The egg ribonuclease SjCP1412 accelerates liver fibrosis caused by Schistosoma japonicum infection involving damage-associated molecular patterns (DAMPs).

Schistosomiasis, a parasite infectious disease caused by Schistosoma japonicum, often leads to egg granuloma and fibrosis due to the inflammatory reaction triggered by egg antigens released in the host liver. This study focuses on the role of the egg antigens CP1412 protein of S. japonicum (SjCP1412) with RNase activity in promoting liver fibrosis. In this study, the recombinant egg ribonuclease SjCP1412, which had RNase activity, was successfully prepared. By analysing the serum of the population, it has been proven that the anti-SjCP1412 IgG in the serum of patients with advanced schistosomiasis was moderately correlated with liver fibrosis, and SjCP1412 may be an important antigen associated with liver fibrosis in schistosomiasis. In vitro, the rSjCP1412 protein induced the human liver cancer cell line Hep G2 and liver sinusoidal endothelial cells apoptosis and necrosis and the release of proinflammatory damage-associated molecular patterns (DAMPs). In mice infected with schistosomes, rSjCP1412 immunization or antibody neutralization of SjCP1412 activity significantly reduced cell apoptosis and necroptosis in liver tissue, thereby reducing inflammation and liver fibrosis. In summary, the SjCP1412 protein plays a crucial role in promoting liver fibrosis during schistosomiasis through mediating the liver cells apoptosis and necroptosis to release DAMPs inducing an inflammatory reaction. Blocking SjCP1412 activity could inhibit its proapoptotic and necrotic effects and alleviate hepatic fibrosis. These findings suggest that SjCP1412 may be served as a promising drug target for managing liver fibrosis in schistosomiasis japonica.

Protective effect of wuzibushen recipe on follicular development via regulating androgen receptor in polycystic ovary syndrome model rats.

OBJECTIVES: The study aimed to explore the role and mechanism of WZBS recipe on PCOS. METHODS: PCOS model was established. After modeling, PCOS rats were intragastrically administered with Diane-35 or WZBS recipe (6.93 g/kg/d). Then, the ovarian and uterine morphology were observed, the estrous cycle was assessed. HE and oil red O staining were conducted for ovarian morphological analysis and counting ovarian follicle and corpora lutea number. Furthermore, the serum content of testosterone (T) and sex-hormone-binding globulin (SHBG) were assessed by ELISA kits. The androgen receptor (AR), CX43 mRNA and protein expression were measured by q-PCR and Western blot. RESULTS: WZBS recipe increased uterine implanted blastocysts, reduced cystic dilated follicles, and normalized estrous cycle in PCOS rats. Meanwhile, WZBS recipe alleviated ovarian injury, increased mature follicles and corpora lutea number in PCOS rats. Moreover, WZBS recipe decreased serum T content, AR expression and increased serum SHBG content, CX43 expression in PCOS rats. CONCLUSIONS: This study reveals that WZBS recipe may attenuate PCOS by protecting follicular development via down-regulating AR.

Few Layer Ti3C2 MXene-Based Label-Free Aptasensor for Ultrasensitive Determination of Chloramphenicol in Milk.

Quantitative detection of veterinary drug residues in animal-derived food is of great significance. In this work, a simple and label-free electrochemical aptasensor for the highly sensitive detection of chloramphenicol (CAP) in milk was successfully developed based on a new biosensing method, where the single- or few-layer Ti3C2 MXene nanosheets functionalized via the specific aptamer by self-assembly were used as electrode modifiers for a glassy carbon electrode (aptamer/Ti3C2 MXene/GCE). Differential pulse voltammetry (DPV), electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV), scanning electron microscopy (SEM), atomic force microscope (AFM), and so on were utilized for electrochemical and morphological characterization. Under the optimized conditions, the constructed aptasensor exhibited excellent performance with a wider linearity to CAP in the range from 10 fM to 1 µM and a low detection limit of 1 fM. Aptamer/Ti3C2 MXene/GCE demonstrated remarkable selectivity over other potentially interfering antibiotics, as well as exceptional reproducibility and stability. In addition, the aptasensor was successfully applied to determine CAP in milk with acceptable recovery values of 96.13% to 108.15% and relative standard deviations below 9%. Therefore, the proposed electrochemical aptasensor is an excellent alternative for determining CAP in food samples.

