Hybrid capture-based next-generation sequencing of new and old world Orthohantavirus strains and wild-type Puumala isolates from humans and bank voles.

Orthohantaviruses, transmitted primarily by rodents, cause hemorrhagic fever with renal syndrome (HFRS) in Eurasia and hantavirus pulmonary syndrome in the Americas. These viruses, with documented human-to-human transmission, exhibit a wide case-fatality rate, 0.5-40 %, depending on the virus species, and no vaccine or effective treatment for severe Orthohantavirus infections exists. In Europe, the Puumala virus (PUUV), carried by the bank vole Myodes glareolus, causes a milder form of HFRS. Despite the reliance on serology and PCR for diagnosis, the three genomic segments of Swedish wild-type PUUV have yet to be completely sequenced. We have developed a targeted hybrid-capture method aimed at comprehensive genomic sequencing of wild-type PUUV isolates and the identification of other Orthohantaviruses. Our custom-designed panel includes >11,200 probes covering the entire Orthohantavirus genus. Using this panel, we sequenced complete viral genomes from bank vole lung tissue, human plasma samples, and cell-cultured reference strains. Analysis revealed that Swedish PUUV isolates belong to the Northern Scandinavian lineage, with nucleotide diversity ranging from 2.8 % to 3.7 % among them. Notably, no significant genotypic differences were observed between the viral sequences from reservoirs and human cases except in the nonstructural protein. Despite the high endemicity of PUUV in Northern Sweden, these are the first complete Swedish wild-type PUUV genomes and substantially increase our understanding of PUUV evolution and epidemiology. The panel's sensitivity enables genomic sequencing of human samples with viral RNA levels reflecting the natural progression of infection and underscores our panel's diagnostic value, and could help to uncover novel Orthohantavirus transmission routes.

Spatial-temporal drivers and incidence heterogeneity of hemorrhagic fever with renal syndrome transmission in Shandong Province, China, 2016-2022.

BACKGROUND: Hemorrhagic fever with renal syndrome (HFRS) signals a recurring risk in Eurasia in recent years owing to its continued rise in case notifications and the extension of geographical distribution. This study was undertaken to investigate the spatiotemporal drivers and incidence heterogeneity of HFRS transmission in Shandong Province. METHODS: The epidemiological data for HFRS, meteorological data and socioeconomic data were obtained from China Information System for Disease Control and Prevention, China Meteorological Data Sharing Service System, and Shandong Statistical Yearbook, respectively. The spatial-temporal multicomponent model was employed to analyze the values of spatial-temporal components and the heterogeneity of HFRS transmission across distinct regions. RESULTS: The total effect values of the autoregressive, epidemic, and endemic components were 0.451, 0.187, and 0.033, respectively, exhibiting significant heterogeneity across various cities. This suggested a pivotal role of the autoregressive component in propelling HFRS transmission in Shandong Province. The epidemic component of Qingdao, Weifang, Yantai, Weihai, and Jining declined sharply at the onset of 2020. The random effect identified distinct incidence levels associated with Qingdao and Weifang, signifying regional variations in HFRS occurrence. CONCLUSIONS: The autoregressive component emerged as a significant driver in the transmission of HFRS in Shandong Province. Targeted preventive measures should be strategically implemented across various regions, taking into account the predominant component influencing the epidemic.

Viral shedding and viraemia of Andes virus during acute hantavirus infection: a prospective study.

