Antiviral Research: At age 41, a transition.

This article marks the transition from Mike Bray to Subhash Vasudevan as editor-in-chief of Antiviral Research, in the journal's 41st year of publication. It reviews AVR's experience since 2011, when the founder and first editor-in-chief, Erik De Clercq, wrote a paper describing the journal's first 30 years. Since that time, the editorial team has doubled in size, with editors now located in 8 countries; half are women. There has been a corresponding increase in the number of published papers, covering research on antiviral drugs, vaccines and pathogenesis for a wide range of endemic and epidemic viral diseases of humans and livestock animals, and there has been a significant rise in the journal's impact factor. AVR's experience during the COVID-19 pandemic is also briefly summarized.

Meeting report: 32nd International Conference on Antiviral Research.

The 32nd International Conference on Antiviral Research (ICAR), sponsored by the International Society for Antiviral Research (ISAR), was held in Baltimore, Maryland, USA, on May 12-15, 2019. This report gives an overview of the conference on behalf of the Society. It provides a general review of the meeting and awardees, summarizing the presentations, and their main conclusions from the perspective of researchers active in many different areas of antiviral research and development. As in past years, ICAR promoted and showcased the most recent progress in antiviral research, and continued to foster collaborations and interactions in drug discovery and development. The 33rd ICAR will be held in Seattle, Washington, USA, March 30th-April 3rd, 2020.

Meeting report: 31st International Conference on Antiviral Research.

The 31st International Conference on Antiviral Research (ICAR) was held in Porto, Portugal from June 11-15, 2018. In this report, volunteer rapporteurs provide their summaries of scientific presentations, hoping to effectively convey the speakers' goals and the results and conclusions of their talks. This report provides an overview of the invited keynote and award lectures and highlights of short oral presentations, from the perspective of experts in antiviral research. Of note, a session on human cytomegalovirus included an update on the introduction to the clinic of letermovir for the prevention of CMV infection and disease. The 31st ICAR successfully promoted new discoveries in antiviral research and drug development. The 32nd ICAR will be held in Baltimore, Maryland, USA, May 6-10, 2019.

Second International Conference on Crimean-Congo Hemorrhagic Fever.

The Second International Conference on Crimean-Congo Hemorrhagic Fever (CCHF) was held in Thessaloniki, Greece, from September 10-13, 2017, and brought together international public health professionals, clinicians, ecologists, and basic laboratory researchers. Nearly 100 participants, representing 24 countries and the World Health Organization (WHO), were in attendance. Meeting sessions covered the epidemiology of CCHF in humans; ticks and virus-tick interactions; wild and domestic animal hosts; molecular virology; taxonomic classification; pathogenesis and animal models; clinical aspects and diagnosis; clinical management and clinical trials; and disease prevention in humans. The concluding session focused on recent WHO recommendations for public health measures and future research. This report summarizes lectures by the invited speakers and highlights advances in the field.

Highlights of the 30th International Conference on Antiviral Research.

The 30th International Conference on Antiviral Research (ICAR) was held in Atlanta, GA, USA from May 18 to 21, 2017. This report provides an account of award lectures, invited keynote addresses and oral presentations during the meeting. The 2017 Gertrude Elion Memorial Lecture Award by Michael Sofia highlighted one of the most important accomplishments in recent drug discovery in antiviral research, the identification of the hepatitis C virus direct-acting antiviral sofosbuvir and new alternatives to combat hepatitis B virus (HBV) infection. The Antonín Holý Lecture Award by David Chu on medicinal chemistry provided an overview of early developments of nucleoside analogs for the treatment of HIV and varicella zoster virus infection and how this knowledge serves to develop new drugs targeting HBV. Priscilla Yang gave the first ISAR Women in Science lecture. She reported on pharmacological validation of new antiviral targets for dengue, Zika and other flaviviruses. The William Prusoff Young Investigator Lecture Award by Maaike Everts described the Alabama Drug Discovery Alliance and the Antiviral Drug Discovery and Development Consortium, and how they are helping to accelerate the development of new antivirals. The 30th ICAR was a success in promoting new discoveries in antiviral drug development and research. The 31st ICAR will be held in Porto, Portugal, June 11-15, 2018.

Small Animal Models for Studying Filovirus Pathogenesis.

