Host range and structural analysis of bat-origin RshSTT182/200 coronavirus binding to human ACE2 and its animal orthologs.

Bat-origin RshSTT182 and RshSTT200 coronaviruses (CoV) from Rhinolophus shameli in Southeast Asia (Cambodia) share 92.6% whole-genome identity with SARS-CoV-2 and show identical receptor-binding domains (RBDs). In this study, we determined the structure of the RshSTT182/200 receptor binding domain (RBD) in complex with human angiotensin-converting enzyme 2 (hACE2) and identified the key residues that influence receptor binding. The binding of the RshSTT182/200 RBD to ACE2 orthologs from 39 animal species, including 18 bat species, was used to evaluate its host range. The RshSTT182/200 RBD broadly recognized 21 of 39 ACE2 orthologs, although its binding affinities for the orthologs were weaker than those of the RBD of SARS-CoV-2. Furthermore, RshSTT182 pseudovirus could utilize human, fox, and Rhinolophus affinis ACE2 receptors for cell entry. Moreover, we found that SARS-CoV-2 induces cross-neutralizing antibodies against RshSTT182 pseudovirus. Taken together, these findings indicate that RshSTT182/200 can potentially infect susceptible animals, but requires further evolution to obtain strong interspecies transmission abilities like SARS-CoV-2.

Molecular basis for receptor recognition and broad host tropism for merbecovirus MjHKU4r-CoV-1.

A novel pangolin-origin MERS-like coronavirus (CoV), MjHKU4r-CoV-1, was recently identified. It is closely related to bat HKU4-CoV, and is infectious in human organs and transgenic mice. MjHKU4r-CoV-1 uses the dipeptidyl peptidase 4 (DPP4 or CD26) receptor for virus entry and has a broad host tropism. However, the molecular mechanism of its receptor binding and determinants of host range are not yet clear. Herein, we determine the structure of the MjHKU4r-CoV-1 spike (S) protein receptor-binding domain (RBD) complexed with human CD26 (hCD26) to reveal the basis for its receptor binding. Measuring binding capacity toward multiple animal receptors for MjHKU4r-CoV-1, mutagenesis analyses, and homology modeling highlight that residue sites 291, 292, 294, 295, 336, and 344 of CD26 are the crucial host range determinants for MjHKU4r-CoV-1. These results broaden our understanding of this potentially high-risk virus and will help us prepare for possible outbreaks in the future.

Complex N-glycans are important for interspecies transmission of H7 influenza A viruses.

Influenza A viruses (IAVs) can overcome species barriers by adaptation of the receptor-binding site of the hemagglutinin (HA). To initiate infection, HAs bind to glycan receptors with terminal sialic acids, which are either N-acetylneuraminic acid (NeuAc) or N-glycolylneuraminic acid (NeuGc); the latter is mainly found in horses and pigs but not in birds and humans. We investigated the influence of previously identified equine NeuGc-adapting mutations (S128T, I130V, A135E, T189A, and K193R) in avian H7 IAVs in vitro and in vivo. We observed that these mutations negatively affected viral replication in chicken cells but not in duck cells and positively affected replication in horse cells. In vivo, the mutations reduced virus virulence and mortality in chickens. Ducks excreted high viral loads longer than chickens, although they appeared clinically healthy. To elucidate why these viruses infected chickens and ducks despite the absence of NeuGc, we re-evaluated the receptor binding of H7 HAs using glycan microarray and flow cytometry studies. This re-evaluation demonstrated that mutated avian H7 HAs also bound to α2,3-linked NeuAc and sialyl-LewisX, which have an additional fucose moiety in their terminal epitope, explaining why infection of ducks and chickens was possible. Interestingly, the α2,3-linked NeuAc and sialyl-LewisX epitopes were only bound when presented on tri-antennary N-glycans, emphasizing the importance of investigating the fine receptor specificities of IAVs. In conclusion, the binding of NeuGc-adapted H7 IAV to tri-antennary N-glycans enables viral replication and shedding by chickens and ducks, potentially facilitating interspecies transmission of equine-adapted H7 IAVs.IMPORTANCEInfluenza A viruses (IAVs) cause millions of deaths and illnesses in birds and mammals each year. The viral surface protein hemagglutinin initiates infection by binding to host cell terminal sialic acids. Hemagglutinin adaptations affect the binding affinity to these sialic acids and the potential host species targeted. While avian and human IAVs tend to bind to N-acetylneuraminic acid (sialic acid), equine H7 viruses prefer binding to N-glycolylneuraminic acid (NeuGc). To better understand the function of NeuGc-specific adaptations in hemagglutinin and to elucidate interspecies transmission potential NeuGc-adapted viruses, we evaluated the effects of NeuGc-specific mutations in avian H7 viruses in chickens and ducks, important economic hosts and reservoir birds, respectively. We also examined the impact on viral replication and found a binding affinity to tri-antennary N-glycans containing different terminal epitopes. These findings are significant as they contribute to the understanding of the role of receptor binding in avian influenza infection.

