Manipulating the Piezoelectric Response of Amino Acid-Based Assemblies by Supramolecular Engineering.

Variation in the molecular architecture significantly affects the electronic and supramolecular structure of biomolecular assemblies, leading to dramatically altered piezoelectric response. However, relationship between molecular building block chemistry, crystal packing and quantitative electromechanical response is still not fully understood. Herein, we systematically explored the possibility to amplify the piezoelectricity of amino acid-based assemblies by supramolecular engineering. We show that a simple change of side-chain in acetylated amino acids leads to increased polarization of the supramolecular arrangements, resulting in significant enhancement of their piezoelectric response. Moreover, compared to most of the natural amino acid assemblies, chemical modification of acetylation increased the maximum piezoelectric tensors. The predicted maximal piezoelectric strain tensor and voltage constant of acetylated tryptophan (L-AcW) assemblies reach 47 pm V-1 and 1719 mV m/N, respectively, comparable to commonly used inorganic materials such as bismuth triborate crystals. We further fabricated an L-AcW crystal-based piezoelectric power nanogenerator that produces a high and stable open-circuit voltage of over 1.4 V under mechanical pressure. For the first time, the illumination of a light-emitting diode (LED) is demonstrated by the power output of an amino acid-based piezoelectric nanogenerator. This work presents the supramolecular engineering toward the systematic modulation of piezoelectric response in amino acid-based assemblies, facilitating the development of high-performance functional biomaterials from simple, readily available, and easily tailored building blocks.

An AIE-Based Probe for Rapid and Ultrasensitive Imaging of Plasma Membranes in Biosystems.

The abnormality of the plasma membrane (PM) is an important biomarker for cell status and many diseases. Hence, visualizing the PM, especially in complex systems, is an emerging field in the life sciences, especially in low-resource settings. Herein, we developed a water-soluble PM-specific probe utilizing electrostatic and hydrophobic interaction strategies with aggregation-induced emission as the signal output. The probe could image the PM with many advanced features (wash-free, ultrafast staining process, excellent PM specificity, and good biocompatibility), which were demonstrated by the PM imaging of neurons. The probe allowed for the first time the imaging of erythrocytes in the complex brain environment through a fluorescence-based method. Moreover, the PM of the epidermal and partial view of the eyeball structure of live zebrafish are also revealed.

High-specificity synthesis of novel monomers by remodeled alcohol hydroxylase.

BACKGROUND: Diols are important monomers for the production of plastics and polyurethanes, which are widely used in our daily life. The medium-chain diols with one hydroxyl group at its subterminal end are able to confer more flexibility upon the synthesized materials. But unfortunately, this type of diols has not been synthesized so far. The strong need for advanced materials impelled us to develop a new strategy for the production of these novel diols. In this study, we use the remodeled P450BM3 for high-specificity production of 1,7-decanediol. RESULTS: The native P450BM3 was capable of converting medium-chain alcohols into corresponding α, ω1-, α, ω2- and α, ω3-diols, with each of them accounting for about one third of the total diols, but it exhibited a little or no activity on the short-chain alcohols. Greatly improved regiospecificity of alcohol hydroxylation was obtained by laboratory evolution of P450BM3. After substitution of 12 amino acid residues (J2-F87A), the ratio of 1,7-decanediol (ω-3 hydroxylation) to total decanediols increased to 86.8 % from 34.0 %. Structure modeling and site-directed mutagenesis demonstrated that the heme end residues such as Ala(78), Phe(87) and Arg(255) play a key role in controlling the regioselectivity of the alcohol hydroxylation, while the residues at the mouth of substrate binding site is not responsible for the regioselectivity. CONCLUSIONS: Herein we employ an engineered P450BM3 for the first time to enable the high-specificity biosynthesis of 1,7-decanediol, which is a promising monomer for the development of advanced materials. Several key amino acid residues that control the regioselectivity of alcohol hydroxylation were identified, providing some new insights into how to improve the regiospecificity of alcohol hydroxylation. This report not only provides a good strategy for the biosynthesis of 1,7-decanediol, but also gives a promising approach for the production of other useful diols.

Single and combined effects of microplastics and cadmium on the sea cucumber Apostichopus japonicus.

