Pathogenesis of lassa fever in cynomolgus macaques

<p>Abstract</p> <p>Background</p> <p>Lassa virus (LASV) infection causes an acute and sometimes fatal hemorrhagic disease in humans and nonhuman primates; however, little is known about the development of Lassa fever. Here, we performed a pilot study to begin to understand the progression of LASV infection in nonhuman primates.</p> <p>Methods</p> <p>Six cynomolgus monkeys were experimentally infected with LASV. Tissues from three animals were examined at an early- to mid-stage of disease and compared with tissues from three animals collected at terminal stages of disease.</p> <p>Results</p> <p>Dendritic cells were identified as a prominent target of LASV infection in a variety of tissues in all animals at day 7 while Kupffer cells, hepatocytes, adrenal cortical cells, and endothelial cells were more frequently infected with LASV in tissues of terminal animals (days 13.5-17). Meningoencephalitis and neuronal necrosis were noteworthy findings in terminal animals. Evidence of coagulopathy was noted; however, the degree of fibrin deposition in tissues was less prominent than has been reported in other viral hemorrhagic fevers.</p> <p>Conclusion</p> <p>The sequence of pathogenic events identified in this study begins to shed light on the development of disease processes during Lassa fever and also may provide new targets for rational prophylactic and chemotherapeutic interventions.</p

Effective Post-Exposure Treatment of Ebola Infection

Ebola viruses are highly lethal human pathogens that have received considerable attention in recent years due to an increasing re-emergence in Central Africa and a potential for use as a biological weapon. There is no vaccine or treatment licensed for human use. In the past, however, important advances have been made in developing preventive vaccines that are protective in animal models. In this regard, we showed that a single injection of a live-attenuated recombinant vesicular stomatitis virus vector expressing the Ebola virus glycoprotein completely protected rodents and nonhuman primates from lethal Ebola challenge. In contrast, progress in developing therapeutic interventions against Ebola virus infections has been much slower and there is clearly an urgent need to develop effective post-exposure strategies to respond to future outbreaks and acts of bioterrorism, as well as to treat laboratory exposures. Here we tested the efficacy of the vesicular stomatitis virus-based Ebola vaccine vector in post-exposure treatment in three relevant animal models. In the guinea pig and mouse models it was possible to protect 50% and 100% of the animals, respectively, following treatment as late as 24 h after lethal challenge. More important, four out of eight rhesus macaques were protected if treated 20 to 30 min following an otherwise uniformly lethal infection. Currently, this approach provides the most effective post-exposure treatment strategy for Ebola infections and is particularly suited for use in accidentally exposed individuals and in the control of secondary transmission during naturally occurring outbreaks or deliberate release

Vesicular Stomatitis Virus-Based Ebola Vaccine Is Well-Tolerated and Protects Immunocompromised Nonhuman Primates

Ebola virus (EBOV) is a significant human pathogen that presents a public health concern as an emerging/re-emerging virus and as a potential biological weapon. Substantial progress has been made over the last decade in developing candidate preventive vaccines that can protect nonhuman primates against EBOV. Among these prospects, a vaccine based on recombinant vesicular stomatitis virus (VSV) is particularly robust, as it can also confer protection when administered as a postexposure treatment. A concern that has been raised regarding the replication-competent VSV vectors that express EBOV glycoproteins is how these vectors would be tolerated by individuals with altered or compromised immune systems such as patients infected with HIV. This is especially important as all EBOV outbreaks to date have occurred in areas of Central and Western Africa with high HIV incidence rates in the population. In order to address this concern, we evaluated the safety of the recombinant VSV vector expressing the Zaire ebolavirus glycoprotein (VSVΔG/ZEBOVGP) in six rhesus macaques infected with simian-human immunodeficiency virus (SHIV). All six animals showed no evidence of illness associated with the VSVΔG/ZEBOVGP vaccine, suggesting that this vaccine may be safe in immunocompromised populations. While one goal of the study was to evaluate the safety of the candidate vaccine platform, it was also of interest to determine if altered immune status would affect vaccine efficacy. The vaccine protected 4 of 6 SHIV-infected macaques from death following ZEBOV challenge. Evaluation of CD4+ T cells in all animals showed that the animals that succumbed to lethal ZEBOV challenge had the lowest CD4+ counts, suggesting that CD4+ T cells may play a role in mediating protection against ZEBOV

