Murray Valley encephalitis virus (MVEV) is a mosquito-borne flavivirus known for causing severe neurological diseases in humans. Despite the rising number of reported infections and high mortality rate among hospitalized patients, no antiviral therapies or licensed vaccines are available. To strengthen preparedness against this reemerging virus, we establish a subgenomic replicon (SGR) platform and a complementary single-round infectious particles (SRIPs) production system, using widely circulating genotype 1 (G1) MVEV as backbone. Stem-loop I(SLI) from 3’UTR stands for the major difference among 4 MVEV genotypes and removal of SLI resulted in mild decrease of genome replication. Through screening a mini anti-flavivirus drug library, we identified that asunaprevir (ASV) and ribavirin (RBV) inhibit MVEV infection independently. Combination of ASV and RBV also showed synergistic activity against MVEV. These results underscore the value of the MVEV replicon system as a versatile tool for evaluating antiviral compounds, supporting the potential of ASV and RBV as a combinatorial therapeutic approach.

3D printing is emerging as a transformative approach in functional materials fabrication, enabling precise control over structural parameters, offering opportunities to custom-design permeability, selectivity, and fouling resistance of membrane materials. The ability to fabricate membranes with near-isoporous pore size distribution further enhances their potential for advanced separation applications. The development of formulation and engineering solutions to support the formation of nanoporous nanocomposites with extremely accurate control over the nano-additives distribution is demonstrated in this study with the incorporation of Zinc phthalocyanine (ZnPc), a visible-light-responsive photosensitizer, to offer reactive oxygen species (ROS)-mediated photodynamic inactivation. This study introduces the development of 3D-printed microfiltration membranes, integrating engineered pore structures with photodynamically active surfaces to enhance filtration and antimicrobial performance. Morphological characterization revealed a structural evolution from globular to sheet-like and rod-like formations, significantly influencing pore size, wettability, and surface charge. Photodynamic assessments validated efficient ROS generation, enhancing methylene blue degradation and E. coli inactivation under irradiation. Filtration trials confirmed ZnPc-enhanced bacterial rejection and biofouling resistance, with the 2 wt% ZnPc membrane achieving 99.5% E. coli rejection under irradiation. Furthermore, virus filtration experiments confirmed the efficacy of 1 wt% ZnPc membrane, achieving a 2.56-log, or 99.72%, reduction in Influenza A virus (IAV) recovery. These findings demonstrate that 3D-printed ZnPc-functionalized membranes offer a dual-function approach, combining precise structural control with photodynamic antimicrobial activity, making them promising candidates for next-generation, light-assisted water treatment systems.

Introduction:Noroviruses are small non-enveloped, single stranded positive-sense RNA viruses that belong to the family Caliciviridae. They are highly contagious and resistant to multiple detergents and are the infectious agents in the majority of viral gastroenteritis in adults. Due to a lack of approved preventive or curative therapy options, intensive research effort is ongoing to better understand the pathogenesis mechanisms of noroviruses. Methods:In this study, using the persistent murine norovirus S99 strain (MNoV_S99), we have investigated the role in the regulation of anti-noroviral responses of ubiquitination, a post-translational modification that covalently adds one or multiple ubiquitin molecules onto lysine residues of target proteins. To that end, we have first generated RAW264.7 cells overexpressing YFP-Ubiquitin_WT, _K29R, _K48R or_K63R constructs. All non-WT constructs encode a ubiquitin fusion protein with one lysine mutated into an arginine residue, thus preventing the formation of the K29-, K48- or K63-dependent polyubiquitin chains respectively. Results:Upon infection of these cells with MNoV_S99, we unexpectedly observed that only cells expressing the YFP-Ubiquitin_K48R protein showed a significantly impaired expression of several viral markers: NS5, NS7, VP1 and the replication intermediate dsRNA. Consequently, the number of viral genome copies or viral titers were also significantly decreased in the YFP-Ubiquitin_K48R cells compared to the YFP-Ubiquitin_WT cells. This negative regulation cannot be explained by perturbed viral entry, but rather a constitutive hypersecretion of the pro-inflammatory cytokine TNF and downstream upregulation of IκBα phosphorylation and the subsequent NF-κB nuclear translocation. Conclusion:Overall, these consequences combined impose a non-permissive environment for MNoV_S99 replication and propagation.

