Submitted to: Journal of Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 6, 2005
Publication Date: October 1, 2005
Citation: Henry, T.M., Duque, H., Baxt, B., Rieder, A.E. 2005. Analysis of Foot-and-Mouth Disease Virus Type A24 Isolate Containing an SGD Receptor Recognition Site In Vitro and Its Pathogenesis in Cattle. Journal of Virology. Vol 79:20, p. 12989-12998. Interpretive Summary: Foot-and-mouth disease virus is a highly contagious viral disease of livestock that has been extensively studied due to its economic impact. The re-emergence of FMD in Europe and South America in recent years has emphasized how little is known about the virus-host interactions in susceptible animals that impact the pathogenesis and virulence of the virus. We have shown that nine successive passages of a parental A24 Cruzeiro strain in cattle selected a variant (A24-B9) which contained an SGD sequence in the cell-receptor binding site with apparent changes in the virus phenotype resulting in low titers and turbid plaques in BHK-21 cells. Propagation of A24-B9 virus in BHK-21 cells resulted in the rapid selection of viruses that grew to high titers and contained an RGD binding motif. Vaccines prepared from tissue culture adapted A24 virus failed potency tests when the A24-B9 virus was used to challenge the vaccinated animals. To examine the role of the SGD in vitro and in vivo the passaged virus together with the parental virus strain were cloned for molecular characterization and the genetic modification responsible for the altered phenotype were determined. The SGD virus could only replicate in COS cells expressing the bovine avb6 integrin. In contrast, tissue culture adapted RGD virus, replicated in cells expressing the bovine aVb1, aVb3, nd aVb6 integrins. A bovine inoculated with the SGD virus developed clinical FMD and transmitted the disease to a naïve steer. Virus isolated from the contact animal maintained the SGD sequence. These results illustrate the need to examine the virus stocks used for production and challenge of vaccines since propagation of FMDV in tissue could result in the selection of viruses with altered cell tropisms.
Technical Abstract: Foot-and-mouth disease virus initiates infection by binding to integrin receptors via an Arg-Gly-Asp (RGD) sequence found in the G-H loop of the structural protein VP1. Following serial passages of a type A24Cruzeiro virus (A24Cru) in bovine via tongue inoculation, a virus was generated which contained a SGD sequence in the cell-receptor binding site and expressed a turbid plaque phenotype in BHK-21 cells. Propagation of this virus in these cells resulted in the rapid selection of viruses that grew to higher titers, produced clear plaques, and now contained an RGD sequence in place of the original SGD. To study the role of the SGD sequence in FMDV receptor recognition and bovine virulence, we assembled an infectious cDNA clone of A24Cru and derived mutant clones containing either SGD with a single nucleotide substitution in the S144 codon, or double substitutions at this position to prevent mutation of the S -> R. The SGD viruses grew poorly in BHK-21 cells and stably maintained the sequence during propagation in BHK cells expressing the bovine aVb6 integrin (BHK3 aVb6) as well as in experimentally infected and contact steers. While all the SGD-containing viruses used only the bovine aVb6 with relatively high efficiency, the revertant RGD viruses utilized bovine aVb1, and, with aVb3 higher efficiency than aVb6 and grew well in BHK-21 cells. Replacing the R at the ' 1 SGD position with either K or E showed that this residue did not contribute to integrin utilization in vitro. These results illustrate the rapid evolution of FMDV with alteration in receptor specificity, and suggest that viruses with sequences other than RGD, but closely related to it, can still infect via integrin receptors, and induce and transmit the disease to susceptible animals.