Custom-designed nuclease technologies such as the clustered regularly interspaced short palindromic repeat (CRISPR)-associated (Cas) system provide attractive genome editing tools for insect functional genetics. The targeted gene mutagenesis mediated by CRISPR/Cas9 has been achieved in several orders including Diptera, Lepidoptera and Coleoptera. However, little success reported agricultural pests due to lack of genomic information embryonic microinjection techniques these species. Here we...

We developed a novel antiviral strategy by combining transposon-based transgenesis and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system for direct cleavage of Bombyx mori nucleopolyhedrovirus (BmNPV) genome DNA to promote virus clearance in silkworms. demonstrate that transgenic silkworms constitutively expressing Cas9 guide RNAs targeting BmNPV immediate early-1 (ie-1) me53 genes effectively induce target-specific subsequent...

Abstract Apolipophorin‐III (ApoLp‐III), a multifunctional protein with lipid transport and immune defense functions, widely exists in insects. Although the function of ApoLp‐III as pattern recognition receptor (PRR) immunity has been relatively studied, response mediated by is still vague. To understand whether involved activation prophenoloxidase‐activating system (PPO‐AS), we examined production nitric oxide (NO), synthesis antimicrobial peptides after recognition. The larvae lepidopteran...

Insect body pigmentation and coloration are critical to adaption the environment. To explore mechanisms that drive pigmentation, we used clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) genome editing system target ebony gene in non-model insect Spodoptera litura. Ebony is crucial melanin synthesis insects. By directly injecting Cas9 messenger RNA ebony-specific guide RNAs into S. litura embryos, successfully induced a typical...

Male fertility is essential for reproduction and population growth in animals. Many factors affect male fertility, such as courtship behavior, sperm quantity, motility, among others. Seminal Fluid Proteins (SFPs) are vital components of seminal fluid the ejaculate, which activation, female ovulation. However, knowledge SFPs insufficient; function many remains unknown, most described functions were mainly characterized Drosophila or other laboratory models. Here, we focus on Serine protease 2...

Abstract BACKGROUND The gene doublesex ( dsx ) plays pivotal roles in sex determination and controls sexually dimorphic development certain insects. Importantly, it also displays a potential candidate target for pest management due to its sex‐specific splicing. Therefore, we used CRISPR/Cas9‐mediated disruption investigate the function of Hyphantria cunea , an invasive forest pest. RESULT In present study, identified from H. which showed sex‐biased expression pattern that was different other...

Sex determination is an important and traditional biological process. In Lepidoptera, Masculinizer (Masc) doublesex (dsx) are the essential genes for sex play critical roles in sexual differentiation development. The functions of Masc dsx have been characterized several model insect species. However, molecular mechanism Ostrinia furnacalis, agricultural pest, still unknown. Here, we successfully used CRISPR/Cas9 genome editing system to knock out OfMasc Ofdsx. Mutation induced male external...

Precise regulation of oogenesis is crucial to female reproduction. Seventy percent pests belong lepidopteran species, so it would be interesting explore the highly conserved genes involved in that do not affect growth and development model, Bombyx mori. This can provide potential target for pest control promote insect sterility technology.In ovarian serine protease (Osp), which encodes a member family, essential oogenesis. In this study, we used transgenic CRISPR/Cas9 technology obtain Osp...

The reproductive process in various species has undergone evolutionary adaptations at both the physiological and molecular levels, playing a significant role maintaining their populations. In lepidopteran insects, spermatophore is unique structure formed female system, which sperm storage activation take place. It known that formation of regulated by seminal fluid proteins derived from males. However, studies investigating genetic mechanisms behind lepidopterans have been limited. this...

Abstract Fusion of the testis occurs in most Lepidoptera insects, including Spodoptera litura , an important polyphagous pest. Testicular fusion S. is advantageous for male reproduction, and molecular mechanism remains unknown. Doublesex influences formation genitalia, behavior courtship, sexually dimorphic traits fruit‐fly silkworm, essential sexual differentiation. However, its purpose The doublesex gene ( Sldsx ) has male‐specific M female‐specific F isoforms, exhibits a higher expression...