Billions of cells undergo apoptosis, a non-inflammatory form of programmed cell death, daily as part of normal development and homeostasis. Apoptotic cells undergo apoptotic cell disassembly to release large extracellular vesicles (EVs) called apoptotic bodies (ApoBDs) to promote dead cell clearance, or otherwise proceed to an inflammatory, lytic outcome (i.e., secondary necrosis). The latter event is regulated by ninjurin-1 (NINJ1), a key executioner of plasma membrane rupture (PMR) through its oligomerisation. However, the precise role of NINJ1 at the intersection of apoptotic cell disassembly and secondary necrosis remain elusive. Here, we show that NINJ1 increasingly oligomerises upon the completion of apoptotic cell disassembly process and that higher-order NINJ1 oligomerisation occurs on ApoBDs. We also demonstrate that NINJ1 regulates PMR of ApoBDs and the release of inflammatory signals and, in part, norovirus particles. Together, our findings provide new insights into NINJ1-mediated PMR and content release-associated functions of ApoBDs.

We previously showed that Polyphenol rich sugarcane extract (PRSE) displayed significant inhibitory effect against influenza A virus (IAV). In this study, we investigated the mechanism of action (MOA) of PRSE against respiratory viruses in human-derived cells. We showed that PRSE treatment does not promote an antiviral state via expression of interferon stimulated genes (ISGs). We subsequently investigated any potential perturbation on the viral entry process and observed that PRSE treatment did not affect caveolin-mediated endocytosis but led to a significant attenuation in clathrin-mediated endocytosis. We confirmed this inhibitory effect on IAV entry, as infection was unaffected by PRSE when IAV fusion was induced at the plasma membrane, instead of endosomal membranes. Based on these findings we observed significant inhibitory effect of PRSE against respiratory syncytial virus and human metapneumovirus, which utilise clathrin-mediated endocytosis, but not human parainfluenza virus type 3, which fuses at the plasma membrane. In conclusion, we show that PRSE has broad antiviral activity and potentially perturbs virus entry via clathrin-mediated endocytosis to inhibit viral replication in vitro.

Inflammasomes are signaling complexes that coordinate inflammation by inducing rapid cytokine secretion and that protect against invading pathogens by initiating death of infected cells. Yet, current methods do not allow us to specifically monitor inflammasome activation in real-time at endogenous protein levels. To overcome this shortcoming, we developed and characterized a novel fluorescent reporter that visualizes inflammasome assembly and reports on the recruitment of the effector protein caspase-1 without impairing downstream signaling. The reporter permits the analysis of inflammasome assembly over time as well as precise quantification of cells with assembled inflammasomes. We have successfully applied the reporter in lentivirus-transduced human and murine cell lines and primary cells but have also applied recombinant viruses that encode the reporter in their genome to detect inflammasome responses in primary cells. Mouse intestinal enteroids expressing the reporter allowed us to visualize how Salmonella infection triggers NAIP/NLRC4 inflammasome assembly, cell death, and expulsion in complex tissues. Lastly, we apply the new tool to gain mechanistic insights and prove that caspase-1 activation on inflammasomes relies on the assembly of filaments of caspase-1CARD, which can be terminated by the CARD-only protein CARD17 to shut down cytokine secretion. We anticipate broad application of the reporter in fundamental and applied research, as it permits the quantitative assessment of inflammasome assembly at both high temporal and spatial resolution or in high throughput. ### Competing Interest Statement F.I. Schmidt is cofounder and consultant of Odyssey Therapeutics. The other authors declare no competing interests.

