Urban habituation, ecological connectivity and epidemic dampening: the emergence of Hendra virus from flying foxes (Pteropus spp.)

Anthropogenic environmental change is often implicated in the emergence of new zoonoses from wildlife; however, there is little mechanistic understanding of these causal links. Here, we examine the transmission dynamics of an emerging zoonotic paramyxovirus, Hendra virus (HeV), in its endemic host, Australian Pteropus bats (fruit bats or flying foxes). HeV is a biosecurity level 4 (BSL-4) pathogen, with a high case-fatality rate in humans and horses. With models parametrized from field and laboratory data, we explore a set of probable contributory mechanisms that explain the spatial and temporal pattern of HeV emergence; including urban habituation and decreased migration-two widely observed changes in flying fox ecology that result from anthropogenic transformation of bat habitat in Australia. Urban habituation increases the number of flying foxes in contact with human and domestic animal populations, and our models suggest that, in addition, decreased bat migratory behaviour could lead to a decline in population immunity, giving rise to more intense outbreaks after local viral reintroduction. Ten of the 14 known HeV outbreaks occurred near urbanized or sedentary flying fox populations, supporting these predictions. We also demonstrate that by incorporating waning maternal immunity into our models, the peak modelled prevalence coincides with the peak annual spill-over hazard for HeV. These results provide the first detailed mechanistic framework for understanding the sporadic temporal pattern of HeV emergence, and of the urban/peri-urban distribution of HeV outbreaks in horses and people.     

Hendra virus infection dynamics in Australian fruit bats

Hendra virus is a recently emerged zoonotic agent in Australia. Since first described in 1994, the virus has spilled from its wildlife reservoir (pteropid fruit bats, or 'flying foxes') on multiple occasions causing equine and human fatalities. We undertook a three-year longitudinal study to detect virus in the urine of free-living flying foxes (a putative route of excretion) to investigate Hendra virus infection dynamics. Pooled urine samples collected off plastic sheets placed beneath roosting flying foxes were screened for Hendra virus genome by quantitative RT-PCR, using a set of primers and probe derived from the matrix protein gene. A total of 1672 pooled urine samples from 67 sampling events was collected and tested between 1 July 2008 and 30 June 2011, with 25% of sampling events and 2.5% of urine samples yielding detections. The proportion of positive samples was statistically associated with year and location. The findings indicate that Hendra virus excretion occurs periodically rather than continuously, and in geographically disparate flying fox populations in the state of Queensland. The lack of any detection in the Northern Territory suggests prevalence may vary across the range of flying foxes in Australia. Finally, our findings suggest that flying foxes can excrete virus at any time of year, and that the apparent seasonal clustering of Hendra virus incidents in horses and associated humans (70% have occurred June to October) reflects factors other than the presence of virus. Identification of these factors will strengthen risk minimization strategies for horses and ultimately humans.     

Hendra Virus Spillover is a Bimodal System Driven by Climatic Factors

Understanding environmental factors driving spatiotemporal patterns of disease can improve risk mitigation strategies. Hendra virus (HeV), discovered in Australia in 1994, spills over from bats (Pteropus sp.) to horses and thence to humans. Below latitude ? 22°, almost all spillover events to horses occur during winter, and above this latitude spillover is aseasonal. We generated a statistical model of environmental drivers of HeV spillover per month. The model reproduced the spatiotemporal pattern of spillover risk between 1994 and 2015. The model was generated with an ensemble of methods for presence–absence data (boosted regression trees, random forests and logistic regression). Presences were the locations of horse cases, and absences per spatial unit (2.7 × 2.7 km pixels without spillover) were sampled with the horse census of Queensland and New South Wales. The most influential factors indicate that spillover is associated with both cold-dry and wet conditions. Bimodal responses to several variables suggest spillover involves two systems: one above and one below a latitudinal area close to ? 22°. Northern spillovers are associated with cold-dry and wet conditions, and southern with cold-dry conditions. Biologically, these patterns could be driven by immune or behavioural changes in response to food shortage in bats and horse husbandry. Future research should look for differences in these traits between seasons in the two latitudinal regions. Based on the predicted risk patterns by latitude, we recommend enhanced preventive management for horses from March to November below latitude 22° south.     

