Novel Oliveros-like Clade C Mammarenaviruses from Rodents in Argentina, 1990-2020.

Following an Argentine Hemorrhagic Fever (AHF) outbreak in the early 1990s, a rodent survey for Junín virus, a New World Clade B arenavirus, in endemic areas of Argentina was conducted. Since 1990, INEVH has been developing eco-epidemiological surveillance of rodents, inside and outside the Argentine Hemorrhagic Fever endemic area. Samples from rodents captured between 1993 and 2019 that were positive for Arenavirus infection underwent Sanger and unbiased, Illumina-based high-throughput sequencing, which yielded 5 complete and 88 partial Mammarenaviruses genomes. Previously, 11 genomes representing four species of New World arenavirus Clade C existed in public records. This work has generated 13 novel genomes, expanding the New World arenavirus Clade C to 24 total genomes. Additionally, two genomes exhibit sufficient genetic diversity to be considered a new species, as per ICTV guidelines (proposed name Mammarenavirus vellosense). The 13 novel genomes exhibited reassortment between the small and large segments in New World Mammarenaviruses. This work demonstrates that Clade C Mammarenavirus infections circulate broadly among Necromys species in the Argentine Hemorrhagic Fever endemic area; however, the risk for Clade C Mammarenavirus human infection is currently unknown.

Design and characterization of BSA-mycophenolic acid nanocomplexes: Antiviral activity exploration.

The interactions between bovine serum albumin (BSA) and mycophenolic acid (MPA) were investigated in silico through molecular docking and in vitro, using fluorescence spectroscopy. Dynamic light scattering and scanning electron microscopy were used to figure out the structure of MPA-Complex (MPA-C). The binding affinity between MPA and BSA was determined, yielding a Kd value of (12.0 ± 0.7) µM, and establishing a distance of 17 Å between the BSA and MPA molecules. The presence of MPA prompted protein aggregation, leading to the formation of MPA-C. The cytotoxicity of MPA-C and its ability to fight Junín virus (JUNV) were tested in A549 and Vero cell lines. It was found that treating infected cells with MPA-C decreased the JUNV yield and was more effective than free MPA in both cell line models for prolonged time treatments. Our results represent the first report of the antiviral activity of this type of BSA-MPA complex against JUNV, as assessed in cell culture model systems. MPA-C shows promise as a candidate for drug formulation against human pathogenic arenaviruses.

Nuclease Activity of the Junín Virus Nucleoprotein C-Terminal Domain.

The mammarenavirus Junín (JUNV) is the causative agent of Argentine hemorrhagic fever, a severe disease of public health concern. The most abundant viral protein is the nucleoprotein (NP), a multifunctional, two-domain protein with the primary role as structural component of the viral nucleocapsids, used as template for viral polymerase RNA synthesis activities. Here, we report that the C-terminal domain (CTD) of the attenuated Candid#1 strain of the JUNV NP can be purified as a stable soluble form with a secondary structure in line with known NP structures from other mammarenaviruses. We show that the JUNV NP CTD interacts with the viral matrix protein Z in vitro, and that the full-length NP and Z interact with each other in cellulo, suggesting that the NP CTD is responsible for this interaction. This domain comprises an arrangement of four acidic residues and a histidine residue conserved in the active site of exoribonucleases belonging to the DEDDh family. We show that the JUNV NP CTD displays metal-ion-dependent nuclease activity against DNA and single- and double-stranded RNA, and that this activity is impaired by the mutation of a catalytic residue within the DEDDh motif. These results further support this activity, not previously observed in the JUNV NP, which could impact the mechanism of the cellular immune response modulation of this important pathogen.

Modulation of the Aryl Hydrocarbon Receptor Signaling Pathway Impacts on Junín Virus Replication.

