[Design and Testing of a Pressure Measurement and Adjustment Device for Fracture Ends].

Objective: To design and test a device which is capable of accurately measuring and dynamically adjusting the axial pressure at the fracture end in real-time. Methods: Upon completion of the design, the pressure measurement and adjustment device was implemented in a canine tibial fracture external fixation model. A pressure sensor was mounted at the fracture end, and the displayed values of the pressure sensor were used as the standard for comparison. The relationship between the displayed values of the measurement and adjustment device and the pressure sensor under identical conditions was examined. Results: The device was utilized in external fixation models of tibial fractures in five beagles. A linear correlation was observed between the displayed values of the device and the pressure sensor at the fracture end. The measurement values from the device could be transformed into fracture end pressure through the application of coefficients, thereby facilitating accurate measurement and dynamic adjustment of the fracture end pressure. Conclusion: The pressure measurement and adjustment device at the fracture end is easy to operate, enabling precise measurement and dynamic regulation of the pressure at the fracture end. It is well-suited for animal experiments aimed at investigating the impact of axial compression on fracture healing, demonstrating promising potential for experimental applications.

Identification of long non-coding RNAs in response to microsporidia infection in Silkworm, Bombyx mori.

Microsporidia Nosema bombycis (Nb) is a cellular parasite responsible for pébrine disease in silkworms, significantly impacting the sericulture industry. Long non-coding RNAs (lncRNAs), which are RNA fragments longer than 200 nucleotides, are pivotal in a range of cellular and physiological functions. However, the potential role of silkworm lncRNAs in response to Nb infection remains unknown. This study conducted transcriptome sequencing on both larvae and Nb-infected midguts of silkworms, identifying 1,440 lncRNAs across all examined midgut samples. Within the Nb-infected group, 42 differentially expressed lncRNAs (DElncRNAs) and 305 differentially expressed mRNAs (DEmRNAs) were detected. Functional annotation and pathway analysis showed that these DEmRNAs are mostly involved in metabolism, apoptosis, autophagy, and other key pathways. The co-expression network of DEmRNAs and DElncRNAs illustrates that 1 gene could be regulated by multiple lncRNAs and 1 lncRNA may target multiple genes, indicating that the regulation of lncRNA is intricate and networked. In addition, the DElncRNA-miRNA-mRNA network showed that some DElncRNAs may be involved in the immune response and metabolism through miRNA. Notably, the study observed an increase in lncRNA MSTRG857.1 following Nb infection, which may promote Nb proliferation. These findings offer insights into the complex interplay between insects and microsporidia.

Comparative analysis of the intestinal flora of BmNPV-resistant and BmNPV-sensitive silkworm varieties.

Bombyx mori nucleopolyhedrovirus (BmNPV) is a very common and infectious virus that affects silkworms and hinders silk production. To investigate the intestinal flora of BmNPV-resistant and BmNPV-sensitive silkworm varieties, 16 S rDNA high-throughput sequencing was performed. The results of the cluster analysis showed that the intestinal flora of the resistant silkworm variety was more abundant than that of the sensitive silkworm variety. This was found even when infection with BmNPV caused a sharp decline in the number of intestinal floral species in both resistant and sensitive silkworm varieties. The abundances of the intestinal flora, including Aureimonas, Ileibacterium, Peptostreptococcus, Pseudomonas, Enterococcus, and Halomonas, in the resistant variety were considerably greater after infection with BmNPV than those in the sensitive variety. After infection with BmNPV, four kinds of important intestinal bacteria, namely, f_Saccharimonadaceae, Peptostreptococcus, Aureirmonas, and f_Rhizobiaceae, were found in the resistant silkworm variety. In the sensitive silkworm variety, only Faecalibaculum was an important intestinal bacterium. The differential or important bacteria mentioned above might be involved in immunoreaction or antiviral activities, especially in the intestines of BmNPV-resistant silkworms. By conducting a functional enrichment analysis, we found that BmNPV infection did not change the abundance of important functional components of the intestinal flora in resistant or sensitive silkworm varieties. However, some functional factors, such as the biosynthesis, transport, and catabolism of secondary metabolites (e.g., terpenoids and polyketides) and lipid transport and metabolism, were more important in the resistant silkworm variety than in the sensitive variety; thus, these factors may increase the resistance of the host to BmNPV. To summarize, we found significant differences in the composition, abundance, and function of the intestinal flora between resistant and sensitive silkworm varieties, especially after infection with BmNPV, which might be closely related to the resistance of resistant silkworm varieties to BmNPV.