Microplastic pollution of the ocean has received extensive attention as plastic pollution increases globally, but the potential ecological risks caused by microplastic interactions with trace metals still require further research. In this study, Apostichopus japonicus was used to explore the individual and combined toxicities of cadmium (Cd) and microplastics and their effects on growth, Cd tissue accumulation, digestive enzymes, and gut microbes. The body weight gain and specific growth rate of animals exposed to a combination of high concentrations of Cd and microplastics decreased. The addition of high concentrations of cadmium to the diet led to an increase in cadmium content in the respiratory tree, digestive tract and body wall. Amylase, lipase and trypsin decreased to different degrees in the group treated with high concentrations of Cd/microplastics. Firmicutes were significantly reduced across multiple treatment groups, with the order Lactobacillales being the most significantly affected. Cd is the pollutant causing the greatest negative impact, but the presence of microplastics undoubtedly increases its toxicity.

Preparation and characterization of silk fibroin as a biomaterial with potential for drug delivery.

BACKGROUND: Degummed silk fibroin from Bombyx mori (silkworm) has potential carrier capabilities for drug delivery in humans; however, the processing methods have yet to be comparatively analyzed to determine the differential effects on the silk protein properties, including crystalline structure and activity. METHODS: In this study, we treated degummed silk with four kinds of calcium-alcohol solutions, and performed secondary structure measurements and enzyme activity test to distinguish the differences between the regenerated fibroins and degummed silk fibroin. RESULTS: Gel electrophoresis analysis revealed that Ca(NO3)2-methanol, Ca(NO3)2-ethanol, or CaCl2-methanol treatments produced more lower molecular weights of silk fibroin than CaCl2-ethanol. X-ray diffraction and Fourier-transform infrared spectroscopy showed that CaCl2-ethanol produced a crystalline structure with more silk I (α-form, type II ß-turn), while the other treatments produced more silk II (ß-form, anti-parallel ß-pleated sheet). Solid-State 13C cross polarization and magic angle spinning-nuclear magnetic resonance measurements suggested that regenerated fibroins from CaCl2-ethanol were nearly identical to degummed silk fibroin, while the other treatments produced fibroins with significantly different chemical shifts. Finally, enzyme activity test indicated that silk fibroins from CaCl2-ethanol had higher activity when linked to a known chemotherapeutic drug, L-asparaginase, than the fibroins from other treatments. CONCLUSIONS: Collectively, these results suggest that the CaCl2-ethanol processing method produces silk fibroin with biomaterial properties that are appropriate for drug delivery.

Incorporating Ag@RF core-shell nanomaterials into the thin film nanocomposite membrane to improve permeability and long-term antibacterial properties for nanofiltration.

Ag@resorcinol-formaldehyde resin (Ag@RF) core-shell nanomaterials were prepared by Stöber method, and introduced into polyamide (PA) selective layer of thin-film nanocomposite (TFN) membranes through the interfacial polymerization (IP) process. Due to the abundant hydroxyl groups on the surface and suitable particle size, Ag@RF nanoparticles (Ag@RFs) could be uniformly dispersed in the piperazine aqueous solution and participate in the IP process to precisely regulate the microstructure of the PA selective layer. The resulting "crater structure" and irregular granular structure enlarged the permeable area and contributed to the surface hydrophilicity. For the nanofiltration application, the water flux of TFN membrane modified by Ag@RFs to Na2SO4 solution reached 150 L·m-2·h-1 which was 87.5% greater than TFC, and salt rejection was maintained. The antibacterial efficiency of the prepared TFN membrane on E. coli reached 99.6% in the antibacterial experiment. In addition, due to the special structure of Ag@RFs, the TFN membrane also showed an expected slow-release capability of Ag+, allowing for long-term anti-biofouling properties. This work demonstrates that Ag@RF core-shell nanoparticles with high compatibility of organic nanoparticles and antibacterial properties of Ag nanoparticles could be used as promising nanofillers for designing functional nanofiltration TFN membranes.

Characterization, expression, and functional analysis of the pathogenesis-related gene PtDIR11 in transgenic poplar.

