Control efficacy and joint toxicity of broflanilide mixed with commercial insecticides to an underground pest, the black cutworm in highland barley.

BACKGROUND: The highland barley, Hordeum vulgare L., is a staple food crop with superior nutritional functions in Xizang, China. It is often damaged by the black cutworm, Agrotis ipsilon (Hufnagel), which is an underground pest and difficult to effectively manage. To introduce a novel insecticide with unique mode of action, broflanilide (BFL) and its binary mixtures with chlorantraniliprole (CAP), fluxametamide, ß-cypermethrin or imidacloprid were screened out as seed treatment to control black cutworm in highland barley in the present study. RESULTS: In the laboratory bioassays, BFL had outstanding insecticidal activity to black cutworm with a median lethal dose (LD50) of 0.07 mg kg-1. The mixture of BFL × CAP at the concentration ratio of 7:40 exhibited the highest synergistic effect with a co-toxicity coefficient of 280.48. In the greenhouse pot experiments, BFL and BFL × CAP seed treatments at 8 g a.i. kg-1 seed could effectively control black cutworm, with a low percentage of injured seedlings <20% and high control efficacies of 93.33-100% during a period of 3-12 days after seed emergence. Moreover, BFL and BFL × CAP seed treatments could promote the seed germination and seedling growth of highland barley at the tested temperatures of 15, 20 and 25 °C. CONCLUSION: Our results indicated that BFL and BFL × CAP were effective and promising insecticides as seed treatment to control black cutworm in highland barley. © 2024 Society of Chemical Industry.

Effects of chlorantraniliprole on the development, fecundity and prey consumption of a non-specific predator, Rhynocoris fuscipes (Hemiptera: Reduviidae).

Transplant treatment with chlorantraniliprole (CAP) is a proactive approach to protect transplanted plants from pests during early establishment and has been comprehensively applied in tobacco fields in Guangdong Province, China. However, it is not known whether the high dose of CAP in transplant treatments has lethal or sublethal effects on the generalist predator Rhynocoris fuscipes Fabricius (Hemiptera: Reduviidae). To address this concern, the mortalities of R. fuscipes were assessed when 2nd instar larvae of R. fuscipes were in direct contact with or consuming CAP and when their eggs were exposed to CAP. Furthermore, 2nd instar nymphs R. fuscipes were long-term exposed to CAP until they reached adulthood, and their life table parameters were determined. After exposure to CAP, the activity of detoxification enzymes (P450, CaeE and GST) and the functional respond of R. fuscipes to their preys Agrotis ipsilon larvae were determined. In this study, CAP at all concentrations did not significantly increase the mortality of 2nd instar of R. fuscipes nymphs in comparison with the control. The detoxification enzyme (P450, CarE and GST) activities and the number of A. ipsilon larvae consumed by R. fuscipes in the transplant treatment were not affected by CAP after 3-d or long-term exposure. These results indicated that CAP was harmless to R. fuscipes according to IOBC protocols. However, during the treatment of 2nd instar nymphs with a label rate of 15 g AI/ha and a 5× label rate of 75 g AI/ha, CAP significantly prolonged the pre-adult and pre-oviposition periods, and treated adults had lower oviposition. Attention should be given to the time interval between transplant treatment and the release of this biocontrol agent into the field to minimize the impact of CAP on the predator R. fuscipes.

Phenolics from Nigella sativa L. straw: Characterization and insecticidal activity against Agrotis ipsilon (Hüfnagel).

The excessive production of food and agro-waste has become a significant problem for society, the economy, and the environment. To meet the growing demand for food free from harmful synthetic insecticides, a recent study has investigated the potential use of an ethanolic extract obtained from the straw of Nigella sativa L., a byproduct of seed collection, as a bioinsecticide. The study also evaluated its in-vitro and in-silico acetylcholinesterase (AChE) inhibitory potential against the Agrotis ipsilon (Hufnagel) moth species, which is known to cause damage to various crops and ornamental plants. The high-performance liquid chromatography examination revealed that the ethanolic N. sativa straw extract contained 18 phenolics, including 3 simple phenols, 8 phenolic acids, and 7 flavonoids. Catechol (330.14 µg/ml), chlorogenic (169.23 µg/ml), and gallic (110.93 µg/ml) acids were the predominant phenolics. On the other hand, catechin (94.07 µg/ml), naringenin (91.99 µg/ml), and rutin (78.16 µg/ml) were the major flavonoids identified in the extract. The insecticidal activity of the extract against the 4th larval instar of A. ipsilon was evaluated using four concentrations (1.25-10 %). The study found that higher extract concentrations led to increased mortality in the larvae. Specifically, the concentration of 10 % resulted in the highest mortality rate of 96.67 %. Lower concentrations of 5 %, 2.5 %, and 1.25 % resulted in mortality rates of 51.11 %, 18.89 %, and 9.17 %, respectively. The extract also showed higher activity against AChE in larval tissue, with an inhibition percentage of 65.2 % after 24 h of treatment. Docking experiments confirmed that ellagic acid and apigenin had higher binding affinity than the control (lanate). These results demonstrate the potential of utilizing agricultural waste like N. sativa straw to create innovative and sustainable bioinsecticides.

