Species-specific responses during Seoul orthohantavirus infection in human and rat lung microvascular endothelial cells.

Seoul orthohantavirus (SEOV) is a rat-borne zoonotic virus that is transmitted via inhalation of aerosolized infectious excreta, and can cause hemorrhagic fever with renal syndrome (HFRS) in humans worldwide. In rats, SEOV predominantly exists as a persistent infection in the absence of overt clinical signs. Lack of disease in rats is attributed to downregulation of pro-inflammatory and upregulation of regulatory host responses. As lung microvascular endothelial cells (LMECs) represent a primary target of infection in both human and rats, infections in these cells provide a unique opportunity to study the central role of LMECs in the dichotomy between pathogenicity in both species. In this study, host responses to SEOV infection in primary human and rat LMECs were directly compared on a transcriptional level. As infection of rat LMECs was more efficient than human LMECs, the majority of anti-viral defense responses were observed earlier in rat LMECs. Most prominently, SEOV-induced processes in both species included responses to cytokine stimulus, negative regulation of innate immune responses, responses to type I and II interferons, regulation of pattern recognition receptor signaling and MHC-I signaling. However, over time, in the rat LMECs, responses shifted from an anti-viral state towards a more immunotolerant state displayed by a PD-L1, B2M-, JAK2-focused interaction network aiding in negative regulation of cytotoxic CD8-positive T cell activation. This suggests a novel mechanism by which species-specific orthohantavirus-induced endothelium and T cell crosstalk may play a crucial role in the development of acute disease in humans and persistence in rodents.

Comparison of Three Detection Methods for Seoul Virus Causing Hemorrhagic Fever with Renal Syndrome.

BACKGROUND: Seoul virus (SEOV) is a significant causative pathogen of hemorrhagic fever with renal syndrome (HFRS). Accurate discrimination of SEOV infection from other viral or bacterial infections holds vital clinical importance. METHODS: Our study utilized quantitative real-time PCR (qRT-PCR), metagenomic next-generation sequencing (mNGS), and immunological assays to identify the pathogen causing HFRS. RESULTS: For the case, mNGS identified SEOV and suspected host or environmental microorganisms at 5 days from symptom onset. qRT-PCR detected SEOV between 5 to 8 days from symptom onset. Anti-hantavirus IgM antibodies reached positive criteria at 7 days and IgG antibodies at 9 days from symptom onset. CONCLUSIONS: qRT-PCR, mNGS, and immunological assays each have merits and drawbacks. Optimal selection depends on laboratory conditions and clinical requirements.

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.

Genetic diversity and molecular evolution of Seoul virus in Hebei province, China.

Seoul virus (SEOV) is a major pathogen which causes hemorrhagic fever with renal syndrome (HFRS), and is present all over the world. However, there are currently few long-term systematic studies of SEOV's phylogenetic and evolutionary mechanisms in epidemic areas. Thus, in this study, we used RT-PCR combined with NGS to obtain the genomes of six SEOV viruses from 1993, as well as 56 Hebei province-specific tissue samples from 1999 to 2022. Phylogenetic analysis showed that the SEOV samples could be divided into seven groups and showed geographic clustering. The geographic region may be the main factor affecting the genetic diversity of SEOV. We also found that SEOV was subject to strong overall purifying selection and positive selection at certain sites during evolution. Recombination events and high nucleotide substitution rates were also shown to accelerate SEOV's evolution. Evolutionary feature of the L segment is more representative of complete genome. Our detailed analysis provides a deeper understanding of the genetic diversity and evolutionary drivers of SEOV within its primary epidemic areas. It will be important to further monitor epidemiological trends and drivers of variation to help increase our understanding of the pathogenicity of SEOV infections.

Orthohantavirus infections in humans and rodents in the Yichun region, China, from 2016 to 2021.

