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Abstract Reverse zoonotic respiratory diseases threaten great apes across Sub-Saharan Africa. Studies of wild chimpanzees have identified the causative agents of most respiratory disease outbreaks as “common cold” paediatric human pathogens, but reverse zoonotic transmission pathways have remained unclear. Between May 2019 and August 2021, we conducted a prospective cohort study of 234 children aged 3–11 years in communities bordering Kibale National Park, Uganda, and 30 adults who were forest workers and regularly entered the park. We collected 2047 respiratory symptoms surveys to quantify clinical severity and simultaneously collected 1989 nasopharyngeal swabs approximately monthly for multiplex viral diagnostics. Throughout the course of the study, we also collected 445 faecal samples from 55 wild chimpanzees living nearby in Kibale in social groups that have experienced repeated, and sometimes lethal, epidemics of human-origin respiratory viral disease. We characterized respiratory pathogens in each cohort and examined statistical associations between PCR positivity for detected pathogens and potential risk factors. Children exhibited high incidence rates of respiratory infections, whereas incidence rates in adults were far lower. COVID-19 lockdown in 2020–2021 significantly decreased respiratory disease incidence in both people and chimpanzees. Human respiratory infections peaked in June and September, corresponding to when children returned to school. Rhinovirus, which caused a 2013 outbreak that killed 10% of chimpanzees in a Kibale community, was the most prevalent human pathogen throughout the study and the only pathogen present at each monthly sampling, even during COVID-19 lockdown. Rhinovirus was also most likely to be carried asymptomatically by adults. Although we did not detect human respiratory pathogens in the chimpanzees during the cohort study, we detected human metapneumovirus in two chimpanzees from a February 2023 outbreak that were genetically similar to viruses detected in study participants in 2019. Our data suggest that respiratory pathogens circulate in children and that adults become asymptomatically infected during high-transmission times of year. These asymptomatic adults may then unknowingly carry the pathogens into forest and infect chimpanzees. This conclusion, in turn, implies that intervention strategies based on respiratory symptoms in adults are unlikely to be effective for reducing reverse zoonotic transmission of respiratory viruses to chimpanzees.

Abstract Proposed mechanisms of zoonotic virus spillover often posit that wildlife transmission and amplification precede human outbreaks. Between 2006 and 2012, the palm Raphia farinifera, a rich source of dietary minerals for wildlife, was nearly extirpated from Budongo Forest, Uganda. Since then, chimpanzees, black-and-white colobus, and red duiker were observed feeding on bat guano, a behavior not previously observed. Here we show that guano consumption may be a response to dietary mineral scarcity and may expose wildlife to bat-borne viruses. Videos from 2017–2019 recorded 839 instances of guano consumption by the aforementioned species. Nutritional analysis of the guano revealed high concentrations of sodium, potassium, magnesium and phosphorus. Metagenomic analyses of the guano identified 27 eukaryotic viruses, including a novel betacoronavirus. Our findings illustrate how “upstream” drivers such as socioeconomics and resource extraction can initiate elaborate chains of causation, ultimately increasing virus spillover risk.

Recreational fisheries involve an intimate connection between people, individual fish, and the environment. Recreational fishers and their health crucially depend on healthy fish and ecosystems. Similarly, fish and ecosystems can be impacted by the activities of people including recreational fishers. Thus, amplified by the global interest in recreational fishing, we posit that recreational fishing is particularly suited as an empirical system to explore a One Health perspective, with a goal of creating pathways to better manage such socio-ecological systems for the benefit of people, fish, and the environment. Although zoonoses are uncommon in fishes, fish can carry pathogens, biotoxins, or contaminants that are harmful to people. When captured and released, fish can experience stress and injuries that may promote pathogen development. Similarly, when humans contribute to environmental degradation, not only are fish impacted but so are the humans that depend on them for nutrition, livelihoods, culture, and well-being. Failure to embrace the One Health perspective for recreational fisheries has the potential to negatively impact the health of fish, fisheries, people, society, and the aquatic environment—especially important since these complex social–ecological systems are undergoing rapid change.

Abstract Protocols for characterizing taxonomic assemblages by deep sequencing of short DNA barcode regions (metabarcoding) have revolutionized our understanding of microbial communities and are standardized for bacteria, archaea, and fungi. Unfortunately, comparable methods for host-associated eukaryotes have lagged due to technical challenges. Despite 54 published studies, issues remain with primer complementarity, off-target amplification, and lack of external validation. Here, we present VESPA (Vertebrate Eukaryotic endoSymbiont and Parasite Analysis) primers and optimized metabarcoding protocol for host-associated eukaryotic community analysis. Using in silico prediction, panel PCR, engineered mock community standards, and clinical samples, we demonstrate VESPA to be more effective at resolving host-associated eukaryotic assemblages than previously published methods and to minimize off-target amplification. When applied to human and non-human primate samples, VESPA enables reconstruction of host-associated eukaryotic endosymbiont communities more accurately and at finer taxonomic resolution than microscopy. VESPA has the potential to advance basic and translational science on vertebrate eukaryotic endosymbiont communities, similar to achievements made for bacterial, archaeal, and fungal microbiomes.

