Tula Virus Research Articles

Replication kinetics of pathogenic Eurasian orthohantaviruses in human mesangial cells

BackgroundEurasian pathogenic orthohantaviruses cause hemorrhagic fever with renal syndrome (HFRS) characterized by acute kidney injury (AKI). The virulence of orthohantaviruses varies enormously and direct infection of different renal cell types contribute to pathogenesis. Glomerular mesangial cells play an essential role in the interplay between kidney cells and proper kidney function. Therefore, we analyzed the replication competence of different orthohantavirus species in primary mesangial cells and a mesangial cell line.MethodsWe tested the suitability of the mesangial cell line CIHGM-1 (conditionally immortalized human glomerular mesangial cells) as cell culture model for orthohantavirus kidney infection by comparison with primary human renal mesangial cells (HRMCs). We analyzed infection with high pathogenic Hantaan virus (HTNV), moderate pathogenic Puumala virus (PUUV) and non-/low-pathogenic Tula virus (TULV).ResultsEffective viral spread was observed for PUUV only, whereas infection with HTNV and TULV was abortive. However, in contrast to TULV, HTNV exhibits an initially high infection rate and declines afterwards. This replication pattern was observed in HRMCs and CIHGM-1 cells. Viability or adhesion was neither impaired for PUUV-infected CIHGM-1 nor HRMCs. A loss of migration capacity was observed in PUUV-infected CIHGM-1 cells, but not in HRMCs.ConclusionsThe identification of differences in the replication competence of pathogenic orthohantavirus strains in renal mesangial cells is of special interest and may provide useful insights in the virus-specific mechanisms of orthohantavirus induced AKI. The use of CIHGM-1 cells will facilitate the research in a relevant cell culture system.

Suppression of Interferon Response and Antiviral Strategies of Bunyaviruses.

The order Bunyavirales belongs to the class of Ellioviricetes and is classified into fourteen families. Some species of the order Bunyavirales pose potential threats to human health. The continuously increasing research reveals that various viruses within this order achieve immune evasion in the host through suppressing interferon (IFN) response. As the types and nodes of the interferon response pathway are continually updated or enriched, the IFN suppression mechanisms and target points of different virus species within this order are also constantly enriched and exhibit variations. For instance, Puumala virus (PUUV) and Tula virus (TULV) can inhibit IFN response through their functional NSs inhibiting downstream factor IRF3 activity. Nevertheless, the IFN suppression mechanisms of Dabie bandavirus (DBV) and Guertu virus (GTV) are mostly mediated by viral inclusion bodies (IBs) or filamentous structures (FSs). Currently, there are no effective drugs against several viruses belonging to this order that pose significant threats to society and human health. While the discovery, development, and application of antiviral drugs constitute a lengthy process, our focus on key targets in the IFN response suppression process of the virus leads to potential antiviral strategies, which provide references for both basic research and practical applications.

Genome-wide support for incipient Tula hantavirus species within a single rodent host lineage.

Evolutionary divergence of viruses is most commonly driven by co-divergence with their hosts or through isolation of transmission after host shifts. It remains mostly unknown, however, whether divergent phylogenetic clades within named virus species represent functionally equivalent byproducts of high evolutionary rates or rather incipient virus species. Here, we test these alternatives with genomic data from two widespread phylogenetic clades in Tula orthohantavirus (TULV) within a single evolutionary lineage of their natural rodent host, the common vole Microtus arvalis. We examined voles from forty-two locations in the contact region between clades for TULV infection by reverse transcription (RT)-PCR. Sequencing yielded twenty-three TULV Central North and twenty-one TULV Central South genomes, which differed by 14.9-18.5 per cent at the nucleotide and 2.2-3.7 per cent at the amino acid (AA) level without evidence of recombination or reassortment between clades. Geographic cline analyses demonstrated an abrupt (<1 km wide) transition between the parapatric TULV clades in continuous landscape. This transition was located within the Central mitochondrial lineage of M. arvalis, and genomic single nucleotide polymorphisms showed gradual mixing of host populations across it. Genomic differentiation of hosts was much weaker across the TULV Central North to South transition than across the nearby hybrid zone between two evolutionary lineages in the host. We suggest that these parapatric TULV clades represent functionally distinct, incipient species, which are likely differently affected by genetic polymorphisms in the host. This highlights the potential of natural viral contact zones as systems for investigating the genetic and evolutionary factors enabling or restricting the transmission of RNA viruses.