Filovirus small animal disease models have so far been developed in laboratory mice, guinea pigs, and hamsters. Since immunocompetent rodents do not exhibit overt signs of disease following infection with wild-type filoviruses isolated from humans, rodent models have been established using adapted viruses produced through sequential passage in rodents. Rodent-adapted viruses target the same cells/tissues as the wild-type viruses, making rodents invaluable basic research tools for studying filovirus pathogenesis. Moreover, comparative analyses using wild-type and rodent-adapted viruses have provided beneficial insights into the molecular mechanisms of pathogenicity and acquisition of species-specific virulence. Additionally, wild-type filovirus infections in immunodeficient rodents have provided a better understanding of the host factors required for resistance to filovirus infection and of the immune response against the infection. This chapter provides comprehensive information on the filovirus rodent models and rodent-adapted filoviruses. Specifically, we summarize the clinical and pathological features of filovirus infections in all rodent models described to date, including the recently developed humanized and collaborative cross (CC) resource recombinant inbred (RI) intercrossed (CC-RIX) mouse models. We also cover the molecular determinants responsible for adaptation and virulence acquisition in a number of rodent-adapted filoviruses. This chapter clearly defines the characteristic and advantages/disadvantages of rodent models, helping to evaluate the practical use of rodent models in future filovirus studies.

A historical perspective on the discovery and elucidation of the hepatitis B virus.

The discovery in 1965 of the "Australia antigen," subsequently identified as the hepatitis B virus surface antigen (HBsAg), was such a watershed event in virology that it is often thought to mark the beginning of hepatitis research, but it is more accurately seen as a critical breakthrough in a long effort to understand the pathogenesis of infectious hepatitis. A century earlier, Virchow provided an authoritative explanation of "catarrhal jaundice," which did not consider an infectious etiology, but the transmission of jaundice by human serum was clearly identified in two outbreaks in 1885, and the distinction between "infectious" and "serum" hepatitis was recognized by the early 1920s. The inability to culture a virus or reproduce either syndrome in laboratory animals led to numerous studies in human volunteers; by the end of World War II, it was known that the diseases were caused by different filterable agents, and the terms "hepatitis A" and "B" were introduced in 1947 (though some long-incubation cases then designated B must in retrospect have been hepatitis C). The development of a number of liver function tests during the 1950s led to the recognition of anicteric infections and the existence of chronic carriers, but little more could be done until an infectious agent had been identified. Once Blumberg and colleagues had found a specific viral marker, the vast amount of accumulated epidemiologic and clinical data, together with huge numbers of stored serum samples, enabled rapid progress in understanding hepatitis B, and revealed the existence of a vast population of chronically infected people in Asia, Oceania and Africa. In this article, we place the identification of the Australia antigen within the historical context of research on viral hepatitis. Following a chronological review from 1865 to 1965, we summarize how the discovery led to improved safety of blood transfusion, the development of a highly effective vaccine and the eventual identification of the hepatitis C, D and E viruses. This article forms part of a symposium in Antiviral Research on "An unfinished story: from the discovery of the Australia antigen to the development of new curative therapies for chronic hepatitis B."

Low-dose ribavirin potentiates the antiviral activity of favipiravir against hemorrhagic fever viruses.

Favipiravir is approved in Japan to treat novel or re-emerging influenza viruses, and is active against a broad spectrum of RNA viruses, including Ebola. Ribavirin is the only other licensed drug with activity against multiple RNA viruses. Recent studies show that ribavirin and favipiravir act synergistically to inhibit bunyavirus infections in cultured cells and laboratory mice, likely due to their different mechanisms of action. Convalescent immune globulin is the only approved treatment for Argentine hemorrhagic fever caused by the rodent-borne Junin arenavirus. We previously reported that favipiravir is highly effective in a number of small animal models of Argentine hemorrhagic fever. We now report that addition of low dose of ribavirin synergistically potentiates the activity of favipiravir against Junin virus infection of guinea pigs and another arenavirus, Pichinde virus infection of hamsters. This suggests that the efficacy of favipiravir against hemorrhagic fever viruses can be further enhanced through the addition of low-dose ribavirin.

The use of nonhuman primates in research on seasonal, pandemic and avian influenza, 1893-2014.

