High-resolution phylogeographical surveillance of Hantaan orthohantavirus using rapid amplicon-based Flongle sequencing, Republic of Korea.

Orthohantaviruses, etiological agents of hemorrhagic fever with renal syndrome (HFRS) and hantavirus cardiopulmonary syndrome, pose a critical public health threat worldwide. Hantaan orthohantavirus (HTNV) outbreaks are particularly endemic in Gyeonggi Province in northern area of the Republic of Korea (ROK). Small mammals were collected from three regions in the Gyeonggi Province during 2017 and 2018. Serological and molecular prevalence of HTNV was 25/201 (12.4%) and 10/25 (40%), respectively. A novel nanopore-based diagnostic assay using a cost-efficient Flongle chip was developed to rapidly and sensitively detect HTNV infection in rodent specimens within 3 h. A rapid phylogeographical surveillance of HTNV at high-resolution phylogeny was established using the amplicon-based Flongle sequencing. In total, seven whole-genome sequences of HTNV were newly obtained from wild rodents collected in Paju-si (Gaekhyeon-ri) and Yeoncheon-gun (Hyeonga-ri and Wangnim-ri), Gyeonggi Province. Phylogenetic analyses revealed well-supported evolutionary divergence and genetic diversity, enhancing the resolution of the phylogeographic map of orthohantaviruses in the ROK. Incongruences in phylogenetic patterns were identified among HTNV tripartite genomes, suggesting differential evolution for each segment. These findings provide crucial insights into on-site diagnostics, genome-based surveillance, and the evolutionary dynamics of orthohantaviruses to mitigate hantaviral outbreaks in HFRS-endemic areas in the ROK.

Clinical and etiological characteristics of severe hemorrhagic fever caused by coinfection of hantaan orthohantavirus and severe fever with thrombocytopenia syndrome virus.

Severe fever with thrombocytopenia syndrome (SFTS) and hemorrhagic fever with renal syndrome (HFRS) usually have different infection routes, and coinfection is relatively rare. This study examines the clinical and etiological characteristics of coinfection by these two pathogens to provide important references for clinical diagnosis and treatment. Blood samples from 22 clinically diagnosed patients with HFRS were collected for molecular detection of HFRS and common tick and mouse borne diseases. Inoculate the blood of six severe and critically patients into cells to isolate and proliferate potential viruses, and retest the cell culture to determine the pathogen. In addition, complete data were collected from these 22 HFRS and concurrent SFTS patients, and white blood cells (WBCs), platelet (PLT), blood urea nitrogen (BUN), creatinine (Cr) and other data were compared and analyzed. A total of 31 febrile patients, including 22 HFRS patients and 9 SFTS patients, were collected from September 2021 to October 2022. Among these HFRS patients, 11 were severe or critical. Severe and critical HFRS patients were characterized by rodent exposure history, pharyngeal and conjunctival hyperemia, abnormal WBC and PLT counts, and elevated BUN and Cr values. Virus isolation and molecular detection on blood samples from 6 patients showed that three of the six severe patients were positive for hantaan virus (HTNV), and two of the three HTNV positives were also positive for SFTS bunyavirus (SFTSV). The two coinfected patients exhibited different clinical and laboratory characteristics compared to those infected by either virus alone. Coinfection of HTNV and SFTSV leads to severe and complex hemorrhagic fever. Laboratory characteristics, such as the indicators of WBC, PLT, BUN, and Cr, may differ between HFRS and SFTS. These findings have implications and provide references for the diagnosis and treatment of coinfected cases.

Insight into the Hantaan virus RNA-dependent RNA polymerase inhibition using in-silico approaches.

