In previous work we have reported the immunization of swine using in vitro-transcribed foot-and-mouth disease virus (FMDV) RNA. With the aim of testing whether RNA-induced immunization can mediate protection against viral infection, a group of Swiss adult mice was inoculated with FMDV infectious transcripts. In most inoculated animals viral RNA was detected in serum at 48-72h postinoculation. A group of the RNA-inoculated mice (11 out of 19) developed significant titers of neutralizing antibodies against FMDV. Among those animals that were successfully challenged with infectious virus (15 out of 19), three out of the eight animals immunized upon RNA inoculation were protected, as infectious virus could not be isolated from sera but specific anti-FMDV antibodies could be readily detected. These results suggest the potential of the inoculation of genetically engineered FMDV RNA for virulence and protection assays in the murine model and allow to explore the suitability of RNA-based FMDV vaccination in natural host animals.