Nephropathia Epidemica Caused by Puumala Virus in Bank Voles, Scania, Southern Sweden.

In 2018, a local case of nephropathia epidemica was reported in Scania, southern Sweden, more than 500 km south of the previously known presence of human hantavirus infections in Sweden. Another case emerged in the same area in 2020. To investigate the zoonotic origin of those cases, we trapped rodents in Ballingslöv, Norra Sandby, and Sörby in southern Sweden during 2020‒2021. We found Puumala virus (PUUV) in lung tissues from 9 of 74 Myodes glareolus bank voles by screening tissues using a hantavirus pan-large segment reverse transcription PCR. Genetic analysis revealed that the PUUV strains were distinct from those found in northern Sweden and Denmark and belonged to the Finnish PUUV lineage. Our findings suggest an introduction of PUUV from Finland or Karelia, causing the human PUUV infections in Scania. This discovery emphasizes the need to understand the evolution, cross-species transmission, and disease outcomes of this newly found PUUV variant.

The impact of wildlife and environmental factors on hantavirus infection in the host and its translation into human risk.

Identifying factors that drive infection dynamics in reservoir host populations is essential in understanding human risk from wildlife-originated zoonoses. We studied zoonotic Puumala orthohantavirus (PUUV) in the host, the bank vole (Myodes glareolus), populations in relation to the host population, rodent and predator community and environment-related factors and whether these processes are translated into human infection incidence. We used 5-year rodent trapping and bank vole PUUV serology data collected from 30 sites located in 24 municipalities in Finland. We found that PUUV seroprevalence in the host was negatively associated with the abundance of red foxes, but this process did not translate into human disease incidence, which showed no association with PUUV seroprevalence. The abundance of weasels, the proportion of juvenile bank voles in the host populations and rodent species diversity were negatively associated with the abundance index of PUUV positive bank voles, which, in turn, showed a positive association with human disease incidence. Our results suggest certain predators, a high proportion of young bank vole individuals, and a diverse rodent community, may reduce PUUV risk for humans through their negative impacts on the abundance of infected bank voles.

Puumala Orthohantavirus Reassortant Genome Variants Likely Emerging in the Watershed Forests.

Hemorrhagic fever with renal syndrome (HFRS) remains a prevalent zoonosis in the Republic of Tatarstan (RT), Russian Federation. Puumala orthohantavirus (PUUV), carried by bank voles (Myodes glareolus), is the principal zoonotic pathogen of HFRS in the RT. In this study, we sought to demonstrate the similarity of the PUUV genetic sequences detected in HFRS case patients and bank vole samples previously collected in some areas of the RT. Furthermore, we intended to identify the reassortant PUUV genomes and locate a potential site for their emergence. During 2019 outbreaks, the PUUV genome sequences of the S and M segments from 42 HFRS cases were analysed and compared with the corresponding sequences from bank voles previously trapped in the RT. Most of the PUUV strains from HFRS patients turned out to be closely related to those isolated from bank voles captured near the site of the human infection. We also found possible reassortant PUUV genomes in five patients while they were absent in bank voles. The location of the corresponding HFRS infection sites suggests that reassortant PUUV genomes could emerge in the bank voles that inhabit the forests on the watershed between the Kazanka River and Myosha River. These findings could facilitate the search for the naturally occurring reassortants of PUUV in bank vole populations.

Hantavirus-Leptospira coinfections in small mammals from central Germany.

European orthohantaviruses (Puumala orthohantavirus (PUUV); Dobrava-Belgrade orthohantavirus (DOBV), genotype Kurkino; Tula orthohantavirus (TULV)), and Leptospira spp. are small mammal-associated zoonotic pathogens that cause diseases with potentially similar symptoms in humans. We investigated the frequency of Leptospira spp. and hantavirus single and double infections in small mammals from 22 sites in Thuringia, central Germany, during 2017. TULV infections were detected at 18 of 22 sites (mean prevalence 13.8%, 93/674). PUUV infections were detected at four of 22 sites (mean prevalence 1.5%, 7/471), and respective PUUV sequences formed a novel phylogenetic clade, but DOBV infections were not detected at all. Leptospira infections were detected at 21 of 22 sites with the highest overall prevalence in field voles (Microtus agrestis) with 54.5% (6/11) and common voles (Microtus arvalis) with 30.3% (205/676). Leptospira-hantavirus coinfections were found in 6.6% (44/671) of common voles but only in two of 395 bank voles. TULV and Leptospira coinfection probability in common voles was driven by individual (age) and population-level factors. Coinfections seemed to be particularly associated with sites where Leptospira spp. prevalence exceeded 35%. Future investigations should evaluate public health consequences of this strong spatial clustering of coinfections.

