Abstract
BackgroundVenezuelan equine encephalitis virus (VEEV) belongs to the Alphavirus group. Several species of this family are also pathogenic to humans and are recognized as potential agents of biological warfare and terrorism. The objective of this work was the generation of recombinant antibodies for the detection of VEEV after a potential bioterrorism assault or an natural outbreak of VEEV.ResultsIn this work, human anti-VEEV single chain Fragments variable (scFv) were isolated for the first time from a human naïve antibody gene library using optimized selection processes. In total eleven different scFvs were identified and their immunological specificity was assessed. The specific detection of the VEEV strains TC83, H12/93 and 230 by the selected antibody fragments was proved. Active as well as formalin inactivated virus particles were recognized by the selected antibody fragments which could be also used for Western blot analysis of VEEV proteins and immunohistochemistry of VEEV infected cells. The anti-VEEV scFv phage clones did not show any cross-reactivity with Alphavirus species of the Western equine encephalitis virus (WEEV) and Eastern equine encephalitis virus (EEEV) antigenic complex, nor did they react with Chikungunya virus (CHIKV), if they were used as detection reagent.ConclusionFor the first time, this study describes the selection of antibodies against a human pathogenic virus from a human naïve scFv antibody gene library using complete, active virus particles as antigen. The broad and sensitive applicability of scFv-presenting phage for the immunological detection and diagnosis of Alphavirus species was demonstrated. The selected antibody fragments will improve the fast identification of VEEV in case of a biological warfare or terroristic attack or a natural outbreak.
Highlights
Venezuelan equine encephalitis virus (VEEV) belongs to the Alphavirus group
All pannings were performed in a biosafety level 3 laboratory and the vaccine strain, TC83, as a medically important and epizootic Alphavirus species was used as antigen
This study showed that single chain Fragments variable (scFv) phage are applicable for a broad range of anti-VEEV diagnosis assays: antigen ELISA on purified virus particles, ELISA on cell lysate and immunoblot
Summary
Venezuelan equine encephalitis virus (VEEV) belongs to the Alphavirus group Several species of this family are pathogenic to humans and are recognized as potential agents of biological warfare and terrorism. Venezuelan equine encephalitis virus (VEEV) belongs to the Alphavirus genus within the Togaviridae family and was first isolated from horses in the end of the 1930s [1,2] These viruses have a natural transmission cycle between rodents and mosquitos [3]. Outcomes are significantly worse for young and elderly patients, with case fatalities ranging from 4 to 35% [12,13] These viruses are highly infectious as aerosols [14,15] and an intentional release of sufficient quantities as inhalable small-particle aerosol is expected to infect a high percentage of individuals within an area of a least 10,000 km2 [16]. They can replicate in cell culture to very high titers and are relatively stable to environmental influences [17]
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