Epsilon Class Glutathione S-Transferase Confers Phoxim Tolerance in the Black Cutworm Agrotis ipsilon (Lepidoptera: Noctuidae).

The black cutworm Agrotis ipsilon is a serious crop pest. Phoxim, an organophosphorus insecticide, has been widely used to control A. ipsilon. When phoxim is extensively applied, the susceptibility of A. ipsilon to insecticide is reduced. However, the mechanisms of tolerance of A. ipsilon to phoxim remain unclear. Herein, we report that an epsilon class glutathione S-transferase, AiGSTE1, confers phoxim tolerance in A. ipsilon. Exposure to a sublethal concentration (LC50) of phoxim caused oxidative stress and activated the transcription of AiGSTe1 genes in A. ipsilon larvae. Recombinant AiGSTE1 expressed in Escherichia coli could metabolize phoxim. Furthermore, E. coli cells overexpressing AiGSTE1 displayed significant tolerance to oxidative stress. Knockdown of AiGSTe1 by RNA interference significantly increased the mortality of A. ipsilon larvae to phoxim. These results demonstrate that AiGSTE1 confers phoxim tolerance in A. ipsilon by metabolizing the insecticide and preventing phoxim-induced oxidative stress.

Impact of gut microbiota composition on black cutworm, Agrotis ipsilon (hufnagel) metabolic indices and pesticide degradation.

Endosymbionts are known to have significant effects on their insect hosts, including nutrition, reproduction, and immunity. Insects gut microbiota is a critical component that affects their physiological and behavioral characteristics. The black cutworm (BCW), Agrotis ipsilon, is an economically important lepidopteran pest that has a diverse gut microbiome composed of nine species belonging to three phyla: Proteobacteria, Actinobacteria, and Firmicutes. This study was conducted to investigate the diversity of gut bacteria isolated from BCW larvae and moths and their effects on metabolism and pesticide degradation. The bacterial isolates were identified using the 16 S rRNA gene. The study showed that the gut microbiome composition significantly affected the metabolism of BCW larvae. Based on the screening results of synthesis of digestive enzymes and pesticide degradation, Brachybacterium conglomeratum and Glutamicibacter sp were selected to perform the remaining experiments as single isolates and consortium. The consortium-fed larvae showed high metabolic indices compared to antibiotic-fed larvae and the control. The gut bacteria were also shown to degrade three pesticide groups. Concerns regarding the health risk of chlorpyrifos have been raised due to its extensive use in agriculture. The isolated B. conglomeratum was more effective in chlorpyrifos degradation than the consortium. Furthermore, the study also examined the presence of sex related endosymbionts (Wolbachia, Spiroplasma, and Rickettsia) in the reproductive tissues of adults. The outcomes demonstrated that none of the examined endosymbionts existed. In conclusion, the study highlights the importance of the gut microbiome in insect physiology and behavior and its potential applications in biotechnology. It provides insights into developing eco-friendly pest control and bioremediation strategies using gut bacteria.

Effects of host plant on the development and reproduction of Agrotis ipsilon (Lepidoptera: Noctuidae) on horticultural crops.

