ICTV Virus Taxonomy Profile: Arenaviridae 2023.

Arenaviridae is a family for ambisense RNA viruses with genomes of about 10.5 kb that infect mammals, snakes, and fish. The arenavirid genome consists of two or three single-stranded RNA segments and encodes a nucleoprotein (NP), a glycoprotein (GP) and a large (L) protein containing RNA-directed RNA polymerase (RdRP) domains; some arenavirids encode a zinc-binding protein (Z). This is a summary of the International Committee on Taxonomy of Viruses (ICTV) report on the family Arenaviridae, which is available at www.ictv.global/report/arenaviridae.

The Key Roles of Interferon Lambda in Human Molecular Defense against Respiratory Viral Infections.

Interferons (IFN) are crucial for the innate immune response. Slightly more than two decades ago, a new type of IFN was discovered: the lambda IFN (type III IFN). Like other IFN, the type III IFN display antiviral activity against a wide variety of infections, they induce expression of antiviral, interferon-stimulated genes (MX1, OAS, IFITM1), and they have immuno-modulatory activities that shape adaptive immune responses. Unlike other IFN, the type III IFN signal through distinct receptors is limited to a few cell types, primarily mucosal epithelial cells. As a consequence of their greater and more durable production in nasal and respiratory tissues, they can determine the outcome of respiratory infections. This review is focused on the role of IFN-λ in the pathogenesis of respiratory viral infections, with influenza as a prime example. The influenza virus is a major public health problem, causing up to half a million lethal infections annually. Moreover, the virus has been the cause of four pandemics over the last century. Although IFN-λ are increasingly being tested in antiviral therapy, they can have a negative influence on epithelial tissue recovery and increase the risk of secondary bacterial infections. Therefore, IFN-λ expression deserves increased scrutiny as a key factor in the host immune response to infection.

Cauliflower Mosaic Virus TAV, a Plant Virus Protein That Functions like Ribonuclease H1 and is Cytotoxic to Glioma Cells.

Recent comparisons between plant and animal viruses reveal many common principles that underlie how all viruses express their genetic material, amplify their genomes, and link virion assembly with replication. Cauliflower mosaic virus (CaMV) is not infectious for human beings. Here, we show that CaMV transactivator/viroplasmin protein (TAV) shares sequence similarity with and behaves like the human ribonuclease H1 (RNase H1) in reducing DNA/RNA hybrids detected with S9.6 antibody in HEK293T cells. We showed that TAV is clearly expressed in the cytosol and in the nuclei of transiently transfected human cells, similar to its distribution in plants. TAV also showed remarkable cytotoxic effects in U251 human glioma cells in vitro. These characteristics pave the way for future analysis on the use of the plant virus protein TAV, as an alternative to human RNAse H1 during gene therapy in human cells.

A Single Dose of Modified Vaccinia Ankara Expressing Lassa Virus-like Particles Protects Mice from Lethal Intra-cerebral Virus Challenge.

Lassa fever surpasses Ebola, Marburg, and all other hemorrhagic fevers except Dengue in its public health impact. Caused by Lassa virus (LASV), the disease is a scourge on populations in endemic areas of West Africa, where reported incidence is higher. Here, we report construction, characterization, and preclinical efficacy of a novel recombinant vaccine candidate GEO-LM01. Constructed in the Modified Vaccinia Ankara (MVA) vector, GEO-LM01 expresses the glycoprotein precursor (GPC) and zinc-binding matrix protein (Z) from the prototype Josiah strain lineage IV. When expressed together, GP and Z form Virus-Like Particles (VLPs) in cell culture. Immunogenicity and efficacy of GEO-LM01 was tested in a mouse challenge model. A single intramuscular dose of GEO-LM01 protected 100% of CBA/J mice challenged with a lethal dose of ML29, a Mopeia/Lassa reassortant virus, delivered directly into the brain. In contrast, all control animals died within one week. The vaccine induced low levels of antibodies but Lassa-specific CD4+ and CD8+ T cell responses. This is the first report showing that a single dose of a replication-deficient MVA vector can confer full protection against a lethal challenge with ML29 virus.

ICTV Virus Taxonomy Profile: Arenaviridae.

Members of the family Arenaviridae produce enveloped virions containing genomes consisting of two or three single-stranded RNA segments totalling about 10.5 kb. Arenaviruses can infect mammals, including humans and other primates, snakes, and fish. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Arenaviridae, which is available at www.ictv.global/report/arenaviridae.