BACKGROUND: Andes virus (ANDV) is a zoonotic Orthohantavirus leading to hantavirus cardiopulmonary syndrome. Although most transmissions occur through environmental exposure to rodent faeces and urine, rare person-to-person transmission has been documented, mainly for close contacts. This study investigates the presence and infectivity of ANDV in body fluids from confirmed cases and the duration of viraemia. METHODS: In this prospective study, 131 participants with confirmed ANDV infection were enrolled in Chile in a prospective study between 2008 and 2022. Clinical samples (buffy coat, plasma, gingival crevicular fluid [GCF], saliva, nasopharyngeal swabs [NPS], and urine) were collected weekly for 3 weeks together with clinical and epidemiological data. Samples were categorised as acute or convalescent (up to and after 16 days following onset of symptoms). Infectivity of positive fluids was assessed after the culture of samples on Vero E6 cells and use of flow cytometry assays to determine the production of ANDV nucleoprotein. FINDINGS: ANDV RNA was detected in 100% of buffy coats during acute phase, declining to 95% by day 17, and to 93% between days 23-29. ANDV RNA in GCF and saliva decreased from 30% and 12%, respectively, during the acute phase, to 12% and 11% during the convalescent phase. Successful infectivity assays of RT-qPCR-positive fluids, including GCF, saliva, NPS, and urine, were observed in 18 (42%) of 43 samples obtained during the acute phase of infection. After re-culture, the capacity to infect Vero E6 cells was maintained in 16 (89%) of 18 samples. Severity was associated with the presence of ANDV RNA in one or more fluids besides blood (odds ratio 2·58 [95% CI 1·42-5·18]). INTERPRETATION: ANDV infection is a systemic and viraemic infection, that affects various organs. The presence of infectious particles in body fluids contributes to our understanding of potential mechanisms for person-to-person transmission, supporting the development of preventive strategies. Detection of ANDV RNA in additional fluids at hospital admission is a predictor of disease severity. FUNDING: None. TRANSLATION: For the Spanish translation of the abstract see Supplementary Materials section.

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.

Case of Human Orthohantavirus Infection, Michigan, USA, 2021.

Orthohantaviruses cause hantavirus cardiopulmonary syndrome; most cases occur in the southwest region of the United States. We discuss a clinical case of orthohantavirus infection in a 65-year-old woman in Michigan and the phylogeographic link of partial viral fragments from the patient and rodents captured near the presumed site of infection.

Visual analysis of global hemorrhagic fever with renal syndrome research from 1980 to 2022: Based on CiteSpace and VOSviewer.

OBJECTIVE: The development and current state of hemorrhagic fever with renal syndrome (HFRS) over the past 40 years are analyzed in this study, along with explored and discovered the hotspots and frontiers in the field, which serve as the foundation for future investigation. METHODS: CiteSpace and VOSviewer analysis software were used to visually analyze the literature data on HFRS from 1980 to 2022, including the annual number of publications, countries and research institutions, authors, co-cited literature and keywords. RESULTS: The number of pertinent papers published in the field of HFRS displayed an overall upward trend from 1980 to 2022. The United States, China, Germany, Sweden, and France are the top 5 countries in terms of publishing volume, with high intermediate centrality mainly concentrated in Europe and the United States. The top 10 co-occurring keywords were hemorrhagic fever, renal syndrome, infection, virus, epidemic, nephropathia epidemical, disease, hantavirus, outbreak, and transmission. According to keyword cluster analysis, there were 4 main research fields. In the HFRS-related study, there were mainly 21 notable keywords and "Korean hemorrhagic fever" had the highest hemorrhagic value (28.87). CONCLUSION: The United States, China, Germany, Sweden and other countries attached great importance to the HFRS-related research. Moreover, the collaboration between authors and institutions in various collaborator clusters should be strengthened. In recent decades, investigations have focused on the study of viral infection and the clinical symptoms and pathophysiology of HFRS. Future research may concentrate on factors affecting host population distribution and density, such as vaccine development and meteorological factors pertaining to virus transmission.

A novel hantavirus identified in bats (Carollia perspicillata) in Brazil.

Bats play an essential role in maintaining ecosystems. Their unique characteristics increase the likelihood of interactions with various species, making them a potential source for the emergence and spread of infectious diseases. Hantaviruses are continuously expanding their range of hosts. This study presents the identification of a partial genome associated with Hantavirus in samples collected from neotropical bats. We conducted a metagenomic study using samples from Carollia perspicillata in Maranhão, Brazil. Tissue fragments were used for RNA extraction and subsequent sequencing. The resulting data was subjected to bioinformatic analysis. A sequence showing an identity of 72.86% with the L gene in the reference genome was obtained. The phylogenetic analysis revealed the study sequence, denoted as Buritiense, clustering within the Mobatvirus clade. The intragroup analysis showed a broader dispersion and were markedly asymmetric. This observation suggests the possibility that Buritiense could potentially represent a new species within the bat-borne hantaviruses, but further analyses are needed to provide additional insights if bats plays a role as reservoirs and the potential for transmission to human populations.

