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1.
FEBS Open Bio ; 11(5): 1452-1464, 2021 05.
Article in English | MEDLINE | ID: mdl-33822489

ABSTRACT

Human pathogenic RNA viruses are threats to public health because they are prone to escaping the human immune system through mutations of genomic RNA, thereby causing local outbreaks and global pandemics of emerging or re-emerging viral diseases. While specific therapeutics and vaccines are being developed, a broad-spectrum therapeutic agent for RNA viruses would be beneficial for targeting newly emerging and mutated RNA viruses. In this study, we conducted a screen of repurposed drugs using Sendai virus (an RNA virus of the family Paramyxoviridae), with human-induced pluripotent stem cells (iPSCs) to explore existing drugs that may present anti-RNA viral activity. Selected hit compounds were evaluated for their efficacy against two important human pathogens: Ebola virus (EBOV) using Huh7 cells and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) using Vero E6 cells. Selective estrogen receptor modulators (SERMs), including raloxifene, exhibited antiviral activities against EBOV and SARS-CoV-2. Pioglitazone, a PPARγ agonist, also exhibited antiviral activities against SARS-CoV-2, and both raloxifene and pioglitazone presented a synergistic antiviral effect. Finally, we demonstrated that SERMs blocked entry steps of SARS-CoV-2 into host cells. These findings suggest that the identified FDA-approved drugs can modulate host cell susceptibility against RNA viruses.


Subject(s)
Antiviral Agents/pharmacology , Drug Repositioning , RNA Viruses/drug effects , RNA, Viral/antagonists & inhibitors , /drug effects , Animals , Cell Line , Chlorocebus aethiops , Drug Repositioning/methods , Ebolavirus/drug effects , Ebolavirus/physiology , Humans , Induced Pluripotent Stem Cells/virology , Microbial Sensitivity Tests/methods , Pioglitazone/pharmacology , RNA Viruses/physiology , Raloxifene Hydrochloride/pharmacology , Selective Estrogen Receptor Modulators/pharmacology , Sendai virus/drug effects , Sendai virus/physiology , Vero Cells
2.
BMC Infect Dis ; 21(1): 265, 2021 Mar 17.
Article in English | MEDLINE | ID: mdl-33731022

ABSTRACT

BACKGROUND: Increasing arbovirus infections have been a global burden in recent decades. Many countries have experienced the periodic emergence of arbovirus diseases. However, information on the prevalence of arboviruses is largely unknown or infrequently updated because of the lack of surveillance studies, especially in Africa. METHODS: A surveillance study was conducted in Gabon, Central Africa, on arboviruses, which are a major public health concern in Africa, including: West Nile virus (WNV), dengue virus (DENV), Zika virus (ZIKV), yellow fever virus (YFV), chikungunya virus (CHIKV), and Rift Valley fever virus (RVFV). Serological and molecular assays were performed to investigate past infection history and the current status of infection, using serum samples collected from healthy individuals and febrile patients, respectively. RESULTS: The overall seroprevalence during 2014-2017 was estimated to be 25.3% for WNV, 20.4% for DENV, 40.3% for ZIKV, 60.7% for YFV, 61.2% for CHIKV, and 14.3% for RVFV. No significant differences were found in the seroprevalence of any of the viruses between the male and female populations. However, a focus on the mean age in each arbovirus-seropositive individual showed a significantly younger age in WNV- and DENV-seropositive individuals than in CHIKV-seropositive individuals, indicating that WNV and DENV caused a relatively recent epidemic in the region, whereas CHIKV had actively circulated before. Of note, this indication was supported by the detection of both WNV and DENV genomes in serum samples collected from febrile patients after 2016. CONCLUSIONS: This study revealed the recent re-emergence of WNV and DENV in Gabon as well as the latest seroprevalence state of the major arboviruses, which indicated the different potential risks of virus infections and virus-specific circulation patterns. This information will be helpful for public health organizations and will enable a rapid response towards these arbovirus infections, thereby preventing future spread in the country.