Reproduction and nutritional stress are risk factors for Hendra virus infection in little red flying foxes (Pteropus scapulatus)

Hendra virus (HeV) is a lethal paramyxovirus which emerged in humans in 1994. Poor understanding of HeV dynamics in Pteropus spp. (flying fox or fruit bat) reservoir hosts has limited our ability to determine factors driving its emergence. We initiated a longitudinal field study of HeV in little red flying foxes (LRFF; Pteropus scapulatus) and examined individual and population risk factors for infection, to determine probable modes of intraspecific transmission. We also investigated whether seasonal changes in host behaviour, physiology and demography affect host-pathogen dynamics. Data showed that pregnant and lactating females had significantly higher risk of infection, which may explain previously observed temporal associations between HeV outbreaks and flying fox birthing periods. Age-specific seroprevalence curves generated from field data imply that HeV is transmitted horizontally via faeces, urine or saliva. Rapidly declining seroprevalence between two field seasons suggests that immunity wanes faster in LRFF than in other flying fox species, and highlights the potentially critical role of this species in interspecific viral persistence. The highest seroprevalence was observed when animals showed evidence of nutritional stress, suggesting that environmental processes that alter flying fox food sources, such as habitat loss and climate change, may increase HeV infection and transmission. These insights into the ecology of HeV in flying fox populations suggest causal links between anthropogenic environmental change and HeV emergence.     

Spatiotemporal Aspects of Hendra Virus Infection in Pteropid Bats (Flying-Foxes) in Eastern Australia

Hendra virus (HeV) causes highly lethal disease in horses and humans in the eastern Australian states of Queensland (QLD) and New South Wales (NSW), with multiple equine cases now reported on an annual basis. Infection and excretion dynamics in pteropid bats (flying-foxes), the recognised natural reservoir, are incompletely understood. We sought to identify key spatial and temporal factors associated with excretion in flying-foxes over a 2300 km latitudinal gradient from northern QLD to southern NSW which encompassed all known equine case locations. The aim was to strengthen knowledge of Hendra virus ecology in flying-foxes to improve spillover risk prediction and exposure risk mitigation strategies, and thus better protect horses and humans. Monthly pooled urine samples were collected from under roosting flying-foxes over a three-year period and screened for HeV RNA by quantitative RT-PCR. A generalised linear model was employed to investigate spatiotemporal associations with HeV detection in 13,968 samples from 27 roosts. There was a non-linear relationship between mean HeV excretion prevalence and five latitudinal regions, with excretion moderate in northern and central QLD, highest in southern QLD/northern NSW, moderate in central NSW, and negligible in southern NSW. Highest HeV positivity occurred where black or spectacled flying-foxes were present; nil or very low positivity rates occurred in exclusive grey-headed flying-fox roosts. Similarly, little red flying-foxes are evidently not a significant source of virus, as their periodic extreme increase in numbers at some roosts was not associated with any concurrent increase in HeV detection. There was a consistent, strong winter seasonality to excretion in the southern QLD/northern NSW and central NSW regions. This new information allows risk management strategies to be refined and targeted, mindful of the potential for spatial risk profiles to shift over time with changes in flying-fox species distribution.     

Climate Change Could Increase the Geographic Extent of Hendra Virus Spillover Risk

Disease risk mapping is important for predicting and mitigating impacts of bat-borne viruses, including Hendra virus (Paramyxoviridae:Henipavirus), that can spillover to domestic animals and thence to humans. We produced two models to estimate areas at potential risk of HeV spillover explained by the climatic suitability for its flying fox reservoir hosts, Pteropus alecto and P. conspicillatus. We included additional climatic variables that might affect spillover risk through other biological processes (such as bat or horse behaviour, plant phenology and bat foraging habitat). Models were fit with a Poisson point process model and a log-Gaussian Cox process. In response to climate change, risk expanded southwards due to an expansion of P. alecto suitable habitat, which increased the number of horses at risk by 175–260% (110,000–165,000). In the northern limits of the current distribution, spillover risk was highly uncertain because of model extrapolation to novel climatic conditions. The extent of areas at risk of spillover from P. conspicillatus was predicted shrink. Due to a likely expansion of P. alecto into these areas, it could replace P. conspicillatus as the main HeV reservoir. We recommend: (1) HeV monitoring in bats, (2) enhancing HeV prevention in horses in areas predicted to be at risk, (3) investigate and develop mitigation strategies for areas that could experience reservoir host replacements.     

