Autophagy promotes replication of Bombyx mori Nucleopolyhedrovirus in insect cells.

BmNPV is a pathogen that infects silkworms exclusively. Although the interaction between BmNPV and the silkworm has been widely noticed and studied, its specific mechanism has still not been elucidated. In this study, we investigated whether BmNPV infection induces the onset of host cell autophagy to enhance viral replication. We observed a significant increase in double- or single-membrane vesicles and an accumulation of enhanced green fluorescent protein eGFP-ATG8 spots in virus-infected cells 72 h after BmNPV infection, accompanied by a conversion of ATG8 to ATG8-PE. In addition, we observed changes in the mitochondrial morphology of BmN cells after BmNPV infection by transmission electron microscopy. By detecting the mitochondrial membrane potential, we found that BmNPV infection resulted in the decrease of mitochondrial membrane potential, and that eGFP-ATG8 was able to co-localise with mitochondria after virus infection of the cells. Moreover, the use of drugs to regulate the occurrence of autophagy affects the replication of cellular BmNPV. Our data demonstrates that BmNPV infection induces host cell autophagy and leads to cellular mitochondrial damage, which in turn may lead to mitochondrial autophagy, and that BmNPV-induced host autophagy promotes its replication in cells. These findings will provide clues for further understanding of host-virus interactions.

Systematic analysis of innate immune-related genes in the silkworm: Application to antiviral research.

The silkworm, a crucial model organism of the Lepidoptera, offers an excellent platform for investigating the molecular mechanisms underlying the innate immune response of insects toward pathogens. Over the years, researchers worldwide have identified numerous immune-related genes in silkworms. However, these identified silkworm immune genes are not well classified and not well known to the scientific community. With the availability of the latest genome data of silkworms and the extensive research on silkworm immunity, it has become imperative to systematically categorize the immune genes of silkworms with different database IDs. In this study, we present a meticulous organization of prevalent immune-related genes in the domestic silkworm, using the SilkDB 3.0 database as a reliable source for updated gene information. Furthermore, utilizing the available data, we classify the collected immune genes into distinct categories: pattern recognition receptors, classical immune pathways, effector genes and others. In-depth data analysis has enabled us to predict some potential antiviral genes. Subsequently, we performed antiviral experiments on selected genes, exploring their impact on Bombyx mori nucleopolyhedrovirus replication. The outcomes of this research furnish novel insights into the immune genes of the silkworm, consequently fostering advancements in the field of silkworm immunity research by establishing a comprehensive classification and functional understanding of immune-related genes in the silkworm. This study contributes to the broader understanding of insect immune responses and opens up new avenues for future investigations in the domain of host-pathogen interactions.

Single-Nucleus Sequencing of Silkworm Larval Brain Reveals the Key Role of Lysozyme in the Antiviral Immune Response in Brain Hemocytes.

INTRODUCTION: The brain is considered as an immune-privileged organ, yet innate immune reactions can occur in the central nervous system of vertebrates and invertebrates. Silkworm (Bombyx mori) is an economically important insect and a lepidopteran model species. The diversity of cell types in the silkworm brain, and how these cell subsets produce an immune response to virus infection, remains largely unknown. METHODS: Single-nucleus RNA sequencing (snRNA-seq), bioinformatics analysis, RNAi, and other methods were mainly used to analyze the cell types and gene functions of the silkworm brain. RESULTS: We used snRNA-seq to identify 19 distinct clusters representing Kenyon cell, glial cell, olfactory projection neuron, optic lobes neuron, hemocyte-like cell, and muscle cell types in the B. mori nucleopolyhedrovirus (BmNPV)-infected and BmNPV-uninfected silkworm larvae brain at the late stage of infection. Further, we found that the cell subset that exerts an antiviral function in the silkworm larvae brain corresponds to hemocytes. Specifically, antimicrobial peptides were significantly induced by BmNPV infection in the hemocytes, especially lysozyme, exerting antiviral effects. CONCLUSION: Our single-cell dataset reveals the diversity of silkworm larvae brain cells, and the transcriptome analysis provides insights into the immune response following virus infection at the single-cell level.

