Reservoir host immune responses to emerging zoonotic viruses.

Zoonotic viruses, such as HIV, Ebola virus, coronaviruses, influenza A viruses, hantaviruses, or henipaviruses, can result in profound pathology in humans. In contrast, populations of the reservoir hosts of zoonotic pathogens often appear to tolerate these infections with little evidence of disease. Why are viruses more dangerous in one species than another? Immunological studies investigating quantitative and qualitative differences in the host-virus equilibrium in animal reservoirs will be key to answering this question, informing new approaches for treating and preventing zoonotic diseases. Integrating an understanding of host immune responses with epidemiological, ecological, and evolutionary insights into viral emergence will shed light on mechanisms that minimize fitness costs associated with viral infection, facilitate transmission to other hosts, and underlie the association of specific reservoir hosts with multiple emerging viruses. Reservoir host studies provide a rich opportunity for elucidating fundamental immunological processes and their underlying genetic basis, in the context of distinct physiological and metabolic constraints that contribute to host resistance and disease tolerance.

Nipah Virus Exposure in Domestic and Peridomestic Animals Living in Human Outbreak Sites, Bangladesh, 2013-2015.

Spillovers of Nipah virus (NiV) from Pteropus bats to humans occurs frequently in Bangladesh, but the risk for spillover into other animals is poorly understood. We detected NiV antibodies in cattle, dogs, and cats from 6 sites where spillover human NiV infection cases occurred during 2013-2015.

Interventions to Reduce Risk for Pathogen Spillover and Early Disease Spread to Prevent Outbreaks, Epidemics, and Pandemics.

The pathogens that cause most emerging infectious diseases in humans originate in animals, particularly wildlife, and then spill over into humans. The accelerating frequency with which humans and domestic animals encounter wildlife because of activities such as land-use change, animal husbandry, and markets and trade in live wildlife has created growing opportunities for pathogen spillover. The risk of pathogen spillover and early disease spread among domestic animals and humans, however, can be reduced by stopping the clearing and degradation of tropical and subtropical forests, improving health and economic security of communities living in emerging infectious disease hotspots, enhancing biosecurity in animal husbandry, shutting down or strictly regulating wildlife markets and trade, and expanding pathogen surveillance. We summarize expert opinions on how to implement these goals to prevent outbreaks, epidemics, and pandemics.

Variations in small-scale movements of, Rousettus aegyptiacus, a Marburg virus reservoir across a seasonal gradient.

BACKGROUND: Bats are increasingly being recognized as important hosts for viruses, some of which are zoonotic and carry the potential for spillover within human and livestock populations. Biosurveillance studies focused on assessing the risk of pathogen transmission, however, have largely focused on the virological component and have not always considered the ecological implications of different species as viral hosts. The movements of known viral hosts are an important component for disease risk assessments as they can potentially identify regions of higher risk of contact and spillover. As such, this study aimed to synthesize data from both virological and ecological fields to provide a more holistic assessment of the risk of pathogen transmission from bats to people. RESULTS: Using radiotelemetry, we tracked the small-scale movements of Rousettus aegyptiacus, a species of bat known to host Marburg virus and other viruses with zoonotic potential, in a rural settlement in Limpopo Province, South Africa. The tracked bats exhibited seasonal variations in their movement patterns including variable usage of residential areas which could translate to contact between bats and humans and may facilitate spillover. We identified a trend for increased usage of residential areas during the winter months with July specifically experiencing the highest levels of bat activity within residential areas. July has previously been identified as a key period for increased spillover risk for viruses associated with R. aegyptiacus from this colony and paired with the increased activity levels, illustrates the risk for spillover to human populations. CONCLUSION: This study emphasizes the importance of incorporating ecological data such as movement patterns with virological data to provide a better understanding of the risk of pathogen spillover and transmission.

Nipah virus dynamics in bats and implications for spillover to humans.