Ostrinia furnacalis (Lepidoptera: Pyralidae) is one of the most destructive agricultural pests in Asia. Traditional pest-management methods include sex pheromone capture, transgenic crops that produce Bacillus thuringiensis toxin, and pesticides. Although these strategies control pest populations effectively, they also cause negative side effects, including dramatically increased pesticide resistance, severe pollution, hazards for human health. Recently developed genome editing tools provide...

(1) Background: Abdominal-A (Abd-A) and Ultrabithorax (Ubx) are homeotic genes that determine the identity morphology of thorax abdomen in insects. The Asian corn borer, Ostrinia furnacalis (Guenée) (Lepidoptera: Pyralidae), is a devastating maize pest throughout Asia, Western Pacific, Australia. Building on previous knowledge, we hypothesized knockout Abd-A Ubx would disrupt abdominal body planning O. furnacalis. (2) Methods: CRISPR/Cas9-targeted mutagenesis was employed to decipher...

Spodoptera frugiperda (J. E. Smith), an emerging invasive pest worldwide, has posed a serious agricultural threat to the newly invaded areas. Although somatic sex differentiation is fundamentally conserved among insects, determination cascade in S. largely unknown. In this study, we cloned and functionally characterized Doublesex (dsx), “molecular switch” modulating sexual dimorphism using male- female-specific isoforms. Given that Lepidoptera recalcitrant RNAi, CRISPR/Cas9-mediated...

Abstract Sex‐specific regulatory elements are key components for developing insect genetic sexing systems. The current system mainly uses a female‐specific modification whereas little success was reported on male‐specific modification. In the silkworm B ombyx mori , lepidopteran model with economic importance, transgene‐based, lethality has been established based sex‐specific alternative splicing factors and promoter BmVgp (vitellogenin promoter) identified. However, no have yet Here we...

The domesticated silkworm is an economically important insect that widely used as a lepidopteran model. Although somatic sex determination in the well characterized, germline not. Here, we transgenic-based CRISPR/Cas9 genome editing system to study function of Ovo gene Bombyx mori. BmOvo homolog factor Drosophila melanogaster. mutants had abnormally shaped eggs were disordered ovarioles, and gonad development was abnormal. Interestingly, wing discs wings did not develop properly, most failed...

Abstract The Masculinizer gene, Masc , encodes a lepidopteran‐specific novel CCCH‐type zinc finger protein, which controls sex determination and dosage compensation in Bombyx mori . Considering the potential application of it pest control, is necessary to investigate function gene Hyphantria cunea globally invasive forest pest. In present study, we identified functionally characterized HcMasc H. Sequence analysis revealed that HcMASC contained conserved domain, nuclear localization signal,...

Abstract As a core member of the Hippo signaling pathway, Hpo plays critical role in regulating growth and development. Previous studies reported that loss function results increased proliferation, reduced apoptosis induction tissue overgrowth Drosophila . In this study, we used CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/Cas9) to study gene ( BmHpo ) lepidopteran insect Bombyx mori , known commonly as silkworm. Sequence analysis revealed an array deletions...

As a typical G protein-coupled receptor, the adipokinetic hormone receptor (AKHR) has seven transmembrane domains (TMDs), and its structure function are similar to gonadotropin-releasing (GnRHR) in vertebrates. However, there is dearth of information on other components AKHR signaling pathway how it functions interaction between insect hosts parasitoids. In this study, we cloned analyzed multifunctional Ostrinia furnacalis (OfAKHR) cDNA (GenBank accession number MF797868). OfAKHR 2206 bp...

Genetic manipulation of sex determination pathways in insects provides the basis for a broad range strategies to benefit agricultural security and human health. The P-element somatic inhibitor (PSI) protein, an exon splicing silencer that promotes male-specific dsx, plays critical role male sexual differentiation development. functions PSI have been characterized lepidopteran model species Bombyx mori. However, molecular mechanism Plutella xylostella, worldwide pest taxonomically basal...