Vesicular stomatitis (VS) outbreaks of unknown origin occur at 8-10-year intervals in the south-western USA with the most recent outbreak beginning in 2004. A previous study has suggested that strains causing US outbreaks are closely related to strains causing outbreaks in Mexico [Rodriguez (2002) Virus Res 85, 211-219]. This study determined the phylogenetic relationships among 116 vesicular stomatitis New Jersey virus (VSNJV) strains obtained from the 2004 outbreak and from endemic areas in Mexico. All 69 US viruses showed little sequence divergence (<or=1.3 %), regardless of their location or time of collection, and clustered with 11 Mexican viruses into a genetic lineage not previously present in the USA. Furthermore, viruses with identical phosphoprotein hypervariable region sequences to those causing the US outbreaks in 1995-1997 and 2004-2005 were found circulating in Mexico between 2002 and 2004. Molecular adaptation analysis provided evidence for positive selection in the phosphoprotein and glycoprotein genes during a south-to-north migration among 69 US viruses collected between the spring and autumn of 2004 and 2005. Phylogenetic data, temporal-spatial distribution and the finding of viral strains identical to those causing major outbreaks in the USA circulating in Mexico demonstrated that VS outbreaks in the south-western USA are the result of the introduction of viral strains from endemic areas in Mexico.