Differences in the Susceptibility of Human Tubular Epithelial Cells for Infection with Orthohantaviruses.

Diseases induced by infection with pathogenic orthohantaviruses are characterized by a pronounced organ-specific manifestation. Pathogenic Eurasian orthohantaviruses cause hemorrhagic fever with renal syndrome (HFRS) with often massive proteinuria. Therefore, the use of a relevant kidney cell culture would be favorable to analyze the underlying cellular mechanisms of orthohantavirus-induced acute kidney injury (AKI). We tested different human tubular epithelial cell lines for their suitability as an in vitro infection model. Permissiveness and replication kinetics of highly pathogenic Hantaan virus (HTNV) and non-/low-pathogenic Tula virus (TULV) were analyzed in tubular epithelial cell lines and compared to human primary tubular epithelial cells. Ana-lysis of the cell line HK-2 revealed the same results for viral replication, morphological and functional effects as observed for HTNV in primary cells. In contrast, the cell lines RPTEC/TERT1 and TH1 demonstrated only poor infection rates after inoculation with HTNV and are unusable as an infection model. While pathogenic HNTV infects primary tubular and HK-2 cells, non-/low-pathogenic TULV infects neither primary tubular cells nor the cell line HK-2. Our results show that permissiveness of renal cells varies between orthohantaviruses with differences in pathogenicity and that HK-2 cells demonstrate a suitable in vitro model to study viral tropism and pathogenesis of orthohantavirus-induced AKI.

Diverse susceptibilities and responses of human and rodent cells to orthohantavirus infection reveal different levels of cellular restriction.

Orthohantaviruses are rodent-borne emerging viruses that may cause severe diseases in humans but no apparent pathology in their small mammal reservoirs. However, the mechanisms leading to tolerance or pathogenicity in humans and persistence in rodent reservoirs are poorly understood, as is the manner in which they spread within and between organisms. Here, we used a range of cellular and molecular approaches to investigate the interactions of three different orthohantaviruses–Puumala virus (PUUV), responsible for a mild to moderate form of hemorrhagic fever with renal syndrome in humans, Tula virus (TULV) with low pathogenicity, and non-pathogenic Prospect Hill virus (PHV)–with human and rodent host cell lines. Besides the fact that cell susceptibility to virus infection was shown to depend on the cell type and virus strain, the three orthohantaviruses were able to infect Vero E6 and HuH7 human cells, but only the former secreted infectious particles. In cells derived from PUUV reservoir, the bank vole (Myodes glareolus), PUUV achieved a complete viral cycle, while TULV did not enter the cells and PHV infected them but did not produce infectious particles, reflecting differences in host specificity. A search for mature virions by electron microscopy (EM) revealed that TULV assembly occurred in part at the plasma membrane, whereas PHV particles were trapped in autophagic vacuoles in cells of the heterologous rodent host. We described differential interactions of orthohantaviruses with cellular factors, as supported by the cellular distribution of viral nucleocapsid protein with cell compartments, and proteomics identification of cellular partners. Our results also showed that interferon (IFN) dependent gene expression was regulated in a cell and virus species dependent manner. Overall, our study highlighted the complexity of the host-virus relationship and demonstrated that orthohantaviruses are restricted at different levels of the viral cycle. In addition, the study opens new avenues to further investigate how these viruses differ in their interactions with cells to evade innate immunity and how it depends on tissue type and host species.

Orthohantaviruses in Reservoir and Atypical Hosts in the Czech Republic: Spillover Infection and Indication of Virus-Specific Tissue Tropism.

