Multimodal evaluation of blood-brain barrier opening in mice in response to low-intensity ultrasound and a claudin-5 binder.

Background: The blood-brain barrier (BBB) is a major bottleneck in delivering therapeutics to the brain. Treatment strategies to transiently open this barrier include focused ultrasound combined with intravenously injected microbubbles (FUS+MB) and targeting of molecules that regulate BBB permeability. Methods: Here, we investigated BBB opening mediated by the claudin-5 binder cCPEm (a microorganismal toxin in a truncated form) and FUS+MB at a centre frequency of 1 MHz, assessing dextran uptake, broadband emission, and endogenous immunoglobulin G (IgG) extravasation. Results: FUS+MB-induced BBB opening was detectable at a pressure ≥0.35 MPa when assessed for leakage of 10 and 70 kDa dextran, and at ≥0.2 MPa for uptake of endogenous IgG. Treating mice with 20 mg/kg cCPEm failed to open the BBB, and pre-treatment with cCPEm followed by FUS+MB at 0.2 and 0.3 MPa did not overtly increase BBB opening compared to FUS+MB alone. Using passive cavitation detection (PCD), we found that broadband emission correlated with the peak negative pressure (PNP) and dextran leakage, indicating the possibility of using broadband emission for developing a feedback controller to monitor BBB opening. Conclusions: Together, our study highlights the challenges in developing combinatorial approaches to open the BBB and presents an additional IgG-based histological detection method for BBB opening.

Premature Progesterone Rise Is Associated with Higher Cumulative Live Birth Rate with Freeze-All Strategy.

Background/Objectives: This paper undertakes an investigation into the implications of premature progesterone rise (PPR) on pregnancy outcomes in freeze-all strategy cycles. Methods: A retrospective cohort study encompassing 675 IVF/ICSI cycles using a freeze-all strategy was enrolled. The cycles were categorized into two groups based on serum progesterone levels at the time of hCG administration: 526 cycles had levels below 1.5 ng/mL, while 149 cycles had levels equal to or above 1.5 ng/mL. Results: The findings revealed a significantly higher number of mature follicles and retrieved oocytes in patients with PPR across all AMH categories. Multiple analyses revealed factors influencing PPR, including the duration of induction and the number of retrieved oocytes. Within the same oocyte retrieval number group, patients with PPR demonstrated non-inferior pregnancy outcomes compared to non-PPR patients. Upon adjustment for age, AMH, and total follicle-stimulating hormone (FSH) dosage, PPR maintained a positive correlation with the cumulative live birth rate (LBR). Conclusions: The study showed that PPR correlates with an increase in retrieved oocytes while maintaining similar embryo quality and oocyte retrieval rates and results in a higher cumulative LBR.

The varicella-zoster virus ORF16 protein promotes both the nuclear transport and the protein abundance of the viral DNA polymerase subunit ORF28.

Although all herpesviruses utilize a highly conserved replication machinery to amplify their viral genomes, different members may have unique strategies to modulate the assembly of their replication components. Herein, we characterize the subcellular localization of seven essential replication proteins of varicella-zoster virus (VZV) and show that several viral replication enzymes such as the DNA polymerase subunit ORF28, when expressed alone, are localized in the cytoplasm. The nuclear import of ORF28 can be mediated by the viral DNA polymerase processivity factor ORF16. Besides, ORF16 could markedly enhance the protein abundance of ORF28. Noteworthily, an ORF16 mutant that is defective in nuclear transport still retained the ability to enhance ORF28 abundance. The low abundance of ORF28 in transfected cells was due to its rapid degradation mediated by the ubiquitin-proteasome system. We additionally reveal that radicicol, an inhibitor of the chaperone Hsp90, could disrupt the interaction between ORF16 and ORF28, thereby affecting the nuclear entry and protein abundance of ORF28. Collectively, our findings imply that the cytoplasmic retention and rapid degradation of ORF28 may be a key regulatory mechanism for VZV to prevent untimely viral DNA replication, and suggest that Hsp90 is required for the interaction between ORF16 and ORF28.

Deep learning-assisted diagnosis of benign and malignant parotid tumors based on ultrasound: a retrospective study.