Development of an Acute and Highly Pathogenic Nonhuman Primate Model of Nipah Virus Infection

Nipah virus (NiV) is an enigmatic emerging pathogen that causes severe and often fatal neurologic and/or respiratory disease in both animals and humans. Amongst people, case fatality rates range between 40 and 75 percent and there are no vaccines or treatments approved for human use. Guinea pigs, hamsters, cats, ferrets, pigs and most recently squirrel monkeys (New World monkey) have been evaluated as animal models of human NiV infection, and with the exception of the ferret, no model recapitulates all aspects of NiV-mediated disease seen in humans. To identify a more viable nonhuman primate (NHP) model, we examined the pathogenesis of NiV in African green monkeys (AGM). Exposure of eight monkeys to NiV produced a severe systemic infection in all eight animals with seven of the animals succumbing to infection. Viral RNA was detected in the plasma of challenged animals and occurred in two of three subjects as a peak between days 7 and 21, providing the first clear demonstration of plasma-associated viremia in NiV experimentally infected animals and suggested a progressive infection that seeded multiple organs simultaneously from the initial site of virus replication. Unlike the cat, hamster and squirrel monkey models of NiV infection, severe respiratory pathology, neurological disease and generalized vasculitis all manifested in NiV-infected AGMs, providing an accurate reflection of what is observed in NiV-infected humans. Our findings demonstrate the first consistent and highly pathogenic NHP model of NiV infection, providing a new and critical platform in the evaluation and licensure of either passive and active immunization or therapeutic strategies for human use

Proteolytic Processing of the Ebola Virus Glycoprotein Is Not Critical for Ebola Virus Replication in Nonhuman Primates

Enveloped viruses often require cleavage of a surface glycoprotein by a cellular endoprotease such as furin for infectivity and virulence. Previously, we showed that Ebola virus glycoprotein does not require the furin cleavage motif for virus replication in cell culture. Here, we show that there are no appreciable differences in disease progression, hematology, serum biochemistry, virus titers, or lethality in nonhuman primates infected with an Ebola virus lacking the furin recognition sequence compared to those infected with wild-type virus. We conclude that glycoprotein cleavage by subtilisin-like endoproteases is not critical for Ebola virus infectivity and virulence in nonhuman primates

The temporal program of peripheral blood gene expression in the response of nonhuman primates to Ebola hemorrhagic fever

Background: Infection with Ebola virus (EBOV) causes a fulminant and often fatal hemorrhagic fever. In order to improve our understanding of EBOV pathogenesis and EBOV-host interactions, we examined the molecular features of EBOV infection in vivo. Results: Using high-density cDNA microarrays, we analyzed genome-wide host expression patterns in sequential blood samples from nonhuman primates infected with EBOV. The temporal program of gene expression was strikingly similar between animals. Of particular interest were features of the data that reflect the interferon response, cytokine signaling, and apoptosis. Transcript levels for tumor necrosis factor-α converting enzyme (TACE)/α-disintegrin and metalloproteinase (ADAM)-17 increased during days 4 to 6 after infection. In addition, the serum concentration of cleaved Ebola glycoprotein (GP2 delta) was elevated in late-stage EBOV infected animals. Of note, we were able to detect changes in gene expression of more than 300 genes before symptoms appeared. Conclusion: These results provide the first genome-wide ex vivo analysis of the host response to systemic filovirus infection and disease. These data may elucidate mechanisms of viral pathogenesis and host defense, and may suggest targets for diagnostic and therapeutic development

Single-Injection Vaccine Protects Nonhuman Primates against Infection with Marburg Virus and Three Species of Ebola Virus▿