Insights into midgut cell types and their crucial role in antiviral immunity in the lepidopteran model Bombyx mori.

The midgut, a vital component of the digestive system in arthropods, serves as an interface between ingested food and the insect's physiology, playing a pivotal role in nutrient absorption and immune defense mechanisms. Distinct cell types, including columnar, enteroendocrine, goblet and regenerative cells, comprise the midgut in insects and contribute to its robust immune response. Enterocytes/columnar cells, the primary absorptive cells, facilitate the immune response through enzyme secretions, while regenerative cells play a crucial role in maintaining midgut integrity by continuously replenishing damaged cells and maintaining the continuity of the immune defense. The peritrophic membrane is vital to the insect's innate immunity, shielding the midgut from pathogens and abrasive food particles. Midgut juice, a mixture of digestive enzymes and antimicrobial factors, further contributes to the insect's immune defense, helping the insect to combat invading pathogens and regulate the midgut microbial community. The cutting-edge single-cell transcriptomics also unveiled previously unrecognized subpopulations within the insect midgut cells and elucidated the striking similarities between the gastrointestinal tracts of insects and higher mammals. Understanding the intricate interplay between midgut cell types provides valuable insights into insect immunity. This review provides a solid foundation for unraveling the complex roles of the midgut, not only in digestion but also in immunity. Moreover, this review will discuss the novel immune strategies led by the midgut employed by insects to combat invading pathogens, ultimately contributing to the broader understanding of insect physiology and defense mechanisms.

Applications and Potentials of a Silk Fibroin Nanoparticle Delivery System in Animal Husbandry.

Silk fibroin (SF), a unique natural polymeric fibrous protein extracted from Bombyx mori cocoons, accounts for approximately 75% of the total mass of silk. It has great application prospects due to its outstanding biocompatibility, biodegradability, low immunogenicity, and mechanical stability. Additionally, it is non-toxic and environmentally friendly. Nanoparticle delivery systems constructed with SF can improve the bioavailability of the carriers, increase the loading rates, control the release behavior of the deliverables, and enhance their action efficiencies. Animal husbandry is an integral part of agriculture and plays a vital role in the development of the rural economy. However, the pillar industry experiences a lot of difficulties, like drug abuse while treating major animal diseases, and serious environmental pollution, restricting sustainable development. Interestingly, the limited use cases of silk fibroin nanoparticle (SF NP) delivery systems in animal husbandry, such as veterinary vaccines and feed additives, have shown great promise. This paper first reviews the SF NP delivery system with regard to its advantages, disadvantages, and applications. Moreover, we describe the application status and developmental prospects of SF NP delivery systems to provide theoretical references for further development in livestock production and promote the high-quality and healthy development of animal husbandry.

Single-nucleus sequencing of silkworm larval midgut reveals the immune escape strategy of BmNPV in the midgut during the late stage of infection.

The midgut is an important barrier against microorganism invasion and proliferation, yet is the first tissue encountered when a baculovirus naturally invades the host. However, only limited knowledge is available how different midgut cell types contribute to the immune response and the clearance or promotion of viral infection. Here, single-nucleus RNA sequencing (snRNA seq) was employed to analyze the responses of various cell subpopulations in the silkworm larval midgut to B. mori nucleopolyhedrovirus (BmNPV) infection. We identified 22 distinct clusters representing enteroendocrine cells (EEs), enterocytes (ECs), intestinal stem cells (ISCs), Goblet cell-like and muscle cell types in the BmNPV-infected and uninfected silkworm larvae midgut at 72 h post infection. Further, our results revealed that the strategies for immune escape of BmNPV in the midgut at the late stage of infection include (1) inhibiting the response of antiviral pathways; (2) inhibiting the expression of antiviral host factors; (3) stimulating expression levels of genes promoting BmNPV replication. These findings suggest that the midgut, as the first line of defense against the invasion of the baculovirus, has dual characteristics of "resistance" and "tolerance". Our single-cell dataset reveals the diversity of silkworm larval midgut cells, and the transcriptome analysis provides insights into the interaction between host and virus infection at the single-cell level.