The Hantaan virus (HTN) is a member of the hantaviridae family. It is a segmented type, negative-strand virus (sNSVs). It causes hemorrhagic fever with renal syndrome, which includes fever, vascular hemorrhage, and renal failure. This illness is one of the most serious hemorrhagic diseases in the world, and it is a major public health concern due to its high mortality rate. The Hantaan virus RNA-dependent RNA polymerase complex (RdRp) is involved in viral RNA transcription and replication for the survival and transmission of this virus. Therefore, it is a primary target for antiviral drug development. Interference with the endonucleolytic "cap-snatching" reaction by the HTN virus RdRp endonuclease domain is a particularly appealing approach for drug discovery against this virus. This RdRp endonuclease domain of the HTN virus has a metal-dependent catalytic activity. We targeted this metal-dependent enzymatic activity to identify inhibitors that can bind and disrupt this endonuclease enzyme activity using in-silico approaches i.e., molecular docking, molecular dynamics simulation, predicted absorption, distribution, metabolism, excretion, toxicity (ADMET) and drug-likeness studies. The docking studies showed that peramivir, and ingavirin compounds can effectively bind with the manganese ions and engage with other active site residues of this protein. Molecular simulations also showed stable binding of these ligands with the active site of HTN RdRp. Simulation analysis showed that they were in constant contact with the active site manganese ions and amino acid residues of the HTN virus endonuclease domain. This study will help in better understanding the HTN and related viruses.

Protective CD8+ T-cell response against Hantaan virus infection induced by immunization with designed linear multi-epitope peptides in HLA-A2.1/Kb transgenic mice.

BACKGROUND: An effective vaccine that prevents disease caused by hantaviruses is a global public health priority, but up to now, no vaccine has been approved for worldwide use. Therefore, novel vaccines with high prophylaxis efficacy are urgently needed. METHODS: Herein, we designed and synthesized Hantaan virus (HTNV) linear multi-epitope peptide consisting of HLA-A*02-restricted HTNV cytotoxic T cell (CTL) epitope and pan HLA-DR-binding epitope (PADRE), and evaluated the immunogenicity, as well as effectiveness, of multi-epitope peptides in HLA-A2.1/Kb transgenic mice with interferon (IFN)-γ enzyme-linked immunospot assay, cytotoxic mediator detection, proliferation assay and HTNV-challenge test. RESULTS: The results showed that a much higher frequency of specific IFN-γ-secreting CTLs, high levels of granzyme B production, and a strong proliferation capacity of specific CTLs were observed in splenocytes of mice immunized with multi-epitope peptide than in those of a single CTL epitope. Moreover, pre-immunization of multi-epitope peptide could reduce the levels of HTNV RNA loads in the liver, spleen and kidneys of mice, indicating that specific CTL responses induced by multi-epitope peptide could reduce HTNV RNA loads in vivo. CONCLUSIONS: This study may provide an important foundation for the development of novel peptide vaccines for HTNV prophylaxis.

Molecular detection of viruses causing hemorrhagic fevers in rodents in the south-west of Korea.

Many pathogens causing hemorrhagic fevers of medical and veterinary importance have been identified and isolated from rodents in the Republic of Korea (ROK). We investigated the occurrence of emerging viruses causing hemorrhagic fevers, such as hemorrhagic fever with renal syndrome (HFRS), severe fever with thrombocytopenia syndrome (SFTS), and flaviviruses, from wild rodents. Striped field mice, Apodemus agrarius (n = 39), were captured during 2014-2015 in the south-west of ROK. Using molecular methods, lung samples were evaluated for SFTS virus, hantavirus, and flavivirus, and seropositivity was evaluated in the blood. A high positive rate of hantavirus (46.2%) was detected in A. agrarius lungs by reverse transcription-nested polymerase chain reaction (RT-N-PCR). The monthly occurrence of hantavirus was 16.7% in October, 86.7% in November, and 25% in August of the following year (p < 0.001). Moreover, 17.9% of blood samples were serologically positive for hantavirus antibodies. The most prevalent strain in A. agrarius was Hantaan virus. All samples were positive for neither SFTS virus nor flavivirus. Hantaan virus was detected in 86.7% of A. agrarius in November (autumn), and thus, virus shedding from A. agrarius can increase the risk of humans contracting HFRS. These findings may help to predict and prevent disease outbreaks in ROK.

Recent Increase of Human Granulocytic Anaplasmosis and Co-Infection with Scrub Typhus or Korean Hemorrhagic Fever with Renal Syndrome in Korea.