T-Cell Response to Viral Hemorrhagic Fevers.

Viral hemorrhagic fevers (VHF) are a group of clinically similar diseases that can be caused by enveloped RNA viruses primarily from the families Arenaviridae, Filoviridae, Hantaviridae, and Flaviviridae. Clinically, this group of diseases has in common fever, fatigue, dizziness, muscle aches, and other associated symptoms that can progress to vascular leakage, bleeding and multi-organ failure. Most of these viruses are zoonotic causing asymptomatic infections in the primary host, but in human beings, the infection can be lethal. Clinical and experimental evidence suggest that the T-cell response is needed for protection against VHF, but can also cause damage to the host, and play an important role in disease pathogenesis. Here, we present a review of the T-cell immune responses to VHF and insights into the possible ways to improve counter-measures for these viral agents.

Improving the Breadth of the Host's Immune Response to Lassa Virus.

In 2017, the global Coalition for Epidemic Preparedness (CEPI) declared Lassa virus disease to be one of the world's foremost biothreats. In January 2018, World Health Organization experts met to address the Lassa biothreat. It was commonly recognized that the diversity of Lassa virus (LASV) isolated from West African patient samples was far greater than that of the Ebola isolates from the West African epidemic of 2013⁻2016. Thus, vaccines produced against Lassa virus disease face the added challenge that they must be broadly-protective against a wide variety of LASV. In this review, we discuss what is known about the immune response to Lassa infection. We also discuss the approaches used to make broadly-protective influenza vaccines and how they could be applied to developing broad vaccine coverage against LASV disease. Recent advances in AIDS research are also potentially applicable to the design of broadly-protective medical countermeasures against LASV disease.

Diagnostics for Lassa Fever: Detecting Host Antibody Responses.

There are two types of viral diagnostics: (1) those that detect components of the pathogen (like viral RNA or proteins) and (2) those that detect host molecules that rise or fall as a consequence of pathogen infection (like anti-viral antibodies or virus-induced inflammatory cytokines). Quantitative PCR to detect Lassa RNA, and clinical chemistry to detect high liver enzymes (AST/ALT) are commonly used to diagnose Lassa fever. Here, we discuss the various types of diagnostics for Lassa fever and the urgent need for early diagnosis. We also describe a protocol for using the attenuated Lassa vaccine candidate, ML29 , as an antigen for detecting Lassa-specific antibodies. Since antibodies are developed late in the progression of Lassa fever disease, this is not an early diagnostic, but is more useful in surveillance of the population to determine the sero-prevalence of antibodies to Lassa virus (LASV ), and to define treatment options for people in close contact with a Lassa-infected person.

A Primate Model for Viral Hemorrhagic Fever.

Lymphocytic choriomeningitis virus strain WE (LCMV-WE), a Risk Group 3 virus, causes a disease in rhesus monkeys that closely resembles human infection with Lassa fever virus, a Risk Group 4 agent. Three stages of disease progression have been defined and profiled in this model: pre-viremic, viremic, and terminal. The earliest or pre-viremic stage reveals changes in the blood profile predictive of the later stages of disease. In order to identify whether specific changes are pathognomonic, it was necessary to perform a parallel infection with an attenuated virus (LCMV-Armstrong). Here we review the use of nonhuman primates to model viral hemorrhagic fever and offer a step-by-step guide to using a rhesus macaque model for Lassa fever.

Past, present, and future of arenavirus taxonomy.

Until recently, members of the monogeneric family Arenaviridae (arenaviruses) have been known to infect only muroid rodents and, in one case, possibly phyllostomid bats. The paradigm of arenaviruses exclusively infecting small mammals shifted dramatically when several groups independently published the detection and isolation of a divergent group of arenaviruses in captive alethinophidian snakes. Preliminary phylogenetic analyses suggest that these reptilian arenaviruses constitute a sister clade to mammalian arenaviruses. Here, the members of the International Committee on Taxonomy of Viruses (ICTV) Arenaviridae Study Group, together with other experts, outline the taxonomic reorganization of the family Arenaviridae to accommodate reptilian arenaviruses and other recently discovered mammalian arenaviruses and to improve compliance with the Rules of the International Code of Virus Classification and Nomenclature (ICVCN). PAirwise Sequence Comparison (PASC) of arenavirus genomes and NP amino acid pairwise distances support the modification of the present classification. As a result, the current genus Arenavirus is replaced by two genera, Mammarenavirus and Reptarenavirus, which are established to accommodate mammalian and reptilian arenaviruses, respectively, in the same family. The current species landscape among mammalian arenaviruses is upheld, with two new species added for Lunk and Merino Walk viruses and minor corrections to the spelling of some names. The published snake arenaviruses are distributed among three new separate reptarenavirus species. Finally, a non-Latinized binomial species name scheme is adopted for all arenavirus species. In addition, the current virus abbreviations have been evaluated, and some changes are introduced to unequivocally identify each virus in electronic databases, manuscripts, and oral proceedings.