BACKGROUND: Rodents are the predominant natural hosts of orthohantavirus and the source of human infection, hemorrhagic fever with renal syndrome (HFRS) caused by orthohantavirus is a severe public health problem in the Yichun region, Jiangxi Province, China. However, little information is known about the infection of orthohantavirus in humans and rodents, and the genetic characteristics of the epidemic orthohantavirus in the region. METHODS: The clinical data of HFRS cases in 2016-2021 was analyzed. Virus infection in rodents was analyzed by orthohantavirus antigen detection using immunofluorescent assay, and the species of orthohantaviruses in rodents and patients were identified by real-time RT-PCR and gene sequencing. The S and M segments of orthohantaviruses from rodents and patients were recovered and analyzed. RESULTS: A total of 1,573 HFRS cases were reported in the Yichun region from 2016 to 2021, including 11 death cases. HFRS cases peaked twice each year: in winter from November to January and early summer from May to June. Farmers constituted the predominant population suffering from HFRS. The orthohantavirus antigen was identified in five species of rodents: Apodemus agrarius (A. agrarius), Rattus norvegicus (R. norvegicus), Sorex araneus, Rattus losea (R. losea), and Niviventer confucianus (N. confucianus). The real-time RT-PCR test and genetic analysis results showed that Hantaan orthohantavirus (HTNV), Seoul orthohantavirus (SEOV), and Dabieshan orthohantavirus (DBSV) were circulated in the rodents. HTNV, SEOV, and DBSV from the rodents were distantly related to other known orthohantaviruses and belonged to novel genetic lineages. SEOV and HTNV were found in HFRS patients, but 97.8% (90/92) of the infections were caused by HTNV. Winter and early summer peaks were both caused by HTNV. The HTNV sequences recovered from HFRS cases were closely related to those from A. agrarius. CONCLUSIONS: In the Yichun region, the orthohantaviruses transmitted in rodents include HTNV, SEOV, and DBSV, which have obvious genetic characteristics and high genetic diversity. At the same time, this region is an HFRS mixed epidemic area dominated by HTNV, with two peaks every year, which deserves our high attention.

A Development of Rapid Whole-Genome Sequencing of Seoul orthohantavirus Using a Portable One-Step Amplicon-Based High Accuracy Nanopore System.

Whole-genome sequencing provides a robust platform for investigating the epidemiology and transmission of emerging viruses. Oxford Nanopore Technologies allows for real-time viral sequencing on a local laptop system for point-of-care testing. Seoul orthohantavirus (Seoul virus, SEOV), harbored by Rattus norvegicus and R. rattus, causes mild hemorrhagic fever with renal syndrome and poses an important threat to public health worldwide. We evaluated the deployable MinION system to obtain high-fidelity entire-length sequences of SEOV for the genome identification of accurate infectious sources and their genetic diversity. One-step amplicon-based nanopore sequencing was performed from SEOV 80-39 specimens with different viral copy numbers and SEOV-positive wild rats. The KU-ONT-SEOV-consensus module was developed to analyze SEOV genomic sequences generated from the nanopore system. Using amplicon-based nanopore sequencing and the KU-ONT-consensus pipeline, we demonstrated novel molecular diagnostics for acquiring full-length SEOV genome sequences, with sufficient read depth in less than 6 h. The consensus sequence accuracy of the SEOV small, medium, and large genomes showed 99.75-100% (for SEOV 80-39 isolate) and 99.62-99.89% (for SEOV-positive rats) identities. This study provides useful insights into on-site diagnostics based on nanopore technology and the genome epidemiology of orthohantaviruses for a quicker response to hantaviral outbreaks.

Development of a novel minigenome and recombinant VSV expressing Seoul hantavirus glycoprotein-based assays to identify anti-hantavirus therapeutics.

Seoul virus (SEOV) is an emerging global health threat that can cause hemorrhagic fever with renal syndrome (HFRS), which results in case fatality rates of ∼2%. There are no approved treatments for SEOV infections. We developed a cell-based assay system to identify potential antiviral compounds for SEOV and generated additional assays to characterize the mode of action of any promising antivirals. To test if candidate antivirals targeted SEOV glycoprotein-mediated entry, we developed a recombinant reporter vesicular stomatitis virus expressing SEOV glycoproteins. To facilitate the identification of candidate antiviral compounds targeting viral transcription/replication, we successfully generated the first reported minigenome system for SEOV. This SEOV minigenome (SEOV-MG) screening assay will also serve as a prototype assay for discovery of small molecules inhibiting replication of other hantaviruses, including Andes and Sin Nombre viruses. Ours is a proof-of-concept study in which we tested several compounds previously reported to have activity against other negative-strand RNA viruses using our newly developed hantavirus antiviral screening systems. These systems can be used under lower biocontainment conditions than those needed for infectious viruses, and identified several compounds with robust anti-SEOV activity. Our findings have important implications for the development of anti-hantavirus therapeutics.