We report 17 patients with human granulocytic anaplasmosis between January 2015 and September 2018 at two tertiary university hospitals in Korea. Monthly incidence peaked in May and June. Among these patients, we identified three who were co-infected with scrub typhus, and one patient with hemorrhagic fever with renal syndrome.

The Long Noncoding RNA NEAT1 Exerts Antihantaviral Effects by Acting as Positive Feedback for RIG-I Signaling.

Hantavirus infection, which causes zoonotic diseases with a high mortality rate in humans, has long been a global public health concern. Over the past decades, accumulating evidence suggests that long noncoding RNAs (lncRNAs) play key regulatory roles in innate immunity. However, the involvement of host lncRNAs in hantaviral control remains uncharacterized. In this study, we identified the lncRNA NEAT1 as a vital antiviral modulator. NEAT1 was dramatically upregulated after Hantaan virus (HTNV) infection, whereas its downregulation in vitro or in vivo delayed host innate immune responses and aggravated HTNV replication. Ectopic expression of NEAT1 enhanced beta interferon (IFN-ß) production and suppressed HTNV infection. Further investigation suggested that NEAT1 served as positive feedback for RIG-I signaling. HTNV infection activated NEAT1 transcription through the RIG-I-IRF7 pathway, whereas NEAT1 removed the transcriptional inhibitory effects of the splicing factor proline- and glutamine-rich protein (SFPQ) by relocating SFPQ to paraspeckles, thus promoting the expression of RIG-I and DDX60. RIG-I and DDX60 had synergic effects on IFN production. Taken together, our findings demonstrate that NEAT1 modulates the innate immune response against HTNV infection, providing another layer of information about the role of lncRNAs in controlling viral infections.IMPORTANCE Hantaviruses have attracted worldwide attention as archetypal emerging pathogens. Recently, increasing evidence has highlighted long noncoding RNAs (lncRNAs) as key regulators of innate immunity; however, their roles in hantavirus infection remain unknown. In the present work, a new unexplored function of lncRNA NEAT1 in controlling HTNV replication was found. NEAT1 promoted interferon (IFN) responses by acting as positive feedback for RIG-I signaling. This lncRNA was induced by HTNV through the RIG-I-IRF7 pathway in a time- and dose-dependent manner and promoted HTNV-induced IFN production by facilitating RIG-I and DDX60 expression. Intriguingly, NEAT1 relocated SFPQ and formed paraspeckles after HTNV infection, which might reverse inhibitive effects of SFPQ on the transcription of RIG-I and DDX60. To the best of our knowledge, this is the first study to address the regulatory role of the lncRNA NEAT1 in host innate immunity after HTNV infection. In summary, our findings provide additional insights regarding the role of lncRNAs in controlling viral infections.

Hemorrhagic fever with renal syndrome accompanied by panhypopituitarism and central diabetes insipidus: a case report.

Central diabetes insipidus (DI) was detected in a patient with hemorrhagic fever with renal syndrome (HFRS) who had been molecularly and serologically diagnosed with Hantaan virus infection. We recommend that clinicians differentiate central DI in HFRS patients with a persistent diuretic phase even when pituitary MRI findings are normal.

Type inactivated bivalent hantavirus vaccines using dual fluorescence quantitative real-time PCR: Establishment and evaluation.