Attempts to reproduce the features of human influenza in laboratory animals date from the early 1890s, when Richard Pfeiffer inoculated apes with bacteria recovered from influenza patients and produced a mild respiratory illness. Numerous studies employing nonhuman primates (NHPs) were performed during the 1918 pandemic and the following decade. Most used bacterial preparations to infect animals, but some sought a filterable agent for the disease. Since the viral etiology of influenza was established in the early 1930s, studies in NHPs have been supplemented by a much larger number of experiments in mice, ferrets and human volunteers. However, the emergence of a novel swine-origin H1N1 influenza virus in 1976 and the highly pathogenic H5N1 avian influenza virus in 1997 stimulated an increase in NHP research, because these agents are difficult to study in naturally infected patients and cannot be administered to human volunteers. In this paper, we review the published literature on the use of NHPs in influenza research from 1893 through the end of 2014. The first section summarizes observational studies of naturally occurring influenza-like syndromes in wild and captive primates, including serologic investigations. The second provides a chronological account of experimental infections of NHPs, beginning with Pfeiffer's study and covering all published research on seasonal and pandemic influenza viruses, including vaccine and antiviral drug testing. The third section reviews experimental infections of NHPs with avian influenza viruses that have caused disease in humans since 1997. The paper concludes with suggestions for further studies to more clearly define and optimize the role of NHPs as experimental animals for influenza research.

Mechanisms of immunity in post-exposure vaccination against Ebola virus infection.

Ebolaviruses can cause severe hemorrhagic fever that is characterized by rapid viral replication, coagulopathy, inflammation, and high lethality rates. Although there is no clinically proven vaccine or treatment for Ebola virus infection, a virus-like particle (VLP) vaccine is effective in mice, guinea pigs, and non-human primates when given pre-infection. In this work, we report that VLPs protect Ebola virus-infected mice when given 24 hours post-infection. Analysis of cytokine expression in serum revealed a decrease in pro-inflammatory cytokine and chemokine levels in mice given VLPs post-exposure compared to infected, untreated mice. Using knockout mice, we show that VLP-mediated post-exposure protection requires perforin, B cells, macrophages, conventional dendritic cells (cDCs), and either CD4+ or CD8+ T cells. Protection was Ebola virus-specific, as marburgvirus VLPs did not protect Ebola virus-infected mice. Increased antibody production in VLP-treated mice correlated with protection, and macrophages were required for this increased production. However, NK cells, IFN-gamma, and TNF-alpha were not required for post-exposure-mediated protection. These data suggest that a non-replicating Ebola virus vaccine can provide post-exposure protection and that the mechanisms of immune protection in this setting require both increased antibody production and generation of cytotoxic T cells.

Virus-like particles activate type I interferon pathways to facilitate post-exposure protection against Ebola virus infection.

Ebola virus (EBOV) causes a severe hemorrhagic disease with high fatality. Virus-like particles (VLPs) are a promising vaccine candidate against EBOV. We recently showed that VLPs protect mice from lethal EBOV infection when given before or after viral infection. To elucidate pathways through which VLPs confer post-exposure protection, we investigated the role of type I interferon (IFN) signaling. We found that VLPs lead to accelerated induction of IFN stimulated genes (ISGs) in liver and spleen of wild type mice, but not in Ifnar-/- mice. Accordingly, EBOV infected Ifnar-/- mice, unlike wild type mice succumbed to death even after VLP treatment. The ISGs induced in wild type mice included anti-viral proteins and negative feedback factors known to restrict viral replication and excessive inflammatory responses. Importantly, proinflammatory cytokine/chemokine expression was much higher in WT mice without VLPs than mice treated with VLPs. In EBOV infected Ifnar-/- mice, however, uninhibited viral replication and elevated proinflammatory factor expression ensued, irrespective of VLP treatment, supporting the view that type I IFN signaling helps to limit viral replication and attenuate inflammatory responses. Further analyses showed that VLP protection requires the transcription factor, IRF8 known to amplify type I IFN signaling in dendritic cells and macrophages, the probable sites of initial EBOV infection. Together, this study indicates that VLPs afford post-exposure protection by promoting expeditious initiation of type I IFN signaling in the host.

Vaccinia virus induces rapid necrosis in keratinocytes by a STAT3-dependent mechanism.

