Spermatogenesis is a key process for the sexual reproduction species. In lepidopteran insects, spermatogenesis produces two different types of sperms, in which eupyrene sperm carry genomic DNA and fertilize eggs, whereas apyrene sperm are necessary for eupyrene sperm to enter eggs. However, functional genetic studies of spermatogenesis in Plutella xylostella remain a longstanding puzzle even though the phenomenon in lepidoptera has been widely documented more than a century. In this study, we particularly focus on the gene Nap1 which belongs to the Nucleosome assembly protein family. Our findings revealed that Nap1 was highly expressed in the testes, and the disruption of PxNap1 induced male sterility in P. xylostella, while the fertility of mutant females was comparable to wild-type females. Additionally, through immunofluorescence staining analysis, we found that the eupyrene sperm bundles presented diffusedly scattered nuclei in PxNap1 mutant males, while the nuclei in the wild-type were clustered together presented as needle shape. We also found that PxNap1 deficiency hinders the transfer of eupyrene sperm to the bursa copulatrix and spermatheca of females. However, the apyrene spermatogenesis was not affected in the PxNap1 mutant. RNA-seq analyses indicated that the defects of eupyrene sperm in PxNap1 mutants were related to energy metabolic such as pentose and glucuronate interconversions, biosynthesis of amino acids, and pentose phosphate pathway. Our study demonstrates that PxNap1 plays crucial function in eupyrene spermatogenesis and eupyrene sperm migration. Our research provides valuable insights for the genetic factors underlying reproductive processes in Lepidopteran insects.

Entomopathogenic fungi (EPFs) are capable of infecting a variety of insect pests and are widely used as biopesticides worldwide. This study intended to isolate indigenous EPFs from cadavers of Protaetia brevitarsis and investigate their effects on the fall armyworm Spodoptera frugiperda (FAW), a globally widespread invasive pest. Morphological and molecular analyses confirmed four native EPF strains all belong to Beauveria bassiana. Pathogenicity tests showed they were virulent toward FAW 1st instar larvae. The application of EPFs either by dipping or spraying significantly increased the larval mortalities compared to the control group, with corrected mortalities ranging from 92% to 73% after dipping in a fungal suspension of 108 conidia/mL, and those ranging from 76% to 35% after spraying with a fungal suspension of 107 conidia/mL. Our findings revealed the infectivity of four strains to FAW larvae significantly changed in a dose- and time-dependent manner. In addition, the combination use of the local B. bassiana strain and parasitoid Microplitis prodeniae resulted in a significantly enhanced S. frugiperda 3rd instar larval mortality compared to a single inoculation with one of them, suggesting an apparent synergistic effect for the joint application of these two biological control agents. The mortality inflicted by B. bassiana was probably promoted by the release of parasitoids, since the parasitoids’ movements and attacks could strengthen the fungal distribution and infection processes. This study underscores the potential of the combination use of EPFs and parasitoids against S. frugiperda early-instar larvae, and provides insights into the consequences of integrating these EPFs into integrated pest management systems.

Uric acid is the end-product of nitrogen metabolism of the silkworm and other lepidopterans. The accumulation of uric acid particles in the epidermis causes the larval silkworm to appear white and opaque. However, the mechanism of uric acid granule formation is still unclear. Silkworm epidermis color is linked to the genes responsible for uric acid particle formation. We first identified two genes in the Bombyx mori genome that encode subunits of the Bloc-1 (Biogenesis of Lysosome-related Organelles Complex-1) by homology to these genes in other eukaryotes, Bmpali and Bmb1. Mutation in these genes caused a transparent phenotype in the silkworm larvae, and the loss of BmBloc-1 subunit gene Bmcap resulted in the same phenotype. These three genes are highly conserved between human and silkworm. We discovered that Bmpali, Bmcap, and Bmb1 localize in the cytoplasm of BmN cells. Yeast two-hybrid assays demonstrated that the Bmpali physically interacts with both Bmcap and Bmb1. Investigating the roles of Bmpali, Bmb1, and Bmcap is essential for uric acid granule formation understanding in Bombyx mori. These mutants present a valuable silkworm model for studying the biogenesis of lysosome-related organelles (LROs).