To establish a dual fluorescence quantitative real-time polymerase chain reaction (DFqRT-PCR) method for typing inactivated bivalent vaccines of Hantaan virus (HTNV, type I) and Seoul virus (SEOV, type II) found in China. Primers and probes were designed based on S segment of the Hantavirus. FAM and HEX were added to the probes for the type I and type II vaccines, respectively. The S fragments were cloned into the pMD18-T vector to create pMD18-T/HTNV and pMD18-T/SEOV plasmids, respectively. The DFqRT-PCR reaction conditions were optimized. The specificity, sensitivity, accuracy, and reproducibility of the DFqRT-PCR method within and among batches of the samples and labs were evaluated. The sensitivity of the DFqRT-PCR method for detecting type I and II vaccines was 4.68 × 101 copies/µL and 5 × 101 copies/µL of pMD18-T/HTNV and pMD18-T/SEOV, respectively. This method distinguished the hantavirus vaccines from several other virus genus and distinguished the type I and II vaccines from each other with coefficients of variation (CVs) of the intra- and inter-batch assay of 0.19-0.66% and 0.09-0.69%, respectively. For bivalent inactivated vaccine, the CVs of the intra- and inter-batch assays, and the intra- and inter-laboratory assays were 0.06-1.82% for the samples from the same batches and different batches from the same manufacturer or the samples from the different manufacturers. The DFqRT-PCR method was simple, fast, and sensitive for identifying and determining the inactivated hantavirus vaccine with high accuracy and reproducibility. Therefore, it can be used for quality control of hantavirus vaccines. J. Med. Virol. 89:10-16, 2017. © 2016 Wiley Periodicals, Inc.

Appearance of renal hemorrhage in adult mice after inoculation of patient-derived hantavirus.

BACKGROUND: Hemorrhagic fever with renal syndrome (HFRS) caused by hantavirus infection is characterized by fever, renal dysfunction and hemorrhage. An animal model mimicking symptoms of HFRS remains to be established. In this study, we evaluated the pathogenicity of an HFRS patient-derived Hantaan virus (HTNV) in adult mice. METHODS: Five clones of HTNV strain KHF 83-61 BL (KHFV) that was derived from blood of an HFRS patient were obtained by plaque cloning. The pathogenicity of the virus clones was evaluated by using 6-week-old female BALB/c mice. Sequence analysis of the viral genome was performed by conventional methods. RESULTS: All of the mice intravenously inoculated with KHFV clone (cl)-1, -2, -3 and -5 showed signs of disease such as transient body weight loss, ruffled fur, reduced activity and remarkably prominent hemorrhage in the renal medulla at 6 to 9 days post-inoculation (dpi) and then recovered. In contrast, mice intravenously inoculated with KHFV cl-4 did not show any signs of disease. We selected KHFV cl-5 and cl-4 as representative of high-pathogenic and low-pathogenic clones, respectively. Quantities of viral RNA in kidneys of KHFV cl-5-infected mice were larger than those in KHFV cl-4-infected mice at any time point examined (3, 6, 9 and 12 dpi). The quantities of viral RNA of KHFV cl-5 and cl-4 peaked at 3 dpi, which was before the onset of disease. Sequence analysis revealed that the amino acid at position 417 in the glycoprotein Gn was the sole difference in viral proteins between KHFV cl-5 and cl-4. The result suggests that amino acid at position 417 in Gn is related to the difference in pathogenicity between KHFV cl-5 and cl-4. When the inoculum of KHFV cl-5 was pretreated with a neutralizing antibody against HTNV strain 76-118, which belongs to the same serotype as KHFV clones, mice did not show any signs of disease, confirming that the disease was caused by KHFV infection. CONCLUSION: We found that an HFRS patient-derived HTNV caused renal hemorrhage in adult mice. We anticipate that this infection model will be a valuable tool for understanding the pathogenesis of HFRS.

Analysis of hemorrhagic fever with renal syndrome and its pathogenic gene sequence based on geographic information system.

This study analyzed the temporal-spatial distribution characteristics, epidemiological characteristics and gene sequences of hemorrhagic fever with renal syndrome (HFRS) in Guangxi, with the intention of providing a theoretical and technical support for the prevention of HFRS. A map of the incidence of HFRS of different cities in Guangxi was drawn up using the Geographic Information System (GIS) to investigate the epidemiological characteristics and infection source of HFRS between 2013 and 2016. Guangxi has a low incidence of HFRS, and autumn and winter are the main high-incidence seasons. Cases of HFRS were reported in all regions in Guangxi except Laibin city between 2013 and 2016. The distribution of cases in the four years suggested that Guilin, Nanning, Hechi and Wuzhou were the main infected regions, especially the local areas in the north of Guilin. The nucleotide and amino acid of S fragment and M fragment of Hantaviruses (HV) detected were highly homologous, and no obvious variation was found. Through analyzing the space-time characteristics, epidemiological characteristics and gene sequence of HFRS in Guangxi, it was found that areas rich in water, grass and moisture, such as paddy fields, are the main active areas for the host of HFRS.

