South American Hemorrhagic Fevers Research Articles

South American Hemorrhagic Fever viruses and the cutting edge of the vaccine and antiviral development

South American Hemorrhagic Fever is caused by the Arenavirus, which belong to the Family Arenaviridae, genus mammarenavirus, infection at South America. South American Hemorrhagic Fever includes 1. Argentinian Hemorrhagic fever caused by Junin virus, 2. Brazilian hemorrhagic fever caused by Sabia virus, 3. Venezuelan Hemorrhagic fever caused by Guanarito virus, 4. Bolivian Hemorrhagic fever caused by Machupo virus, and 5. Unassigned hemorrhagic fever caused by Chapare virus. These viruses are classified in New World (NW) Arenavirus, which is different from Old World Arenavirus (ex. Lassa virus), based on phylogeny, serology, and geographic differences. In this review, the current knowledge of the biology and the development of the vaccines and antivirals of NW Arenaviruses which cause South American Hemorrhagic Fever will be described.

Rodent borne zoonoses: A brief review

Rodents are the most abundant and diversified order of living mammals in the world. Since the middle ages they are known to contribute to human diseases, eg. black rats associated with distribution of plague. In modern times also rodents form a threat for public health and a number of pathogens are directly or indirectly transmitted by rodents. Of 2277 extant rodent species, 217 species are reservoirs harboring 66 zoonoses caused by viruses, bacteria, fungi, helminths, and protozoa and play a major role in their transmission and spread in different ways. Among the most important diseases in terms of public health are salmonellosis, plague, leptospirosis, leishmaniasis, toxoplasmosis, rat-bit fever, taeniasis-like Capillaria hepatica, zoonotic babesiosis, Lassa fever, hemorrhagic fever with renal syndrome (HFRS), and the hantavirus cardiopulmonary syndrome (HCPS). In addition, Arenaviruses are responsible for South American Hemorrhagic Fevers (SAHF). Rodents may harbor different complex bacteria, such as Mycobacterium tuberculosis and Mycobacterium microti, Escherichia coli, agents of Tularemia, Tick-borne relapsing fever, Bartonellosis, Listeriosis, Lyme disease, Q fever, Ehrlichiosis and others. Recent changes in the global climate and ecosystem composition led to the spread of rodents and rodent-borne pathogens globally. Rodent control in rural environments relies mainly on the use of rodenticides. Other management methods which can be applied are trapping, habitat management (e.g., proofing, sanitation), and biocontrol (e.g., wild or domestic predators, rodent pathogens).

Antibody-Based Inhibition of Pathogenic New World Hemorrhagic Fever Mammarenaviruses by Steric Occlusion of the Human Transferrin Receptor 1 Apical Domain.

ABSTRACTPathogenic clade B New World mammarenaviruses (NWM) can cause Argentine, Venezuelan, Brazilian, and Bolivian hemorrhagic fevers. Sequence variability among NWM glycoproteins (GP) poses a challenge to the development of broadly neutralizing therapeutics against the entire clade of viruses. However, blockade of their shared binding site on the apical domain of human transferrin receptor 1 (hTfR1/CD71) presents an opportunity for the development of effective and broadly neutralizing therapeutics. Here, we demonstrate that the murine monoclonal antibody OKT9, which targets the apical domain of hTfR1, can sterically block cellular entry by viral particles presenting clade B NWM glycoproteins (GP1-GP2). OKT9 blockade is also effective against viral particles pseudotyped with glycoproteins of a recently identified pathogenic Sabia-like virus. With nanomolar affinity for hTfR1, the OKT9 antigen binding fragment (OKT9-Fab) sterically blocks clade B NWM-GP1s and reduces infectivity of an attenuated strain of Junin virus. Binding of OKT9 to the hTfR1 ectodomain in its soluble, dimeric state produces stable assemblies that are observable by negative-stain electron microscopy. A model of the OKT9-sTfR1 complex, informed by the known crystallographic structure of sTfR1 and a newly determined structure of the OKT9 antigen binding fragment (Fab), suggests that OKT9 and the Machupo virus GP1 share a binding site on the hTfR1 apical domain. The structural basis for this interaction presents a framework for the design and development of high-affinity, broadly acting agents targeting clade B NWMs.IMPORTANCE Pathogenic clade B NWMs cause grave infectious diseases, the South American hemorrhagic fevers. Their etiological agents are Junin (JUNV), Guanarito (GTOV), Sabiá (SABV), Machupo (MACV), Chapare (CHAV), and a new Sabiá-like (SABV-L) virus recently identified in Brazil. These are priority A pathogens due to their high infectivity and mortality, their potential for person-to-person transmission, and the limited availability of effective therapeutics and vaccines to curb their effects. While low homology between surface glycoproteins of NWMs foils efforts to develop broadly neutralizing therapies targeting NWMs, this work provides structural evidence that OKT9, a monoclonal antibody targeting a single NWM glycoprotein binding site on hTfR1, can efficiently prevent their entry into cells.

