Hantaan virus replication is promoted via AKT activated mitochondria OXPHOS

Oxidative phosphorylation (OXPHOS) is a vital pathway provides ATP for intracellular activities. Here, we found that Hantaan virus (HTNV) exploited mitochondria OXPHOS to assist its replication in host cells and Protein Kinase B/AKT played a major function in this process. Inhibiting AKT activation by BEZ treatment can inhibit HTNV replication and prevent the increase of OXPHOS level caused by HTNV infection. We also found that HTNV infection can promote AKT translocation to mitochondria, where AKT phosphorylates Polynucleotide phosphorylase (PNPT). Taken together, our research demonstrates that HTNV replication exploits OXPHOS in host cells and it increases OXPHOS function by AKT-PNPT interaction in mitochondria.

Differential Macrophage Phenotype Rewired by Hantaan Virus Constrains the Magnitude of Inflammatory Responses in Murine versus Humans

Hantaan virus (HTNV) is principally maintained and transmitted by rodents in nature, the infection of which is non-pathogenic in the field or laboratory mouse, but can cause hemorrhagic fever with renal syndrome (HFRS) in human beings, a severe systemic inflammatory disease with high mortality. It remains obscure how HTNV infection leads to disparate outcomes in distinct species. Here, we revealed a differential immune status in murine versus humans post HTNV infection, which was orchestrated by the macrophage reprogramming process and characterized by late-phase inactivation of NF-κB signaling. In HFRS patients, the immoderate and continuous activation of inflammatory monocyte/macrophage (M1) launched TNFα-centered cytokine storm and aggravated host immunopathologic injury, which can be life-threatening; however, in field or laboratory mice, the M1 activation and TNFα release were significantly suppressed at the late infection stage of HTNV, restricting excessive inflammation and blocking viral disease process, which also protected mice from secondary LPS challenge or polymicrobial sepsis. Mechanistically, we found that murine macrophage phenotype was dynamically manipulated by HTNV via the Notch-lncRNA-p65 axis. At the early stage of HTNV infection, the intracellular domain of Notch receptor (NICD) was activated by viral nucleocapsid (NP) stimulation and potentiated the NF-κB pathway by associating with and facilitating the interaction between IKKβ and p65. At the late stage, Notch signaling launched the expression of diverse murine-specific long non-coding RNAs (lncRNAs) and attenuated M1 polarization. Among them, lncRNA 30740.1 (termed as lnc-ip65, an inhibitor of p65) bound to p65 and hindered its phosphorylation, exerting negative feedback on the NF-κB pathway. Genetic ablation of lnc-ip65 shifted the balance of macrophage polarization from a pro-resolution to an inflammatory phenotype, leading to superabundant production of pro-inflammatory cytokines and increasing mice susceptibility to HTNV infection or bacterial sepsis. Collectively, our findings identify an immune braking function and mechanism for murine lncRNAs in inhibiting p65-mediated M1 activation, opening a novel therapeutic avenue of controlling the magnitude of immune responses for HFRS and other inflammatory diseases.

Coumarin Derivative N6 as a Novel anti-hantavirus Infection Agent Targeting AKT

Hantaviruses are globally emerging zoonotic viruses that can cause hemorrhagic fever with renal syndrome (HFRS) in Asia and Europe, which is primarily caused by Hantaan virus (HTNV) infection, results in profound morbidity and mortality. However, no specific treatment is available for this disease. Coumarin derivatives have been reported as antiviral molecules, while studies about the bioactivity of coumarin derivatives against HTNV infection are limited. To study the potential antiviral activity of coumarin derivatives, 126 coumarin derivatives are synthesized, and their inhibitory activity against HTNV is analyzed in vitro. Among these compounds, N6 inhibits HTNV with relatively high selectivity index at 10.9, and the viral titer of HTNV is reduced significantly after 5, 10, and 20 μM N6 treatments. Furthermore, the administration of N6 at the early stage of HTNV infection can inhibit the replication and production of infectious HTNV in host cell, this therapeutic efficacy is confirmed in HTNV-infected newborn mice at the early stage of infection. The molecular docking results show that N6 forms interactions with the key amino acid residues at its active site, and reveals several molecular interactions responsible for the observed affinity, and the treatment of N6 can inhibit the expression of p (Ser473)Akt and HTNV nucleocapsid protein significantly. As such, these observations demonstrate that coumarin derivative N6 might be used as a potential agent against HTNV infection.