Junín virus (JUNV), a member of the family Arenaviridae, is the etiological agent of the Argentine hemorrhagic fever, an endemic disease in the rural region of Argentina lacking a specific chemotherapy. Aryl hydrocarbon receptor (AHR) is expressed in several mammalian tissues and has been indicated as a sensor of ligands from variable sources and a modulator of the cell immune response. Interestingly, recent studies have suggested that the activation or depression of the AHR signaling pathway may play a role in the outcome of diverse human viral infections. In the present report, the effect of the pharmacological modulation of AHR on JUNV in vitro infection was analyzed. An initial microarray screening showed that the AHR pathway was overexpressed in JUNV-infected hepatic cells. Concomitantly, the infection of Vero and Huh-7 cells with the JUNV strains IV4454 and Candid#1 was significantly inhibited in a dose-dependent manner by treatment with CH223191, a specific AHR antagonist, as detected by infectivity assays, real-time RT-PCR and immunofluorescence detection of viral proteins. Furthermore, the pro-viral role of AHR in JUNV infection appears to be independent of the IFN-I pathway. Our findings support the promising perspectives of the pharmacological modulation of AHR as a potential target for the control of AHF.

Recent developments on Junin virus, a causative agent for Argentine haemorrhagic fever.

Junin virus consists of ribonucleic acid as the genome and is responsible for a rapidly changing tendency of the virus. The virus is accountable for ailments in the human body and causes Argentine Haemorrhagic Fever (AHF). The infection is may be transmitted through contact between an infected animal/host and a person, and later between person to person. Prevention of outbreaks of AHF in humans can be a tough practice, as their occurrence is infrequent and unpredictable. In this review, recent information from the past 5 years available on the Junin virus including the risk of its emergence, infectious agents, its pathogenesis in humans, available diagnostic and therapeutic approaches, and disease management has been summarised. Altogether, this article would be highly significant in understanding the mechanistic basis behind virus interaction and other processes during the life cycle. Currently, no specific therapeutic options are available to treat the Junin virus infection. The information covered in this review could be important for finding possible treatment options for Junin virus infections.

The Virus-Host Interplay in Junín Mammarenavirus Infection.

Junín virus (JUNV) belongs to the Arenaviridae family and is the causative agent of Argentine hemorrhagic fever (AHF), a severe human disease endemic to agricultural areas in Argentina. At this moment, there are no effective antiviral therapeutics to battle pathogenic arenaviruses. Cumulative reports from recent years have widely provided information on cellular factors playing key roles during JUNV infection. In this review, we summarize research on host molecular determinants that intervene in the different stages of the viral life cycle: viral entry, replication, assembly and budding. Alongside, we describe JUNV tight interplay with the innate immune system. We also review the development of different reverse genetics systems and their use as tools to study JUNV biology and its close teamwork with the host. Elucidating relevant interactions of the virus with the host cell machinery is highly necessary to better understand the mechanistic basis beyond virus multiplication, disease pathogenesis and viral subversion of the immune response. Altogether, this knowledge becomes essential for identifying potential targets for the rational design of novel antiviral treatments to combat JUNV as well as other pathogenic arenaviruses.

[Co-circulation of Junín virus and other mammarenaviruses in a geographical area without confirmed cases of Argentine Hemorrhagic Fever]. / Cocirculación de virus Junin y otros mammarenavirus en área geográfica sin casos confirmados de Fiebre Hemorrágica Argentina.

Since the identification of Junin virus in the 1950s, many studies were carried out in wild rodents within the endemic area of the Argentine Hemorrhagic Fever (AHF) that recorded also the activity of the lymphocytic choriomeningitis virus (LCMV) and the Latino virus (LATV). The absence of confirmed cases of AHF since the 1990s in the department of Rio Cuarto, Córdoba province, promoted ecoepidemiological surveillance of infection of Calomys musculinus (Junin virus reservoir) and the search of reservoirs of the other mammarenaviruses. During two years of seasonal sampling, with a capture, mark and release system, 857 rodents were captured, corresponding 57.3% to the rodent reservoirs: C. musculinus, C. venustus and Mus musculus, being the first the most abundant species. Antibodies were detected and the three viral agents were molecularly characterized, showing a prevalence of infection of 3.5% (9/254) for Junin virus, 100% (3/3) for LCMV and 24.1% (21/87) for LATV. In conclusion, we demonstrated Junin virus circulation in its rodent reservoir in a region considered historic for AHF with potential risk for the population and the spatio-temporal co-circulation of the three mammarenaviruses in the central region of Argentina.