Expression of influenza A virus glycan receptor candidates in mallard, chicken, and tufted duck.

Influenza A virus (IAV) pandemics result from interspecies transmission events within the avian reservoir and further into mammals including humans. Receptor incompatibility due to differently expressed glycan structures between species has been suggested to limit zoonotic IAV transmission from the wild bird reservoir as well as between different bird species. Using glycoproteomics, we have studied the repertoires of expressed glycan structures with focus on putative sialic acid-containing glycan receptors for IAV in mallard, chicken and tufted duck; three bird species with different roles in the zoonotic ecology of IAV. The methodology used pinpoints specific glycan structures to specific glycosylation sites of identified glycoproteins and was also used to successfully discriminate α2-3- from α2-6-linked terminal sialic acids by careful analysis of oxonium ions released from glycopeptides in tandem MS/MS (MS2), and MS/MS/MS (MS3). Our analysis clearly demonstrated that all three bird species can produce complex N-glycans including α2-3-linked sialyl Lewis structures, as well as both N- and O- glycans terminated with both α2-3- and α2-6-linked Neu5Ac. We also found the recently identified putative IAV receptor structures, Man-6P N-glycopeptides, in all tissues of the three bird species. Furthermore, we found many similarities in the repertoires of expressed receptors both between the bird species investigated and to previously published data from pigs and humans. Our findings of sialylated glycan structures, previously anticipated to be mammalian specific, in all three bird species may have major implications for our understanding of the role of receptor incompatibility in interspecies transmission of IAV.

Chicken or Porcine Aminopeptidase N Mediates Cellular Entry of Pseudoviruses Carrying Spike Glycoprotein from the Avian Deltacoronaviruses HKU11, HKU13, and HKU17.