RATIONALE: Humans with a dominant negative mutation in STAT3 are susceptible to severe skin infections, suggesting an essential role for STAT3 signaling in defense against cutaneous pathogens. METHODS: To focus on innate antiviral defenses in keratinocytes, we used a standard model of cutaneous infection of severe combined immunodeficient mice with the current smallpox vaccine, ACAM-2000. In parallel, early events post-infection with the smallpox vaccine ACAM-2000 were investigated in cultured keratinocytes of human and mouse origin. RESULTS: Mice treated topically with a STAT3 inhibitor (Stattic) developed larger vaccinia lesions with higher virus titers and died more rapidly than untreated controls. Cultured human and murine keratinocytes infected with ACAM-2000 underwent rapid necrosis, but when treated with Stattic or with inhibitors of RIP1 kinase or caspase-1, they survived longer, produced higher titers of virus, and showed reduced activation of type I interferon responses and inflammatory cytokines release. Treatment with inhibitors of RIP1 kinase and STAT3, but not caspase-1, also reduced the inflammatory response of keratinocytes to TLR ligands. Vaccinia growth properties in Vero cells, which are known to be defective in some antiviral responses, were unaffected by inhibition of RIP1K, caspase-1, or STAT3. CONCLUSIONS: Our findings indicate that keratinocytes suppress the replication and spread of vaccinia virus by undergoing rapid programmed cell death, in a process requiring STAT3. These data offer a new framework for understanding susceptibility to skin infection in patients with STAT3 mutations. Interventions which promote prompt necroptosis/pyroptosis of infected keratinocytes may reduce risks associated with vaccination with live vaccinia virus.

The cyanobacterial lectin scytovirin displays potent in vitro and in vivo activity against Zaire Ebola virus.

The cyanobacterial lectin scytovirin (SVN) binds with high affinity to mannose-rich oligosaccharides on the envelope glycoprotein (GP) of a number of viruses, blocking entry into target cells. In this study, we assessed the ability of SVN to bind to the envelope GP of Zaire Ebola virus (ZEBOV) and inhibit its replication. SVN interacted specifically with the protein's mucin-rich domain. In cell culture, it inhibited ZEBOV replication with a 50% virus-inhibitory concentration (EC50) of 50 nM, and was also active against the Angola strain of the related Marburg virus (MARV), with a similar EC50. Injected subcutaneously in mice, SVN reached a peak plasma level of 100 nm in 45 min, but was cleared within 4h. When ZEBOV-infected mice were given 30 mg/kg/day of SVN by subcutaneous injection every 6h, beginning the day before virus challenge, 9 of 10 animals survived the infection, while all infected, untreated mice died. When treatment was begun one hour or one day after challenge, 70-90% of mice survived. Quantitation of infectious virus and viral RNA in samples of serum, liver and spleen collected on days 2 and 5 postinfection showed a trend toward lower titers in treated than control mice, with a significant decrease in liver titers on day 2. Our findings provide further evidence of the potential of natural lectins as therapeutic agents for viral infections.

Ebola virus-like particles stimulate type I interferons and proinflammatory cytokine expression through the toll-like receptor and interferon signaling pathways.

Ebola viruses (EBOV) can cause severe hemorrhagic disease with high case fatality rates. Currently, no vaccines or therapeutics are approved for use in humans. Ebola virus-like particles (eVLP) comprising of virus protein (VP40), glycoprotein, and nucleoprotein protect rodents and nonhuman primates from lethal EBOV infection, representing as a candidate vaccine for EBOV infection. Previous reports have shown that eVLP stimulate the expression of proinflammatory cytokines in dendritic cells (DCs) and macrophages (MΦs) in vitro. However, the molecular mechanisms and signaling pathways through which eVLP induce innate immune responses remain obscure. In this study, we show that eVLP stimulate not only the expression of proinflammatory cytokines but also the expression of type I interferons (IFNs) and IFN-stimulated genes (ISGs) in murine bone marrow-derived DCs (BMDCs) and MΦs. Our data indicate that eVLP trigger host responses through toll-like receptor (TLR) pathway utilizing 2 distinct adaptors, MyD88 and TRIF. More interestingly, eVLP activated the IFN signaling pathway by inducing a set of potent antiviral ISGs. Last, eVLP and synthetic adjuvants, Poly I:C and CpG DNA, cooperatively increased the expression of cytokines and ISGs. Further supporting this synergy, eVLP when administered together with Poly I:C conferred mice enhanced protection against EBOV infection. These results indicate that eVLP stimulate early innate immune responses through TLR and type I IFN signaling pathways to protect the host from EBOV infection.

Crimean-Congo hemorrhagic fever: history, epidemiology, pathogenesis, clinical syndrome and genetic diversity.