Desde la identificación del virus Junin en la década del 50, se realizaron numerosos estudios en roedores silvestres dentro del área endémica de la Fiebre Hemorrágica Argentina (FHA) que permitieron registrar, además, actividad del virus de la coriomeningitis linfocitaria (LCMV) y del virus Latino (LATV). La ausencia de casos confirmados de FHA desde la década del 90 en el departamento Río Cuarto, provincia de Córdoba, promovió la vigilancia ecoepidemiológica y de infección del Calomys musculinus (reservorio del virus Junin) y la búsqueda de reservorios e infección de los otros mammarenavirus. Durante dos años de muestreo estacional, con un sistema de captura, marcación y liberación capturamos 857 roedores, que correspondieron 57.3% a los reservorios: C. musculinus (especie más abundante), C. venustus y Mus musculus. Detectamos anticuerpos y caracterizamos molecularmente los tres agentes virales. Observamos una prevalencia de infección de 3.5% (9/254) para virus Junin, 100% (3/3) para LCMV y 24.1% (21/87) para LATV. En conclusión, demostramos circulación de virus Junin en su roedor reservorio, en una región considerada histórica para FHA con riesgo potencial para la población y cocirculación espacio-temporal de los tres mammarenavirus en la región central de Argentina.

Promyelocytic leukemia protein is a restriction factor for Junín virus independently of Z matrix protein.

The New World (NW) mammarenavirus Junín (JUNV) is the etiological agent of Argentine hemorrhagic fever, a human endemic disease of Argentina. Promyelocytic leukemia protein (PML) has been reported as a restriction factor for several viruses although the mechanism/s behind PML-mediated antiviral effect may be diverse and are a matter of debate. Previous studies have reported a nuclear to cytoplasm translocation of PML during the murine Old World mammarenavirus lymphocytic choriomeningitis virus (LCMV) infection. This translocation was found to be mediated by the viral Z protein. Here, we show that PML restricts JUNV infection in human A549 cells. However, in contrast to LCVM, JUNV infection enhances PML expression and PML is not translocated to the cytoplasm neither it colocalizes with JUNV Z protein. Our study demonstrates that a NW mammarenavirus as JUNV interacts differently with the antiviral protein PML than LCMV.

Machupo Virus with Mutations in the Transmembrane Domain and Glycosylation Sites of the Glycoprotein Is Attenuated and Immunogenic in Animal Models of Bolivian Hemorrhagic Fever.

Several highly pathogenic mammarenaviruses cause severe hemorrhagic and neurologic disease in humans for which vaccines and antivirals are limited or unavailable. New World (NW) mammarenavirus Machupo virus (MACV) infection causes Bolivian hemorrhagic fever in humans. We previously reported that the disruption of specific N-linked glycan sites on the glycoprotein (GPC) partially attenuates MACV in an interferon alpha/beta and gamma (IFN-α/ß and -γ) receptor knockout (R-/-) mouse model. However, some capability to induce neurological pathology still remained. The highly pathogenic Junin virus (JUNV) is another NW arenavirus closely related to MACV. An F427I substitution in the GPC transmembrane domain (TMD) rendered JUNV attenuated in a lethal mouse model after intracranial inoculation. In this study, we rationally designed and rescued a MACV containing mutations at two glycosylation sites and the corresponding F438I substitution in the GPC TMD. The MACV mutant is fully attenuated in IFN-α/ß and -γ R-/- mice and outbred guinea pigs. Furthermore, inoculation with this mutant MACV completely protected guinea pigs from wild-type MACV lethal challenge. Last, we found the GPC TMD F438I substitution greatly impaired MACV growth in neuronal cell lines of mouse and human origins. Our results highlight the critical roles of the glycans and the TMD on the GPC in arenavirus virulence, which provide insight into the rational design of potential vaccine candidates for highly pathogenic arenaviruses. IMPORTANCE For arenaviruses, the only vaccine available is the live attenuated Candid#1 vaccine, a JUNV vaccine approved in Argentina. We and others have found that the glycans on GPC and the F427 residue in the GPC TMD are important for virulence of JUNV. Nevertheless, mutating either of them is not sufficient for full and stable attenuation of JUNV. Using reverse genetics, we disrupted specific glycosylation sites on MACV GPC and also introduced the corresponding F438I substitution in the GPC TMD. This MACV mutant is fully attenuated in two animal models and protects animals from lethal infection. Thus, our studies highlight the feasibility of rational attenuation of highly pathogenic arenaviruses for vaccine development. Another important finding from this study is that the F438I substitution in GPC TMD could substantially affect MACV replication in neurons. Future studies are warranted to elucidate the underlying mechanism and the implication of this mutation in arenavirus neural tropism.