The Function of Different Subunits of the Molecular Chaperone CCT in the Microsporidium Nosema bombycis: NbCCTζ Interacts with NbCCTα.

Chaperonin containing tailless complex polypeptide 1 (CCT) is a molecular chaperone protein that consists of eight completely different subunits and assists in the folding of newly synthesized peptides. The zeta subunit of CCT is a regulatory factor for the folding and assembly of cytoskeletal proteins as individuals or complexes. In this study, the zeta subunit of Nosema bombycis (NbCCTζ) is identified for the first time. The complete ORF of the NbCCTζ gene is 1533 bp in length and encodes a 510 amino acid polypeptide. IFA results indicate that NbCCTζ is colocalized with actin and ß-tubulin in the cytoplasm during the proliferative phase and that NbCCTζ is completely colocalized with NbCCTα in the cytoplasm of N. bombycis throughout the entire life cycle. Furthermore, the yeast two-hybrid assay revealed that the NbCCTζ interacts with NbCCTα. The transcriptional level of NbCCTζ is significantly downregulated by knocking down the NbCCTα gene, while the transcriptional level of NbCCTα is downregulated after knocking down the NbCCTζ gene. These results suggest that NbCCTζ may play a vital role in the proliferation of N. bombycis by coordinating with NbCCTα.

Identifying immune-related prognostic biomarkers in osteosarcoma: Development and validation of the tumor immune microenvironment risk model based on analysis of TCGA.

OBJECTIVE: Osteosarcoma is a rare and aggressive malignancy with limited effective therapeutic options. This study aimed to identify immune-related prognostic biomarkers and develop a prognostic model for osteosarcoma. METHODS: We performed integrated analysis of transcriptomic data and immune cell infiltration profiles of 84 osteosarcoma samples from the Cancer Genome Atlas (TCGA) database. Time-dependent receiver operating characteristic (ROC) curve analysis was used to assess the prognostic value of the TIMErisk model. We also performed functional annotation and pathway enrichment analyses to explore the potential mechanisms underlying the TIMErisk model. RESULTS: We identified a seven-gene TIMErisk model (C2, APBB1IP, BST2, TRPV2, CCL5, GBP1, and F13A1) that was independently associated with overall survival of osteosarcoma patients. The TIMErisk model showed significant associations with immune cell infiltration and immunosuppressive gene expression. In addition, the TIMErisk model was associated with drug sensitivity, and we found that several immune checkpoint genes were significantly differentially expressed between high- and low-TIMErisk groups. Functional annotation and pathway enrichment analyses revealed that the TIMErisk model was associated with multiple immune-related pathways, including antigen processing and presentation, cytokine-cytokine receptor interaction, and T cell receptor signaling pathway. CONCLUSION: Our study identified a novel TIME-based prognostic model for osteosarcoma that incorporates immune-related genes and can be used to predict patient prognosis and response to immunotherapy. Our findings highlight the importance of the TIME microenvironment in osteosarcoma progression and suggest that immune-related biomarkers may have clinical significance in the management of osteosarcoma.

Turnovers of Sex-Determining Mutation in the Golden Pompano and Related Species Provide Insights into Microevolution of Undifferentiated Sex Chromosome.

The suppression of recombination is considered a hallmark of sex chromosome evolution. However, previous research has identified undifferentiated sex chromosomes and sex determination by single SNP in the greater amberjack (Seriola dumerili). We observed the same phenomena in the golden pompano (Trachinotus ovatus) of the same family Carangidae and discovered a different sex-determining SNP within the same gene Hsd17b1. We propose an evolutionary model elucidating the turnover of sex-determining mutations by highlighting the contrasting dynamics between purifying selection, responsible for maintaining W-linked Hsd17b1, and neutral evolution, which drives Z-linked Hsd17b1. Additionally, sporadic loss-of-function mutations in W-linked Hsd17b1 contribute to the conversion of W chromosomes into Z chromosomes. This model was directly supported by simulations, closely related species, and indirectly by zebrafish mutants. These findings shed new light on the early stages of sex chromosome evolution.

Sensory brand experience and brand loyalty: Mediators and gender differences.