Subject(s)
Arboviruses/isolation & purification , Dengue/epidemiology , Zika Virus Infection/epidemiology , Adolescent , Animals , Arbovirus Infections/diagnosis , Arbovirus Infections/epidemiology , Arboviruses/classification , Child , Child, Preschool , Communicable Diseases, Emerging , Dengue/diagnosis , Female , Fever/epidemiology , Fever/virology , Gabon/epidemiology , Humans , Infant , Male , Public Health , Seroepidemiologic Studies , Zika Virus Infection/diagnosis
3.
Int J Infect Dis ; 105: 452-459, 2021 Apr.
Article in English | MEDLINE | ID: mdl-33667697

ABSTRACT

OBJECTIVES: Lymphocytic choriomeningitis virus (LCMV), a human pathogenic arenavirus, is distributed worldwide. However, no human cases have been reported in Africa. This study aimed to investigate the current situation and potential risks of LCMV infection in Gabon, Central Africa. METHODS: A total of 492 human samples were screened to detect LCMV genome RNA and anti-LCMV IgG antibodies using reverse transcription-quantitative PCR and enzyme-linked immunosorbent assay (ELISA), respectively. ELISA-positive samples were further examined using a neutralization assay. Viral RNAs and antibodies were also analyzed in 326 animal samples, including rodents, shrews, and bushmeat. RESULTS: While no LCMV RNA was detected in human samples, the overall seroprevalence was 21.5% and was significantly higher in male and adult populations. The neutralization assay identified seven samples with neutralizing activity. LCMV RNA was detected in one species of rodent (Lophuromys sikapusi) and a porcupine, and anti-LCMV IgG antibodies were detected in four rodents and three shrews. CONCLUSIONS: This study determined for the first time the seroprevalence of LCMV in Gabon, and revealed that local rodents, shrews, and porcupines in areas surrounding semi-urban cities posed an infection risk. Hence, LCMV infection should be considered a significant public health concern in Africa.

4.
Biochem Biophys Res Commun ; 545: 203-207, 2021 03 19.
Article in English | MEDLINE | ID: mdl-33571909

ABSTRACT

The current COVID-19 pandemic requires urgent development of effective therapeutics. 5-amino levulinic acid (5-ALA) is a naturally synthesized amino acid and has been used for multiple purposes including as an anticancer therapy and as a dietary supplement due to its high bioavailability. In this study, we demonstrated that 5-ALA treatment potently inhibited infection of SARS-CoV-2, a causative agent of COVID-19, in cell culture. The antiviral effects could be detected in both human and non-human cells, without significant cytotoxicity. Therefore, 5-ALA is worth to be further investigated as an antiviral drug candidate for COVID-19.


Subject(s)
Antiviral Agents/pharmacology , Levulinic Acids/pharmacology , Animals , Antiviral Agents/administration & dosage , /virology , Caco-2 Cells , Chlorocebus aethiops , Dose-Response Relationship, Drug , Drug Evaluation, Preclinical/methods , Ferrous Compounds/pharmacology , Humans , Levulinic Acids/administration & dosage , Vero Cells
5.
Cell Struct Funct ; 2020 Nov 13.
Article in English | MEDLINE | ID: mdl-33191384

ABSTRACT

The smallest arenavirus gene product, Z protein, plays critical roles in the virus life cycle. Z is the major driving force of budding and particle production because of a unique property that defines self-assembly. In addition to the roles in budding, Z also participates in the suppression of type I interferon production to evade host antiviral immunity. Therefore, Z and its assembled form are an attractive drug target for arenaviral hemorrhagic fever, such as Lassa fever. Here, we developed a biosensor that enabled the evaluation of the prototype arenavirus, lymphocytic choriomeningitis virus (LCMV), Z assembly using the principle of Förster resonance energy transfer (FRET). This FRET biosensor consisted of three tandem Z that were sandwiched between super-enhanced cyan-emitting fluorescent protein and variant of a yellow-emitting mutant of green fluorescent protein so that Z-Z intermolecular binding via the really interesting new gene finger domain increased the emission ratio. To identify novel anti-arenavirus compounds, the FRET biosensor was employed to screen the PathogenBox400 for inhibitors of Z assembly in a 96-well plate format. The assay performed well, with a Z'-factor of 0.89, and identified two compounds that decreased the emission ratio of the FRET biosensor in a dose-dependent manner. Of them, the compound, 5,6,7,8-tetrahydro-7-(benzyl) -pyrido[4',3':4,5]thieno[2,3-d]pyrimidin-2,4-diamine, was found to significantly inhibit LCMV propagation in infected cells. Thereby, the present study demonstrated that a novel FRET biosensor incorporating Z assembly built on FRET and named Zabton, was a valuable screening tool to identify anti-arenavirus compounds in the context of inhibition of Z assembly.Key words: Arenavirus, Förster resonance energy transfer, anti-viral drugs, Z protein.