Investigation of the climatic and environmental context of Hendra virus spillover events 1994-2010

Hendra virus is a recently emerged bat-borne zoonotic agent with high lethality in horses and humans in Australia. This is a rare disease and the determinants of bat to horse transmission, including the factors that bring these hosts together at critical times, are poorly understood. In this cross-disciplinary study climatic and vegetation primary productivity variables are compared for the dispersed and heterogenic 1994–2010 outbreak sites. The significant occurrence of spillover events within the dry season (p =  0.013, 95% CI (0.57–0.98)) suggests seasonal forcing of transmission across species, or seasonal forcing of virus excretion by the reservoir host. We explore the evidence for both. Preliminary investigations of the spatial determinants of Hendra disease locations are also presented. We find that postal areas in the Australian state of Queensland in which pteropid fruit bat (flying fox) roosts occur are approximately forty times more likely (OR = 40.5, (95% CI (5.16, 317.52)) to be the location of Hendra spillover events. This appears to be independent of density of horses at these locations. We consider issues of scale of host resource use, land use change and limitations of existing data that challenge analysis and limit further conclusive outcomes. This investigation of a broad range of potential climatic and environmental influences provides a good base for future investigations. Further understanding of cross-species Hendra virus transmission requires better understanding of flying fox resource use in the urban-rural landscape.     

Routes of Hendra Virus Excretion in Naturally-Infected Flying-Foxes: Implications for Viral Transmission and Spillover Risk

Pteropid bats or flying-foxes (Chiroptera: Pteropodidae) are the natural host of Hendra virus (HeV) which sporadically causes fatal disease in horses and humans in eastern Australia. While there is strong evidence that urine is an important infectious medium that likely drives bat to bat transmission and bat to horse transmission, there is uncertainty about the relative importance of alternative routes of excretion such as nasal and oral secretions, and faeces. Identifying the potential routes of HeV excretion in flying-foxes is important to effectively mitigate equine exposure risk at the bat-horse interface, and in determining transmission rates in host-pathogen models. The aim of this study was to identify the major routes of HeV excretion in naturally infected flying-foxes, and secondarily, to identify between-species variation in excretion prevalence. A total of 2840 flying-foxes from three of the four Australian mainland species (Pteropus alecto, P. poliocephalus and P. scapulatus) were captured and sampled at multiple roost locations in the eastern states of Queensland and New South Wales between 2012 and 2014. A range of biological samples (urine and serum, and urogenital, nasal, oral and rectal swabs) were collected from anaesthetized bats, and tested for HeV RNA using a qRT-PCR assay targeting the M gene. Forty-two P. alecto (n = 1410) had HeV RNA detected in at least one sample, and yielded a total of 78 positive samples, at an overall detection rate of 1.76% across all samples tested in this species (78/4436). The rate of detection, and the amount of viral RNA, was highest in urine samples (>serum, packed haemocytes >faecal >nasal >oral), identifying urine as the most plausible source of infection for flying-foxes and for horses. Detection in a urine sample was more efficient than detection in urogenital swabs, identifying the former as the preferred diagnostic sample. The detection of HeV RNA in serum is consistent with haematogenous spread, and with hypothesised latency and recrudesence in flying-foxes. There were no detections in P. poliocephalus (n = 1168 animals; n = 2958 samples) or P. scapulatus (n = 262 animals; n = 985 samples), suggesting (consistent with other recent studies) that these species are epidemiologically less important than P. alecto in HeV infection dynamics. The study is unprecedented in terms of the individual animal approach, the large sample size, and the use of a molecular assay to directly determine infection status. These features provide a high level of confidence in the veracity of our findings, and a sound basis from which to more precisely target equine risk mitigation strategies.     

Pollination by the locally endangered island flying fox (Pteropus hypomelanus) enhances fruit production of the economically important durian (Durio zibethinus)