Isolation and characterization of new Puumala orthohantavirus strains from Germany.

Orthohantaviruses are re-emerging rodent-borne pathogens distributed all over the world. Here, we report the isolation of a Puumala orthohantavirus (PUUV) strain from bank voles caught in a highly endemic region around the city Osnabrück, north-west Germany. Coding and non-coding sequences of all three segments (S, M, and L) were determined from original lung tissue, after isolation and after additional passaging in VeroE6 cells and a bank vole-derived kidney cell line. Different single amino acid substitutions were observed in the RNA-dependent RNA polymerase (RdRP) of the two stable PUUV isolates. The PUUV strain from VeroE6 cells showed a lower titer when propagated on bank vole cells compared to VeroE6 cells. Additionally, glycoprotein precursor (GPC)-derived virus-like particles of a German PUUV sequence allowed the generation of monoclonal antibodies that allowed the reliable detection of the isolated PUUV strain in the immunofluorescence assay. In conclusion, this is the first isolation of a PUUV strain from Central Europe and the generation of glycoprotein-specific monoclonal antibodies for this PUUV isolate. The obtained virus isolate and GPC-specific antibodies are instrumental tools for future reservoir host studies.

Phylogenetic analysis of Puumala virus strains from Central Europe highlights the need for a full-genome perspective on hantavirus evolution.

Puumala virus (PUUV), carried by bank voles (Myodes glareolus), is the medically most important hantavirus in Central and Western Europe. In this study, a total of 523 bank voles (408 from Germany, 72 from Slovakia, and 43 from Czech Republic) collected between the years 2007-2012 were analyzed for the presence of hantavirus RNA. Partial PUUV genome segment sequences were obtained from 51 voles. Phylogenetic analyses of all three genome segments showed that the newfound strains cluster with other Central and Western European PUUV strains. The new sequences from Sumava (Bohemian Forest), Czech Republic, are most closely related to the strains from the neighboring Bavarian Forest, a known hantavirus disease outbreak region. Interestingly, the Slovak strains clustered with the sequences from Bohemian and Bavarian Forests only in the M but not S segment analyses. This well-supported topological incongruence suggests a segment reassortment event or, as we analyzed only partial sequences, homologous recombination. Our data highlight the necessity of sequencing all three hantavirus genome segments and of a broader bank vole screening not only in recognized endemic foci but also in regions with no reported human hantavirus disease cases.

Small mammal community composition impacts bank vole (Clethrionomys glareolus) population dynamics and associated seroprevalence of Puumala orthohantavirus.

Rodents are important reservoirs for zoonotic pathogens that cause diseases in humans. Biodiversity is hypothesized to be closely related to pathogen prevalence through multiple direct and indirect pathways. For example, the presence of non-host species can reduce contact rates of the main reservoir host and thus reduce the risk of transmission ("dilution effect"). In addition, an overlap in ecological niches between two species could lead to increased interspecific competition, potentially limiting host densities and reducing density-dependent pathogen transmission processes. In this study, we investigated the relative impact of population-level regulation of direct and indirect drivers of the prevalence of Puumala orthohantavirus (PUUV) in bank voles (Clethrionomys glareolus) during years with high abundance. We compiled data on small mammal community composition from four regions in Germany between 2010 and 2013. Structural equation modeling revealed a strong seasonality in PUUV control mechanisms in bank voles. The abundance of shrews tended to have a negative relationship with host abundance, and host abundance positively influenced PUUV seroprevalence, while at the same time increasing the abundance of competing non-hosts like the wood mouse (Apodemus sylvaticus) and the yellow-necked field mouse (Apodemus flavicollis) were associated with reduced PUUV seroprevalence in the host. These results indicate that for PUUV in bank voles, dilution is associated with increased interspecific competition. Anthropogenic pressures leading to the decline of Apodemus spp. in a specific habitat could lead to the amplification of mechanisms promoting PUUV transmission within the host populations.