Purpose: This work aims to develop a novel distortion-free 3D-EPI acquisition and image reconstruction technique for fast and robust, high-resolution, whole-brain imaging as well as quantitative T2* mapping. Methods: 3D-Blip-Up and -Down Acquisition (3D-BUDA) sequence is designed for both single- and multi-echo 3D GRE-EPI imaging using multiple shots with blip-up and -down readouts to encode B0 field map information. Complementary k-space coverage is achieved using controlled aliasing in parallel imaging (CAIPI) sampling across the shots. For image reconstruction, an iterative hard-thresholding algorithm is employed to minimize the cost function that combines field map information informed parallel imaging with the structured low-rank constraint for multi-shot 3D-BUDA data. Extending 3D-BUDA to multi-echo imaging permits T2* mapping. For this, we propose constructing a joint Hankel matrix along both echo and shot dimensions to improve the reconstruction. Results: Experimental results on in vivo multi-echo data demonstrate that, by performing joint reconstruction along with both echo and shot dimensions, reconstruction accuracy is improved compared to standard 3D-BUDA reconstruction. CAIPI sampling is further shown to enhance the image quality. For T2* mapping, T2* values from 3D-Joint-CAIPI-BUDA and reference multi-echo GRE are within limits of agreement as quantified by Bland-Altman analysis. Conclusions: The proposed technique enables rapid 3D distortion-free high-resolution imaging and T2* mapping. Specifically, 3D-BUDA enables 1-mm isotropic whole-brain imaging in 22 s at 3 T and 9 s on a 7 T scanner. The combination of multi-echo 3D-BUDA with CAIPI acquisition and joint reconstruction enables distortion-free whole-brain T2* mapping in 47 s at 1.1x1.1x1.0 mm3 resolution.

The black soldier fly (BSF), Hermetia illucens, is a promising insect for mitigating solid waste problems as its larvae are able to bioconvert organic waste into valuable biomass. We recently reported a high-quality genome assembly of the BSF; analysis of this genome sequence will further the understanding of insect biology and identify genes that can be manipulated to improve efficiency of bioconversion. To enable genetic manipulation of the BSF, we have established the first transgenic methods for this economically important insect. We cloned and identified the ubiquitous actin5C promoter (Hiactin5C-p3k) and 3 endogenous U6 promoters (HiU6:1, HiU6:2, and HiU6:3). The Hiactin5C promoter was used to drive expression of a hyperactive variant of the piggyBac transposase, which exhibited up to 6-fold improvement in transformation rate when compared to the wild-type transposase. Furthermore, we evaluated the 3 HiU6 promoters using this transgenic system. HiU6:1 and HiU6:2 promoters provided the highest knockdown efficiency with RNAi and are thus promising candidates for future Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) development. Overall, our findings provide valuable genetic engineering toolkits for basic research and genetic manipulation of the BSF.

The reproductive process in various species has undergone evolutionary adaptations at both the physiological and molecular levels, playing a significant role in maintaining their populations. In lepidopteran insects, the spermatophore is a unique structure formed in the female reproductive system, in which sperm storage and activation take place. It is known that the formation of the spermatophore is regulated by seminal fluid proteins derived from males. However, studies investigating the genetic mechanisms behind spermatophore formation in lepidopterans have been limited. In this study, our focus was on SPSL1, a gene that encodes a trypsin-type seminal fluid protein in Spodoptera frugiperda, a pest species with global invasive tendencies. Our findings revealed that SPSL1 expression was predominantly observed in the male reproductive tracts, and the disruption of this gene resulted in male sterility. Surprisingly, fluorescence analysis indicated that the absence of SPSL1 did not affect spermatogenesis or sperm migration within the male reproductive system. However, when females mated with SPSL1-mutant males, several defects were observed. These included disruptions in spermatophore formation, sperm activation in the copulatory bursae, and sperm migration into the spermathecae. Additionally, mass spectrometry analysis highlighted reduced levels of energy-related metabolites, suggesting that SPSL1 plays an essential role in promoting hydrolysis reactions during copulation. Consequently, our study demonstrates that SPSL1 is crucial for male fertility due to its functions in spermatophore formation and sperm activation. This research provides valuable insights into the genetic factors underlying reproductive processes in lepidopteran insects and sheds light on potential strategies for controlling invasive pest populations.