IntroductionRespiratory illness is the most common childhood disease globally, especially in developing countries. Previous studies have detected viruses in approximately 70-80% of respiratory illnesses.MethodsIn a prospective cohort study of 234 young children (ages 3-11 years) and 30 adults (ages 22-51 years) in rural Western Uganda sampled monthly from May 2019 to August 2021, only 24.2% of nasopharyngeal swabs collected during symptomatic disease had viruses detectable by multiplex PCR diagnostics and metagenomic sequencing. In the remaining 75.8% of swabs from symptomatic participants, we measured detection rates of respiratory bacteria Haemophilus influenzae, Moraxella catarrhalis, and Streptococcus pneumoniae by quantitative PCR.Results100% of children tested positive for at least one bacterial species. Detection rates were 87.2%, 96.8%, and 77.6% in children and 10.0%, 36.7%, and 13.3% for adults for H. influenzae, M. catarrhalis, and S. pneumoniae, respectively. In children, 20.8% and 70.4% were coinfected with two and three pathogens, respectively, and in adults 6.7% were coinfected with three pathogens but none were coinfected with two. Detection of any of the three pathogens was not associated with season or respiratory symptoms severity, although parsing detection status by symptoms was challenged by children experiencing symptoms in 80.3% of monthly samplings, whereas adults only reported symptoms 26.6% of the time. Pathobiont colonization in children in Western Uganda was significantly more frequent than in children living in high-income countries, including in a study of age-matched US children that utilized identical diagnostic methods. Detection rates were, however, comparable to rates in children living in other Sub-Saharan African countries.DiscussionOverall, our results demonstrate that nonviral colds contribute significantly to respiratory disease burden among children in rural Uganda and that high rates of respiratory pathobiont colonization may play a role. These conclusions have implications for respiratory health interventions in the area, such as increasing childhood immunization rates and decreasing air pollutant exposure.

IntroductionRespiratory disease is a major cause of morbidity and mortality in the developing world, but prospective studies of temporal patterns and risk factors are rare.MethodsWe studied people in rural Western Uganda, where respiratory disease is pervasive. We followed 30 adults (ages 22–51 years; 534 observations) and 234 children (ages 3–11 years; 1,513 observations) between May 2019 and July 2022 and collected monthly data on their respiratory symptoms, for a total of 2,047 case records. We examined associations between demographic and temporal factors and respiratory symptoms severity.ResultsThe timing of our study (before, during, and after the emergence of COVID-19) allowed us to document the effects of public health measures instituted in the region. Incidence rates of respiratory symptoms before COVID-19 lockdown were 568.4 cases per 1,000 person-months in children and 254.2 cases per 1,000 person-months in adults. These rates were 2.6 times higher than the 2019 global average for children but comparable for adults. Younger children (ages 3–6 years) had the highest frequencies and severities of respiratory symptoms. Study participants were most likely to experience symptoms in February, which is a seasonal pattern not previously documented. Incidence and severity of symptoms in children decreased markedly during COVID-19 lockdown, illustrating the broad effects of public health measures on the incidence of respiratory disease.DiscussionOur results demonstrate that patterns of respiratory disease in settings such as Western Uganda resemble patterns in developed economies in some ways (age-related factors) but not in others (increased incidence in children and seasonal pattern). Factors such as indoor air quality, health care access, timing of school trimesters, and seasonal effects (rainy/dry seasons) likely contribute to the differences observed.

Freshwater mussels (order: Unionida) are highly imperiled globally and are increasingly the focus of captive propagation efforts to protect and restore wild populations. The Upper Tennessee River Basin (UTRB) in Virginia is a freshwater biodiversity hotspot hosting at least 45 of North America’s ~300 species of freshwater mussels, including 21 threatened and endangered species listed under the U.S. Endangered Species Act. Recent studies have documented that viruses and other microbes have contributed to freshwater mussel population declines in the UTRB. We conducted a multi-year longitudinal study of captive-reared hatchery mussels released to restoration sites throughout the UTRB to evaluate their viromes and compare them to captive hatchery environments. We documented 681 viruses from 27 families. The hatchery mussels had significantly less viruses than those deployed to wild sites, with only 20 viruses unique to the hatchery mussels. After the hatchery mussels were released into the wild, their number of viruses initially spiked and then increased steadily over time, with 451 viruses in total unique to the mussels in the wild. We found Clinch densovirus 1 (CDNV-1), a virus previously associated with mass mortality events in the Clinch River, in all samples, but the wild site mussels consistently had significantly higher CDNV-1 levels than those held in the hatchery. Our data document substantial differences between the viruses in the mussels in the hatchery and wild environments and rapid virome shifts after the mussels are released to the wild sites. These findings indicate that mussel release programs might benefit from acclimatization periods or other measures to mitigate the potential negative effects of rapid exposure to infectious agents found in natural environments.

