Disease In Nonhuman Primates Research Articles

Modelling the Effect of Post-mortem Contact on the Spread of Ebola with Quarantine As an Intervention

Ebola virus disease (EVD) is a severe, often fatal disease in humans and other non-human primates caused by infection with any of the four identified Ebola virus species of the family Filoviridae. This paper develops the SEIR and the SEIHDR epidemic models that investigate the effects of the ante-mortem contact and post-mortem contact on the spread of the disease. The reproduction number of the models are determined. The equilibria and conditions for the existence of the equilibria are also determined. The models are solved numerically and the numerical simulations implemented to elucidate various scenarios. The results of the models are then compared to WHO data of confirmed cases for the 2014 Ebola outbreak in Liberia. It is observed that the SEIHDR model agrees better with the data than the SEIR model. Moreover, a new model, the SEIQDR model (a modification of the SEIHDR Model) is formulated which incorporates quarantine as an intervention. Again, this SEIQDR model is compared to the WHO data of confirmed cases for the 2014 Ebola outbreak in Liberia. The results of the SEIQDR model is found to agree better than those of the other models especially in respect of the latter stages of the disease outbreak. Finally, the effect of vaccination on both the SEIHDR and the SEIQDR models is investigated. Different rates of vaccination using numerical simulations in order to predict the effect of vaccination on the infected individuals over time is also discussed. The SEIQDR model with vaccination indicates a lower threshold which should not be less than 25\% as compared to the SEIHDR model for which vaccination should not be less than 65\%. It is observed that vaccination as an additional strategy helps to control the disease more effectively.

Accumulating necrotic cell debris and damaged mitochondria contribute to sepsis-like syndrome during late-phase tularemia.

Abstract Infection with Francisella tularensis SchuS4 causes a severe pulmonary disease in mice, non-human primates, and patients known as tularemia. F. tularensis subverts host cell death programs to facilitate their unfettered replication within macrophages and other cell types, a common feature of early respiratory infection (≤ 72 h). The mechanism(s) by which the complex process of cell death is initiated remains incompletely-defined as does the impact of F. tularensis on mitochondria, the host cell’s organellar ‘canary in a coal mine’. Herein we reveal that host cells undergo Bax-dependent mitochondrial apoptosis via the intrinsic pathway. These events encompass: i) decreased mitochondrial membrane potential and consequent mitochondrial oncosis, ii) increased mitochondrial superoxide generation, and iii) release of damaged mitochondria and other damage-associated molecular patterns into the lung parenchyma, which contributes to a sepsis-like syndrome. Notably, host recognition of and response to accumulating necrotic cell debris elicits the TNF, IL-1β, IL-6, IL-12p70, and IFN-γ which are absent during early infection, but present during late-phase septic disease (≥ 72 h).

Status of vaccine research and development for Campylobacter jejuni

Campylobacter jejuni is one of the leading causes of bacterial diarrhea worldwide and is associated with a number of sequelae, including Guillain–Barre Syndrome, reactive arthritis, irritable bowel syndrome and growth stunting/malnutrition. Vaccine development against C. jejuni is complicated by its antigenic diversity, a lack of small animal models, and a poor understanding of the bacterium's pathogenesis. Vaccine approaches have been limited to recombinant proteins, none of which have advanced beyond Phase I testing. Genomic analyses have revealed the presence of a polysaccharide capsule on C. jejuni. Given the success of capsule-conjugate vaccines for other mucosal pathogens of global importance, efforts to evaluate this established approach for C. jejuni are also being pursued. A prototypical capsule-conjugate vaccine has demonstrated efficacy against diarrheal disease in non-human primates and is currently in Phase I testing. In addition to proof of concept studies, more data on the global prevalence of capsular types, and a better understanding of the acute and chronic consequences of C. jejuni are needed to inform investments for a globally relevant vaccine.

Specific Detection of Two Divergent Simian Arteriviruses Using RNAscope In Situ Hybridization.

