Different populations of A(H1N1)pdm09 viruses in a patient with hemolytic-uremic syndrome.

Respiratory viral infections may have different impacts ranging from infection without symptoms to severe disease or even death though the reasons are not well characterized. A patient (age group 5-15 years) displaying symptoms of hemolytic uremic syndrome died one day after hospitalization. qPCR, next generation sequencing, virus isolation, antigenic characterization, resistance analysis was performed and virus replication kinetics in well-differentiated airway cells were determined. Autopsy revealed hemorrhagic pneumonia as major pathological manifestation. Lung samples harbored a large population of A(H1N1)pdm09 viruses with the polymorphism H456H/Y in PB1 polymerase. The H456H/Y viruses replicated much faster to high viral titers than upper respiratory tract viruses in vitro. H456H/Y-infected air-liquid interface cultures of differentiated airway epithelial cells did reflect a more pronounced loss of ciliated cells. A different pattern of virus quasispecies was found in the upper airway samples where substitution S263S/F (HA1) was observed. The data support the notion that viral quasispecies had evolved locally in the lung to support high replicative fitness. This change may have initiated further pathogenic processes leading to rapid dissemination of inflammatory mediators followed by development of hemorrhagic lung lesions and fatal outcome.

Infection of porcine enteroids and 2D differentiated intestinal epithelial cells with rotavirus A to study cell tropism and polarized immune response.

Intestinal epithelial cell interactions with enteric pathogens have been incompletely elucidated owing to the lack of model systems that recapitulate the cellular diversity, architecture and functionality of the intestine. To analyze rotavirus (RV) infection and the subsequent innate immune response, we established cultures of differentiated porcine intestinal epithelial cells in three different variations: basolateral-out enteroids, apical-out enteroids and two-dimensional (2D) filter-grown intestinal epithelial cells. Application of specific antibodies for fluorescent staining indicated that enteroids and enteroid-derived cell cultures contain multiple intestinal epithelial cell types. Infection studies indicated that both apical-out enteroids and 2D intestinal epithelial cells are susceptible to porcine RV infection. However, 2D intestinal epithelial cells are more useful for a detailed characterization and comparison of apical and basolateral infection than apical-out enteroids. Virus-induced apoptosis was observed in apical-out enteroids at 24 h post infection but not at earlier time points after infection. RV infected not only enterocytes but also goblet cells and Paneth cells in apical-out enteroids and 2D intestinal epithelial cells. Interestingly, despite the lack of significant differences in the efficiency of infection after apical and basolateral infection of 2D intestinal epithelial cells, stronger innate immune and inflammatory responses were observed after basolateral infection as compared to infection via the apical route. Therefore, apical-out enteroids and 2D intestinal epithelial cells provide useful primary cell culture models that can be extended to analyze invasion and replication strategies of agents implicated in enteric diseases or to study immune and inflammatory responses of the host induced by enteric pathogens.

Phenotypic and Transcriptional Changes of Pulmonary Immune Responses in Dogs Following Canine Distemper Virus Infection.

Canine distemper virus (CDV), a morbillivirus within the family Paramyxoviridae, is a highly contagious infectious agent causing a multisystemic, devastating disease in a broad range of host species, characterized by severe immunosuppression, encephalitis and pneumonia. The present study aimed at investigating pulmonary immune responses of CDV-infected dogs in situ using immunohistochemistry and whole transcriptome analyses by bulk RNA sequencing. Spatiotemporal analysis of phenotypic changes revealed pulmonary immune responses primarily driven by MHC-II+, Iba-1+ and CD204+ innate immune cells during acute and subacute infection phases, which paralleled pathologic lesion development and coincided with high viral loads in CDV-infected lungs. CD20+ B cell numbers initially declined, followed by lymphoid repopulation in the advanced disease phase. Transcriptome analysis demonstrated an increased expression of transcripts related to innate immunity, antiviral defense mechanisms, type I interferon responses and regulation of cell death in the lung of CDV-infected dogs. Molecular analyses also revealed disturbed cytokine responses with a pro-inflammatory M1 macrophage polarization and impaired mucociliary defense in CDV-infected lungs. The exploratory study provides detailed data on CDV-related pulmonary immune responses, expanding the list of immunologic parameters potentially leading to viral elimination and virus-induced pulmonary immunopathology in canine distemper.