Nipah virus (NiV) is an emerging bat-borne zoonotic virus that causes near-annual outbreaks of fatal encephalitis in South Asia-one of the most populous regions on Earth. In Bangladesh, infection occurs when people drink date-palm sap contaminated with bat excreta. Outbreaks are sporadic, and the influence of viral dynamics in bats on their temporal and spatial distribution is poorly understood. We analyzed data on host ecology, molecular epidemiology, serological dynamics, and viral genetics to characterize spatiotemporal patterns of NiV dynamics in its wildlife reservoir, Pteropus medius bats, in Bangladesh. We found that NiV transmission occurred throughout the country and throughout the year. Model results indicated that local transmission dynamics were modulated by density-dependent transmission, acquired immunity that is lost over time, and recrudescence. Increased transmission followed multiyear periods of declining seroprevalence due to bat-population turnover and individual loss of humoral immunity. Individual bats had smaller host ranges than other Pteropus species (spp.), although movement data and the discovery of a Malaysia-clade NiV strain in eastern Bangladesh suggest connectivity with bats east of Bangladesh. These data suggest that discrete multiannual local epizootics in bat populations contribute to the sporadic nature of NiV outbreaks in South Asia. At the same time, the broad spatial and temporal extent of NiV transmission, including the recent outbreak in Kerala, India, highlights the continued risk of spillover to humans wherever they may interact with pteropid bats and the importance of limiting opportunities for spillover throughout Pteropus's range.

Nipah Virus Detection at Bat Roosts after Spillover Events, Bangladesh, 2012-2019.

Knowledge of the dynamics and genetic diversity of Nipah virus circulating in bats and at the human-animal interface is limited by current sampling efforts, which produce few detections of viral RNA. We report a series of investigations at Pteropus medius bat roosts identified near the locations of human Nipah cases in Bangladesh during 2012-2019. Pooled bat urine was collected from 23 roosts; 7 roosts (30%) had >1 sample in which Nipah RNA was detected from the first visit. In subsequent visits to these 7 roosts, RNA was detected in bat urine up to 52 days after the presumed exposure of the human case-patient, although the probability of detection declined rapidly with time. These results suggest that rapidly deployed investigations of Nipah virus shedding from bat roosts near human cases could increase the success of viral sequencing compared with background surveillance and could enhance understanding of Nipah virus ecology and evolution.

Possibility for reverse zoonotic transmission of SARS-CoV-2 to free-ranging wildlife: A case study of bats.

The COVID-19 pandemic highlights the substantial public health, economic, and societal consequences of virus spillover from a wildlife reservoir. Widespread human transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) also presents a new set of challenges when considering viral spillover from people to naïve wildlife and other animal populations. The establishment of new wildlife reservoirs for SARS-CoV-2 would further complicate public health control measures and could lead to wildlife health and conservation impacts. Given the likely bat origin of SARS-CoV-2 and related beta-coronaviruses (ß-CoVs), free-ranging bats are a key group of concern for spillover from humans back to wildlife. Here, we review the diversity and natural host range of ß-CoVs in bats and examine the risk of humans inadvertently infecting free-ranging bats with SARS-CoV-2. Our review of the global distribution and host range of ß-CoV evolutionary lineages suggests that 40+ species of temperate-zone North American bats could be immunologically naïve and susceptible to infection by SARS-CoV-2. We highlight an urgent need to proactively connect the wellbeing of human and wildlife health during the current pandemic and to implement new tools to continue wildlife research while avoiding potentially severe health and conservation impacts of SARS-CoV-2 "spilling back" into free-ranging bat populations.

Classification of new morbillivirus and jeilongvirus sequences from bats sampled in Brazil and Malaysia.

As part of a broad One Health surveillance effort to detect novel viruses in wildlife and people, we report several paramyxovirus sequences sampled primarily from bats during 2013 and 2014 in Brazil and Malaysia, including seven from which we recovered full-length genomes. Of these, six represent the first full-length paramyxovirid genomes sequenced from the Americas, including two that are the first full-length bat morbillivirus genome sequences published to date. Our findings add to the vast number of viral sequences in public repositories, which have been increasing considerably in recent years due to the rising accessibility of metagenomics. Taxonomic classification of these sequences in the absence of phenotypic data has been a significant challenge, particularly in the subfamily Orthoparamyxovirinae, where the rate of discovery of novel sequences has been substantial. Using pairwise amino acid sequence classification (PAASC), we propose that five of these sequences belong to members of the genus Jeilongvirus and two belong to members of the genus Morbillivirus. We also highlight inconsistencies in the classification of Tupaia virus and Mòjiang virus using the same demarcation criteria and suggest reclassification of these viruses into new genera. Importantly, this study underscores the critical importance of sequence length in PAASC analysis as well as the importance of biological characteristics such as genome organization in the taxonomic classification of viral sequences.

