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Construction of mutant FMD viruses to facilitate analyses on virus host cell interactions

Background: In order to determine the importance of particular integrins - and indeed heparan sulphate entities on cell surfaces - for infection of host cells by FMDV, it is necessary to possess the appropriate viral tools. To this end, it is necessary to construct FMD viruses with mutations in the RGD receptor binding site within the GH loop of the viral VP1. Accordingly, the aim was to generate FMDV viruses with mutations in the amino acids flanking this RGD receptor binding site, for application in studies on integrin receptor binding specificity of the virus.

Results: Recombinant FMD viruses were generated from plasmid DNA. The integrin binding loop (the G-H loop of the viral VP1) was sequenced for these rescued viruses. This allowed description of the integrin expressed on porcine and bovine cells targeted by FMDV. Consequently, the integrin specificity of the recombinant FMD viruses was established with cells derived from the natural hosts for the virus.

This integrin specificity was also correlated with infectivity in vivo. RNA inoculation experiments in suckling mice identified the sequence requirements in the VP1 G-H loop for viral entry. Certain amino acid positions were seen to be particularly relevant for interaction with the integrins, while others were dispensable. Actually, in some cases the capsid sequence changed to accommodate the introduced alterations in the G-H loop. These changes introduced into the G-H loop modified virus infectivity in cultured cells and in animals the level of modified infectivity was dependent on the particular change introduced into the G-H loop.

This work has allowed conclusions to be drawn on integrin usage by FMDV, as well as modulation of virus-integrin interaction, in terms of virus infection in cell culture and in vivo.

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