Lignins and lignans are important for plant resistance to pathogens. Dirigent (DIR) proteins control the regio- and stereo-selectivity of coniferyl alcohol in lignan and lignin biosynthesis. DIR genes have been implicated in defense-related responses in several plant species, but their role in poplar immunity is unclear. We cloned PtDIR11 from Populus trichocarpa; we found that overexpression of PtDIR11 in poplar improved the lignan biosynthesis and enhanced the resistance of poplar to Septotis populiperda. PtDIR11 has a typical DIR domain; it belongs to the DIR-b/d family and is expressed in the cell membrane. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis showed that PtDIR11 expression was highest in stems, followed by leaves and roots. Furthermore, PtDIR11 expression was induced by S. populiperda, salicylic acid (SA), jasmonate (JA), and ethylene (ET) stresses. The recombinant PtDIR11 protein inhibited the growth of S. populiperda in vitro. Overexpressing (OE) PtDIR11 in "Nanlin 895" poplar enhanced growth. The OE lines exhibited minimal changes in lignin content, but their total lignan and flavonoid contents were significantly greater than in the wild-type (WT) lines. Overexpression of PtDIR11 affected multiple biological pathways of poplar, such as phenylpropanoid biosynthesis. The methanol extracts of OE-PtDIR11 lines showed greater anti-S. populiperda activity than did lignin extracts from the WT lines. Furthermore, OE-PtDIR11 lines upregulated genes that were related to phenylpropanoid biosynthesis and genes associated with the JA and ET signal transduction pathways; it downregulated genes that were related to SA signal transduction compared with the WT line under S. populiperda stress. Therefore, the OE transgenic plants analysis revealed that PtDIR11 is a good candidate gene for breeding of disease resistant poplar.

Construction of a fungal consortium for effective degradation of rice straw lignin and potential application in bio-pulping.

To improve the lignin degradation efficiency, we established a co-culture consortium (LDFC) consisting of Trametes hirsuta BYL-3, Trametes versicolor BYL-7 and Trametes hirsuta BYL-8. The testing results showed that the constructed consortium showed improved the lignin degradation rate by fungi. The optimal cultivation conditions were mixture at 1:1:1 vol ratio of each fungus, 7% (w/v) of inoculum amount, culture temperature at 26 °C, pH was 6.9 and 10 days of culturing time. Under these conditions, the degradation rate of lignin was 39.7%, which was 9.3% higher than those before optimization (30.4%). Using rice straw for treatment by LDFC to papermaking, the paper tensile strength was 8 N, and the ring pressure index was 2.46 N·m/g, which meets the standards for the production of corrugated paper for packaging. These results indicate that LDFC has potential application value to convert rice straw resources for bio-pulping to make papers.

Plastic mulching reduces nitrogen footprint of food crops in China: A meta-analysis.

Sustainably feeding the growing population amid rising global temperatures and dwindling resources is a grand challenge facing mankind. Plastic mulching (PM) is widely used in China aiming to the increase of crop productivity. However, the impact of PM on reactive nitrogen (Nr) emissions and nitrogen (N) footprint has not been explicitly described. In this study, we collected 4051 observations from 394 published papers for potato (Solanum tuberosum L.), maize (Zea mays L.), and wheat (Triticum aestivum L.), and used meta-analysis to investigate how PM affected crop yield, net economic return, Nr emissions, and N footprints including nitrogen footprint per unit of output energy (NFo) and nitrogen footprint per unit of net economic return (NFe) at regional scale and across a range of precipitation and N fertilization gradients in China. The meta-analysis showed that compared to non-PM practice, PM increased grain yield by 25, 27, and 20% in potato, maize, and wheat, respectively, and enhanced net economic return by 19, 29, and 22%, respectively, with corresponding reduction in NFo of 24, 36, and 18% and NFe of 19, 37, and 19%, respectively. Potato and maize had greater energy output and net economic return than wheat. Plastic mulching was more effective in improving net economic return (or energy output) and reducing N footprints (i.e., NFe and NFo) in the semiarid region (i.e., annual precipitation <600 mm) when N was applied at 100-200 kg N ha-1, especially in potato and maize. Our analysis suggests that the use of PM enhanced grain yield and net economic return while lowering the N footprint without increasing Nr emission. Therefore, PM has great potential to mitigate Nr loss in China when crop species, N fertilization rate, and local environmental factors (i.e., growing region and annual precipitation) are appropriately considered.

In vitro and in vivo investigations of a-C/a-C:Ti nanomultilayer coated Ti6Al4V alloy as artificial femoral head.

