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Astroviruses (AstVs) infect numerous mammalian species including reservoirs such as bats. Peptides encoded by the genes of the highly polymorphic Major Histocompatibility Complex (MHC) form the first line of host defence against pathogens. Aside from direct involvement in mounting adaptive immune responses, MHC class II genes are hypothesized to regulate gut commensal diversity and shape the production of immune‐modulatory substances by microbes, indirectly affecting host susceptibility. Despite initial empirical evidence for the link between host MHC and the microbiota, associations among these factors remain largely unknown. To fill this gap, we examined MHC allelic diversity and constitution, the gut bacterial community and abundance pattern of a wild population of a neotropical bat (Artibeus jamaicensis) challenged by AstV infections. First, we show an age‐dependent relationship between the host MHC class II diversity and constitution and the gut microbiota in AstV‐uninfected bats. Crucially, these associations changed in AstV‐infected bats. Additionally, we identify changes in the abundance of specific bacterial taxa linked to the presence of certain MHC supertypes and AstV infection. We suggest changes in the microbiota to be either a result of AstV infection or the MHC‐mediated modulation of microbial communities. The latter could subsequently affect microbe‐mediated immunity and resistance against AstV infection. Our results emphasize that the reciprocal nature of host immune genetics, gut microbial diversity and pathogen infection require attention, which are particularly important given their repercussions for disease susceptibility and severity in wild animal populations with a history of zoonotic spillover and frequent human contact., publishedVersion

Shrews, insectivorous small mammals, pertain to an ancient mammalian order. We screened 693 European and African shrews for hepatitis B virus (HBV) homologs to elucidate the enigmatic genealogy of HBV. Shrews host HBVs at low prevalence (2.5%) across a broad geographic and host range. The phylogenetically divergent shrew HBVs comprise separate species termed crowned shrew HBV (CSHBV) and musk shrew HBV (MSHBV), each containing distinct genotypes. Recombination events across host orders, evolutionary reconstructions, and antigenic divergence of shrew HBVs corroborated ancient origins of mammalian HBVs dating back about 80 million years. Resurrected CSHBV replicated in human hepatoma cells, but human- and tupaia-derived primary hepatocytes were resistant to hepatitis D viruses pseudotyped with CSHBV surface proteins. Functional characterization of the shrew sodium taurocholate cotransporting polypeptide (Ntcp), CSHBV/MSHBV surface peptide binding patterns, and infection experiments revealed lack of Ntcp-mediated entry of shrew HBV. Contrastingly, HBV entry was enabled by the shrew Ntcp. Shrew HBVs universally showed mutations in their genomic preCore domains impeding hepatitis B e antigen (HBeAg) production and resembling those observed in HBeAg-negative human HBV. Deep sequencing and in situ hybridization suggest that HBeAg-negative shrew HBVs cause intense hepatotropic monoinfections and low within-host genomic heterogeneity. Geographical clustering and low MSHBV/CSHBV-specific seroprevalence suggest focal transmission and high virulence of shrew HBVs. HBeAg negativity is thus an ancient HBV infection pattern, whereas Ntcp usage for entry is not evolutionarily conserved. Shrew infection models relying on CSHBV/MSHBV revertants and human HBV will allow comparative assessments of HBeAg-mediated HBV pathogenesis, entry, and species barriers.

The tent-making bat hepatitis B virus (TBHBV) is a hepadnavirus closely related to human hepatitis B virus. The ecology of TBHBV is unclear. We show that it is widespread and highly diversified in Peters’ tent-making bats (Uroderma bilobatum) within Panama, while local prevalence varied significantly between sample sites, ranging from 0 to 14.3%. Females showed significantly higher prevalence than males, and pregnant females were more often acutely infected than non-reproductive ones. The distribution of TBHBV in bats was significantly affected by forest cover, with higher infection rates in areas with lower forest cover. Our data indicate that loss of natural habitat may lead to positive feedback on the biotic factors driving infection possibility. These results underline the necessity of multidisciplinary studies for a better understanding of mechanisms in pathogen–host relationships and for predictions in disease ecology. Electronic supplementary material The online version of this article (10.1007/s10393-018-1387-5) contains supplementary material, which is available to authorized users.