Sensory brand experience is a fundamental and central brand experience. Its influencing mechanism on brand outcome variables still needs to be clarified. This study proposed a new SEM model to explore the influencing mechanism from sensory brand experience to brand loyalty and tested it using SmartPLS 4 software. Data were gathered through a web-based questionnaire survey of 304 Chinese consumers. The results show that sensory brand experience is vital in fostering brand loyalty. It impacts brand loyalty both directly and indirectly. The mediating effect of brand attitude is much higher than the mediating effect of brand-self connection, which means that in some situations, a strong emotional bond is not essential for brand success. The results also reveal the moderating effect of gender on relationships in the model. The key gender difference is that the male group does not have a mediation path through brand-self connection, while the female group has. This study expands the research on brand experience and sheds light on developing brand loyalty. Gender differences should be considered when developing sensory marketing strategies. Especially for women, creating sensory brand experiences that elicit strong emotional connections has positive effects on brand loyalty.

Functional characterization of Nosema bombycis (microsporidia) trehalase 3.

Nosema bombycis, an obligate intracellular parasite, is a single-celled eukaryote known to infect various tissues of silkworms, leading to the manifestation of pebrine. Trehalase, a glycosidase responsible for catalyzing the hydrolysis of trehalose into two glucose molecules, assumes a crucial role in thermal stress tolerance, dehydration, desiccation stress, and asexual development. Despite its recognized importance in these processes, the specific role of trehalase in N. bombycis remains uncertain. This investigation focused on exploring the functions of trehalase 3 in N. bombycis (NbTre3). Immunofluorescence analysis of mature (dormant) spores indicated that NbTre3 primarily localizes to the spore membrane or spore wall, suggesting a potential involvement in spore germination. Reverse transcription-quantitative polymerase chain reaction results indicated that the transcriptional level of NbTre3 peaked at 6 h post N. bombycis infection, potentially contributing to energy storage for proliferation. Throughout the life cycle of N. bombycis within the host cell, NbTre3 was detected in sporoplasm during the proliferative stage rather than the sporulation stage. RNA interference experiments revealed a substantial decrease in the relative transcriptional level of NbTre3, accompanied by a certain reduction in the relative transcriptional level of Nb16S rRNA. These outcomes suggest that NbTre3 may play a role in the proliferation of N. bombycis. The application of the His pull-down technique identified 28 proteins interacting with NbTre3, predominantly originating from the host silkworm. This finding implies that NbTre3 may participate in the metabolism of the host cell, potentially utilizing the host cell's energy resources.

Recombinase-aided amplification coupled with lateral flow dipstick for efficient and accurate detection of Bombyx mori nucleopolyhedrovirus.

The infection of Bombyx mori nucleopolyhedrovirus (BmNPV) is one of the main causes of economic losses in sericulture. Thus, it is essential to establish rapid and effective method for BmNPV detection. In the present study, we have developed a recombinase-aided amplification (RAA) to amplify the BmNPV genomic DNA at 37 °C within 30 min, and achieved a rapid detection method by coupling with a lateral flow dipstick (LFD). The RAA-LFD method had a satisfactory detection limit of 6 copies/µL of recombinant plasmid pMD19-T-IE1, and BmNPV infection of silkworm can be detected 12 h post-infection. This method was highly specific for BmNPV, and without cross-reactivity to other silkworm pathogens. In contrast to conventional polymerase chain reaction (PCR), the RAA-LFD assay showed higher sensitivity, cost-saving, and especially is apt to on-site detection of BmNPV infection in the sericulture production.

Ssn6 Interacts with Polar Tube Protein 2 and Transcriptional Repressor for RNA Polymerase II: Insight into Its Involvement in the Biological Process of Microsporidium Nosema bombycis.

Nosema bombycis is a representative species of Microsporidia, and is the pathogen that causes pebrine disease in silkworms. In the process of infection, the polar tube of N. bombycis is injected into the host cells. During proliferation, N. bombycis recruits the mitochondria of host cells. The general transcriptional corepressor Ssn6 contains six tetratricopeptide repeats (TPR) and undertakes various important functions. In this study, we isolated and characterized Nbssn6 of the microsporidium N. bombycis. The Nbssn6 gene contains a complete ORF of 1182 bp in length that encodes a 393 amino acid polypeptide. Indirect immunofluorescence assay showed that the Ssn6 protein was mainly distributed in the cytoplasm and nucleus at the proliferative phase of N. bombycis. We revealed the interaction of Nbssn6 with polar tube protein 2 (Nbptp2) and the transcriptional repressor for RNA polymerase II (Nbtrrp2) by Co-IP and yeast two-hybrid assays. Results from RNA interference further confirmed that the transcriptional level of Nbptp2 and Nbtrrp2 was regulated by Nbssn6. These results suggest that Nbssn6 impacts the infection and proliferation of N. bombycis via interacting with the polar tube protein and transcriptional repressor for RNA polymerase II.