Fruit bats provide valuable pollination services to humans through a unique coevolutionary relationship with chiropterophilous plants. However, chiropterophily in the Old World and the pollination roles of large bats, such as flying foxes (Pteropus spp., Acerodon spp., Desmalopex spp.), are still poorly understood and require further elucidation. Efforts to protect these bats have been hampered by a lack of basic quantitative information on their role as ecosystem service providers. Here, we investigate the role of the locally endangered island flying fox Pteropus hypomelanus in the pollination ecology of durian (Durio zibethinus), an economically important crop in Southeast Asia. On Tioman Island, Peninsular Malaysia, we deployed 19 stations of paired infrared camera and video traps across varying heights at four individual flowering trees in a durian orchard. We detected at least nine species of animal visitors, but only bats had mutualistic interactions with durian flowers. There was a clear vertical stratification in the feeding niches of flying foxes and nectar bats, with flying foxes feeding at greater heights in the trees. Flying foxes had a positive effect on mature fruit set and therefore serve as important pollinators for durian trees. As such, semi‐wild durian trees—particularly tall ones—may be dependent on flying foxes for enhancing reproductive success. Our study is the first to quantify the role of flying foxes in durian pollination, demonstrating that these giant fruit bats may have far more important ecological, evolutionary, and economic roles than previously thought. This has important implications and can aid efforts to promote flying fox conservation, especially in Southeast Asian countries.     

Flying-Fox Roost Disturbance and Hendra Virus Spillover Risk

Bats of the genus Pteropus (flying-foxes) are the natural host of Hendra virus (HeV) which periodically causes fatal disease in horses and humans in Australia. The increased urban presence of flying-foxes often provokes negative community sentiments because of reduced social amenity and concerns of HeV exposure risk, and has resulted in calls for the dispersal of urban flying-fox roosts. However, it has been hypothesised that disturbance of urban roosts may result in a stress-mediated increase in HeV infection in flying-foxes, and an increased spillover risk. We sought to examine the impact of roost modification and dispersal on HeV infection dynamics and cortisol concentration dynamics in flying-foxes. The data were analysed in generalised linear mixed models using restricted maximum likelihood (REML). The difference in mean HeV prevalence in samples collected before (4.9%), during (4.7%) and after (3.4%) roost disturbance was small and non-significant (P = 0.440). Similarly, the difference in mean urine specific gravity-corrected urinary cortisol concentrations was small and non-significant (before = 22.71 ng/mL, during = 27.17, after = 18.39) (P= 0.550). We did find an underlying association between cortisol concentration and season, and cortisol concentration and region, suggesting that other (plausibly biological or environmental) variables play a role in cortisol concentration dynamics. The effect of roost disturbance on cortisol concentration approached statistical significance for region, suggesting that the relationship is not fixed, and plausibly reflecting the nature and timing of disturbance. We also found a small positive statistical association between HeV excretion status and urinary cortisol concentration. Finally, we found that the level of flying-fox distress associated with roost disturbance reflected the nature and timing of the activity, highlighting the need for a ‘best practice’ approach to dispersal or roost modification activities. The findings usefully inform public discussion and policy development in relation to Hendra virus and flying-fox management.     

Flying foxes as carriers of pathogenic Leptospira species

Recent serologic studies have identified flying foxes (Pteropus spp.) as carriers of leptospirosis; however, little is known about the role of flying foxes as carriers of pathogenic Leptospira spp. To determine if Australian Pteropus spp. are carriers of pathogenic Leptospira spp., TaqMan real-time polymerase chain reaction (PCR) was used to detect leptospiral DNA in kidney and urine specimens from four species of flying fox, including the spectacled flying fox (Pteropus conspicillatus), black flying fox (Pteropus alecto), grey-headed flying fox (Pteropus poliocephalus), and little red flying fox (Pteropus scapulatus). Of the 173 kidney samples tested, 19 (11%) were positive for leptospiral DNA. Positive individuals were detected in all four species; significant differences in prevalence were not detected between species, between species within the same geographic area, or between geographically separated samples from the same species. Of the 46 urine samples tested, 18 (39%) tested positive by PCR, confirming that flying foxes shed leptospires into the environment. The detection of leptospiral DNA in the kidneys and urine of flying foxes suggests that flying foxes are carriers of pathogenic Leptospira spp. No evidence collected in the present study, however, suggests that flying foxes pose a significant risk of leptospirosis to the wider community or that humans who are in regular, close contact with flying foxes are at risk for leptospirosis.     