Phylogenetic analysis of variants of the Puumala virus (Hantaviridae: Orthohantavirus) circulating in the Saratov region.

The objective is to determine the complete nucleotide sequence and conduct a phylogenetic analysis of genome variants of the Puumala virus isolated in the Saratov region. MATERIALS AND METHODS: The samples for the study were field material collected in the Gagarinsky (formerly Saratovsky), Engelssky, Novoburassky and Khvalynsky districts of the Saratov region in the period from 2019 to 2022. To specifically enrich the Puumala virus genome in the samples, were used PCR and developed a specific primer panel. Next, the resulting PCR products were sequenced and the fragments were assembled into one sequence for each segment of the virus genome. To construct phylogenetic trees, the maximum parsimony algorithm was used. RESULTS: Genetic variants of the Puumala virus isolated in the Saratov region have a high degree of genome similarity to each other, which indicates their unity of origin. According to phylogenetic analysis, they all form a separate branch in the cluster formed by hantaviruses from other subjects of the Volga Federal District. The virus variants from the Republics of Udmurtia and Tatarstan, as well as from the Samara and Ulyanovsk regions, are closest to the samples from the Saratov region. CONCLUSION: The data obtained show the presence of a pronounced territorial confinement of strains to certain regions or areas that are the natural biotopes of their carriers. This makes it possible to fairly accurately determine the territory of possible infection of patients and/or the circulation of carriers of these virus variants based on the sequence of individual segments of their genome.

Impact of climate change on vector- and rodent-borne infectious diseases.

Background: Endemic and imported vector- and rodent-borne infectious agents can be linked to high morbidity and mortality. Therefore, vector- and rodent-borne human diseases and the effects of climate change are important public health issues. Methods: For this review, the relevant literature was identified and evaluated according to the thematic aspects and supplemented with an analysis of surveillance data for Germany. Results: Factors such as increasing temperatures, changing precipitation patterns, and human behaviour may influence the epidemiology of vector- and rodent-borne infectious diseases in Germany. Conclusions: The effects of climatic changes on the spread of vector- and rodent-borne infectious diseases need to be further studied in detail and considered in the context of climate adaptation measures.

Genetic features of the Puumala virus (Hantaviridae: Orthohantavirus) identified in the Moscow region.

INTRODUCTION: Puumala virus (family Hantaviridae, genus Orthohantavirus) is distributed in most regions of the European part of Russia. However, information about its genetic variants circulating on the territory of the Central Federal District is extremely scarce. MATERIALS AND METHODS: Rodents' tissue samples were tested after reverse transcription by PCR for the presence of hantaviral RNA. The amplified fragments of the L segment were sequenced by the Sanger method. For two samples, sequences of all three segments were obtained using the NGS method. Phylogenetic trees were built in the MEGA-X software. RESULTS: Puumala virus was found in six samples. Based on the phylogenetic analysis of sequences of three segments, the obtained genetic variants belong to the sublineage previously designated as W-RUS. CONCLUSION: A genetic variant of the Puumala virus, belonging to the subline W-RUS, circulates on the territory of the Volokolamsk district of Moscow region.

Puumala orthohantavirus circulation in its wild reservoir, the bank vole, during the 2021 outbreak of hemorrhagic fever with renal syndrome in Jura, France.

OBJECTIVE: A large and unprecedented outbreak of an attenuated form of hemorrhagic fever with renal syndrome called nephropathia epidemica (NE) and caused by Puumala virus (PUUV) occurred in 2021 in the southern Jura Mountains (France) leading to numerous hospitalizations. The aim of this study was to investigate the circulation of PUUV in its animal reservoir at the time of this outbreak. METHODS: We conjointly surveyed bank vole relative abundance, small mammal community composition, and PUUV circulation in bank voles (seroprevalence and genetic diversity) in the Jura NE epidemic area, between 2020 and 2022. RESULTS: Trapping results showed a higher relative abundance of bank voles in 2021 compared to 2020 and 2022. Extremely high levels of PUUV seroprevalence in bank voles were found at the time of the human NE epidemic with seropositive animals trapped in almost all trap lines as of spring 2021. Genetic analyses of PUUV (S segment) gathered in 2021 at two sampling sites revealed a strong clustering of these strains within the "Jura" clade. No significant genetic variation was detected compared to what was already known to be circulating in the Jura region. CONCLUSION: These results underline a need for enhanced monitoring of PUUV circulation in host reservoir populations in NE endemic areas. This would enable the relevant actors to better inform and sensitize the public on this zoonotic risk, and to implement prevention strategies in collaboration with physicians.