Pet Rats as the Likely Reservoir for Human Seoul Orthohantavirus Infection.

Seoul orthohantavirus (SEOV) is a rat-associated zoonotic pathogen with an almost worldwide distribution. In 2019, the first autochthonous human case of SEOV-induced hemorrhagic fever with renal syndrome was reported in Germany, and a pet rat was identified as the source of the zoonotic infection. To further investigate the SEOV reservoir, additional rats from the patient and another owner, all of which were purchased from the same vendor, were tested. SEOV RNA and anti-SEOV antibodies were found in both of the patient's rats and in two of the three rats belonging to the other owner. The complete coding sequences of the small (S), medium (M), and large (L) segments obtained from one rat per owner exhibited a high sequence similarity to SEOV strains of breeder rat or human origin from the Netherlands, France, the USA, and Great Britain. Serological screening of 490 rats from breeding facilities and 563 wild rats from Germany (2007-2020) as well as 594 wild rats from the Netherlands (2013-2021) revealed 1 and 6 seropositive individuals, respectively. However, SEOV RNA was not detected in any of these animals. Increased surveillance of pet, breeder, and wild rats is needed to identify the origin of the SEOV strain in Europe and to develop measures to prevent transmission to the human population.

First study of Seoul virus (SEOV) in urban rodents from newly urbanized areas of Gran La Plata, Argentina.

Alterations of ecosystems have deep effects on the distribution of parasites. Big cities of Argentina present structural features that favor the presence of synanthropic species, acting as source of zoonotic diseases, for example in urban rodents: the Norway rat (Rattus norvegicus) and the black rat (R. rattus). One of the important zoonotic pathogens related are the RNA virus Hantavirus, with high prevalence rates in South America. The aim of this study was to explore and identify the presence of Hantavirus in urban rodents from Gran La Plata, Argentina. The presence of anti-hantavirus IgG antibodies was determined by the Enzyme-Linked Immunosorbent Assay. Six samples turned out positive for Seoul virus (SEOV, p = 14.3%). These are the first records of SEOV in urban rodents in Gran La Plata. It represents the first report in R. rattus in Argentina, and in America. This situation underscores the inequality and historical forgetfulness of a portion of society, calling for urgent action to be taken in this regard.

Central nervous system infection with Seoul Orthohantavirus in a child after hematopoietic stem cell transplantation: a case report.

BACKGROUND: Patients with allogeneic hematopoietic stem cell transplantation (allo-HSCT) are prone to complicate viral infection. Central nervous system (CNS) involvement caused by the viruses is rare but with poor prognosis. Hantavirus, which usually cause hemorrhagic fever with renal syndrome (HFRS), and none case has been reported about these infection in allo-HSCT patients. CASE PRESENTATION: In August 2021, a 13-year-old male child developed intermittent fever and refractory hypotension after allo-HSCT. Magnetic resonance imaging of the head revealed abnormal signal foci in the left midbrain cerebral peduncle and bilateral thalamus. His family reported traces of mouse activity in the patient's home kitchen. HFRS was suspected, but with no significant kidney damage. The specific immunoglobulin (Ig) G and M of hantavirus were negative. The metagenomic next-generation sequencing (mNGS) detected Seoul Orthohantavirus (SEOV) sequences directly in cerebrospinal fluid and blood. CONCLUSIONS: Allo-HSCT patients are a high-risk group for infection. Usually the causative agent of infection is difficult to determine, and sometimes the site of infection is concealed. This report highlights the importance of suspecting hantavirus infection in allo-HSCT patients with CNS symptoms despite the absence of renal syndromes. The mNGS is a powerful tool for detecting pathogens. CNS infection with Seoul orthohantavirus in transplant patients is rare but possible as demonstrated in this case. To the best of our knowledge, this is the first reported case employing mNGS to diagnose SEOV caused CNS infection in an allo-HSCT patient.

Seoul Virus Associated with Pet Rats, Scotland, UK, 2019.

We describe a case of hemorrhagic fever with renal syndrome caused by Seoul virus in a woman in Scotland, UK. Whole-genome sequencing showed the virus belonged to a lineage characterized by recent international expansion, probably driven by trade in pet rats.

Genetic Characterization of Seoul Virus in the Seaport of Cotonou, Benin.