6.
PLoS Negl Trop Dis ; 14(11): e0008855, 2020 11.
Article in English | MEDLINE | ID: mdl-33147214

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly pathogenic novel coronavirus that has caused a worldwide outbreak. Here we describe a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay that uses a portable device for efficient detection of SARS-CoV-2. This RT-LAMP assay specifically detected SARS-CoV-2 without cross-reacting with the most closely related human coronavirus, SARS-CoV. Clinical evaluation of nasal swab samples from suspected SARS-CoV-2 pneumonia (COVID-19) patients showed that the assay could detect over 23.7 copies within 15 min with a 100% probability. Since the RT-LAMP assay can be performed with a portable battery-supported device, it is a rapid, simple, and sensitive diagnostic assay for COVID-19 that can be available at point-of-care. We also developed the RT-LAMP assay without the RNA extraction step-Direct RT-LAMP, which could detect more than 1.43 x 103 copies within 15 min with a 100% probability in clinical evaluation test. Although the Direct RT-LAMP assay was less sensitive than the standard RT-LAMP, the Direct RT-LAMP assay can be available as the rapid first screening of COVID-19 in poorly equipped areas, such as rural areas in developing countries.


Subject(s)
Betacoronavirus/genetics , Coronavirus Infections/diagnosis , Molecular Diagnostic Techniques/methods , Nucleic Acid Amplification Techniques/methods , Pneumonia, Viral/diagnosis , Humans , Pandemics , Point-of-Care Systems , Time Factors
7.
Front Microbiol ; 11: 562814, 2020.
Article in English | MEDLINE | ID: mdl-33117310

ABSTRACT

Several arenaviruses are highly pathogenic to humans, causing hemorrhagic fever. Discovery of anti-arenavirus drug candidates is urgently needed, although the molecular basis of the host- and organ-specific pathogenicity remains to be fully elucidated. The arenavirus Z protein facilitates production of virus-like particles (VLPs), providing an established method to assess virus budding. In this study, we examined the efficiency of VLP production by solely expressing Z protein of several different arenaviruses. In addition, we analyzed the role of the late (L)-domain of the arenavirus Z protein, which is essential for the interaction with ESCRT proteins, in VLP production among different cell lines. VLP assay was performed using Z proteins of Junín virus (JUNV), Machupo virus (MACV), Tacaribe virus (TCRV), Latino virus (LATV), Pichinde virus (PICV), and Lassa virus (LASV) in six different cell lines: HEK293T, Huh-7, A549, Vero76, BHK-21, and NIH3T3 cells. JUNV, MACV, and LASV Z proteins efficiently produced VLPs in all tested cell lines, while the efficiencies of VLP production by the other arenavirus Z proteins were cell type-dependent. The contribution of the L-domain(s) within Z protein to VLP production also highly depended on the cell type. These results suggested that each arenavirus has its own particle-production mechanism, which is different among the cell types.