Members of deltacoronavirus (DCoV) have mostly been identified in diverse avian species as natural reservoirs, though the porcine DCoV (PDCoV) is a major swine enteropathogenic virus with global spread. The important role of aminopeptidase N (APN) orthologues from various mammalian and avian species in PDCoV cellular entry and interspecies transmission has been revealed recently. In this study, comparative analysis indicated that three avian DCoVs, bulbul DCoV HKU11, munia DCoV HKU13, and sparrow DCoV HKU17 (Chinese strain), and PDCoV in the subgenera Buldecovirus are grouped together at whole-genome levels; however, the spike (S) glycoprotein and its S1 subunit of HKU17 are more closely related to night heron DCoV HKU19 in Herdecovirus. Nevertheless, the S1 protein of HKU11, HKU13, or HKU17 bound to or interacted with chicken APN (chAPN) or porcine APN (pAPN) by flow cytometry analysis of cell surface expression of APN and by coimmunoprecipitation in APN-overexpressing cells. Expression of chAPN or pAPN allowed entry of pseudotyped lentiviruses with the S proteins from HKU11, HKU13 and HKU17 into nonsusceptible cells and natural avian and porcine cells, which could be inhibited by the antibody against APN or anti-PDCoV-S1. APN knockdown by siRNA or knockout by CRISPR/Cas9 in chicken or swine cell lines significantly or almost completely blocked infection of these pseudoviruses. Hence, we demonstrate that HKU11, HKU13, and HKU17 with divergent S genes likely engage chAPN or pAPN to enter the cells, suggesting a potential interspecies transmission from wild birds to poultry and from birds to mammals by certain avian DCoVs. IMPORTANCE The receptor usage of avian deltacoronaviruses (DCoVs) has not been investigated thus far, though porcine deltacoronavirus (PDCoV) has been shown to utilize aminopeptidase N (APN) as a cell receptor. We report here that chicken or porcine APN also mediates cellular entry by three avian DCoV (HKU11, HKU13, and HKU17) spike pseudoviruses, and the S1 subunit of three avian DCoVs binds to APN in vitro and in the surface of avian and porcine cells. The results fill the gaps in knowledge about the avian DCoV receptor and elucidate important insights for the monitoring and prevention of potential interspecies transmission of certain avian DCoVs. In view of the diversity of DCoVs, whether this coronavirus genus will cause novel virus to emerge in other mammals from birds, are worthy of further surveillance and investigation.

Characteristics of two zoonotic swine influenza A(H1N1) viruses isolated in Germany from diseased patients.

Interspecies transmission of influenza A viruses (IAV) from pigs to humans is a concerning event as porcine IAV represent a reservoir of potentially pandemic IAV. We conducted a comprehensive analysis of two porcine A(H1N1)v viruses isolated from human cases by evaluating their genetic, antigenic and virological characteristics. The HA genes of those human isolates belonged to clades 1C.2.1 and 1C.2.2, respectively, of the A(H1N1) Eurasian avian-like swine influenza lineage. Antigenic profiling revealed substantial cross-reactivity between the two zoonotic H1N1 viruses and human A(H1N1)pdm09 virus and some swine viruses, but did not reveal cross-reactivity to H1N2 and earlier human seasonal A(H1N1) viruses. The solid-phase direct receptor binding assay analysis of both A(H1N1)v showed a predominant binding to α2-6-sialylated glycans similar to human-adapted IAV. Investigation of the replicative potential revealed that both A(H1N1)v viruses grow in human bronchial epithelial cells to similar high titers as the human A(H1N1)pdm09 virus. Cytokine induction was studied in human alveolar epithelial cells A549 and showed that both swine viruses isolated from human cases induced higher amounts of type I and type III IFN, as well as IL6 compared to a seasonal A(H1N1) or a A(H1N1)pdm09 virus. In summary, we demonstrate a remarkable adaptation of both zoonotic viruses to propagate in human cells. Our data emphasize the needs for continuous monitoring of people and regions at increased risk of such trans-species transmissions, as well as systematic studies to quantify the frequency of these events and to identify viral molecular determinants enhancing the zoonotic potential of porcine IAV.

Genetic diversity and cross-species transmissibility of bat-associated picornaviruses from Spain.

BACKGROUND: Emerging zoonotic diseases arise from cross-species transmission events between wild or domesticated animals and humans, with bats being one of the major reservoirs of zoonotic viruses. Viral metagenomics has led to the discovery of many viruses, but efforts have mainly been focused on some areas of the world and on certain viral families. METHODS: We set out to describe full-length genomes of new picorna-like viruses by collecting feces from hundreds of bats captured in different regions of Spain. Viral sequences were obtained by high-throughput Illumina sequencing and analyzed phylogenetically to classify them in the context of known viruses. Linear discriminant analysis (LDA) was performed to infer likely hosts based on genome composition. RESULTS: We found five complete or nearly complete genomes belonging to the family Picornaviridae, including a new species of the subfamily Ensavirinae. LDA suggested that these were true vertebrate viruses, rather than viruses from the bat diet. Some of these viruses were related to picornaviruses previously found in other bat species from distant geographical regions. We also found a calhevirus genome that most likely belongs to a proposed new family within the order Picornavirales, and for which genome composition analysis suggested a plant host. CONCLUSIONS: Our findings describe new picorna-like viral species and variants circulating in the Iberian Peninsula, illustrate the wide geographical distribution and interspecies transmissibility of picornaviruses, and suggest new hosts for calheviruses.