One of cosmopolitan pest, Agrotis ipsilon (Lepidoptera: Noctuidae), causes serious economic damages in horticultural crops. This pest is difficult to manage and causes irreversible damage because its larvae stay in the ground at day and cut the plant stems at night. Thus, this study compared the host fitness of A. ipsilon among nine major horticultural crops in Korea. Among the nine crops, the population of A. ipsilon failed to complete its development in spinach, cucumber, melon, and kidney bean. The host effects on development and reproduction of A. ipsilon were further investigated in the remained five crops (i.e., napa cabbage, soybean, perilla, corn, and pepper). Host plants significantly (P < 0.05) affected the development-related factors (i.e., developmental time, survivorship, and weight) of A. ipsilon eggs, larvae, and pupae. They also affected the adult reproduction-related factors including preoviposition period, oviposition period and number, and longevity except for the prepupa stage. A positive relationship was found between biological factors (i.e., development- and reproduction-related factors). Among the nine crops in this study, napa cabbage showed the highest suitability for the A. ipsilon populations. These findings in this study would be helpful to understand the ecology and develop the management tactics of A. ipsilon in horticultural crops.

Diet impacts the reproductive system's maturation in the male moth Agrotis ipsilon (Noctuidae, Lepidoptera).

In male moth Agrotis ipsilon, sexual maturation occurs between the third and the fifth day of adult life and is characterized by the development of the reproductive organs such as testes and accessory sex glands. Since sexual maturation requires considerable energy investment, we hypothesized that diet would be an essential regulatory factor in this developmental process. Indeed, the links between the male diet and reproductive physiology have not been described as in females. To test the previous hypothesis, we offered male moths diets corresponding to different flower nectars found in nature, and measured morphological and functional changes in the testes and accessory sex glands. In comparison to a diet composed of sucrose only, males fed with a diet composed of diverse sugars, including glucose, supplemented with sodium led to an earlier increase in the length and the protein content of accessory sex glands, as well as a reduction of the testicular volume accompanied by an acceleration of the sperm bundle transfer from the testes to the duplex. These results show that these specific diets accelerate the maturation of the reproductive system in male moth Agrotis ipsilon.

Identification and Functional Characterization of Two Chitin Synthases in the Black Cutworm, Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae).

The black cutworm, Agrotis ipsilon (Hufnagel), a seasonal migrant and a prolific generalist, can feed on nearly all vegetables and grain crops, causing considerable economic impacts on a global scale. Given its cryptic nature, A. ipsilon management has been extremely challenging. Chitin synthase (CHS), a key enzyme involved in chitin biosynthetic pathway and crucially important for the growth and development of insects, is the molecular target of chitin synthesis inhibitors, a group of broad-spectrum insecticides that is compatible with Integrated Pest Management practices. In this study, we investigated the potential of targeting chitin synthases to control A. ipsilon. As a result, two chitin synthases, AiCHS1 and AiCHS2, were identified and cloned from A. ipsilon. The temporal-spatial distribution study showed that AiCHS1 was predominantly expressed at the pupal stage and most abundant among tissues of head capsule and integument, while AiCHS2 was mainly expressed at the sixth instar larval stage and tissues of foregut and midgut. RNAi-based functional study confirmed gene silencing caused significant reduction in the expression levels of the corresponding mRNA, as well as resulted in abnormal pupation and mortality, respectively. Furthermore, under the treatment of lufenuron, a chitin synthesis inhibitor, A. ipsilon responded with an elevated expression in AiCHS1 and AiCHS2, while larvae showed difficulty in shedding old cuticle, and a cumulative mortality of 69.24% at 48 h. In summary, chitin synthases are crucial for chitin biosynthesis in A. ipsilon and can be targeted for the control (e.g., RNAi-based biopesticides) of this devastating insect pest.

Methoprene-tolerant and Krüppel homolog 1 are actors of juvenile hormone-signaling controlling the development of male sexual behavior in the moth Agrotis ipsilon.

In insects, juvenile hormone (JH) is critical for the orchestration of male reproductive maturation. For instance, in the male moth, Agrotis ipsilon, the behavioral response and the neuronal sensitivity within the primary olfactory centers, the antennal lobes (ALs), to the female-emitted sex pheromone increase with fertility during adulthood and the coordination between these events is governed by JH. However, the molecular basis of JH action in the development of sexual behavior remains largely unknown. Here, we show that the expression of the paralogous JH receptors, Methoprene-tolerant 1 and 2 (Met1, Met2) and of the JH-inducible transcription factor, Krüppel homolog 1 (Kr-h1) within ALs raised from the third day of adult life and this dynamic is correlated with increased behavioral responsiveness to sex pheromone. Met1-, Met2- and Kr-h1-depleted sexually mature males exhibited altered sex pheromone-guided orientation flight. Moreover, injection of JH-II into young males enhanced the behavioral response to sex pheromone with increased AL Met1, Met2 and Kr-h1 mRNA levels. By contrast, JH deficiency suppressed the behavioral response to sex pheromone coupled with reduced AL Met1, Met2 and Kr-h1 mRNA levels in allatectomized old males and these inhibitions were compensated by an injection of JH-II in operated males. Our results demonstrated that JH acts through Met-Kr-h1 signaling pathway operating in ALs, to promote the pheromone information processing and consequently the display of sexual behavior in synchronization with fertility to optimize male reproductive fitness. Thus, this study provides insights into the molecular mechanisms underlying the hormonal regulation of reproductive behavior in insects.