Unravelling the Complex LiOH-Based Cathode Chemistry in Lithium-Oxygen Batteries.

The LiOH-based cathode chemistry has demonstrated potential for high-energy Li-O2 batteries. However, the understanding of such complex chemistry remains incomplete. Herein, we use the combined experimental methods with ab initio calculations to study LiOH chemistry. We provide a unified reaction mechanism for LiOH formation during discharge via net 4 e- oxygen reduction, in which Li2 O2 acts as intermediate in low water-content electrolyte but LiHO2 as intermediate in high water-content electrolyte. Besides, LiOH decomposes via 1 e- oxidation during charge, generating surface-reactive hydroxyl species that degrade organic electrolytes and generate protons. These protons lead to early removal of LiOH, followed by a new high-potential charge plateau (1 e- water oxidation). At following cycles, these accumulated protons lead to a new high-potential discharge plateau, corresponding to water formation. Our findings shed light on understanding of 4 e- cathode chemistries in metal-air batteries.

Structure-guided rational design of the Geobacillus thermoglucosidasius feruloyl esterase GthFAE to improve its thermostability.

Feruloyl esterases are indispensable biocatalysts catalyzing the cleavage of ester bonds between polysaccharides and their hydroxycinnamoyl cross-links. GthFAE from Geobacillus thermoglucosidasius was identified as a thermophilic alkaline feruloyl esterase with potential applications in paper manufacturing. To improve the enzymatic properties rationally and efficiently, the structure of GthFAE was solved at 1.9 Å, revealing a core domain of classical α/ß hydrolase fold and an inserted α/ß cap domain. In silico analysis based on it helped us to investigate whether the residues at the active center have positive effects on the stability, and how. Several site-directed mutations were conducted, of which substitutions at residues T41 and T150 apparently improved the thermostability. The combination mutant T41N/T150R exhibited an optimal temperature of 65 °C, a 6.4 °C higher Tm compared to wild type by 80 °C, and a 35-fold longer in half-life (201 min) at 70 °C. Molecular dynamics simulations further illustrated that the structure of T41N/T150R was more stable than the wild type and T150R stabilized the cap domain by introducing salt bridges to the region with E154 and D164. This study not only highlighted residues within the active center on their thermostability improving effects, but also contributed to the prospective industrial application of GthFAE.

Structural insights into the CP312R protein of the African swine fever virus.

African swine fever (ASF) is a lethal hemorrhagic disease that affects domestic pigs and wild boars. There is no medication available for ASF to date. The ability to mount antigen-specific responses to viral vectored CP312R makes it a crucial potential target for designing vaccines or drugs. This study determined the crystal structure of ASFV CP312R at 2.32 Å and found it to be a monomer with a single-stranded DNA binding core domain with a clear five-strands ß-barrel OB-fold architecture. Electrophoretic mobility shift assay and size-exclusion chromatography characterization assay further confirmed the single-stranded DNA (ssDNA)-binding property of ASFV CP312R. This study revealed the structure and preliminary ssDNA interaction mechanisms of ASFV CP312R, providing new clues for developing new antiviral strategies.

In Situ Halogen-Ion Leaching Regulates Multiple Sites on Tandem Catalysts for Efficient CO2 Electroreduction to C2+ Products.

Tandem catalysts can divide the reaction into distinct steps by local multiple sites and thus are attractive to trigger CO2 RR to C2+ products. However, the evolution of catalysts generally exists during CO2 RR, thus a closer investigation of the reconstitution, interplay, and active origin of dual components in tandem catalysts is warranted. Here, taking AgI-CuO as a conceptual tandem catalyst, we uncovered the interaction of two phases during the electrochemical reconstruction. Multiple operando techniques unraveled that in situ iodine ions leaching from AgI restrained the entire reduction of CuO to acquire stable active Cu0 /Cu+ species during the CO2 RR. This way, the residual iodine species of the Ag matrix accelerated CO generation and iodine-induced Cu0 /Cu+ promotes C-C coupling. This self-adaptive dual-optimization endowed our catalysts with an excellent C2+ Faradaic efficiency of 68.9 %. Material operando changes in this work offer a new approach for manipulating active species towards enhancing C2+ products.

Improvement of solution-processed Zn-Sn-O active-layer thin film transistors by novel high valence Mo doping.