Seoul virus is a zoonotic pathogen carried by the brown rat Rattus norvegicus. Information on its circulation in Africa is limited. In this study, the virus was detected in 37.5% of brown rats captured in the Autonomous Port of Cotonou, Benin. Phylogenetic analyses place this virus in Seoul virus lineage 7.

Bayesian Binary Mixture Models as a Flexible Alternative to Cut-Off Analysis of ELISA Results, a Case Study of Seoul Orthohantavirus.

Serological assays, such as the enzyme-linked immunosorbent assay (ELISA), are popular tools for establishing the seroprevalence of various infectious diseases in humans and animals. In the ELISA, the optical density is measured and gives an indication of the antibody level. However, there is variability in optical density values for individuals that have been exposed to the pathogen of interest, as well as individuals that have not been exposed. In general, the distribution of values that can be expected for these two categories partly overlap. Often, a cut-off value is determined to decide which individuals should be considered seropositive or seronegative. However, the classical cut-off approach based on a putative threshold ignores heterogeneity in immune response in the population and is thus not the optimal solution for the analysis of serological data. A binary mixture model does include this heterogeneity, offers measures of uncertainty and the direct estimation of seroprevalence without the need for correction based on sensitivity and specificity. Furthermore, the probability of being seropositive can be estimated for individual samples, and both continuous and categorical covariates (risk-factors) can be included in the analysis. Using ELISA results from rats tested for the Seoul orthohantavirus, we compared the classical cut-off method with a binary mixture model set in a Bayesian framework. We show that it performs similarly or better than cut-off methods, by comparing with real-time quantitative polymerase chain reaction (RT-qPCR) results. We therefore recommend binary mixture models as an analysis tool over classical cut-off methods. An example code is included to facilitate the practical use of binary mixture models in everyday practice.

Seoul Virus in Pet and Feeder Rats in The Netherlands.

Seoul virus (SEOV) is a zoonotic orthohantavirus carried by rats. In humans, SEOV can cause hemorrhagic fever with renal syndrome. Recent human SEOV cases described in the USA, United Kingdom, France and the Netherlands were associated with contact with pet or feeder rats. The prevalence of SEOV in these types of rats is unknown. We collected 175 pet and feeder rats (Rattus norvegicus) from private owners, ratteries and commercial breeders/traders in the Netherlands. Lung tissue of the rats was tested using a SEOV real-time RT-qPCR and heart fluid was tested for the presence of antibodies against SEOV. In all three investigated groups, RT-qPCR-positive rats were found: in 1/29 rats from private owners (3.6%), 2/56 rats from ratteries (3.4%) and 11/90 rats from commercial breeders (12.2%). The seroprevalence was largely similar to the prevalence calculated from RT-qPCR-positive rats. The SEOV sequences found were highly similar to sequences previously found in domesticated rats in Europe. In conclusion, SEOV is spread throughout different populations of domesticated rats.

Hemorrhagic Fever With Renal Syndrome in Vladivostok City, Russia.

Hemorrhagic fever with renal syndrome (HFRS) is a public health problem in Vladivostok city, Russia. From 1997 to 2019, a study of hantaviruses in Norway rats (Rattus norvegicus), a natural reservoir of Seoul virus (SEOV), and in HFRS patients was conducted. We demonstrated the presence of SEOV in the local population of Norway rats and detected SEOV in 10, Amur virus (AMRV) in 4 and Hantaan virus (HTNV) in 1 out of 15 HFRS patients. Genetic analysis based on partial S, M and L segment sequences revealed that the Russian SEOV strains were related most closely to strains from Cambodia and Vietnam. We postulate that the SEOV strains found in the port city of Vladivostok have been spread from South-East Asia as a result of distribution of rats during standard shipping trade activities. Moreover, we suggest that city residents may have acquired AMRV and HTNV infection during visits to rural areas.

Human infection with Seoul orthohantavirus in Korea, 2019.