Regulation of feeding dynamics by the circadian clock, light and sex in an adult nocturnal insect.

Animals invest crucial resources in foraging to support development, sustenance, and reproduction. Foraging and feeding behaviors are rhythmically expressed by most insects. Rhythmic behaviors are modified by exogenous factors like temperature and photoperiod, and internal factors such as the physiological status of the individual. However, the interactions between these factors and the circadian clock to pattern feeding behavior remains elusive. As Drosophila, a standard insect model, spends nearly all its life on food, we rather chose to focus on the adults of a non-model insect, Agrotis ipsilon, a nocturnal cosmopolitan crop pest moth having structured feeding activity. Our study aimed to explore the impact of environmental cues on directly measured feeding behavior rhythms. We took advantage of a new experimental set-up, mimicking an artificial flower, allowing us to specifically monitor feeding behavior in a naturalistic setting, e.g., the need to enter a flower to get food. We show that the frequency of flower visits is under the control of the circadian clock in males and females. Feeding behavior occurs only during the scotophase, informed by internal clock status and external photic input, and females start to visit flowers earlier than males. Shorter duration visits predominate as the night progresses. Importantly, food availability reorganizes the microstructure of feeding behavior, revealing its plasticity. Interestingly, males show a constant number of daily visits during the 5 days of adult life whereas females decrease visitations after the third day of adult life. Taken together, our results provide evidence that the rhythmicity of feeding behavior is sexually dimorphic and controlled by photoperiodic conditions through circadian clock-dependent and independent pathways. In addition, the use of the new experimental set-up provides future opportunities to examine the regulatory mechanisms of feeding behavior paving the way to investigate complex relationships between feeding, mating, and sleep-wake rhythms in insects.

Identification and RNAi-based functional analysis of chitinase family genes in Agrotis ipsilon.

BACKGROUND: Chitin is a major component in the extracellular matrix of insects, and its metabolism largely affects insect development and molting. As essential degradative enzymes, chitinases are encoded by multiple genes that differ in size, expression pattern and function in insects. However, our limited knowledge on the functions of different chitinases in Agrotis ipsilon has prevented our application of new technologies to target these genes as new pest management strategies. RESULTS: We revealed 11 full-length complementary DNA sequences of chitinase genes (AiChts) from A. ipsilon transcriptome. Although the domain architecture of these chitinases varied greatly, they all contained at least one chitinase catalytic domain. Developmental stage- and tissue-dependent expression profiles showed that most AiChts had the highest expression in the pupal stage. Furthermore, AiCht2, AiCht6, AiCht7 and AiCht10 were mainly expressed in the integument, whereas AiCht8 and AiCht-h had the highest expression in the midgut. The RNA interference (RNAi) experiment revealed that knockdown of AiCht10 or the imaginal disc growth factor gene (AiIDGF) induced high larval mortality. Larvae failed to shed the old cuticle during molting after the injection of double-stranded RNA targeting AiCht10 (dsAiCht10), whereas the larval bodies shrunk and blackened after the injection of dsRNA targeting AiIDGF (dsAiIDGF). CONCLUSION: Our results revealed for the first time the important functions of AiCht10 and AiIDGF in A. ipsilon. These genes are essential for larval development, and can potentially serve as new targets for RNAi-based pest management. © 2022 Society of Chemical Industry.

The Novel Agrotis ipsilon Nora Virus Confers Deleterious Effects to the Fitness of Spodoptera frugiperda (Lepidoptera: Noctuidae).