URSID γ-HERPESVIRUS TYPE 1-RELATED VIRUS IN CAPTIVE BORNEAN SUN BEARS (HELARCTOS MALAYANUS EURYSPILUS) IN SABAH, MALAYSIA.

The Bornean sun bear (Helarctos malayanus euryspilus) is the smallest subspecies of sun bear. Their numbers are declining, and more research is needed to better understand their health and biology. Forty-four bears housed at the Bornean Sun Bear Conservation Centre (BSBCC) in Sabah, Malaysia, were screened for known and novel viruses in November 2018. Ursid γ-herpesvirus type 1 (UrHV-1) is a herpesvirus that has been detected from swab samples of clinically healthy sun bears and biopsy samples of oral squamous cell carcinoma in sun bears. We detected an UrHV-1-related virus from throat and rectal swabs by molecular viral screening in samples from 15.9% of the sun bears at BSBCC. None of the bears with the UrHV-1-related virus in this study had oral lesions. There is no known report of UrHV-1 detection in the wild sun bear population, and its association with oral squamous cell carcinoma is not fully understood. Finding an UrHV-1-related virus in a rehabilitation center is a concern because conditions in captivity may contribute to spreading this virus, and there is the potential of introducing it into wild populations when a bear is released. This study demonstrates an urgent need to carry out similar surveillance for sun bears in captivity as well as those in the wild, to better understand the impact of captivity on the prevalence and spread of UrHV-1-related viruses. Positive bears also should be monitored for oral lesions to better understand whether there is a causal relationship.

Serological Evidence for Henipa-like and Filo-like Viruses in Trinidad Bats.

Bat-borne zoonotic pathogens belonging to the family Paramxyoviridae, including Nipah and Hendra viruses, and the family Filoviridae, including Ebola and Marburg viruses, can cause severe disease and high mortality rates on spillover into human populations. Surveillance efforts for henipaviruses and filoviruses have been largely restricted to the Old World; however, recent studies suggest a potentially broader distribution for henipaviruses and filoviruses than previously recognized. In the current study, we screened for henipaviruses and filoviruses in New World bats collected across 4 locations in Trinidad near the coast of Venezuela. Bat tissue samples were screened using previously established reverse-transcription polymerase chain reaction assays. Serum were screened using a multiplex immunoassay to detect antibodies reactive with the envelope glycoprotein of viruses in the genus Henipavirus and the family Filoviridae. Serum samples were also screened by means of enzyme-linked immunosorbent assay for antibodies reactive with Nipah G and F glycoproteins. Of 84 serum samples, 28 were reactive with ≥1 henipavirus glycoprotein by ≥1 serological method, and 6 serum samples were reactive against ≥1 filovirus glycoproteins. These data provide evidence of potential circulation of viruses related to the henipaviruses and filoviruses in New World bats.

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.

Isolation and Full-Genome Characterization of Nipah Viruses from Bats, Bangladesh.

Despite molecular and serologic evidence of Nipah virus in bats from various locations, attempts to isolate live virus have been largely unsuccessful. We report isolation and full-genome characterization of 10 Nipah virus isolates from Pteropus medius bats sampled in Bangladesh during 2013 and 2014.

Isolation and characterization of a bat SARS-like coronavirus that uses the ACE2 receptor.