BACKGROUND:Norovirus causes an estimated 699 million cases of gastroenteritis and 219,000 deaths each year. Historically, novel strains with a genogroup II genotype 4 (GII.4) capsid have emerged every 3-5 years to cause gastroenteritis pandemics. Contrary to historical trends, viruses with aGII.4 Sydney 2012 capsid have extended the timeframe of capsid circulation, well beyond the usual 3-5 years, through genetic recombination to obtain new non-structural regions, for example, a GII.P16 ORF1. OBJECTIVES AND METHODS:The molecular evolution in the GII.4 capsid of strains in New South Wales (NSW), Australia and New Zealand (NZ) before and into the COVID-19 pandemic (2018-20) was investigated by sequencing noroviruses from clinical specimens and wastewater. RESULTS:A continued high prevalence of GII.4 Sydney 2012 [P16] was observed (NSW: 23.0%; NZ: 24.2%), albeit co-dominant with GII.2 [P16] (NSW: 20.2%; NZ: 29.4%). Unlike the historical trends, the GII.4 Sydney 2012 capsid has been in circulation for eight years. Circulating norovirus in the community was disrupted by COVID-19 control measures; lockdowns reduced viral concentration in wastewater by >90% (1.4 × 105 genome copies (gc)/L) from May to September 2020 compared to equivalent timeframes in 2018 (1.6 × 106gc/L) and 2019 (1.9 × 106gc/L). The relaxation of lockdown measures in late-2020 coincided with a strong resurgence of GII.2[P16] prevalence both clinically and in wastewater in NSW and Melbourne, accompanied by a decline in the diversity of circulating noroviruses. Conclusion: In summary, COVID-19 disrupted the strain diversity and levels of norovirus in Australia and New Zealand.

The Australasian Virology Society (AVS) holds premier biennial virology meetings that foster multidisciplinary research and collaboration and promote equity and inclusion of early-career researchers. The 12th AVS meeting (AVS12), convened by M. Tate, J. Fraser, and G. Moseley, was held from 2 to 5 December 2024 on Dja Dja Wurrung country at the RACV Goldfields Resort in Creswick, Victoria, Australia. In this report, we give a brief overview of the history of AVS and outline the current and developing priorities for the society. We provide a summary of the insightful panel discussions held to address career development and Indigenous virology, highlight the presentations given by international plenary speakers Joe Grove and Chantal Abergel, and celebrate the recipients of the numerous awards.

Respiratory infections cause significant morbidity and mortality, yet it is unclear why some individuals succumb to severe disease. In patients hospitalized with avian A(H7N9) influenza, we investigated early drivers underpinning fatal disease. Transcriptomics strongly linked oleoyl-acyl-carrier-protein (ACP) hydrolase (OLAH), an enzyme mediating fatty acid production, with fatal A(H7N9) early after hospital admission, persisting until death. Recovered patients had low OLAH expression throughout hospitalization. High OLAH levels were also detected in patients hospitalized with life-threatening seasonal influenza, COVID-19, respiratory syncytial virus (RSV), and multisystem inflammatory syndrome in children (MIS-C) but not during mild disease. In olah-/-- /- mice, lethal influenza infection led to survival and mild disease as well as reduced lung viral loads, tissue damage, infection-driven pulmonary cell infiltration, and inflammation. This was underpinned by differential lipid droplet dynamics as well as reduced viral replication and virus-induced inflammation in macrophages. Supplementation of oleic acid, the main product of OLAH, increased influenza replication in macrophages and their inflammatory potential. Our findings define how the expression of OLAH drives life-threatening viral disease.

Human norovirus is the leading cause of acute gastroenteritis worldwide, however despite the significance of this pathogen, we have a limited understanding of how noroviruses cause disease, and modulate the innate immune response. Programmed cell death (PCD) is an important part of the innate response to invading pathogens, but little is known about how specific PCD pathways contribute to norovirus replication. Here, we reveal that murine norovirus (MNV) virus-induced PCD in macrophages correlates with the release of infectious virus. We subsequently show, genetically and chemically, that MNV-induced cell death and viral replication occurs independent of the activity of inflammatory mediators. Further analysis revealed that MNV infection promotes the cleavage of apoptotic caspase-3 and PARP. Correspondingly, pan-caspase inhibition, or BAX and BAK deficiency, perturbed viral replication rates and delayed virus release and cell death. These results provide new insights into how MNV harnesses cell death to increase viral burden.

Norovirus infection is characterised by a rapid onset of disease and the development of debilitating symptoms including projectile vomiting and diffuse diarrhoea. Vaccines and antivirals are sorely lacking and developments in these areas are hampered by the lack of an adequate cell culture system to investigate human norovirus replication and pathogenesis. Herein, we describe how the model norovirus, Mouse norovirus (MNV), produces a viral protein, NS3, with the functional capacity to attenuate host protein translation which invokes the activation of cell death via apoptosis. We show that this function of NS3 is conserved between human and mouse viruses and map the protein domain attributable to this function. Our study highlights a critical viral protein that mediates crucial activities during replication, potentially identifying NS3 as a worthy target for antiviral drug development.