Inhibitors of the p38 cell signaling pathway as antiviral compounds against Junín virus.

In the present study, we analyzed the modulation of p38 cell signaling by Junín virus (JUNV) and evaluated the antiviral activity of p38 inhibitors against JUNV. While JUNV induced a progressive activation of p38 throughout the infection in Vero cells, a partial downregulation of p38 phosphorylation was observed in HEK293 and HeLa cells. The compounds SB203580 and SB202190, which are selective inhibitors of p38, significantly reduced viral protein expression and viral yield in the cell lines examined, indicating that the p38 signaling pathway might be a promising antiviral target against JUNV infection.

Therapy for Argentine hemorrhagic fever in nonhuman primates with a humanized monoclonal antibody.

The COVID-19 pandemic has reemphasized the need to identify safe and scalable therapeutics to slow or reverse symptoms of disease caused by newly emerging and reemerging viral pathogens. Recent clinical successes of monoclonal antibodies (mAbs) in therapy for viral infections demonstrate that mAbs offer a solution for these emerging biothreats. We have explored this with respect to Junin virus (JUNV), an arenavirus classified as a category A high-priority agent and the causative agent of Argentine hemorrhagic fever (AHF). There are currently no Food and Drug Administration-approved drugs available for preventing or treating AHF, although immune plasma from convalescent patients is used routinely to treat active infections. However, immune plasma is severely limited in quantity, highly variable in quality, and poses significant safety risks including the transmission of transfusion-borne diseases. mAbs offer a highly specific and consistently potent alternative to immune plasma that can be manufactured at large scale. We previously described a chimeric mAb, cJ199, that provided protection in a guinea pig model of AHF. To adapt this mAb to a format more suitable for clinical use, we humanized the mAb (hu199) and evaluated it in a cynomolgus monkey model of AHF with two JUNV isolates, Romero and Espindola. While untreated control animals experienced 100% lethality, all animals treated with hu199 at 6 d postinoculation (dpi) survived, and 50% of animals treated at 8 dpi survived. mAbs like hu199 may offer a safer, scalable, and more reproducible alternative to immune plasma for rare viral diseases that have epidemic potential.

Clinical Management of Argentine Hemorrhagic Fever using Ribavirin and Favipiravir, Belgium, 2020.

We report a case of Argentine hemorrhagic fever diagnosed in a woman in Belgium who traveled from a disease-endemic area. Patient management included supportive care and combination therapy with ribavirin and favipiravir. Of 137 potential contacts, including friends, relatives, and healthcare and laboratory workers, none showed development of clinical symptoms of this disease.

Junin Virus Triggers Macrophage Activation and Modulates Polarization According to Viral Strain Pathogenicity.

The New World arenavirus Junin (JUNV) is the etiological agent of Argentine hemorrhagic fever (AHF). Previous studies of human macrophage infection by the Old-World arenaviruses Mopeia and Lassa showed that while the non-pathogenic Mopeia virus replicates and activates human macrophages, the pathogenic Lassa virus replicates but fails to activate human macrophages. Less is known in regard to the impact of New World arenavirus infection on the human macrophage immune response. Macrophage activation is critical for controlling infections but could also be usurped favoring immune evasion. Therefore, it is crucial to understand how the JUNV infection modulates macrophage plasticity to clarify its role in AHF pathogenesis. With this aim in mind, we compared infection with the attenuated Candid 1 (C#1) or the pathogenic P strains of the JUNV virus in human macrophage cultures. The results showed that both JUNV strains similarly replicated and induced morphological changes as early as 1 day post-infection. However, both strains differentially induced the expression of CD71, the receptor for cell entry, the activation and maturation molecules CD80, CD86, and HLA-DR and selectively modulated cytokine production. Higher levels of TNF-α, IL-10, and IL-12 were detected with C#1 strain, while the P strain induced only higher levels of IL-6. We also found that C#1 strain infection skewed macrophage polarization to M1, whereas the P strain shifted the response to an M2 phenotype. Interestingly, the MERTK receptor, that negatively regulates the immune response, was down-regulated by C#1 strain and up-regulated by P strain infection. Similarly, the target genes of MERTK activation, the cytokine suppressors SOCS1 and SOCS3, were also increased after P strain infection, in addition to IRF-1, that regulates type I IFN levels, which were higher with C#1 compared with P strain infection. Together, this differential activation/polarization pattern of macrophages elicited by P strain suggests a more evasive immune response and may have important implications in the pathogenesis of AHF and underpinning the development of new potential therapeutic strategies.