Studies of marine fish have revealed distant relatives of viruses important to global fish and animal health, but few such studies exist for freshwater fish. To investigate whether freshwater fish also host such viruses, we characterized the viromes of five wild species of freshwater fish in Wisconsin, USA: bluegill (Lepomis macrochirus), brown trout (Salmo trutta), lake sturgeon (Acipenser fulvescens), northern pike (Esox lucius), and walleye (Sander vitreus). We analyzed 103 blood serum samples collected during a state-wide survey from 2016 to 2020 and used a metagenomic approach for virus detection to identify known and previously uncharacterized virus sequences. We then characterized viruses phylogenetically and quantified prevalence, richness, and relative abundance for each virus. Within these viromes, we identified 19 viruses from 11 viral families: Amnoonviridae, Circoviridae, Coronaviridae, Hepadnaviridae, Peribunyaviridae, Picobirnaviridae, Picornaviridae, Matonaviridae, Narnaviridae, Nudnaviridae, and Spinareoviridae, 17 of which were previously undescribed. Among these viruses was the first fish-associated coronavirus from the Gammacoronavirus genus, which was present in 11/15 (73%) of S. vitreus. These results demonstrate that, similar to marine fish, freshwater fish also harbor diverse relatives of viruses important to the health of fish and other animals, although it currently remains unknown what effect, if any, the viruses we identified may have on fish health.

As deforestation progresses in the tropics, wildlife populations are increasingly restricted to forest fragments. Here we study genetic population structure in the endangered Ashy red colobus (Piliocolobus tephrosceles) population in the forest fragments surrounding Kibale National Park, Uganda. Subsequently, we use landscape features (elevation, road data and distance to the park) to design a feasible strategy to restore forest in a fashion suitable for both the dispersal patterns of the species and land use practices of the local people. A lack of association between geographic distance and pairwise genetic relatedness among localities, the presence of first degree relatives across localities, and a low global Fst value suggest that red colobus individuals have migrated across this landscape in the recent past. Thus, a series of “stepping stone” forests from the fragments to the park will likely maintain viability of red colobus fragment populations. In this area, low-lying valleys are legally protected to prevent flooding and are considered of low-economic value to local people. We identify such valleys for development of community-based forest restoration efforts that will aid in red colobus conservation and provide various ecosystem services. Our study outlines how genetics and community-based restoration can be integrated to provide realistic conservation solutions.

We report an unknown taxon of bot fly (Diptera: Oestridae: Oestrinae) in red river hogs (Potamochoerus porcus Linnaeus, 1758) in Cross River State, Nigeria. From direct observation and interviews with local hunters, we document that, remarkably, the parasite typically occurs within the intracranial supra-meningeal space – i.e., between the inner wall of the skull and the brain – but without causing visible inflammation or clinical signs. The parasite is most similar (up to 87.9%) to Rhinoestrus usbekistanicus based on cytochrome oxidase subunit 1 DNA sequencing but is sufficiently divergent phylogenetically to represent a new or previously un-sequenced taxon. Morphologically, the parasite shares some, but not all, features with R. nivarleti. Local cultural belief systems attribute aspects of red river hog behavior (e.g. intelligence, elusiveness) to the parasite, suggesting a prolonged presence in the red river hog population. The parasite's unusual anatomic location may be aberrant, or it may be a protective adaptation to life in red river hogs, which forage vigorously with their snouts.

Abstract Background Understanding how and why people interact with animals is important for the prevention and control of zoonoses. To date, studies have primarily focused on the most visible forms of human-animal contact (e.g., hunting and consumption), thereby blinding One Health researchers and practitioners to the broader range of human-animal interactions that can serve as cryptic sources of zoonotic diseases. Zootherapy, the use of animal products for traditional medicine and cultural practices, is widespread and can generate opportunities for human exposure to zoonoses. Existing research examining zootherapies omits details necessary to adequately assess potential zoonotic risks. Methods We used a mixed-methods approach, combining quantitative and qualitative data from questionnaires, key informant interviews, and field notes to examine the use of zootherapy in nine villages engaged in wildlife hunting, consumption, and trade in Cross River State, Nigeria. We analyzed medicinal and cultural practices involving animals from a zoonotic disease perspective, by including details of animal use that may generate pathways for zoonotic transmission. We also examined the sociodemographic, cultural, and environmental contexts of zootherapeutic practices that can further shape the nature and frequency of human-animal interactions. Results Within our study population, people reported using 44 different animal species for zootherapeutic practices, including taxonomic groups considered to be “high risk” for zoonoses and threatened with extinction. Variation in use of animal parts, preparation norms, and administration practices generated a highly diverse set of zootherapeutic practices (n = 292) and potential zoonotic exposure risks. Use of zootherapy was patterned by demographic and environmental contexts, with zootherapy more commonly practiced by hunting households (OR = 2.47, p

Pathogen surveillance for great ape health monitoring has typically been performed on non-invasive samples, primarily feces, in wild apes and blood in sanctuary-housed apes. However, many important primate pathogens, including known zoonoses, are shed in saliva and transmitted via oral fluids. Using metagenomic methods, we identified viruses in saliva samples from 46 wild-born, sanctuary-housed chimpanzees at two African sanctuaries in Republic of Congo and Uganda. In total, we identified 20 viruses. All but one, an unclassified CRESS DNA virus, are classified in five families: Circoviridae, Herpesviridae, Papillomaviridae, Picobirnaviridae, and Retroviridae. Overall, viral prevalence ranged from 4.2% to 87.5%. Many of these viruses are ubiquitous in primates and known to replicate in the oral cavity (simian foamy viruses, Retroviridae; a cytomegalovirus and lymphocryptovirus; Herpesviridae; and alpha and gamma papillomaviruses, Papillomaviridae). None of the viruses identified have been shown to cause disease in chimpanzees or, to our knowledge, in humans. These data suggest that the risk of zoonotic viral disease from chimpanzee oral fluids in sanctuaries may be lower than commonly assumed.