Transmission of Norwegian reindeer CWD to sheep by intracerebral inoculation results in an unusual phenotype and prion distribution.

Chronic wasting disease (CWD), a prion disease affecting cervids, has been known in North America (NA) since the 1960s and emerged in Norway in 2016. Surveillance and studies have revealed that there are different forms of CWD in Fennoscandia: contagious CWD in Norwegian reindeer and sporadic CWD in moose and red deer. Experimental studies have demonstrated that NA CWD prions can infect various species, but thus far, there have been no reports of natural transmission to non-cervid species. In vitro and laboratory animal studies of the Norwegian CWD strains suggest that these strains are different from the NA strains. In this work, we describe the intracerebral transmission of reindeer CWD to six scrapie-susceptible sheep. Detection methods included immunohistochemistry (IHC), western blot (WB), enzyme-linked immunosorbent assay (ELISA), real-time quaking-induced conversion (RT-QuIC) and protein misfolding cyclic amplification (PMCA). In the brain, grey matter vacuolation was limited, while all sheep exhibited vacuolation of the white matter. IHC and WB conventional detection techniques failed to detect prions; however, positive seeding activity with the RT-QuIC and PMCA amplification techniques was observed in the central nervous system of all but one sheep. Prions were robustly amplified in the lymph nodes of all animals, mainly by RT-QuIC. Additionally, two lymph nodes were positive by WB, and one was positive by ELISA. These findings suggest that sheep can propagate reindeer CWD prions after intracerebral inoculation, resulting in an unusual disease phenotype and prion distribution with a low amount of detectable prions.

Pigs in southern Italy are exposed to three ruminant pathogens: an analysis of seroprevalence and risk factors analysis study.

BACKGROUND: Pigs are susceptible to several ruminant pathogens, including Coxiella burnetti, Schmallenberg virus (SBV) and bovine viral diarrhea virus (BVDV). These pathogens have already been described in the pig population, although the dynamics of the infection and the impact on pig farms are currently unclear. The aim of this work was to evaluate the presence of these infections in the pig population of the Campania region, southern Italy, and to evaluate the risk factors associated with a greater risk of exposure. RESULTS: A total of 414 serum samples belonging to 32 herds were tested for the presence of antibodies against SBV, Coxiella, and BVD using commercial multispecies ELISA kits. SBV (5.3%) was the most prevalent pathogen, followed by Coxiella (4.1%) and BVD (3%). The risk factors included in the study (age, sex, province, farming system, ruminant density and major ruminant species) had no influence on the probability of being exposed to BVD and Coxiella, except for the location, in fact more pigs seropositive to Coxiella were found in the province of Caserta. However, the univariate analysis highlighted the influence of age, location, and sex on exposure to SBV. The subsequent multivariate analysis statistically confirmed the importance of these factors. The presence of neutralizing antibodies for SBV and BVDV, or antibodies directed towards a specific phase of infection for Coxiella was further confirmed with virus-neutralization assays and phase-specific ELISAs in a large proportion of positive samples. The presence of high neutralizing antibody titers (especially for SBV) could indicate recent exposures. Twelve of the 17 positive samples tested positive for antibodies against Coxiella phase I or II antigens, indicating the presence of both acute and chronic infections (one animal tested positive for both phases antibodies). CONCLUSIONS: Our study indicates a non-negligible exposure of pigs from southern Italy to the above pathogens. Further studies are necessary to fully understand the dynamics of these infections in pigs, the impact on productivity, and the public health consequences in the case of Coxiella.