Genomic profiling of host responses to Lassa virus: therapeutic potential from primate to man.

Lassa virus infection elicits distinctive changes in host gene expression and metabolism. We focus on changes in host gene expression that may be biomarkers that discriminate individual pathogens or may help to provide a prognosis for disease. In addition to assessing mRNA changes, functional studies are also needed to discriminate causes of disease from mechanisms of host resistance. Host responses that drive pathogenesis are likely to be targets for prevention or therapy. Host responses to Lassa or its related arenaviruses have been monitored in cell culture, in animal models of hemorrhagic fever, in Lassa-infected nonhuman primates and, to a limited extent, in infected human beings. Here, we describe results from those studies and discuss potential targets for reducing virus replication and mitigating disease.

Modulation of SIV and HIV DNA vaccine immunity by Fas-FasL signaling.

Signaling through the Fas/Apo-1/CD95 death receptor is known to affect virus-specific cell-mediated immune (CMI) responses. We tested whether modulating the Fas-apoptotic pathway can enhance immune responses to DNA vaccination or lymphocytic choriomeningitis virus (LCMV) infection. Mice were electroporated with plasmids expressing a variety of pro- or anti-apoptotic molecules related to Fas signaling and then either LCMV-infected or injected with plasmid DNA expressing SIV or HIV antigens. Whereas Fas or FasL knockout mice had improved CMI, down-regulation of Fas or FasL by shRNA or antibody failed to improve CMI and was accompanied by increases in regulatory T cells (Treg). Two "adjuvant" plasmids were discovered that significantly enhanced plasmid immunizations. The adjuvant effects of Fas-associated death domain (FADD) and of cellular FLICE-inhibitory protein (cFLIP) were consistently accompanied by increased effector memory T lymphocytes and increased T cell proliferation. This adjuvant effect was also observed when comparing murine infections with LCMV-Armstrong and its persisting variant LCMV-Clone 13. LCMV-Armstrong was cleared in 100% of mice nine days after infection, while LCMV-Clone 13 persisted in all mice. However, half of the mice pre-electroporated with FADD or cFLIP plasmids were able to clear LCMV-Clone 13 by day nine, and, in the case of cFLIP, increased viral clearance was accompanied by higher CMI. Our studies imply that molecules in the Fas pathway are likely to affect a number of events in addition to the apoptosis of cells involved in immunity.

The role of platelets in the pathogenesis of viral hemorrhagic fevers.

Viral hemorrhagic fevers (VHF) are acute zoonotic diseases that, early on, seem to cause platelet destruction or dysfunction. Here we present the four major ways viruses affect platelet development and function and new evidence of molecular factors that are preferentially induced by the more pathogenic members of the families Flaviviridae, Bunyaviridae, Arenaviridae, and Filoviridae. A systematic search was performed through the main medical electronic databases using as parameters all current findings concerning platelets in VHF. Additionally, the review contains information from conference proceedings.

Genetic variation in vitro and in vivo of an attenuated Lassa vaccine candidate.