Simian hemorrhagic fever (SHF) is an often lethal disease of Asian macaques. Simian hemorrhagic fever virus (SHFV) is one of at least three distinct simian arteriviruses that can cause SHF, but pathogenesis studies using modern methods have been scarce. Even seemingly straightforward studies, such as examining viral tissue and cell tropism in vivo, have been difficult to conduct due to the absence of standardized SHFV-specific reagents. Here we report the establishment of an in situ hybridization assay for the detection of SHFV and distantly related Kibale red colobus virus 1 (KRCV-1) RNA in cell culture. In addition, we detected SHFV RNA in formalin-fixed, paraffin-embedded tissues from an infected rhesus monkey (Macaca mulatta). The assay is easily performed and can clearly distinguish between SHFV and KRCV-1. Thus, if further developed, this assay may be useful during future studies evaluating the mechanisms by which a simian arterivirus with a restricted cell tropism can cause a lethal nonhuman primate disease similar in clinical presentation to human viral hemorrhagic fevers.

Designing Efficacious Vesicular Stomatitis Virus-Vectored Vaccines Against Ebola Virus.

Infection with the Ebola virus (EBOV) causes an aggressive hemorrhagic disease in humans and nonhuman primates. Traditional approaches, such as vaccination with inactivated virion preparations, have had limited efficacy, whereas immunization with live-attenuated EBOV is not feasible due to the highly lethal nature of the pathogen. This has necessitated the development of other approaches towards an effective EBOV vaccine. Over the past decade, recombinant viruses expressing the EBOV glycoprotein (GP) have constituted the most promising platforms, as evidenced by their ability to protect naïve nonhuman primates from a lethal EBOV challenge. The vesicular stomatitis virus (VSV) is one such vector and is currently progressing through the clinical pipeline. This chapter presents methodologies for the design, cloning, rescue, and preparation of live, recombinant VSV vaccines expressing GP for research purposes.

Abstract CN04-03: DNA vaccines as treatment for prostate cancer - understanding mechanisms of resistance

Abstract Anti-tumor vaccines have demonstrated clinical benefit in patients with advanced prostate cancer, and the first vaccine approved by FDA as a treatment for human cancer was a vaccine targeting the prostate tumor antigen prostatic acid phosphatase (PAP). Plasmid DNA, as another method of antigen delivery, offers significant potential safety and manufacturing advantages over other immunization approaches. DNA vaccines have been approved by USDA as treatments for diseases in fish, horses, dogs, and non-human primates. However, DNA vaccines have generally been perceived as less immunologically potent than other methods of antigen delivery in human clinical trials, leading to efforts to increase their immunogenicity. Most of these efforts have focused on increasing the delivery of vaccine to antigen-presenting cells using different carriers or by electroporation. Our group has evaluated repetitive immunization with plasmid DNA vaccines in rodent models, and has translated this approach to early clinical trials in men with low-volume recurrent prostate cancer with a DNA vaccine encoding PAP. We have demonstrated that persistent Th1-biased PAP-specific T-cell immune responses can be elicited in men with prostate cancer. Moreover, the development of persistent immunity appeared to be associated with favorable changes in tumor growth rate. A randomized phase II trial using this approach is currently ongoing in men with minimal residual prostate cancer to determine whether immunization can delay the time to metastatic disease progression (NCT01341652). In an effort to further improve the immunogenicity of DNA vaccines, we have explored methods to increase antigen presentation by encoding epitopes with increased MHC class I affinity or by increasing the duration of target antigen expression. While such “optimized” vaccines elicited increased numbers of antigen-specific CD8+ T cells, these T cells had less anti-tumor activity in vivo. We determined that immunization with plasmid DNA elicited antigen-specific IFNγ-secreting T cells that led to increased expression of PD-L1 on antigen-expressing tumor cells. However, immunization with the vaccine encoding high affinity epitopes resulted in antigen-specific CD8+ T cells with higher PD-1 expression. In the presence of PD-L1-expressing tumors these CD8+ were less effective. The anti-tumor activity was restored in the presence of PD-1 or PD-L1 blockade. Conversely, immunization with plasmid DNA engineered to increase the duration of antigen expression led to CD8+ T cells with normal PD-1 expression but higher LAG-3 expression. The anti-tumor activity was similarly restored in the presence of LAG-3 blockade. We are currently studying the mechanisms by which different T cell regulatory ligands are expressed following immunization. Notwithstanding, such findings suggest that subtle differences in the method of immunization can affect these different regulatory mechanisms, and hence the choice of optimal combination strategies may depend on identifying the specific mechanisms of resistance to immunization. In the case of patients immunized with a DNA vaccine encoding PAP, we have recently found that PD-L1 expression on circulating tumor (CD45-EpCAM+) cells was increased following immunization, analogous to our murine studies. In addition, PAP-specific secretion of IFNγ and granzyme B by T cells obtained from patients after immunization was increased in the presence of PD-1/PD-L1 blockade. Finally, using a trans vivo delayed type hypersensitivity (tvDTH) assay, we observed that PAP-specific immune responses in T cells obtained from patients after immunization with a DNA vaccine encoding PAP could be “uncovered” with PD-1 blockade but not blockade with anti-LAG-3 or anti-TIM-3. Collectively, these findings suggest that the PD-1/PD-L1 pathway and mechanism of resistance is most relevant to a DNA vaccine encoding PAP, and the efficacy of this vaccine might be augmented with PD-1 blockade. A clinical trial testing this approach in men with advanced prostate cancer, using this DNA vaccine and pembrolizumab, delivered either concurrently or sequentially, is currently underway (NCT02499835). Citation Format: Brian T. Rekoske, Viswa T. Colluru, Douglas G. McNeel. DNA vaccines as treatment for prostate cancer - understanding mechanisms of resistance. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr CN04-03.

