An immunoinformatics-derived DNA vaccine encoding human class II T cell epitopes of Ebola virus, Sudan virus, and Venezuelan equine encephalitis virus is immunogenic in HLA transgenic mice

Immunoinformatics tools were used to predict human leukocyte antigen (HLA) class II-restricted T cell epitopes within the envelope glycoproteins and nucleocapsid proteins of Ebola virus (EBOV) and Sudan virus (SUDV) and the structural proteins of Venezuelan equine encephalitis virus (VEEV). Selected epitopes were tested for binding to soluble HLA molecules representing 5 class II alleles (DRB1*0101, DRB1*0301, DRB1*0401, DRB1*0701, and DRB1*1501). All but one of the 25 tested peptides bound to at least one of the DRB1 alleles, and 4 of the peptides bound at least moderately or weakly to all 5 DRB1 alleles. Additional algorithms were used to design a single “string-of-beads” expression construct with 44 selected epitopes arranged to avoid creation of spurious junctional epitopes. Seventeen of these 44 predicted epitopes were conserved between the major histocompatibility complex (MHC) of humans and mice, allowing initial testing in mice. BALB/c mice vaccinated with the multi-epitope construct developed statistically significant cellular immune responses to EBOV, SUDV, and VEEV peptides as measured by interferon (IFN)-γ ELISpot assays. Significant levels of antibodies to VEEV, but not EBOV, were also detected in vaccinated BALB/c mice. To assess immunogenicity in the context of a human MHC, HLA-DR3 transgenic mice were vaccinated with the multi-epitope construct and boosted with a mixture of the 25 peptides used in the binding assays. The vaccinated HLA-DR3 mice developed significant cellular immune responses to 4 of the 25 (16%) tested individual class II peptides as measured by IFN-γ ELISpot assays. In addition, these mice developed antibodies against EBOV and VEEV as measured by ELISA. While a low but significant level of protection was observed in vaccinated transgenic mice after aerosol exposure to VEEV, no protection was observed after intraperitoneal challenge with mouse-adapted EBOV. These studies provide proof of concept for the use of an informatics approach to design a multi-agent, multi-epitope immunogen and provide a basis for further testing aimed at focusing immune responses toward desired protective T cell epitopes

Construction and Nonclinical Testing of a Puumala Virus Synthetic M Gene-Based DNA Vaccine

Puumala virus (PUUV) is a causative agent of hemorrhagic fever with renal syndrome (HFRS). Although PUUV-associated HFRS does not result in high case-fatality rates, the social and economic impact is considerable. There is no licensed vaccine or specific therapeutic to prevent or treat HFRS. Here we report the synthesis of a codon-optimized, full-lengthMsegment open reading frame and its cloning into a DNA vaccine vector to produce the plasmid pWRG/PUU-M(s2). pWRG/PUU-M(s2) delivered by gene gun produced high-titer neutralizing antibodies in hamsters and nonhuman primates. Vaccination with pWRG/ PUU-M(s2) protected hamsters against infection with PUUV but not against infection by related HFRS-associated hantaviruses. Unexpectedly, vaccination protected hamsters in a lethal disease model of Andes virus (ANDV) in the absence of ANDV crossneutralizing antibodies. This is the first evidence that an experimental DNA vaccine for HFRS can provide protection in a hantavirus lethal disease model

Construction and Nonclinical Testing of a Puumala Virus Synthetic M Gene-Based DNA Vaccine

Puumala virus (PUUV) is a causative agent of hemorrhagic fever with renal syndrome (HFRS). Although PUUV-associated HFRS does not result in high case-fatality rates, the social and economic impact is considerable. There is no licensed vaccine or specific therapeutic to prevent or treat HFRS. Here we report the synthesis of a codon-optimized, full-lengthMsegment open reading frame and its cloning into a DNA vaccine vector to produce the plasmid pWRG/PUU-M(s2). pWRG/PUU-M(s2) delivered by gene gun produced high-titer neutralizing antibodies in hamsters and nonhuman primates. Vaccination with pWRG/ PUU-M(s2) protected hamsters against infection with PUUV but not against infection by related HFRS-associated hantaviruses. Unexpectedly, vaccination protected hamsters in a lethal disease model of Andes virus (ANDV) in the absence of ANDV crossneutralizing antibodies. This is the first evidence that an experimental DNA vaccine for HFRS can provide protection in a hantavirus lethal disease model