Significance Hepatitis delta virus (HDV) aggravates hepatitis B virus (HBV) infection of liver cells. Although the viruses are evolutionarily unrelated, HDV depends on HBV because it requires the HBV envelope protein for its transmission. HDV is only described in humans, which has triggered diverse hypotheses regarding its evolution and origins. Here we show that spiny rats (Proechimys semispinosus) carry a counterpart to HDV that surprisingly does not cause hepatitis and is not linked to HBV. The rodent deltavirus finding alone, but also taken together with the recent deltavirus findings in snakes and other vertebrates and invertebrates, suggests that a deltavirus precursor may have infected mammals before it acquired dependence on HBV as seen in humans., Hepatitis delta virus (HDV) is a human hepatitis-causing RNA virus, unrelated to any other taxonomic group of RNA viruses. Its occurrence as a satellite virus of hepatitis B virus (HBV) is a singular case in animal virology for which no consensus evolutionary explanation exists. Here we present a mammalian deltavirus that does not occur in humans, identified in the neotropical rodent species Proechimys semispinosus. The rodent deltavirus is highly distinct, showing a common ancestor with a recently described deltavirus in snakes. Reverse genetics based on a tandem minus-strand complementary DNA genome copy under the control of a cytomegalovirus (CMV) promoter confirms autonomous genome replication in transfected cells, with initiation of replication from the upstream genome copy. In contrast to HDV, a large delta antigen is not expressed and the farnesylation motif critical for HBV interaction is absent from a genome region that might correspond to a hypothetical rodent large delta antigen. Correspondingly, there is no evidence for coinfection with an HBV-related hepadnavirus based on virus detection and serology in any deltavirus-positive animal. No other coinfecting viruses were detected by RNA sequencing studies of 120 wild-caught animals that could serve as a potential helper virus. The presence of virus in blood and pronounced detection in reproductively active males suggest horizontal transmission linked to competitive behavior. Our study establishes a nonhuman, mammalian deltavirus that occurs as a horizontally transmitted infection, is potentially cleared by immune response, is not focused in the liver, and possibly does not require helper virus coinfection.

Toll-like receptors (TLRs) form part of the innate immune system and can recognize structurally conserved pathogen-associated molecular pattern (PAMP) molecules. Their functional importance in the resistance to pathogens has been documented in laboratory experimental settings and in humans. TLR diversity, however, has been rarely investigated in wildlife species. How the genetic diversity of TLRs is associated with various pathogens and how it is shaped by habitat disturbance are understudied. Therefore, we investigated the role of genetic diversity in the functionally important parts of TLR4 and TLR7 genes in resistance towards gastrointestinal nematodes and Hepacivirus infection. We chose a generalist study species, the rodent Proechimys semispinosus, because it is highly abundant in three Panamanian landscapes that differ in their degree of anthropogenic modification. We detected only two TLR7 haplotypes that differed by one synonymous single-nucleotide polymorphism (SNP) position. The TLR4 variability was higher, and we detected four TLR4 haplotypes that differed at one synonymous SNP and at three amino acid positions within the leucine-rich repeat region. Only TLR4 haplotypes had different frequencies in each landscape. Using generalized linear models, we found evidence that nematode loads and virus prevalence were influenced by both specific TLR4 haplotypes and landscape. Here, the variable “landscape” served as a surrogate for the important influential ecological factors distinguishing landscapes in our study, i.e. species diversity and host population density. Individuals carrying the common TLR4_Ht1 haplotype were less intensely infected by the most abundant strongyle nematode. Individuals carrying the rare TLR4_Ht3 haplotype were all Hepacivirus-positive, where those carrying the rare haplotype TLR4_Ht4 were less often infected by Hepacivirus than individuals with other haplotypes. Our study highlights the role of TLR diversity in pathogen resistance and the importance of considering immune genetic as well as ecological factors in order to understand the effects of anthropogenic changes on wildlife health., publishedVersion

Anthropogenic environmental change can impact community and population traits such as species diversity and population densities, which have been shown to influence the prevalence of viruses in wildlife reservoirs. In particular, host species resilient to changes in their natural habitat may increase in numbers, which in turn can affect the prevalence of directly transmitted viruses. We have carried out a survey of small mammal communities in three tropical landscapes differing in their degree of environmental change in Central Panama and investigated the effects of community changes on Hepacivirus prevalence. The modification of continuous habitat into partly connected or isolated habitat patches during the past century was linked to changes in species diversity and species assemblages, which was further associated with shifts in the abundance of generalist marsupial (Didelphis marsupialis, Philander opossum) and rodent (Proechimys semispinosus) species. The latter has become dominant in isolated habitat patches and was the only identified Hepacivirus host in our study system. Our analyses suggest that, in addition to the effects of host age and sex, host population density in interaction with sex ratio is a crucial predictor of infection probability. Although we found no significant relationships between species diversity per se and infection probability, the lowest prevalence detected in the landscape with the highest species diversity indicates that shifts in species assemblages (e.g. changes in the presence and abundance of marsupial predators) impact the host's intraspecific contact rates, the probability of virus transmission and, thus, the virus prevalence. Our study additionally provides important data on the influence of human-induced landscape changes on infection probability and, therefore, on virus prevalence in wildlife and emphasizes the importance of a landscape-scale approach with concomitant consideration of the complex interactions between ecological factors.