Bivalent VSV Vectors Mediate Rapid and Potent Protection from Andes Virus Challenge in Hamsters.

Orthohantaviruses may cause hemorrhagic fever with renal syndrome or hantavirus cardiopulmonary syndrome. Andes virus (ANDV) is the only orthohantavirus associated with human-human transmission. Therefore, emergency vaccination would be a valuable public health measure to combat ANDV-derived infection clusters. Here, we utilized a promising vesicular stomatitis virus (VSV)-based vaccine to advance the approach for emergency applications. We compared monovalent and bivalent VSV vectors containing the Ebola virus (EBOV), glycoprotein (GP), and ANDV glycoprotein precursor (GPC) for protective efficacy in pre-, peri- and post-exposure immunization by the intraperitoneal and intranasal routes. Inclusion of the EBOV GP was based on its favorable immune cell targeting and the strong innate responses elicited by the VSV-EBOV vaccine. Our data indicates no difference of ANDV GPC expressing VSV vectors in pre-exposure immunization independent of route, but a potential benefit of the bivalent VSVs following peri- and post-exposure intraperitoneal vaccination.

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.

Modulations in the host cell proteome by the hantavirus nucleocapsid protein.

Hantaviruses have evolved a unique translation strategy to boost the translation of viral mRNA in infected cells. Hantavirus nucleocapsid protein (NP) binds to the viral mRNA 5' UTR and the 40S ribosomal subunit via the ribosomal protein S19. NP associated ribosomes are selectively loaded on viral transcripts to boost their translation. Here we demonstrate that NP expression upregulated the steady-state levels of a subset of host cell factors primarily involved in protein processing in the endoplasmic reticulum. Detailed investigation of Valosin-containing protein (VCP/p97), one of the upregulated host factors, in both transfected and virus infected cells revealed that NP with the assistance of VCP mRNA 5' UTR facilitates the translation of downstream VCP ORF. The VCP mRNA contains a 5' UTR of 987 nucleotides harboring six unusual start codons upstream of the correct start codon for VCP which is located at 988th position from the 5' cap. In vitro translation of a GFP reporter transcript harboring the VCP mRNA 5' UTR generated both GFP and a short polypeptide of ~14 KDa by translation initiation from start codon located in the 5' UTR at 542nd position from the 5' cap. The translation initiation from 542nd AUG in the UTR sequence was confirmed in cells using a dual reporter construct expressing mCherry and GFP. The synthesis of 14KDa polypeptide dramatically inhibited the translation of the ORF from the downstream correct start codon at 988th position from the 5' cap. We report that purified NP binds to the VCP mRNA 5' UTR with high affinity and NP binding site is located close to the 542ndAUG. NP binding shuts down the translation of 14KDa polypeptide which then facilitates the translation initiation at the correct AUG codon. Knockdown of VCP generated lower levels of poorly infectious hantavirus particle in the cellular cytoplasm whose egress was dramatically inhibited in human umbilical vein endothelial cells. We demonstrated that VCP binds to the hantavirus glycoprotein Gn before its incorporation into assembled virions and facilitates viral spread to neighboring cells during infection. Our results suggest that ribosome engagement at the 542nd AUG codon in the 5' UTR likely regulates the endogenous steady state levels of VCP in cells. Hantaviruses interrupt this regulatory mechanism to enhance the steady state levels of VCP in virus infected cells. This augmentation facilitates virus replication, supports the transmission of the virus to adjacent cells, and promotes the release of infectious virus particles from the host cell.

Viral Pneumonias.

Viral pneumonia is usually community acquired and caused by influenza, parainfluenza, respiratory syncytial virus, human metapneumovirus, and adenovirus. Many of these infections are airway centric and chest imaging demonstrates bronchiolitis and bronchopneumonia, With the exception of adenovirus infections, the presence of lobar consolidation usually suggests bacterial coinfection. Community-acquired viral pathogens can cause more severe pneumonia in immunocompromised hosts, who are also susceptible to CMV and varicella infection. These latter 2 pathogens are less likely to manifest the striking airway-centric pattern. Airway-centric pattern is distinctly uncommon in Hantavirus pulmonary syndrome, a rare environmentally acquired infection with high mortality.