Freshwater mussels (Unionida) are globally imperiled, in part due to largely unexplained mass mortality events (MMEs). While recent studies have begun to investigate the possibility that mussel MMEs in the Eastern USA may be caused by infectious diseases, mussels in the Western USA have received relatively little attention in this regard. We conducted a two-year epidemiologic investigation of the role of viruses in ongoing MMEs of the Western pearlshell (Margaritifera falcata) and the Western ridged mussel (Gonidea angulata) in the Chehalis River and Columbia River watersheds in the Western USA. We characterized viromes of mussel hemolymph from 5 locations in 2018 and 2020 using metagenomic methods and identified 557 viruses based on assembled contiguous sequences, most of which are novel. We also characterized the distribution and diversity of a previously identified mussel Gammarhabdovirus related to pathogenic finfish viruses. Overall, we found few consistent associations between viruses and mussel health status. Variation in mussel viromes was most strongly driven by location, with little influence from date, species, or health status, though these variables together only explained ~1/3 of variation in virome composition. Our results demonstrate that Western freshwater mussels host remarkably diverse viromes, but no single virus or combination of viruses appears to be associated with morbidity or mortality during MMEs. Our findings have implications for the conservation of imperiled freshwater mussels, including efforts to enhance natural populations through captive propagation.

Infections with bacteria of the genus Sarcina are associated with gastric diseases of unclear etiology. Here, Owens et al. show that infection with a distinct Sarcina species is strongly associated with a lethal disease that affects sanctuary chimpanzees in Sierra Leone.

ABSTRACT White-nose syndrome (WNS), a fungal disease that has caused catastrophic population declines of bats in eastern North America, is rapidly spreading across the continent and now threatens previously unexposed bat species in western North America. The causal agent of WNS, the fungus Pseudogymnoascus destructans, can infect many species of hibernating bats, but susceptibility to WNS varies by host species. We previously reported that certain traits of the skin microbiome, particularly yeast diversity and abundance, of bat species in eastern North America are strongly associated with resistance to WNS. Using these traits, we developed models to predict WNS susceptibility of 13 species of western North American bats. Based on models derived from yeast species diversity, only one bat species, Myotis velifer, was predicted to be WNS resistant (i.e., may develop the disease, but with low mortality rates). We also screened yeasts found on western bats for P. destructans-antagonistic properties by spore germination and growth inhibition/competition assays and found the ability of yeasts to inhibit P. destructans in vitro to be strain specific. Similar to results of inhibition assays performed with yeasts isolated from bats in eastern North America, few yeasts isolated from bats in western North America inhibited P. destructans in vitro. Continued monitoring of western bat populations will serve to validate the accuracy of the mycobiome analysis in predicting WNS susceptibility, document population and susceptibility trends, and identify additional predictors to assess the vulnerability of naive bat populations to WNS. IMPORTANCE White-nose syndrome is one of the most devastating wildlife diseases ever documented. Some bat species are resistant to or tolerant of the disease, and we previously reported that certain traits of the skin mycobiome of bat species in eastern North America are strongly associated with resistance to WNS. Predicting which western bat species will be most susceptible to WNS would be of great value for establishing conservation priorities. Based on models derived from yeast species diversity, only one bat species was predicted to be WNS resistant. High susceptibility to WNS would pose a significant conservation threat to bats in western North America.

Freshwater mussels (Unionida) are among the world’s most imperiled taxa, but the relationship between freshwater mussel mortality events and infectious disease is largely unstudied. We surveyed viromes of a widespread and abundant species (mucket, Actinonaias ligamentina; syn: Ortmanniana ligamentina) experiencing a mortality event of unknown etiology in the Huron River, Michigan, in 2019–2020 and compared them to viromes from mucket in a healthy population in the St. Croix River, Wisconsin and a population from the Clinch River, Virginia and Tennessee, where a mortality event was affecting the congeneric pheasantshell (Actinonaias pectorosa; syn: Ortmanniana pectorosa) population. We identified 38 viruses, most of which were associated with mussels collected during the Huron River mortality event. Viral richness and cumulative viral read depths were significantly higher in moribund mussels from the Huron River than in healthy controls from each of the three populations. Our results demonstrate significant increases in the number and intensity of viral infections for freshwater mussels experiencing mortality events, whereas individuals from healthy populations have a substantially reduced virome comprising a limited number of species at low viral read depths.

ABSTRACTRespiratory viruses of human origin infect wild apes across Africa, sometimes lethally. Here we report simultaneous outbreaks of two distinct human respiratory viruses, human metapneumovirus (MPV; Pneumoviridae: Metapneumovirus) and human respirovirus 3 (HRV3; Paramyxoviridae; Respirovirus, formerly known as parainfluenza virus 3), in two chimpanzee (Pan troglodytes schweinfurthii) communities in the same forest in Uganda in December 2016 and January 2017. The viruses were absent before the outbreaks, but each was present in ill chimpanzees from one community during the outbreak period. Clinical signs and gross pathologic changes in affected chimpanzees closely mirrored symptoms and pathology commonly observed in humans for each virus. Epidemiologic modelling showed that MPV and HRV3 were similarly transmissible (R0 of 1.27 and 1.48, respectively), but MPV caused 12.2% mortality mainly in infants and older chimpanzees, whereas HRV3 caused no direct mortality. These results are consistent with the higher virulence of MPV than HRV3 in humans, although both MPV and HRV3 cause a significant global disease burden. Both viruses clustered phylogenetically within groups of known human variants, with MPV closely related to a lethal 2009 variant from mountain gorillas (Gorilla beringei beringei), suggesting two independent and simultaneous reverse zoonotic origins, either directly from humans or via intermediary hosts. These findings expand our knowledge of human origin viruses threatening wild chimpanzees and suggest that such viruses might be differentiated by their comparative epidemiological dynamics and pathogenicity in wild apes. Our results also caution against assuming common causation in coincident outbreaks.

