Identification of T cell responses to the nonstructural glycoproteins in survivors of Crimean-Congo hemorrhagic fever in South Africa.

Crimean-Congo hemorrhagic fever orthonairovirus (CCHFV) is listed as a priority pathogen by the World Health Organization due to the severity of disease, propensity for spread to nonendemic regions, and absence of a vaccine or specific treatment. The immune correlates of protection are not clearly defined and hence the importance of investigating host immune responses in survivors. We have previously shown that survivors generate memory T cell responses that are long-lived and this study aimed to further define specific viral proteins targeted by the T cell response. The NSM , GP38, highly variable mucin-like domain, and N-terminus of GC regions in CCHFV are considered immunogenic regions and were investigated using peptide libraries representing regions of interest. An interferon gamma ELISpot assay was used to identify responses in peripheral blood mononuclear cells isolated from 12 survivors of laboratory confirmed CCHFV infections. IFN-γ responses were detected from eight survivors, against nine peptides, including four peptides located in the NSM region and five peptides located in the GP38 protein. No response was detected against peptides representing the mucin-like domain. In conclusion, the results suggest the presence of a long-lasting T cell memory response upon stimulation with viral epitopes in survivors of infection.

Chikungunya Virus Diagnosis: A Review of Current Antigen Detection Methods.

Chikungunya is a mosquito-borne viral disease caused by the chikungunya virus (CHIKV). CHIKV is expanding at an alarming rate, potentially spreading and establishing endemicity in new areas where competent vectors are present. The dramatic spread of CHIKV in recent years highlights the urgent need to take precautionary measures and investigate options for control. It is crucial in developing nations where diagnostic tools are limited, and symptoms are similar to other prevalent diseases such as malaria and dengue. The most reliable method for diagnosing chikungunya virus is viral gene detection by RT-PCR. Alternative methods like detecting human antibody and viral antigen can also be used, especially in areas where resources are limited. In this review, we summarize the limited data on antigen detection immunoassays. We further explain the essential structural elements of the virus to help comprehend the scientific concepts underlying the testing methods, as well as future methods and diagnostic approaches under investigation.

Perspective Technologies of Vaccination: Do We Still Need Old Vaccines?

Until December 2019, we were living in the world of successfully functioning vaccines and vaccination programs [...].

Rapid reverse transcriptase recombinase polymerase amplification assay for flaviviruses using non-infectious in vitro transcribed RNA as positive controls.

West Nile virus (WNV) and Wesselsbron virus (WSLV) are mosquito-borne viruses belonging to the Flavivirus genus, family Flaviviridae and cause outbreaks in southern Africa after heavy rain. Isothermal assays have been proposed for application in field situations as well as low resource settings and hence we developed a reverse-transcriptase recombinase polymerase amplification (RT-RPA) to detect WNV and WSLV known to occur in South Africa, causing sporadic outbreaks usually associated with good rainfall favouring mosquito breeding. Infectious virus can only be handled within a biosafety level (BSL) 3 facility, hence we opted to validate the assay with transcribed RNA. Specific RT-RPA primers and probes were designed for detection of WNV and WSLV and products detected using a rapid lateral flow device. The assay was performed in 30 min and detected 1.9 × 10¹ copies of WNV and 3.5 × 10° copies WSLV using noninfectious transcribed RNA controls. In addition, the assay was not inhibited by the presence of mosquito extracts in spiked samples. Mismatches between the WNV and WSLV probes and other flaviviruses will likely prevent cross reactivity. The sensitivity, low RPA incubation temperature and rapid processing time makes assay systems based on RPA technology ideally suited for fieldable diagnostics.

Immunogenicity of a DNA-Based Sindbis Replicon Expressing Crimean-Congo Hemorrhagic Fever Virus Nucleoprotein.

Crimean-Congo hemorrhagic fever virus (CCHFV) infrequently causes hemorrhagic fever in humans with a case fatality rate of 30%. Currently, there is neither an internationally approved antiviral drug nor a vaccine against the virus. A replicon based on the Sindbis virus vector encoding the complete open reading frame of a CCHFV nucleoprotein from a South African isolate was prepared and investigated as a possible candidate vaccine. The transcription of CCHFV RNA and recombinant protein production by the replicon were characterized in transfected baby hamster kidney cells. A replicon encoding CCHFV nucleoprotein inserted in plasmid DNA, pSinCCHF-52S, directed transcription of CCHFV RNA in the transfected cells. NIH-III heterozygous mice immunized with pSinCCHF-52S generated CCHFV IgG specific antibodies with notably higher levels of IgG2a compared to IgG1. Splenocytes from mice immunized with pSinCCHF-52S secreted IFN-γ and IL-2, low levels of IL-6 or IL-10, and no IL-4. No specific cytokine production was registered in splenocytes of mock-immunized mice (p < 0.05). Thus, our study demonstrated the expression of CCHFV nucleoprotein by a Sindbis virus vector and its immunogenicity in mice. The spectrum of cytokine production and antibody profile indicated predominantly Th1-type of an anti-CCHFV immune response. Further studies in CCHFV-susceptible animals are necessary to determine whether the induced immune response is protective.

Crimean-Congo Hemorrhagic Fever Virus: Advances in Vaccine Development.

Crimean-Congo hemorrhagic fever (CCHF) is a severe human disease with mortality rates of up to 30%. The disease is widespread in Africa, Asia, the Middle East and Eastern Europe. The last few years have seen disease emergence in Spain for the first time and disease re-emergence in other regions of the world after periods of inactivity. Factors, such as climate change, movement of infected ticks, animals, and changes in human activity, are likely to broaden endemic foci. There are therefore concerns that CCHF might emerge in currently nonendemic regions. The absence of approved vaccines or therapies heightens these concerns; thus Crimean-Congo hemorrhagic fever virus (CCHFV) is listed by the World Health Organization as a priority organism. However, the current sporadic nature of CCHF cases may call for targeted vaccination of risk groups as opposed to mass vaccinations. CCHF vaccine development has accelerated in recent years, partly because of the discovery of CCHF animal models. In this review, we discuss CCHF risk groups who are most likely to benefit from vaccine development, the merits and demerits of available CCHF animal models, and the various approaches which have been explored for CCHF vaccine development. Lastly, we present concluding remarks and research areas which can be further explored to enhance the available CCHFV vaccine data.

Epitope-mapping of the glycoprotein from Crimean-Congo hemorrhagic fever virus using a microarray approach.

Crimean-Congo hemorrhagic fever virus (CCHFV) causes severe acute human disease with lethal outcome. The knowledge about the immune response for this human health threat is highly limited. In this study, we have screened the glycoprotein of CCHFV for novel linear B-cell epitopic regions using a microarray approach. The peptide library consisted of 168 synthesized 20mer peptides with 10 amino acid overlap covering the entire glycoprotein. Using both pooled and individual human sera from survivors of CCHF disease in Turkey five peptide epitopes situated in the mucin-like region and GP 38 (G15-515) and GN G516-1037 region of the glycoprotein were identified as epitopes for a CCHF immune response. An epitope walk of the five peptides revealed a peptide sequence located in the GN region with high specificity and sensitivity. This peptide sequence, and a sequence downstream, reacted also against sera from survivors of CCHF disease in South Africa. The cross reactivity of these peptides with samples from a geographically distinct region where genetically diverse strains of the virus circulate, enabled the identification of unique peptide epitopes from the CCHF glycoprotein that could have application in development of diagnostic tools. In this study clinical samples from geographically distinct regions were used to identify conserved linear epitopic regions of the glycoprotein of CCHF.