Medial Septal Glutamatergic Neurons Modulate States of Consciousness during Sevoflurane Anesthesia in Mice.

BACKGROUND: Multiple neural structures involved in maintaining wakefulness have been found to promote arousal from general anesthesia. The medial septum is a critical region that modulates arousal behavior. This study hypothesized that glutamatergic neurons in the medial septum play a crucial role in regulating states of consciousness during sevoflurane general anesthesia. METHODS: Adult male mice were used in this study. The effects of sevoflurane anesthesia on neuronal activity were determined by fiber photometry. Lesions and chemogenetic manipulations were used to study the effects of the altered activity of medial septal glutamatergic neurons on anesthesia induction, emergence, and sensitivity to sevoflurane. Optogenetic stimulation was used to observe the role of acute activation of medial septal glutamatergic neurons on cortical activity and behavioral changes during sevoflurane-induced continuous steady state of general anesthesia and burst suppression state. RESULTS: The authors found that medial septal glutamatergic neuronal activity decreased during sevoflurane anesthesia induction and recovered in the early period of emergence. Chemogenetic activation of medial septal glutamatergic neurons prolonged the induction time (mean ± SD, hM3Dq-clozapine N-oxide vs. hM3Dq-saline, 297.5 ± 60.1 s vs. 229.4 ± 29.9 s, P < 0.001, n = 11) and decreased the emergence time (53.2 ± 11.8 s vs. 77.5 ± 33.5 s, P = 0.025, n = 11). Lesions or chemogenetic inhibition of these neurons produced the opposite effects. During steady state of general anesthesia and deep anesthesia-induced burst suppression state, acute optogenetic activation of medial septal glutamatergic neurons induced cortical activation and behavioral emergence. CONCLUSIONS: The study findings reveal that activation of medial septal glutamatergic neurons has arousal-promoting effects during sevoflurane anesthesia in male mice. The activation of these neurons prolongs the induction and accelerates the emergence of anesthesia.

Microwave-Responsive Flexible Room-Temperature Phosphorescence Materials Based on Poly(vinylidene fluoride) Polymer.

The development of flexible, room-temperature phosphorescence (RTP) materials remains challenging owing to the quenching of their unstable triplet excitons via molecular motion. Therefore, a polymer matrix with Tg higher than room temperature is required to prevent polymer segment movement. In this study, a RTP material was developed by incorporating a 4-biphenylboronic acid (BPBA) phosphor into a poly(vinylidene fluoride) (PVDF) matrix (Tg =-27.1 °C), which exhibits a remarkable UV-light-dependent oxygen consumption phosphorescence with a lifetime of 1275.7 ms. The adjustable RTP performance is influenced by the crystallinity and polymorph (α, ß, and γ phases) fraction of PVDF, therefore, the low Tg of the PVDF matrix enables the polymeric segmental motion upon microwave irradiation. Consequently, a reduction in the crystallinity and an increase in the α phase fraction in PVDF film induces RTP after 2.45 GHz microwave irradiation. These findings open up new avenues for constructing crystalline and phase-dependent RTP materials while demonstrating a promising approach toward microwave detection.

Medial septum glutamatergic neurons modulate nociception in chronic neuropathic pain via projections to lateral hypothalamus.

The medial septum (MS) contributes in pain processing and regulation, especially concerning persistent nociception. However, the role of MS glutamatergic neurons in pain and the underlying neural circuit mechanisms in pain remain poorly understood. In this study, chronic constrictive injury of the sciatic nerve (CCI) surgery was performed to induce thermal and mechanical hyperalgesia in mice. The chemogenetic activation of MS glutamatergic neurons decreased pain thresholds in naïve mice. In contrast, inhibition or ablation of these neurons has improved nociception thresholds in naïve mice and relieved thermal and mechanical hyperalgesia in CCI mice. Anterograde viral tracing revealed that MS glutamatergic neurons had projections to the lateral hypothalamus (LH) and supramammillary nucleus (SuM). We further demonstrated that MS glutamatergic neurons regulate pain thresholds by projecting to LH but not SuM, because the inhibition of MS-LH glutamatergic projections suppressed pain thresholds in CCI and naïve mice, yet, optogenetic activation or inhibition of MS-SuM glutamatergic projections had no effect on pain thresholds in naïve mice. In conclusion, our results reveal that MS glutamatergic neurons play a significant role in regulating pain perception and decipher that MS glutamatergic neurons modulate nociception via projections to LH.