Involvement of Pro-Inflammatory Macrophages in Liver Pathology of Pirital Virus-Infected Syrian Hamsters.

New World arenaviruses cause fatal hemorrhagic disease in South America. Pirital virus (PIRV), a mammarenavirus hosted by Alston’s cotton rat (Sigmodon alstoni), causes a disease in Syrian golden hamsters (Mesocricetus auratus) (biosafety level-3, BSL-3) that has many pathologic similarities to the South American hemorrhagic fevers (BSL-4) and, thus, is considered among the best small-animal models for human arenavirus disease. Here, we extend in greater detail previously described clinical and pathological findings in Syrian hamsters and provide evidence for a pro-inflammatory macrophage response during PIRV infection. The liver was the principal target organ of the disease, and signs of Kupffer cell involvement were identified in mortally infected hamster histopathology data. Differential expression analysis of liver mRNA revealed signatures of the pro-inflammatory response, hematologic dysregulation, interferon pathway and other host response pathways, including 17 key transcripts that were also reported in two non-human primate (NHP) arenavirus liver-infection models, representing both Old and New World mammarenavirus infections. Although antigen presentation may differ among rodent and NHP species, key hemostatic and innate immune-response components showed expression parallels. Signatures of pro-inflammatory macrophage involvement in PIRV-infected livers included enrichment of Ifng, Nfkb2, Stat1, Irf1, Klf6, Il1b, Cxcl10, and Cxcl11 transcripts. Together, these data indicate that pro-inflammatory macrophage M1 responses likely contribute to the pathogenesis of acute PIRV infection.

Importation of rare but life-threatening and highly contagious diseases. Current situation and outlook

ZusammenfassungAls Internist muss man damit rechnen, Erstbehandler eines Patienten mit selten importierter hochkontagiöser, lebensbedrohlicher Erkrankung zu sein. Dies kommt sicherlich nur äußerst selten vor, hat aber enorme Konsequenzen. Man sollte daher grundsätzlich die viralen hämorrhagischen Fieber kennen: Filoviren verursachen das Ebola- und Marburg-Fieber, Arenaviren das Lassa-Fieber und südamerikanische hämorrhagische Fieber, die Bunya-Viren u. a. das hämorrhagische Krim-Kongo-Fieber. Weiterhin sollte man einige hochkontagiöse respiratorische Infektionen wie das „hantavirus pulmonary syndrome“, die Lungenpest und das „Middle East respiratory syndrome“ kennen.