Primary harbour seal (Phoca vitulina) airway epithelial cells show high susceptibility to infection by a seal-derived influenza A virus (H5N8).

Highly pathogenic avian influenza viruses of the H5N8 subtype have been circulating in Europe and Asia since 2016, causing huge economic losses to the poultry industry. A new wave of H5Nx infections has begun in 2020. The viruses mainly infect wild birds and waterfowl; from there they spread to poultry and cause diseases. Previous studies have shown that the H5N8 viruses have seldom spread to mammals; however, reports in early 2021 indicate that humans may be infected, and some incident reports indicate that H5Nx clade 2.3.4.4B virus may be transmitted to wild mammals, such as red foxes and seals. In order to get more information on how the H5N8 virus affects seals and other marine animals, here, we used primary cultures to analyze the cell tropism of the H5N8 virus, which was isolated from an infected grey seal (H5N8/Seal-2016). Primary tracheal epithelial cells were readily infected by H5N8/Seal -2016 virus; in contrast, the commonly used primary seal kidney cells required the presence of exogenous trypsin to initiate virus infection. When applied to an ex vivo precision-cut lung slice model, compared with recombinant human H3N2 virus or H9N2 LPAI virus, the H5N8/Seal-2016 virus replicated to a high titre and caused a strong detrimental effect; with these characteristics, the virus was superior to a human H3N2 virus and to an H9N2 LPAI virus. By using well-differentiated air-liquid interface (ALI) cultures, we have observed that ALI cultures of canines, ferrets, and harbour seals are more sensitive to H5N8/Seal-2016 virus than are human or porcine ALI cultures, which cannot be fully explained by sialic acid distribution. Our results indicate that the airway epithelium of carnivores may be the main target of H5N8 viruses. Consideration should be given to an increased monitoring of the distribution of highly pathogenic avian influenza viruses in wild animals.

Time-dependent viral interference between influenza virus and coronavirus in the infection of differentiated porcine airway epithelial cells.

Coronaviruses and influenza viruses are circulating in humans and animals all over the world. Co-infection with these two viruses may aggravate clinical signs. However, the molecular mechanisms of co-infections by these two viruses are incompletely understood. In this study, we applied air-liquid interface (ALI) cultures of well-differentiated porcine tracheal epithelial cells (PTECs) to analyze the co-infection by a swine influenza virus (SIV, H3N2 subtype) and porcine respiratory coronavirus (PRCoV) at different time intervals. Our results revealed that in short-term intervals, prior infection by influenza virus caused complete inhibition of coronavirus infection, while in long-term intervals, some coronavirus replication was detectable. The influenza virus infection resulted in (i) an upregulation of porcine aminopeptidase N, the cellular receptor for PRCoV and (ii) in the induction of an innate immune response which was responsible for the inhibition of PRCoV replication. By contrast, prior infection by coronavirus only caused a slight inhibition of influenza virus replication. Taken together, the timing and the order of virus infection are important determinants in co-infections. This study is the first to show the impact of SIV and PRCoV co- and super-infection on the cellular level. Our results have implications also for human viruses, including potential co-infections by SARS-CoV-2 and seasonal influenza viruses.

Infection of polarized bovine respiratory epithelial cells by bovine viral diarrhea virus (BVDV).