Afidopyropen suppresses silkworm growth and vitality by affecting carbohydrate metabolism and immune function.

Afidopyropen has strong insecticidal toxicity to sucking pests by silencing the vanilloid-type transient receptor potential (TRPV) channels. However, the toxicity of afidopyropen to the Lepidoptera model insect silkworm remain unknown. In this study, the LC50 of afidopyropen to the silkworm at 72 h exposure was 256.82 mg/L. This indicates that afidopyropen is moderately toxic to the silkworm. Long-term exposure to concentrations of 100 mg/L, or less, of afidopyropen, significantly reduced silkworm growth, vitality, silk protein synthesis, and fecundity. A total of 220 differentially expressed genes (DEGs) were detected by transcriptome sequencing, among which 166 were downregulated and 54 were upregulated. Gene Ontology (GO) enrichment analysis showed that the DEGs were enriched in the immune system, immune response and carbohydrate metabolism. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis indicated that DEGs were primarily concentrated in carbohydrate metabolism and biosynthesis of neomycin, kanamycin and gentamicin. Genes related to carbohydrate metabolism and immune system pathways in silkworm were detected by quantitative real-time PCR. The results showed that the genes related to carbohydrate metabolism, silk protein synthesis, and immune response were significantly downregulated. These genes included BCL-6 corepressor-like protein 1 (BCORL1), hexokinase type 2 (HEXO2), phosphoserine aminotransferase 1 (PSAT1), relish (Rel), peptidoglycan recognition protein 2 (PGRP2) and 27 kda glycoprotein precursor (P27K). The data demonstrated the toxic effects of afidopyropen against the silkworm and its regulation of genes responsible for immune function and abscissa carbohydrate metabolism.

Dexmedetomidine after deep brain stimulation for prevention of delirium in elderly patients with Parkinson's disease: protocol for a single-centre, randomised, double-blind, placebo-controlled trial in China.

INTRODUCTION: Parkinson's disease is one of the most common neurodegenerative diseases. Deep brain stimulation (DBS) can improve motor symptoms in patients with middle and late Parkinson's disease, reduce the use of levodopa, and thus reduce drug-related side effects. Postoperative delirium can significantly reduce the short-term and long-term quality of life in elderly patients, which can be alleviated by dexmedetomidine (DEX). However, whether prophylactic DEX could reduce the incidence of postoperative delirium in patients with Parkinson's disease was still unknown. METHODS AND ANALYSIS: This is a single-centre, randomised, double-blinded, placebo-controlled group trial. A total of 292 patients aged 60 years and above elected for DBS will be stratified according to DBS procedure, subthalamic nucleus or globus pallidus interna, then randomly allocated to the DEX group or the placebo control group with a 1:1 ratio, respectively. In the DEX group, patients will be injected with the DEX continuously with an electronic pump at a rate of 0.1 µg/kg/hour for 48 hours at the beginning of general anaesthesia induction. In the control group, normal saline will be administered at the same rate for patients as in the DEX group. The primary endpoint is the incidence of postoperative delirium within 5 days after surgery. Postoperative delirium is assessed by the combination of the Richmond Anxiety Scale and the Confusion Assessment Method (CAM) for the intensive care unit or the 3-minute diagnostic interview for CAM as applicable. The secondary endpoints include the incidence of adverse events and non-delirium complications, the length of stay in the intensive care unit and hospital and all-cause 30-day mortality after the operation. ETHICS AND DISSEMINATION: The protocol has been approved by the Ethics Committee of Beijing Tiantan Hospital of Capital Medical University (KY2022-003-03). The results of this study will be disseminated through presentation at scientific conferences and publication in scientific journals. TRIAL REGISTRATION NUMBER: NCT05197439.

Metabolomic analysis of lipid changes in Bombyx mori infected with Nosema bombycis.

