| 1. |
- Hepojoki, Satu, et al.
(författare)
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Competitive Homogeneous Immunoassay for Rapid Serodiagnosis of Hantavirus Disease
- 2015
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Ingår i: Journal of Clinical Microbiology. - 0095-1137 .- 1098-660X. ; 53:7, s. 2292-2297
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Tidskriftsartikel (refereegranskat)abstract
- In this study, we describe a competitive homogeneous immunoassay that makes use of Forster resonance energy transfer (FRET) in rapid detection of pathogen-specific antibodies. The assay principle is based on competition between a monoclonal antibody (MAb) and serum antibodies to a given antigen. In the assay, named competitive FRET immunoassay (CFRET-IA), the FRET signal is induced if MAb carrying a donor label binds to an acceptor-labeled antigen. Specific antibodies in serum compete for antigen binding, resulting in reduced FRET signal. The proof-of-principle for the assay was obtained using donor-labeled Puumala virus nucleocapsid protein (PUUV-N) and acceptor-labeled anti-PUUV-N MAb. The assay was evaluated by analyzing 329 clinical samples comprising 101 from individuals with acute PUUV infection, 42 from individuals with past infection, and 186 from individuals with PUUV-seronegative sera, and the results were compared to those of reference tests. The rapid serodiagnostic test we introduced herein performed with 100% sensitivity and 99% specificity for diagnosing acute hantavirus disease.
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| 2. |
- Iheozor-Ejiofor, Rommel, et al.
(författare)
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Neutralizing Antibody Titers in Hospitalized Patients with Acute Puumala Orthohantavirus Infection Do Not Associate with Disease Severity
- 2022
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Ingår i: Viruses. - : MDPI. - 1999-4915. ; 14:5
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Tidskriftsartikel (refereegranskat)abstract
- Nephropathia epidemica (NE), a mild form of haemorrhagic fever with renal syndrome (HFRS), is an acute febrile illness caused by Puumala orthohantavirus (PUUV). NE manifests typically with acute kidney injury (AKI), with a case fatality rate of about 0.1%. The treatment and management of hantavirus infections are mainly supportive, although neutralizing monoclonal antibodies and immune sera therapeutics are under investigation. In order to assess the potential use of antibody therapeutics in NE, we sought to determine the relationship between circulating PUUV neutralizing antibodies, PUUV nucleocapsid protein (N) IgG antibodies, and viral loads with markers of disease severity. The study included serum samples of extensively characterized patient cohorts (n = 116) from Tampere University Hospital, Finland. The results showed that upon hospitalization, most patients already had considerable neutralizing and anti-PUUV-N IgG antibody levels. However, contrary to expectations, neutralizing antibody titers from the first day of hospitalization did not appear to protect from AKI or correlate with more favorable disease outcomes. This indicates that further studies are needed to investigate the applicability of neutralizing antibodies as a therapy for hospitalized NE patients.
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| 3. |
- Iheozor-Ejiofor, Rommel Paneth, et al.
(författare)
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Vaccinia virus-free rescue of fluorescent replication-defective vesicular stomatitis virus and pseudotyping with Puumala virus glycoproteins for use in neutralization tests
- 2016
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Ingår i: Journal of General Virology. - : Microbiology Society. - 0022-1317 .- 1465-2099. ; 97, s. 1052-1059
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Tidskriftsartikel (refereegranskat)abstract
- Puumala virus (PUUV) grows slowly in cell culture. To study antigenic properties of PUUV, an amenable method for their expression would be beneficial. To achieve this, a replication-defective recombinant vesicular stomatitis virus, rVSV Delta G*EGFP, was rescued using BSRT7/5 and encephalomyocarditis virus (EMCV) internal ribosomal entry site (IRES)-enabled rescue plasmids. Using these particles, pseudotypes bearing PUUV Sotkamo strain glycoproteins were produced, with titres in the range 10(5)-10(8), and were used in pseudotype focus reduction neutralization tests (pFRNTs) with neutralizing monoclonal antibodies and patient sera. The results were compared with those from orthodox focus reduction neutralization tests (oFRNTs) using native PUUV with the same samples and showed a strong positive correlation (r(s)=0.82) between the methods. While developing the system we identified three amino acids which were mutated in the Vero E6 cell culture adapted PUUV prototype Sotkamo strain sequence, and changing these residues was critical for expression and neutralizing antibody binding of PUUV glycoproteins.
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| 4. |
- Rissanen, Ilona, et al.
(författare)
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Molecular rationale for antibody-mediated targeting of the hantavirus fusion glycoprotein
- 2020
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Ingår i: eLIFE. - : eLife Sciences Publications, Ltd. - 2050-084X. ; 9
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Tidskriftsartikel (refereegranskat)abstract
- The intricate lattice of Gn and Gc glycoprotein spike complexes on the hantavirus envelope facilitates host-cell entry and is the primary target of the neutralizing antibody-mediated immune response. Through study of a neutralizing monoclonal antibody termed mAb P-4G2, which neutralizes the zoonotic pathogen Puumala virus (PUUV), we provide a molecular-level basis for antibody-mediated targeting of the hantaviral glycoprotein lattice. Crystallographic analysis demonstrates that P-4G2 binds to a multi-domain site on PUUV Gc and may preclude fusogenic rearrangements of the glycoprotein that are required for host-cell entry. Furthermore, cryoelectron microscopy of PUUV-like particles in the presence of P-4G2 reveals a lattice-independent configuration of the Gc, demonstrating that P-4G2 perturbs the (Gn-Gc)(4) lattice. This work provides a structure-based blueprint for rationalizing antibody-mediated targeting of hantaviruses.
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| 5. |
- Strandin, Tomas, et al.
(författare)
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Interferons Induce STAT1-Dependent Expression of Tissue Plasminogen Activator, a Pathogenicity Factor in Puumala Hantavirus Disease
- 2016
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Ingår i: Journal of Infectious Diseases. - : Oxford University Press (OUP). - 0022-1899 .- 1537-6613. ; 213:10, s. 1632-1641
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Tidskriftsartikel (refereegranskat)abstract
- Hantaviruses are zoonotic viruses that show various degrees of vasculopathy in humans. In this study, we analyzed the regulation of 2 fibrinolytic parameters, tissue plasminogen activator (tPA) and its physiological inhibitor, plasminogen activator inhibitor 1 (PAI-1), in Puumala hantavirus (PUUV)-infected patients and in human microvascular endothelial cells. We detected strong upregulation of tPA in the acute phase of illness and in PUUV-infected macaques and found the tPA level to positively correlate with disease severity. The median levels of PAI-1 during the acute stage did not differ from those during the recovery phase. In concordance, hantaviruses induced tPA but not PAI-1 in microvascular endothelial cells, and the induction was demonstrated to be dependent on type I interferon. Importantly, type I and II interferons directly upregulated tPA through signal transducer and activator of transcription 1 (STAT1), which regulated tPA gene expression via a STAT1-responsive enhancer element. These results suggest that tPA may be a general factor in the immunological response to viruses.
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