ABSTRACTOrthohantaviruses (genus Orthohantavirus) are a diverse group of viruses that are closely associated with their natural hosts (rodents, shrews, and moles). Several orthohantaviruses cause severe disease in humans. Central and western Europe are areas with emerging orthohantavirus occurrences. In our study, several orthohantaviruses, including the pathogenic Kurkino virus (KURV), were detected in their natural hosts trapped at several study sites in the Czech Republic. KURV was detected mainly in its typical host, the striped field mouse (Apodemus agrarius). Nevertheless, spillover infections were also detected in wood mice (Apodemus sylvaticus) and common voles (Microtus arvalis). Similarly, Tula virus (TULV) was found primarily in common voles, and events of spillover to rodents of other host species, including Apodemus spp., were recorded. In addition, unlike most previous studies, different tissues were sampled and compared to assess their suitability for orthohantavirus screening and possible tissue tropism. Our data suggest possible virus-specific tissue tropism in rodent hosts. TULV was most commonly detected in the lung tissue, whereas KURV was more common in the liver, spleen, and brain. Moreover, Seewis and Asikkala viruses were detected in randomly found common shrews (Sorex araneus). In conclusion, we have demonstrated the presence of human-pathogenic KURV and the potentially pathogenic TULV in their typical hosts as well as their spillover to atypical host species belonging to another family. Furthermore, we suggest the possibility of virus-specific tissue tropism of orthohantaviruses in their natural hosts.IMPORTANCE Orthohantaviruses (genus Orthohantavirus, family Hantaviridae) are a diverse group of globally distributed viruses that are closely associated with their natural hosts. Some orthohantaviruses are capable of infecting humans and causing severe disease. Orthohantaviruses are considered emerging pathogens due to their ever-increasing diversity and increasing numbers of disease cases. We report the detection of four different orthohantaviruses in rodents and shrews in the Czech Republic. Most viruses were found in their typical hosts, Kurkino virus (KURV) in striped field mice (Apodemus agrarius), Tula virus (TULV) in common voles (Microtus arvalis), and Seewis virus in common shrews (Sorex araneus). Nevertheless, spillover infections of atypical host species were also recorded for KURV, TULV, and another shrew-borne orthohantavirus, Asikkala virus. In addition, indications of virus-specific patterns of tissue tropism were observed. Our results highlight the circulation of several orthohantaviruses, including KURV, which is pathogenic to humans, among rodents and shrews in the Czech Republic.

Molecular Characterisation and Phylogeny of Tula Virus in Kazakhstan.

Orthohantaviruses are zoonotic pathogens that play a significant role in public health. These viruses can cause haemorrhagic fever with renal syndrome in Eurasia. In the Republic of Kazakhstan, the first human cases were registered in the year 2000 in the West Kazakhstan region. Small mammals can be reservoirs of orthohantaviruses. Previous studies showed orthohantavirus antigens in wild-living small mammals in four districts of West Kazakhstan. Clinical studies suggested that there might be further regions with human orthohantavirus infections in Kazakhstan, but genetic data of orthohantaviruses in natural foci are limited. The aim of this study was to investigate small mammals for the presence of orthohantaviruses by molecular biological methods and to provide a phylogenetic characterization of the circulating strains in Kazakhstan. Small mammals were trapped at 19 sites in West Kazakhstan, four in Almaty region and at seven sites around Almaty city during all seasons of 2018 and 2019. Lung tissues of small mammals were homogenized and RNA was extracted. Orthohantavirus RT-PCR assays were applied for detection of partial S and L segment sequences. Results were compared to published fragments. In total, 621 small mammals from 11 species were analysed. Among the collected small mammals, 2.4% tested positive for orthohantavirus RNA, one sample from West Kazakhstan and 14 samples from Almaty region. None of the rodents caught in Almaty city were infected. Sequencing parts of the small (S) and large (L) segments specified Tula virus (TULV) in these two regions. Our data show that geographical distribution of TULV is more extended as previously thought. The detected sequences were found to be split in two distinct genetic clusters of TULV in West Kazakhstan and Almaty region. TULV was detected in the common vole (Microtus arvalis) and for the first time in two individuals of the forest dormouse (Dryomys nitedula), interpreted as a spill-over infection in Kazakhstan.

Expression Profile of Human Renal Mesangial Cells Is Altered by Infection with Pathogenic Puumala Orthohantavirus.

Acute kidney injury (AKI) with proteinuria is a hallmark of infections with Eurasian orthohantaviruses. Different kidney cells are identified as target cells of hantaviruses. Mesangial cells may play a central role in the pathogenesis of AKI by regulation of inflammatory mediators and signaling cascades. Therefore, we examined the characteristics of hantavirus infection on human renal mesangial cells (HRMCs). Receptor expression and infection with pathogenic Puumala virus (PUUV) and low-pathogenic Tula virus (TULV) were explored. To analyze changes in protein expression in infected mesangial cells, we performed a proteome profiler assay analyzing 38 markers of kidney damage. We compared the proteome profile of in vitro-infected HRMCs with the profile detected in urine samples of 11 patients with acute hantavirus infection. We observed effective productive infection of HRMCs with pathogenic PUUV, but only poor abortive infection for low-pathogenic TULV. PUUV infection resulted in the deregulation of proteases, adhesion proteins, and cytokines associated with renal damage. The urinary proteome profile of hantavirus patients demonstrated also massive changes, which in part correspond to the alterations observed in the in vitro infection of HRMCs. The direct infection of mesangial cells may induce a local environment of signal mediators that contributes to AKI in hantavirus infection.