Modifying PTAA/Perovskite Interface via 4-Butanediol Ammonium Bromide for Efficient and Stable Inverted Perovskite Solar Cells.

Inverted perovskite solar cells (IPSCs) have witnessed an impressive development in recent years. However, their efficiency is still significantly behind theoretical limits, and device instabilities hinder their commercialization. Two main obstacles to further enhancing their performance via one-step deposition are: 1) the unsatisfactory film quality of perovskite and 2) the poor surface contact. To address the above issues, 4-butanediol ammonium Bromide (BD) is utilized to passivate Pb2+ defects by forming PbN bonds and fill vacancies of formamidinium ions at the buried surface of perovskite. The wettability of poly [bis (4-phenyl) (2,4,6-triMethylphenyl) amine] films is also improved due to the formation of hydrogen bonds between PTAA and BD molecules, resulting in better surface contacts and enhanced perovskite crystallinity. As a result, BD-modified perovskite thin films show a significant increase in the mean grain size, as well as a dramatic enhancement in the PL decay lifetime. The BD-treated device exhibits an efficiency of up to 21.26%, considerably higher than the control device. Moreover, the modified devices show dramatically enhanced thermal and ambient stability compared to the control ones. This methodology paves the way to obtain high-quality perovskite films for fabricating high-performance IPSCs.

Single-Nucleus Sequencing of Fat Body Reveals Distinct Metabolic and Immune Response Landscapes in Silkworm Larvae after Bombyx mori Nucleopolyhedrovirus Infection.

The fat body plays a central role in the regulation of the life cycle of insects and acts as the major site for detoxification, nutrient storage, energy metabolism, and innate immunity. However, the diversity of cell types in the fat body, as well as how these cell subsets respond to virus infection, remains largely unknown. We used single-nucleus RNA sequencing to identify 23 distinct clusters representing adipocyte, hemocyte, epithelial cell, muscle cell, and glial cell types in the fat body of silkworm larvae. Further, by analysis of viral transcriptomes in each cell subset, we reveal that all fat body cells could be infected by Bombyx mori nucleopolyhedrovirus (BmNPV) at 72 h postinfection, and that the majority of infected cells carried at least a medium viral load, whereas most cells infected by BmNPV at 24 h postinfection had only low levels of infection. Finally, we characterize the responses occurring in the fat body cell clusters on BmNPV infection, which, on one hand, mainly reduce their metabolic functions, involving energy, carbohydrates, lipids, and amino acids, but, on the other hand, initiate a strong antiviral response. Our single-nucleus RNA sequencing analysis reveals the diversity of insect fat body cells and provides a resource of gene expression profiles for a systems-level understanding of their response to virus infection.

Alteration of neural activity and neuroinflammatory factors in the insular cortex of mice with corneal neuropathic pain.

Dry eye disease (DED) affects nearly 55% of people worldwide; several studies have proposed that central sensitization and neuroinflammation may contribute to the developing corneal neuropathic pain of DED, while the underlying mechanisms of this contribution remain to be investigated. Excision of extra orbital lacrimal glands established the dry eye model. Corneal hypersensitivity was examined through chemical and mechanical stimulation, and open field test measured the anxiety levels. Restingstate fMRI is a method of functional magnetic resonance imaging (rs-fMRI) was performed for anatomical involvement of the brain regions. The amplitude of low-frequency fluctuation (ALFF) determined brain activity. Immunofluorescence testing and Quantitative real-time polymerase chain reaction were also performed to further validate the findings. Compared with the Sham group, ALFF signals in the supplemental somatosensory area, secondary auditory cortex, agranular insular cortex, temporal association areas, and ectorhinal cortex brain areas were increased in the dry eye group. This change of ALFF in the insular cortex was linked with the increment in corneal hypersensitivity (p < 0.01), c-Fos (p < 0.001), brain-derived neurotrophic factor (p < 0.01), TNF-α, IL-6, and IL-1ß (p < 0.05). In contrast, IL-10 levels (p < 0.05) decreased in the dry eye group. DED-induced corneal hypersensitivity and upregulation of inflammatory cytokines could be blocked by insular cortex injection of Tyrosine Kinase receptor B agonist cyclotraxin-B (p < 0.01) without affecting anxiety levels. Our study reveals that the functional activity of the brain associated with corneal neuropathic pain and neuroinflammation in the insular cortex might contribute to dry eye-related corneal neuropathic pain.