Population genetics of fruit bat reservoir informs the dynamics,distribution and diversity of Nipah virus

The structure and connectivity of wildlife host populations may influence zoonotic disease dynamics, evolution and therefore spillover risk to people. Fruit bats in the genus Pteropus, or flying foxes, are the primary natural reservoir for henipaviruses—a group of emerging paramyxoviruses that threaten livestock and public health. In Bangladesh, Pteropus medius is the reservoir for Nipah virus—and viral spillover has led to human fatalities nearly every year since 2001. Here, we use mitochondrial DNA and nuclear microsatellite markers to measure the population structure, demographic history and phylogeography of P. medius in Bangladesh. We combine this with a phylogeographic analysis of all known Nipah virus sequences and strains currently available to better inform the dynamics, distribution and evolutionary history of Nipah virus. We show that P. medius is primarily panmictic, but combined analysis of microsatellite and morphological data shows evidence for differentiation of two populations in eastern Bangladesh, corresponding to a divergent strain of Nipah virus also found in bats from eastern Bangladesh. Our demographic analyses indicate that a large, expanding population of flying foxes has existed in Bangladesh since the Late Pleistocene, coinciding with human population expansion in South Asia, suggesting repeated historical spillover of Nipah virus likely occurred. We present the first evidence of mitochondrial introgression, or hybridization, between P. medius and flying fox species found in South‐East Asia (P. vampyrus and P. hypomelanus), which may help to explain the distribution of Nipah virus strains across the region.     

Disease Risk Perception and Safety Practices: A Survey of Australian Flying Fox Rehabilitators

Interactions with flying foxes pose disease transmission risks to volunteer rehabilitators (carers) who treat injured, ill, and orphaned bats. In particular, Australian bat lyssavirus (ABLV) can be transmitted directly from flying foxes to humans in Australia. Personal protective equipment (PPE) and rabies vaccination can be used to protect against lyssavirus infection. During May and June 2014, active Australian flying fox carers participated in an online survey (SOAR: Survey Of Australian flying fox Rehabilitators) designed to gather demographic data, assess perceptions of disease risk, and explore safety practices. Responses to open-ended questions were analysed thematically. A logistic regression was performed to assess whether rehabilitators’ gender, use of PPE, threat perception, and years of experience predicted variation in their odds of being bitten or scratched. Eligible responses were received from 122 rehabilitators located predominantly on the eastern coast of Australia. Eighty-four percent of respondents were female. Years of experience ranged from <1 to 30 years (median 5 years). Respondents were highly educated. All rehabilitators were vaccinated against rabies and 94% received a rabies titre check at least every two years. Sixty-three percent of carers did not perceive viruses in flying foxes as a potential threat to their health, yet 74% of carers reported using PPE when handling flying foxes. Eighty-three percent of rehabilitators had received a flying fox bite or scratch at some point during their career. Carers provide an important community service by rescuing and rehabilitating flying foxes. While rehabilitators in this study have many excellent safety practices, including a 100% vaccination rate against rabies, there is room for improvement in PPE use. We recommend 1) the establishment of an Australia-wide set of guidelines for safety when caring for bats and 2) that the responsible government agencies in Australia support carers who rescue potentially ABLV-infected bats by offering compensation for PPE.     

Ecological dynamics of emerging bat virus spillover

Viruses that originate in bats may be the most notorious emerging zoonoses that spill over from wildlife into domestic animals and humans. Understanding how these infections filter through ecological systems to cause disease in humans is of profound importance to public health. Transmission of viruses from bats to humans requires a hierarchy of enabling conditions that connect the distribution of reservoir hosts, viral infection within these hosts, and exposure and susceptibility of recipient hosts. For many emerging bat viruses, spillover also requires viral shedding from bats, and survival of the virus in the environment. Focusing on Hendra virus, but also addressing Nipah virus, Ebola virus, Marburg virus and coronaviruses, we delineate this cross-species spillover dynamic from the within-host processes that drive virus excretion to land-use changes that increase interaction among species. We describe how land-use changes may affect co-occurrence and contact between bats and recipient hosts. Two hypotheses may explain temporal and spatial pulses of virus shedding in bat populations: episodic shedding from persistently infected bats or transient epidemics that occur as virus is transmitted among bat populations. Management of livestock also may affect the probability of exposure and disease. Interventions to decrease the probability of virus spillover can be implemented at multiple levels from targeting the reservoir host to managing recipient host exposure and susceptibility.     

Coexistence and Conflict between the Island Flying fox (<Emphasis Type="Italic">Pteropus hypomelanus</Emphasis>) and Humans on Tioman Island,Peninsular Malaysia

As tropical landscapes become increasingly human-dominated, conflicts between people and wildlife threaten ecological processes. Old World fruit bats such as flying foxes are especially susceptible to extinction risk because there is low interest in their conservation, particularly when they are considered pests. In order to arrest fruit bat declines, there is an urgent need to understand human-bat conflict and its implications. On a tropical island in Peninsular Malaysia, we conducted a questionnaire survey to investigate coexistence between people and the island flying fox (Pteropus hypomelanus). Among 119 respondents, knowledge of ecosystem services provided by flying foxes was extremely low. Most respondents held negative attitudes towards the bats, and older male locals were more likely to support killing them. This was also true for older owners of fruit trees who derived income from selling fruit, and experienced flying fox raids. Our results can be used to design appropriate interventions to support conservation efforts, and has important implications for managing conflicts between humans and synanthropic wildlife.     