In the present study, we identified a novel, positive-sense single-stranded RNA virus in the Chinese black cutworm, Agrotis ipsilon. It has a genome length of 11,312 nucleotides, excluding the poly(A) tails, and contains five open reading frames. The ORF2 encodes the conserved domains of RNA helicase and RNA-dependent RNA polymerase, while ORF4 and 5 encode three viral proteins. Herein, the A. ipsilon virus was clustered with a Helicoverpa armigera Nora virus and was thus provisionally named "Agrotis ipsilon Nora virus" (AINV). AINV was successfully transmitted into a novel host, Spodoptera frugiperda, through injection, causing a stable infection. This found the possibility of horizontal AINV transmission among moths belonging to the same taxonomic family. Nonetheless, AINV infection was deleterious to S. frugiperda and mainly mediated by antiviral and amino acid metabolism-related pathways. Furthermore, the infection significantly increased the S. frugiperda larval period but significantly reduced its moth eclosion rate. It suggests that AINV is probably to be a parasitic virus of S. frugiperda.

The genome of the black cutworm Agrotis ipsilon.

The black cutworm (BCW), Agrotis ipsilon, is a worldwide polyphagous and underground pest that causes a high level of economic loss to a wide range of crops through the damage of roots. This species performs non-directed migration throughout East and Southeast Asia seasonally. Lack of a genome information has limited further studies on its unique biology and the development of novel management approaches. In this study, we present a 476 Mb de novo assembly of BCW, along with a consensus gene set of 14,801 protein-coding gene models. Quality controls show that both genome assembly and annotations are high-quality and mostly complete. We focus manual annotation and comparative genomics on gene families that related to the unique attributes of this species, such as nocturnality, long-distance migration, and host adaptation. We find that the BCW genome encodes a similar gene repertoire in various migration-related gene families to the diural migratory butterfly Danaus plexiipus, with additional copies of long wavelength opsin and two eye development-related genes. On the other hand, we find that the genomes of BCW and many other polyphagous lepidopterans encode many more gustatory receptor genes, particularly the lineage-specific expanded bitter receptor genes, than the mono- or oligo-phagous species, suggesting a common role of gustatory receptors (GRs) expansion in host range expansion. The availability of a BCW genome provides valuable resources to study the molecular mechanisms of non-directed migration in lepidopteran pests and to develop novel strategies to control migratory nocturnal pests.

Insecticidal Activity of Lemongrass Essential Oil as an Eco-Friendly Agent against the Black Cutworm Agrotis ipsilon (Lepidoptera: Noctuidae).

BACKGROUND: The destructive insect pest Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae) is a polyphagous species targeting many economically important plants. The extensive and arbitrary use of insecticides has resulted in the build-up of insecticide resistance and pesticide residues accumulating in food. Therefore, it is becoming evident that alternative pest management tools are needed to reduce risks to humans, the environment, and non-target organisms, and at the same time, they should be used in field application at the lowest cost. METHODS: In view of this objective, the present study demonstrates the toxicity of lemongrass (Cymbopogon citratus (DC.) Stapf) essential oil (EO), against the black cutworm A. ipsilon under controlled laboratory conditions in terms of measuring the activity of peroxidase and detoxification enzymes. The chemical components of the EO were analyzed using GC-MS. RESULTS: The results show that after 96 h post treatment, the LC15 and LC50 values were 427.67 and 2623.06 mg/L, respectively, of C. citratus EO on second-instar larvae of A. ipsilon. A slight significance in elongation of the larval duration with LC15 and LC50 value was found with control. By GC-MS analysis, the main compounds identified in the EO were α-citral and ß-citral with percentages of 35.91%, and 35%, respectively. The oxidative stress indicates a significant increase in CAT and lipid peroxidase enzyme activity after 96 h post treatment at the LC15 and LC50. Conversely, the detoxification enzyme activity shows an inhibition of CarE and GST enzymes of larvae exposed to LC15 and LC50 values in response to C. citratus EO. CONCLUSIONS: The present data show that lemongrass EO has insecticidal activity against the black cutworm, A. ipsilon.

Sublethal Exposure Effects of the Neonicotinoid Clothianidin Strongly Modify the Brain Transcriptome and Proteome in the Male Moth Agrotis ipsilon.