Key mutations on spike protein altering ACE2 receptor utilization and potentially expanding host range of emerging SARS-CoV-2 variants.

Increasing evidence supports inter-species transmission of SARS-CoV-2 variants from humans to domestic or wild animals during the ongoing COVID-19 pandemic, which is posing great challenges to epidemic control. Clarifying the host range of emerging SARS-CoV-2 variants will provide instructive information for the containment of viral spillover. The spike protein (S) of SARS-CoV-2 is the key determinant of receptor utilization, and therefore amino acid mutations on S will probably alter viral host range. Here, to evaluate the impact of S mutations, we tested 27 pseudoviruses of SARS-CoV-2 carrying different spike mutants by infecting Hela cells expressing different angiotensin-converting enzyme 2 (ACE2) orthologs from 20 animals. Of these 27 pseudoviruses, 20 bear single mutation and the other 7 were cloned from emerging SARS-CoV-2 variants, including D614G, Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Delta (B.1.617.2), Lambda (B.1.429), and Mu (B.1.621). Using pseudoviral reporter assay, we identified that the substitutions of T478I and N501Y enabled the pseudovirus to utilize chicken ACE2, indicating potential infectivity to avian species. Furthermore, the S mutants of real SARS-CoV-2 variants comprising N501Y showed significantly acquired abilities to infect cells expressing mouse ACE2, indicating a critical role of N501Y in expanding SARS-CoV-2 host range. In addition, A262S and T478I significantly enhanced the utilization of various mammal ACE2. In summary, our results indicated that T478I and N501Y substitutions were two S mutations important for receptor adaption of SARS-CoV-2, potentially contributing to the spillover of the virus to many other animal hosts. Therefore, more attention should be paid to SARS-CoV-2 variants with these two mutations.

The origin and evolution of emerged swine acute diarrhea syndrome coronavirus with zoonotic potential.

Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a newly discovered alphacoronavirus with zoonotic potential that causes diarrhea and vomiting mainly in piglets. Having emerged suddenly in 2017, the prevailing opinion is that the virus originated from HKU2, an alphacoronavirus whose primary host is bats, and at some unknown point achieved interspecies transmission via some intermediate. Here, we further explore the evolutionary history and possible cross-species transmission event for SADS-CoV. Coevolutionary analysis demonstrated that HKU2 may have achieved host switch via SADS-related (SADSr)-CoV, which was isolated from the genus Rhinolophus in 2017. SADS-CoV, HKU2, and SADSr-CoV share similar codon usage patterns and showed a lower tendency to use CpG, which may reflect a method of immune escape. The analyses of virus-host coevolution and recombination support SADSr-CoV is the direct source of SADS-CoV that may have undergone recombination events during its formation. Structure-based spike glycoprotein variance analysis revealed a more nuanced evolutionary pathway to receptor recognition for host switch. We did not find a possible positive selection site, and the dN/dS of the S gene was only 0.29, which indicates that the current SADS-CoV is slowly evolving. These results provide new insights that may help predict future cross-species transmission, and possibly surveil future zoonotic outbreaks and associated public health emergencies.

[Transmission routes of neuropathogenic pathogens-Possible mechanisms of neuroinvasion]. / Übertragungswege neuropathogener Erreger ­ mögliche Mechanismen der Neuroinvasion.