Interferon response in a hamster model of arenavirus hemorrhagic disease

A difficulty in studying hemorrhagic arenavirus pathogenesis is a lack of animal models that recapitulate human disease and which can be manipulated at BSL-3 or lower. Pirital virus (PIRV), a South American arenavirus hosted by Alston’s cotton rats ( Sigmodon alstoni ), has not been shown to cause disease in humans and is considered a BSL-3 agent. Infection of Syrian golden hamsters ( Mesocricetus auratus ) causes a disease that shares many features of the South American hemorrhagic fevers and Lassa fever. Significantly, neurological manifestations occur, a feature commonly found in Argentine hemorrhagic fever and which is absent in most other models of arenavirus disease. We examined the pathogenesis of PIRV infection by clinical and molecular methods, with a focus on gene expression of the antiviral type I interferon response using RNA-Seq. More than 3,000 genes were differentially expressed in the livers of infected hamsters, of which 86 are involved with antiviral responses. Several of these genes participate in apoptosis and autophagy, which may suggest a mechanism of pathogenesis observed in damaged livers.

Serological evidence that Tacaribe virus is circulating among bats in Trinidad and Tobago

Tacaribe virus (TCRV) is a bisegmented, ambisense, RNA virus within the genus Arenavirus . Arenaviruses are grouped into Old World lymphocytic choriomeningitis-Lassa virus complex and the New World Tacaribe complex viruses. TCRV is placed within the Tacaribe complex along with the South American hemorrhagic fever viruses: Chapare, Guanarito, Junin, Machupo, and Sabia viruses. The only isolates of TCRV were from 11 artibeus bats collected by investigators at the Trinidad Regional Virology Laboratory in the Republic of Trinidad in the 1950 s. TCRV has not been isolated since, although serological data from the 1970 s suggested it may circulate among Caribbean bats. Only one isolate remains, TRVL-11573, and it has been passaged in suckling mice and Vero cells. We sought to determine if TCRV is still circulating in bat populations in Trinidad through serological investigation. We developed an ELISA and western blot assay using His-tagged recombinant TCRV nucleocapsid antigen. Serum from Artibeus jamaicensis that had been experimentally infected with TCRV was used as a positive control, and serum collected from an uninfected A. jamaicensis used as a negative control. ELISA screen of bloods from 84 bats of various species captured in Trinidad identified several, mostly artibeus bats, as seropositive for antibodies to TCRV. Some of these were tested by western blot. Four were negative, eight were weakly positive, and five were strongly positive. These results suggest that TCRV or other arenaviruses continue to circulate among bats in Trinidad.

Multiplex degenerate primer design for targeted whole genome amplification of many viral genomes.

Background. Targeted enrichment improves coverage of highly mutable viruses at low concentration in complex samples. Degenerate primers that anneal to conserved regions can facilitate amplification of divergent, low concentration variants, even when the strain present is unknown. Results. A tool for designing multiplex sets of degenerate sequencing primers to tile overlapping amplicons across multiple whole genomes is described. The new script, run_tiled_primers, is part of the PriMux software. Primers were designed for each segment of South American hemorrhagic fever viruses, tick-borne encephalitis, Henipaviruses, Arenaviruses, Filoviruses, Crimean-Congo hemorrhagic fever virus, Rift Valley fever virus, and Japanese encephalitis virus. Each group is highly diverse with as little as 5% genome consensus. Primer sets were computationally checked for nontarget cross reactions against the NCBI nucleotide sequence database. Primers for murine hepatitis virus were demonstrated in the lab to specifically amplify selected genes from a laboratory cultured strain that had undergone extensive passage in vitro and in vivo. Conclusions. This software should help researchers design multiplex sets of primers for targeted whole genome enrichment prior to sequencing to obtain better coverage of low titer, divergent viruses. Applications include viral discovery from a complex background and improved sensitivity and coverage of rapidly evolving strains or variants in a gene family.