Bovine viral diarrhea virus (BVDV) is affecting cattle populations all over the world causing acute disease, immunosuppressive effects, respiratory diseases, gastrointestinal, and reproductive failure in cattle. The virus is taken up via the oronasal route and infection of epithelial and immune cells contributes to the dissemination of the virus throughout the body. However, it is not known how the virus gets across the barrier of epithelial cells encountered in the airways. Here, we analyzed the infection of polarized primary bovine airway epithelial cells (BAEC). Infection of BAEC by a non-cytopathogenic BVDV was possible via both the apical and the basolateral plasma membrane, but the infection was most efficient when the virus was applied to the basolateral plasma membrane. Irrespective of the site of infection, BVDV was efficiently released to the apical site, while only minor amounts of virus were detected in the basal medium. This indicates that the respiratory epithelium can release large amounts of BVDV to the environment and susceptible animals via respiratory fluids and aerosols, but BVDV cannot cross the airway epithelial cells to infect subepithelial cells and establish systemic infection. Further experiments showed that the receptor, bovine CD46, for BVDV is expressed predominantly on the apical membrane domain of the polarized epithelial cells. In a CD46 blocking experiment, the addition of an antibody directed against CD46 almost completely inhibited apical infection, whereas basolateral infection was not affected. While CD46 serves as a receptor for apical infection of BAEC by BVDV, the receptor for basolateral infection remains to be elucidated.

Infection of bovine well-differentiated airway epithelial cells by Pasteurella multocida: actions and counteractions in the bacteria-host interactions.

Pasteurella (P.) multocida is a zoonotic pathogen, which is able to cause respiratory disorder in different hosts. In cattle, P. multocida is an important microorganism involved in the bovine respiratory disease complex (BRDC) with a huge economic impact. We applied air-liquid interface (ALI) cultures of well-differentiated bovine airway epithelial cells to analyze the interaction of P. multocida with its host target cells. The bacterial pathogen grew readily on the ALI cultures. Infection resulted in a substantial loss of ciliated cells. Nevertheless, the epithelial cell layer maintained its barrier function as indicated by the transepithelial electrical resistance and the inability of dextran to get from the apical to the basolateral compartment via the paracellular route. Analysis by confocal immunofluorescence microscopy confirmed the intactness of the epithelial cell layer though it was not as thick as the uninfected control cells. Finally, we chose the bacterial neuraminidase to show that our infection model is a sustainable tool to analyze virulence factors of P. multocida. Furthermore, we provide an explanation, why this microorganism usually is a commensal and becomes pathogenic only in combination with other factors such as co-infecting microorganisms.

The Cell Tropism of Porcine Respiratory Coronavirus for Airway Epithelial Cells Is Determined by the Expression of Porcine Aminopeptidase N.

Porcine respiratory coronavirus (PRCoV) infects the epithelial cells in the respiratory tract of pigs, causing a mild respiratory disease. We applied air-liquid interface (ALI) cultures of well-differentiated porcine airway cells to mimic the respiratory tract epithelium in vitro and use it for analyzing the infection by PRCoV. As reported for most coronaviruses, virus entry and virus release occurred mainly via the apical membrane domain. A novel finding was that PRCoV preferentially targets non-ciliated and among them the non-mucus-producing cells. Aminopeptidase N (APN), the cellular receptor for PRCoV was also more abundantly expressed on this type of cell suggesting that APN is a determinant of the cell tropism. Interestingly, differentiation-dependent differences were found both in the expression of pAPN and the susceptibility to PRCoV infection. Cells in an early differentiation stage express higher levels of pAPN and are more susceptible to infection by PRCoV than are well-differentiated cells. A difference in the susceptibility to infection was also detected when tracheal and bronchial cells were compared. The increased susceptibility to infection of bronchial epithelial cells was, however, not due to an increased abundance of APN on the cell surface. Our data reveal a complex pattern of infection in porcine differentiated airway epithelial cells that could not be elucidated with immortalized cell lines. The results are expected to have relevance also for the analysis of other respiratory viruses.

Fusogenicity of the Ghana Virus (Henipavirus: Ghanaian bat henipavirus) Fusion Protein is Controlled by the Cytoplasmic Domain of the Attachment Glycoprotein.