The silkworm (Bombyx mori) is a model species of lepidopteran insect. Microsporidium spp. are obligate intracellular eukaryotic parasites. Infection by the microsporidian Nosema bombycis (Nb) results in an outbreak of Pébrine disease in silkworms and causes substantial losses to the sericulture industry. It has been suggested that Nb depends on nutrients from host cells for spore growth. However, little is known about changes in lipid levels after Nb infection. In this study, ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) was performed to analyze the effect of Nb infection on lipid metabolism in the midgut of silkworms. A total of 1601 individual lipid molecules were detected in the midgut of silkworms, of which 15 were significantly decreased after Nb challenge. Classification, chain length, and chain saturation analysis revealed that these 15 differential lipids can be classified into different lipid subclasses, of which 13 belong to glycerol phospholipid lipids and two belong to glyceride esters. The results indicated that Nb uses the host lipids to complete its own replication, and the acquisition of host lipid subclasses is selective; not all lipid subclasses are required for microsporidium growth or proliferation. Based on lipid metabolism data, phosphatidylcholine (PC) was found to be an important nutrient for Nb replication. Diet supplementation with lecithin substantially promoted the replication of Nb. Knockdown and overexpression of the key enzyme phosphatidate phosphatase (PAP) and phosphatidylcholine (Bbc) for PC synthesis also confirmed that PC is necessary for Nb replication. Our results showed that most lipids in the host midgut decreased when silkworms were infected with Nb. Reduction of or supplementation with PC may be a strategy to suppress or promote microsporidial replication.

Exorista sorbillans (Diptera: Tachinidae) parasitism shortens host larvae growth duration by regulating ecdysone and juvenile hormone titers in Bombyx mori (Lepidoptera: Bombycidae).

The tachinid fly, Exorista sorbillans, is a notorious ovolarviparous endoparasitoid of the silkworm, Bombyx mori, causing severe damage to silkworm cocoon industry. Silkworm larvae show typically precocious wandering behavior after being parasitized by E. sorbillans; however, the underlying molecular mechanism remains unexplored. Herein, we investigated the changes in the levels of 20-hydroxyecdysone (20E) and juvenile hormone (JH) titer, and they both increased in the hemolymph of parasitized silkworms. Furthermore, we verified the expression patterns of related genes, which showed an upregulation of 20E signaling and biosynthesis genes but a significant downregulation of ecdysone oxidase (EO), a 20E inactivation enzyme, in parasitized silkworms. In addition, related genes of the JH signaling were activated in parasitized silkworms, while related genes of the JH degradation pathway were suppressed, resulting in an increase in JH titer. Notably, the precocious wandering behavior of parasitized silkworms was partly recoverable by silencing the transcriptions of BmCYP302A1 or BmCYP307A1 genes. Our findings suggest that the developmental duration of silkworm post parasitism could be shortened by regulation of 20E and JH titers, which may help silkworm to resist the E. sorbillans infestation. These findings provide a basis for deeper insight into the interplay between silkworms and E. sorbillans and may serve as a reference for the development of a novel approach to control silkworm myiasis.

Intraoperative infusion of dexmedetomidine for prevention of postoperative delirium in elderly patients undergoing craniotomy: a protocol of randomised clinical trial.

INTRODUCTION: Postoperative delirium (POD) is a common surgical complication. The incidence is 19% in neurological procedures, and advanced age is a risk factor for neurological procedures. Many studies have shown that dexmedetomidine (DEX) reduced the incidence of delirium after non-cardiac surgery in elderly patients. However, there are few studies focus on the effect of DEX on POD in elderly patients undergoing neurosurgery. METHODS AND ANALYSIS: This is a randomised, double-blinded, paralleled-group and controlled trial. Patients older than 65 years and scheduled for elective craniotomy will be randomly assigned to the DEX group and the control group. After endotracheal intubation, patients in the DEX group will be administered with continuous DEX infusion at rate of 0.4 µg/kg/hour until the surgical haemostasis. In the control group, patients will receive the identical volume of normal saline in the same setting. The primary outcome is the incidence of POD during the first 5 days. Delirium will be evaluated through a combination of three methods, including the Richmond Agitation Sedation Scale (RASS), the confusion assessment method for ICU (CAM-ICU) and the 3 min diagnostic interview for CAM (3D-CAM). The RASS, CAM-ICU and 3D-CAM will be evaluated two times per day (08:00-10:00 and 18:00-20:00 hours) during the first postoperative 5 days. Secondary outcomes include pain severity score, quality of recovery, quality of sleep, cognitive function, psychological health state, intraoperative data, physiological status, length of stay in ICU and hospital, hospitalisation costs, non-delirium complications, and 30-day all-cause mortality. ETHICS AND DISSEMINATION: The protocol (V.4.0) has been approved by the medical ethics committee of Beijing Tiantan Hospital, Capital Medical University (KY2021-194-03). The findings of the study will be disseminated in a peer-reviewed journal and at a scientific conference. TRIAL REGISTRATION NUMBER: NCT05168280.