BACKGROUND: To develop a deep learning(DL) model utilizing ultrasound images, and evaluate its efficacy in distinguishing between benign and malignant parotid tumors (PTs), as well as its practicality in assisting clinicians with accurate diagnosis. METHODS: A total of 2211 ultrasound images of 980 pathologically confirmed PTs (Training set: n = 721; Validation set: n = 82; Internal-test set: n = 89; External-test set: n = 88) from 907 patients were retrospectively included in this study. The optimal model was selected and the diagnostic performance evaluation is conducted by utilizing the area under curve (AUC) of the receiver-operating characteristic(ROC) based on five different DL networks constructed at varying depths. Furthermore, a comparison of different seniority radiologists was made in the presence of the optimal auxiliary diagnosis model. Additionally, the diagnostic confusion matrix of the optimal model was calculated, and an analysis and summary of misjudged cases' characteristics were conducted. RESULTS: The Resnet18 demonstrated superior diagnostic performance, with an AUC value of 0.947, accuracy of 88.5%, sensitivity of 78.2%, and specificity of 92.7% in internal-test set, and with an AUC value of 0.925, accuracy of 89.8%, sensitivity of 83.3%, and specificity of 90.6% in external-test set. The PTs were subjectively assessed twice by six radiologists, both with and without the assisted of the model. With the assisted of the model, both junior and senior radiologists demonstrated enhanced diagnostic performance. In the internal-test set, there was an increase in AUC values by 0.062 and 0.082 for junior radiologists respectively, while senior radiologists experienced an improvement of 0.066 and 0.106 in their respective AUC values. CONCLUSIONS: The DL model based on ultrasound images demonstrates exceptional capability in distinguishing between benign and malignant PTs, thereby assisting radiologists of varying expertise levels to achieve heightened diagnostic performance, and serve as a noninvasive imaging adjunct diagnostic method for clinical purposes.

Silanol-Assisted High-Yield Nanofabrication of SnO2 Single Crystals with Highly Tunable and Ordered Mesoporosity.

Highly ordered mesoporous materials with a single-crystalline structure have attracted broad interest due to their wide applications from catalysis to energy conversion/storage, but constructing them with good controllability and high yields remains a highly daunting task. Herein, we construct a new class of three-dimensionally ordered mesoporous SnO2 single crystals (3DOm-SnO2) with well-defined facets and excellent mesopore tunability. Mechanism studies demonstrate that the silanol groups on ordered silica nanospheres (3DO-SiO2) can induce the efficient heterogeneous crystallization of uniform SnO2 single crystals in its periodic voids by following the hard and soft acid and base theory, affording a much higher yield of ∼96% for 3DOm-SnO2 than that of its solid counterpart prepared in the absence of 3DO-SiO2 (∼1.5%). Benefiting from its permanent ordered mesopores and favorable electronic structure, Pd-supported 3DOm-SnO2 can efficiently catalyze the unprecedented sequential hydrogenation of 4-nitrophenylacetylene to produce 4-nitrostyrene, then 4-nitroethylbenzene, and finally 4-aminoethylbenzene. DFT calculations further reveal the favorable synergistic effect between Pd and 3DOm-SnO2 via moderate electron transfer for realizing this sequential hydrogenation reaction. Our work underlines the crucial role of silanol groups in inducing the high-yield heterogeneous crystallization of 3DOm-SnO2, shedding light on the rational design and construction of various 3DO single crystals that are of great practical significance.

Macrophage exosomes modified by miR-365-2-5p promoted osteoblast osteogenic differentiation by targeting OLFML1.

In the bone immune microenvironment, immune cells can regulate osteoblasts through a complex communication network. Macrophages play a central role in mediating immune osteogenesis, exosomes derived from them have osteogenic regulation and can be used as carriers in bone tissue engineering. However, there are problems with exosomal therapy alone, such as poor targeting, and the content of loaded molecules cannot reach the therapeutic concentration. In this study, macrophage-derived exosomes modified with miR-365-2-5p were developed to accelerate bone healing. MC3T3-E1 cells were incubated with the culture supernatants of M0, M1 and M2 macrophages, and it was found that the culture medium of M2 macrophages had the most significant effects in contributing to osteogenesis. High-throughput sequencing identified that miR-365-2-5p was significantly expressed in exosomes derived from M2 macrophages. We incubated MC3T3-E1 with exosomes overexpressing or knocking down miR-365-2-5p to examine the biological function of exosome miR-365-2-5p on MC3T3-E1 differentiation. These findings suggested that miR-365-2-5p secreted by exosomes increased the osteogenesis of MC3T3-E1. Moreover, miR-365-2-5p had a direct influence over osteogenesis for MC3T3-E1. Sequencing analysis combined with dual luciferase detection indicated that miR-365-2-5p binded to the 3'-UTR of OLFML1. In summary, exosomes secreted by M2 macrophages targeted OLFML1 through miR-365-2-5p to facilitate osteogenesis.