De novo transcriptome reconstruction and annotation of the Egyptian rousette bat.

BackgroundThe Egyptian Rousette bat (Rousettus aegyptiacus), a common fruit bat species found throughout Africa and the Middle East, was recently identified as a natural reservoir host of Marburg virus. With Ebola virus, Marburg virus is a member of the family Filoviridae that causes severe hemorrhagic fever disease in humans and nonhuman primates, but results in little to no pathological consequences in bats. Understanding host-pathogen interactions within reservoir host species and how it differs from hosts that experience severe disease is an important aspect of evaluating viral pathogenesis and developing novel therapeutics and methods of prevention.ResultsProgress in studying bat reservoir host responses to virus infection is hampered by the lack of host-specific reagents required for immunological studies. In order to establish a basis for the design of reagents, we sequenced, assembled, and annotated the R. aegyptiacus transcriptome. We performed de novo transcriptome assembly using deep RNA sequencing data from 11 distinct tissues from one male and one female bat. We observed high similarity between this transcriptome and those available from other bat species. Gene expression analysis demonstrated clustering of expression profiles by tissue, where we also identified enrichment of tissue-specific gene ontology terms. In addition, we identified and experimentally validated the expression of novel coding transcripts that may be specific to this species.ConclusionWe comprehensively characterized the R. aegyptiacus transcriptome de novo. This transcriptome will be an important resource for understanding bat immunology, physiology, disease pathogenesis, and virus transmission.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2124-x) contains supplementary material, which is available to authorized users.

Role of surface charge in determining the biological effects of CdSe/ZnS quantum dots.

The growing potential of quantum dots (QDs) in biomedical applications has provoked the urgent need to thoroughly address their interaction with biological systems. However, only limited studies have been performed to explore the effects of surface charge on the biological behaviors of QDs. In the present study, three commercially available QDs with different surface coatings were used to systematically evaluate the effects of surface charge on the cellular uptake, cytotoxicity, and in vivo biodistribution of QDs. Our results demonstrated that charged QDs entered both cancer cells and macrophages more efficiently than neutral ones, while negative QDs internalized mostly. Upon entry into cells, QDs were localized in different subcellular compartments (eg, cytoplasm and lysosomes) depending on the surface charge. Interestingly, inconsistent with the result of internalization, positive QDs but not negative QDs exhibited severe cytotoxicity, which was likely due to their disruption of cell membrane integrity, and production of reactive oxygen species. Biodistribution studies demonstrated that negative and neutral QDs preferentially distributed in the liver and the spleen, whereas positive QDs mainly deposited in the kidney with obvious uptake in the brain. In general, surface charge plays crucial roles in determining the biological interactions of QDs.