The natural history of Hendra and Nipah viruses

Pteropid bats (flying foxes), species of which are the probable natural host of both Hendra and Nipah viruses, occur in overlapping populations from India to Australia. Ecological changes associated with land use and with animal husbandry practices appear most likely to be associated with the emergence of these two agents.     

Population structure of two rabies hosts relative to the known distribution of rabies virus variants in Alaska

For pathogens that infect multiple species, the distinction between reservoir hosts and spillover hosts is often difficult. In Alaska, three variants of the arctic rabies virus exist with distinct spatial distributions. We tested the hypothesis that rabies virus variant distribution corresponds to the population structure of the primary rabies hosts in Alaska, arctic foxes (Vulpes lagopus) and red foxes (Vulpes vulpes) to possibly distinguish reservoir and spillover hosts. We used mitochondrial DNA (mtDNA) sequence and nine microsatellites to assess population structure in those two species. mtDNA structure did not correspond to rabies virus variant structure in either species. Microsatellite analyses gave varying results. Bayesian clustering found two groups of arctic foxes in the coastal tundra region, but for red foxes it identified tundra and boreal types. Spatial Bayesian clustering and spatial principal components analysis identified 3 and 4 groups of arctic foxes, respectively, closely matching the distribution of rabies virus variants in the state. Red foxes, conversely, showed eight clusters comprising two regions (boreal and tundra) with much admixture. These results run contrary to previous beliefs that arctic fox show no fine‐scale spatial population structure. While we cannot rule out that the red fox is part of the maintenance host community for rabies in Alaska, the distribution of virus variants appears to be driven primarily by the arctic fox. Therefore, we show that host population genetics can be utilized to distinguish between maintenance and spillover hosts when used in conjunction with other approaches.     

A Novel Model of Lethal Hendra Virus Infection in African Green Monkeys and the Effectiveness of Ribavirin Treatment