Tula Virus as Causative Agent of Hantavirus Disease in Immunocompetent Person, Germany.

We report molecular evidence of Tula virus infection in an immunocompetent patient from Germany who had typical signs of hantavirus disease. Accumulating evidence indicates that Tula virus infection, although often considered nonpathogenic, represents a threat to human health.

Effectiveness Regarding Hantavirus Detection in Rodent Tissue Samples and Urine.

The natural hosts of Orthohantaviruses are rodents, soricomorphs and bats, and it is well known that they may cause serious or even fatal diseases among humans worldwide. The virus is persistent among animals and it is shed via urine, saliva and feces throughout the entirety of their lives. We aim to identify the effectiveness of hantavirus detection in rodent tissue samples and urine originating from naturally infected rodents. Initially, animals were trapped at five distinct locations throughout the Transdanubian region in Hungary. Lung, liver, kidney and urine samples were obtained from 163 deceased animals. All organs and urine were tested using nested reverse transcription polymerase chain reaction (nRT-PCR). Furthermore, sera were examined for IgG antibodies against Dobrava–Belgrade virus (DOBV) and Puumala virus (PUUV) by Western blot assay. IgG antibodies against hantaviruses and/or nucleic acid were detected in 25 (15.3%) cases. Among Apodemus, Myodes, and Microtus rodent species, DOBV, PUUV and Tula virus (TULV) were clearly identified. Amid the PCR-positive samples, the nucleic acid of the viruses was detected most effectively in the kidney (100%), while only 55% of screened lung tissues were positive. Interestingly, only three out of 20 rodent urine samples were positive when tested using nRT-PCR. Moreover, five rodents were seropositive without detectable virus nucleic acid in any of the tested organs.

The RNA Replication Site of Tula Orthohantavirus Resides within a Remodelled Golgi Network.

The family Hantaviridae within the Bunyavirales order comprises tri-segmented negative sense RNA viruses, many of which are rodent-borne emerging pathogens associated with fatal human disease. In contrast, hantavirus infection of corresponding rodent hosts results in inapparent or latent infections, which can be recapitulated in cultured cells that become persistently infected. In this study, we used Tula virus (TULV) to investigate the location of hantavirus replication during early, peak and persistent phases of infection, over a 30-day time course. Using immunofluorescent (IF) microscopy, we showed that the TULV nucleocapsid protein (NP) is distributed within both punctate and filamentous structures, with the latter increasing in size as the infection progresses. Transmission electron microscopy of TULV-infected cell sections revealed these filamentous structures comprised aligned clusters of filament bundles. The filamentous NP-associated structures increasingly co-localized with the Golgi and with the stress granule marker TIA-1 over the infection time course, suggesting a redistribution of these cellular organelles. The analysis of the intracellular distribution of TULV RNAs using fluorescent in-situ hybridization revealed that both genomic and mRNAs co-localized with Golgi-associated filamentous compartments that were positive for TIA. These results show that TULV induces a dramatic reorganization of the intracellular environment, including the establishment of TULV RNA synthesis factories in re-modelled Golgi compartments.

Replication in the Mononuclear Phagocyte System (MPS) as a Determinant of Hantavirus Pathogenicity.

