Abstract:
Foot-and-mouth disease (FMD) affects cloven-hoofed domestic and wildlife animals and
an outbreak can cause severe losses in milk production, reduction in meat production
and death amongst young animals. Several parts of Asia, most of Africa, and the
Middle East remain endemic, thus emphasis on improved FMD vaccines, diagnostic
assays, and control measures are key research areas. FMD virus (FMDV) populations
are quasispecies, which pose serious implications in vaccine design and efficacy where
an effective vaccine should include multiple independent neutralizing epitopes to elicit
an adequate immune response. Further investigation of the residues that comprise the
antigenic determinants of the virus will allow the identification of mutations in outbreak
strains that potentially lessen the efficacy of a vaccine. Additionally, of utmost importance
in endemic regions, is the accurate diagnosis of FMDV infection for the control and
eradication of the disease. To this end, a phage display library was explored to identify
FMDV epitopes for recombinant vaccines and for the generation of reagents for improved
diagnostic FMD enzyme-linked immunosorbent assays (ELISAs). A naïve semi-synthetic
chicken single chain variable fragment (scFv) phage display library i.e., the Nkuku®
library was used for bio-panning against FMD Southern-African Territories (SAT) 1, SAT3,
and serotype A viruses. Biopanning yielded one unique scFv against SAT1, two for
SAT3, and nine for A22. SAT1 and SAT3 specific scFvs were exploited as capturing
and detecting reagents to develop an improved diagnostic ELISA for FMDV. The SAT1
soluble scFv showed potential as a detecting reagent in the liquid phase blocking ELISA
(LPBE) as it reacted specifically with a panel of SAT1 viruses, albeit with different ELISA
absorbance signals. The SAT1svFv1 had little or no change on its paratope when
coated on polystyrene plates whilst the SAT3scFv’s paratope may have changed. SAT1 and SAT3 soluble scFvs did not neutralize the SAT1 and SAT3 viruses; however, three of the nine A22 binders i.e., A22scFv1, A22scFv2, and A22scFv8 were able to neutralize
A22 virus. Following the generation of virus escape mutants through successive virus
passage under scFv pressure, FMDV epitopes were postulated i.e., RGD+3 and +4
positions respectively, proving the epitope mapping potential of scFvs.