Detection of Hantaan virus RNA from anti-Hantaan virus IgG seronegative rodents in an area of high endemicity in Republic of Korea.

Hantaan virus (HTNV), of the family Bunyaviridae, causes hemorrhagic fever with renal syndrome (HFRS) in humans. Although the majority of epidemiologic studies have found that rodents are seropositive for hantavirus-specific immunoglobulin, the discovery of hantavirus RNA in seronegative hosts has led to an investigation of the presence of HTNV RNA in rodents captured in HFRS endemic areas. HTNV RNA was detected in seven (3.8%) of 186 anti-HTNV IgG seronegative rodents in Republic of Korea (ROK) during 2013-2014. RT-qPCR for HTNV RNA revealed dynamic virus-host interactions of HTNV in areas of high endemicity, providing important insights into the epidemiology of hantaviruses.

Genetic characterization of hantaviruses isolated from rodents in the port cities of Heilongjiang, China, in 2014.

BACKGROUND: Hantavirus is a tripartite negative-sense RNA virus. It can infect humans through contaminated rodent excreta and causes two types of fatal human diseases: hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS). China exhibits the highest HFRS occurrence rate in the world, and the Heilongjiang area is one of the most severely infected regions. RESULTS: To obtain additional insights into the genetic characteristics of hantaviruses in the port cities of the Heilongjiang area in China, a molecular epidemiological investigation of hantaviruses isolated from rodents was performed in 2014. A total of 649 rodents (11 murine species and 1 shrew species) were caught in 12 port cities in Heilongjiang. Among these rodents, the most common species was A. agrarius, and the second-most common was R. norvegicus. A viral gene PCR assay revealed the presence of two specific genotypes of hantavirus, referred to as Hantaan virus (HTNV) and Seoul virus (SEOV), and the positive SEOV infection rate was higher than that for HTNV. A genetic analysis based on partial M segment sequences indicated that all of the isolates belonging to SEOV could be assigned to two genetic lineages, whereas the isolate belonging to HTNV could be assigned to only one genetic lineage. CONCLUSIONS: These results suggested that HTNV and SEOV are circulating in A. agrarius and R. norvegicus in the port cities in the area of Heilongjiang, but SEOV may be the dominant common hantavirus.

The murine model for Hantaan virus-induced lethal disease shows two distinct paths in viral evolutionary trajectory with and without ribavirin treatment.

In vitro, ribavirin acts as a lethal mutagen in Hantaan virus (HTNV)-infected Vero E6 cells, resulting in an increased mutation load and viral population extinction. In this study, we asked whether ribavirin treatment in the lethal, suckling mouse model of HTNV infection would act similarly. The HTNV genomic RNA (vRNA) copy number and infectious virus were measured in lungs of untreated and ribavirin-treated mice. In untreated, HTNV-infected mice, the vRNA copy number increased for 10 days postinfection (dpi) and thereafter remained constant through 26 dpi. Surprisingly, in ribavirin-treated, HTNV-infected mice, vRNA levels were similar to those in untreated mice between 10 and 26 dpi. Infectious virus levels, however, were different: in ribavirin-treated mice, the amount of infectious HTNV was significantly decreased relative to that in untreated mice, suggesting that ribavirin reduced the specific infectivity of the virus (amount of infectious virus produced per vRNA copy). Mutational analysis revealed a ribavirin-associated elevation in mutation frequency in HTNV vRNA similar to that previously reported in vitro. Codon-based analyses of rates of nonsynonymous (dN) and synonymous (dS) substitutions in the S segment revealed a positive selection for codons within the HTNV N protein gene in the ribavirin-treated vRNA population. In contrast, the vRNA population in untreated, HTNV-infected mice showed a lower level of diversity, reflecting purifying selection for the wild-type genome. In summary, these experiments show two different evolutionary paths that Hantavirus may take during infection in a lethal murine model of disease, as well as the importance of the in vivo host environment in the evolution of the virus, which was not apparent in our prior in vitro model system.