Impact Of the 2014/2015 Ebola Virus Disease Epidemic on Malaria Cases and Control in Some West African Countries. A Mini Review

Ebola hemorrhagic Disease (EHD) is a severe, often fatal disease in humans and nonhuman primates (monkeys, gorillas, and chimpanzees) that has appeared sporadically since its initial recognition in 1976. Malaria is a life-threatening disease caused by Plasmodium that is transmitted to people through the bites of infected mosquitoes. Increased malaria prevention and control measures are dramatically reducing the malaria burden in many places. Disease outbreaks and catastrophes can affect countries at any time, causing substantial human suffering, deaths, and economic losses. Ebola virus disease outbreak in Western African countries; Guinea, Liberia and Sierra Leone caused the cases of malaria to increase and death rate from malaria also increased in that region. Capacity health systems and external health related aid to Guinea, Liberia and Sierra Leone increased in combating the disease neglecting malaria. The lack of balanced investment in the health systems contributed to the challenges of controlling and combating malaria cases. Ebola caused a spike in malaria cases in the three West African countries heavily affected by the disease which have undermined previous efforts to curb malaria. Thousands of malaria cases went untreated in Guinea in 2014 as people with symptoms avoided health clinics for the fear of being sent into isolation at Ebola treatment centers as the early symptoms of malaria mimic those of Ebola virus disease. Ebola overwhelmed health care systems in the year 2014, making adequate care for malaria impossible and disrupted the distribution of bed nets in the affected areas. Rather than label one disease as more important than the others, equal attention or near equal attention should be given to all the diseases of the public health concern to lower the death tolls for all of them. Malaria control efforts must be kept on track during an Ebola epidemic so that progress made in malaria control is not jeopardized.

Outbreak of Tuberculosis in a Colony of Rhesus Monkeys (Macaca mulatta) after Possible Indirect Contact with a Human TB Patient

Simian tuberculosis is one of the most important bacterial diseases of non-human primates. Outbreaks of tuberculosis have been reported in primate colonies almost as long as these animals have been used experimentally or kept in zoological gardens. Significant progress has been made in reducing the incidence of tuberculosis in captive non-human primates, but despite reasonable precautions, outbreaks continue to occur. The most relevant reason is the high incidence of tuberculosis (TB) amongst the human population, in which tuberculosis is regarded as an important re-emerging disease. Furthermore, many non-human primate species originate from countries with a high burden of human TB. Therefore, Mycobacterium tuberculosis remains a significant threat in animals imported from countries with high rates of human infection. We report an outbreak of tuberculosis among a group of rhesus monkeys (Macaca mulatta) living in a closed, long-term colony. The outbreak coincided with reactivation of a TB infection in a co-worker who never had direct access to the animal house or laboratories. Eleven of 26 rhesus monkeys developed classical chronic active tuberculosis with typical caseous granulomata of varying size within different organs. The main organ system involved was the lung, suggesting an aerosol route of infection. Such an outbreak has significant economic consequences due to animal loss, disruption of research and costs related to disease control. Precautionary measures must be improved in order to avoid TB in non-human primate colonies.

Modeling the Disease Course of Zaire ebolavirus Infection in the Outbred Guinea Pig.

Rodent models that accurately reflect human filovirus infection are needed as early screens for medical countermeasures. Prior work in rodents with the Zaire species of Ebola virus (ZEBOV) primarily used inbred mice and guinea pigs to model disease. However, these inbred species do not show some of the important features of primate ZEBOV infection, most notably, coagulation abnormalities. Thirty-six outbred guinea pigs were infected with guinea pig-adapted ZEBOV and examined sequentially over an 8-day period to investigate the pathologic events that lead to death. Features of disease in ZEBOV-infected outbred guinea pigs were largely consistent with disease in humans and nonhuman primates and included early infection of macrophages and dendritiform cells, apoptosis of bystander lymphocytes, and increases in levels of proinflammatory cytokines. Most importantly, dysregulation of circulating levels of fibrinogen, protein C activity, and antifibrinolytic proteins and deposition of fibrin in tissues demonstrated both biochemical and microscopic evidence of disseminated intravascular coagulation. These findings suggest that the outbred guinea pig model recapitulates ZEBOV infection of primates better than inbred rodent models, is useful for dissecting key events in the pathogenesis of ZEBOV, and is useful for evaluating candidate interventions prior to assessment in primates.

