 h had an impact on larval body weight acquisition, leading to a decrease in cocoon and pupae weight. Cry1F exposure disrupted the intestinal integrity of silkworms by affecting the columnar cells of the midgut. The activity of detoxification enzymes (CarE, AChE, and GST) as well as antioxidant enzymes (SOD, CAT, and POD) were also affected by Cry1F. After 96 h Cry1F exposure, the evenness of the bacterial community was disrupted, resulting in alterations in the structure of the intestinal microbiota. Additionally, Cry1F exposure affected the relative expression levels of the peritrophic membrane (PM) protein and the corresponding immune pathways genes of silkworms. Most of the immune-related gene expressions were inhibited after exposure to Cry1F toxin but increased with prolonged treatment. This study demonstrates that non-lethal Cry1F exposure can affect the growth, immune response, and intestinal microbiota of silkworm.

Aflatoxin levels and Aspergillus species in maize in the Province of Isabela, Philippines

AbstractThe province of Isabela is the top maize producer in the Philippines. The intensive cultivation and the tropical climate in the region may favour fungal growth and aflatoxin contamination in maize grains. Thus, the study aimed to determine the occurrence of aflatoxin and mycotoxigenic Aspergillus species in maize varieties in this region. Samples were obtained in six municipalities from hybrid (Bt) maize (n = 101) and open‐pollinated varieties (n = 6) during the dry season (March and April) of 2019. Aflatoxin levels were quantified through enzyme‐linked immunosorbent assay, and Aspergillus species were identified through cultural and molecular methods. Aflatoxin was detected in 50.5% of maize samples; 49.5% of samples were less than the limit of detection (3 μg/kg), 16.8% with 3–20 μg/kg, 10.9% with 21–50 μg/kg and 22.8% above 50 μg/kg. Samples within the acceptable level were 66.3% for food (&lt;20 μg/kg) and 77.2% for animal feed (&lt;50 μg/kg), while 22.8% of samples were above the acceptable level of the Philippine National Standard for raw maize grains. More than 90% of Aspergillus species detected were A. flavus. Other species identified were A. tamarii and A. terreus. Despite the dry production season in the province with low relative humidity during harvesting, inadequate post‐harvest practices and the presence of A. flavus elevated the level of aflatoxin in sample grains. Additional work involving multi‐year surveys is needed to confirm the results and conclusions of this study.

Gut bacterial community and gene expression alterations induced by transgenic Bt maize contribute to insecticidal activity against Mythimna separata

Gut bacterial community and gene expression alterations induced by transgenic Bt maize contribute to insecticidal activity against Mythimna separata

Impact of Seed Blend and Structured Maize Refuge on Helicoverpa zea (Lepidoptera: Noctuidae) Potential Phenological Resistance Development Parameters in Pupae and Adults.

Helicoverpa zea, an economic pest in the southeastern U.S., has evolved practical resistance to Bt Cry toxins in maize and cotton. Insect resistance management (IRM) programs have historically required planting of structured non-Bt maize, but because of its low adoption, the use of seed blends has been considered. To generate knowledge on target pest biology and ecology to help improve IRM strategies, nine field trials were conducted in 2019 and 2020 in Florida, Georgia, North Carolina, and South Carolina to evaluate the impact of Bt (Cry1Ab + Cry1F or Cry1Ab + Cry1F + Vip3A) and non-Bt maize plants in blended and structured refuge treatments on H. zea pupal survival, weight, soil pupation depth, adult flight parameters, and adult time to eclosion. From a very large sample size and geography, we found a significant difference in pupal mortality and weight among treatments in seed blends with Vip3A, implying that cross-pollination occurred between Bt and non-Bt maize ears. There was no treatment effect for pupation depth, adult flight distance, and eclosion time. Results of this study demonstrate the potential impact of different refuge strategies on phenological development and survival of an important pest species of regulatory concern. This article is protected by copyright. All rights reserved.

Assessment of genetically modified maize Bt11 × MIR162 × MIR604 × MON 89034 × 5307 × GA21 and 30 subcombinations, for food and feed uses, underRegulation (EC) No 1829/2003 (application EFSA-GMO-DE-2018-149).

Genetically modified maize Bt11 × MIR162 × MIR604 × MON 89034 × 5307 × GA21 was developed by crossing to combine six single events: Bt11, MIR162, MIR604, MON 89034, 5307 and GA21, the GMO Panel previously assessed the 6 single maize events and 27 out of the 56 possible subcombinations and did not identify safety concerns. No new data on the single maize events or the assessed subcombinations were identified that could lead to modification of the original conclusions on their safety. The molecular characterisation, comparative analysis (agronomic, phenotypic and compositional characteristics) and the outcome of the toxicological, allergenicity and nutritional assessment indicate that the combination of the single maize events and of the newly expressed proteins in the six-event stack maize does not give rise to food and feed safety and nutritional concerns. The GMO Panel concludes that six-event stack maize, as described in this application, is as safe as the conventional counterpart and non-GM maize varieties tested, and no post-market monitoring of food/feed is considered necessary. In the case of accidental release of viable six-event stack maize grains into the environment, this would not raise environmental safety concerns. The GMO Panel assessed the likelihood of interactions among the single events in 29 of the maize subcombinations not previously assessed and covered by the scope of this application and concludes that these are expected to be as safe as the single events, the previously assessed subcombinations and the six-event stack maize. The post-market environmental monitoring plan and reporting intervals are in line with the intended uses of maize Bt11 × MIR162 × MIR604 × MON 89034 × 5307 × GA21. The GMO Panel concludes that six-event stack maize and the 30 subcombinations covered by the scope of the application are as safe as its conventional counterpart and the tested non-GM maize varieties with respect to potential effects on human and animal health and the environment.