The Ghana virus (GhV) is phylogenetically related to the zoonotic henipaviruses Nipah (NiV) and Hendra virus. Although GhV uses the highly conserved receptor ephrin-B2, the fusogenicity is restricted to cell lines of bat origin. Furthermore, the surface expression of the GhV attachment glycoprotein (G) is reduced compared to NiV and most of this protein is retained in the endoplasmic reticulum (ER). Here, we generated truncated as well as chimeric GhV G proteins and investigated the influence of the structural domains (cytoplasmic tail, transmembrane domain, ectodomain) of this protein on the intracellular transport and the fusogenicity following coexpression with the GhV fusion protein (F). We demonstrate that neither the cytoplasmic tail nor the transmembrane domain is responsible for the intracellular retention of GhV G. Furthermore, the cytoplasmic tail of GhV G modulates the fusogenicity of GhV F and therefore controls the species-restricted fusogenicity of the GhV surface glycoproteins.

Viral Coinfection Replaces Effects of Suilysin on Streptococcus suis Adherence to and Invasion of Respiratory Epithelial Cells Grown under Air-Liquid Interface Conditions.

Streptococcus suis is an important zoonotic pathogen which can infect humans and pigs worldwide, posing a potential risk to global public health. Suilysin, a pore-forming cholesterol-dependent cytolysin, is considered to play an important role in the pathogenesis of S. suis infections. It is known that infection with influenza A viruses may favor susceptibility to secondary bacterial infection, resulting in more severe disease and increased mortality. However, the molecular mechanisms underlying these coinfections are incompletely understood. Applying highly differentiated primary porcine respiratory epithelial cells grown under air-liquid interface (ALI) conditions, we analyzed the contribution of swine influenza viruses (SIV) to the virulence of S. suis, with a special focus on its cytolytic toxin, suilysin. We found that during secondary bacterial infection, suilysin of S. suis contributed to the damage of well-differentiated respiratory epithelial cells in the early stage of infection, whereas the cytotoxic effects induced by SIV became prominent at later stages of infection. Prior infection by SIV enhanced the adherence to and colonization of porcine airway epithelial cells by a wild-type (wt) S. suis strain and a suilysin-negative S. suis mutant in a sialic acid-dependent manner. A striking difference was observed with respect to bacterial invasion. After bacterial monoinfection, only the wt S. suis strain showed an invasive phenotype, whereas the mutant remained adherent. When the epithelial cells were preinfected with SIV, the suilysin-negative mutant also showed an invasion capacity. Therefore, we propose that coinfection with SIV may compensate for the lack of suilysin in the adherence and invasion process of suilysin-negative S. suis.

Infection Studies in Pigs and Porcine Airway Epithelial Cells Reveal an Evolution of A(H1N1)pdm09 Influenza A Viruses Toward Lower Virulence.

We analyzed the virulence of pandemic H1N1 2009 influenza A viruses in vivo and in vitro. Selected viruses isolated in 2009, 2010, 2014, and 2015 were assessed using an aerosol-mediated high-dose infection model for pigs as well as air-liquid interface cultures of differentiated airway epithelial cells. Using a dyspnea score, rectal temperature, lung lesions, and viral load in the lung as parameters, the strains from 2014-2015 were significantly less virulent than the strains isolated in 2009-2010. In vitro, the viruses from 2009-2010 also differed from the 2014-2015 viruses by increased release of infectious virus, a more pronounced loss of ciliated cells, and a reduced thickness of the epithelial cell layer. Our in vivo and in vitro results reveal an evolution of A(H1N1)pdm09 viruses toward lower virulence. Our in vitro culture system can be used to predict the virulence of influenza viruses.

The Sialic Acid Binding Activity of Human Parainfluenza Virus 3 and Mumps Virus Glycoproteins Enhances the Adherence of Group B Streptococci to HEp-2 Cells.