Tropism of Puumala orthohantavirus and Endoparasite Coinfection in the Bank Vole Reservoir.

In Europe, most cases of human hantavirus disease are caused by Puumala orthohantavirus (PUUV) transmitted by bank voles (Clethrionomys glareolus, syn. Myodes glareolus), in which PUUV causes inconspicuous infection. Little is known about tropism and endoparasite coinfections in PUUV-infected reservoir and spillover-infected rodents. Here, we characterized PUUV tropism, pathological changes and endoparasite coinfections. The voles and some non-reservoir rodents were examined histologically, immunohistochemically, by in situ hybridization, indirect IgG enzyme-linked immunosorbent assay and reverse transcription-polymerase chain reaction. PUUV RNA and anti-PUUV antibodies were detected simultaneously in a large proportion of the bank voles, indicating persistent infection. Although PUUV RNA was not detected in non-reservoir rodents, the detection of PUUV-reactive antibodies suggests virus contact. No specific gross and histological findings were detected in the infected bank voles. A broad organ tropism of PUUV was observed: kidney and stomach were most frequently infected. Remarkably, PUUV was detected in cells lacking the typical secretory capacity, which may contribute to the maintenance of virus persistence. PUUV-infected wild bank voles were found to be frequently coinfected with Hepatozoon spp. and Sarcocystis (Frenkelia) spp., possibly causing immune modulation that may influence susceptibility to PUUV infection or vice versa. The results are a prerequisite for a deeper understanding of virus-host interactions in natural hantavirus reservoirs.

Hantavirus Pulmonary Syndrome Caused by Puumala Orthohantavirus-A Case Report and Literature Review.

In this article, we report on a rare case of acute respiratory distress syndrome (ARDS) caused by the Puumala orthohantavirus (PUUV), which is typically associated with hemorrhagic fever with renal syndrome (HFRS). This is the first documented case of PUUV-associated ARDS in Southeast Europe. The diagnosis was confirmed by serum RT-PCR and serology and corroborated by phylogenetic analysis and chemokine profiling. The patient was a 23-year-old male from Zagreb, Croatia, who had recently traveled throughout Europe. He presented with fever, headache, abdominal pain, and sudden onset of ARDS. Treatment involved high-flow nasal cannula oxygen therapy and glucocorticoids, which resulted in a full recovery. A systematic literature review identified 10 cases of hantavirus pulmonary syndrome (HPS) caused by PUUV in various European countries and Turkey between 2002 and 2023. The median age of patients was 53 years (range 24-73), and six of the patients were male. Most patients were treated in intensive care units, but none received antiviral therapy targeting PUUV. Eight patients survived hospitalization. The presented case highlights the importance of considering HPS in the differential diagnosis of ARDS, even in areas where HFRS is the dominant form of hantavirus infection.

Climate change accelerates winter transmission of a zoonotic pathogen.

Many zoonotic diseases are weather sensitive, raising concern how their distribution and outbreaks will be affected by climate change. At northern high latitudes, the effect of global warming on especially winter conditions is strong. By using long term monitoring data (1980-1986 and 2003-2013) from Northern Europe on temperature, precipitation, an endemic zoonotic pathogen (Puumala orthohantavirus, PUUV) and its reservoir host (the bank vole, Myodes glareolus), we show that early winters have become increasingly wet, with a knock-on effect on pathogen transmission in its reservoir host population. Further, our study is the first to show a climate change effect on an endemic northern zoonosis, that is not induced by increased host abundance or distribution, demonstrating that climate change can also alter transmission intensity within host populations. Our results suggest that rainy early winters accelerate PUUV transmission in bank voles in winter, likely increasing the human zoonotic risk in the North.