Particle-to-PFU ratio of Ebola virus influences disease course and survival in cynomolgus macaques.

This study addresses the role of Ebola virus (EBOV) specific infectivity in virulence. Filoviruses are highly lethal, enveloped, single-stranded negative-sense RNA viruses that can cause hemorrhagic fever. No approved vaccines or therapies exist for filovirus infections, and infectious virus must be handled in maximum containment. Efficacy testing of countermeasures, in addition to investigations of pathogenicity and immune response, often requires a well-characterized animal model. For EBOV, an obstacle in performing accurate disease modeling is a poor understanding of what constitutes an infectious dose in animal models. One well-recognized consequence of viral passage in cell culture is a change in specific infectivity, often measured as a particle-to-PFU ratio. Here, we report that serial passages of EBOV in cell culture resulted in a decrease in particle-to-PFU ratio. Notably, this correlated with decreased potency in a lethal cynomolgus macaque (Macaca fascicularis) model of infection; animals were infected with the same viral dose as determined by plaque assay, but animals that received more virus particles exhibited increased disease. This suggests that some particles are unable to form a plaque in a cell culture assay but are able to result in lethal disease in vivo. These results have a significant impact on how future studies are designed to model EBOV disease and test countermeasures. Ebola virus (EBOV) can cause severe hemorrhagic disease with a high case-fatality rate, and there are no approved vaccines or therapies. Specific infectivity can be considered the total number of viral particles per PFU, and its impact on disease is poorly understood. In stocks of most mammalian viruses, there are particles that are unable to complete an infectious cycle or unable to cause cell pathology in cultured cells. We asked if these particles cause disease in nonhuman primates by infecting monkeys with equal infectious doses of genetically identical stocks possessing either high or low specific infectivities. Interestingly, some particles that did not yield plaques in cell culture assays were able to result in lethal disease in vivo. Furthermore, the number of PFU needed to induce lethal disease in animals was very low. Our results have a significant impact on how future studies are designed to model EBOV disease and test countermeasures.

Are adaptive randomised trials or non-randomised studies the best way to address the Ebola outbreak in west Africa?

The Ebola outbreak that has devastated parts of west Africa represents an unprecedented challenge for research and ethics. Estimates from the past three decades emphasise that the present effort to contain the epidemic in the three most affected countries (Guinea, Liberia, and Sierra Leone) has been insufficient, with more than 24 900 cases and about 10 300 deaths, as of March 25, 2015. Faced with such an exceptional event and the urgent response it demands, the use of randomised controlled trials (RCT) for Ebola-related research might be both unethical and infeasible and that potential interventions should be assessed in non-randomised studies on the basis of compassionate use. However, non-randomised studies might not yield valid conclusions, leading to large residual uncertainty about how to interpret the results, and can also waste scarce intervention-related resources, making them profoundly unethical. Scientifically sound and rigorous study designs, such as adaptive RCTs, could provide the best way to reduce the time needed to develop new interventions and to obtain valid results on their efficacy and safety while preserving the application of ethical precepts. We present an overview of clinical studies registered at present at the four main international trial registries and provide a simulation on how adaptive RCTs can behave in this context, when mortality varies simultaneously in either the control or the experimental group.

Interaction between TIM-1 and NPC1 Is Important for Cellular Entry of Ebola Virus.