The Binding of the SARS-CoV-2 Spike Protein to Platelet Factor 4: A Proposed Mechanism for the Generation of Pathogenic Antibodies.

Pathogenic platelet factor 4 (PF4) antibodies contributed to the abnormal coagulation profiles in COVID-19 and vaccinated patients. However, the mechanism of what triggers the body to produce these antibodies has not yet been clarified. Similar patterns and many comparable features between the COVID-19 virus and heparin-induced thrombocytopenia (HIT) have been reported. Previously, we identified a new mechanism of autoimmunity in HIT in which PF4-antibodies self-clustered PF4 and exposed binding epitopes for other pathogenic PF4/eparin antibodies. Here, we first proved that the SARS-CoV-2 spike protein (SP) also binds to PF4. The binding was evidenced by the increase in mass and optical intensity as observed through quartz crystal microbalance and immunosorbent assay, while the switching of the surface zeta potential caused by protein interactions and binding affinity of PF4-SP were evaluated by dynamic light scattering and isothermal spectral shift analysis. Based on our results, we proposed a mechanism for the generation of PF4 antibodies in COVID-19 patients. We further validated the changes in zeta potential and interaction affinity between PF4 and SP and found that their binding mechanism differs from ACE2-SP binding. Importantly, the PF4/SP complexes facilitate the binding of anti-PF4/Heparin antibodies. Our findings offer a fresh perspective on PF4 engagement with the SARS-CoV-2 SP, illuminating the role of PF4/SP complexes in severe thrombotic events.

Analyzing the Diversity of MYB Family Response Strategies to Drought Stress in Different Flax Varieties Based on Transcriptome Data.

The MYB transcription factor family has numerous members, and is involved in biological activities, such as ABA signaling, which plays an important role in a plant's resistance to abiotic stresses such as drought. However, the diversity of MYB members that respond to drought stress and their regulatory mechanisms in different flax varieties were unclear. In this study, we obtained 855.69 Gb of clean data from 120 flax root samples from 20 flax (Linum usitatissimum L.) varieties, assembled 92,861 transcripts, and identified 434 MYB family members in each variety. The expression profiles of the MYB transcription factor family from 20 flax varieties under drought stress were analyzed. The results indicated that there are four strategies by which the MYB family responds to drought stress in these 20 flax varieties, each of which has its own specific processes, such as development, reproduction, and localization processes. The four strategies also include common biological processes, such as stimulus responses, metabolic processes, and biological regulation. The WGCNA method was subsequently employed to identify key members of the MYB family involved in response strategies to drought stress. The results demonstrated that a 1R-MYB subfamily gene co-expression network is significantly related to the gibberellin response and cytokinin-activated signaling pathway processes in the 'Strategy 4' for MYB family response to drought, identifying core genes such as Lus.scaffold70.240. Our results showed a diversity of MYB family responses to drought stress within flax varieties, and these results contribute to deciphering the mechanisms of the MYB family regulation of drought resistance. This will promote the more accurate breeding development of flax to adapt to agricultural production under drought conditions.

ESCRT-I protein UBAP1 controls ventricular expansion and cortical neurogenesis via modulating adherens junctions of radial glial cells.

Intricate cerebral cortex formation is orchestrated by the precise behavior and division dynamics of radial glial cells (RGCs). Endocytosis functions in the recycling and remodeling of adherens junctions (AJs) in response to changes in RGC activity and function. Here, we show that conditional disruption of ubiquitin-associated protein 1 (UBAP1), a component of endosomal sorting complex required for transport (ESCRT), causes severe brain dysplasia and prenatal ventriculomegaly. UBAP1 depletion disrupts the AJs and polarity of RGCs, leading to failure of apically directed interkinetic nuclear migration. Accordingly, UBAP1 knockout or knockdown results in reduced proliferation and precocious differentiation of neural progenitor cells. Mechanistically, UBAP1 regulates the expression and surface localization of cell adhesion molecules, and ß-catenin over-expression significantly rescues the phenotypes of Ubap1 knockdown in vivo. Our study reveals a critical physiological role of the ESCRT machinery in cortical neurogenesis by regulating AJs of RGCs.