Junín virus induces autophagy in human A549 cells.

Autophagy, a highly regulated degradative process that promotes cellular homeostasis, is increasingly recognised as a fundamental component of the cellular response against viral infection. In this study, we investigated the role of autophagy during Junín virus (JUNV) multiplication using human A549 cells. We found that JUNV infection induces an increment of the LC3-II/LC3-I ratio, an accumulation of punctate pattern in RFP-LC3-transfected cells and the colocalisation of viral nucleoprotein and LC3 protein, suggesting autophagosome formation. JUNV infection also induced the degradation of the autophagy receptor p62, suggesting that complete autophagic flux was triggered. In addition, we showed that inhibition of autophagy with bafilomycin A1 or 3-methyladenine significantly reduces viral multiplication. Moreover, viral yield was increased when autophagy was induced using rapamycin. Furthermore, JUNV infection induced the colocalisation of p62, ATG16, RAB5, RAB7A and LAMP1 with the autophagosomal LC3 protein. That suggests that phagosomes undergo the maturation process during viral infection. Finally, we demonstrated that siRNA experiments targeting essential autophagy genes (ATG5, ATG7 and Beclin 1) reduce viral protein synthesis and viral yield. Overall, our results indicate that JUNV activates host autophagy machinery enhancing its multiplication.

Infection of Rat Osteoblasts with Junin Virus Promotes the Expression of Bone Morphogenetic Protein 6, an Osteogenic Differentiation Inducer.

BACKGROUND: The arenavirus Junin virus (JUNV), causative agent of the argentine hemorrhagic fever, is able to modulate several signaling pathways involved in cell survival and multiplication. OBJECTIVES: We aimed to characterize the infection of rat osteoblasts (OBCs) with JUNV and its consequence on the modulation of osteogenic genes expression, thus studying the ability of this virus to induce cell differentiation. In addition, we evaluated the effect of purinergic agonists on viral replication. METHOD: Quantification of infectivity by plaque forming unit (PFU) assay, synthesis of viral proteins by western blot and immunofluorescence, and expression of osteogenic differentiation markers (ODM) by quantitative real-time polymerase chain reaction were employed. RESULTS: Infection of OBCs with JUNV (MOI 0.01 PFU/cell) showed a peak of infectivity, reaching 1.5 × 105 PFU/mL at the second day post-infection (p.i.). A marked restriction in multiplication was detected at day 7 p.i. that did not impair the establishment of a persistent stage of infection in OBCs. Analysis of mRNAs corresponding to ODM such as alkaline phosphatase, bone sialo-protein, and bone morphogenetic proteins (BMPs) 4 and 6 revealed that only the levels of BMP-6 were significantly higher in infected cells. Treatment with the purinergic agonists ATPγS, UTP, ADP, or UDP diminished viral titer and reduced the expression of the viral nucleoprotein. Also, treatment with 10 µM ATPγS reduced the stimulation of BMP-6 expression induced by the infection. CONCLUSIONS: These data demonstrate that JUNV is capable of infecting OBCs and point out BMP-6 as a key factor during this process.

Modified ribavirin analogues as antiviral agents against Junín virus.

In this work, several ribavirin analogues were synthesized and incorporated into a multivalent arrangement. Both were subsequently modified by the addition of polyhydroxylated residues. Their antiviral activity was tested against Junín virus, etiological agent responsible of Argentine hemorrhagic fever. Some compounds inhibited Junín virus in the range of 13.2-389.1 µM. Two modified ribavirin analogues presented an effective concentration comparable to ribavirin but with a higher selectivity index.