Analysis of Puumala orthohantavirus Genome Variants Identified in the Territories of Volga Federal District.

Hemorrhagic fever with renal syndrome (HFRS) is a zoonotic disease commonly diagnosed in the Volga Federal District (VFD). HFRS is caused by Puumala orthohantavirus (PUUV), and this virus is usually detected in bank voles as its natural host (Myodes glareolus). The PUUV genome is composed of the single-stranded, negative-sense RNA containing three segments. The goal of the current study is to identify genome variants of PUUV strains circulating in bank voles captured in the Udmurt Republic (UR) and Ulyanovsk region (ULR). The comparative and phylogenetic analysis of PUUV strains revealed that strains from Varaksino site UR are closely related to strains previously identified in the Pre-Kama area of the Republic of Tatarstan (RT), whilst strains from Kurlan and Mullovka sites ULR are similar to strains from the Trans-Kama area of the RT. It was also found that Barysh ULR strains form a separate distinct group phylogenetically equidistant from Varaksino and Kurlan−Mullovka groups. The identified groups of strains can be considered as separate sub-lineages in the PUUV Russian genetic lineage. In addition, the genomes of the strains from the UR, most likely, were formed as a result of reassortment.

Clinical and genomic characterisation of a fatal Puumala orthohantavirus case with low levels of neutralising antibodies.

BACKGROUND: Orthohantaviruses are rodent-borne emerging viruses that cause haemorrhagic fever with renal syndrome (HFRS) in Eurasia and hantavirus pulmonary syndrome in America. Transmission between humans have been reported and the case-fatality rate ranges from 0.4% to 40% depending on virus strain. There is no specific and efficient treatment for patients with severe HFRS. Here, we characterised a fatal case of HFRS and sequenced the causing Puumala orthohantavirus (PUUV). METHODS: PUUV RNA and virus specific neutralising antibodies were quantified in plasma samples from the fatal case and other patients with non-fatal PUUV infection. To investigate if the causing PUUV strain was different from previously known strains, Sanger sequencing was performed directly from the patient's plasma. Biopsies obtained from autopsy were stained for immunohistochemistry. RESULTS: The patient had approximately tenfold lower levels of PUUV neutralising antibodies and twice higher viral load than was normally seen for patients with less severe PUUV infection. We could demonstrate unique mutations in the S and M segments of the virus that could have had an impact on the severity of infection. Due to the severe course of infection, the patient was treated with the bradykinin receptor inhibitor icatibant to reduce bradykinin-mediated vessel permeability and maintain vascular circulation. CONCLUSIONS: Our data suggest that bradykinin receptor inhibitor may not be highly efficient to treat patients that are at an advanced stage of HFRS. Low neutralising antibodies and high viral load at admission to the hospital were associated with the fatal outcome and may be useful for future predictions of disease outcome.

Cluster of human Puumala orthohantavirus infections due to indoor exposure?-An interdisciplinary outbreak investigation.

Puumala orthohantavirus (PUUV) is the most important hantavirus species in Europe, causing the majority of human hantavirus disease cases. In central and western Europe, the occurrence of human infections is mainly driven by bank vole population dynamics influenced by beech mast. In Germany, hantavirus epidemic years are observed in 2- to 5-year intervals. Many of the human infections are recorded in summer and early autumn, coinciding with peaks in bank vole populations. Here, we describe a molecular epidemiological investigation in a small company with eight employees of whom five contracted hantavirus infections in late 2017. Standardized interviews with employees were conducted to assess the circumstances under which the disease cluster occurred, how the employees were exposed and which counteractive measures were taken. Initially, two employees were admitted to hospital and serologically diagnosed with hantavirus infection. Subsequently, further investigations were conducted. By means of a self-administered questionnaire, three additional symptomatic cases could be identified. The hospital patients' sera were investigated and revealed in one patient a partial PUUV L segment sequence, which was identical to PUUV sequences from several bank voles collected in close proximity to company buildings. This investigation highlights the importance of a One Health approach that combines efforts from human and veterinary medicine, ecology and public health to reveal the origin of hantavirus disease clusters.

Cocirculation of Leptospira spp. and multiple orthohantaviruses in rodents, Lithuania, Northern Europe.