8.
J Viral Hepat ; 27(11): 1234-1242, 2020 Nov.
Article in English | MEDLINE | ID: mdl-32564517

ABSTRACT

Although a high seroprevalence of antibodies against hepatitis A virus (HAV) has been estimated in Central Africa, the current status of both HAV infections and seroprevalence of anti-HAV antibodies remains unclear due to a paucity of surveillance data available. We conducted a serological survey during 2015-2017 in Gabon, Central Africa, and confirmed a high seroprevalence of anti-HAV antibodies in all age groups. To identify the currently circulating HAV strains and to reveal the epidemiological and genetic characteristics of the virus, we conducted molecular surveillance in a total of 1007 patients presenting febrile illness. Through HAV detection and sequencing, we identified subgenotype IIA (HAV-IIA) infections in the country throughout the year. A significant prevalence trend emerged in the young child population, presenting several infection peaks which appeared to be unrelated to dry or rainy seasons. Whole-genome sequencing and phylogenetic analyses revealed local HAV-IIA evolutionary events in Central Africa, indicating the circulation of HAV-IIA strains of a region-specific lineage. Recombination analysis of complete genome sequences revealed potential recombination events in Gabonese HAV strains. Interestingly, Gabonese HAV-IIA possibly acquired the 5'-untranslated region (5'-UTR) of the rare subgenotype HAV-IIB in recent years, suggesting the present existence of HAV-IIB in Central Africa. These findings indicate a currently stable HAV-IIA circulation in Gabon, with a high risk of infections in children aged under 5 years. Our findings will enhance the understanding of the current status of HAV infections in Central Africa and provide new insight into the molecular epidemiology and evolution of HAV genotype II.

9.
J Gen Virol ; 101(6): 573-586, 2020 06.
Article in English | MEDLINE | ID: mdl-32375950

ABSTRACT

Bone marrow stromal cell antigen-2 (BST-2), also known as tetherin, is an interferon-inducible membrane-associated protein. It effectively targets enveloped viruses at the release step of progeny viruses from host cells, thereby restricting the further spread of viral infection. Junin virus (JUNV) is a member of Arenaviridae, which causes Argentine haemorrhagic fever that is associated with a high rate of mortality. In this study, we examined the effect of human BST-2 on the replication and propagation of JUNV. The production of JUNV Z-mediated virus-like particles (VLPs) was significantly inhibited by over-expression of BST-2. Electron microscopy analysis revealed that BST-2 functions by forming a physical link that directly retains VLPs on the cell surface. Infection using JUNV showed that infectious JUNV production was moderately inhibited by endogenous or exogenous BST-2. We also observed that JUNV infection triggers an intense interferon response, causing an upregulation of BST-2, in infected cells. However, the expression of cell surface BST-2 was reduced upon infection. Furthermore, the expression of JUNV nucleoprotein (NP) partially recovered VLP production from BST-2 restriction, suggesting that the NP functions as an antagonist against antiviral effect of BST-2. We further showed that JUNV NP also rescued the production of Ebola virus VP40-mediated VLP from BST-2 restriction as a broad spectrum BST-2 antagonist. To our knowledge, this is the first report showing that an arenavirus protein counteracts the antiviral function of BST-2.


Subject(s)
Antigens, CD/metabolism , Host-Pathogen Interactions/physiology , Junin virus/physiology , Nucleoproteins/metabolism , Viral Core Proteins/metabolism , Virus Release/physiology , A549 Cells , Antiviral Agents/pharmacology , Cell Line , Cell Line, Tumor , GPI-Linked Proteins/metabolism , HEK293 Cells , HeLa Cells , Host-Pathogen Interactions/drug effects , Humans , Interferons/pharmacology , Junin virus/drug effects , Virus Release/drug effects , Virus Replication/drug effects , Virus Replication/genetics
10.
Virus Res ; 278: 197868, 2020 03.
Article in English | MEDLINE | ID: mdl-31962066

ABSTRACT

Recent reports have shown that rat hepatitis E virus (HEV) is capable of infecting humans. We also successfully propagated rat HEV into human PLC/PRF/5 cells, raising the possibility of a similar mechanism shared by human HEV and rat HEV. Rat HEV has the proline-rich sequence, PxYPMP, in the open reading frame 3 (ORF3) protein that is indispensable for its release. However, the release mechanism remains unclear. The overexpression of dominant-negative (DN) mutant of vacuolar protein sorting (Vps)4A or Vps4B decreased rat HEV release to 23.9 % and 18.0 %, respectively. The release of rat HEV was decreased to 8.3 % in tumor susceptibility gene 101 (Tsg101)-depleted cells and to 31.5 % in apoptosis-linked gene 2-interacting protein X (Alix)-depleted cells. Although rat HEV ORF3 protein did not bind to Tsg101, we found a 90-kDa protein capable of binding to wild-type rat HEV ORF3 protein but not to ORF3 mutant with proline to leucine mutations in the PxYPMP motif. Rat HEV release was also decreased in Ras-associated binding 27A (Rab27A)- or hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs)-depleted cells (to 20.1 % and 18.5 %, respectively). In addition, the extracellular rat HEV levels in the infected PLC/PRF/5 cells were increased after treatment with Bafilomycin A1 and decreased after treatment with GW4869. These results indicate that rat HEV utilizes multivesicular body (MVB) sorting for its release and that the exosomal pathway is required for rat HEV egress. A host protein alternative to Tsg101 that can bind to rat HEV ORF3 should be explored in further study.