Virtual agents among participants with methamphetamine use disorders: Acceptability and usability study.

INTRODUCTION: While the potential future role of virtual agents (VAs) in treating addiction is promising, participants' attitudes toward the use of VAs in psychotherapy remain insufficiently investigated. This lack of investigation could pose barriers to the adoption of VA-led psychotherapy for people with substance use disorders (SUD). This research aims to explore the acceptability and usability of VAs for people with methamphetamine use disorder. METHODS: Following a single session of psychotherapy led by VAs through the Echo-app, a group of 49 individuals actively seeking treatment for current DSM-V substance dependence (with a mean age of 39.06 ± 8.02) completed self-administered questionnaires and participated in focus group interviews. These questionnaires aimed to investigate participants' preference regarding the type of psychotherapy and their willingness to engage in VA-led psychotherapy, taking into account their diverse psychological needs. RESULTS: Quantitative data were subjected to analysis through both descriptive and inferential statistical methods. Interestingly, participants exhibited a significantly higher acceptability for traditional face-to-face psychotherapy compared to email-based psychotherapy (p = 0.042), but there was no statistically significant difference between their acceptance of traditional psychotherapy and VA-led psychotherapy (p = 0.059). The questionnaire outcomes indicated participants' willingness to engage in VA-led psychotherapy for purposes such as relapse prevention intervention, addressing emotional issues, managing somatic experiences, and facilitating social and family functional recovery. Furthermore, the participants' attitudes toward VA-led psychotherapy were predicted by factors including the need for anxiety-focused psychotherapy (p = 0.027; OR [95%CI] = 0.14[0.03,0.80]), the presence of chronic somatic diseases (p = 0.017; OR [95%CI] = 13.58[1.59,116.03]), and marital status (p = 0.031; OR [95%CI] = 5.02[1.16,21.79]). DISCUSSION: Through the interviews, the study uncovered the factors that either supported or hindered participants' experiences with VA-led psychotherapy, while also gathering suggestions for future improvements. This research highlights the willingness and practicality of individuals with SUD in embracing VA-led psychotherapy. The findings are anticipated to contribute to the refinement of VA-led tools to better align with the preferences and needs of the users.

Extending the Coverage of Lys-C/Trypsin-Based Bottom-up Proteomics by Cysteine S-Aminoethylation.

To improve the coverage in bottom-up proteomics, S-aminoethylation of cysteine residues (AE-Cys) was carried out with 2-bromoethylamine, followed by cleavage with lysyl endopeptidase (Lys-C) or Lys-C/trypsin. A model study with bovine serum albumin showed that the C-terminal side of AE-Cys was successfully cleaved by Lys-C. The frequency of side reactions at amino acids other than Cys was less than that in the case of carbamidomethylation of Cys with iodoacetamide. Proteomic analysis of A549 cell extracts in the data-dependent acquisition mode after AE-Cys modification afforded a greater number of identified protein groups, especially membrane proteins. In addition, label-free quantification of proteins in mouse nonsmall cell lung cancer (NSCLC) tissue in the data-independent acquisition mode after AE-Cys modification showed improved NSCLC pathway coverage and greater reproducibility. Furthermore, the AE-Cys method could identify an epidermal growth factor receptor peptide containing the T790 M mutation site, a well-established lung-cancer-related mutation site that has evaded conventional bottom-up methods. Finally, AE-Cys was found to fully mimic Lys in terms of collision-induced dissociation fragmentation, ion mobility separation, and cleavage by Lys-C/trypsin, except for sulfoxide formation during sample preparation.

Improving Sensitivity and Reproducibility of Surface-Enhanced Raman Scattering Biochips Utilizing Magnetoplasmonic Nanoparticles and Statistical Methods.