Development of an immunochromatography strip test based on truncated nucleocapsid antigens of three representative hantaviruses.

BACKGROUND: Hantaviruses are causative agents of hemorrhagic fever with renal syndrome (HFRS) and nephropathia epidemica (NE) in the Old World and hantavirus pulmonary syndrome (HPS) in the New World. There is a need for time-saving diagnostic methods. In the present study, recombinant N antigens were used as antigens in an immunochromatography strip (ICG) test to detect specific IgG antibodies. METHODS: The N-terminal 103 amino acids (aa) of Hantaan virus (HTNV), Puumala virus (PUUV) and Andes virus (ANDV) nucleocapsid (N) protein were expressed in E. coli as representative antigens of three groups (HFRS, NE and HPS-causing viruses) of hantavirus. Five different types of ICG test strips, one antigen line on one strip for each of the three selected hantaviruses (HTNV, PUUV and ANDV), three antigen lines on one strip and a mixed antigen line on one strip, were developed and sensitivities were compared. RESULTS: A total of 87 convalescent-phase patient sera, including sera from 35 HFRS patients, 36 NE patients and 16 HPS patients, and 25 sera from healthy seronegative people as negative controls were used to evaluate the ICG test. Sensitivities of the three-line strip and mixed-line strip were similar to those of the single antigen strip (97.2 to 100%). On the other hand, all of the ICG test strips showed high specificities to healthy donors. CONCLUSION: These results indicated that the ICG test with the three representative antigens is an effective serodiagnostic tool for screening and typing of hantavirus infection in humans.

Hantaan virus inhibits type I interferon response by targeting RLR signaling pathways through TRIM25.

Hantaan virus (HTNV) is responsible for hemorrhagic fever with renal syndrome (HFRS), primarily due to its ability to inhibit host innate immune responses, such as type I interferon (IFN-I). In this study, we conducted a transcriptome analysis to identify host factors regulated by HTNV nucleocapsid protein (NP) and glycoprotein. Our findings demonstrate that NP and Gc proteins inhibit host IFN-I production by manipulating the retinoic acid-induced gene I (RIG-I)-like receptor (RLR) pathways. Further analysis reveals that HTNV NP and Gc proteins target upstream molecules of MAVS, such as RIG-I and MDA-5, with Gc exhibiting stronger inhibition of IFN-I responses than NP. Mechanistically, NP and Gc proteins interact with tripartite motif protein 25 (TRIM25) to competitively inhibit its interaction with RIG-I/MDA5, suppressing RLR signaling pathways. Our study unveils a cross-talk between HTNV NP/Gc proteins and host immune response, providing valuable insights into the pathogenic mechanism of HTNV.

The Role of Long Noncoding RNA Negative Regulator of Interferon Response in the Regulation of Hantaan Virus Infection.

Hantaan virus (HTNV) is prevalent in Eurasia. It causes hemorrhagic fever with renal syndrome (HFRS). Long noncoding RNAs (lncRNAs) play key roles in regulating innate immunity. Among these, lncRNA negative regulator of interferon response (NRIR) was reported as an inhibitor of several interferon (IFN)-stimulated genes. Our results showed that: NRIR expression was upregulated by HTNV infection in a type I IFN-dependent manner. The expression of NRIR in CD14+ monocytes from HFRS patients in acute phase was significantly higher than that in convalescent phase and healthy controls. HTNV infection in some HTNV-compatible cells was promoted by NRIR. NRIR negatively regulated innate immunity, especially IFITM3 expression. Localized in the nucleus, NRIR bound with HNRNPC, and knockdown of HNRNPC significantly weakened the effect of NRIR in promoting HTNV infection and restored IFITM3 expression. These results indicated that NRIR regulates the innate immune response against HTNV infection possibly through its interaction with HNRNPC and its influence on IFITM3.

Development of an all-in-one real-time PCR assay for simultaneous detection of spotted fever group rickettsiae, severe fever with thrombocytopenia syndrome virus and hantaan virus prevalent in central China.