Sloths are genetically and physiologically divergent mammals. Phleboviruses are major arthropod‐borne viruses (arboviruses) causing disease in humans and other animals globally. Sloths host arboviruses, but virus detections are scarce. A phlebovirus termed Anhanga virus (ANHV) was isolated from a Brazilian Linnaeus's two‐toed sloth (Choloepus didactylus) in 1962. Here, we investigated the presence of phleboviruses in sera sampled in 2014 from 74 Hoffmann's two‐toed (Choloepus hoffmanni, n = 65) and three‐toed (Bradypus variegatus, n = 9) sloths in Costa Rica by broadly reactive RT‐PCR. A clinically healthy adult Hoffmann's two‐toed sloth was infected with a phlebovirus. Viral load in this animal was high at 8.5 × 107 RNA copies/ml. The full coding sequence of the virus was determined by deep sequencing. Phylogenetic analyses and sequence distance comparisons revealed that the new sloth virus, likely representing a new phlebovirus species, provisionally named Penshurt virus (PEHV), was most closely related to ANHV, with amino acid identities of 93.1%, 84.6%, 94.7% and 89.0% in the translated L, M, N and NSs genes, respectively. Significantly more non‐synonymous mutations relative to ANHV occurred in the M gene encoding the viral glycoproteins and in the NSs gene encoding a putative interferon antagonist compared to L and N genes. This was compatible with viral adaptation to different sloth species and with micro‐evolutionary processes associated with immune evasion during the genealogy of sloth‐associated phleboviruses. However, gene‐wide mean dN/dS ratios were low at 0.02–0.15 and no sites showed significant evidence for positive selection, pointing to comparable selection pressures within sloth‐associated viruses and genetically related phleboviruses infecting hosts other than sloths. The detection of a new phlebovirus closely‐related to ANHV, in sloths from Costa Rica fifty years after and more than 3,000 km away from the isolation of ANHV confirmed the host associations of ANHV‐related phleboviruses with the two extant species of two‐toed sloths.