In the complex microenvironment of the human respiratory tract, different kinds of microorganisms may synergistically interact with each other resulting in viral-bacterial co-infections that are often associated with more severe diseases than the respective mono-infections. Human respiratory paramyxoviruses, for example parainfluenza virus type 3 (HPIV3), are common causes of respiratory diseases both in infants and a subset of adults. HPIV3 recognizes sialic acid (SA)-containing receptors on host cells. In contrast to human influenza viruses which have a preference for α2,6-linked sialic acid, HPIV3 preferentially recognize α2,3-linked sialic acids. Group B streptococci (GBS) are colonizers in the human respiratory tract. They contain a capsular polysaccharide with terminal sialic acid residues in an α2,3-linkage. In the present study, we report that HPIV3 can recognize the α2,3-linked sialic acids present on GBS. The interaction was evident not only by the binding of virions to GBS in a co-sedimentation assay, but also in the GBS binding to HPIV3-infected cells. While co-infection by GBS and HPIV3 had a delaying effect on the virus replication, it enhanced GBS adherence to virus-infected cells. To show that other human paramyxoviruses are also able to recognize the capsular sialic acid of GBS we demonstrate that GBS attaches in a sialic acid-dependent way to transfected BHK cells expressing the HN protein of mumps virus (MuV) on their surface. Overall, our results reveal a new type of synergism in the co-infection by respiratory pathogens, which is based on the recognition of α2,3-linked sialic acids. This interaction between human paramyxoviruses and GBS enhances the bacterial adherence to airway cells and thus may result in more severe disease.

Ciliostasis of airway epithelial cells facilitates influenza A virus infection.

Porcine precision-cut lung slices (PCLS) were used to analyze the effect of the ciliary activity on infection of airway epithelial cells by influenza viruses. Treatment of slices with 2% NaCl for 30 min resulted in reversible ciliostasis. When PCLS were infected by a swine influenza virus of the H3N2 subtype under ciliostatic conditions, the viral yield was about twofold or threefold higher at 24 or 48 h post-infection, respectively, as compared to slices with ciliary activity. Therefore, the cilia beating not only transports the mucus out of the airways, it also impedes virus infection.

Increased virulence of a PB2/HA mutant of an avian H9N2 influenza strain after three passages in porcine differentiated airway epithelial cells.

We analyzed the adaptation of influenza viruses to growth in differentiated airway epithelial cells of a new host by passaging an avian H9N2 virus three times in porcine precision-cut lung slices (PCLS). Sequence analysis revealed four mutations: one each in the PB2 and NS1 proteins, and two in the HA protein. In this study, we characterized the PB2 mutation G685R by generating recombinant H9N2 viruses containing the PB2 single mutation alone or in combination with one of the HA mutations (A190V or T212I). When analyzed in porcine cells - a tracheal cell line (NPTr) or PCLS - the PB2-685 mutant did not provide a growth advantage and had no effect on the ciliary activity which is a virulence marker of swine influenza viruses. Pathogenicity for mice was also not increased by the single PB2 mutation. However, both double mutants (HA-190+PB2-685 and HA-212+PB2-685) showed significantly increased virulence in mice. Therefore, the mutations in the HA and PB2 proteins may confer early adaptation of an avian H9N2 virus to a mammalian host. In conclusion, we expect that a broader ensemble of mutations will be required to render an H9N2 virus virulent for pigs.

The differentiated airway epithelium infected by influenza viruses maintains the barrier function despite a dramatic loss of ciliated cells.

Virus-host interactions in the respiratory epithelium during long term influenza virus infection are not well characterized. Therefore, we developed an air-liquid interface culture system for differentiated porcine respiratory epithelial cells to study the effect of virus-induced cellular damage. In our well-differentiated cells, α2,6-linked sialic acid is predominantly expressed on the apical surface and the basal cells mainly express α2,3-linked sialic acid. During the whole infection period, release of infectious virus was maintained at a high titre for more than seven days. The infected epithelial cells were subject to apoptosis resulting in the loss of ciliated cells together with a thinner thickness. Nevertheless, the airway epithelium maintained trans-epithelial electrical resistance and retained its barrier function. The loss of ciliated cells was compensated by the cells which contained the KRT5 basal cell marker but were not yet differentiated into ciliated cells. These specialized cells showed an increase of α2,3-linked sialic acid on the apical surface. In sum, our results help to explain the localized infection of the airway epithelium by influenza viruses. The impairment of mucociliary clearance in the epithelial cells provides an explanation why prior viral infection renders the host more susceptible to secondary co-infection by another pathogen.

Recombinant mumps viruses expressing the batMuV fusion glycoprotein are highly fusion active and neurovirulent.