Of various rodent-borne hantaviruses, Seoul orthohantavirus (SEOV) causes haemorrhagic fever with renal syndrome (HFRS), as does Hantaan orthohantavirus (HTNV). Given global-scale of cases of human infection with SEOV, it is of great clinical importance to distinguish SEOV from other HFRS-causing hantaviruses. In May 2019, a middle-aged patient who had lived in a suburban area of Chungcheong Province, Republic of Korea and enjoyed outdoor activities was transferred to Asan Medical Center in Seoul, Republic of Korea with HFRS; his symptoms included high fever and generalized myalgia. The rapid diagnostic test performed immediately after his transfer detected HTNV-specific antibodies, and the patient was treated accordingly. However, two consecutive IFAs performed at ten-day intervals showed no HTNV-specific immunoglobulin (Ig) G. During continuous supportive care, next-generation sequencing successfully identified viral genomic sequences in the patient's serum, which were SEOV and not HTNV. Phylogenetic analysis grouped the L, M, and S genes of this SEOV strain together with those of rat- or human-isolated Korean strains reported previously. Given global outbreaks and public health threats of zoonotic hantaviruses, a causative pathogen of hantavirus HFRS should be identified correctly at the time of diagnosis and by point-of-care testing.

Rare case of intracranial hemorrhage associated with seoul virus infection diagnosed by metagenomic next-generation sequencing.

BACKGROUND: Seoul virus (SEOV) is a Hantavirus and the causative pathogen of Hemorrhagic Fever with Renal Syndrome (HFRS). Diagnosing SEOV infection is difficult because the clinical presentations are often undistinguishable from other viral or bacterial infections. In addition, diagnostic tools including serological and molecular assays are not readily available in the clinical settings. CASE REPORT: A 57-year-old male presented with fever and a sudden loss of consciousness in November 2019. Computed tomography (CT) scan showed subdural hematoma, subfalcine herniation, and brain infarction. He developed thrombocytopenia and elevated transaminases, but no rashes or obvious kidney damage. He reported having a rat bite. HFRS was suspected. The Hantavirus IgG was positive, and the metagenomic next-generation sequencing (mNGS) detected SEOV sequences directly in the blood. CONCLUSION: This report highlights the importance of suspecting SEOV infection in febrile patients with thrombocytopenia and elevated liver enzymes despite the absence of hemorrhagic manifestations of skin and renal syndromes. Next-generation sequencing is a powerful tool for pathogen detection. Intracranial hemorrhage and brain infarction as extrarenal manifestations of HFRS are rare but possible as demonstrated in this case.

Autochthonous Ratborne Seoul Virus Infection in Woman with Acute Kidney Injury.

Outside Asia, Seoul virus (SEOV) is an underestimated pathogen. In Germany, autochthonous SEOV-associated hantavirus disease has not been unequivocally diagnosed. We found clinical and molecular evidence for SEOV infection in a young woman; her pet rat was the source of infection.

Seoul Orthohantavirus in Wild Black Rats, Senegal, 2012-2013.

Hantaviruses cause hemorrhagic fever in humans worldwide. However, few hantavirus surveillance campaigns occur in Africa. We detected Seoul orthohantavirus in black rats in Senegal, although we did not find serologic evidence of this disease in humans. These findings highlight the need for increased surveillance of hantaviruses in this region.

Seoul Virus Infection and Spread in United States Home-Based Ratteries: Rat and Human Testing Results From a Multistate Outbreak Investigation.

BACKGROUND: During 2017, a multistate outbreak investigation occurred after the confirmation of Seoul virus (SEOV) infections in people and pet rats. A total of 147 humans and 897 rats were tested. METHODS: In addition to immunoglobulin (Ig)G and IgM serology and traditional reverse-transcription polymerase chain reaction (RT-PCR), novel quantitative RT-PCR primers/probe were developed, and whole genome sequencing was performed. RESULTS: Seventeen people had SEOV IgM, indicating recent infection; 7 reported symptoms and 3 were hospitalized. All patients recovered. Thirty-one facilities in 11 US states had SEOV infection, and among those with ≥10 rats tested, rat IgG prevalence ranged 2%-70% and SEOV RT-PCR positivity ranged 0%-70%. Human laboratory-confirmed cases were significantly associated with rat IgG positivity and RT-PCR positivity (P = .03 and P = .006, respectively). Genomic sequencing identified >99.5% homology between SEOV sequences in this outbreak, and these were >99% identical to SEOV associated with previous pet rat infections in England, the Netherlands, and France. Frequent trade of rats between home-based ratteries contributed to transmission of SEOV between facilities. CONCLUSIONS: Pet rat owners, breeders, and the healthcare and public health community should be aware and take steps to prevent SEOV transmission in pet rats and to humans. Biosecurity measures and diagnostic testing can prevent further infections.