To examine the diagnostic performance of real-time reverse transcription (RT)-polymerase chain reaction (PCR) assays for Zika virus detection. We compared seven published real-time RT-PCR assays and two new assays that we have developed. To determine the analytical sensitivity of each assay, we constructed a synthetic universal control ribonucleic acid (uncRNA) containing all of the assays' target regions on one RNA strand and spiked human blood or urine with known quantities of African or Asian Zika virus strains. Viral loads in 33 samples from Zika virus-infected patients were determined by using one of the new assays. Oligonucleotides of the published real-time RT-PCR assays, showed up to 10 potential mismatches with the Asian lineage causing the current outbreak, compared with 0 to 4 mismatches for the new assays. The 95% lower detection limit of the seven most sensitive assays ranged from 2.1 to 12.1 uncRNA copies/reaction. Two assays had lower sensitivities of 17.0 and 1373.3 uncRNA copies/reaction and showed a similar sensitivity when using spiked samples. The mean viral loads in samples from Zika virus-infected patients were 5 × 104 RNA copies/mL of blood and 2 × 104 RNA copies/mL of urine. We provide reagents and updated protocols for Zika virus detection suitable for the current outbreak strains. Some published assays might be unsuitable for Zika virus detection, due to the limited sensitivity and potential incompatibility with some strains. Viral concentrations in the clinical samples were close to the technical detection limit, suggesting that the use of insensitive assays will cause false-negative results. Étudier la performance diagnostique des tests basés sur l'amplification en chaîne par polymérase (PCR) en temps réel après transcription inverse (RT) pour détecter le virus Zika. Nous avons comparé sept tests publiés utilisant la RT-PCR en temps réel et deux nouveaux tests développés par nos soins. Afin de déterminer la sensibilité analytique de chaque test, nous avons conçu un acide ribonucléique synthétique de contrôle universel (ARNcun) contenant toutes les régions ciblées par les tests sur une hélice ARN et enrichi de l’urine ou du sang humain de quantités connues de souches africaines ou asiatiques du virus Zika. Les charges virales dans 33 échantillons provenant de patients infectés par le virus Zika ont été déterminées à l'aide de l'un des nouveaux tests. Les oligonucléotides des tests publiés utilisant la RT-PCR en temps réel ont présenté jusqu'à 10 mauvais appariements potentiels avec la lignée asiatique provoquant l'épidémie actuelle, alors qu'on a constaté de 0 à 4 mauvais appariements dans le cas des nouveaux tests. La limite de détection inférieure à 95% des sept tests les plus sensibles variait de 2,1 à 12,1 copies d'ARNcun/réaction. Deux tests présentaient des sensibilités plus basses de 17,0 et 1373,3 copies d'ARNcun/réaction et montraient une sensibilité similaire lorsque des échantillons enrichis étaient utilisés. Les charges virales moyennes dans les échantillons provenant de patients infectés par le virus Zika étaient de 5 × 104 copies d'ARN/mL de sang et de 2 × 104 copies d'ARN/mL d'urine. Nous proposons pour détecter le virus Zika des réactifs et des protocoles actualisés, adaptés aux souches responsables de la flambée actuelle. Tous les tests publiés ne permettent pas de détecter le virus Zika en raison d'une sensibilité limitée et d'une incompatibilité potentielle avec certaines souches. Les concentrations virales dans les échantillons cliniques étaient proches de la limite de détection technique, ce qui laisse penser que l'utilisation de tests insensibles donnera des résultats faussement négatifs. Examinar el rendimiento del diagnóstico de las pruebas de reacción en cadena de la polimerasa de transcriptasa inversa (RT-PCR) en tiempo real para la detección del virus de Zika. Se compararon siete pruebas de RT-PCR en tiempo real publicadas y dos pruebas nuevas que se han desarrollado. Con el fin de determinar la sensibilidad analítica de cada prueba, se construyó un ácido ribonucleico sintético de control universal (uncRNA) que contenía todas las regiones objetivo de las pruebas en una hebra de RNA y se añadió a sangre u orina humana con cantidades conocidas de cepas de virus de Zika de Asia o África. Se determinaron las cargas víricas en 33 muestras procedentes de pacientes infectados por el virus de Zika a través de una de las pruebas nuevas. Los oligonucleótidos de las pruebas de RT-PCR en tiempo real publicadas mostraron hasta 10 posibles discordancias con el linaje de Asia causante de los brotes actuales, en comparación con 0 de 4 discordancias en el caso de las pruebas nuevas. El límite de detección inferior del 95% de las siete pruebas más sensibles abarcó de 2,1 a 12,1 copias/reacción de uncRNA. Dos pruebas mostraron sensibilidades inferiores de 17,0 y 1373,3 copias/reacción de uncRNA y presentaron una sensibilidad similar al usar muestras infectadas. Las cargas víricas medias