11.
Int J Infect Dis ; 91: 129-136, 2020 Feb.
Article in English | MEDLINE | ID: mdl-31821892

ABSTRACT

OBJECTIVES: Dengue outbreaks, mainly caused by dengue virus serotype 2 (DENV-2), occurred in 2007 and in 2010 in Gabon, Central Africa. However, information on DENV infections has been insufficient since 2010. The aim of this study was to investigate the current DENV infection scenario and the risk of repeated infections in Gabon. METHODS: During 2015-2017, serum samples were collected from enrolled febrile participants and were tested for DENV infection using RT-qPCR. DENV-positive samples were analyzed for a history of previous DENV infection(s) using ELISA. The complete DENV genome was sequenced to analyze the phylogeny of Gabonese DENV strains. RESULTS: DENV-3 was exclusively detected, with a high rate of anti-DENV IgG seropositivity among DENV-3-positive participants. DENV-3 showed higher infection rates in adults and the infection was seasonal with peaks in the rainy seasons. Phylogenetic analysis revealed that Gabonese DENV-3 originated from West African strains and has been circulating continuously in Gabon since at least 2010, when the first DENV-3 case was reported. CONCLUSIONS: These findings indicate stable DENV-3 circulation and the risk of repeated DENV infections in Gabon, highlighting the need for continuous monitoring to control DENV infections.


Subject(s)
Dengue Virus/isolation & purification , Dengue/epidemiology , Adolescent , Adult , Aged , Aged, 80 and over , Child , Child, Preschool , Dengue/virology , Dengue Virus/classification , Dengue Virus/genetics , Female , Gabon/epidemiology , Humans , Infant , Male , Middle Aged , Phylogeny , Real-Time Polymerase Chain Reaction , Seasons , Seroepidemiologic Studies , Serogroup , Young Adult
12.
J Med Virol ; 92(2): 251-256, 2020 02.
Article in English | MEDLINE | ID: mdl-31538666

ABSTRACT

Hepatitis B virus (HBV) infection remains to be a major public health issue worldwide, although there is currently a safe vaccine and effective antiviral treatments. In surveillance of infectious diseases in Gabon, HBV viremia was detected in patients with febrile. Whole-genome sequencing was conducted to characterize the HBV strains currently circulating in Gabon and to investigate HBV genome diversity during viremia. Phylogenetic analysis revealed the existence of former subgenotype A5, which exhibits a particular pattern of distribution from several West and Central African countries to Haiti. Furthermore, sequencing analysis identified two similar HBV strains mixed in one sample, and a very rare 1-base pair insertion in the viral precore region. This insertion caused a frameshift mutation, indicating the production of an aberrant fusion protein of the HBV x and e antigens. Our data showed that the detected HBV strain was possibly in an "evolving" state during viremia, a phase of active replication.

13.
J Gen Virol ; 100(7): 1099-1111, 2019 07.
Article in English | MEDLINE | ID: mdl-31184566

ABSTRACT

Ebola virus (EBOV) VP40 is a major driving force of nascent virion production and a negative regulator of genome replication/transcription. Here, we showed that the YIGL sequence at the C-terminus of EBOV VP40 is important for virus-like particle (VLP) production and the regulation of genome replication/transcription. Accordingly, a mutation in the YIGL sequence caused defects in VLP production and genome replication/transcription. The residues I293 and L295 in the YIGL sequence were particularly critical for VLP production. Furthermore, an in silico analysis indicated that the amino acids surrounding the YIGL sequence contribute to intramolecular interactions within VP40. Among those surrounding residues, F209 was shown to be critical for VLP production. These results suggested that the VP40 YIGL sequence regulates two different viral replication steps, VLP production and genome replication/transcription, and the nearby residue F209 influences VLP production.