Members of different virus families including Hantaviridae cause viral hemorrhagic fevers (VHFs). The decisive determinants of hantavirus-associated pathogenicity are still enigmatic. Pathogenic hantavirus species, such as Puumala virus (PUUV), Hantaan virus (HTNV), Dobrava-Belgrade virus (DOBV), and Sin Nombre virus (SNV), are associated with significant case fatality rates. In contrast, Tula virus (TULV) only sporadically causes mild disease in immunocompetent humans and Prospect Hill virus (PHV) so far has not been associated with any symptoms. They are thus defined here as low pathogenic/apathogenic hantavirus species. We found that productive infection of cells of the mononuclear phagocyte system (MPS), such as monocytes and dendritic cells (DCs), correlated well with the pathogenicity of hantavirus species tested. HTNV (intermediate case fatality rates) replicated more efficiently than PUUV (low case fatality rates) in myeloid cells, whereas low pathogenic/apathogenic hantavirus species did not produce any detectable virus titers. Analysis of PHPUV, a reassortant hantavirus derived from a pathogenic (PUUV) and an apathogenic (PHV) hantavirus species, indicated that the viral glycoproteins are not decisive for replication in MPS cells. Moreover, blocking acidification of endosomes with chloroquine decreased the number of TULV genomes in myeloid cells suggesting a post-entry block for low pathogenic/apathogenic hantavirus species in myeloid cells. Intriguingly, pathogenic but not low pathogenic/apathogenic hantavirus species induced conversion of monocytes into inflammatory DCs. The proinflammatory programming of MPS cells by pathogenic hantavirus species required integrin signaling and viral replication. Our findings indicate that the capacity to replicate in MPS cells is a prominent feature of hantaviral pathogenicity.

Puumala and Tula Virus Differ in Replication Kinetics and Innate Immune Stimulation in Human Endothelial Cells and Macrophages.

Old world hantaviruses cause hemorrhagic fever with renal syndrome (HFRS) upon zoonotic transmission to humans. In Europe, the Puumala virus (PUUV) is the main causative agent of HFRS. Tula virus (TULV) is also widely distributed in Europe, but there is little knowledge about the pathogenicity of TULV for humans, as reported cases are rare. We studied the replication of TULV in different cell types in comparison to the pathogenic PUUV and analyzed differences in stimulation of innate immunity. While both viruses replicated to a similar extent in interferon (IFN)-deficient Vero E6 cells, TULV replication in human lung epithelial (A549) cells was slower and less efficient when compared to PUUV. In contrast to PUUV, no replication of TULV could be detected in human microvascular endothelial cells and in macrophages. While a strong innate immune response towards PUUV infection was evident at 48 h post infection, TULV infection triggered only a weak IFN response late after infection of A549 cells. Using appropriate in vitro cell culture models for the orthohantavirus infection, we could demonstrate major differences in host cell tropism, replication kinetics, and innate immune induction between pathogenic PUUV and the presumably non- or low-pathogenic TULV that are not observed in Vero E6 cells and may contribute to differences in virulence.

Revisiting the genetic diversity of emerging hantaviruses circulating in Europe using a pan-viral resequencing microarray

Hantaviruses are zoonotic agents transmitted from small mammals, mainly rodents, to humans, where they provoke diseases such as Hemorrhagic fever with Renal Syndrome (HFRS) and its mild form, Nephropathia Epidemica (NE), or Hantavirus Cardio-Pulmonary Syndrome (HCPS). Hantaviruses are spread worldwide and monitoring animal reservoirs is of primary importance to control the zoonotic risk. Here, we describe the development of a pan-viral resequencing microarray (PathogenID v3.0) able to explore the genetic diversity of rodent-borne hantaviruses endemic in Europe. Among about 800 sequences tiled on the microarray, 52 correspond to a tight molecular sieve of hantavirus probes covering a large genetic landscape. RNAs from infected animal tissues or from laboratory strains have been reverse transcribed, amplified, then hybridized to the microarray. A classical BLASTN analysis applied to the sequence delivered through the microarray allows to identify the hantavirus species up to the exact geographical variant present in the tested samples. Geographical variants of the most common European hantaviruses from France, Germany, Slovenia and Finland, such as Puumala virus, Dobrava virus and Tula virus, were genetically discriminated. Furthermore, we precisely characterized geographical variants still unknown when the chip was conceived, such as Seoul virus isolates, recently emerged in France and the United Kingdom.