The piRNA pathway is required for nucleopolyhedrovirus replication in Lepidoptera.

The Piwi-interacting RNA (piRNA) pathway has been shown to be involved in the antiviral defense against RNA viruses, especially in mosquitoes, but its universality has been questioned. Here, we used the Bombyx mori nucleopolyhedrovirus (BmNPV) -infected silkworm as a model to explore the effects of the key factors of piRNA pathway, BmAgo3 and Siwi, on replication of a large DNA virus (belonging to the family of Baculoviridae). We demonstrated that BmAgo3 and Siwi could promote the replication of BmNPV through both overexpression and knockdown experiments in BmN cell lines and silkworm larvae. In addition, we also studied the effect of PIWI-class genes on Autographa californica nucleopolyhedrovirus (AcMNPV) replication in the Spodoptera frugiperda cell line Sf9. By knocking down the expression of PIWI-class genes in Sf9, we found that Piwi-like-1 and Piwi-like-2-3 could inhibit AcMNPV replication, while Piwi-like-4-5 promoted virus replication. Our study provides compelling evidence that the piRNA pathway affects host infection by exogenous viruses in Lepidoptera. Also, our results reflect the diversity of the roles of PIWI-class genes in virus infection of the host across species. This study is the first to explore the interaction of PIWI-class proteins with DNA viruses, providing new insights into the functional roles of the piRNA pathway.

BmCH25H, a vertebrate interferon-stimulated gene(ISG) homolog, inhibits BmNPV infection dependent on its hydroxylase activity in Bombyx mori.

Cholesterol-25-hydroxylase (CH25H) has been identified as an interferon-stimulated gene (ISG) in mammals that exerts its antiviral effects by catalyzing the conversion of cholesterol to 25-hydroxycholesterol (25HC). However, invertebrates lack an antiviral system homologous to vertebrate interferons (IFNs) because the genomes of invertebrates do not encode IFN-like cytokines. Nevertheless, CH25H is present in insect genomes and it therefore deserves further study of whether and by which mechanism it could exert an antiviral effect in invertebrates. In this study, the Bombyx mori CH25H (BmCH25H) gene, of which the encoded protein has high homology with other lepidopteran species, was identified and located on chromosome 9. Interestingly, we found that the expression of BmCH25H was significantly upregulated in B. mori nucleopolyhedrovirus (BmNPV) -infected BmN cells and silkworm (B. mori) larvae at the early infection stage. The inhibitory effect of BmCH25H on BmNPV replication was further demonstrated to depend on its catalytic residues to convert cholesterol to 25HC. More importantly, we demonstrated that during BmNPV infection, BmCH25H expression was increased through the Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway, similar to the induction of ISGs following virus infection in vertebrates. This is the first report that CH25H has antiviral effects in insects; the study also elucidates the regulation of its expression and its mechanism of action.

Apolipoprotein D facilitate the proliferation of BmNPV.

The silkworm, Bombyx mori, a model Lepidopteran specie, is an important economic insect. It is specifically infected by Bombyx mori nucleopolyhedrovirus (BmNPV), causing huge losses to the sericulture industry. Therefore, the understandings of the interaction mechanism between BmNPV and the host will help to provide the theoretical basis for the sericulture industry to control BmNPV. Apolipoprotein D (ApoD) is a member of lipid transport family and capable of binding to a variety of lipophilic ligands. ApoD is mainly used in neurodegenerative disease research in mammals, and there is little research on ApoD against viruses. Here, we explored the effects of Bombyx mori Apolipoprotein D (BmApoD) on BmNPV replication. We knocked out and overexpressed BmApoD in BmN cells and infected them with Bombyx mori nucleopolyhedrovirus (BmNPV). The results showed that BmApoD promote the replication of BmNPV in BmN cells. It was also confirmed that BmApoD promote the replication of BmNPV after knocking down BmApoD in silkworm larvae. This study is the first to explore the role of ApoD in insect-virus interactions, providing new insights into the functional role of ApoD.