The filoviruses Marburg virus and Ebola virus cause severe hemorrhagic fever with high mortality in humans and nonhuman primates. Among the most promising filovirus vaccines under development is a system based on recombinant vesicular stomatitis virus (VSV) that expresses a single filovirus glycoprotein (GP) in place of the VSV glycoprotein (G). Here, we performed a proof-of-concept study in order to determine the potential of having one single-injection vaccine capable of protecting nonhuman primates against Sudan ebolavirus (SEBOV), Zaire ebolavirus (ZEBOV), Cote d'Ivoire ebolavirus (CIEBOV), and Marburgvirus (MARV). In this study, 11 cynomolgus monkeys were vaccinated with a blended vaccine consisting of equal parts of the vaccine vectors VSVΔG/SEBOVGP, VSVΔG/ZEBOVGP, and VSVΔG/MARVGP. Four weeks later, three of these animals were challenged with MARV, three with CIEBOV, three with ZEBOV, and two with SEBOV. Three control animals were vaccinated with VSV vectors encoding a nonfilovirus GP and challenged with SEBOV, ZEBOV, and MARV, respectively, and five unvaccinated control animals were challenged with CIEBOV. Importantly, none of the macaques vaccinated with the blended vaccine succumbed to a filovirus challenge. As expected, an experimental control animal vaccinated with VSVΔG/ZEBOVGP and challenged with SEBOV succumbed, as did the positive controls challenged with SEBOV, ZEBOV, and MARV, respectively. All five control animals challenged with CIEBOV became severely ill, and three of the animals succumbed on days 12, 12, and 14, respectively. The two animals that survived CIEBOV infection were protected from subsequent challenge with either SEBOV or ZEBOV, suggesting that immunity to CIEBOV may be protective against other species of Ebola virus. In conclusion, we developed an immunization scheme based on a single-injection vaccine that protects nonhuman primates against lethal challenge with representative strains of all human pathogenic filovirus species

Cross-Protection against Marburg Virus Strains by Using a Live, Attenuated Recombinant Vaccine

Marburg virus (MARV) has been associated with sporadic episodes of hemorrhagic fever, including a recent highly publicized outbreak in Angola that produced severe disease and significant mortality in infected patients. MARV is also considered to have potential as a biological weapon. Recently, we reported the development of a promising attenuated, replication-competent vaccine against MARV based on recombinant vesicular stomatitis virus (VSV) expressing the glycoprotein of the Musoke strain of MARV (VSVΔG/MARVGP-Musoke). We used this vaccine to demonstrate complete protection of cynomolgus monkeys against a homologous MARV challenge. While these results are highly encouraging, an effective vaccine would need to confer protection against all relevant strains of MARV. Here, we evaluated the protective efficacy of the VSVΔG/MARVGP-Musoke vaccine against two heterologous MARV strains, the seemingly more pathogenic Angola strain and the more distantly related Ravn strain. In this study, seven cynomolgus monkeys were vaccinated with the VSVΔG/MARVGP-Musoke vector. Three of these animals were challenged with the Angola strain, three with the Ravn strain, and a single animal with the Musoke strain of MARV. Two animals served as controls and were each injected with a nonspecific VSV vector; these controls were challenged with the Angola and Ravn strains, respectively. Both controls succumbed to challenge by day 8. However, none of the specifically vaccinated animals showed any evidence of illness either from the vaccination or from the MARV challenges and all of these animals survived. These data suggest that the VSVΔG/MARVGP-Musoke vaccine should be sufficient to protect against all known MARV strains

The temporal program of peripheral blood gene expression in the response of non-human primates to Ebola hemorrhagic fever

Background: Infection with Ebola virus (EBOV) causes a fulminant and often fatal hemorrhagic fever. In order to improve our understanding of EBOV pathogenesis and EBOV-host interactions, we examined the molecular features of EBOV infection in vivo. Results: Using high-density cDNA microarrays, we analyzed genome-wide host expression patterns in sequential blood samples from nonhuman primates infected with EBOV. The temporal program of gene expression was strikingly similar between animals. Of particular interest were features of the data that reflect the interferon response, cytokine signaling, and apoptosis. Transcript levels for tumor necrosis factor-α converting enzyme (TACE)/α-disintegrin and metalloproteinase (ADAM)-17 increased during days 4 to 6 after infection. In addition, the serum concentration of cleaved Ebola glycoprotein (GP2 delta) was elevated in late-stage EBOV infected animals. Of note, we were able to detect changes in gene expression of more than 300 genes before symptoms appeared. Conclusion: These results provide the first genome-wide ex vivo analysis of the host response to systemic filovirus infection and disease. These data may elucidate mechanisms of viral pathogenesis and host defense, and may suggest targets for diagnostic and therapeutic development.National Institutes of Health (U.S.) (NIH grant AI54922)United States. Defense Advanced Research Projects Agency (DARPA grant N65236-99-1-5428)Horn FoundationHoward Hughes Medical InstituteUnited States. Defense Threat Reduction AgencyU.S. Army Medical Research Institute of Infectious DiseasesChemical and Biological Defense Program (U.S.)United States. Army Medical Research and Materiel Command (project number 02-4-4J-081