A recent study reported the detection of a bat-derived virus (BatPV/Epo_spe/AR1/DCR/2009, batMuV) with phylogenetic relatedness to human mumps virus (hMuV). Since all efforts to isolate infectious batMuV have reportedly failed, we generated recombinant mumps viruses (rMuVs) in which the open reading frames (ORFs) of the fusion (F) and haemagglutinin-neuraminidase (HN) glycoproteins of an hMuV strain were replaced by the corresponding ORFs of batMuV. The batMuV F and HN proteins were successfully incorporated into viral particles and the resultant chimeric virus was able to mediate infection of Vero cells. Distinct differences were observed between the fusogenicity of rMuVs expressing one or both batMuV glycoproteins: viruses expressing batMuV F were highly fusogenic, regardless of the origin of HN. In contrast, rMuVs expressing human F and bat-derived HN proteins were less fusogenic compared to hMuV. The growth kinetics of chimeric MuVs expressing batMuV HN in combination with either hMuV or batMuV F were similar to that of the backbone virus, whereas a delay in virus replication was obtained for rMuVs harbouring batMuV F and hMuV HN. Replacement of the hMuV F and HN genes or the HN gene alone by the corresponding batMuV genes led to a slight reduction in neurovirulence of the highly neurovirulent backbone strain. Neutralizing antibodies inhibited infection mediated by all recombinant viruses generated. Furthermore, group IV anti-MuV antibodies inhibited the neuraminidase activity of bat-derived HN. Our study reports the successful generation of chimeric MuVs expressing the F and HN proteins of batMuV, providing a means for further examination of this novel batMuV.

Mycoplasma hyopneumoniae does not affect the interferon-related anti-viral response but predisposes the pig to a higher level of inflammation following swine influenza virus infection.

In pigs, influenza A viruses and Mycoplasma hyopneumoniae (Mhp) are major contributors to the porcine respiratory disease complex. Pre-infection with Mhp was previously shown experimentally to exacerbate the clinical outcomes of H1N1 infection during the first week after virus inoculation. In order to better understand the interactions between these pathogens, we aimed to assess very early responses (at 5, 24 and 48 h) after H1N1 infection in pigs pre-infected or not with Mhp. Clinical signs and macroscopic lung lesions were similar in both infected groups at early times post-H1N1 infection; and Mhp pre-infection affected neither the influenza virus replication nor the IFN-induced antiviral responses in the lung. However, it predisposed the animals to a higher inflammatory response to H1N1 infection, as revealed by the massive infiltration of neutrophils and macrophages into the lungs and the increased production of pro-inflammatory cytokines (IL-6, IL-1ß and TNF-α). Thus, it seems it is this marked inflammatory state that would play a role in exacerbating the clinical signs subsequent to H1N1 infection.

Porcine epidemic diarrhea virus inhibits dsRNA-induced interferon-ß production in porcine intestinal epithelial cells by blockade of the RIG-I-mediated pathway.

BACKGROUND: The lack of optimal porcine cell lines has severely impeded the study and progress in elucidation of porcine epidemic diarrhea virus (PEDV) pathogenesis. Vero cell, an African green monkey kidney cell line, was often used to isolate and propagate PEDV. Nonetheless, the target cells of PEDV in vivo are intestinal epithelial cells, during infection, intestinal epithelia would be damaged and resulted in digestive disorders. The immune functions of porcine epithelial cells and interactions with other immune cell populations display a number of differences compared to other species. Type I interferon (IFN) plays an important role in antiviral immune response. Limited reports showed that PEDV could inhibit type I interferon production. In this study, porcine small intestinal epithelial cells (IECs), the target cells of PEDV, were used as the infection model in vitro to identify the possible molecular mechanisms of PEDV-inhibition IFN-ß production. RESULTS: PEDV not only failed to induce IFN-ß expression, but also inhibited dsRNA-mediated IFN-ß production in IECs. As the key IFN-ß transcription factors, we found that dsRNA-induced activation of IFN regulatory factor 3 (IRF-3) was inhibited after PEDV infection, but not nuclear factor-kappaB (NF-κB). To identify the mechanism of PEDV intervention with dsRNA-mediated IFN-ß expression more accurately, the role of individual molecules of RIG-I signaling pathway were investigated. In the upstream of IRF-3, TANK-binding kinase 1 (TBK1)-or inhibitor of κB kinase-ε (IKKε)-mediated IFN-ß production was not blocked by PEDV, while RIG-I-and its adapter molecule IFN-ß promoter stimulator 1 (IPS-1)-mediated IFN-ß production were completely inhibited after PEDV infection. CONCLUSION: Taken together, our data demonstrated for the first time that PEDV infection of its target cell line, IECs, inhibited dsRNA-mediated IFN-ß production by blocking the activation of IPS-1 in RIG-I-mediated pathway. Our studies offered new visions in understanding of the interaction between PEDV and host innate immune system.