Crimean-Congo hemorrhagic fever (CCHF) is the most important tick-borne viral disease of humans, causing sporadic cases or outbreaks of severe illness across a huge geographic area, from western China to the Middle East and southeastern Europe and throughout most of Africa. CCHFV is maintained in vertical and horizontal transmission cycles involving ixodid ticks and a variety of wild and domestic vertebrates, which do not show signs of illness. The virus circulates in a number of tick genera, but Hyalomma ticks are the principal source of human infection, probably because both immature and adult forms actively seek hosts for the blood meals required at each stage of maturation. CCHF occurs most frequently among agricultural workers following the bite of an infected tick, and to a lesser extent among slaughterhouse workers exposed to the blood and tissues of infected livestock and medical personnel through contact with the body fluids of infected patients. CCHFV is the most genetically diverse of the arboviruses, with nucleotide sequence differences among isolates ranging from 20% for the viral S segment to 31% for the M segment. Viruses with diverse sequences can be found within the same geographic area, while closely related viruses have been isolated in far distant regions, suggesting that widespread dispersion of CCHFV has occurred at times in the past, possibly by ticks carried on migratory birds or through the international livestock trade. Reassortment among genome segments during co-infection of ticks or vertebrates appears to have played an important role in generating diversity, and represents a potential future source of novel viruses. In this article, we first review current knowledge of CCHFV, summarizing its molecular biology, maintenance and transmission, epidemiology and geographic range. We also include an extensive discussion of CCHFV genetic diversity, including maps of the range of the virus with superimposed phylogenetic trees. We then review the features of CCHF, including the clinical syndrome, diagnosis, treatment, pathogenesis, vaccine development and laboratory animal models of CCHF. The paper ends with a discussion of the possible future geographic range of the virus. For the benefit of researchers, we include a Supplementary Table listing all published reports of CCHF cases and outbreaks in the English-language literature, plus some principal articles in other languages, with total case numbers, case fatality rates and all CCHFV strains on GenBank.

Mouse models for filovirus infections.

The filoviruses marburg- and ebolaviruses can cause severe hemorrhagic fever (HF) in humans and nonhuman primates. Because many cases have occurred in geographical areas lacking a medical research infrastructure, most studies of the pathogenesis of filoviral HF, and all efforts to develop drugs and vaccines, have been carried out in biocontainment laboratories in non-endemic countries, using nonhuman primates (NHPs), guinea pigs and mice as animal models. NHPs appear to closely mirror filoviral HF in humans (based on limited clinical data), but only small numbers may be used in carefully regulated experiments; much research is therefore done in rodents. Because of their availability in large numbers and the existence of a wealth of reagents for biochemical and immunological testing, mice have become the preferred small animal model for filovirus research. Since the first experiments following the initial 1967 marburgvirus outbreak, wild-type or mouse-adapted viruses have been tested in immunocompetent or immunodeficient mice. In this paper, we review how these types of studies have been used to investigate the pathogenesis of filoviral disease, identify immune responses to infection and evaluate antiviral drugs and vaccines. We also discuss the strengths and weaknesses of murine models for filovirus research, and identify important questions for further study.

Eczema vaccinatum.

Eczema vaccinatum (EV) is a complication of smallpox vaccination that can occur in persons with eczema/atopic dermatitis (AD), in which vaccinia virus disseminates to cause an extensive rash and systemic illness. Because persons with eczema are deferred from vaccination, only a single, accidentally transmitted case of EV has been described in the medical literature since military vaccination was resumed in the United States in 2002. To enhance understanding of EV, we review its history during the era of universal vaccination and discuss its relationship to complications in persons with other diseases or injuries of the skin. We then discuss current concepts of the pathophysiology of AD, noting how defective skin barrier function, epidermal hyperplasia, and abnormal immune responses favor the spread of poxviral infection, and identify a number of unanswered questions about EV. We conclude by considering how its occurrence might be minimized in the event of a return to universal vaccination.

Computer-assisted detection of infectious lung diseases: a review.

Respiratory tract infections are a leading cause of death and disability worldwide. Although radiology serves as a primary diagnostic method for assessing respiratory tract infections, visual analysis of chest radiographs and computed tomography (CT) scans is restricted by low specificity for causal infectious organisms and a limited capacity to assess severity and predict patient outcomes. These limitations suggest that computer-assisted detection (CAD) could make a valuable contribution to the management of respiratory tract infections by assisting in the early recognition of pulmonary parenchymal lesions, providing quantitative measures of disease severity and assessing the response to therapy. In this paper, we review the most common radiographic and CT features of respiratory tract infections, discuss the challenges of defining and measuring these disorders with CAD, and propose some strategies to address these challenges.