A growing body of evidence demonstrates the importance of forests and wild animal-based foods for diets within tropical environments. However, deforestation and associated land-use changes can have competing effects on nutrition and food security as communities reorient from wild food use and subsistence-based agriculture to import/export markets. This research examines dietary differences and associated changes in food security during intermediate stages of deforestation and market integration in the agriculture-forest frontier of Cross River State, Nigeria. We used participant responses to mixed-methods interviews (n = 528) in six communities to measure individual dietary diversity, household food access, and short-term nutritional status, with specific attention to animal-based foods and the cultural and economic values attached to them, in two interior forest (n = 177) and four forest-edge (n = 351) communities. Multivariate analysis of dietary compositions revealed differences in food categories and types of meat consumed between forest environments. People in forest-edge communities reported consuming less bushmeat and dark green leafy vegetables, and more pulses, domestic meat, fish, eggs, dairy, other vegetables, sweets, condiments, and non-red palm oil compared to interior forest communities. Bushmeat was highly preferred and had more economic value than other animal-based foods, regardless of location. Forest-edge communities had fewer households involved in bushmeat related activities, and fewer hunters per household. However, traders in forest-edge communities sold a larger proportion of meat to people outside of the community than did traders in interior forest communities. Measures of nutrition and food security, but not wealth, improved in relation to dietary patterns in forest-edge communities compared to interior forest communities. Our results may reflect a “best of both worlds” scenario during the intermediate stages of deforestation and agricultural expansion near forested areas, where people have access to forest resources, increased ability to capitalize on forest goods, and access to market goods as they become integrated into market economies. Understanding the dietary consequences of environmental change is important, as food-related experiences may shape the trajectories of livelihood practices and landscape changes in tropical forests of biodiversity significance.

We describe a lethal respiratory outbreak among wild chimpanzees in Uganda in 2013 for which molecular and epidemiologic analyses implicate human rhinovirus C as the cause. Postmortem samples from an infant chimpanzee yielded near-complete genome sequences throughout the respiratory tract; other pathogens were absent. Epidemiologic modeling estimated the basic reproductive number (R0) for the epidemic as 1.83, consistent with the common cold in humans. Genotyping of 41 chimpanzees and examination of 24 published chimpanzee genomes from subspecies across Africa showed universal homozygosity for the cadherin-related family member 3 CDHR3-Y529 allele, which increases risk for rhinovirus C infection and asthma in human children. These results indicate that chimpanzees exhibit a species-wide genetic susceptibility to rhinovirus C and that this virus, heretofore considered a uniquely human pathogen, can cross primate species barriers and threatens wild apes. We advocate engineering interventions and prevention strategies for rhinovirus infections for both humans and wild apes.

Freshwater mussels (Unionida) are suffering mass mortality events worldwide, but the causes remain enigmatic. Here, we describe an analysis of bacterial loads, community structure, and inferred metabolic pathways in the hemolymph of pheasantshells (Actinonaias pectorosa) from the Clinch River, USA, during a multi-year mass mortality event. Bacterial loads were approximately 2 logs higher in moribund mussels (cases) than in apparently healthy mussels (controls). Bacterial communities also differed between cases and controls, with fewer sequence variants (SVs) and higher relative abundances of the proteobacteria Yokenella regensburgei and Aeromonas salmonicida in cases than in controls. Inferred bacterial metabolic pathways demonstrated a predominance of degradation, utilization, and assimilation pathways in cases and a predominance of biosynthesis pathways in controls. Only two SVs correlated with Clinch densovirus 1, a virus previously shown to be strongly associated with mortality in this system: Deinococcota and Actinobacteriota, which were associated with densovirus-positive and densovirus-negative mussels, respectively. Overall, our results suggest that bacterial invasion and shifts in the bacterial microbiome during unionid mass mortality events may result from primary insults such as viral infection or environmental stressors. If so, bacterial communities in mussel hemolymph may be sensitive, if generalized, indicators of declining mussel health.