In this letter, the performance of Zn-Sn-O (ZTO) thin film transistors (TFTs) has been greatly improved by Mo doping as an oxygen vacancy to control the residual electrons. The results show that the TFT with 3 at% Mo doping exhibits the best electrical characteristics with a high saturation mobility of 26.53 cm2 V-1 s-1, a threshold voltage of 0.18 V, a subthreshold swing of 0.32 V dec-1 and a large switching ratio of 2 × 106. The saturation mobility and switching ratio of Mo-doped Zn-Sn-O (MZTO, 3 at%) TFTs improved almost five and two orders of magnitude compared with ZTO TFTs, respectively. Therefore, the MZTO TFT has much potential for future electrical applications with its excellent properties.

Runjing Decoction alleviated cyclophosphamide-induced oligoasthenospermia rats by inhibiting cell apoptosis via RXFP1/AKT/FOXO1 pathway.

Runjing Decoction (RJD) is a prescription of traditional Chinese medicine for the treatment of oligoasthenospermia. However, the molecular mechanism of RJD on oligoasthenospermia still remains unknown. A model of oligoasthenospermia was induced in 30 Sprague Dawley rats by intraperitoneal injection of cyclophosphamide at 35 mg/kg per day for 5 days and treated by intragastric RJD (13.5 g/kg) or L-carnitine (100 mg/kg) for 14 days. The body weight, testis and epididymis weight, grade A spermatozoa, grade B spermatozoa, the percentage of sperm forward motility (PR%), the sperm activity rate and the sperm density of rats were evaluated before and after RJD treatment. The testis apoptosis was determined by TUNEL staining. The expressions of RXFP1, FoxO1, PI3K, Akt, Bax and Bcl-2 were determined by qRT-PCR and Western blot, respectively. After RJD treatment, the grade A spermatozoa, sperm PR%, sperm activity and sperm density were significantly increased relative to those in model rats. Cell apoptosis of testis tissue was reversed by RJD. RJD suppressed cell apoptosis, inhibited the expression of RXFP1, FOXO1, PI3K, AKT and Bax, and promoted the expression levels of Bcl-2 in testicular tissue of oligoasthenospermia rats. RJD could alleviate sperm quality and testis damage in oligoasthenospermia rats by inhibiting RXFP1/AKT/FOXO1 pathway.

PKCλ/ι regulates Th17 differentiation and house dust mite-induced allergic airway inflammation.

Asthma is a chronic airway inflammation in which Th2 and Th17 cells play critical roles in its pathogenesis. We have reported that atypical protein kinase (PKC) λ/ι is a new regulator for Th2 differentiation and function. However, the role of PKCλ/ι for Th17 cells remains elusive. In this study, we explored the effect of PKCλ/ι on Th17 cells in the context of ex vivo cell culture systems and an in vivo murine model of allergic airway inflammation with the use of activated T cell-specific conditional PKCλ/ι-deficient mice. Our findings indicate that PKCλ/ι regulates Th17 cells. The secretion of Th17 effector cytokines, including IL-17, IL-21 and IL-22, were inhibited from PKCλ/ι-deficient T cells under non-skewing or Th17-skewing culture conditions. Moreover, the impaired Th17 differentiation and function by the PKCλ/ι-deficiency was associated with the downregulation of Stat3 and Rorγt, key Th17 transcription factors. We developed a model of Th17 and neutrophil-involved allergic airway inflammation by intratracheal inoculation of house dust mites. PKCλ/ι-deficiency significantly inhibited airway inflammations. The infiltrating cells in the lungs and bronchoalveolar lavage fluids were significantly reduced in conditional PKCλ/ι-deficient mice. Th17 effector cytokines were reduced in the bronchoalveolar lavage fluids and lungs at protein and mRNA levels. Thus, PKCλ/ι emerges as a critical regulator of Th17 differentiation and allergic airway hyperresponsiveness.

B-cell-activating factor code and human cytomegalovirus infection in renal transplant recipients.