Urinary genome detection and tracking of Hantaan virus from hemorrhagic fever with renal syndrome patients using multiplex PCR-based next-generation sequencing

Background Hantavirus infection occurs through the inhalation of aerosolized excreta, including urine, feces, and saliva of infected rodents. The presence of Hantaan virus (HTNV) RNA or infectious particles in urine specimens of patient with hemorrhagic fever with renal syndrome (HFRS) remains to be investigated. Methodology/Principal findings We collected four urine and serum specimens of Republic of Korea Army (ROKA) patients with HFRS. We performed multiplex PCR-based next-generation sequencing (NGS) to obtain the genome sequences of clinical HTNV in urine specimens containing ultra-low amounts of viral genomes. The epidemiological and phylogenetic analyses of HTNV demonstrated geographically homogenous clustering with those in Apodemus agrarius captured in highly endemic areas, indicating that phylogeographic tracing of HTNV genomes reveals the potential infection sites of patients with HFRS. Genetic exchange analyses showed a genetic configuration compatible with HTNV L segment exchange in nature. Conclusion/Significance Our results suggest that whole or partial genome sequences of HTNV from the urine enabled to track the putative infection sites of patients with HFRS by phylogeographically linking to the zoonotic HTNV from the reservoir host captured at endemic regions. This report raises awareness among physicians for the presence of HTNV in the urine of patients with HFRS.

Comparison of transcriptional responses between pathogenic and nonpathogenic hantavirus infections in Syrian hamsters using NanoString

Background Syrian hamsters infected with Andes virus (ANDV) develop a disease that recapitulates many of the salient features of human hantavirus pulmonary syndrome (HPS), including lethality. Infection of hamsters with Hantaan virus (HTNV) results in an asymptomatic, disseminated infection. In order to explore this dichotomy, we examined the transcriptome of ANDV- and HTNV-infected hamsters. Results Using NanoString technology, we examined kinetic transcriptional responses in whole blood collected from ANDV- and HTNV-infected hamsters. Of the 770 genes analyzed, key differences were noted in the kinetics of type I interferon sensing and signaling responses, complement activation, and apoptosis pathways between ANDV- and HTNV-infected hamsters. Conclusions Delayed activation of type I interferon responses in ANDV-infected hamsters represents a potential mechanism that ANDV uses to subvert host immune responses and enhance disease. This is the first genome-wide analysis of hantavirus-infected hamsters and provides insight into potential avenues for therapeutics to hantavirus disease.

Integrative Analysis of HTNV Glycoprotein Derived MHC II Epitopes by In Silico Prediction and Experimental Validation

Hantaan virus (HTNV), the causative pathogen of hemorrhagic fever with renal syndrome (HFRS), is a negative RNA virus belonging to the Orthohantaviridae family. HTNV envelope glycoprotein (GP), encoded by the genomic medium segment, is immunogenic and is therefore a promising vaccine candidate. Major histocompatibility complex class I (MHC-I) epitopes derived from HTNV has been extensively studied, but little is known of MHC-II epitopes. In silico predictions based on four databases indicated that the full-length HTNV GP has 1121 15-mer epitopes, of which 289 had a high score for binding to the human and murine MHC-II superfamily. It found that epitope ILTVLKFIANIFHTS could potentially bind most MHC-II molecules covering human and murine haplotypes. Dominant epitopes were validated by enzyme-linked immunospot assay of splenocytes from immunized mice; 6 of 10 epitopes supported the predictions including TATYSIVGPANAKVP, TKTLVIGQCIYTITS, FSLLPGVAHSIAVEL, CETYKELKAHGVSCP, CGLYLDRLKPVGSAY, and NLGENPCKIGLQTSS. Conservation analysis of dominant epitopes revealed host–virus interactions without geographic stratification, thus meeting the requirements of candidate vaccines for large-population prophylaxis. These findings provide insight into hantavirus antigenicity and suggest that vaccines targeting MHC-II could provide immune protection in large population to complement symptomatic therapies for the treatment of HFRS.