en las muestras procedentes de pacientes infectados por el virus de Zika fueron de 5 × 104 copias de RNA/mL de sangre y de 2 × 104 copias de RNA/ml de orina. Se proporcionan reactivos y protocolos actualizados para la detección adecuada del virus de Zika en el caso de las cepas de brotes actuales. Algunas pruebas publicadas pueden no ser adecuadas para la detección del virus de Zika, debido a la limitada sensibilidad y a la posible incompatibilidad con algunas cepas. Las concentraciones víricas en las muestras clínicas se acercaron al límite de detección técnico, lo que sugería que el uso de pruebas intensivas causaría resultados falsos negativos. دراسة الأداء التشخيصي لاختبارات تفاعل البوليميراز المتسلسل اللحظي (PCR) مع إنزيم النسخ العكسي (RT) لاكتشاف الإصابة بفيروس زيكا. قمنا بإجراء مقارنة بين سبعة اختبارات منشورة لتفاعل البوليميراز المتسلسل اللحظي مع إنزيم النسخ العكسي (RT‑PCR) واختبارين جديدين تم إعدادهما من جانبنا. ولتحديد الحساسية التحليلية لكل اختبار، قمنا بتكوين الحمض النووي الريبوزي الاصطناعي للمكافحة العامة (uncRNA) والذي يحتوي على جميع المناطق المستهدفة في الاختبار في شريط RNA واحد ويحتوي على مؤشرات للدم أو البول البشري الذي يحتوي على كميات معروفة من سلالات فيروس زيكا في منطقة أفريقيا أو آسيا. وتم تحديد الحمولات الفيروسية في 33 عينة صادرة من مرضى مصابين بفيروس زيكا باستخدام أحد الاختبارات الجديدة. أظهرت الأوليغونيكليوتيدات الخاصة باختبارات تفاعل البوليميراز المتسلسل اللحظي مع إنزيم النسخ العكسي ما يصل إلى 10 حالات محتملة من عدم التطابق مع السلالة الآسيوية المسببة للعدوى الحالية، بالمقارنة مع حالات عدم التطابق بالنسبة للاختبارات الجديدة بمعدل 0 إلى 4. تراوح حد الاكتشاف المنخفض بنسبة 95‏% في الاختبارات السبعة الأكثر حساسية من 2.1 إلى 12.1 من نسخ سلالات uncRNA لتفاعل كل حالة. وُجد لدى اثنين من الاختبارات درجات منخفضة من الحساسية بمقدار 17.0 و1373.3 لنسخ سلالات uncRNA لتفاعل كل حالة وأظهرت وجود درجة حساسية مماثلة عند استخدام عينات المؤشرات. بلغ متوسط الحمولات الفيروسية في العينات الصادرة عن المرضى المصابين بفيروس زيكا 5 × 10 4 نسخ من RNA لكل ملليلتر من الدم و 2 × 10 4 نسخ من RNA لكل ملليلتر من البول. نحن نوفر الكاشفات وأحدث البروتوكولات لاكتشاف فيروس زيكا والتي تناسب سلالات العدوى الحالية. وقد تكون بعض الاختبارات المنشورة غير مناسبة لاكتشاف فيروس زيكا نظرًا لوجود درجة محدودة من الحساسية واحتمالية عدم توافقها مع بعض السلالات. كانت نسب تركيز الفيروسات في العينات التحليلية قريبة من حد الاكتشاف التقني، مما يشير إلى أن استخدام الاختبارات التي تفتقد إلى الحساسية سيؤدي إلى إصدار نتائج خاطئة وسلبية. 旨在检查针对寨卡病毒检测的实时逆转录 (RT)-聚合酶链反应 (PCR) 试验的诊断性能。. 我们比较了 7 种公布的实时 RT-PCR 试验和我们开发的两种新试验。 为了确定各种试验的分析灵敏度，我们构建了在一个 RNA 链上包含所有试验目标区域的合成通用对照核糖核酸 (uncRNA) 以及带有已知数量的非洲或亚洲寨卡病毒株的加标人体血液或尿液。 通过使用一种新的试验测定 33 个寨卡病毒感染患者样本的病毒载量。. 已公布的实时 RT-PCR 试验的寡核苷酸，显示出与亚洲血统潜在的失配率高达 10，相比之下，新试验的失配率为 0 至 4。 七种最敏感试验的 95% 检测下限范围为 2.1 至 12.1 uncRNA 拷贝/反应。 两个试验中 17.0 和 1373.3 uncRNA 复制/反应的灵敏度较低，使用加标样本时表现出类似的灵敏度。 寨卡病毒感染患者的样本中，血液的平均病毒载量为 5 × 104 RNA 拷贝/毫升，尿液的平均病毒载量为 2 × 104 拷贝/毫升。. 我们为适合当前疫情的寨卡病毒检测提供试剂和更新方案。 由于有限的灵敏度以及与一些菌株潜在的不兼容性，因此一些公布的试验可能不适合寨卡病毒检测。 临床样本中的病毒含量接近于技术检测极限，这表明使用不敏感试验将会引起假阴性结果。. Изучить диагностические возможности выявления вируса Зика с помощью полимеразной цепной реакции (ПЦР) с обратной транскриптазой (ОТ) в режиме реального времени. Авторы сравнили семь методов ОТ-ПЦР, сведения о которых были опубликованы в литературе, и два новых метода, разработанные авторами. Для выявления аналитической чувствительности каждого метода были сконструированы синтетические универсальные контрольные рибонуклеиновые кислоты (ункРНК), содержащие все целевые участки на одной из цепей РНК. Эти РНК были добавлены к образцам крови или мочи пациентов, содержащих вирус Зика африканского или азиатского происхождения в известных количествах. Одним из новых методов определялась вирусная нагрузка для 33 образцов пациентов, зараженных вирусом Зика. Олигонуклеотиды для опубликованных методов ОТ-ПЦР показали до 10 потенциальных несовпадений при определении вируса азиатского происхождения, вызвавшего нынешнюю вспышку заболеваемости, тогда как новые методы показали от 0 до 4 несовпадений. Для семи наиболее чувствительных анализов 95%-й нижний порог определения составлял от 2,1 до 12,1 копии ункРНК на реакцию. Два метода продемонстрировали меньшую чувствительность (от 17,0 до 1373,3 копии ункРНК на анализ) и сходную чувствительность при использовании образцов с добавлением ункРНК. Средняя величина вирусной нагрузки в образцах пациентов, инфицированных вирусом Зика, составила 5 × 104 копий РНК/мл для крови и 2 × 104 копийРНК/мл для мочи. Авторы предлагают реактивы и обновленные протоколы для выявления вируса Зика, вызвавшего нынешнюю вспышку заболевания. Некоторые из методов, по которым имеются опубликованные литературные данные, могут быть неподходящими для текущей ситуации из-за ограниченной чувствительности и потенциальной несовместимости с некоторыми штаммами вируса. Концентрация вируса в клинических образах была близка к техническому пределу определения, что позволяет предположить, что использование нечувствительных методов может приводить к получению ложноотрицательных результатов