Norovirus infections are a leading cause of gastroenteritis worldwide. Despite the substantial global health burden and economic impact, there are currently no approved antiviral therapeutics or vaccines. Additionally, much of our knowledge of norovirus comes from experiments using surrogate viruses, such as murine norovirus and feline calicivirus. The challenge surrounding human norovirus research arises from a lack of robust cell culture systems and efficient animal models. In this review, we explore recent advances in the in vitro cultivation of human norovirus and reverse genetics systems and discuss commonly used in vivo models. We summarize the current understanding of both innate and adaptive immune responses to norovirus infection and provide an overview of vaccine strategies and the current clinical trial landscape, with a focus on the only vaccine candidate that has reached phase III clinical development stage.

Existing mRNA COVID-19 vaccines have shown efficacy in reducing severe cases and fatalities. However, their effectiveness against infection caused by emerging SARS-CoV-2 variants has waned considerably, necessitating the development of variant vaccines. Ideally, next-generation vaccines will be capable of eliciting broader and more sustained immune responses to effectively counteract new variants. Additionally, in vitro assays that more closely represent virus neutralization in humans would greatly assist in the analysis of protective vaccine-induced antibody responses. Here, we present findings from a SARS-CoV-2 VLP vaccine encompassing three key structural proteins: Spike (S), Envelope (E), and Membrane (M). The VLP vaccine effectively produced neutralizing antibodies as determined by surrogate virus neutralization test, and induced virus-specific T-cell responses: predominantly CD4+, although CD8+ T cell responses were detected. T cell responses were more prominent with vaccine delivered with AddaVax compared to vaccine alone. The adjuvanted vaccine was completely protective against live virus challenge in mice. Furthermore, we utilized air–liquid-interface (ALI)-differentiated human nasal epithelium (HNE) as an in vitro system, which authentically models human SARS-CoV-2 infection and neutralization. We show that immune sera from VLP-vaccinated mice completely neutralized SARS-CoV-2 virus infection, demonstrating the potential of ALI-HNE to assess vaccine induced Nab.

The European Union (EU) regulations mandate 10% of all food packaging to be reusable by 2030. United States (U.S.) exporters of specialty crops face new challenges in ensuring microbiological food safety using reusable packaging. A novel antimicrobial formulation consisting of ammonium carboxylate salt of capric acid and L-arginine (GS-2) was recently developed as a spray coating chemical for food packaging materials. In this study, we evaluated the antimicrobial efficacy of GS-2 against microbial strains representing three foodborne bacterial pathogens (Escherichia coli O157:H7, Listeria monocytogenes, Salmonella enterica), one fungal spoilage organism (Aspergillus niger), and one surrogate viral pathogen (murine norovirus) on three reusable plastic materials (acrylonitrile butadiene styrene, high-density polyethylene, and polypropylene) and one cardboard packaging material, respectively. Different chemical concentrations, exposure times, and storage conditions were individually evaluated for the relative antimicrobial efficacies of GS-2 against these microorganisms. Our results showed that GS-2 was highly effective for inactivating bacterial pathogens on both plastic and cardboard surfaces. For instance, 3% GS-2 achieved a >5 log CFU/in2 reduction in E. coli O157:H7, L. monocytogenes, and S. enterica on tested plastic surfaces at an exposure time of 60 min. However, its efficacy against A. niger and murine norovirus was less optimal, resulting in a ≤1 log CFU/in2 reduction on all tested surfaces. Based on our study, GS-2 demonstrated a strong potential as an antibacterial coating reagent for reusable food packaging materials to minimize pathogen contamination and ensure the safety of the specialty crops.

Influenza A virus (IAV) is one of the major global public health concerns but the emerging resistance of IAV to currently available antivirals requires the need to identify potential alternatives. Polyphenol rich sugarcane extract (PRSE) is an extract prepared from the sugarcane plant Saccharum Officinarum. Herein we aimed to determine if PRSE had antiviral activity against IAV. We showed that treatment of IAV-infected cells with PRSE results in a dose-dependent inhibition of virus infection at concentrations that were non-cytotoxic. PRSE treatment limited the early stages of infection, reducing viral genome replication, mRNA transcription and viral protein expression. PRSE did not affect the ability of IAV to bind sialic acid or change the morphology of viral particles. Additionally, PRSE treatment attenuated the replication of multiple IAV strains of the H3N2 and H1N1 subtype. In conclusion, we show that PRSE displays antiviral activity against a broad range of IAV strains, in vitro.