Subject(s)
Ebolavirus/physiology , Hemorrhagic Fever, Ebola/virology , Viral Matrix Proteins/chemistry , Viral Matrix Proteins/metabolism , Virion/physiology , Virus Replication , Amino Acid Motifs , Amino Acid Sequence , Ebolavirus/chemistry , Ebolavirus/genetics , Genome, Viral , Humans , Sequence Alignment , Viral Matrix Proteins/genetics , Virion/chemistry , Virion/genetics , Virus Release
14.
J Virol Methods ; 269: 30-37, 2019 07.
Article in English | MEDLINE | ID: mdl-30974179

ABSTRACT

Lassa virus (LASV) causes Lassa fever (LF), a viral hemorrhagic fever endemic in West Africa. LASV strains are clustered into six lineages according to their geographic location. To confirm a diagnosis of LF, a laboratory test is required. Here, a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay using a portable device for the detection of LASV in southeast and south-central Nigeria using three primer sets specific for strains clustered in lineage II was developed. The assay detected in vitro transcribed LASV RNAs within 23 min and was further evaluated for detection in 73 plasma collected from suspected LF patients admitted into two health settings in southern Nigeria. The clinical evaluation using the conventional RT-PCR as the reference test revealed a sensitivity of 50% in general with 100% for samples with a viral titer of 9500 genome equivalent copies (geq)/mL and higher. The detection limit was estimated to be 4214 geq/mL. The assay showed 98% specificity with no cross-reactivity to other viruses which cause similar symptoms. These results suggest that this RT-LAMP assay is a useful molecular diagnostic test for LF during the acute phase, contributing to early patient management, while using a convenient device for field deployment and in resource-poor settings.


Subject(s)
Lassa Fever/diagnosis , Lassa virus/isolation & purification , Molecular Diagnostic Techniques/methods , Nucleic Acid Amplification Techniques/methods , Reverse Transcription , DNA Primers/genetics , Genome, Viral , Humans , Lassa Fever/blood , Limit of Detection , Nigeria , Nucleic Acid Amplification Techniques/instrumentation , RNA, Viral/genetics , Sensitivity and Specificity , Temperature , Viral Load
15.
J Virol ; 93(10)2019 05 15.
Article in English | MEDLINE | ID: mdl-30814285

ABSTRACT

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a novel emerging virus that has been identified in China, South Korea, and Japan, and it induces thrombocytopenia and leukocytopenia in humans with a high case fatality rate. SFTSV is pathogenic to humans, while immunocompetent adult mice and golden Syrian hamsters infected with SFTSV never show apparent symptoms. However, mice deficient for the gene encoding the α chain of the alpha- and beta-interferon receptor (Ifnar1-/- mice) and golden Syrian hamsters deficient for the gene encoding signal transducer and activator of transcription 2 (Stat2-/- hamsters) are highly susceptible to SFTSV infection, with infection resulting in death. The nonstructural protein (NSs) of SFTSV has been reported to inhibit the type I IFN response through sequestration of human STAT proteins. Here, we demonstrated that SFTSV induces lethal acute disease in STAT2-deficient mice but not in STAT1-deficient mice. Furthermore, we discovered that NSs cannot inhibit type I IFN signaling in murine cells due to an inability to bind to murine STAT2. Taken together, our results imply that the dysfunction of NSs in antagonizing murine STAT2 can lead to inefficient replication and the loss of pathogenesis of SFTSV in mice.IMPORTANCE Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by SFTSV, which has been reported in China, South Korea, and Japan. Here, we revealed that mice lacking STAT2, which is an important factor for antiviral innate immunity, are highly susceptible to SFTSV infection. We also show that SFTSV NSs cannot exert its anti-innate immunity activity in mice due to the inability of the protein to bind to murine STAT2. Our findings suggest that the dysfunction of SFTSV NSs as an IFN antagonist in murine cells confers a loss of pathogenicity of SFTSV in mice.