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.

Hantavirus Disease Cluster Caused by Seoul Virus, Germany.

A cluster of 3 persons in Germany experienced hantavirus disease with renal insufficiency. Reverse transcription PCR-based genotyping revealed infection by Seoul hantavirus transmitted from pet rats. Seoul virus could be responsible for disease clusters in Europe, and infected pet rats should be considered a health threat.

HantaNet: A New MicrobeTrace Application for Hantavirus Classification, Genomic Surveillance, Epidemiology and Outbreak Investigations.

Hantaviruses zoonotically infect humans worldwide with pathogenic consequences and are mainly spread by rodents that shed aerosolized virus particles in urine and feces. Bioinformatics methods for hantavirus diagnostics, genomic surveillance and epidemiology are currently lacking a comprehensive approach for data sharing, integration, visualization, analytics and reporting. With the possibility of hantavirus cases going undetected and spreading over international borders, a significant reporting delay can miss linked transmission events and impedes timely, targeted public health interventions. To overcome these challenges, we built HantaNet, a standalone visualization engine for hantavirus genomes that facilitates viral surveillance and classification for early outbreak detection and response. HantaNet is powered by MicrobeTrace, a browser-based multitool originally developed at the Centers for Disease Control and Prevention (CDC) to visualize HIV clusters and transmission networks. HantaNet integrates coding gene sequences and standardized metadata from hantavirus reference genomes into three separate gene modules for dashboard visualization of phylogenetic trees, viral strain clusters for classification, epidemiological networks and spatiotemporal analysis. We used 85 hantavirus reference datasets from GenBank to validate HantaNet as a classification and enhanced visualization tool, and as a public repository to download standardized sequence data and metadata for building analytic datasets. HantaNet is a model on how to deploy MicrobeTrace-specific tools to advance pathogen surveillance, epidemiology and public health globally.

Presence and Persistence of Andes Virus RNA in Human Semen.

When infecting humans, Andes orthohantavirus (ANDV) may cause a severe disease called hantavirus cardiopulmonary syndrome (HCPS). Following non-specific symptoms, the infection may progress to a syndrome of hemorrhagic fever combined with hyper-acute cardiopulmonary failure. The case fatality rate ranges between 25-40%, depending on the outbreak. In this study, we present the follow-up of a male patient who recovered from HCPS six years ago. We demonstrate that the ANDV genome persists within the reproductive tract for at least 71 months. Genome sequence analysis early and late after infection reveals a low number of mutations (two single nucleotide variants and one deletion), suggesting limited replication activity. We can exclude the integration of the viral genome into the host genome, since the treatment of the specimen with RNAse led to a loss of signal. We demonstrate a long-lasting, strong neutralizing antibody response using pseudovirions expressing the ANDV glycoprotein. Taken together, our results show that ANDV has the potential for sexual transmission.

Evaluating the zoonotic potential of RNA viromes of rodents provides new insight into rodent-borne zoonotic pathogens in Guangdong, China.

Emerging and re-emerging infectious diseases have been on the rise, with a significant proportion being zoonotic. Rodents, as the natural reservoirs of numerous diverse zoonotic viruses, pose a substantial threat to human health. To investigate the diversity of known and unknown viruses harbored by rodents in Guangdong (southern province of China), we conducted a comprehensive analysis of viral genomes through metagenomic sequencing of organs from 194 rodents. Our analysis yielded 2163 viral contigs that were assigned to 25 families known to infect a wide range of hosts, including vertebrates, invertebrates, amoebas, and plants. The viral compositions vary considerably among different organs, but not in rodent species. We also assessed and prioritized zoonotic potential of those detected viruses. Ninety-two viral species that are either known to infect vertebrates and invertebrates or only vertebrates were identified, among which 21 are considered high-risk to humans. The high-risk viruses included members of the Hantavirus, Picobirnaviruses, Astroviruses and Pestivirus. The phylogenetic trees of four zoonotic viruses revealed features of novel viral genomes that seem to fit evolutionarily into a zone of viruses that potentially pose a risk of transmission to humans. Recognizing that zoonotic diseases are a One Health issue, we approached the problem of identifying the zoonotic risk from rodent-transmitted disease in the Guangdong province by performing next-generation sequencing to look for potentially zoonotic viruses in these animals.