Multiple host molecules are known to be involved in the cellular entry of filoviruses, including Ebola virus (EBOV); T-cell immunoglobulin and mucin domain 1 (TIM-1) and Niemann-Pick C1 (NPC1) have been identified as attachment and fusion receptors, respectively. However, the molecular mechanisms underlying the entry process have not been fully understood. We found that TIM-1 and NPC1 colocalized and interacted in the intracellular vesicles where EBOV glycoprotein (GP)-mediated membrane fusion occurred. Interestingly, a TIM-1-specific monoclonal antibody (MAb), M224/1, prevented GP-mediated membrane fusion and also interfered with the binding of TIM-1 to NPC1, suggesting that the interaction between TIM-1 and NPC1 is important for filovirus membrane fusion. Moreover, MAb M224/1 efficiently inhibited the cellular entry of viruses from all known filovirus species. These data suggest a novel mechanism underlying filovirus membrane fusion and provide a potential cellular target for antiviral compounds that can be universally used against filovirus infections. Filoviruses, including Ebola and Marburg viruses, cause rapidly fatal diseases in humans and nonhuman primates. There are currently no approved vaccines or therapeutics for filovirus diseases. In general, the cellular entry step of viruses is one of the key mechanisms to develop antiviral strategies. However, the molecular mechanisms underlying the entry process of filoviruses have not been fully understood. In this study, we demonstrate that TIM-1 and NPC1, which serve as attachment and fusion receptors for filovirus entry, interact in the intracellular vesicles where Ebola virus GP-mediated membrane fusion occurs and that this interaction is important for filovirus infection. We found that filovirus infection and GP-mediated membrane fusion in cultured cells were remarkably suppressed by treatment with a TIM-1-specific monoclonal antibody that interfered with the interaction between TIM-1 and NPC1. Our data provide new insights for the development of antiviral compounds that can be universally used against filovirus infections.

Sustained Delivery of a Broadly Neutralizing Antibody in Nonhuman Primates Confers Long-Term Protection against Simian/Human Immunodeficiency Virus Infection.

Pathogen-specific neutralizing antibodies protect against many viral infections and can potentially prevent human immunodeficiency virus (HIV) transmission in humans. However, neutralizing antibodies have so far only been shown to protect nonhuman primates (NHP) against lentiviral infection when given shortly before challenge. Thus, the clinical utility and feasibility of passive antibody transfer to confer long-term protection against HIV-1 are still debated. Here, we investigate the potential of a broadly neutralizing HIV-1 antibody to provide long-term protection in a NHP model of HIV-1 infection. A human antibody was simianized to avoid immune rejection and used to sustain therapeutic levels for ∼5 months. Two months after the final antibody administration, animals were completely protected against viral challenge. These findings demonstrate the feasibility and potential of long-term passive antibody for protection against HIV-1 in humans and provide a model to test antibody therapies for other diseases in NHP. Antibodies against HIV are potential drugs that may be able to prevent HIV infection in humans. However, the long-term protective capacity of antibodies against HIV has not been assessed. Here, we repetitively administered a macaque version of a human anti-HIV antibody to monkeys, after which the antibody persisted in the blood for >5 months. Moreover, the antibody could be sustained at protective levels for 108 days, conferring protection 52 days after the last dose in a monkey model of HIV infection. Thus, passive antibody transfer can provide durable protection against infection by viruses that cause AIDS in primates.

A single phosphorodiamidate morpholino oligomer targeting VP24 protects rhesus monkeys against lethal Ebola virus infection.

ABSTRACTEbola viruses (EBOV) cause severe disease in humans and nonhuman primates with high mortality rates and continue to emerge in new geographic locations, including several countries in West Africa, the site of a large ongoing outbreak. Phosphorodiamidate morpholino oligomers (PMOs) are synthetic antisense molecules that are able to target mRNAs in a sequence-specific fashion and suppress translation through steric hindrance. We previously showed that the use of PMOs targeting a combination of VP35 and VP24 protected rhesus monkeys from lethal EBOV infection. Surprisingly, the present study revealed that a PMOplus compound targeting VP24 alone was sufficient to confer protection from lethal EBOV infection but that a PMOplus targeting VP35 alone resulted in no protection. This study further substantiates recent data demonstrating that VP24 may be a key virulence factor encoded by EBOV and suggests that VP24 is a promising target for the development of effective anti-EBOV countermeasures.

Progress in the Diagnosis and Control of Ebola Disease.