Abstract Bats are natural reservoir hosts of highly virulent pathogens such as Marburg virus, Nipah virus, and SARS coronavirus. However, little is known about the role of bat ectoparasites in transmitting and maintaining such viruses. The intricate relationship between bats and their ectoparasites suggests that ectoparasites might serve as viral vectors, but evidence to date is scant. Bat flies, in particular, are highly specialized obligate hematophagous ectoparasites that incidentally bite humans. Using next-generation sequencing, we discovered a novel ledantevirus (mononegaviral family Rhabdoviridae, genus Ledantevirus) in nycteribiid bat flies infesting pteropodid bats in western Uganda. Mitochondrial DNA analyses revealed that both the bat flies and their bat hosts belong to putative new species. The coding-complete genome of the new virus, named Kanyawara virus (KYAV), is only distantly related to that of its closest known relative, Mount Elgon bat virus, and was found at high titers in bat flies but not in blood or on mucosal surfaces of host bats. Viral genome analysis indicates unusually low CpG dinucleotide depletion in KYAV compared to other ledanteviruses and rhabdovirus groups, with KYAV displaying values similar to rhabdoviruses of arthropods. Our findings highlight the possibility of a yet-to-be-discovered diversity of potentially pathogenic viruses in bat ectoparasites.

Bat flies (Hippoboscoidea: Nycteribiidae and Streblidae) are obligate hematophagous ectoparasites of bats. We collected streblid bat flies from the New World (México) and the Old World (Uganda), and used metagenomics to identify their viruses. In México, we found méjal virus (Rhabdoviridae; Vesiculovirus), Amate virus (Reoviridae: Orbivirus), and two unclassified viruses of invertebrates. Méjal virus is related to emerging zoonotic encephalitis viruses and to the agriculturally important vesicular stomatitis viruses (VSV). Amate virus and its sister taxon from a bat are most closely related to mosquito- and tick-borne orbiviruses, suggesting a previously unrecognized orbivirus transmission cycle involving bats and bat flies. In Uganda, we found mamucuso virus (Peribunyaviridae: Orthobunyavirus) and two unclassified viruses (a rhabdovirus and an invertebrate virus). Mamucuso virus is related to encephalitic viruses of mammals and to viruses from nycteribiid bat flies and louse flies, suggesting a previously unrecognized orthobunyavirus transmission cycle involving hippoboscoid insects. Bat fly virus transmission may be neither strictly vector-borne nor strictly vertical, with opportunistic feeding by bat flies occasionally leading to zoonotic transmission. Many “bat-associated” viruses, which are ecologically and epidemiologically associated with bats but rarely or never found in bats themselves, may actually be viruses of bat flies or other bat ectoparasites.

Simian hemorrhagic fever virus (SHFV) causes acute, lethal disease in macaques. We developed a single-plasmid cDNA-launch infectious clone of SHFV (rSHFV) and modified the clone to rescue an enhanced green fluorescent protein-expressing rSHFV-eGFP that can be used for rapid and quantitative detection of infection. SHFV has a narrow cell tropism in vitro, with only the grivet MA-104 cell line and a few other grivet cell lines being susceptible to virion entry and permissive to infection. Using rSHFV-eGFP, we demonstrate that one cricetid rodent cell line and three ape cell lines also fully support SHFV replication, whereas 55 human cell lines, 11 bat cell lines, and three rodent cells do not. Interestingly, some human and other mammalian cell lines apparently resistant to SHFV infection are permissive after transfection with the rSHFV-eGFP cDNA-launch plasmid. To further demonstrate the investigative potential of the infectious clone system, we introduced stop codons into eight viral open reading frames (ORFs). This approach suggested that at least one ORF, ORF 2b’, is dispensable for SHFV in vitro replication. Our proof-of-principle experiments indicated that rSHFV-eGFP is a useful tool for illuminating the understudied molecular biology of SHFV.

A Correction to this paper has been published: https://doi.org/10.1038/s41467-021-22494-5

We report the discovery and sequence-based molecular characterization of a novel virus, lanama virus (LNMV), in blood samples obtained from two wild vervet monkeys (Chlorocebus pygerythrus), sampled near Lake Nabugabo, Masaka District, Uganda. Sequencing of the complete viral genomes and subsequent phylogenetic analysis identified LNMV as a distinct member of species Kunsagivirus C, in the undercharacterized picornavirid genus Kunsagivirus.

We previously reported fatal infection of a captive Bornean orangutan with metacestodes of a novel taeniid tapeworm, Versteria sp. New data implicate mustelids as definitive hosts of these tapeworms in North America. At least 2 parasite genetic lineages circulate in North America, representing separate introductions from Eurasia.

Eggs of the lung fluke genus Paragonimus were detected in red-capped mangabeys (Cercocebus torquatus) in Nigeria. We assess the role of these primates as potential sylvatic hosts and the clinical effects of the parasite on monkeys. DNA sequenced from eggs in feces were 100% identical in the ITS2 region to Paragonimus africanus sequences from humans in Cameroon. Paragonimus-positive monkeys coughed more than uninfected monkeys. Experimental de-worming led to reduction in parasite intensity and a corresponding reduction of coughing to baseline levels in infected monkeys. This report provides the first evidence of Paragonimus sp. in C. torquatus, of P. africanus in Nigerian wildlife, and the first molecular evidence of the parasite in African wildlife. Coughing, sometimes interpreted as a communication behavior in primates, can actually indicate infection with lung parasites. Observations of coughing in primates may, in turn, provide a useful mechanism for surveillance of Paragonimus spp, which are re-emerging human pathogens, in wildlife reservoirs.