Subject(s)
Bunyaviridae Infections/metabolism , Phlebovirus/metabolism , STAT2 Transcription Factor/metabolism , Animals , Antiviral Agents/metabolism , Bunyaviridae Infections/virology , Glycoproteins/metabolism , HEK293 Cells , Host Microbial Interactions/genetics , Host Microbial Interactions/physiology , Humans , Immunity, Innate/immunology , Mice , Mice, Inbred C57BL , Mice, Knockout , Phlebotomus Fever/virology , Phlebovirus/pathogenicity , Phosphorylation , Receptor, Interferon alpha-beta/metabolism , Signal Transduction/physiology , Species Specificity , Thrombocytopenia/metabolism , Viral Nonstructural Proteins/metabolism , Virulence
16.
PLoS Negl Trop Dis ; 12(11): e0006971, 2018 11.
Article in English | MEDLINE | ID: mdl-30500827

ABSTRACT

Lassa virus (LASV) is endemic in parts of West Africa where it causes Lassa fever (LF), a viral hemorrhagic fever with frequent fatal outcomes. The diverse LASV strains are grouped into six major lineages based on the geographical location of the isolated strains. In this study, we have focused on the lineage II strains from southern Nigeria. We determined the viral sequences from positive cases of LF reported at tertiary hospitals in Ebonyi and Enugu between 2012 and 2016. Reverse transcription-polymerase chain reaction (RT-PCR) showed that 29 out of 123 suspected cases were positive for the virus among which 11 viral gene sequences were determined. Phylogenetic analysis of the complete coding sequences of the four viral proteins revealed that lineage II strains are broadly divided into two genetic clades that diverged from a common ancestor 195 years ago. One clade, consisting of strains from Ebonyi and Enugu, was more conserved than the other from Irrua, although the four viral proteins were evolving at similar rates in both clades. These results suggested that the viruses of these clades have been distinctively evolving in geographically separate parts of southern Nigeria. Furthermore, the epidemiological data of the 2014 outbreak highlighted the role of human-to-human transmission in this outbreak, which was supported by phylogenetic analysis showing that 13 of the 16 sequences clustered together. These results provide new insights into the evolution of LASV in southern Nigeria and have important implications for vaccine development, diagnostic assay design, and LF outbreak management.


Subject(s)
Lassa Fever/virology , Lassa virus/genetics , Lassa virus/isolation & purification , Evolution, Molecular , Genetic Variation , Humans , Lassa Fever/epidemiology , Lassa virus/classification , Nigeria/epidemiology , Phylogeny , Viral Proteins/genetics
17.
J Am Chem Soc ; 140(48): 16834-16841, 2018 12 05.
Article in English | MEDLINE | ID: mdl-30475615

ABSTRACT

Immunosensing is a bioanalytical technique capable of selective detections of pathogens by utilizing highly specific and strong intermolecular interactions between recognition probes and antigens. Here, we exploited the molecular mechanism in artificial nanopores for selective single-virus identifications. We designed hemagglutinin antibody mimicking oligopeptides with a weak affinity to influenza A virus. By functionalizing the pore wall surface with the synthetic peptides, we rendered specificity to virion-nanopore interactions. The ligand binding thereof was found to perturb translocation dynamics of specific viruses in the nanochannel, which facilitated digital typing of influenza by the resistive pulse bluntness. As amino acid sequence degrees of freedom can potentially offer variety of recognition ability to the molecular probes, this peptide nanopore approach can be used as a versatile immunosensor with single-particle sensitivity that promises wide applications in bioanalysis including bacterial and viral screening to infectious disease diagnosis.


Subject(s)
Influenza A Virus, H1N1 Subtype/isolation & purification , Nanopores , Animals , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/immunology , Chickens , Gold/chemistry , Humans , Influenza A Virus, H1N1 Subtype/immunology , Peptide Fragments/chemistry , Peptide Fragments/immunology , Silicon Compounds/chemistry , Viral Load/methods
18.
PLoS Pathog ; 14(7): e1007172, 2018 07.
Article in English | MEDLINE | ID: mdl-30028868