Insect pest management relies mainly on neurotoxic insecticides, including neonicotinoids such as clothianidin. The residual accumulation of low concentrations of these insecticides can have positive effects on target pest insects by enhancing various life traits. Because pest insects often rely on sex pheromones for reproduction and olfactory synaptic transmission is cholinergic, neonicotinoid residues could indeed modify chemical communication. We recently showed that treatments with low doses of clothianidin could induce hormetic effects on behavioral and neuronal sex pheromone responses in the male moth, Agrotis ipsilon. In this study, we used high-throughput RNAseq and proteomic analyses from brains of A. ipsilon males that were intoxicated with a low dose of clothianidin to investigate the molecular mechanisms leading to the observed hormetic effect. Our results showed that clothianidin induced significant changes in transcript levels and protein quantity in the brain of treated moths: 1229 genes and 49 proteins were differentially expressed upon clothianidin exposure. In particular, our analyses highlighted a regulation in numerous enzymes as a possible detoxification response to the insecticide and also numerous changes in neuronal processes, which could act as a form of acclimatization to the insecticide-contaminated environment, both leading to enhanced neuronal and behavioral responses to sex pheromone.

Involvement of Methoprene-tolerant and Krüppel homolog 1 in juvenile hormone-signaling regulating the maturation of male accessory glands in the moth Agrotis ipsilon.

Male accessory glands (MAGs) produce seminal fluid proteins that are essential for the fertility and also influence the reproductive physiology and behavior of mated females. In many insect species, and especially in the moth Agrotis ipsilon, juvenile hormone (JH) promotes the maturation of the MAGs but the underlying molecular mechanisms in this hormonal regulation are not yet well identified. Here, we examined the role of the JH receptor, Methoprene-tolerant (Met) and the JH-inducible transcription factor, Krüppel homolog 1 (Kr-h1) in transmitting the JH signal that upregulates the growth and synthetic activity of the MAGs in A. ipsilon. We cloned two full length cDNAs encoding Met1 and Met2 which are co-expressed with Kr-h1 in the MAGs where their expression levels increase with age in parallel with the length and protein content of the MAGs. RNAi-mediated knockdown of either Met1, Met2, or Kr-h1 resulted in reduced MAG length and protein amount. Moreover, injection of JH-II into newly emerged adult males induced the transcription of Met1, Met2 and Kr-h1 associated to an increase in the length and protein content of the MAGs. By contrast, JH deficiency decreased Met1, Met2 and Kr-h1 mRNA levels as well as the length and protein reserves of the MAGs of allatectomized old males and these declines were partly compensated by a combined injection of JH-II in operated males. Taken together, our results highlighted an involvement of the JH-Met-Kr-h1 signaling pathway in the development and secretory activity of the MAGs in A. ipsilon.

Toxicity, residue and risk assessment of tetraniliprole in soil-earthworm microcosms.

Maize seed treatment with chemicals to control underground pests is a common agricultural practice, but inappropriate use of insecticides poses a considerable threat to plant development and soil nontarget organisms. In this study, the availability of tetraniliprole seed dressing to control the black cutworm Agrotis ipsilon (Lepidoptera: Noctuidae) in the maize seeding stage and its safety to earthworms (Eisenia fetida) were investigated. The selective toxicity (ST) of tetraniliprole between E. fetida and A. ipsilon was greater than 4000. No significant adverse effect of tetraniliprole seed treatment on the germination of maize seeds was observed at concentrations of 2.4-9.6 g a.i. /kg seed. Compared with the untreated control, seed treatment with tetraniliprole at 9.6 g a.i. /kg seed greatly reduced the percentage of damaged plants from 88.73% to 26.67%, and achieved the highest control effect of 69.91%. Tetraniliprole of 2.4 g a.i. /kg seed can effectively inhibit A. ipsilon until 14 days after seed germination, with the lowest mortality rate of 44.44%. During the entire exposure period, the maximum residual concentration of tetraniliprole detected in the soil (5.86 mg/kg) was considerably lower than the LC50 value of tetraniliprole to E. fetida (>4000 mg/kg). According to the low-tier risk assessment, the highest risk quotient (RQ) of tetraniliprole seed treatment to earthworms at test concentrations was 2.8 × 10-3, which was evaluated as acceptable. This study provided data support for tetraniliprole seed treatment to control underground pests in maize fields.

Insecticidal Activity of 11 Bt toxins and 3 Transgenic Maize Events Expressing Vip3Aa19 to Black Cutworm, Agrotis ipsilon (Hufnagel).