Identification and characterization of a novel subtype of Tula virus in Microtus arvalis obscurus voles sampled from Xinjiang, China

Although most of Arvicolinae associated hantaviruses can not cause disease in humans, hemorrhagic fever with renal syndrome (HFRS) cases caused by Tula virus (TULV) have been described in Europe since 2002. In addition to Europe, TULV was also identified in the Microtus arvalis obscurus voles sampled from Kazakhstan, which shares borders with China. To gain more insight into the molecular epidemiology of TULV, a total of 365 rodents representing 7 species of 4 subfamily (Arvicolinae, Murinae, Gerbillinae, and Cricetinae) were captured in Qapqal county, Xinjiang, northwest China. Hantavirus RNA was recovered from 40 lung tissue samples of M. arvalis obscurus, with the prevalence of 10.96%. Genetic analysis revealed that all recovered viral sequences were most closely related to those of TULV, but exhibited >11% nucleotide differences from all currently known TULV, suggesting that they may represent a new subtype of TULV. In the S tree, the newly identified viruses formed a distinct lineage and showed a close evolutionary relationship with those sampled from Southwestern Siberia and Kazakhstan. However, they exhibited a different clustering pattern in both the M and the L trees, suggesting the possibility of genetic reassortment. Finally, the recombination event was also observed in Xinjiang TULV viruses. In sum, all these data reveal a complex evolutionary history of TULV in Central Asia.

Tula orthohantavirus nucleocapsid protein is cleaved in infected cells and may sequester activated caspase-3 during persistent infection to suppress apoptosis.

The family Hantaviridae mostly comprises rodent-borne segmented negative-sense RNA viruses, many of which are capable of causing devastating disease in humans. In contrast, hantavirus infection of rodent hosts results in a persistent and inapparent infection through their ability to evade immune detection and inhibit apoptosis. In this study, we used Tula hantavirus (TULV) to investigate the interplay between viral and host apoptotic responses during early, peak and persistent phases of virus infection in cell culture. Examination of early-phase TULV infection revealed that infected cells were refractory to apoptosis, as evidenced by the complete lack of cleaved caspase-3 (casp-3C) staining, whereas in non-infected bystander cells casp-3C was highly abundant. Interestingly, at later time points, casp-3C was abundant in infected cells, but the cells remained viable and able to continue shedding infectious virus, and together these observations were suggestive of a TULV-associated apoptotic block. To investigate this block, we viewed TULV-infected cells using laser scanning confocal and wide-field deconvolution microscopy, which revealed that TULV nucleocapsid protein (NP) colocalized with, and sequestered, casp-3C within cytoplasmic ultrastructures. Consistent with casp-3C colocalization, we showed for the first time that TULV NP was cleaved in cells and that TULV NP and casp-3C could be co-immunoprecipitated, suggesting that this interaction was stable and thus unlikely to be solely confined to NP binding as a substrate to the casp-3C active site. To account for these findings, we propose a novel mechanism by which TULV NP inhibits apoptosis by spatially sequestering casp-3C from its downstream apoptotic targets within the cytosol.

Molecular evidence of Tula virus in Microtus obscurus in the region of Yili, Xinjiang, China

BackgroundHantaviruses are important zoonotic pathogens, and they pose a profound risk to public health. So far, there has been no evidence showing that Tula virus (TULV), one species of hantavirus, is endemic in China. In this study, we captured rodents and found that the Tula virus had infected voles in Yili region, Xinjiang, China.MethodsRodents were captured by flooding their burrows in mountain pasture areas in Narati, Xinyuan County, Xinjiang, China. Hantavirus L gene fragments were amplified by nest RT-PCR using genus-specific primers. Positive samples were further identified by sequencing of RT-PCR products of S gene fragment for species identification. To identify the species of captured small mammals, the rodents’ cytochrome b (Cytb) was amplified by PCR and sequenced. Phylogenetic analysis was used to show the clustering and evolution relationship of the viral nucleic acids.ResultsHere, 31 out of 198 voles captured (16%) were infected with TULV. Host sequencing analysis showed these voles were Microtus obscurus (M. obscurs). Alignment and phylogenetic analysis of the exon region (1191 bp) of the hantavirus S gene confirmed that all of the detected amplicons were TULV, which was similar to one strain of TULV identified in Kazakhstan.ConclusionThis is the first identification of Tula virus in China, and we found that M. obscurus acts as a natural reservoir for carrying the virus. Although the infection rate in the local human population remains unknown, the high prevalence of TULV in the small mammals in the region constitutes a risk that this putative pathogen may spread to the local population.