The objective of the present study was to explore the correlation between the BAFF signal and HCMV-TLR activation in RTx recipients complicated by HCMV. Peripheral blood (anticoagulated by EDTA-Na2 ) and urine of 113 RTx recipients were collected; healthy volunteers were controlled. Urine HCMV-DNA was detected by real-time PCR. Recipients were classified into a positive group (>10,000 copies/mL urine) and a negative group (<10,000 copies/mL urine). ELISA results showed that sBAFF, sera anti-HCMV pp65 immunoglobulin (Ig)G antibody, and total IgG all significantly increased in recipients with positive HCMV-DNA (>10,000 copies/mL urine) (P < 0.05) compared with negative recipients (<10,000 copies/mL urine). In the positive group, HCMV-DNA copies and total IgG positively correlated with sBAFF (r = 0.988 and 0.625, respectively) (P < 0.05). Luminex assay results suggested that the incidence of anti-HLA I and II and MICA antibody obviously increased in positive recipients. The expression level of BAFF and BAFF-R increased in positive recipients. A total of 88 particular genes-involved in TLR signaling pathways, NF-κB signaling pathways, and cytokine-cytokine receptor signaling pathways-were detected in real-time PCR chip assay. A total of 46 genes were differentially expressed greater than two-fold, and the expression characteristic of BAFF-R was concordant with FACS results. Our findings are that activation of HCMV would induce or enhance the activation of BAFF code in RTx recipients, which may independently or cooperatively participate in renal allograft injury and decrease the long-term outcome of renal allografts.

Blocking BAFF Alleviates Hepatic Fibrosis in Schistosoma japonicum-Infected Mice.

Schistosomiasis is an immunopathogenic disease characterized by egg granuloma and fibrosis. The hepatic fibrosis of schistosomiasis is caused by the coordinated action of local immune cells, liver-resident cells and related cytokines around the eggs of the liver. B-cell-activating factor (BAFF), expressed in many cells, is an essential factor for promoting the survival, differentiation, and maturation of cells. The overexpression of BAFF is closely related to many autoimmune diseases and fibrosis, but has not been reported to play a role in liver fibrosis caused by schistosomiasis. In the study, we found that, during Schistosoma japonicum (S. japonicum) infection in mice, the level of BAFF and its receptor BAFF-R progressively increased, then decreased with the extension of infection time, which was consistent with the progression of hepatic granuloma and fibrosis. Anti-BAFF treatment attenuated the histopathological damage in the liver of infected mice. The average areas of individual granulomas and liver fibrosis in anti-BAFF treatment mice were significantly lower than those in control mice, respectively. Anti-BAFF treatment increased the IL-10, decreased IL-4, IL-6, IL-17A, TGF-ß, and downregulated the antibody level against S. japonicum antigens. These results suggested that BAFF acts a strong player in the immunopathology of schistosomiasis. Anti-BAFF treatment may influence Th2 and Th17 responses, and reduce the inflammatory reaction and fibrosis of schistosomiasis liver egg granuloma. It is suggested that BAFF might be a prospective target for the development of new methods to treat schistosomiasis liver fibrosis.

Toward High-Performance Metal-Organic-Framework-Based Quasi-Solid-State Electrolytes: Tunable Structures and Electrochemical Properties.

Metal-organic frameworks (MOFs) have been reported as promising materials for electrochemical applications owing to their tunable porous structures and ion-sieving capability. However, it remains challenging to rationally design MOF-based electrolytes for high-energy lithium batteries. In this work, by combining advanced characterization and modeling tools, a series of nanocrystalline MOFs is designed, and the effects of pore apertures and open metal sites on ion-transport properties and electrochemical stability of MOF quasi-solid-state electrolytes are systematically studied. It isdemonstrated that MOFs with non-redox-active metal centers can lead to a much wider electrochemical stability window than those with redox-active centers. Furthermore, the pore aperture of MOFs is found to be a dominating factor that determines the uptake of lithium salt and thus ionic conductivity. The ab initio molecular dynamics simulations further demonstrate that open metal sites of MOFs can facilitate the dissociation of lithium salt and immobilize anions via Lewis acid-base interaction, leading to good lithium-ion mobility and high transference number. The MOF quasi-solid-state electrolyte demonstrates great battery performance with commercial LiFePO4 and LiCoO2 cathodes at 30 °C. This work provides new insights into structure-property relationships between tunable structure and electrochemical properties of MOFs that can lead to the development of advanced quasi-solid-state electrolytes for high-energy lithium batteries.

Luminescent MOFs constructed by using butterfly-like AIE ligands.