Transcriptome Sequencing and Annotation for the Jamaican Fruit Bat (Artibeus jamaicensis)

The Jamaican fruit bat (Artibeus jamaicensis) is one of the most common bats in the tropical Americas. It is thought to be a potential reservoir host of Tacaribe virus, an arenavirus closely related to the South American hemorrhagic fever viruses. We performed transcriptome sequencing and annotation from lung, kidney and spleen tissues using 454 and Illumina platforms to develop this species as an animal model. More than 100,000 contigs were assembled, with 25,000 genes that were functionally annotated. Of the remaining unannotated contigs, 80% were found within bat genomes or transcriptomes. Annotated genes are involved in a broad range of activities ranging from cellular metabolism to genome regulation through ncRNAs. Reciprocal BLAST best hits yielded 8,785 sequences that are orthologous to mouse, rat, cattle, horse and human. Species tree analysis of sequences from 2,378 loci was used to achieve 95% bootstrap support for the placement of bat as sister to the clade containing horse, dog, and cattle. Through substitution rate estimation between bat and human, 32 genes were identified with evidence for positive selection. We also identified 466 immune-related genes, which may be useful for studying Tacaribe virus infection of this species. The Jamaican fruit bat transcriptome dataset is a resource that should provide additional candidate markers for studying bat evolution and ecology, and tools for analysis of the host response and pathology of disease.

An Antibody Recognizing the Apical Domain of Human Transferrin Receptor 1 Efficiently Inhibits the Entry of All New World Hemorrhagic Fever Arenaviruses

Five New World (NW) arenaviruses cause human hemorrhagic fevers. Four of these arenaviruses are known to enter cells by binding human transferrin receptor 1 (hTfR1). Here we show that the fifth arenavirus, Chapare virus, similarly uses hTfR1. We also identify an anti-hTfR1 antibody, ch128.1, which efficiently inhibits entry mediated by the glycoproteins of all five viruses, as well as replication of infectious Junín virus. Our data indicate that all NW hemorrhagic fever arenaviruses utilize a common hTfR1 apical-domain epitope and suggest that therapeutic agents targeting this epitope, including ch128.1 itself, can be broadly effective in treating South American hemorrhagic fevers.

Vaccine Platforms to Control Arenaviral Hemorrhagic Fevers

Arenaviruses are rodent-borne emerging human pathogens. Diseases caused by these viruses, e.g., Lassa fever (LF) in West Africa and South American hemorrhagic fevers (HFs), are serious public health problems in endemic areas. We have employed replication-competent and replication-deficient strategies to design vaccine candidates potentially targeting different groups "at risk". Our leader LF vaccine candidate, the live reassortant vaccine ML29, is safe and efficacious in all tested animal models including non-human primates. In this study we showed that treatment of fatally infected animals with ML29 two days after Lassa virus (LASV) challenge protected 80% of the treated animals. In endemic areas, where most of the target population is poor and many live far from health care facilities, a single-dose vaccination with ML29 would be ideal solution. Once there is an outbreak, a fast-acting vaccine or post-exposure prophylaxis would be best. The 2(nd) vaccine technology is based on Yellow Fever (YF) 17D vaccine. We designed YF17D-based recombinant viruses expressing LASV glycoproteins (GP) and showed protective efficacy of these recombinants. In the current study we developed a novel technology to clone LASV nucleocapsid within YF17D C gene. Low immunogenicity and stability of foreign inserts must be addressed to design successful LASV/YFV bivalent vaccines to control LF and YF in overlapping endemic areas of West Africa. The 3(rd) platform is based on the new generation of alphavirus replicon virus-like-particle vectors (VLPV). Using this technology we designed VLPV expressing LASV GP with enhanced immunogenicity and bivalent VLPV expressing cross-reactive GP of Junin virus (JUNV) and Machupo virus (MACV), causative agents of Argentinian and Bolivian HF, respectively. A prime-boost regimen required for VLPV immunization might be practical for medical providers, military, lab personnel, and visitors in endemic areas.