Graphical abstract, Summary Hepatitis viruses are major threats to human health. During the last decade, highly diverse viruses related to human hepatitis viruses were found in animals other than primates. Herein, we describe both surprising conservation and striking differences of the unique biological properties and infection patterns of human hepatitis viruses and their animal homologues, including transmission routes, liver tropism, oncogenesis, chronicity, pathogenesis and envelopment. We discuss the potential for translation of newly discovered hepatitis viruses into preclinical animal models for drug testing, studies on pathogenesis and vaccine development. Finally, we re-evaluate the evolutionary origins of human hepatitis viruses and discuss the past and present zoonotic potential of their animal homologues.

Astroviruses (AstV) are a major cause of diarrhoea in children. Interestingly, some wildlife species, including bats, remain phenotypically asymptomatic after infection. Disease symptoms, however, may only be less visible in bats and enteric viruses may indeed perturb their gut microbial communities. Gut microbiomes represent an important driver of immune defence mechanisms but potential effects of enteric virus-host microbiome interactions are largely unexplored. Using bats as a natural model system, we show that AstV-infections affect the gut microbiome, with the strength of the effect depending on host age. The gut microbial α- and β-diversity and the predicted microbial functional orthologs decreased in young bats but surprisingly increased in adult AstV + bats. The abundance of bacterial taxa characteristic for healthy microbiomes was strongly reduced in young AstV+ bats, possibly attributable to their immature immune system. Regardless of age, pathogen-containing genera exhibited negative interactions with several commensal taxa and increased after AstV-infection, leading to pathobiont-like shifts in the gut microbiome of all infected bats. Thus, in apparently healthy bats, AstV-infections disturb gut bacterial homeostasis, possibly increasing previously suppressed health risks by promoting co-infections. If similar processes are present in humans, the effects of enteric virus infections might have longer-term impacts extending beyond the directly observed symptoms.

Hepatitis A virus (HAV) is an ancient and ubiquitous human pathogen recovered previously only from primates. The sole species of the genus Hepatovirus, existing in both enveloped and nonenveloped forms, and with a capsid structure intermediate between that of insect viruses and mammalian picornaviruses, HAV is enigmatic in its origins. We conducted a targeted search for hepatoviruses in 15,987 specimens collected from 209 small mammal species globally and discovered highly diversified viruses in bats, rodents, hedgehogs, and shrews, which by pairwise sequence distance comprise 13 novel Hepatovirus species. Near-complete genomes from nine of these species show conservation of unique hepatovirus features, including predicted internal ribosome entry site structure, a truncated VP4 capsid protein lacking N-terminal myristoylation, a carboxyl- terminal pX extension of VP1, VP2 late domains involved in membrane envelopment, and a cis-acting replication element within the 3D(pol) sequence. Antibodies in some bat sera immunoprecipitated and neutralized human HAV, suggesting conservation of critical antigenic determinants. Limited phylogenetic cosegregation among hepatoviruses and their hosts and recombination patterns are indicative of major hepatovirus host shifts in the past. Ancestral state reconstructions suggest a Hepatovirus origin in small insectivorous mammals and a rodent origin of human HAV. Patterns of infection in small mammals mimicked those of human HAV in hepatotropism, fecal shedding, acute nature, and extinction of the virus in a closed host population. The evolutionary conservation of hepatovirus structure and pathogenesis provide novel insight into the origins of HAV and highlight the utility of analyzing animal reservoirs for risk assessment of emerging viruses.

The hepatitis C virus (HCV) is a major human pathogen. Genetically related viruses in animals suggest a zoonotic origin of HCV. The closest relative of HCV is found in horses (termed equine hepacivirus [EqHV]). However, low EqHV genetic diversity implies relatively recent acquisition of EqHV by horses, making a derivation of HCV from EqHV unlikely. To unravel the EqHV evolutionary history within equid sister species, we analyzed 829 donkeys and 53 mules sampled in nine European, Asian, African, and American countries by molecular and serologic tools for EqHV infection. Antibodies were found in 278 animals (31.5%), and viral RNA was found in 3 animals (0.3%), all of which were simultaneously seropositive. A low RNA prevalence in spite of high seroprevalence suggests a predominance of acute infection, a possible difference from the mostly chronic hepacivirus infection pattern seen in horses and humans. Limitation of transmission due to short courses of infection may explain the existence of entirely seronegative groups of animals. Donkey and horse EqHV strains were paraphyletic and 97.5 to 98.2% identical in their translated polyprotein sequences, making virus/host cospeciation unlikely. Evolutionary reconstructions supported host switches of EqHV between horses and donkeys without the involvement of adaptive evolution. Global admixture of donkey and horse hepaciviruses was compatible with anthropogenic alterations of EqHV ecology. In summary, our findings do not support EqHV as the origin of the significantly more diversified HCV. Identification of a host system with predominantly acute hepacivirus infection may enable new insights into the chronic infection pattern associated with HCV. IMPORTANCE The evolutionary origins of the human hepatitis C virus (HCV) are unclear. The closest animal-associated relative of HCV occurs in horses (equine hepacivirus [EqHV]). The low EqHV genetic diversity implies a relatively recent acquisition of EqHV by horses, limiting the time span for potential horse-to-human infections in the past. Horses are genetically related to donkeys, and EqHV may have cospeciated with these host species. Here, we investigated a large panel of donkeys from various countries using serologic and molecular tools. We found EqHV to be globally widespread in donkeys and identify potential differences in EqHV infection patterns, with donkeys potentially showing enhanced EqHV clearance compared to horses. We provide strong evidence against EqHV cospeciation and for its capability to switch hosts among equines. Differential hepacivirus infection patterns in horses and donkeys may enable new insights into the chronic infection pattern associated with HCV.