Black cutworm (BCW), Agrotis ipsilon (Hufnagel), is an occasional pest of maize that can cause considerable economic loss and injury to corn seedlings. This research mainly assessed the susceptibility of BCW neonates to 11 Bt toxins (Cry1Ab, Cry1Ac, Cry1Ah, Cry1F, Cry1Ie, Cry1B, Cry2Aa, Vip3_ch1, Vip3_ch4, Vip3Ca2, Vip3Aa19) by exposing neonates to an artificial diet containing Bt toxins and evaluated the efficacy of three transgenic maize events (C008, C009, C010) expressing Vip3Aa19 toxin against BCW. The toxin-diet bioassay data indicated that Vip3Aa19 protein (LC50 = 0.43 µg/g) was the most active against BCW. Chimeric protein Vip3_ch1 (LC50 = 5.53 µg/g), Cry1F (LC50 = 83.62 µg/g) and Cry1Ac (LC50 = 184.77 µg/g) were less toxic. BCW was very tolerant to the other Bt toxins tested, with LC50 values more than 200 µg/g. Greenhouse studies were conducted with artificial infestations at the whorl stage by placing second-instar BCW larvae into whorl leaf and the fourth-instar larvae at the base of maize seedings. These results suggest that these transgenic maize events expressing Vip3Aa19 can provide effective control for BCW.

Assessing the Efficacy of Bacillus thuringiensis (Bt) Pyramided Proteins Cry1F, Cry1A.105, Cry2Ab2, and Vip3Aa20 Expressed in Bt Maize Against Lepidopteran Pests in Brazil.

Field studies across four states in maize-producing areas of Brazil were conducted to characterize the efficacy of a new pyramided Bacillus thuringiensis (Bt) Berliner technology in maize, Zea mays L., and compare it to existing single and pyramided commercial Bt technologies, to control Helicoverpa zea Boddie (Lepidoptera: Noctuidae), Elasmopalpus lignosellus Zeller (Lepidoptera: Pyralidae), Agrotis ipsilon (Hufnagel) (Lepidoptera: Noctuidae), and Diatraea saccharalis F. (Lepidoptera: Crambidae). Bt maize expressing Vip3Aa20 protein and pyramided Bt maize expressing proteins Cry1F + Cry1A.105 + Cry2Ab2 + Vip3Aa20 provided excellent protection against kernel feeding by H. zea compared to Bt technologies expressing only Cry1F or Cry1A.105 + Cry2Ab2. Bt maize expressing Cry1F, Cry1A.105 + Cry2Ab2, Cry1F + Cry1A.105 + Cry2Ab2, and Cry1F + Cry1A.105 + Cry2Ab2 + Vip3Aa20 resulted in less than 5% of plants injured by E. lignosellus, significantly less than Bt maize expressing only Vip3Aa20 and a non-Bt maize iso-hybrid with and without a thiamethoxam seed treatment. The highest protection against plant cutting injury caused by A. ipsilon was observed in the pyramid Bt maize technology expressing Cry1F + Cry1A.105 + Cry2Ab2 + Vip3Aa20. However, it did not differ statistically from the Bt maize expressing Vip3Aa20, Cry1F, or Cry1F + Cry1A.105 + Cry2Ab2. All Bt maize hybrids evaluated in our study were highly effective in reducing tunneling injury caused by D. saccharalis. These results show that a new maize technology expressing pyramided Bt proteins Cry1F + Cry1A.105 + Cry2Ab2 + Vip3Aa20 offers a higher level of protection from feeding by the above lepidopteran pest complex compared to maize with a single Bt protein or double pyramided Bt proteins.

The Masc gene product controls masculinization in the black cutworm, Agrotis ipsilon.

Sex determination has been studied in the model lepidopteran species Bombyx mori, but it remains poorly understood in lepidopteran pests. In the present study, we identified and characterized the Masculinizer (Masc) gene in a Noctuidae pest species, Agrotis ipsilon. Sequence analysis revealed that AiMasc encodes a protein of 658 amino acids that has two CCCH-type zinc finger domains and two conserved cysteine residues (Cys-277 and Cys-280). We assessed the masculinizing activity of AiMasc in BmN cells and found that AiMasc induced expression of the male-specific doublesex isoform. Disruption of Masc via clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) in A. ipsilon caused abnormalities in abdominal segments and external genitalia, resulting in male-specific sterility. These results suggest that Masc participates in the process of sex determination in A. ipsilon. Successful identification of sex-determination gene in a pest species may enable the development of novel genetic approaches for pest control.