The extremely high field has significant advantages in imaging quality and analyzing the fine structure of substances. However, its excellent performance requires the support of a higher-performance shim technique. In this paper, a novel structural design pattern of the shim coil for a 27 T extremely high field superconducting magnet is proposed. According to the contours of the stream function, we designed and optimized the shim coil pattern and engineering processing. The novel design was realized by using the contours as the centerline, and the wire spacing was controlled at 1 mm. The performance of the novel pattern was compared with those of alternative winding schemes. The results indicate that the novel design can improve coil performance, achieving higher fidelity and lower power dissipation.

MicroRNAs (miRNAs) are important regulators of nearly all aspects of biological processes in eukaryotes. During the biogenesis of miRNAs, the RNase III enzyme Dicer processes double-strand precursor miRNAs into mature miRNAs and promotes the assembly of RNA-induced silencing complexes (RISCs). Dicer has been reported to participate in a wide range of physiological processes, including development and immunity, in some insect species. However, the physiological roles of Dicer in lepidopterans remain poorly understood. In this study, we investigated the function of Bombyx mori Dicer1. We first performed sequence alignment and found that the sequence of functional domains of Dicer1 are varied among Lepidoptera, Diptera, Coleoptera, Blattaria, and Orthoptera. Using a binary clustered regularly interspaced palindromic repeats (CRISPR) / CRISPR-associated protein 9 genome editing approach, we showed that BmDicer1 mutants have arrested development from the 3rd instar into the 4th instar. RNA sequencing analysis indicated that the defects in BmDicer1 mutants are due to dysregulation of genes that encode proteins involved in metabolism, protein degradation, absorption, and renin-angiotensin pathways. Analysis using quantitative real-time polymerase chain reaction showed that mutation of BmDicer1 altered expression of miRNAs and their target genes. Therefore, our study demonstrates the critical roles of BmDicer1 in miRNA biogenesis and larval development in silkworm.

The silkworm Bombyx mori is an economically important insect. The sericulture industry is seriously affected by pathogen infections. Of these pathogens, Bombyx mori nucleopolyhedrovirus (BmNPV) causes approximately 80% of the total economic losses due to pathogen infections. We previously constructed a BmNPV-specific CRISPR/Cas9 silkworm line with significantly enhanced resistance to BmNPV. In order to optimize the resistance properties and minimize its impact on economic traits, we constructed an inducible CRISPR/Cas9 system for use in transgenic silkworms. We used the 39k promoter, which is induced by viral infection, to express Cas9 and the U6 promoter to express four small guide RNA targeting the genes encoding BmNPV late expression factors 1 and 3 (lef-1 and lef-3, respectively), which are essential for viral DNA replication. The system was rapidly activated when the silkworm was infected and showed considerably higher resistance to BmNPV infection than the wild-type silkworm. The inducible system significantly reduced the development effects due to the constitutive expression of Cas9. No obvious differences in developmental processes or economically important characteristics were observed between the resulting transgenic silkworms and wild-type silkworms. Adoption of this accurate and highly efficient inducible CRISPR/Cas9 system targeting BmNPV DNA replication will result in enhanced antivirus measures during sericulture, and our work also provides insights into the broader application of the CRISPR/Cas9 system in the control of infectious diseases and insect pests.

The evolution of feeding habits leads to speciation in insects. Bactrocera true fruit flies display diverse feeding habits across species. We combine behavioral and functional genomic studies to probe the divergence between the specialist B. minax and the generalist B. dorsalis. We find that both vision and olfaction contribute to their respective host preferences, with a dominant effect of vision over the olfaction in short range. Correspondingly, host location-related genes are significantly enriched in the phototransduction pathway, of which the long-wavelength rhodopsin confers the color preference in both species and has been subject to selection in the specialist. We also find a massive expansion of olfactory receptors in the generalist, along with signatures of conditional expression and positive selection. The phylogenetic context suggests an ancestrally important role of vision in the host location of Bactrocera, as well as the increased performance and plasticity of olfaction alongside the arising of generalism.

