Analysis of SAT Type Foot-And-Mouth Disease Virus Capsid Proteins and the Identification of Putative Amino Acid Residues Affecting Virus Stability

Abstract

Foot-and-mouth disease virus (FMDV) initiates infection by adhering to integrin receptors on target cells, followed by cell entry and disassembly of the virion through acidification within endosomes. Mild heating of the virions also leads to irreversible dissociation into pentamers, a characteristic linked to reduced vaccine efficacy. In this study, the structural stability of intra- and inter-serotype chimeric SAT2 and SAT3 virus particles to various conditions including low pH, mild temperatures or high ionic strength, was compared. Our results demonstrated that while both the SAT2 and SAT3 infectious capsids displayed different sensitivities in a series of low pH buffers, their stability profiles were comparable at high temperatures or high ionic strength conditions. Recombinant vSAT2 and intra-serotype chimeric viruses were used to map the amino acid differences in the capsid proteins of viruses with disparate low pH stabilities. Four His residues at the inter-pentamer interface were identified that change protonation states at pH 6.0. Of these, the H145 of VP3 appears to be involved in interactions with A141 in VP3 and K63 in VP2, and may be involved in orientating H142 of VP3 for interaction at the inter-pentamer interfaces.

Intra-serotype SAT2 chimeric foot-and-mouth disease vaccine protects cattle against FMDV challenge

“…Virus stocks were titrated by plaque assays in BHK-21 cells as described previously [22]. The BHK-21 cells were also used for RNA transfection and virus recovery of the chimeric pSAT2 ZIM14 -SAT2virus [34]. The plasmid pSAT2 ZIM14 -SAT2 containing the outer capsid-coding region of SAT2/ZIM/14/90 in the pSAT2 backbone ( Fig.…”

“…The plasmid pSAT2 ZIM14 -SAT2 containing the outer capsid-coding region of SAT2/ZIM/14/90 in the pSAT2 backbone ( Fig. 1) was previously constructed [34].…”

“…The rescue of vSAT2 ZIM14 -SAT2 virus from a chimeric genomelength cDNA plasmid has been described before [34]. Culture fluids from SAT2/ZIM/14/90 and vSAT2 ZIM14 -SAT2 infected BHK-21 cells were harvested, inactivated with 5mM binary ethyleneimine (BEI) for 26 h at 25 • C, clarified by centrifugation, concentrated with 8% PEG (w/v) and resolved on 10-50% (w/v) sucrose density gradients (SDG) by rate zonal centrifugation at 36,000 g for 16 h at 4 • C. The gradients were fractionated and analyzed spectrophotometrically by measuring the absorbance at 260 nm.…”

“…The construction of the intra-serotype chimeric vSAT2 ZIM14 -SAT2 virus and its antigenic profiling, growth kinetics, plaque morphologies and biophysical stability have been described elsewhere [34]. Although titres produced in BHK-21 cells were relatively low, i.e.…”

“…We assessed the serological profile gen-erated in cattle, using liquid phase blocking ELISA (LPBE) and the gold standard virus neutralisation (VN) assays as in vitro mark-ers of protection. The intra-serotype vaccine was prepared from a chimeric virus containing the external capsid-coding region of a field SAT2 isolate, ZIM/14/90, inserted in the genetic background of a SAT2 infectious clone [34]. The SAT2/ZIM/14/90 virus was iso-lated from buffalo which originated from Doma Safari area (16 • 20 , 30 • 15 ) in Zimbabwe and was selected for its potential use as a vaccine strain based on (i) initial data showing the ability to elicit an immune response that cross-reacts broadly against the three SAT2 topotype viruses from southern Africa [28] and (ii) the 9.7%amino acid differences in the capsid proteins compare to the SAT2 infectious clone shows genetic divergence from the clone [28].…”