Emerging infectious diseases (EIDs) are considered a major threat to global health. Most EIDs appear to result from increased contact between wildlife and humans, especially when humans encroach into formerly pristine habitats. Habitat deterioration may also negatively affect the physiology and health of wildlife species, which may eventually lead to a higher susceptibility to infectious agents and/or increased shedding of the pathogens causing EIDs. Bats are known to host viruses closely related to important EIDs. Here, we tested in a paleotropical forest with ongoing logging and fragmentation, whether habitat disturbance influences the occurrence of astro- and coronaviruses in eight bat species. In contrast to our hypothesis, anthropogenic habitat disturbance was not associated with corona- and astrovirus detection rates in fecal samples. However, we found that bats infected with either astro- or coronaviruses were likely to be coinfected with the respective other virus. Additionally, we identified two more risk factors influencing astrovirus shedding. First, the detection rate of astroviruses was higher at the beginning of the rainy compared to the dry season. Second, there was a trend that individuals with a poor body condition had a higher probability of shedding astroviruses in their feces. The identification of risk factors for increased viral shedding that may potentially result in increased interspecies transmission is important to prevent viral spillovers from bats to other animals, including humans.

A new hepatitis E virus (HEV-7) was recently found in dromedaries and 1 human from the United Arab Emirates. We screened 2,438 dromedary samples from Pakistan, the United Arab Emirates, and 4 African countries. HEV-7 is long established, diversified and geographically widespread. Dromedaries may constitute a neglected source of zoonotic HEV infections.

Hepatitis E virus (HEV) is one of the most common causes of acute hepatitis in tropical and temperate climates. Tropical genotypes 1 and 2 are associated with food-borne and waterborne transmission. Zoonotic reservoirs (mainly pigs, wild boar, and deer) are considered for genotypes 3 and 4, which exist in temperate climates. In view of the association of several zoonotic viruses with bats, we analyzed 3,869 bat specimens from 85 different species and from five continents for hepevirus RNA. HEVs were detected in African, Central American, and European bats, forming a novel phylogenetic clade in the family Hepeviridae . Bat hepeviruses were highly diversified and comparable to human HEV in sequence variation. No evidence for the transmission of bat hepeviruses to humans was found in over 90,000 human blood donations and individual patient sera. Full-genome analysis of one representative virus confirmed formal classification within the family Hepeviridae . Sequence- and distance-based taxonomic evaluations suggested that bat hepeviruses constitute a distinct genus within the family Hepeviridae and that at least three other genera comprising human, rodent, and avian hepeviruses can be designated. This may imply that hepeviruses invaded mammalian hosts nonrecently and underwent speciation according to their host restrictions. Human HEV-related viruses in farmed and peridomestic animals might represent secondary acquisitions of human viruses, rather than animal precursors causally involved in the evolution of human HEV.

The origin of primate HBV (family Hepadnaviridae) is unknown. Hepadnaviruses are ancient pathogens and may have been associated with old mammalian lineages like bats for prolonged time. Indeed, the genetic diversity of bat hepadnaviruses exceeds that of extant hepadnaviruses in other host orders, suggesting a long evolution of hepadnaviruses in bats. Strikingly, a recently detected New World bat hepadnavirus is antigenically related to HBV and can infect human hepatocytes. Together with genetically diverse hepadnaviruses from New World rodents and a non-human primate, these viruses argue for a New World origin of ancestral orthohepadnaviruses. Multiple host switches of bat and primate viruses are evident and bats are likely sources of ancestral hepadnaviruses acquired by primates.

Bats are likely natural hosts for a range of zoonotic viruses such as Marburg, Ebola, Rabies, as well as for various Corona-and Paramyxoviruses. In 2009/10, researchers discovered RNA of two novel influenza virus subtypes - H17N10 and H18N11 - in Central and South American fruit bats. The identification of bats as possible additional reservoir for influenza A viruses raises questions about the role of this mammalian taxon in influenza A virus ecology and possible public health relevance. As molecular testing can be limited by a short time window in which the virus is present, serological testing provides information about past infections and virus spread in populations after the virus has been cleared. This study aimed at screening available sera from 100 free-ranging, frugivorous bats (Eidolon helvum) sampled in 2009/10 in Ghana, for the presence of antibodies against the complete panel of influenza A haemagglutinin (HA) types ranging from H1 to H18 by means of a protein microarray platform. This technique enables simultaneous serological testing against multiple recombinant HA-types in 5 mu l of serum. Preliminary results indicate serological evidence against avian influenza subtype H9 in about 30% of the animals screened, with low-level cross-reactivity to phylogenetically closely related subtypes H8 and H12. To our knowledge, this is the first report of serological evidence of influenza A viruses other than H17 and H18 in bats. As avian influenza subtype H9 is associated with human infections, the implications of our findings from a public health context remain to be investigated.