The Rift Valley fever accessory proteins NSm and P78/NSm-GN are distinct determinants of virus propagation in vertebrate and invertebrate hosts.: Role of NSm-related proteins in RVFV infection

International audienceRift Valley fever virus (RVFV) is an enzootic virus circulating in Africa that is transmitted to its vertebrate host by a mosquito vector and causes severe clinical manifestations in humans and ruminants. RVFV has a tripartite genome of negative or ambisense polarity. The M segment contains five in-frame AUG codons that are alternatively used for the synthesis of two major structural glycoproteins, GN and GC, and at least two accessory proteins, NSm, a 14-kDa cytosolic protein, and P78/NSm-GN, a 78-kDa glycoprotein. To determine the relative contribution of P78 and NSm to RVFV infectivity, AUG codons were knocked out to generate mutant viruses expressing various sets of the M-encoded proteins. We found that, in the absence of the second AUG codon used to express NSm, a 13-kDa protein corresponding to an N-terminally truncated form of NSm, named NSm', was synthesized from AUG 3. None of the individual accessory proteins had any significant impact on RVFV virulence in mice. However, a mutant virus lacking both NSm and NSm' was strongly attenuated in mice and grew to reduced titers in murine macrophages, a major target cell type of RVFV. In contrast, P78 was not associated with reduced viral virulence in mice, yet it appeared as a major determinant of virus dissemination in mosquitoes. This study demonstrates how related accessory proteins differentially contribute to RVFV propagation in mammalian and arthropod hosts

Temporal trend and climate factors of hemorrhagic fever with renal syndrome epidemic in Shenyang City, China

<p>Abstract</p> <p>Background</p> <p>Hemorrhagic fever with renal syndrome (HFRS) is an important infectious disease caused by different species of hantaviruses. As a rodent-borne disease with a seasonal distribution, external environmental factors including climate factors may play a significant role in its transmission. The city of Shenyang is one of the most seriously endemic areas for HFRS. Here, we characterized the dynamic temporal trend of HFRS, and identified climate-related risk factors and their roles in HFRS transmission in Shenyang, China.</p> <p>Methods</p> <p>The annual and monthly cumulative numbers of HFRS cases from 2004 to 2009 were calculated and plotted to show the annual and seasonal fluctuation in Shenyang. Cross-correlation and autocorrelation analyses were performed to detect the lagged effect of climate factors on HFRS transmission and the autocorrelation of monthly HFRS cases. Principal component analysis was constructed by using climate data from 2004 to 2009 to extract principal components of climate factors to reduce co-linearity. The extracted principal components and autocorrelation terms of monthly HFRS cases were added into a multiple regression model called principal components regression model (PCR) to quantify the relationship between climate factors, autocorrelation terms and transmission of HFRS. The PCR model was compared to a general multiple regression model conducted only with climate factors as independent variables.</p> <p>Results</p> <p>A distinctly declining temporal trend of annual HFRS incidence was identified. HFRS cases were reported every month, and the two peak periods occurred in spring (March to May) and winter (November to January), during which, nearly 75% of the HFRS cases were reported. Three principal components were extracted with a cumulative contribution rate of 86.06%. Component 1 represented MinRH₀, MT₁, RH₁, and MWV₁; component 2 represented RH₂, MaxT₃, and MAP₃; and component 3 represented MaxT₂, MAP₂, and MWV₂. The PCR model was composed of three principal components and two autocorrelation terms. The association between HFRS epidemics and climate factors was better explained in the PCR model (<it>F </it>= 446.452, <it>P </it>< 0.001, adjusted <it>R</it>²= 0.75) than in the general multiple regression model (<it>F </it>= 223.670, <it>P </it>< 0.000, adjusted <it>R</it>²= 0.51).</p> <p>Conclusion</p> <p>The temporal distribution of HFRS in Shenyang varied in different years with a distinctly declining trend. The monthly trends of HFRS were significantly associated with local temperature, relative humidity, precipitation, air pressure, and wind velocity of the different previous months. The model conducted in this study will make HFRS surveillance simpler and the control of HFRS more targeted in Shenyang.</p

Development of a highly protective combination monoclonal antibody therapy against Chikungunya virus

Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes global epidemics of a debilitating polyarthritis in humans. As there is a pressing need for the development of therapeutic agents, we screened 230 new mouse anti-CHIKV monoclonal antibodies (MAbs) for their ability to inhibit infection of all three CHIKV genotypes. Four of 36 neutralizing MAbs (CHK-102, CHK-152, CHK-166, and CHK-263) provided complete protection against lethality as prophylaxis in highly susceptible immunocompromised mice lacking the type I IFN receptor (Ifnar−/−) and mapped to distinct epitopes on the E1 and E2 structural proteins. CHK-152, the most protective MAb, was humanized, shown to block viral fusion, and require Fc effector function for optimal activity in vivo. In post-exposure therapeutic trials, administration of a single dose of a combination of two neutralizing MAbs (CHK-102+CHK-152 or CHK-166+CHK-152) limited the development of resistance and protected immunocompromised mice against disease when given 24 to 36 hours before CHIKV-induced death. Selected pairs of highly neutralizing MAbs may be a promising treatment option for CHIKV in humans

Milder winters in northern Scandinavia may contribute to larger outbreaks of haemorrhagic fever virus

The spread of zoonotic infectious diseases may increase due to climate factors such as temperature, humidity and precipitation. This is also true for hantaviruses, which are globally spread haemorrhagic fever viruses carried by rodents. Hantaviruses are frequently transmitted to humans all over the world and regarded as emerging viral diseases. Climate variations affect the rodent reservoir populations and rodent population peaks coincide with increased number of human cases of hantavirus infections. In northern Sweden, a form of haemorrhagic fever called nephropathia epidemica (NE), caused by the Puumala hantavirus (PUUV) is endemic and during 2006–2007 an unexpected, sudden and large outbreak of NE occurred in this region. The incidence was 313 cases/100,000 inhabitants in the most endemic areas, and from January through March 2007 the outbreak had a dramatic and sudden start with 474 cases in the endemic region alone. The PUUV rodent reservoir is bank voles and immediately before and during the peak of disease outbreak the affected regions experienced extreme climate conditions with a record-breaking warm winter, registering temperatures 6–9°C above normal. No protective snow cover was present before the outbreak and more bank voles than normal came in contact with humans inside or in close to human dwellings. These extreme climate conditions most probably affected the rodent reservoir and are important factors for the severity of the outbreak

The severity of Puumala hantavirus induced nephropathia epidemica can be better evaluated using plasma interleukin-6 than C-reactive protein determinations

<p>Abstract</p> <p>Background</p> <p>Nephropathia epidemica (NE) is a Scandinavian type of hemorrhagic fever with renal syndrome caused by Puumala hantavirus. The clinical course of the disease varies greatly in severity. The aim of the present study was to evaluate whether plasma C-reactive protein (CRP) and interleukin (IL)-6 levels associate with the severity of NE.</p> <p>Methods</p> <p>A prospectively collected cohort of 118 consecutive hospital-treated patients with acute serologically confirmed NE was examined. Plasma IL-6, CRP, and creatinine, as well as blood cell count and daily urinary protein excretion were measured on three consecutive days after admission. Plasma IL-6 and CRP levels higher than the median were considered high.</p> <p>Results</p> <p>We found that high IL-6 associated with most variables reflecting the severity of the disease. When compared to patients with low IL-6, patients with high IL-6 had higher maximum blood leukocyte count (11.9 <it>vs </it>9.0 × 10⁹/l, <it>P </it>= 0.001) and urinary protein excretion (2.51 <it>vs </it>1.68 g/day, <it>P </it>= 0.017), as well as a lower minimum blood platelet count (55 <it>vs </it>80 × 10⁹/l, <it>P </it>< 0.001), hematocrit (0.34 <it>vs </it>0.38, <it>P </it>= 0.001), and urinary output (1040 <it>vs </it>2180 ml/day, <it>P </it>< 0.001). They also stayed longer in hospital than patients with low IL-6 (8 <it>vs </it>6 days, <it>P </it>< 0.001). In contrast, high CRP did not associate with severe disease.</p> <p>Conclusions</p> <p>High plasma IL-6 concentrations associate with a clinically severe acute Puumala hantavirus infection, whereas high plasma CRP as such does not reflect the severity of the disease.</p