Despite natural protective barriers, many human pathogens can penetrate the nervous system. The pathogens have developed sophisticated mechanisms to overturn the privileged immune status of the nervous system. The central nervous system (CNS) has natural barriers and immunological protective mechanisms, such as the blood-brain barrier (BBB) and the blood-cerebrospinal fluid barrier (BCSFB), that prevent the invasion of pathogens. Bacteria enter the CNS, for example, through transcellular penetration, paracellular entry or via infected leucocytes from the peripheral circulation. Viruses can enter the CNS by hematogenous routes, by direct infection of endothelial cells or by paracellular passage between disrupted endothelial cells (cribriform plate). Also, a few "enhanced barriers", such as the cribriform plate and circumventricular organs (CVO) can serve as entry points for viruses. Viruses can also gain access to the CNS by infection of peripheral nerves. Importantly, most successful neurotropic pathogens are not necessarily restricted to a CNS entry portal. The majority of viral pathogens are of animal origin. Through sometimes sophisticated immune mechanisms for example in bats, highly pathogenic pathogens arose through cross-species transmission. In the last 50 years, various viruses, such as West Nile, Ebola, Marburg, Zika, Nipah and Hendra viruses have also been transmitted from animals to humans. Knowledge of these biological strategies is crucial to hinder, contain or prevent CNS infections.

Progress on viral and host determinants of influenza A virus species specificity.

Influenza A viruses have a wide range of hosts and are highly infectious, which can cause zoonotic diseases and pose a serious public health threat to human safety. An influenza pandemic could outbreak if new strains gain the ability of human-to-human transmission, either by genetic mutation or by gene reassortment. It is an urgent issue for the scientific community to reveal the genetic basis and molecular mechanisms underlying the interspecies transmission of influenza viruses, which will provide important implications for the effective monitoring and prevention of potential influenza pandemics. In this review, we summarize the molecular determinants of influenza viruses for host adaptation, and highlight the advances in the gene mutations of the virus itself and the interaction between virus and host factors. This will help to make a theoretical reserve for the next influenza pandemic and find new strategies to fight against influenza.

Characterization of an avian rotavirus A strain isolated from a velvet scoter (Melanitta fusca): implication for the role of migratory birds in global spread of avian rotaviruses.

Avian G18P[17] rotaviruses with similar complete genome constellation, including strains that showed pathogenicity in mammals, have been detected worldwide. However, it remains unclear how these strains spread geographically. In this study, to investigate the role of migratory birds in the dispersion of avian rotaviruses, we analysed whole genetic characters of the rotavirus strain RK1 that was isolated from a migratory species of birds [velvet scoter (Melanitta fusca)] in Japan in 1989. Genetic analyses revealed that the genotype constellation of the RK1 strain, G18-P[17]-I4-R4-C4-M4-A21-N4-T4-E4-H4, was highly consistent with those of other G18P[17] strains detected in various parts of the world, supporting the possibility that the G18P[17] strains spread via migratory birds that move over a wide area. Furthermore, the RK1 strain induced diarrhoea in suckling mice after oral gastric inoculation, indicating that at least some of the rotaviruses that originated from migratory birds are infectious to and pathogenic in mammals. In conclusion, it was demonstrated that migratory birds may contribute to the global spread of avian rotaviruses that are pathogenic in mammalian species.

Recent advances and public health implications for environmental exposure to Chlamydia abortus: from enzootic to zoonotic disease.

Environmental transmission of Chlamydia abortus as a result of enzootic disease or disease outbreaks and the threats posed by this pathogen has been previously reported, however a state-of-the-science review of these reports and the identification of future research priorities in this area is still lacking. This study provides an overview of the current knowledge of host-pathogen-environment interactions, addressing public health risks and identifying critical questions and research gaps. We performed a systematic PubMed and Web of Science search for publications related to Chlamydia abortus in the past four decades, and we reviewed and combined the evidence critically discussing and commenting the results. A total of 182 studies, 5 chapters of specific books and the "OIE terrestrial manual" were included in this review. There were substantial variations between the studies in topic addressed and experimental design. Overall, the literature largely supports the crucial role played by environmental exposure on the acquisition of zoonotic disease caused by Chlamydia abortus. We also identify the paucity of information related to interspecies transmission and pathogen adaptation in relation to environmental dissemination and zoonotic risk. This analysis further highlights the need for additional research given that environmental transmission represents a serious risk not only to susceptible patients (pregnant women and immunocompromised individuals), but also for other species including wildlife.