Significant Effects of Associated Microorganisms on the Community of Photosynthetic Picoeukaryotes.

Photosynthetic picoeukaryotes (PPEs) form associations with other microorganisms. However, whether and how the associated microbes affect PPE communities remain unknown. We used flow cytometric cell sorting combined with parallel high-throughput sequencing of the 18S and 16S rRNA genes to simultaneously investigate PPEs and their associated microbial communities in the Yangtze-connected Lake Dongting. The lake harbors a great diversity of PPEs. PPE communities exhibited significant temporal rather than spatial variations. Two distinct PPE taxa affiliated with Discostella nipponica and Poterioochromonas malhamensis were dominant during winter/spring and summer, respectively. Parallel high-throughput sequencing revealed a great diversity of associated bacteria and non-pigmented eukaryotes (NPEs) in PPEs sorts. Proteobacteria, Actinobacteria, Bacteroidetes, and Cyanobacteria among the associated bacteria and fungi among the associated NPEs were dominant. PPEs were more apparently associated with bacteria than with NPEs. The co-occurrence network of PPEs and associated microbes formed five major modules, which exhibited distinct temporal patterns, being specific to a certain period. Variations in PPEs communities were significantly correlated with both environmental factors and associated microbial communities. In variation partitioning analysis, the associated bacteria explained the greatest variations in PPE communities, and associated bacteria and NPEs co-explained a large portion of environmental effects on PPE communities. Our results highlight the significance of associated microbes in shaping PPE communities.

Identification and subcellular colocalization of protein transport protein Sec61α and Sec61γ in Nosema bombycis.

The main function of Sec61 complex is participating in the transport of polypeptide chains across the endoplasmic reticulum. The Sec61α subunit is the largest subunit of the Sec61 complex and shows high degree of conservation. In this study, we identified the NbSec61α and NbSec61γ genes in the microsporidian Nosema bombycis for the first time. Multiple sequence alignment showed that the sequence similarity between NbSec61α and homologous proteins of other microsporidia was greater than 48 %. NbSec61α contains a "plug" domain (amino acids 40-74) unique to the Sec61/SecY complex. Phylogenetic analysis based on NbSec61α and NbSec61γ indicated that the N. bombycis was closely related to Nosema granulosis, Nosema ceranae and Nosema apis. Indirect immunfluorescence assay showed that NbSec61α and NbSec61γ were mainly distributed in the perinuclear region of N. bombycis in different developmental phases. qRT-PCR results revealed that the expression level of NbSec61α gene increased in the early stage and reached the highest at 48 h, then decreased in the late stages. After knockdown of NbSec61α, the expression of NbSec61α, NbSec61γ and NbssrRNA genes were all significantly down-regulated. These results suggest that the NbSec61α and NbSec61γ may play an important role in the intracellular development of N. bombycis.

The spirotetramat inhibits growth and reproduction of silkworm by interfering with the fatty acid metabolism.

Spirotetramat is a novel insecticide and acaricide that can effectively control many species of piercing-sucking pests by inhibiting lipid synthesis. The silkworm is an economically important insect and a model organism for genetics and biochemical research. However, the toxic effect on their development and reproduction remain unclear. In this study, we demonstrated the negative effects of spirotetramat on the development, vitality, silk protein synthesis, and fecundity of silkworm. We also compared expression changes of silkworm genes using digital gene expression (DGE). A total of 1567 differentially expressed genes (DEGs) were detected, of which 874 genes were downregulated and 693 genes were upregulated. Gene Ontology (GO) enrichment analysis showed that the DEGs were enriched in the oxidation-reduction process, oxidoreductase activity, and fatty-acyl-CoA reductase activity. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that the DEGs were mainly enriched in fatty acid metabolism and lysosome pathways. We detected the relative expression of silkworm genes related to fatty acid synthesis and decomposition pathways and the degradation pathway of juvenile hormone by quantitative real-time PCR. The expression levels of Acetyl CoA carboxylase (ACC), fatty acyl-CoA reductase (FACR), Enoyl-CoA hydratase (ECH), very-long-chain (3R)-3-hydroxyacyl-CoA dehydratase (LCHAD), juvenile hormone epoxide hydrolase (JHEH), and phytanoyl-CoA dioxygenase (PCD) genes were downregulated. These data demonstrate the effects of spirotetramat on silkworm and its effects on genes involved in fatty acid metabolism.