The attenuated Lassa vaccine candidate ML29 is a laboratory-produced reassortant between Lassa and Mopeia viruses, two Old World arenaviruses that differ by 40% in nucleic acid sequence. In our previous studies, ML29 elicited sterilizing immunity against Lassa virus challenge in guinea pigs and marmosets and virus-specific cell-mediated immunity in both simian immunodeficiency virus (SIV)-infected and uninfected rhesus macaques. Here, we show that ML29 is stable after 12 passages in vitro without losing its plaque morphology or its attenuated phenotype in suckling mice. Additionally, we used deep sequencing to characterize the viral population comprising the original stock of ML29, the stock of ML29 after 12 passages in Vero cells, and the ML29 isolates obtained from vaccinated animals. Twenty-seven isolates bore approximately 77 mutations that exceeded 20% of the single-nucleotide polymorphism (SNP) changes at any single locus. Of these 77 mutations, 5 appeared to be host specific, for example, appearing in mice but not in primates. None of these mutations were reversions of ML29 to the sequences of the parental Lassa and Mopeia viruses. The host-specific mutations indicate viral adaptations to virus-host interactions, and such interactions make reasonable targets for antiviral approaches. Variants capable of chronic infection did not emerge from any of the primate infections, even in immune-deficient animals, indicating that the ML29 reassortant is reasonably stable in vivo. In conclusion, the preclinical studies of ML29 as a Lassa virus vaccine candidate have been advanced, showing high levels of protection in nonhuman primates and acceptable stability both in vitro and in vivo.

Transcriptome analysis of human peripheral blood mononuclear cells exposed to Lassa virus and to the attenuated Mopeia/Lassa reassortant 29 (ML29), a vaccine candidate.

Lassa virus (LASV) is the causative agent of Lassa Fever and is responsible for several hundred thousand infections and thousands of deaths annually in West Africa. LASV and the non-pathogenic Mopeia virus (MOPV) are both rodent-borne African arenaviruses. A live attenuated reassortant of MOPV and LASV, designated ML29, protects rodents and primates from LASV challenge and appears to be more attenuated than MOPV. To gain better insight into LASV-induced pathology and mechanism of attenuation we performed gene expression profiling in human peripheral blood mononuclear cells (PBMC) exposed to LASV and the vaccine candidate ML29. PBMC from healthy human subjects were exposed to either LASV or ML29. Although most PBMC are non-permissive for virus replication, they remain susceptible to signal transduction by virus particles. Total RNA was extracted and global gene expression was evaluated during the first 24 hours using high-density microarrays. Results were validated using RT-PCR, flow cytometry and ELISA. LASV and ML29 elicited differential expression of interferon-stimulated genes (ISG), as well as genes involved in apoptosis, NF-kB signaling and the coagulation pathways. These genes could eventually serve as biomarkers to predict disease outcomes. The remarkable differential expression of thrombomodulin, a key regulator of inflammation and coagulation, suggests its involvement with vascular abnormalities and mortality in Lassa fever disease.

An attenuated Lassa vaccine in SIV-infected rhesus macaques does not persist or cause arenavirus disease but does elicit Lassa virus-specific immunity.

BACKGROUND: Lassa hemorrhagic fever (LHF) is a rodent-borne viral disease that can be fatal for human beings. In this study, an attenuated Lassa vaccine candidate, ML29, was tested in SIV-infected rhesus macaques for its ability to elicit immune responses without instigating signs pathognomonic for arenavirus disease. ML29 is a reassortant between Lassa and Mopeia viruses that causes a transient infection in non-human primates and confers sterilizing protection from lethal Lassa viral challenge. However, since the LHF endemic area of West Africa also has high HIV seroprevalence, it is important to determine whether vaccination could be safe in the context of HIV infection. RESULTS: SIV-infected and uninfected rhesus macaques were vaccinated with the ML29 virus and monitored for specific humoral and cellular immune responses, as well as for classical and non-classical signs of arenavirus disease. Classical disease signs included viremia, rash, respiratory distress, malaise, high liver enzyme levels, and virus invasion of the central nervous system. Non-classical signs, derived from profiling the blood transcriptome of virulent and non-virulent arenavirus infections, included increased expression of interferon-stimulated genes (ISG) and decreased expression of COX2, IL-1ß, coagulation intermediates and nuclear receptors needed for stress signaling. All vaccinated monkeys showed ML29-specific antibody responses and ML29-specific cell-mediated immunity. CONCLUSION: SIV-infected and uninfected rhesus macaques responded similarly to ML29 vaccination, and none developed chronic arenavirus infection. Importantly, none of the macaques developed signs, classical or non-classical, of arenavirus disease.

Arenavirus variations due to host-specific adaptation.

Arenavirus particles are enveloped and contain two single-strand RNA genomic segments with ambisense coding. Genetic plasticity of the arenaviruses comes from transcription errors, segment reassortment, and permissive genomic packaging, and results in their remarkable ability, as a group, to infect a wide variety of hosts. In this review, we discuss some in vitro studies of virus genetic and phenotypic variation after exposure to selective pressures such as high viral dose, mutagens and antivirals. Additionally, we discuss the variation in vivo of selected isolates of Old World arenaviruses, particularly after infection of different animal species. We also discuss the recent emergence of new arenaviruses in the context of our observations of sequence variations that appear to be host-specific.