Countermeasures against Zika virus (ZIKV) epidemics are urgently needed. In this study we generated a ZIKV virus-like particle (VLP) based vaccine candidate and assessed the immunogenicity of these particles in mice. The ZIKV-VLPs were morphologically similar to ZIKV by electron microscopy and were recognized by anti-Flavivirus neutralising antibodies. We observed that a single dose of unadjuvanted ZIKV-VLPs, or inactivated ZIKV, generated an immune response that lasted over 6 months, but did not neutralize ZIKV infection of cells in vitro. However, when we co-administered the ZIKV VLPs with either Aluminium hydroxide (Alhydrogel®; Alum), AddaVax or Pam2Cys we observed that Alum was the most effective in a single dose regime, since it not only produced antibodies that neutralized the virus, but also generated a greater number of antigen-specific memory B cells. We additionally observed that the generation of the neutralising antibodies persisted for up to 6 months. Our results suggest that a single dose ZIKV VLPs could be a suitable single dose vaccine candidate for use in outbreak settings.

Murine norovirus (MNV) is a positive-sense, plus-stranded RNA virus in the Caliciviridae family. Viruses in this family replicate in the intestine and are transmitted by the fecal-oral route. MNV is related to the human noroviruses, which cause the majority of nonbacterial gastroenteritis worldwide. Given the technical challenges in studying human norovirus, MNV is often used to study mechanisms in norovirus biology since it combines the availability of a cell culture and reverse genetics system with the ability to study infection in the native host. Adding to our previous protocol collection, here we describe additional techniques that have since been developed to study MNV biology. (c) 2023 The Authors. Current Protocols published by Wiley Periodicals LLC.Basic Protocol 1: Indirect method for measuring cell cytotoxicity and antiviral activityBasic Protocol 2: Measuring murine norovirus genome titers by RT-qPCRSupport Protocol 1: Preparation of standardBasic Protocol 3: Generation of recombinant murine norovirus with minimal passagingBasic Protocol 4: Generation of recombinant murine norovirus via circular polymerase extension reaction (CPER)Basic Protocol 5: Expression of norovirus NS1-2 in insect cell suspension cultures using a recombinant baculovirusSupport Protocol 2: Isotope labelling of norovirus NS1-2 in insect cellsSupport Protocol 3: Purification of the norovirus NS1-2 proteinSupport Protocol 4: Expression of norovirus NS1-2 in mammalian cells by transduction with a recombinant baculovirusBasic Protocol 6: Infection of enteroids in transwell inserts with murine norovirusSupport Protocol 5: Preparation of conditioned medium for enteroids cultureSupport Protocol 6: Isolation of crypts for enteroids generationSupport Protocol 7: Enteroid culture passaging and maintenanceBasic Protocol 7: Quantification of murine norovirus-induced diarrhea using neonatal mouse infectionsAlternate Protocol 1: Intragastric inoculation of neonatal miceAlternate Protocol 2: Scoring colon contents

AbstractThe insect endosymbiotic bacteriumWolbachia pipientisis being utilised as a biocontrol tool to reduce the incidence ofAedes aegypti-transmitted viral diseases like dengue. However, the precise mechanisms underpinningWolbachia’s antiviral activity are not well defined. Here we generated a panel ofAe. aegypti-derived cell lines infected with antiviral strainswMel andwAlbB or the non-antiviral strainwPip to understand host cell morphological changes specifically induced by antiviral strains. Antiviral strains were frequently found to be entirely wrapped by the host endoplasmic reticulum (ER) membrane, whilewPip bacteria clustered separately in the host cell cytoplasm. ER-derived lipid droplets (LDs) increased in volume inwMel-andwAlbB-infected cell lines and mosquito tissues compared to cells infected withwPip orWolbachia-free controls. Inhibition of fatty acid synthase (required for triacylglycerol biosynthesis) reduced LD formation and significantly restored ER-associated dengue virus replication in cells occupied bywMel. Together, this suggests that antiviralWolbachiastrains may specifically alter the lipid composition of the ER to preclude the establishment of DENV replication complexes. DefiningWolbachia’s antiviral mechanisms will support the application and longevity of this effective biocontrol tool that is already being used at scale.ImportanceAedes aegyptitransmits a range of important human pathogenic viruses like dengue. However, infection ofAe. aegyptiwith the insect endosymbiotic bacterium,Wolbachia, reduces the risk of mosquito to human viral transmission.Wolbachiais being utilized at field sites across more than 13 countries to reduce the incidence of viruses like dengue, but it is not well understood howWolbachiainduces its antiviral effects. To examine this at the subcellular level, we compared how different strains ofWolbachiawith varying antiviral strengths, associate with and modify host cell structures. Strongly antiviral strains were found to specifically associate with the host endoplasmic reticulum and induce striking impacts on host cell lipid distribution. InhibitingWolbachia-induced lipid redistribution partially restored dengue virus replication demonstrating this is a contributing role forWolbachia’s antiviral activity. These findings provide new insights into how antiviralWolbachiastrains associate with and modifyAe. aegyptihost cells.