Factors Associated with Prolonged Extubation after Total Intravenous Anesthesia in Patients Undergoing Vestibular Schwannoma Resection.

OBJECTIVE: To identify factors associated with prolonged tracheal extubation after vestibular schwannoma resection in patients receiving propofol-remifentanil-based total intravenous anesthesia (TIVA). STUDY DESIGN: Single-center retrospective study of vestibular schwannoma resection performed by a single neurosurgeon between July 2018 and September 2021. SETTING: Tertiary academic medical center. PATIENTS: Adults receiving TIVA for vestibular schwannoma resection, classified according to extubation time: non-prolonged extubation (<15 min) and prolonged extubation (≥15 min). MAIN OUTCOME MEASURES: Time from end of surgery to extubation, demographic parameters, intraoperative variables, and familiarity between the anesthesia provider and the neurosurgeon were analyzed. Predictors for prolonged extubation were analyzed via multivariate analysis. The primary outcome was the incidence of prolonged extubation. The secondary outcome was factors associated with prolonged tracheal extubation. RESULTS: A total of 234 cases were analyzed. The median (interquartile range) extubation time was 9.4 minutes (7.2, 12.2 min). Extubation was prolonged in 39 patients (16.7%). Factors predicting prolonged extubation were significant blood loss (odds ratio [OR], 12.8; 95% confidence interval [CI], 2.6-61.7; p = 0.002), intraoperative neuromuscular blocking drug infusion (OR, 6.6; 95% CI, 2.8-15.7; p < 0.001), and lack of familiarity between the anesthesia provider and neurosurgeon (OR, 4.4; 95% CI, 1.5-12.3; p = 0.005). CONCLUSION: Significant blood loss, intraoperative neuromuscular blocking drug infusion, and lack of familiarity between anesthesia provider and neurosurgeon were associated with prolonged extubation following TIVA for vestibular schwannoma resection.

Sirt5 Inhibits BmNPV Replication by Promoting a Relish-Mediated Antiviral Pathway in Bombyx mori.

Silent information regulators (Sirtuins) belong to the family of nicotinamide adenine dinucleotide (NAD+)-dependent histone deacetylases (HDACs) that have diverse functions in cells. Mammalian Sirtuins have seven isoforms (Sirt1-7) which have been found to play a role in viral replication. However, Sirtuin members of insects are very different from mammals, and the function of insect Sirtuins in regulating virus replication is unclear. The silkworm, Bombyx mori, as a model species of Lepidoptera, is also an important economical insect. B. mori nucleopolyhedrovirus (BmNPV) is a major pathogen that specifically infects silkworms and causes serious losses in the sericulture industry. Here, we used the infection of the silkworm by BmNPV as a model to explore the effect of Sirtuins on virus replication. We initially knocked down all silkworm Sirtuins, and then infected with BmNPV to analyze its replication. Sirt2 and Sirt5 were found to have potential antiviral functions in the silkworm. We further confirmed the antiviral function of silkworm Sirt5 through its effects on viral titers during both knockdown and overexpression experiments. Additionally, Suramin, a Sirt5 inhibitor, was found to promote BmNPV replication. In terms of molecular mechanism, it was found that silkworm Sirt5 might promote the immune pathway mediated by Relish, thereby enhancing the host antiviral response. This study is the first to explore the role of Sirtuins in insect-virus interactions, providing new insights into the functional role of members of the insect Sirtuin family.

Added Syrup as a Flavoring to Oral Midazolam Intravenous Solution Premedication Improves Tolerability in Pediatric Patients.