Pteropine orthoreovirus (PRV; Reoviridae: Spinareovirinae) is an emerging bat-borne zoonotic virus that causes influenza-like illness (ILI). PRV has thus far been found only in Australia and Asia, where diverse old-world fruit bats (Pteropodidae) serve as hosts. In this study, we report the discovery of PRV in Africa, in an Angolan soft-furred fruit bat (Lissonycteris angolensis ruwenzorii) from Bundibugyo District, Uganda. Metagenomic characterization of a rectal swab yielded 10 dsRNA genome segments, revealing this virus to cluster within the known diversity of PRV variants detected in bats and humans in Southeast Asia. Phylogeographic analyses revealed a correlation between geographic distance and genetic divergence of PRVs globally, which suggests a geographic continuum of PRV diversity spanning Southeast Asia to sub-Saharan Africa. The discovery of PRV in an African bat dramatically expands the geographic range of this zoonotic virus and warrants further surveillance for PRVs outside of Southeast Asia.

Obligate hematophagous ectoparasitic flies of the superfamily Hippoboscoidea are distributed worldwide, but their role as vectors and reservoirs of viruses remains understudied. We examined hippoboscoid bat flies (family Nycteribiidae) parasitizing Angolan soft-furred fruit bats (Lissonycteris angolensis ruwenzorii) from Bundibugyo District, Uganda. Using metagenomic methods, we detected 21 variants of the rhabdovirid genus Ledantevirus, which contains medically important “bat-associated” viruses. These 21 viruses, representing at least two divergent viral lineages, infected 26 bat flies from 8 bats in a single roost. Cophylogenetic analyses of viruses and bat flies resulted in strong evidence of virus-host codivergence, indicating vertical transmission of bat fly ledanteviruses. Examination of oral swabs from bats revealed ledantevirus RNA in the saliva of 1 out of 11 bats, with no evidence of insect genetic material in the mouth of this bat. These data demonstrate that bat flies can harbor diverse ledanteviruses even in a single roost and that the predominant mode of transmission is likely vertical (among bat flies), but that bats can become infected and shed viruses orally. In conclusion, bat flies may serve as ectoparasitic reservoirs of “bat-associated” viruses that only transiently or sporadically infect bats.

Respiratory illnesses have caused significant mortality in African great ape populations. While much effort has been given to identifying the responsible pathogens, little is known about the factors that influence disease transmission or individual susceptibility. In the Kanyawara community of wild chimpanzees, respiratory illness has been the leading cause of mortality over 31 years, contributing to 27% of deaths. Deaths were common in all age groups except juveniles. Over 22 years of health observations, respiratory signs were rare among infants and most common among older adults of both sexes. Respiratory signs were also common among males during the transition to adulthood (ages 10–20 years), particularly among those of low rank. Respiratory signs peaked conspicuously in March, a pattern that we could not explain after modelling climatic factors, group sizes, diet or exposure to humans. Furthermore, rates of respiratory illness in the chimpanzees did not track seasonal rates of illness in the nearby village. Our data indicate that the epidemiology of chimpanzee respiratory illness warrants more investigation but clearly differs in important ways from humans. Findings on individual susceptibility patterns suggest that respiratory signs are a robust indicator for investigating immunocompetence in wild chimpanzees.

A captive juvenile Bornean orangutan (Pongo pygmaeus) died from an unknown disseminated parasitic infection. Deep sequencing of DNA from infected tissues, followed by gene-specific PCR and sequencing, revealed a divergent species within the newly proposed genus Versteria (Cestoda: Taeniidae). Versteria may represent a previously unrecognized risk to primate health.

Foamy viruses (FVs) are complex retroviruses present in many mammals, including nonhuman primates, where they are called simian foamy viruses (SFVs). SFVs can zoonotically infect humans, but very few complete SFV genomes are available, hampering the design of diagnostic assays. Gibbons are lesser apes widespread across Southeast Asia that can be infected with SFV, but only two partial SFV sequences are currently available. We used a metagenomics approach with next-generation sequencing of nucleic acid extracted from the cell culture of a blood specimen from a lesser ape, the pileated gibbon (Hylobates pileatus), to obtain the complete SFVhpi_SAM106 genome. We used Bayesian analysis to co-infer phylogenetic relationships and divergence dates. SFVhpi_SAM106 is ancestral to other ape SFVs with a divergence date of ~20.6 million years ago, reflecting ancient co-evolution of the host and SFVhpi_SAM106. Analysis of the complete SFVhpi_SAM106 genome shows that it has the same genetic architecture as other SFVs but has the longest recorded genome (13,885-nt) due to a longer long terminal repeat region (2,071 bp). The complete sequence of the SFVhpi_SAM106 genome fills an important knowledge gap in SFV genetics and will facilitate future studies of FV infection, transmission, and evolutionary history.