Hydrogen-free a-C/a-C:Ti nanomultilayer (a-C NM) films were deposited on medical Ti6Al4V by the magnetron sputtering technique under bias-graded voltage. Cell tests and implantations were performed for the a-C NM films coated Ti6Al4V with the uncoated Ti6Al4V as the control. The canine total hip arthroplasty (THA) surgeries were conducted for 12 dogs using the coated femoral heads, with the CoCr heads as the control. Results of cell tests showed that the coated Ti6Al4V had no cytotoxicity, and there was no statistical difference of the cell attachment rates between the coated and uncoated sample (P = 0.091). No significant difference of the tissue response around the coated and uncoated implants were observed after the intramuscular (P = 0.679) and intraosseous implantations (P = 0.122). After two years of successful canine THA, the polyethylene wear particles isolated from periprosthetic soft tissue showed similar sizes, shapes and counts in the two groups (all of the P values >0.05). The retrieved femoral heads showed slightly change of the surface roughness, but no statistical differences between groups (P = 0.696). However, the systemic metal ion analysis indicated that the content of Co and Cr ions released in the coated group (Co: 0.71 ±â€¯0.06 µg/L, Cr: 0.52 ±â€¯0.05 µg/L) were significant lower than that in the control (Co: 1.98 ±â€¯0.16 µg/L, Cr: 1.17 ±â€¯0.19 µg/L) (both P < 0.005). Histological analysis of the periprosthetic tissue in CoCr group showed a severer histiocyte response than that in the coated group (P = 0.029). The head-taper interfaces showed galvanic corrosion attack in the CoCr group, but not in the coated Ti6Al4V group. Therefore, the a-C NM films coated Ti6Al4V exhibited good biocompatibility as an implant material. Compared with the CoCr, the coated Ti6Al4V femoral head could provide comparable in vivo wear properties, release less harmful metal ions and reduce the inflammatory response in periprosthetic tissue, which may help to prolong the longevity of prostheses.

Mechanical Properties and in Vitro and in Vivo Biocompatibility of a-C/a-C:Ti Nanomultilayer Films on Ti6Al4V Alloy as Medical Implants.

Hydrogen-free a-C/a-C:Ti nanomultilayer films are deposited on medical Ti6Al4V alloy using a closed field unbalanced magnetron sputtering under graded bias voltage. The mechanical and tribological properties of the nanomultilayer films are performed on the nanoindentor, Rockwell and scratch tests, and ball-on-disk tribometer. The biological properties are evaluated by cell cytotoxicity, genotoxicity, subchronic systemic toxicity and implant. The hard a-C/a-C:Ti nanomultilayer films on medical alloy exhibit high adhesion strength and excellent tribological properties in both ambient air and Hank's solution. Biocompatibility results reveal the film no cytotoxity, no genotoxicity, no subchronic systemic toxicity and no contraindications in implant systems. Because of excellent mechanical properties and biosafety, the carbon-based films on medical alloy unveils a prospective application in medical implants.

Associations Between Ventilator Bundle Components and Outcomes.