Analysis of Ebola Virus Entry Into Macrophages.

Ebolaviruses constitute a public health threat, particularly in Central and Western Africa. Host cell factors required for spread of ebolaviruses may serve as targets for antiviral intervention. Lectins, TAM receptor tyrosine kinases (Tyro3, Axl, Mer), T cell immunoglobulin and mucin domain (TIM) proteins, integrins, and Niemann-Pick C1 (NPC1) have been reported to promote entry of ebolaviruses into certain cellular systems. However, the factors used by ebolaviruses to invade macrophages, major viral targets, are poorly defined. Here, we show that mannose-specific lectins, TIM-1 and Axl augment entry into certain cell lines but do not contribute to Ebola virus (EBOV)-glycoprotein (GP)-driven transduction of macrophages. In contrast, expression of Mer, integrin αV, and NPC1 was required for efficient GP-mediated transduction and EBOV infection of macrophages. These results define cellular factors hijacked by EBOV for entry into macrophages and, considering that Mer and integrin αV promote phagocytosis of apoptotic cells, support the concept that EBOV relies on apoptotic mimicry to invade target cells.

Functional properties and genetic relatedness of the fusion and hemagglutinin-neuraminidase proteins of a mumps virus-like bat virus.

UNLABELLED: A bat virus with high phylogenetic relatedness to human mumps virus (MuV) was identified recently at the nucleic acid level. We analyzed the functional activities of the hemagglutinin-neuraminidase (HN) and the fusion (F) proteins of the bat virus (batMuV) and compared them to the respective proteins of a human isolate. Transfected cells expressing the F and HN proteins of batMuV were recognized by antibodies directed against these proteins of human MuV, indicating that both viruses are serologically related. Fusion, hemadsorption, and neuraminidase activities were demonstrated for batMuV, and either bat-derived protein could substitute for its human MuV counterpart in inducing syncytium formation when coexpressed in different mammalian cell lines, including chiropteran cells. Cells expressing batMuV glycoproteins were shown to have lower neuraminidase activity. The syncytia were smaller, and they were present in lower numbers than those observed after coexpression of the corresponding glycoproteins of a clinical isolate of MuV (hMuV). The phenotypic differences in the neuraminidase and fusion activity between the glycoproteins of batMuV and hMuV are explained by differences in the expression level of the HN and F proteins of the two viruses. In the case of the F protein, analysis of chimeric proteins revealed that the signal peptide of the bat MuV fusion protein is responsible for the lower surface expression. These results indicate that the surface glycoproteins of batMuV are serologically and functionally related to those of hMuV, raising the possibility of bats as a reservoir for interspecies transmission. IMPORTANCE: The recently described MuV-like bat virus is unique among other recently identified human-like bat-associated viruses because of its high sequence homology (approximately 90% in most genes) to its human counterpart. Although it is not known if humans can be infected by batMuV, the antigenic relatedness between the bat and human forms of the virus suggests that humans carrying neutralizing antibodies against MuV are protected from infection by batMuV. The close functional relationship between MuV and batMuV is demonstrated by cooperation of the respective HN and F proteins to induce syncytium formation in heterologous expression studies. An interesting feature of the glycoproteins of batMuV is the downregulation of the fusion activity by the signal peptide of F, which has not been reported for other paramyxoviruses. These results are important contributions for risk assessment and for a better understanding of the replication strategy of batMuV.