[Spatial difference in the incidence of Argentine Hemorrhagic Fever and composition and abundance of rodents in the assemblage]. / Diferencia espacial de la incidencia de fiebre hemorrágica argentina y la composición y abundancia de roedores en el ensamble.

BACKGROUND: The Argentine Hemorrhagic Fever (AHF) is a zoonotic disease endemic in a wide area of the humid pampa of Argentina. The etiologic agent is the Junin virus that is maintained in the wild by the rodent Calomys musculinus and transmitted to humans, mainly, through aerosols generated from secretions and excretions. AIMS: To characterize and compare the assemblages of small rodent composition and diversity inside the epidemic, historic and non-endemic zone of AHF and to register C. musculinus abundance in each zone and in each area within each zone, registering the prevalence of infection in rodent populations. METHOD: One central and two peripheral areas were delimited to sample rodents in each zone with different incidence of AHF. Thus, 18 localities were selected to do the sampling in two years. Host abundance between zones and among areas inside each zone and among nearby areas between zones were compared applying nested ANOVA's. RESULTS: In each zone, the rodent assemblage showed differences in composition, diversity and numeric representation of C. musculinus. The epidemic zone was the richest of the three, registering also great host abundance; meanwhile in the historic zone, A. azarae was the dominant numeric species with less number of other species. Regarding the non-endemic zone, the assemblage composition and C. musculinus abundance varied respect the sampled year. Junin virus infection was only detected in C. musculinus individuals corresponding to the epidemic zone, with a prevalence of 2.7 and 1.1% for the years 2007 and 2008, respectively. CONCLUSION: In this system, the abundance of C. musculinus could be impacting over the pathogen dynamic, rather than the assemblage diversity or the A. azarae presence.

Diferencia espacial de la incidencia de fiebre hemorrágica argentina y la composición y abundancia de roedores en el ensamble / Spatial difference in the incidence of Argentine Hemorrhagic Fever and composition and abundance of rodents in the assemblage

Resumen La fiebre hemorrágica argentina (FHA) es una enfermedad zoonótica endémica en una amplia zona de la pampa húmeda de Argentina. El agente etiológico es el virus Junin que es mantenido en la naturaleza por el roedor Calomys musculinus y transmitido, principalmente, al humano a través de aerosoles generados de las secreciones y excreciones. Objetivos: Caracterizar la composición y diversidad de los ensambles de pequeños roedores, determinar la abundancia del hospedador C. musculinus y la prevalencia del virus de la FHA en las zonas epidémica, histórica y no endémica de dicha enfermedad en Argentina. Métodos: Para el muestreo de roedores en cada una de las zonas se demarcaron un área central y dos periféricas para 18 localidades de la región central de Argentina (incluyendo las provincias de Córdoba, Buenos Aires y Santa Fe) muestreadas en dos años. Se comparó la abundancia de C. musculinus entre zonas y entre las áreas dentro de cada zona y áreas cercanas entre zonas, utilizando modelos de análisis de varianza anidados. Resultados. Dentro de cada zona, el ensamble de roedores mostró diferencia espacial en la composición específica, diversidad y abundancia de C. musculinus. La zona epidémica registró mayor número de especies y mayor abundancia del hospedador. En zona histórica se capturó el menor número de especies (de roedores) y Akodon azarae fue la más abundante. En zona no endémica la composición del ensamble y la abundancia de C. musculinus variaron entre los dos años. Sólo se detectó infección por virus Junin en C. musculinus correspondientes a la zona epidémica con una prevalencia de 2,7 y 1,1% para los años 2007 y 2008, respectivamente. Conclusión: En este sistema, la abundancia del hospedador estaría afectando la dinámica espacial de este virus, más que la diversidad del ensamble o la presencia de A. azarae.