Ebola hemorrhagic fever is one of numerous viral hemorrhagic fevers. It is a severe, often fatal disease in humans and nonhuman primates (gorillas and chimpanzees). This article discusses the history of Ebola disease, already known routes of infection together with defining prevention methods and treatment trials. The importance of increasing awareness of the risk of disease among people who do not inhabit endemic regions is emphasized. This risk is associated especially with the increasing popularity of tourism to African countries, even to those where the virus is endemic. The research conducted over the years shows that three species of frugivorous bats are subjected to contamination by Ebola, but the infection is asymptomatic in them. It is believed that the saliva of these mammals and other body fluids may be a potential source of infection for primates and humans. In the laboratory, infection through small-particle aerosols has been demonstrated in primates, and airborne spread among humans is strongly suspected, although it has not yet been conclusively demonstrated. The importance of this route of transmission remains unclear. Poor hygienic conditions can aid the spread of the virus. These observations suggest approaches to the study of routes of transmission to and among humans.

Ebola: are we prepared for recurring infectious threats?

Ebola: are we prepared for recurring infectious threats?

The VP40 protein of Marburg virus exhibits impaired budding and increased sensitivity to human tetherin following mouse adaptation.

The Marburg virus VP40 protein is a viral matrix protein that spontaneously buds from cells. It also functions as an interferon (IFN) signaling antagonist by targeting Janus kinase 1 (JAK1). A previous study demonstrated that the VP40 protein of the Ravn strain of Marburg virus (Ravn virus [RAVV]) failed to block IFN signaling in mouse cells, whereas the mouse-adapted RAVV (maRAVV) VP40 acquired the ability to inhibit IFN responses in mouse cells. The increased IFN antagonist function of maRAVV VP40 mapped to residues 57 and 165, which were mutated during the mouse adaptation process. In the present study, we demonstrate that maRAVV VP40 lost the capacity to efficiently bud from human cell lines, despite the fact that both parental and maRAVV VP40s bud efficiently from mouse cell lines. The impaired budding in human cells corresponds with the appearance of protrusions on the surface of maRAVV VP40-expressing Huh7 cells and with an increased sensitivity of maRAVV VP40 to restriction by human tetherin but not mouse tetherin. However, transfer of the human tetherin cytoplasmic tail to mouse tetherin restored restriction of maRAVV VP40. Residues 57 and 165 were demonstrated to contribute to the failure of maRAVV VP40 to bud from human cells, and residue 57 was demonstrated to alter VP40 oligomerization, as assessed by coprecipitation assay, and to determine sensitivity to human tetherin. This suggests that RAVV VP40 acquired, during adaptation to mice, changes in its oligomerization potential that enhanced IFN antagonist function. However, this new capacity impaired RAVV VP40 budding from human cells. Filoviruses, which include Marburg viruses and Ebola viruses, are zoonotic pathogens that cause severe disease in humans and nonhuman primates but do not cause similar disease in wild-type laboratory strains of mice unless first adapted to these animals. Although mouse adaptation has been used as a method to develop small animal models of pathogenesis, the molecular determinants associated with filovirus mouse adaptation are poorly understood. Our study demonstrates how genetic changes that accrued during mouse adaptation of the Ravn strain of Marburg virus have impacted the budding function of the viral VP40 matrix protein. Strikingly, we find impairment of mouse-adapted VP40 budding function in human but not mouse cell lines, and we correlate the impairment with an increased sensitivity of VP40 to restriction by human but not mouse tetherin and with changes in VP40 oligomerization. These data suggest that there are functional costs associated with filovirus adaptation to new hosts and implicate tetherin as a filovirus host restriction factor.

Reversion of advanced Ebola virus disease in nonhuman primates with ZMapp.

Without an approved vaccine or treatment, Ebola outbreak management has been limited to palliative care and barrier methods to prevent transmission. These approaches, however, have yet to end the 2014 outbreak of Ebola after its prolonged presence in West Africa. Here we show that a combination of monoclonal antibodies (ZMapp™), optimized from two previous antibody cocktails, is able to rescue 100% of rhesus macaques when treatment is initiated up to 5 days post-challenge. High fever, viremia, and abnormalities in blood count and chemistry were evident in many animals before ZMapp™ intervention. Advanced disease, as indicated by elevated liver enzymes, mucosal hemorrhages and generalized petechia could be reversed, leading to full recovery. ELISA and neutralizing antibody assays indicate that ZMapp™ is cross-reactive with the Guinean variant of Ebola. ZMapp™ currently exceeds all previous descriptions of efficacy with other therapeutics, and results warrant further development of this cocktail for clinical use.