Pathogenic Old World arenaviruses inhibit TLR2/Mal-dependent proinflammatory cytokines in vitro.

Lymphocytic choriomeningitis virus (LCMV), the prototype arenavirus, and Lassa virus (LASV), the causative agent of Lassa fever (LF), have extensive strain diversity and significant variations in pathogenicity for humans and experimental animals. The WE strain of LCMV (LCMV-WE), but not the Armstrong (Arm) strain, induces a fatal LF-like disease in rhesus macaques. We also demonstrated that LASV infection of human macrophages and endothelial cells resulted in reduced levels of proinflammatory cytokines. Here we have shown that cells infected with LASV or with LCMV-WE suppressed Toll-like receptor 2 (TLR2)-dependent proinflammatory cytokine responses. The persisting isolate LCMV clone 13 (CL13) also failed to stimulate interleukin-6 (IL-6) in macrophages. In contrast, nonpathogenic Mopeia virus, which is a genetic relative of LASV and LCMV-Arm induced robust responses that were TLR2/Mal dependent, required virus replication, and were enhanced by CD14. Superinfection experiments demonstrated that the WE strain of LCMV inhibited the Arm-mediated IL-8 response during the early stage of infection. In cells transfected with the NF-κB-luciferase reporter, infection with LCMV-Arm resulted in the induction of NF-κB, but cells infected with LCMV-WE and CL13 did not. These results suggest that pathogenic arenaviruses suppress NF-κB-mediated proinflammatory cytokine responses in infected cells.

Evaluation of Lassa virus vaccine immunogenicity in a CBA/J-ML29 mouse model.

Lassa fever (LF) is one of the most prevalent viral hemorrhagic fevers in West Africa responsible for thousands of deaths annually. The BSL-4 containment requirement and lack of small animal model to evaluate Lassa virus (LASV)-specific cell-mediated immunity (CMI) complicate development of effective LF vaccines. Here we have described a CBA/J-ML29 model allowing evaluation of LASV-specific CMI responses in mice. This model is based on Mopeia virus reassortant clone ML29, an attractive immunogenic surrogate for LASV. A single intraperitoneal (i.p.) immunization of CBA/J mice with ML29 protected animals against a lethal homologous intracerebral (i.c.) challenge with 588 LD(50). The ML29-immunized mice displayed negligible levels of LASV-specific antibody titers, but LASV-specific CMI responses were detectable early and peaked on day 8-10 after immunization. A T cell cytotoxicity assay in vivo showed a correlation between LASV-specific cytotoxicity and the timing of protection induced by the ML29 immunization. Notably, CBA/J mice that received CD8+ T cell-depleted splenocytes from ML29-immunized donors all succumbed to a lethal i.c. challenge, demonstrating that CD8+ T cells are critical in protection. The CBA/J-ML29 model can be useful immunological tool for the preliminary evaluation of immunogenicity and efficacy of vaccine candidates against LASV outside of BSL-4 containment facilities.

Lymphocytic choriomeningitis virus (LCMV) infection of macaques: a model for Lassa fever.

Arenaviruses such as Lassa fever virus (LASV) and lymphocytic choriomeningitis virus (LCMV) are benign in their natural reservoir hosts, and can occasionally cause severe viral hemorrhagic fever (VHF) in non-human primates and in human beings. LCMV is considerably more benign for human beings than Lassa virus, however certain strains, like the LCMV-WE strain, can cause severe disease when the virus is delivered as a high-dose inoculum. Here we describe a rhesus macaque model for Lassa fever that employs a virulent strain of LCMV. Since LASV must be studied within Biosafety Level-4 (BSL-4) facilities, the LCMV-infected macaque model has the advantage that it can be used at BSL-3. LCMV-induced disease is rarely as severe as other VHF, but it is similar in cases where vascular leakage leads to lethal systemic failure. The LCMV-infected macaque has been valuable for describing the course of disease with differing viral strains, doses and routes of infection. By monitoring system-wide changes in physiology and gene expression in a controlled experimental setting, it is possible to identify events that are pathognomonic for developing VHF and potential treatment targets.