As an effective and reliable method of obtaining extremely high magnetic fields, rare-earth barium copper oxide (REBCO) magnets have contributed significantly to the development of condensed matter physics, chemistry, life sciences, and materials research. However, large screening currents in REBCO magnets can significantly degrade the magnetic field quality. Recent experiments on the inner insert of a 30 T superconducting magnet at the Institute of Electrical Engineering, Chinese Academy of Sciences showed that the measured magnetic field is highly asymmetric single-peaked field instead of the designed saddle-shaped fields, and it differs significantly from the values calculated based on the commonly-used uniform and symmetric screening current model. Therefore, a simulation model based on the T – A formulation with consideration of accurate dimensions and critical currents was developed. The simulation results with this new developed model indicate that the angle dependence can cause more than an 8.3% difference in the critical currents of the symmetrically located double pancake (DP) coils of the magnet. The average relative error between the measured and calculated magnetic field values was sharply reduced by a maximum of 44.4% after considering the non-uniform critical currents. In consideration of manufacturing uncertainties and non-uniform critical currents, better magnetic field quality of REBCO magnets can be achieved by arranging the location of each DP with this model.

Male fertility is essential for reproduction and population growth in animals. Many factors affect male fertility, such as courtship behavior, sperm quantity, and sperm motility, among others. Seminal Fluid Proteins (SFPs) are vital components of seminal fluid in the male ejaculate, which affect male fertility, sperm activation, and female ovulation. However, the knowledge of SFPs is insufficient; the function of many SFPs remains unknown, and most described functions were mainly characterized in Drosophila or other laboratory models. Here, we focus on the Serine protease 2 (Ser2) gene in the lepidopteran pest Spodoptera litura. The Ser2 gene was specifically expressed in male adults. Disruption of the Ser2 gene mediated by CRISPR/Cas9 induced male sterility but females remained fertile. PCR-based detection of the next-generation mutants showed that male sterility was stably inherited. The qRT-PCR analysis of SlSer2 mutants showed that motor protein family genes and structural protein family genes were down-regulated, while protein modification family genes were up-regulated, suggesting that SlSer2 may be involved in sperm movement and activity. These results demonstrate that Ser2 is an important component of SFPs in seminal fluid and was identified for a useful sterile gene for pest control that may lead to new control strategies for lepidopteran insect pests such as S. litura.

3D-BUDA data acquired from a 3T Siemens scanner and a 7T Siemens scanner

Proton therapy (PT) is a precise and efficient radiotherapy method in modern medical treatment, which can be focused on the lesion location to kill the tumor cells with little affection on the normal tissues and thus largely reduce the potential side effects. However, the proton therapy instrument is usually very huge with a large occupation in the space. In order to reduce the instrument scale, a petawatt level laser proton accelerator project is being conducted in China. This paper will report a curved Canted-Cosine-Theta (CCT) magnet which is a critical component in the proposed laser proton accelerator system. The CCT superconducting magnet was designed with a curved dipole magnet that had an equivalent arc π/4 with radius 1 m, and two short quadrupole magnets were nested symmetrically at the dipole magnet ends. The warm bore of the magnet was 72 mm, where a homogeneous transverse magnetic field that amounted to 2.5 T was required along the magnet bore and homogeneous magnetic field gradients 25 T/m were required at the magnet inlet and outlet. NbTi superconducting wire will be used on the coil winding and immersed in liquid helium environment. The structural design of the magnet will be emphatically introduced in this paper.