The 2002-3 pandemic caused by severe acute respiratory syndrome coronavirus (SARS-CoV) was one of the most significant public health events in recent history. An ongoing outbreak of Middle East respiratory syndrome coronavirus suggests that this group of viruses remains a key threat and that their distribution is wider than previously recognized. Although bats have been suggested to be the natural reservoirs of both viruses, attempts to isolate the progenitor virus of SARS-CoV from bats have been unsuccessful. Diverse SARS-like coronaviruses (SL-CoVs) have now been reported from bats in China, Europe and Africa, but none is considered a direct progenitor of SARS-CoV because of their phylogenetic disparity from this virus and the inability of their spike proteins to use the SARS-CoV cellular receptor molecule, the human angiotensin converting enzyme II (ACE2). Here we report whole-genome sequences of two novel bat coronaviruses from Chinese horseshoe bats (family: Rhinolophidae) in Yunnan, China: RsSHC014 and Rs3367. These viruses are far more closely related to SARS-CoV than any previously identified bat coronaviruses, particularly in the receptor binding domain of the spike protein. Most importantly, we report the first recorded isolation of a live SL-CoV (bat SL-CoV-WIV1) from bat faecal samples in Vero E6 cells, which has typical coronavirus morphology, 99.9% sequence identity to Rs3367 and uses ACE2 from humans, civets and Chinese horseshoe bats for cell entry. Preliminary in vitro testing indicates that WIV1 also has a broad species tropism. Our results provide the strongest evidence to date that Chinese horseshoe bats are natural reservoirs of SARS-CoV, and that intermediate hosts may not be necessary for direct human infection by some bat SL-CoVs. They also highlight the importance of pathogen-discovery programs targeting high-risk wildlife groups in emerging disease hotspots as a strategy for pandemic preparedness.

Middle East Respiratory Syndrome Coronavirus Antibodies in Dromedary Camels, Bangladesh, 2015.

Dromedary camels are bred domestically and imported into Bangladesh. In 2015, of 55 camels tested for Middle East respiratory syndrome coronavirus in Dhaka, 17 (31%) were seropositive, including 1 bred locally. None were PCR positive. The potential for infected camels in urban markets could have public health implications and warrants further investigation.

Convergence of Humans, Bats, Trees, and Culture in Nipah Virus Transmission, Bangladesh.

Preventing emergence of new zoonotic viruses depends on understanding determinants for human risk. Nipah virus (NiV) is a lethal zoonotic pathogen that has spilled over from bats into human populations, with limited person-to-person transmission. We examined ecologic and human behavioral drivers of geographic variation for risk of NiV infection in Bangladesh. We visited 60 villages during 2011-2013 where cases of infection with NiV were identified and 147 control villages. We compared case villages with control villages for most likely drivers for risk of infection, including number of bats, persons, and date palm sap trees, and human date palm sap consumption behavior. Case villages were similar to control villages in many ways, including number of bats, persons, and date palm sap trees, but had a higher proportion of households in which someone drank sap. Reducing human consumption of sap could reduce virus transmission and risk for emergence of a more highly transmissible NiV strain.

Interdisciplinary approaches to understanding disease emergence: the past, present, and future drivers of Nipah virus emergence.

Emerging infectious diseases (EIDs) pose a significant threat to human health, economic stability, and biodiversity. Despite this, the mechanisms underlying disease emergence are still not fully understood, and control measures rely heavily on mitigating the impact of EIDs after they have emerged. Here, we highlight the emergence of a zoonotic Henipavirus, Nipah virus, to demonstrate the interdisciplinary and macroecological approaches necessary to understand EID emergence. Previous work suggests that Nipah virus emerged due to the interaction of the wildlife reservoir (Pteropus spp. fruit bats) with intensively managed livestock. The emergence of this and other henipaviruses involves interactions among a suite of anthropogenic environmental changes, socioeconomic factors, and changes in demography that overlay and interact with the distribution of these pathogens in their wildlife reservoirs. Here, we demonstrate how ecological niche modeling may be used to investigate the potential role of a changing climate on the future risk for Henipavirus emergence. We show that the distribution of Henipavirus reservoirs, and therefore henipaviruses, will likely change under climate change scenarios, a fundamental precondition for disease emergence in humans. We assess the variation among climate models to estimate where Henipavirus host distribution is most likely to expand, contract, or remain stable, presenting new risks for human health. We conclude that there is substantial potential to use this modeling framework to explore the distribution of wildlife hosts under a changing climate. These approaches may directly inform current and future management and surveillance strategies aiming to improve pathogen detection and, ultimately, reduce emergence risk.