Aujeszky's disease in hunting dogs after the ingestion of wild boar raw meat in Sicily (Italy): clinical, diagnostic and phylogenetic features.

BACKGROUND: Aujeszky's disease is caused by Suid Herpes Virus-1 and species belonging to the genus Sus scrofa are the main reservoir hosts. This virus, however, is capable of infecting and causing severe disease, with an almost constant fatal outcome in other species, both domestic and wild (carnivores, monogastric herbivores and ruminants). Moreover, the possibility of transmission to humans has been demonstrated. This study reports and describes the clinical, diagnostic, pathological and phylogenetic aspects of two cases of Aujeszky's disease in two hunting dogs following the ingestion of infected wild boar raw meat. These cases are contextualized in the province of Messina (Sicily), where a high prevalence of Aujeszky's disease has been recorded (average of 12,20% in the period 2010-2019) in farmed pig, and with evidence of spread to other species. A severe outbreak in cattle has recently been reported in these areas. Nevertheless, cases of Aujeszky's disease in dogs are rarely reported and this study represents the first well-documented report in this species in Sicily. CASE PRESENTATION: After a wild boar hunt, two dogs showed neurological symptoms and intense itching unresponsive to therapy. Diagnosis of Aujeszky's disease was made based on clinical suspicion, anamnestic information and confirmed by the isolation of the virus from the brain of both dogs. In addition, molecular typing, sequencing and phylogenetic analysis of the Real-Time PCR products were performed. The sequences studied were placed in the Italian Clade 1 along with the sequences obtained from wild boars and hunting dogs from Italy and France. CONCLUSIONS: The finding of this disease in non-natural hosts in Sicilian multi-host epidemiological contexts suggests that the risk of inter-species transmission is concrete and that attention should be paid to developing disease control programs in these territories. The data obtained from genome sequencing of the two SuHV-1 isolates contribute to the enrichment of the GenBank with unknown sequences and the phylogenetic analysis implementation.

Molecular characterization of the first human G15 rotavirus strain of zoonotic origin from the bovine species.

Group A rotaviruses (RVAs) infect a wide variety of mammalian and avian species. Animals act as a potential reservoir to RVA human infections by direct virion transmission or by contributing genes to reassortants. Here, we report the molecular characterization of a rare human RVA strain Ni17-46 with a genotype G15P[14], isolated in Japan in 2017 during rotavirus surveillance in a paediatric outpatient clinic. The genome constellation of this strain was G15-P[14]-I2-R2-C2-M2-A13-N2-T9-E2-H3. This is the first report of an RVA with G15 genotype in humans, and sequencing and phylogenetic analysis results suggest that human infection with this strain has zoonotic origin from the bovine species. Given the fact that this strain was isolated from a patient with gastroenteritis and dehydration symptoms, we must take into account the virulence of this strain in humans.

Genomic analysis reveals G3P[13] porcine rotavirus A interspecific transmission to human from pigs in a swine farm with diarrhoea outbreak.

Rotavirus A (RVA) is a major diarrhoea-causing pathogen in young animals and children. The zoonotic potential of RVA has received extensive attention in recent years. In May 2018, an outbreak of diarrhoea among piglets occurred on a swine farm in Sichuan province, PR China. RVA was detected in 95.7 % (22/23) of piglet samples, 60 % (9/15) of sow samples and 100 % (3/3) of pig-breeder faecal samples. The predominant RVA genotype on this swine farm was G3P[13], and G3P[13] RVA was also detected in the three breeder faecal samples. Three G3P[13] RVA strains were isolated from a piglet faecal sample, a sow faecal sample and a pig-breeder faecal sample, and were named SCLS-X1, SCLS-3 and SCLS-R3, respectively. The complete sequences of 11 gene segments of these three isolates were derived. Phylogenetic analysis showed that ten gene segments (VP7, VP4, VP1-VP3 and NSP1-NSP5) of pig-breeder isolate SCLS-R3 were closely related to pig isolates SCLS-X1 and SCLS-3 from this farm. Only the VP6 gene shared higher homology with human RVA strain I321. Therefore, a G3P[13] porcine RVA strain most likely infected pig breeders. These results provided the first complete epidemiological link demonstrating interspecies transmission of G3P[13] RVA from pigs to human. Our data contribute to an improved understanding of the genetic evolution and interspecies transmission of RVA.