ABSTRACT Simian hemorrhagic fever (SHF) is a highly lethal disease in captive macaques. Three distinct arteriviruses are known etiological agents of past SHF epizootics, but only one, simian hemorrhagic fever virus (SHFV), has been isolated in cell culture. The natural reservoir(s) of the three viruses have yet to be identified, but African nonhuman primates are suspected. Eleven additional divergent simian arteriviruses have been detected recently in diverse and apparently healthy African cercopithecid monkeys. Here, we report the successful isolation in MARC-145 cell culture of one of these viruses, Kibale red colobus virus 1 (KRCV-1), from serum of a naturally infected red colobus (Procolobus [Piliocolobus] rufomitratus tephrosceles) sampled in Kibale National Park, Uganda. Intramuscular (i.m.) injection of KRCV-1 into four cynomolgus macaques (Macaca fascicularis) resulted in a self-limiting nonlethal disease characterized by depressive behavioral changes, disturbance in coagulation parameters, and liver enzyme elevations. In contrast, i.m. injection of SHFV resulted in typical lethal SHF characterized by mild fever, lethargy, lymphoid depletion, lymphoid and hepatocellular necrosis, low platelet counts, increased liver enzyme concentrations, coagulation abnormalities, and increasing viral loads. As hypothesized based on the genetic and presumed antigenic distance between KRCV-1 and SHFV, all four macaques that had survived KRCV-1 injection died of SHF after subsequent SHFV injection, indicating a lack of protective heterotypic immunity. Our data indicate that SHF is a disease of macaques that in all likelihood can be caused by a number of distinct simian arteriviruses, although with different severity depending on the specific arterivirus involved. Consequently, we recommend that current screening procedures for SHFV in primate-holding facilities be modified to detect all known simian arteriviruses. IMPORTANCE Outbreaks of simian hemorrhagic fever (SHF) have devastated captive Asian macaque colonies in the past. SHF is caused by at least three viruses of the family Arteriviridae: simian hemorrhagic fever virus (SHFV), simian hemorrhagic encephalitis virus (SHEV), and Pebjah virus (PBJV). Nine additional distant relatives of these three viruses were recently discovered in apparently healthy African nonhuman primates. We hypothesized that all simian arteriviruses are potential causes of SHF. To test this hypothesis, we inoculated cynomolgus macaques with a highly divergent simian arterivirus (Kibale red colobus virus 1 [KRCV-1]) from a wild Ugandan red colobus. Despite being only distantly related to red colobuses, all of the macaques developed disease. In contrast to SHFV-infected animals, KRCV-1-infected animals survived after a mild disease presentation. Our study advances the understanding of an important primate disease. Furthermore, our data indicate a need to include the full diversity of simian arteriviruses in nonhuman primate SHF screening assays.

The local abundance of Culex mosquitoes is a central factor adding to the risk of West Nile virus transmission, and vector abundance data influence public health decisions. This study evaluated differences in abundance estimates from mosquitoes trapped using two common methods: CO 2 - baited CDC light traps and infusion-baited gravid traps in suburban, Chicago, Illinois. On a weekly basis, the two methods were modestly correlated ( r = 0.219) across 71 weeks over 4 years. Lagged weather conditions of up to four weeks were associated with the number of mosquitoes collected in light and gravid traps. Collections in light traps were higher with higher temperature in the same week, higher precipitation one, two, and four weeks before the week of trapping, and lower maximum average wind speed. Collections in gravid traps were higher with higher temperature in the same week and one week earlier, lower temperature four weeks earlier, and with higher precipitation two and four weeks earlier. Culex abundance estimates from light traps were significantly higher in semi-natural areas compared to residential areas, but abundance estimates from gravid traps did not vary by the landscape type. These results highlight the importance of the surveillance methods used in the assessment of local Culex abundance estimates. Measures of risk of exposure to West Nile virus should assess carefully how mosquito abundance has been estimated and integrated into assessments of transmission risk.

Bird-facilitated introduction of ticks and associated pathogens is postulated to promote invasion of tick-borne zoonotic diseases into urban areas. Results of a longitudinal study conducted in suburban Chicago, Illinois, USA, during 2005–2010 show that 1.6% of 6,180 wild birds captured in mist nets harbored ticks. Tick species in order of abundance were Haemaphysalis leporispalustris, Ixodes dentatus, and I. scapularis, but 2 neotropical tick species of the genus Amblyomma were sampled during the spring migration. I. scapularis ticks were absent at the beginning of the study but constituted the majority of ticks by study end and were found predominantly on birds captured in areas designated as urban green spaces. Of 120 ticks, 5 were infected with Borrelia burgdorferi, spanning 3 ribotypes, but none were infected with Anaplasma phagocytophilum. Results allow inferences about propagule pressure for introduction of tick-borne diseases and emphasize the large sample sizes required to estimate this pressure.

We conducted a prospective study of bacterial transmission among humans, nonhuman primates (primates hereafter), and livestock in western Uganda. Humans living near forest fragments harbored Escherichia coli bacteria that were ≈75% more similar to bacteria from primates in those fragments than to bacteria from primates in nearby undisturbed forests. Genetic similarity between human/livestock and primate bacteria increased ≈3-fold as anthropogenic disturbance within forest fragments increased from moderate to high. Bacteria harbored by humans and livestock were approximately twice as similar to those of red-tailed guenons, which habitually enter human settlements to raid crops, than to bacteria of other primate species. Tending livestock, experiencing gastrointestinal symptoms, and residing near a disturbed forest fragment increased genetic similarity between a participant’s bacteria and those of nearby primates. Forest fragmentation, anthropogenic disturbance within fragments, primate ecology, and human behavior all influence bidirectional, interspecific bacterial transmission. Targeted interventions on any of these levels should reduce disease transmission and emergence.

Enzyme-linked immunosorbent assay, Western blot, and virus neutralization assays indicated that red colobus monkeys in Kibale National Park, western Uganda, had antibodies to a virus that was similar, but not identical, to known orthopoxviruses. The presence of a novel poxvirus in this endangered primate raises public health and conservation concerns.