Bats are a major natural reservoir for hepaciviruses and pegiviruses.

Although there are over 1,150 bat species worldwide, the diversity of viruses harbored by bats has only recently come into focus as a result of expanded wildlife surveillance. Such surveys are of importance in determining the potential for novel viruses to emerge in humans, and for optimal management of bats and their habitats. To enhance our knowledge of the viral diversity present in bats, we initially surveyed 415 sera from African and Central American bats. Unbiased high-throughput sequencing revealed the presence of a highly diverse group of bat-derived viruses related to hepaciviruses and pegiviruses within the family Flaviridae. Subsequent PCR screening of 1,258 bat specimens collected worldwide indicated the presence of these viruses also in North America and Asia. A total of 83 bat-derived viruses were identified, representing an infection rate of nearly 5%. Evolutionary analyses revealed that all known hepaciviruses and pegiviruses, including those previously documented in humans and other primates, fall within the phylogenetic diversity of the bat-derived viruses described here. The prevalence, unprecedented viral biodiversity, phylogenetic divergence, and worldwide distribution of the bat-derived viruses suggest that bats are a major and ancient natural reservoir for both hepaciviruses and pegiviruses and provide insights into the evolutionary history of hepatitis C virus and the human GB viruses.

Macacine Herpesvirus 1 in Long-Tailed Macaques, Malaysia, 2009-2011.

Macacine herpesvirus 1 (MaHV1; B virus) naturally infects macaques (Macaca spp.) and can cause fatal encephalitis in humans. In Peninsular Malaysia, wild macaques are abundant, and translocation is used to mitigate human-macaque conflict. Most adult macaques are infected with MaHV1, although the risk for transmission to persons who handle them during capture and translocation is unknown. We investigated MaHV1 shedding among 392 long-tailed macaques (M. fascicularis) after capture and translocation by the Department of Wildlife and National Parks in Peninsular Malaysia, during 2009-2011. For detection of MaHV1 DNA, PCR was performed on urogenital and oropharyngeal swab samples. Overall, 39% of macaques were shedding MaHV1 DNA; rates of DNA detection did not differ between sample types. This study demonstrates that MaHV1 was shed by a substantial proportion of macaques after capture and transport and suggests that persons handling macaques under these circumstances might be at risk for exposure to MaHV1.

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.

Diversity of coronavirus in bats from Eastern Thailand.

BACKGROUND: Bats are reservoirs for a diverse range of coronaviruses (CoVs), including those closely related to human pathogens such as Severe Acute Respiratory Syndrome (SARS) CoV and Middle East Respiratory Syndrome CoV. There are approximately 139 bat species reported to date in Thailand, of which two are endemic species. Due to the zoonotic potential of CoVs, standardized surveillance efforts to characterize viral diversity in wildlife are imperative. FINDINGS: A total of 626 bats from 19 different bat species were individually sampled from 5 provinces in Eastern Thailand between 2008 and 2013 (84 fecal and 542 rectal swabs). Samples collected (either fresh feces or rectal swabs) were placed directly into RNA stabilization reagent, transported on ice within 24 hours and preserved at -80°C until further analysis. CoV RNA was detected in 47 specimens (7.6%), from 13 different bat species, using broadly reactive consensus PCR primers targeting the RNA-Dependent RNA Polymerase gene designed to detect all CoVs. Thirty seven alphacoronaviruses, nine lineage D betacoronaviruses, and one lineage B betacoronavirus (SARS-CoV related) were identified. Six new bat CoV reservoirs were identified in our study, namely Cynopterus sphinx, Taphozous melanopogon, Hipposideros lekaguli, Rhinolophus shameli, Scotophilus heathii and Megaderma lyra. CONCLUSIONS: CoVs from the same genetic lineage were found in different bat species roosting in similar or different locations. These data suggest that bat CoV lineages are not strictly concordant with their hosts. Our phylogenetic data indicates high diversity and a complex ecology of CoVs in bats sampled from specific areas in eastern regions of Thailand. Further characterization of additional CoV genes may be useful to better describe the CoV divergence.