Background. The Argentine Hemorrhagic Fever (AHF) is a zoonotic disease endemic in a wide area of the humid pampa of Argentina. The etiologic agent is the Junin virus that is maintained in the wild by the rodent Calomys musculinus and transmitted to humans, mainly, through aerosols generated from secretions and excretions. Aims: To characterize and compare the assemblages of small rodent composition and diversity inside the epidemic, historic and non-endemic zone of AHF and to register C. musculinus abundance in each zone and in each area within each zone, registering the prevalence of infection in rodent populations. Method: One central and two peripheral areas were delimited to sample rodents in each zone with different incidence of AHF. Thus, 18 localities were selected to do the sampling in two years. Host abundance between zones and among areas inside each zone and among nearby areas between zones were compared applying nested ANOVA's. Results: In each zone, the rodent assemblage showed differences in composition, diversity and numeric representation of C. musculinus. The epidemic zone was the richest of the three, registering also great host abundance; meanwhile in the historic zone, A. azarae was the dominant numeric species with less number of other species. Regarding the non-endemic zone, the assemblage composition and C. musculinus abundance varied respect the sampled year. Junin virus infection was only detected in C. musculinus individuals corresponding to the epidemic zone, with a prevalence of 2.7 and 1.1% for the years 2007 and 2008, respectively. Conclusion: In this system, the abundance of C. musculinus could be impacting over the pathogen dynamic, rather than the assemblage diversity or the A. azarae presence.

Effective Population Size Differences in Calomys musculinus, the Host of Junín Virus: Their Relationship with the Epidemiological History of Argentine Hemorrhagic Fever.

Argentine hemorrhagic fever (AHF) is a serious endemic disease in Argentina, produced by Junín virus, whose host is the Sigmodontinae rodent Calomys musculinus. Within the endemic area, human incidence and proportion of infected rodents remains high for 5-10 years after the first appearance of the disease (epidemic [E] zone) and then gradually declines to sporadic cases (historic [H] zone). We tested the hypothesis that host populations within the E zone are large and well connected by gene flow, facilitating the transmission and maintenance of the virus, whereas those in the H and nonendemic (NE) zones are small and isolated, with the opposite effect. We estimated parameters affected by levels of gene flow and population size in 14 populations of C. musculinus: population effective size (Ne), genetic variability, and mean relatedness. Our hypothesis was not supported: the lowest levels of variability and of Ne and the highest genetic relatedness among individuals were found in the H zone. Populations from the NE zone displayed opposite results, whereas those in the E zone showed intermediate values. If we consider that populations are first NE, then E, and finally H, a correlative decrease in Ne was observed. Chronically infected females have a low reproductive success. We propose that this would lower Ne because each cohort would originate from a fraction of females of the previous generation, and affect other factors such as proportion of individuals that develop acute infection, probability of viral transmission, and evolution of virulence, which would explain, at least partly, the changing incidence of AHF.

Antiviral activity of A771726, the active metabolite of leflunomide, against Junín virus.

The aim of this study was to investigate the effect of A771726, the active metabolite of leflunomide, (CONICET-UBA), Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Ciudad against the infection with Junín virus (JUNV), agent of Argentine hemorrhagic fever (AHF). The treatment with non-cytotoxic concentrations of A771726 of Vero and A549 cells infected with JUNV inhibited virus replication in a dose-dependent manner, as determined by virus yield reduction assay. The antiviral effectiveness of A771726 was not importantly affected by the multiplicity of infection and the virus strain. Moreover, the combination of A771726 and ribavirin had a significantly more potent antiviral activity than each single drug treatment. Mechanistic studies showed that the main action of A771726 is exerted before 6 h of JUNV infection. Accordingly, inhibition of viral RNA synthesis was detected in treated infected cells by real time RT-PCR. The exogenous addition of uridine or orotic acid produced a partial reversal of the inhibitory effect of A771726 on infective virus production whereas a total reversion was detected on JUNV RNA synthesis, probably by restoration of the enzymatic activity of dihydroorotate dehydrogenase (DHODH) and the intracellular pyrimidine pools. In conclusion, these results suggest that the antiviral target would be viral RNA synthesis through pyrimidine depletion, but any other effect of the compound on JUNV infection cannot be excluded. This study opens the possibility of the therapeutic application of a wide spectrum host-targeted compound alone or in combination with ribavirin to combat AHF as well as other human pathogenic arenaviruses.