Purpose: To investigate the tolerability of midazolam intravenous (IV) solution by masking its bitter taste through the addition of syrup. Methods: In the waiting area before surgery at our institution, Eye and ENT Hospital, from May to November of 2021, pediatric patients scheduled for anterior ear fistula removal or eardrum catheterization were randomly assigned to receive a mixture of IV midazolam and water (Group C) or IV midazolam and syrup (Group S) in equal volumes. We measured the difference between the groups in terms of drug tolerability with a 5-point facial hedonic scale given to every patient, parent, and nurse involved in the study. Secondary outcomes included sedation and anxiety levels, which were assessed at separation from the parents and general anesthesia induction via a mask. Results: In total, 180 patients were enrolled. The facial hedonic scales were higher in Group S as evaluated by the children (p=0.046), parents (p=0.020), and nurses (p=0.026). More patients were willing to take the same solution again in Group S than in Group C (p=0.024). The levels of sedation and anxiety at two timepoints were similar between the groups (all p>0.05). No adverse events were noted preoperatively. Conclusion: The tolerability of IV midazolam administered orally was increased in a pediatric population by adding syrup. The sedative and anxiolytic effects were comparable for both midazolam mixtures. Trial Registry Number: China Clinical Research Information Service, ChiCTR2000040229.

Global Metabolic Profiling of Baculovirus Infection in Silkworm Hemolymph Shows the Importance of Amino-Acid Metabolism.

Viruses rely on host cell metabolism to provide the necessary energy and biosynthetic precursors for successful viral replication. Infection of the silkworm, Bombyx mori, by Bombyx mori nucleopolyhedrovirus (BmNPV), has been studied extensively in the past to unravel interactions between baculoviruses and their lepidopteran hosts. To understand the interaction between the host metabolic responses and BmNPV infection, we analyzed global metabolic changes associated with BmNPV infection in silkworm hemolymph. Our metabolic profiling data suggests that amino acid metabolism is strikingly altered during a time course of BmNPV infection. Amino acid consumption is increased during BmNPV infection at 24 h post infection (hpi), but their abundance recovered at 72 hpi. Central carbon metabolism, on the other hand, particularly glycolysis and glutaminolysis, did not show obvious changes during BmNPV infection. Pharmacologically inhibiting the glycolytic pathway and glutaminolysis also failed to reduce BmNPV replication, revealing that glycolysis and glutaminolysis are not essential during BmNPV infection. This study reveals a unique amino acid utilization process that is implemented during BmNPV infection. Our metabolomic analysis of BmNPV-infected silkworm provides insights as to how baculoviruses induce alterations in host metabolism during systemic infection.

Identification of Silkworm Hemocyte Subsets and Analysis of Their Response to Baculovirus Infection Based on Single-Cell RNA Sequencing.

A wide range of hemocyte types exist in insects but a full definition of the different subclasses is not yet established. The current knowledge of the classification of silkworm hemocytes mainly comes from morphology rather than specific markers, so our understanding of the detailed classification, hemocyte lineage and functions of silkworm hemocytes is very incomplete. Bombyx mori nucleopolyhedrovirus (BmNPV) is a representative member of the baculoviruses and a major pathogen that specifically infects silkworms (Bombyx mori) and causes serious losses in sericulture industry. Here, we performed single-cell RNA sequencing (scRNA-seq) of hemocytes in BmNPV and mock-infected larvae to comprehensively identify silkworm hemocyte subsets and determined specific molecular and cellular characteristics in each hemocyte subset before and after viral infectmadion. A total of 20 cell clusters and their potential marker genes were identified in silkworm hemocytes. All of the hemocyte clusters were infected by BmNPV at 3 days after inoculation. Interestingly, BmNPV infection can cause great changes in the distribution of hemocyte types. The cells appearing in the infection group mainly belong to prohemocytes (PR), while plasmatocytes (PL) and granulocytes (GR) are very much reduced. Furthermore, we found that BmNPV infection suppresses the RNA interference (RNAi) and immune response in the major hemocyte types. In summary, our results revealed the diversity of silkworm hemocytes and provided a rich resource of gene expression profiles for a systems-level understanding of their functions in the uninfected condition and as a response to BmNPV.