Surface-enhanced Raman scattering (SERS) technology has been widely recognized for its remarkable sensitivity in biochip development. This study presents a novel sandwich immunoassay that synergizes SERS with magnetoplasmonic nanoparticles (MPNs) to improve sensitivity. By taking advantage of the unique magnetism of these nanoparticles, we further enhance the detection sensitivity of SERS biochips through the applied magnetic field. Despite the high sensitivity, practical applications of SERS biochips are often limited by the issues of stability and reproducibility. In this study, we introduced a straightforward statistical method known as "Gaussian binning", which involves initially binning the two-dimensional Raman mapping data and subsequently applying Gaussian fitting. This approach enables a more consistent and reliable interpretation of data by reducing the variability inherent in Raman signal measurements. Based on our method, the biochip, targeting for C-reactive protein (CRP), achieves an impressive detection limit of 5.96 fg/mL, and with the application of a 3700 G magnetic field, it further enhances the detection limit by 5.7 times, reaching 1.05 fg/mL. Furthermore, this highly sensitive and magnetically tunable SERS biochip is easily designed for versatile adaptability, enabling the detection of other proteins. We believe that this innovation holds promise in enhancing the clinical applicability of SERS biochips.

Atomically Dispersed ZnN4 Sites Anchored on P-Functionalized Carbon with Hierarchically Ordered Porous Structures for Boosted Electroreduction of CO2.

Tuning the coordination structures of metal sites is intensively studied to improve the performances of single-atom site catalysts (SASC). However, the pore structure of SASC, which is highly related to the accessibility of active sites, has received little attention. In this work, single-atom ZnN4 sites embedded in P-functionalized carbon with hollow-wall and 3D ordered macroporous structure (denoted as H-3DOM-ZnN4 /P-C) are constructed. The creation of hollow walls in ordered macroporous structures can largely increase the external surface area to expose more active sites. The introduction of adjacent P atoms can optimize the electronic structure of ZnN4 sites through long-rang regulation to enhance the intrinsic activity and selectivity. In the electrochemical CO2 reduction reaction, H-3DOM-ZnN4 /P-C exhibits high CO Faradaic efficiency over 90% in a wide potential window (500 mV) and a large turnover frequency up to 7.8 × 104  h-1 at -1.0 V versus reversible hydrogen electrode, much higher than its counterparts without the hierarchically ordered structure or P-functionalization.

A direct healthcare cost analysis of recombinant LH versus hMG supplementation on FSH during controlled ovarian hyperstimulation in the GnRH-antagonist protocol.

PURPOSE: We have previously published a retrospective matched-case control study comparing the effect of recombinant LH (r-hLH) versus highly purified human menopausal gonadotropin (hMG) supplementation on the follicle-stimulating hormone (FSH) during controlled ovarian hyperstimulation (COH) in the GnRH-antagonist protocol. The result from that study showed that the cumulative live birth rate (CLBR) was significantly higher in the r-hLH group (53% vs. 64%, p = 0.02). In this study, we aim to do a cost analysis between these two groups based on our previous study. METHODS: The analysis consisted of 425 IVF and ICSI cycles in our previous study. There were 259 cycles in the r-hFSH + hMG group and 166 cycles in the r-hFSH + r-hLH group. The total cost related to the treatment of each patient was recorded. Probabilistic sensitivity analysis (PSA) and a cost-effectiveness acceptability curve (CEAC) were performed and created. RESULTS: The total treatment cost per patient was significantly higher in the r-hFSH + r-hLH group than in the r-hFSH + hMG group ($4550 ± 798.86 vs. $4290 ± 734.6, p = 0.003). However, the mean cost per live birth in the r-hFSH + hMG group was higher at $8052, vs. $7059 in the r-hFSH + r-hLH group. The CEAC showed that treatment with hFSH + r-hLH proved to be more cost-effective than treatment with r-hFSH + hMG. Willingness-to-pay was evident when considering a hypothetical threshold of $18,513, with the r-hFSH + r-hLH group exhibiting a 99% probability of being considered cost-effective. CONCLUSION: The cost analysis showed that recombinant LH is more cost-effective than hMG supplementation on r-hFSH during COH in the GnRH-antagonist protocol.

Modulating SARS-CoV-2 Spike Protein Reactivity through Moderate Electric Fields: A Pathway to Innovative Therapies.