The henipaviruses, Hendra virus (HeV) and Nipah virus (NiV), are emerging zoonotic paramyxoviruses that can cause severe and often lethal neurologic and/or respiratory disease in a wide variety of mammalian hosts, including humans. There are presently no licensed vaccines or treatment options approved for human or veterinarian use. Guinea pigs, hamsters, cats, and ferrets, have been evaluated as animal models of human HeV infection, but studies in nonhuman primates (NHP) have not been reported, and the development and approval of any vaccine or antiviral for human use will likely require efficacy studies in an NHP model. Here, we examined the pathogenesis of HeV in the African green monkey (AGM) following intratracheal inoculation. Exposure of AGMs to HeV produced a uniformly lethal infection, and the observed clinical signs and pathology were highly consistent with HeV-mediated disease seen in humans. Ribavirin has been used to treat patients infected with either HeV or NiV; however, its utility in improving outcome remains, at best, uncertain. We examined the antiviral effect of ribavirin in a cohort of nine AGMs before or after exposure to HeV. Ribavirin treatment delayed disease onset by 1 to 2 days, with no significant benefit for disease progression and outcome. Together our findings introduce a new disease model of acute HeV infection suitable for testing antiviral strategies and also demonstrate that, while ribavirin may have some antiviral activity against the henipaviruses, its use as an effective standalone therapy for HeV infection is questionable.Hendra virus (HeV) and Nipah virus (NiV) are members of the genus Henipavirus (family Paramyxoviridae) that can cause severe respiratory illness and/or encephalitis in a wide variety of mammals, including horses, pigs, and humans (7, 23). HeV was identified as the causative agent of an acute respiratory disease in horses in 1994 in Queensland, Australia (23), and to date there have been 14 outbreaks in Australia since, with at least one occurrence per year since 2006, most recently in May 2010 (ProMed-mail no. 20100522.1699 [International Society for Infectious Diseases, http://www.promedmail.org]). Every outbreak of HeV has involved horses as the initial infected host, and there have been a total of seven human cases arising from exposure to infected horses. Four human fatalities have occurred (22), with the most recent occurring in August of 2009 (ProMed-mail no. 20090826.2998 and 20090903.3098). All patients initially presented with influenza-like illnesses (ILIs) after an incubation period of 7 to 16 days. While two individuals recovered from ILI, one patient developed pneumonitis and died from multiorgan failure. Three of the lethal cases developed encephalitic manifestations (mild confusion and ataxia), with two patients experiencing seizures (22, 23, 27).Data on the histopathology of fatal human HeV cases are limited, but the pathology includes small necrotic plaques in the cerebrum and cerebellum, in addition to mild parenchymal inflammation (21, 27). Severe parenchymal inflammation and necrosis were observed in the lungs. More extensive histopathologic data are available from 32 autopsies of fatal human NiV cases (28). Similarly to the HeV cases, pathology was characterized by systemic vasculitis and parenchymal necrosis in the central nervous system (CNS), while in the lung, pathological findings mainly included vasculitis, fibrinoid necrosis, alveolar hemorrhage, pulmonary edema, and aspiration pneumonia. Other organs that were affected included heart, kidney, and spleen and showed generally mild or focal inflammation. The development of syncytial multinucleated endothelial cells is characteristic of both HeV and NiV (27, 28). At present, the details of the pathogenesis and histopathological changes mediated by either HeV or NiV infection in humans are naturally derived from only the late phases of the disease course, and therefore a relevant animal model is needed that mimics the disease progression seen in humans.Pteropid fruit bats, commonly known as flying foxes in the family Pteropodidae, are the principle natural reservoirs for both NiV and HeV (reviewed in reference 3). However, these henipaviruses display a broad species tropism, and in addition to bats, horses and humans, natural and/or experimental infection of HeV has been demonstrated in guinea pigs, hamsters, pigs, cats, and ferrets (25). Experimental infections of Syrian hamsters with HeV is lethal, and animals show disease similar to that of human cases, including respiratory and neurological symptoms, depending on the dose (11; unpublished data). In this model, viral RNA can be detected in various organs of infected hamsters, including brain, lung, kidney, heart, liver, and spleen. The main histopathological findings included parenchymal infection in various organs, including the brain, with vasculitis and syncytial multinucleated endothelial cells in many blood vessels (11). While this model is useful in studying pathogenesis, it is limited in the availability of reagents to do so.There are currently no vaccines or treatments licensed for human use. Several in vitro studies have shown that ribavirin is effective against both HeV and NiV infection (1, 2, 29). An open-label ribavirin treatment trial was run during an outbreak of NiV in Malaysia in 1998 and reported to reduce mortality by 36% (6). Of the seven recorded human HeV cases, three patients were treated with ribavirin, one of whom survived (22). In the most recent outbreak of HeV in Australia, three additional people received ribavirin treatment in combination with chloroquine after suspected exposure to HeV-contaminated secretions from infected horses. While all three individuals survived, infection was not confirmed, and therefore it remains unknown whether the treatment had any beneficiary effect (ProMed-mail no. 20090826.2998). In addition, two animal studies in hamsters showed that ribavirin treatment delays but does not prevent death from NiV or HeV infection (8, 10). Therefore, an animal model with greater relevance to humans and that recapitulates the disease processes seen in human cases of HeV is needed to get a better answer to whether ribavirin might be effective against henipavirus infections. In addition, the U.S. FDA implemented the “Animal Efficacy Rule,” which specifically applies to the development of therapeutic products when human efficacy studies are not possible or ethical, such as is often the case with highly virulent pathogens like HeV (24). Essentially, this rule allows for the evaluation of vaccines or therapeutics using data derived from studies carried out in at least two animal models. The licensure of any therapeutic modalities for HeV will require a thorough evaluation of HeV pathogenesis in nonhuman primates (NHPs).In the present study, we report the development and characterization of a new nonhuman primate (NHP) model of lethal HeV infection in the African green monkey (AGM). The pathogenesis and disease progression in the AGM upon HeV infection essentially mirrored the lethal disease episodes seen among human cases of HeV. Using this new model, the efficacy of ribavirin treatment against lethal challenge with HeV was examined. Here we have shown that ribavirin treatment can significantly delay but not prevent death of AGMs from lethal HeV infection. In addition to severe respiratory symptoms in all animals, prolonged disease progression in ribavirin-treated animals was also marked by the appearance of neurological symptoms.