Land-cover, land-use and human hantavirus infection risk: a systematic review.

Previous studies suggest that the risk of human infection by hantavirus, a family of rodent-borne viruses, might be affected by different environmental determinants such as land cover, land use and land use change. This study examined the association between land-cover, land-use, land use change, and human hantavirus infection risk. PubMed and Scopus databases were interrogated using terms relative to land use (change) and human hantavirus disease. Screening and selection of the articles were completed by three independent reviewers. Classes of land use assessed by the different studies were categorized into three macro-categories of exposure ('Agriculture', 'Forest Cover', 'Urban Areas') to qualitatively synthesize the direction of the association between exposure variables and hantavirus infection risk in humans. A total of 25 articles were included, with 14 studies (56%) conducted in China, 4 studies (16%) conducted in South America and 7 studies (28%) conducted in Europe. Most of the studies (88%) evaluated land cover or land use, while 3 studies (12%) evaluated land use change, all in relation to hantavirus infection risk. We observed that land cover and land-use categories could affect hantavirus infection incidence. Overall, agricultural land use was positively associated with increased human hantavirus infection risk, particularly in China and Brazil. In Europe, a positive association between forest cover and hantavirus infection incidence was observed. Studies that assessed the relationship between built-up areas and hantavirus infection risk were more variable, with studies reporting positive, negative or no associations.

The prevalence of rodent-borne zoonotic pathogens in the South Gobi desert region of Mongolia.

The alpine ecosystems and communities of central Asia are currently undergoing large-scale ecological and socio-ecological changes likely to affect wildlife-livestock-human disease interactions and zoonosis transmission risk. However, relatively little is known about the prevalence of pathogens in this region. Between 2012 and 2015 we screened 142 rodents in Mongolia's Gobi desert for exposure to important zoonotic and livestock pathogens. Rodent seroprevalence to Leptospira spp. was >1/3 of tested animals, Toxoplasma gondii and Coxiella burnetii approximately 1/8 animals, and the hantaviruses being between 1/20 (Puumala-like hantavirus) and <1/100 (Seoul-like hantavirus). Gerbils trapped inside local dwellings were one of the species seropositive to Puumala-like hantavirus, suggesting a potential zoonotic transmission pathway. Seventeen genera of zoonotic bacteria were also detected in the faeces and ticks collected from these rodents, with one tick testing positive to Yersinia. Our study helps provide baseline patterns of disease prevalence needed to infer potential transmission between source and target populations in this region, and to help shift the focus of epidemiological research towards understanding disease transmission among species and proactive disease mitigation strategies within a broader One Health framework.

Human-to-Human Transmission of Andes Virus Modeled in Syrian Hamsters.

Several occurrences of human-to-human transmission of Andes virus, an etiological agent of hantavirus cardiopulmonary syndrome, are documented. Syrian hamsters consistently model human hantavirus cardiopulmonary syndrome, yet neither transmission nor shedding has been investigated. We demonstrate horizontal virus transmission and show that Andes virus is shed efficiently from both inoculated and contact-infected hamsters.

Zoonotic Hantaviridae with Global Public Health Significance.

Hantaviridae currently encompasses seven genera and 53 species. Multiple hantaviruses such as Hantaan virus, Seoul virus, Dobrava-Belgrade virus, Puumala virus, Andes virus, and Sin Nombre virus are highly pathogenic to humans. They cause hemorrhagic fever with renal syndrome (HFRS) and hantavirus cardiopulmonary syndrome or hantavirus pulmonary syndrome (HCPS/HPS) in many countries. Some hantaviruses infect wild or domestic animals without causing severe symptoms. Rodents, shrews, and bats are reservoirs of various mammalian hantaviruses. Recent years have witnessed significant advancements in the study of hantaviruses including genomics, taxonomy, evolution, replication, transmission, pathogenicity, control, and patient treatment. Additionally, new hantaviruses infecting bats, rodents, shrews, amphibians, and fish have been identified. This review compiles these advancements to aid researchers and the public in better recognizing this zoonotic virus family with global public health significance.