The insect endosymbiotic bacterium Wolbachia pipientis is being utilised as a biocontrol tool to reduce the incidence of Aedes aegypti -transmitted viral diseases like dengue. However, the precise mechanisms underpinning Wolbachia ’s antiviral activity are not well defined. Here we generated a panel of Ae. aegypti -derived cell lines infected with antiviral strains w Mel and w AlbB or the non-antiviral strain w Pip to understand host cell morphological changes specifically induced by antiviral strains. Antiviral strains were frequently found to be entirely wrapped by the host endoplasmic reticulum (ER) membrane, while w Pip bacteria clustered separately in the host cell cytoplasm. ER-derived lipid droplets (LDs) increased in volume in w Mel-and w AlbB-infected cell lines and mosquito tissues compared to cells infected with w Pip or Wolbachia -free controls. Inhibition of fatty acid synthase (required for triacylglycerol biosynthesis) reduced LD formation and significantly restored ER-associated dengue virus replication in cells occupied by w Mel. Together, this suggests that antiviral Wolbachia strains may specifically alter the lipid composition of the ER to preclude the establishment of DENV replication complexes. Defining Wolbachia ’s antiviral mechanisms will support the application and longevity of this effective biocontrol tool that is already being used at scale.Importance Aedes aegypti transmits a range of important human pathogenic viruses like dengue. However, infection of Ae. aegypti with the insect endosymbiotic bacterium, Wolbachia , reduces the risk of mosquito to human viral transmission. Wolbachia is being utilized at field sites across more than 13 countries to reduce the incidence of viruses like dengue, but it is not well understood how Wolbachia induces its antiviral effects. To examine this at the subcellular level, we compared how different strains of Wolbachia with varying antiviral strengths, associate with and modify host cell structures. Strongly antiviral strains were found to specifically associate with the host endoplasmic reticulum and induce striking impacts on host cell lipid distribution. Inhibiting Wolbachia -induced lipid redistribution partially restored dengue virus replication demonstrating this is a contributing role for Wolbachia ’s antiviral activity. These findings provide new insights into how antiviral Wolbachia strains associate with and modify Ae. aegypti host cells.### Competing Interest StatementThe authors have declared no competing interest.

Virus infections of the central nervous system (CNS) cause important diseases of humans and animals. As in other tissues, innate antiviral responses mediated by type I interferons (IFNs) are crucially important in controlling CNS virus infections. The maturity of neuronal populations is an established critical factor determining the outcome of CNS virus infection. Using primary cultures of mouse cortical neurons, we investigated the relationships between neuronal maturation, type I IFN responses, and the outcome of Semliki Forest virus infection. The virus replicated better, infected more cells, and produced higher titres of infectious viruses in immature neurons. Complete transcriptome analysis demonstrated that resting immature neurons have low transcriptional competence to mount antiviral responses. They had no detectable transcription of the genes Ddx58 and Ifih1, which encode key RNA virus cytoplasmic sensors RIG-I and MDA5, and very low expression of genes encoding key regulators of associated signalling pathways. Upon infection, immature neurons failed to mount an antiviral response as evidenced by their failure to produce chemokines, IFNs, and other cytokines. Treatment of immature neurons with exogenous IFNβ prior to infection resulted in antiviral responses and lower levels of virus replication and infectious virus production. In contrast, resting mature neurons generated a robust antiviral response. This was augmented by pretreatment with IFNβ. Infection of mature neurons derived from IFNAR−/− mice did not make an antiviral response and replicated virus to high levels.