Bombyx mori nucleopolyhedrovirus (BmNPV) is a pathogen that causes severe disease in silkworms. In a previous study, we demonstrated that by using the CRISPR/Cas9 system to disrupt the BmNPV ie-1 and me53 genes, transgenic silkworms showed resistance to BmNPV infection. Here, we used the same strategy to simultaneously target lef8 and lef9, which are essential for BmNPV replication. A PCR assay confirmed that double-stranded breaks were induced in viral DNA at targeted sequences in BmNPV-infected transgenic silkworms that expressed small guide RNAs (sgRNAs) and Cas9. Bioassays and qPCR showed that replication of BmNPV and mortality were significantly reduced in the transgenic silkworms in comparison with the control groups. Microscopy showed degradation of midgut cells in the BmNPV-infected wild type silkworms, but not in the transgenic silkworms. These results demonstrated that transgenic silkworms using the CRISPR/Cas9 system to disrupt BmNPV lef8 and lef9 genes could successfully prevent BmNPV infection. Our research not only provides more alternative targets for the CRISPR antiviral system, but also aims to provide new ideas for the application of virus infection research and the control of insect pests.

在昆虫与植物协同进化过程中,昆虫成功定位寄主植物决定其食性及生态位.实蝇类昆虫是世界范围内分布的重要果蔬害虫,在定位寄主植物过程中,实蝇视觉和嗅觉发挥重要作用,利用寄主颜色和气味模拟的诱捕装置能够有效诱杀实蝇,但诱杀效果参差不齐.因此,本文对实蝇类昆虫视觉、嗅觉定位寄主植物的行为和相关分子基础进行了阐述,介绍了有关的应用成果,并对未来的方向进行了展望,以期为建立实蝇类害虫可持续的绿色防控技术奠定基础.

The magnetic dam (MD) is a promising passive protection method for high-temperature superconducting (HTS) magnets. Urgent problems remain to be resolved in order to put it into practice. This article deals with the deformation problem of the magnetic dam in the background magnet quench. A novel copper–steel composite structure was proposed, which could improve the MD’s mechanical properties. As a comparison criterion, the electromagnetic stress and deformation of the magnetic dam during the background field magnet quench are simulated and analyzed. The effect of different thicknesses of stainless steel on the strength and the protection performance of the magnetic dam is also discussed. The results revealed that the original copper magnetic dam does not possess sufficient strength to survive from the low-temperature superconducting (LTS) magnet quench in the HTS/LTS magnetic system. The composite structure is more apt to be used as a passive protection method for the insert HTS magnets.

Yolk proteins are the main source of nutrients during embryonic and early larval development in oviparous animals. Therefore, vitellogenesis is crucial for reproduction. The silkworm, Bombyx mori, is a model lepidopteran insect in which there are three yolk proteins: vitellin, 30-kDa protein, and egg-specific protein (Esp). In this study, we explored the gene function of Esp through transgenic clustered regularly interspaced palindromic repeats (CRISPR) / CRISPR-associated protein 9 technology-mediated mutations in the silkworm. We found that Esp mutation resulted in female sterility but had no effect on male fertility. Female mutants could lay eggs after mating, but the eggs were smaller and lighter colored than those laid by wild-type females. The most important finding is that the eggs laid by female mutants did not hatch. Furthermore, we observed stable inheritance of female sterility caused by Esp mutation through successive generations. Thus, Esp encodes a yolk protein that is crucial for female reproductive success and is a potential target for pest control.

Sex determination is an important and traditional biological process. In Lepidoptera, Masculinizer (Masc) and doublesex (dsx) are the essential genes for sex determination and play critical roles in sexual differentiation and development. The functions of Masc and dsx have been characterized in several model insect species. However, the molecular mechanism and sex determination functions of Masc and dsx in Ostrinia furnacalis, an agricultural pest, are still unknown. Here, we successfully used the CRISPR/Cas9 genome editing system to knock out OfMasc and Ofdsx. Mutation of OfMasc induced male external genital defects and sterility. Disruptions of the Ofdsx common region caused sex-specific defects in the external genitals and adult sterility. In addition, we found that OfMasc and Ofdsx can regulate the pigmentation genes that control wing pigmentation patterns. These results demonstrate that OfMasc and Ofdsx play key roles in the sex determination of O. furnacalis, and suggest novel genetic control approaches for the management of pests, including O. furnacalis.