Nlrc3 Knockout Mice Showed Renal Pathological Changes After HTNV Infection

Hantaan virus (HTNV) infects humans and causes hemorrhagic fever with renal syndrome (HFRS). The development of well-characterized animal models of HFRS could accelerate the testing of vaccine candidates and therapeutic agents and provide a useful tool for studying the pathogenesis of HFRS. Because NLRC3 has multiple immunoregulatory roles, we investigated the susceptibility of Nlrc3−/− mice to HTNV infection in order to establish a new model of HFRS. Nlrc3−/− mice developed weight loss, renal hemorrhage, and tubule dilation after HTNV infection, recapitulating many clinical symptoms of human HFRS. Moreover, infected Nlrc3−/− mice showed higher viral loads in serum, spleen, and kidney than wild type C57BL/6 (WT) mice, and some of them manifested more hematological disorders and significant pathological changes within multiple organs than WT mice. Our results identify that HTNV infected Nlrc3−/− mice can develop clinical symptoms and pathological changes resembling patients with HFRS, suggesting a new model for studying the pathogenesis and testing of candidate vaccines and therapeutics.

Acute Appendicitis Associated with Hantaan Virus Infection

Hantaviruses are Bunyaviridae viruses that cause hemorrhagic fever with renal syndrome (HFRS). Appendicitis caused by Hantaan virus has not been reported previously. An 81-year-old man who underwent laparoscopic appendectomy for suspected appendicitis based on abdominal pain, fever, hypotension, and computed tomography findings. Based on a suspicion of hemorrhagic fever with renal syndrome, the patient’s plasma was simultaneously analyzed using an indirect immunofluorescent antibody assay and nested reverse transcription–polymerase chain reaction (RT-PCR). The appendix tissue was also analyzed using nested RT-PCR and immunohistochemical (IHC) staining to identify the presence of Hantaan virus. Nested RT-PCR detected the presence of Hantaan virus, and indirect immunofluorescent antibody assay results revealed the presence of elevated antibody levels. Furthermore, IHC staining of the appendix tissue confirmed Hantaan virus antigens in the peripheral nerve bundle. Based on these findings, we confirmed the nerve tropism of the Hantaan virus. Hantaan virus in plasma and appendix tissue samples was confirmed using PCR and phylogenetic tree analysis. Moreover, we detected hypertrophy of the submucosa and periappendiceal adipose tissue nerve bundle along with Hantaan virus antigens in peripheral nerve bundles using IHC staining. Hence, we report that Hantaan virus infection may be accompanied by appendicitis.

Structural Basis for a Neutralizing Antibody Response Elicited by a Recombinant Hantaan Virus Gn Immunogen

The spillover of pathogenic hantaviruses from rodent reservoirs into the human population poses a continued threat to human health. Here, we show that a recombinant form of the Hantaan virus (HTNV) surface-displayed glycoprotein, Gn, elicits a neutralizing antibody response in rabbits.

Antiviral Efficacy of Ribavirin and Favipiravir against Hantaan Virus

Ecological changes, population movements and increasing urbanization promote the expansion of hantaviruses, placing humans at high risk of virus transmission and consequent diseases. The currently limited therapeutic options make the development of antiviral strategies an urgent need. Ribavirin is the only antiviral used currently to treat hemorrhagic fever with renal syndrome (HFRS) caused by Hantaan virus (HTNV), even though severe side effects are associated with this drug. We therefore investigated the antiviral activity of favipiravir, a new antiviral agent against RNA viruses. Both ribavirin and favipiravir demonstrated similar potent antiviral activity on HTNV infection. When combined, the efficacy of ribavirin is enhanced through the addition of low dose favipiravir, highlighting the possibility to provide better treatment than is currently available.