IMPORTANCE: Ventilator bundles, including head-of-bed elevation, sedative infusion interruptions, spontaneous breathing trials, thromboprophylaxis, stress ulcer prophylaxis, and oral care with chlorhexidine gluconate, are ubiquitous, but the absolute and relative value of each bundle component is unclear. OBJECTIVE: To evaluate associations between individual and collective ventilator bundle components and ventilator-associated events, time to extubation, ventilator mortality, time to hospital discharge, and hospital death. DESIGN, SETTING, AND PARTICIPANTS: This retrospective cohort study included all 5539 consecutive patients who underwent mechanical ventilation for at least 3 days from January 1, 2009, to December 31, 2013, at Brigham and Women's Hospital. EXPOSURES: Head-of-bed elevation, sedative infusion interruptions, spontaneous breathing trials, thromboprophylaxis, stress ulcer prophylaxis, and oral care with chlorhexidine. MAIN OUTCOMES AND MEASURES: Hazard ratios (HRs) for ventilator-associated events, extubation alive vs ventilator mortality, and hospital discharge vs hospital death. Effects were modeled using Cox proportional hazards regression and Fine-Gray competing risk models adjusted for patients' demographic characteristics, comorbidities, unit type, severity of illness, recent procedures, process measure contraindications, day-to-day markers of clinical status, and calendar year. RESULTS: Of 5539 consecutive patients undergoing mechanical ventilation, 3208 were male (57.9%), 2331 female (42.1%), and the mean (SD) age was 61.2 (16.1) years. Sedative infusion interruptions were associated with less time to extubation (HR, 1.81; 95% CI, 1.54-2.12; P < .001) and a lower hazard for ventilator mortality (HR, 0.51, 95% CI, 0.38-0.68; P < .001). Similar associations were found for spontaneous breathing trials (HR for extubation, 2.48; 95% CI 2.23-2.76; P < .001; HR for mortality, 0.28; 95% CI, 0.20-0.38; P = .001). Spontaneous breathing trials were also associated with lower hazards for ventilator-associated events (HR, 0.55; 95% CI, 0.40-0.76; P < .001). Associations with less time to extubation were found for head-of-bed elevation (HR, 1.38, 95% CI, 1.14-1.68; P = .001) and thromboembolism prophylaxis (HR, 2.57; 95% CI, 1.80-3.66; P < .001) but not ventilator mortality. Oral care with chlorhexidine was associated with an increased risk for ventilator mortality (HR, 1.63; 95% CI, 1.15-2.31; P = .006), and stress ulcer prophylaxis was associated with an increased risk for ventilator-associated pneumonia (HR, 7.69; 95% CI, 1.44-41.10; P = .02). CONCLUSIONS AND RELEVANCE: Standard ventilator bundle components vary in their associations with patient-centered outcomes. Head-of-bed elevation, sedative infusion interruptions, spontaneous breathing trials, and thromboembolism prophylaxis appear beneficial, whereas daily oral care with chlorhexidine and stress ulcer prophylaxis may be harmful in some patients.

A low-cost mullite-titania composite ceramic hollow fiber microfiltration membrane for highly efficient separation of oil-in-water emulsion.

Oil-in-water (O/W) emulsion is considered to be difficult to treat. In this work, a low-cost multi-layer-structured mullite-titania composite ceramic hollow fiber microfiltration membrane was fabricated and utilized to efficiently remove fine oil droplets from (O/W) emulsion. In order to reduce membrane cost, coal fly ash was effectively recycled for the first time to fabricate mullite hollow fiber with finger-like and sponge-like structures, on which a much more hydrophilic TiO2 layer was further deposited. The morphology, crystalline phase, mechanical and surface properties were characterized in details. The filtration capability of the final composite membrane was assessed by the separation of a 200 mg·L(-1) synthetic (O/W) emulsion. Even with this microfiltration membrane, a TOC removal efficiency of 97% was achieved. Dilute NaOH solution backwashing was used to effectively accomplish membrane regeneration (∼96% flux recovery efficiency). This study is expected to guide an effective way to recycle waste coal fly ash not only to solve its environmental problems but also to produce a high-valued mullite hollow fiber membrane for highly efficient separation application of O/W emulsion with potential simultaneous functions of pure water production and oil resource recovery.

Highly luminescent and biocompatible near-infrared core-shell CdSeTe/CdS/C quantum dots for probe labeling tumor cells.

In this study, double shelled NIR CdSeTe/CdS/C quantum dots (QDs) were synthesized by a liquid phase method. The as-prepared QDs showed low cytotoxicity and good biocompatibility due to the formation of carbon shell. The imaging of targeted Human cervical carcinoma cells (HeLa cells) indicates that the CdSeTe/CdS/C QDs have excellent optical properties and cell viability. These results clearly shows that the CdSeTe/CdS/C QDs can be a good candidate for bioapplications.

Reversible pH-responsive fluorescence of water-soluble polyfluorenes and their application in metal ion detection.

A novel water-soluble conjugated polymer poly{(4,4'-azobenzene)-2,7-[9,9-bis(6'-N,N,N,-trimethylammonium)hexyl fluorene]dibromide} (PFAB) has been designed and synthesized via Suzuki cross-coupling the fluorene units and azobenzene units. Through simple photoreduction, the azo group of the nonfluorescent PFAB to hydrazine group using UV light, polyfluorene PFAB-L with turn-on fluorescence in aqueous solution is obtained. The optical measurements illustrate that the generation of the flexible hydrazine group induces face-to-face arrangement of phenyl-fluorene-phenyl moieties. Therefore, the excimer formation of phenyl-fluorene-phenyl moieties was induced in PFAB-L. And the fluorescence of PFAB-L can be controlled through modulating the protonation of the -NH-NH- group in solution with different pH. The pH-responsive property is reversible. Moreover, the Fe(3+) ions can selectively quench the fluorescence of the PFAB-L. This new polymer PFAB-L could be used for selective and sensitive sensing Fe(3+) ions in aqueous solution.