Central China has been reported to be one of the most important endemic areas of zoonotic infection by spotted fever group rickettsiae (SFGR), severe fever with thrombocytopenia syndrome virus (SFTSV) and hantaan virus (HTNV). Due to similar clinical symptoms, it is challenging to make a definite diagnosis rapidly and accurately in the absence of microbiological tests. In the present study, an all-in-one real-time PCR assay was developed for the simultaneous detection of nucleic acids from SFGR, SFTSV and HTNV. Three linear standard curves for determining SFGR-ompA, SFTSV-L and HTNV-L were obtained within the range of 101-106 copies/µL, with the PCR amplification efficiencies ranging from 93.46% to 96.88% and the regression coefficients R2 of >0.99. The detection limit was 1.108 copies/µL for SFGR-ompA, 1.075 copies/µL for SFTSV-L and 1.006 copies/µL for HTNV-L, respectively. Both the within-run and within-laboratory coefficients of variation on the cycle threshold (Ct) values were within the range of 0.53%-2.15%. It was also found there was no statistical difference in the Ct values between single template and multiple templates (PSFGR-ompA = 0.186, PSFTSV-L = 0.612, PHTNV-L = 0.298). The sensitivity, specificity, positive and negative predictive value were all 100% for determining SFGR-ompA and SFTSV-L, 97%, 100%, 100% and 99.6% for HTNV-L, respectively. Therefore, the all-in-one real-time PCR assay appears to be a reliable, sensitive, rapid, high-throughput and low cost-effective method to diagnose the zoonotic infection by SFGR, SFTSV and HTNV.

Isolation and characterization of genetic variants of Orthohantavirus hantanense from clinical cases of HFRS in Jiangxi Province, China.

BACKGROUND: Hemorrhagic fever with renal syndrome (HFRS) is a severe public health problem in Jiangxi province, China. Previous studies reported genetic variants of Orthohantavirus hantanense (Hantaan virus, HTNV) in rodents in this area. However, the relationship between HTNV variants and human infection needs to be confirmed. This study aimed to identify the HTNV variants in patients and to understand the clinical characteristics of HFRS caused by these variants. METHODS: Samples were collected from hospitalized suspected cases of HFRS during the acute phase. HFRS cases were confirmed using quantitative real-time RT-PCR. Peripheral blood mononuclear cells (PBMC) from patients with HFRS were inoculated into Vero-E6 cells for viral isolation. The genomic sequences of HTNV from patients were obtained by amplicon-based next-generation sequencing. A retrospective analysis was conducted on the clinical characteristics of the patients. RESULTS: HTNV RNA was detected in 53 of 183 suspected HFRS patients. Thirteen HTNVs were isolated from 32 PBMCs of HFRS cases. Whole genome sequences of 14 HTNVs were obtained, including 13 isolates in cell culture from 13 patients, and one from plasma of the fatal case which was not isolated successfully in cell culture. Genetic analysis revealed that the HTNV sequence from the 14 patients showed significant variations in nucleotide and amino acid to the HTNV strains found in other areas. Fever (100%, 53/53), thrombocytopenia (100%, 53/53), increased serum aspartate aminotransferase (100%, 53/53), and increased lactate dehydrogenase (96.2%, 51/53) were the most common characteristics. Severe acute kidney injury was observed in 13.2% (7/53) of cases. Clinical symptoms, such as pain, petechiae, and gastrointestinal or respiratory symptoms were uncommon. CONCLUSION: The HTNV genetic variants cause human infections in Jiangxi. The clinical symptoms of HFRS caused by the HTNV genetic variant during the acute phase are atypical. In addition to renal dysfunction, attention should be paid to the common liver injuries caused by these genetic variants.

Complete genome sequence of an amur virus isolated from Apodemus peninsulae in Northeastern China.

Amur virus was recently identified as the causative agent of hemorrhagic fever with renal syndrome. Here we report the complete genome sequence of an Amur virus isolated from Apodemus peninsulae in Northeastern China. The sequence information provided here is critical for the molecular epidemiology and evolution of Amur virus in China.