BSL4 Facilities in Anti-Infectious Disease Measures

Pathogens are divided into biosafety levels (BSL) 1 to 4 based on multiple factors such as virulence, transmissibility, environment effect, and treatment availability. BSL1 pathogens are the least virulent and BSL4 the most. BSL4 pathogens include ebolavirus, marburgvirus, Crimean-Congo hemorrhagic fever virus, lassa virus, variolla virus, and South American hemorrhagic fever viruses, as detailed in Table 1. Pathogens at each of the 4 BSLs must be handled in equivalently physically contained laboratories, graded P1-4. BSL4 pathogens do not exist in nature in Japan, which currently has no equivalent physical containment facilities, but the possibility exists that they may be brought into the country unintentionally by those infected in endemic areas or intentionally by bioterrorists.

Site 1 Protease Is Required for Proteolytic Processing of the Glycoproteins of the South American Hemorrhagic Fever Viruses Junin, Machupo, and Guanarito

The cellular proprotein convertase site 1 protease (S1P) has been implicated in the proteolytic processing of the glycoproteins (GPs) of Old World arenaviruses. Here we report that S1P is also involved in the processing of the GPs of the genetically more-distant South American hemorrhagic fever viruses Guanarito, Machupo, and Junin. Efficient cleavage of Guanarito virus GP, whose protease recognition sites deviate from the reported S1P consensus sequence, indicates a broader specificity of S1P than anticipated. Lack of GP processing of Junin virus dramatically reduced production of infectious virus and prevented cell-to-cell propagation. Infection of S1P-deficient cells resulted in viral persistence over several weeks without the emergence of escape variants able to use other cellular proteases for GP processing.

Absence of CTL Responses to Early Viral Antigens Facilitates Viral Persistence

CD8+ T cells are crucial for the control of intracellular pathogens such as viruses and some bacteria. Using lymphocytic choriomeningitis virus (LCMV) infection of mice--the prototypic arenavirus evolutionarily closely related to human Lassa fever and South American hemorrhagic fever viruses, we have shown previously that the kinetics of Ag presentation determine immunodominance of the LCMV-specific CTL response due to progressive exhaustion of LCMV nucleoprotein (NP)-specific CTL upon increasing viral load. In this study, we provide evidence that CTL against early LCMV NP-derived epitopes are more important in virus control than those against late glycoprotein-derived epitopes. We show that mice that are tolerant to all NP-derived T cell epitopes are severely compromised in their ability to control larger inocula of LCMV, supporting our hypothesis that CD8+ T cells specific for early viral Ags play a major role in acute virus control. Thus, the kinetics with which virus-derived T cell epitopes are presented has a strong impact on the efficacy of the antiviral immunity. This aspect should be taken into consideration for the development of vaccines.

Viral exanthems in the tropics

Viral exanthems are a common problem in tropical regions, particularly affecting children. Most exanthems are transient and harmless, but some are potentially very dangerous. Pregnant women and malnourished or immunocompromised infants carry the greatest risk of adverse outcome. In this article, parvovirus B19; dengue and yellow fever; West Nile, Barmah Forest, Marburg, and Ebola viruses, and human herpesviruses; asymmetric periflexural exanthema of childhood; measles; rubella; enteroviruses; Lassa fever; and South American hemorrhagic fevers will be discussed.

Characterization of the cellular receptors for the South American hemorrhagic fever viruses Junin, Guanarito, and Machupo

The New World arenaviruses Junin, Machupo, and Guanarito are the causative agents of hemorrhagic fevers (HF) with high mortality in humans. The cellular receptor for Old World arenaviruses and one subgroup of the New World arenaviruses (Clade C) have been identified as α-dystroglycan (α-DG). In contrast, the receptor(s) of the South American HF viruses, which belong to the Clade B New World arenaviruses, are currently unknown. To begin to characterize the cellular receptors used by these pathogens, we generated recombinant retroviral pseudotypes with the glycoproteins of Guanarito, Junin, and Machupo. Infection with the South American HF viruses is independent of α-DG and functional receptors for Guanarito, Junin, and Machupo were found on most human cell types and cells derived from non-human primate and rodents. Guanarito, Junin, and Machupo share a common receptor, which is distinct from the receptor(s) used by the closely related non-pathogenic Clade B virus Amapari, and the genetically more distant Clade A and C New World arenaviruses. We show that the cellular receptor(s) for the South American HF viruses are proteins or protein-linked entities and that infection is not dependent on protein-linked N-glycans, O-glycans, or glycosaminoglycans.