Penetration enhancement of lidocaine hydrochlorid by a novel chitosan coated elastic liposome for transdermal drug delivery.

An effective transdermal delivery system for local anaesthetic was developed with lidocaine hydrochloride (LID) as model drug. Chitosan coated elastic liposome (CCEL) were proposed and its in vitro/in vivo skin permeation properties were evaluated. Elastic liposome composed of soya lecithin with sodium deoxycholate (SDC) as edge activator, was prepared by rotary evaporation-sonication method. Chitosan (CH) (0.1-0.5%, w/v) coated elastic liposome by electrostatic attraction of negative elastic liposome and positive CH. CH coating changed the elastic liposome surface charge and increased the vesicle size. The drug encapsulation efficiency (EE) decreased with the increase of CH content. CH coated elastic liposome demonstrated an improved physicochemical stability at 4 degrees C in a 3 months storage period. After coated, CCEL displayed a prolonged drug release profile in vitro release study. The in vitro/in vivo studies showed that CCEL were able to give a statistically significant improvement of skin permeation of LID and significantly reduced pain in comparison with elastic liposome and CH solution.

Enhanced electrochemiluminescence of CdSe quantum dots composited with CNTs and PDDA for sensitive immunoassay.

Electrochemiluminescence (ECL) of CdSe quantum dots (QDs) was greatly enhanced by the combination of carbon nanotubes (CNTs) and poly (diallyldimethylammonium chloride) (PDDA) in the CdSe QDs film, and could successfully be used to develop a sensitive ECL immunosensor for the detection of human IgG (Ag). The novel CdSe QDs-CNTs composites exhibited high ECL intensity, good biocompatibility, and high stability, which held great promise for the fabrication of the ECL biosensors with improved sensitivity. After PDDA as a binding linker was conjugated to the CdSe QDs-CNTs composite film on the electrode, the ECL signal was significantly enhanced. Subsequently, gold nanoparticles (GNPs) assembled onto the CdSe QDs-CNTs/PDDA modified electrode could amplify the ECL signal once again. After antibody (Ab) was immobilized onto the electrode through GNPs, the ECL immunosensor was successfully fabricated. It is for the first time that the unique function of PDDA for enhancing QDs ECL was explored and used to develop an ECL biosensor. The principle of ECL detection for target Ag is based on the increment of steric hindrance after immunoreaction, which resulted in the decrease of ECL intensity. The Ag concentration was determined in the linear range of 0.002-500 ng L(-1) with a detection limit of 0.6 pg mL(-1). The sensor showed good fabrication and detection reproducibility, and the assay results were in acceptable agreement with the clinical sera tests, showing a promising clinical application. This work opened the new avenues for applying QDs ECL in highly sensitive bioassays.

Autographa californica multiple nucleopolyhedrovirus ac53 plays a role in nucleocapsid assembly.

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) orf53 (ac53) is a highly conserved gene existing in all sequenced Lepidoptera and Hymenoptera baculoviruses, but its function remains unknown. To investigate its role in the baculovirus life cycle, an ac53 deletion virus (vAc(ac53KO-PH-GFP)) was generated through homologous recombination in Escherichia coli. Fluorescence and light microscopy and titration analysis revealed that vAc(ac53KO-PH-GFP) could not produce infectious budded virus in infected Sf9 cells. Real-time PCR demonstrated that the ac53 deletion did not affect the levels of viral DNA replication. Electron microscopy showed that many lucent tubular shells devoid of the nucleoprotein core are present in the virogenic stroma and ring zone, indicating that the ac53 knockout affected nucleocapsid assembly. With a recombinant virus expressing an Ac53-GFP fusion protein, we observed that Ac53 was distributed within the cytoplasm and nucleus at 24 h post-infection, but afterwards accumulated predominantly near the nucleus-cytoplasm boundary. These data demonstrate that ac53 is involved in nucleocapsid assembly and is an essential gene for virus production.