In this work, a series of butterfly-like isomers named oxacalix[2]naphthalene[2]pyrazine (ONP) were conveniently synthesized by a one-step catalyst-free reaction in a facile manner, and they exhibit typical characteristics of aggregation-induced emission (AIE). The mechanism study shows that restriction of intramolecular vibration (RIV) is the reason for their AIE properties. The pyrazine groups endow ONP molecules with good coordination ability, which makes them ideal ligands for constructing metal-organic frameworks (MOFs). Thus, three ONP-based luminescent MOFs were constructed, and they exhibit intense emission with lifetimes in the order of microseconds. More importantly, different ONP isomers have different binding capacities, and thus only one kind of MOF can be obtained even when using an isomer mixture of ONP ligands. This result suggested that the conformation of ONPs is an important determining factor for their application as bridging ligands. This work not only reports a series of new RIV-type AIEgens, but also offers a new platform for the construction of luminescent MOFs.

Advances in Lithium-Oxygen Batteries Based on Lithium Hydroxide Formation and Decomposition.

The rechargeable lithium-oxygen (Li-O2) batteries have been considered one of the promising energy storage systems owing to their high theoretical energy density. As an alternative to Li-O2 batteries based on lithium peroxide (Li2O2) cathode, cycling Li-O2 batteries via the formation and decomposition of lithium hydroxide (LiOH) has demonstrated great potential for the development of practical Li-O2 batteries. However, the reversibility of LiOH-based cathode chemistry remains unclear at the fundamental level. Here, we review the recent advances made in Li-O2 batteries based on LiOH formation and decomposition, focusing on the reaction mechanisms occurring at the cathode, as well as the stability of Li anode and cathode binder. We also provide our perspectives on future research directions for high-performance, reversible Li-O2 batteries.

Deficiency of PKCλ/ι alleviates the liver pathologic impairment of Schistosoma japonicum infection by thwarting Th2 response.

BACKGROUND: The activation of immune response driven by the eggs of Schistosoma japonicum and the subsequent secretions is the culprit behind granulomatous inflammation and liver fibrosis. Evidence suggests that PKCλ/ι participates in a variety of physiological and pathological processes, including the regulation of metabolism, growth, proliferation and differentiation of cells. However, the role of PKCλ/ι in liver disease caused by Schistosoma japonicum remains unclear. METHODS: In the present study, we observe the pathological changes of egg-induced granulomatous inflammation and fibrosis in the liver of mice infected by Schistosoma japonicum by using conditional PKCλ/ι-knockout mice and wild-type control. Immune cytokines and fibrogenic factors were analyzed by performing flow cytometry and real-time fluorescence quantitative PCR. RESULTS: The results of H&E and Masson staining show that the degree of granulomatous lesions and fibrosis in the liver of the infected PKCλ/ι-knockout mice was significantly reduced compared with those of the infected wild-type mice. The mean area of single granuloma and hepatic fibrosis in the PKCλ/ι-knockout mice was significantly lower than that of the wild-type mice (85,295.10 ± 5399.30 µm2 vs. 1,433,702.04 ± 16,294.01 µm2, P < 0.001; 93,778.20 ± 8949.05 µm2 vs. 163,103.01 ± 11,103.20 µm2, P < 0.001), respectively. Serological analysis showed that the ALT content was significantly reduced in the infected knockout mice compared with infected wild-type mice. RT-PCR analysis showed that IL-4 content in knockout mice was significantly increased after Schistosoma japonicum infection, yet the increase was less than that in infected wild-type mice (P < 0.05). PKCλ/ι deficiency led to reduced expression of fibrosis-related factors, including TGF-ß1, Col-1, Col-3, α-SMA and liver DAMP factor HMGB1. Flow cytometry analysis showed that the increasing percentage of Th2 cells, which mainly secrete IL-4 cytokines in spleen cells, was significantly lower in PKCλ/ι-deficient mice compared with wild-type mice after infection (P < 0.05). CONCLUSIONS: Our data demonstrate that PKCλ/ι deficiency alleviating granulomatous inflammation and fibrosis in the liver of mice with S. japonicum infection by downregulating Th2 immune response is the potential molecular mechanism behind the role of PKCλ/ι in schistosomiasis.

Highly dispersed Cr oxygenated species on Pt nanowire assemblies for enhanced electrocatalytic methanol oxidation.