Cells expressing the RING finger Z protein are resistant to arenavirus infection.

Arenaviruses include Lassa fever virus (LFV) and the South American hemorrhagic fever viruses. These viruses cause severe human disease, and they pose a threat as agents of bioterrorism. Arenaviruses are enveloped viruses with a bisegmented negative-strand RNA genome whose proteomic capability is limited to four polypeptides: nucleoprotein (NP); surface glycoprotein (GP), which is proteolytically processed into GP1 and GP2; polymerase (L); and a small (11-kDa) RING finger protein (Z). Our investigators have previously shown that Z has a strong inhibitory activity on RNA synthesis mediated by the polymerase of the prototypic arenavirus, lymphocytic choriomeningitis virus (LCMV). In this report we show that cells transduced with a replication-deficient recombinant adenovirus expressing Z (rAd-Z) are resistant to LCMV and LFV infection. Virus cell entry mediated by LCMV or LFV GP was not affected in rAd-Z-transduced cells, but both virus transcription and replication were strongly and specifically inhibited, which resulted in a dramatic reduction in production of infectious virus. These findings open new avenues for developing antiviral strategies to combat the highly pathogenic human arenaviruses, including LFV.

Junin virus vaccines.

Argentine hemorrhagic fever (AHF) was recognized as a new clinical entity from the richest farming region of Argentina in the 1950s (Arribalzaga 1955). The etiologic agent of this disease, Junin virus (JUN), was isolated in 1958 (Parodi et al. 1958; Piroski et al. 1959). JUN belongs to the Arenaviridae family, which includes other rodent-borne pathogens which are important causes of hemorrhagic fever in Africa, (Lassa) and South America (Machupo, Guanarito, and Sabia viruses). The Arenaviridae comprises at least 20 recognized members. Arenaviruses are enveloped RNA viruses, and are divided into two groups, with low level antigenic relatedness: the Old World group, and New World group or Tacaribe complex. Two antigenic subgroups were defined within the New World arenaviruses. JUN is contained into the first group, together with Amapari, Latino, Machupo and Tacaribe viruses. Phylogenetic analysis have shown that Old World and New World arenaviruses occupied two distinct clades, and that New World arenaviruses comprise three evolutionary lineages, named A, B, and C. JUN is contained in lineage B, together with the other three agents causing South American hemorrhagic fevers (Enria et al. 1999a).

RODENT-BORNE EMERGING VIRAL ZOONOSIS: Hemorrhagic Fevers and Hantavirus Infections in South America

Hantaviruses and arenaviruses are naturally occurring viruses of rodents. Four South American hemorrhagic fevers caused by arenaviruses have emerged in the last 5 decades. All have similar clinical manifestations, with a case-fatality rate as high as 15% to 30%. Hantavirus infections have been increasingly recognized in South America since the description in 1993 of Hantavirus pulmonary syndrome. Given the diversity of rodent species in the region, it can be foreseen that many other viruses will be discovered, and some of them will be causing human illnesses of high public health impact.

VIRAL HEMORRHAGIC FEVERS AND HANTAVIRUS INFECTIONS IN THE AMERICAS

Several arenaviruses and hantaviruses have been isolated in the Americas during the last 4 decades. These are rodent-borne viruses responsible for the South American hemorrhagic fevers (SAHF) and hantavirus pulmonary syndrome (HPS). Although rare, SAHF and HPS are serious illnesses with high mortality rates. Most viral isolates found in the Americas represent New World lineages of their respective viral families. Their presence in the Western hemisphere is likely ancient, their relationship with their rodent hosts is likely coevolutionary, and their recent detection forebodes the likelihood of detecting additional arena- and hantaviral species in the Americas.