Sar1 Interacts with Sec23/Sec24 and Sec13/Sec31 Complexes: Insight into Its Involvement in the Assembly of Coat Protein Complex II in the Microsporidian Nosema bombycis.

Microsporidia, as unicellular eukaryotes, also have an endomembrane system for transporting proteins, which is essentially similar to those of other eukaryotes. In eukaryotes, coat protein complex II (COPII) consists of Sar1, Sec23, Sec24, Sec13, and Sec31 and mediates protein transport from the endoplasmic reticulum (ER) to the Golgi apparatus. Sar1 is the central player in the regulation of coat protein complex II vesicle formation in the endoplasmic reticulum. In this study, we successfully cloned the NbSar1, NbSec23-1, NbSec23-2, NbSec24-1, NbSec24-2, NbSec13, NbSec31-1, and NbSec31-2 genes and prepared NbSar1 polyclonal antibody. We found that NbSar1 was localized mainly in the perinuclear cytoplasm of Nosema bombycis by immunofluorescence analysis (IFA). Yeast two-hybrid assays demonstrated that NbSar1 interacts with NbSec23-2, NbSec23-2 interacts with NbSec24-1 or NbSec24-2, NbSec23-1 interacts with NbSec31, and NbSec31 interacts with NbSec13. Moreover, the silencing of NbSar1 by RNA interference resulted in the aberrant expression of NbSar1, NbSec23-1, NbSec24-1, NbSec24-2, NbSec13, NbSec31-1, and NbSec31-2 and significantly inhibited the proliferation of N. bombycis. Altogether, these findings indicated that the subunits of coat protein complex II work together to perform functions in the proliferation of N. bombycis and that NbSar1 may play a crucial role in coat protein complex II vesicle formation. IMPORTANCE As eukaryotes, microsporidia have retained the endomembrane system for transporting and sorting proteins throughout their evolution. Whether the microsporidia form coat protein complex II (COPII) vesicles to transport cargo proteins and whether they play other roles besides cargo transport are not fully explained at present. Our results showed that NbSar1, NbSec23-1/NbSec23-2, NbSec24-1/NbSec24-2, NbSec13, and NbSec31 might be assembled to form COPII in the ER of N. bombycis, and the functions of COPII are also closely related to the proliferation of N. bombycis, this may be a new target for the prevention of pébrine disease of the silkworm.

Pleistocene climate and geomorphology drive the evolution and phylogeographic pattern of Triplophysa robusta (Kessler, 1876).

Montane systems provide excellent opportunities to study the rapid radiation influenced by geological and climatic processes. We assessed the role of Pleistocene climatic oscillations and mountain building on the evolution history of Triplophysa robusta, a cold-adapted species restricted to high elevations in China. We found seven differentiated sublineages of T. robusta, which were established during the Mid Pleistocene 0.87-0.61 Mya. The species distribution modeling (SDM) showed an expansion of T. robusta during the Last Glacial Maximum (LGM) and a considerable retraction during the Last Interglacial (LIG). The deep divergence between Clade I distributed in Qinling Mountains and Clade II in Northeastern Qinghai-Tibet Plateau (QTP) was mainly the result of a vicariance event caused by the rapid uplifting of Qinling Mountains during the Early Pleistocene. While the middling to high level of historical gene flow among different sublineages could be attributed to the dispersal events connected to the repetition of the glacial period during the Pleistocene. Our findings suggested that frequent range expansions and regressions due to Pleistocene glaciers likely have been crucial for driving the phylogeographic pattern of T. robusta. Finally, we urge a burning question in future conservation projection on the vulnerable cold-adapted species endemic to high elevations, as they would be negatively impacted by the recent rapid climate warming.