The mystery of the phylogeographic structural pattern in rodent-borne hantaviruses

Hantaviruses (order Bunyavirales, family Hantaviridae) are important zoonotic pathogens. Because of the great diversity of their reservoir hosts, hantaviruses are excellent models to evaluate the dynamics of virus-host co-evolution. To understand the mechanisms behind the evolutionary history of hantaviruses through virus-reservoir interactions, it is important to know how the radiation and diversity of hantaviruses occurred. In this paper, we evaluate the pattern of hantavirus diversification based on a complete S segment representing major groups of hantaviruses found in the Americas. Phylogenetic analyses revealed a high degree of phylogeographic structure and a surprising pattern of geographical distribution of New World hantaviruses. The available data suggest that hantaviruses related to the Arvicolinae rodent subfamily in North America probably emerged and initially adapted from a shared common ancestor of the Tula virus. The first clade of hantaviruses associated with Neotominae occupied a stem lineage, especially those that emerged in Central America or Mexico. Hantaviruses from Central America and Mexico found in Neotominae rodents spread northward and probably gave rise to the first phylogroup of hantaviruses associated with Sigmodontinae in North America. Two preferential host-switching transmissions in hantaviruses apparently gave rise to two different paraphyletic group in Neotominae and Sigmodontinae. Our study supports a probable epicenter of diversification in Central America and/or Mexico for hantaviruses related to both the Neotominae and Sigmodontinae subfamilies.

Orthohantaviruses belonging to three phylogroups all inhibit apoptosis in infected target cells

Orthohantaviruses, previously known as hantaviruses, are zoonotic viruses that can cause hantavirus pulmonary syndrome (HPS) and hemorrhagic fever with renal syndrome (HFRS) in humans. The HPS-causing Andes virus (ANDV) and the HFRS-causing Hantaan virus (HTNV) have anti-apoptotic effects. To investigate if this represents a general feature of orthohantaviruses, we analysed the capacity of six different orthohantaviruses – belonging to three distinct phylogroups and representing both pathogenic and non-pathogenic viruses – to inhibit apoptosis in infected cells. Primary human endothelial cells were infected with ANDV, HTNV, the HFRS-causing Puumala virus (PUUV) and Seoul virus, as well as the putative non-pathogenic Prospect Hill virus and Tula virus. Infected cells were then exposed to the apoptosis-inducing chemical staurosporine or to activated human NK cells exhibiting a high cytotoxic potential. Strikingly, all orthohantaviruses inhibited apoptosis in both settings. Moreover, we show that the nucleocapsid (N) protein from all examined orthohantaviruses are potential targets for caspase-3 and granzyme B. Recombinant N protein from ANDV, PUUV and the HFRS-causing Dobrava virus strongly inhibited granzyme B activity and also, to certain extent, caspase-3 activity. Taken together, this study demonstrates that six different orthohantaviruses inhibit apoptosis, suggesting this to be a general feature of orthohantaviruses likely serving as a mechanism of viral immune evasion.

Parallel Survey of Two Widespread Renal Syndrome-Causing Zoonoses: Leptospira spp. and Hantavirus in Urban Environment, Hungary.

Rodents are important reservoir hosts for several zoonotic pathogens that cause significant morbidity and mortality in humans. Among others, leptospirosis is one of the most widespread zoonotic diseases worldwide and has the similar clinical manifestation with hantavirus infection in humans. Despite the fact that both pathogens have great epidemiological significance in Europe, no epizootiological data exist for urbanized areas so far. Therefore, this study was conducted to investigate the occurrence and prevalence of Leptospira spp. and hantaviruses in small wild rodents living in close proximity to humans. Altogether, 338 small rodents representing five different species (Apodemus agrarius, A. flavicollis, A. sylvaticus, Microtus arvalis, and Myodes glareolus) were captured in the city of Pécs (Hungary) and screened for pathogens by different types of PCR methods (TaqMan-based real-time PCR/PCR, RT-PCR/PCR). A total of 18.3% of the rodents were positive for Leptospira kirschneri, L. interrogans, and L. borgpetersenii. Nucleic acid of Tula hantavirus and human pathogen Dobrava-Belgrade orthohantavirus were detected in 8% of tested specimens. Furthermore, dual infections with both Leptospira spp. and hantaviruses were shown in 2.6% of animals, suggesting that the same rodent host can be infected with several pathogens at the same time, therefore, representing a serious threat to public health. Overall, this study provides important surveillance data on the prevalence of Leptospira spp. and hantaviruses from rodents in urbanized environment for the first time in Hungary and emphasizes the importance of further ecoepidemiological investigations.