In Europe, zoonotic Leptospira spp. and orthohantaviruses are mainly associated with specific rodent hosts. These pathogens cause febrile human diseases with similar symptoms and disease progression. In Lithuania, the presence of Dobrava-Belgrade orthohantavirus (DOBV), Tula orthohantavirus (TULV) and Leptospira spp. in rodent reservoirs is still unknown, and Puumala orthohantavirus (PUUV) was detected in bank voles (Clethrionomys glareolus) at only one site. Therefore, we collected and screened 1617 rodents and insectivores from Lithuania for zoonotic (re-)emerging Leptospira and orthohantaviruses. We detected Leptospira DNA in six rodent species, namely striped field mouse (Apodemus agrarius), yellow-necked mouse (Apodemus flavicollis), bank vole, common vole (Microtus arvalis), field vole (Microtus agrestis) and root vole (Microtus oeconomus). Leptospira DNA was detected with an overall mean prevalence of 4.4% (range 3.7%-7.9% per rodent species). We detected DOBV RNA in 5.6% of the striped field mice, PUUV RNA in 1% of bank voles and TULV RNA in 4.6% of common voles, but no Leptospira DNA in shrews and no hantavirus-Leptospira coinfections in rodents. Based on the complete coding sequences of the three genome segments, two distant DOBV phylogenetic lineages in striped field mice, one PUUV strain in bank voles and two TULV strains in common voles were identified. The Leptospira prevalence for striped field mice and yellow-necked mice indicated a significant negative effect of the distance to water points. The detection of (re-)emerging human pathogenic Leptospira and three orthohantaviruses in rodent reservoirs in Lithuania calls for increased awareness of public health institutions and allows the improvement of molecular diagnostics for pathogen identification.

Seroprevalence and incidence of Puumala orthohantavirus in its bank vole (Myodes glareolus) host population in northeastern France: Between-site and seasonal variability.

Given the difficulty of measuring pathogen transmission in wildlife, epidemiological studies frequently rely on cross-sectional seroprevalence. However, seropositivity indicates only exposure to a pathogen at an unknown time. By allowing to obtain repeated test results from individuals sampled multiple times over an extended period, longitudinal data help reduce this uncertainty. We used capture-mark-recapture data on bank vole (Myodes glareolus) individuals collected at four sites over ten years in northeastern France to investigate the impact of environmental variables on seroprevalence and incidence of Puumala orthohantavirus (PUUV). PUUV causes a chronic infection without apparent symptoms, that may however impair survival of its rodent host in the wild. Viral transmission between rodents may occur through direct contact or via the environment. Principal component analysis was used to deal with multicollinearity among environmental variables. Incidence and seroprevalence were investigated with either generalized estimating equations or Poisson regression models depending on the number of observations for each season. In spring, only the factor site was found to be significant for seroprevalence, while a principal component including meteorological conditions of the previous winter and the normalized difference vegetation index (NDVI) of both the previous winter and spring had a significant effect on incidence. In autumn, only the factor site was significant for incidence, while two principal components, including either the meteorological conditions of the autumn and previous spring or NDVI of the autumn significantly affected seroprevalence. We discuss these results in light of the particular demography of small mammals. We encourage other researchers to investigate the relationships between demographic parameters of wild host populations and the environment, by using both incidence and seroprevalence.

Genetic Diversity of Puumala orthohantavirus in Rodents and Human Patients in Austria, 2012-2019.

Puumala orthohantavirus (PUUV) has a wide distribution throughout Europe. Distinctive temporal patterns of spillover into the human population are related to population dynamics of the reservoir host, the bank vole (Clethrionomys glareolus). As the rodent host is tied to specific habitats with small individual ranges, PUUV genetic diversity is also highly correlated with geographic distance. Using sequenced portions of viral S and M segments, we determined whether geographic clusters were supported. Human cases of PUUV infections are concentrated in southeastern Austria. We detected four distinct genotypes: two genotypes of the Alpe-Adria (ALAD) lineage typically associated with southeast Europe, and two sublineages of the Central Europe (CE) lineage. One cluster of CE genotypes represents a phylogenetically distinct sublineage compared to previously reported CE clades, and extends the boundary of the CE lineage further south than previously reported.