Nature Communications 3: Article number: 796 (2012); Published: 24 April 2012; Updated: 23 January 2014. The authors inadvertently omitted Veronika M. Cottontail and Mirjam Knornschild, who collected samples in Panama and Costa Rica, from the author list. This has now been corrected in both the PDF and HTML versions of the Article.

Bats host a broad diversity of coronaviruses (CoVs), including close relatives of human pathogens. There is only limited data on neotropical bat CoVs. We analysed faecal, blood and intestine specimens from 1562 bats sampled in Costa Rica, Panama, Ecuador and Brazil for CoVs by broad-range PCR. CoV RNA was detected in 50 bats representing nine different species, both frugivorous and insectivorous. These bat CoVs were unrelated to known human or animal pathogens, indicating an absence of recent zoonotic spill-over events. Based on RNA-dependent RNA polymerase (RdRp)-based grouping units (RGUs) as a surrogate for CoV species identification, the 50 viruses represented five different alphacoronavirus RGUs and two betacoronavirus RGUs. Closely related alphacoronaviruses were detected in Carollia perspicillata and C. brevicauda across a geographical distance exceeding 5600 km. Our study expands the knowledge on CoV diversity in neotropical bats and emphasizes the association of distinct CoVs and bat host genera.

The hepatitis B virus (HBV), family Hepadnaviridae, is one of most relevant human pathogens. HBV origins are enigmatic, and no zoonotic reservoirs are known. Here, we screened 3,080 specimens from 54 bat species representing 11 bat families for hepadnaviral DNA. Ten specimens (0.3%) from Panama and Gabon yielded unique hepadnaviruses in coancestral relation to HBV. Full genome sequencing allowed classification as three putative orthohepadnavirus species based on genome lengths (3,149–3,377 nt), presence of middle HBV surface and X-protein genes, and sequence distance criteria. Hepatic tropism in bats was shown by quantitative PCR and in situ hybridization. Infected livers showed histopathologic changes compatible with hepatitis. Human hepatocytes transfected with all three bat viruses cross-reacted with sera against the HBV core protein, concordant with the phylogenetic relatedness of these hepadnaviruses and HBV. One virus from Uroderma bilobatum, the tent-making bat, cross-reacted with monoclonal antibodies against the HBV antigenicity determining S domain. Up to 18.4% of bat sera contained antibodies against bat hepadnaviruses. Infectious clones were generated to study all three viruses in detail. Hepatitis D virus particles pseudotyped with surface proteins of U. bilobatum HBV, but neither of the other two viruses could infect primary human and Tupaia belangeri hepatocytes. Hepatocyte infection occurred through the human HBV receptor sodium taurocholate cotransporting polypeptide but could not be neutralized by sera from vaccinated humans. Antihepadnaviral treatment using an approved reverse transcriptase inhibitor blocked replication of all bat hepadnaviruses. Our data suggest that bats may have been ancestral sources of primate hepadnaviruses. The observed zoonotic potential might affect concepts aimed at eradicating HBV.

The large virus family Paramyxoviridae includes some of the most significant human and livestock viruses, such as measles-, distemper-, mumps-, parainfluenza-, Newcastle disease-, respiratory syncytial virus and metapneumoviruses. Here we identify an estimated 66 new paramyxoviruses in a worldwide sample of 119 bat and rodent species (9,278 individuals). Major discoveries include evidence of an origin of Hendra- and Nipah virus in Africa, identification of a bat virus conspecific with the human mumps virus, detection of close relatives of respiratory syncytial virus, mouse pneumonia- and canine distemper virus in bats, as well as direct evidence of Sendai virus in rodents. Phylogenetic reconstruction of host associations suggests a predominance of host switches from bats to other mammals and birds. Hypothesis tests in a maximum likelihood framework permit the phylogenetic placement of bats as tentative hosts at ancestral nodes to both the major Paramyxoviridae subfamilies (Paramyxovirinae and Pneumovirinae). Future attempts to predict the emergence of novel paramyxoviruses in humans and livestock will have to rely fundamentally on these data., The large virus family, Paramyxoviridae, includes several human and livestock viruses. This study, testing 119 bat and rodent species distributed globally, identifies novel putative paramyxovirus species, providing data with potential uses in predictions of the emergence of novel paramyxoviruses in humans and livestock.