In this paper, Cr oxygenated species dispersed on Pt nanowire assemblies (PtCrOX NWs) were successfully prepared for the methanol oxidation reaction by a metal precursor dilution strategy. Notably, the PtCrOX NWs catalyst exhibits excellent performance for electrocatalytic methanol oxidation. Density functional theory results revealed that the doped Cr oxygenated species, which moderated the electronic structure of the Pt atoms, can significantly decrease the free energy of COOH* formation, thus leading to superior methanol oxidation performance.

Roles of ERRα and TGF-ß signaling in stemness enhancement induced by 1 µM bisphenol A exposure via human neural stem cells.

Bisphenol A (BPA) is a common industrial chemical widely used to produce various plastics and is known to impair neural stem cells (NSCs). However, the effects of low-dose BPA exposure on the stemness maintenance and differentiation fate of NSCs remain unclear in the infant brain. The present study demonstrated that 1 µM BPA promoted human NSC proliferation and stemness, without significantly increasing apoptosis. The Chip-seq experiments demonstrated that both the cell cycle and the TGF-ß signaling pathway were accelerated after treatment with 1 µM BPA. Subsequently, estrogen-related receptor α (ERRα) gene knockout cell lines were constructed using CRISPR/Cas9. Further western blotting and chromatin immunoprecipitation-PCR experiments demonstrated that BPA maintained cell stemness by binding to an EERα receptor and activating the TGF-ß1 signaling pathway, including the downstream factors Aurora kinases B and Id2. In conclusion, the stemness of NSCs could be maintained by BPA at 1 µM through the activation of the ERRα and TGF-ß1 signaling pathways and could restrain the differentiation of NSCs into neurons. The present research further clarified the mechanism of BPA toxicity on NSCs from the novel perspective of ERRα and TGF-ß1 signaling pathways regulated by BPA and provided insights into potential novel methods of prevention and therapy for neurogenic diseases.

Crystal Structure of the Disease-Specific Protein of Rice Stripe Virus.

The rice stripe virus (RSV) is responsible for devastating effects in East Asian rice-producing areas. The disease-specific protein (SP) level in rice plants determines the severity of RSV symptoms. Isothermal titration calorimetry (ITC) and bimolecular fluorescence complementation (BiFC) assays confirmed the interaction between an R3H domain-containing host factor, OsR3H3, and RSV SP in vitro and in vivo. This study determined the crystal structure of SP at 1.71 Å. It is a monomer with a clear shallow groove to accommodate host factors. Docking OsR3H3 into the groove generates an SP/OsR3H3 complex, which provides insights into the protein-binding mechanism of SP. Furthermore, SP's protein-binding properties and model-defined recognition residues were assessed using mutagenesis, ITC, and BiFC assays. This study revealed the structure and preliminary protein interaction mechanisms of RSV SP, shedding light on the molecular mechanism underlying the development of RSV infection symptoms.

Structural insight into African swine fever virus I73R protein reveals it as a Z-DNA binding protein.

African Swine Fever (ASF) is a highly contagious viral haemorrhagic disease of swine, leading to enormous economic losses in the swine industry. However, vaccines and drugs to treat ASF have yet to be developed. African swine fever virus (ASFV) encodes more than 150 proteins, but 50% of them have unknown functions. Here, we present the crystal structure of the ASFV I73R protein at a resolution of 2.0 Å. Similar search tools based solely on amino acid sequence shows that it has no relationships to any proteins of known function. Interestingly, the overall structure of the I73R protein shares a winged helix-turn-helix fold, structural similarity with the Z-DNA binding domain (Zα). In accordance with this result, the I73R is capable of binding to a CpG repeats DNA duplex, which has a high propensity for forming Z-DNA during the DNA binding assays. In addition, the I73R protein was shown to be expressed at both early and late stages of ASFV post-infection in PAM cells as an 8.9 kDa protein. Immunofluorescence studies revealed that the I73R protein is expressed in the nucleus at early times post-infection and gradually translocated from the nucleus to the cytoplasm. Taken together, these data indicate that the I73R could be a member of Zα family that is important in host-pathogen interaction, which paves the way for the design of inhibitors to target this severe pathogen. Further exploring the biological role of I73R during ASFV infection in vitro and in vivo will provide new clues for development of new antiviral strategies.