In the quest for effective COVID-19 treatments and vaccines, traditional biochemical methods have been paramount, yet the challenge of accommodating diverse viral mutants persists. Recent simulations propose an innovative physical strategy involving an external electric field applied to the SARS-CoV-2 spike protein, demonstrating a reduced viral binding potential. However, limited empirical knowledge exists regarding the characteristics of the spike protein after E-field treatment. Our study addresses this gap by employing diverse analytical techniques to elucidate the impact of low/moderate E-field intensity on the binding of the SARS-CoV-2 spike protein to the ACE2 receptor. Through comprehensive analysis, we unveil a substantial reduction in the spike protein binding capacity validated via enzyme-linked immunosorbent assay and quartz crystal microbalance experiments. Remarkably, the E-field exposure induces significant protein structure rearrangement, leading to an enhanced negative surface zeta potential confirmed by dynamic light scattering. Circular dichroism spectroscopy corroborates these structural changes, showing alterations in the secondary protein structures. This study provides insights into SARS-CoV-2 spike protein modification under an E-field pulse, potentially paving the way for nonbiochemical strategies to mitigate viral reactivity and opening avenues for innovative therapeutic and preventive approaches against COVID-19 and its evolving variants.

Clinical features and imaging examination assessment of cervical lymph nodes for thyroid carcinoma.

BACKGROUNDS: The purpose of this study is to investigate the relationship between clinical characteristics and cervical lymph node metastasis (LNM) in patients with thyroid carcinoma, as well as estimate the preoperative diagnosis values of ultrasound (US) and contrast enhanced computed tomography (CECT) examinations on the neck for detection of cervical LNM in thyroid carcinoma. METHODS: A retrospective analysis of 3 026 patients with surgically proven thyroid carcinoma was conducted. Patients' clinical characteristics, including gender, age, tumor size, bilateral lesions, multifocality, adenomatous nodules, Hashimoto's thyroiditis (HT), and extrathyroidal extension, were collected to explore their association with cervical LNM in thyroid carcinoma. Preoperative assessments for central lymph node metastasis (CLNM) and lateral lymph node metastasis (LLNM) were conducted through US and CECT. The diagnostic value of US, CECT and US combined with CECT for detection of LNM located in various cervical compartments was estimated based on the pathological results. RESULTS: The risk of cervical LNM was higher in thyroid cancer patients who were male, age < 55 years old, tumor size > 10 mm, bilateral lesions, and extrathyroidal extension, while multifocality, adenomatous nodules and HT had no significant effect on LNM. US, CECT and US combined with CECT all had a higher sensitivity to LLNM (93.1%, 57.8%, 95.4%) than to CLNM (32.3%, 29.0%, 43.4%). US and CECT had a high specificity to both CLNM and LLNM (94.3-97.8%). CONCLUSION: Preoperative clinical characteristics and imaging examinations on patients with thyroid carcinoma are crucial to the evaluation of cervical lymph nodes and conducive to individualizing surgical treatments by clinicians. US combined with CECT are superior to single US or CECT alone in detection of CLNM and LLNM.

AI diagnosis of Bethesda category IV thyroid nodules.

Thyroid nodules are a common disease, and fine needle aspiration cytology (FNAC) is the primary method to assess their malignancy. For the diagnosis of follicular thyroid nodules, however, FNAC has limitations. FNAC can classify them only as Bethesda IV nodules, leaving their exact malignant status and pathological type undetermined. This imprecise diagnosis creates difficulties in selecting the follow-up treatment. In this retrospective study, we collected ultrasound (US) image data of Bethesda IV thyroid nodules from 2006 to 2022 from five hospitals. Then, US image-based artificial intelligence (AI) models were trained to identify the specific category of Bethesda IV thyroid nodules. We tested the models using two independent datasets, and the best AI model achieved an area under the curve (AUC) between 0.90 and 0.95, demonstrating its potential value for clinical application. Our research findings indicate that AI could change the diagnosis and management process of Bethesda IV thyroid nodules.

Interaction and assembly of the DNA replication core proteins of Kaposi's sarcoma-associated herpesvirus.

IMPORTANCE: Eukaryotic DNA replication is a highly regulated process that requires multiple replication enzymes assembled onto DNA replication origins. Due to the complexity of the cell's DNA replication machinery, most of what we know about cellular DNA replication has come from the study of viral systems. Herein, we focus our study on the assembly of the Kaposi's sarcoma-associated herpesvirus core replication complex and propose a pairwise protein-protein interaction network of six highly conserved viral core replication proteins. A detailed understanding of the interaction and assembly of the viral core replication proteins may provide opportunities to develop new strategies against viral propagation.