Molecular Basis of Binding between Middle East Respiratory Syndrome Coronavirus and CD26 from Seven Bat Species.

Continued reports of Middle East respiratory syndrome coronavirus (MERS-CoV) infecting humans have occurred since the identification of this virus in 2012. MERS-CoV is prone to cause endemic disease in the Middle East, with several dozen spillover infections to other continents. It is hypothesized that MERS-CoV originated from bat coronaviruses and that dromedary camels are its natural reservoir. Although gene segments identical to MERS-CoV were sequenced from certain species of bats and one species experimentally shed the virus, it is still unknown whether other bats can transmit the virus. Here, at the molecular level, we found that all purified bat CD26s (bCD26s) from a diverse range of species interact with the receptor binding domain (RBD) of MERS-CoV, with equilibrium dissociation constant values ranging from several to hundreds at the micromolar level. Moreover, all bCD26s expressed in this study mediated the entry of pseudotyped MERS-CoV to receptor-expressing cells, indicating the broad potential engagement of bCD26s as MERS-CoV receptors. Further structural analysis indicated that in the bat receptor, compared to the human receptor, substitutions of key residues and their adjacent amino acids leads to decreased binding affinity to the MERS-RBD. These results add more evidence to the existing belief that bats are the original source of MERS-CoV and suggest that bCD26s in many species can mediate the entry of the virus, which has significant implications for the surveillance and control of MERS-CoV infection.IMPORTANCE In this study, we found that bat CD26s (bCD26s) from different species exhibit large diversities, especially in the region responsible for binding to the receptor binding domain (RBD) of Middle East respiratory syndrome coronavirus (MERS-CoV). However, they maintain the interaction with MERS-RBD at varied affinities and support the entry of pseudotyped MERS-CoV. These bat receptors polymorphisms seem to confer evolutionary pressure for the adaptation of CD26-binding virus, such as the ancestor of MERS-CoV, and led to the generation of diversified CD26-engaging CoV strains. Thus, our data add more evidence to support that bats are the reservoir of MERS-CoV and similar viruses, as well as further emphasize the necessity to survey MERS-CoV and other CoVs among bats.

Dogs are resistant to prion infection, due to the presence of aspartic or glutamic acid at position 163 of their prion protein.

Unlike other species, prion disease has never been described in dogs even though they were similarly exposed to the bovine spongiform encephalopathy (BSE) agent. This resistance prompted a thorough analysis of the canine PRNP gene and the presence of a negatively charged amino acid residue in position 163 was readily identified as potentially fundamental as it differed from all known susceptible species. In the present study, the first transgenic mouse model expressing dog prion protein (PrP) was generated and challenged intracerebrally with a panel of prion isolates, none of which could infect them. The brains of these mice were subjected to in vitro prion amplification and failed to find even minimal amounts of misfolded prions providing definitive experimental evidence that dogs are resistant to prion disease. Subsequently, a second transgenic model was generated in which aspartic acid in position 163 was substituted for asparagine (the most common in prion susceptible species) resulting in susceptibility to BSE-derived isolates. These findings strongly support the hypothesis that the amino acid residue at position 163 of canine cellular prion protein (PrPC ) is a major determinant of the exceptional resistance of the canidae family to prion infection and establish this as a promising therapeutic target for prion diseases.