ABSTRACT

The interferon inducible protein, BST-2 (or, tetherin), plays an important role in the innate antiviral defense system by inhibiting the release of many enveloped viruses. Consequently, viruses have evolved strategies to counteract the anti-viral activity of this protein. While the mechanisms by which BST-2 prevents viral dissemination have been defined, less is known about how this protein shapes the early viral distribution and immunological defense against pathogens during the establishment of persistence. Using the lymphocytic choriomeningitis virus (LCMV) model of infection, we sought insights into how the in vitro antiviral activity of this protein compared to the immunological defense mounted in vivo. We observed that BST-2 modestly reduced production of virion particles from cultured cells, which was associated with the ability of BST-2 to interfere with the virus budding process mediated by the LCMV Z protein. Moreover, LCMV does not encode a BST-2 antagonist, and viral propagation was not significantly restricted in cells that constitutively expressed BST-2. In contrast to this very modest effect in cultured cells, BST-2 played a crucial role in controlling LCMV in vivo. In BST-2 deficient mice, a persistent strain of LCMV was no longer confined to the splenic marginal zone at early times post-infection, which resulted in an altered distribution of LCMV-specific T cells, reduced T cell proliferation / function, delayed viral control in the serum, and persistence in the brain. These data demonstrate that BST-2 is important in shaping the anatomical distribution and adaptive immune response against a persistent viral infection in vivo.


Subject(s)
Antigens, CD/immunology , Lymphocytic Choriomeningitis/immunology , T-Lymphocytes/immunology , Animals , Antigens, CD/metabolism , Cell Proliferation , GPI-Linked Proteins/immunology , GPI-Linked Proteins/metabolism , Humans , Lymphocyte Activation , Lymphocytic Choriomeningitis/metabolism , Lymphocytic choriomeningitis virus/immunology , Mice , Mice, Inbred C57BL
19.
Biochem Biophys Res Commun ; 503(2): 631-636, 2018 09 05.
Article in English | MEDLINE | ID: mdl-29906459

ABSTRACT

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by the SFTS virus (SFTSV), which has a high mortality rate. Currently, no licensed vaccines or therapeutic agents have been approved for use against SFTSV infection. Here, we report that the cholesterol, fatty acid, and triglyceride synthesis pathways regulated by S1P is involved in SFTSV replication, using CHO-K1 cell line (SRD-12B) that is deficient in site 1 protease (S1P) enzymatic activity, PF-429242, a small compound targeting S1P enzymatic activity, and Fenofibrate and Lovastatin, which inhibit triglyceride and cholesterol synthesis, respectively. These results enhance our understanding of the SFTSV replication mechanism and may contribute to the development of novel therapies for SFTSV infection.


Subject(s)
Cholesterol/metabolism , Fatty Acids/metabolism , Phlebotomus Fever/metabolism , Phlebovirus/physiology , Proprotein Convertases/metabolism , Serine Endopeptidases/metabolism , Triglycerides/metabolism , Virus Replication , Animals , Biosynthetic Pathways , CHO Cells , Cell Line , Cricetulus , Humans , Phlebotomus Fever/enzymology
20.
J Gen Virol ; 99(2): 181-186, 2018 02.
Article in English | MEDLINE | ID: mdl-29300152

ABSTRACT

Ebola virus (EBOV), which belongs to the genus Ebolavirus, causes a severe and often fatal infection in primates, including humans, whereas Reston virus (RESTV) only causes lethal disease in non-human primates. Two amino acids (aa) at positions 82 and 544 of the EBOV glycoprotein (GP) are involved in determining viral infectivity. However, it remains unclear how these two aa residues affect the infectivity of Ebolavirus species in various hosts. Here we performed viral pseudotyping experiments with EBOV and RESTV GP derivatives in 10 cell lines from 9 mammalian species. We demonstrated that isoleucine at position 544/545 increases viral infectivity in all host species, whereas valine at position 82/83 modulates viral infectivity, depending on the viral and host species. Structural modelling suggested that the former residue affects viral fusion, whereas the latter residue influences the interaction with the viral entry receptor, Niemann-Pick C1.


Subject(s)
Ebolavirus/genetics , Glycoproteins/genetics , Hemorrhagic Fever, Ebola/virology , Host-Pathogen Interactions , Models, Structural , Niemann-Pick C1 Protein/metabolism , Amino Acid Substitution , Animals , Cell Line , Ebolavirus/pathogenicity , Glycoproteins/metabolism , Humans , Mammals , Mutation , Niemann-Pick C1 Protein/genetics , Primates , Sequence Alignment , Viral Fusion Proteins/genetics , Viral Fusion Proteins/metabolism , Viral Proteins/genetics , Viral Proteins/metabolism
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