Estimates from radiocarbon-dated packrat middens indicate that the high elevation woodland communities of the Madrean Sky Islands were continuous as recently as 8,000 to 12,000 years ago. A number of population studies on a diverse collection of taxa have investigated the extent to which the Madrean Sky Island system has limited gene flow among mountain ranges. The results of several of these studies indicate that population divergences may be more ancient than the Holocene. Yarrow’s spiny lizards, Sceloporus jarrovii, were sampled from eight sites representing seven mountain ranges. The populations of S. jarrovii are host to the malarial parasite, Plasmodium chiricahuae. DNA sequences from the lizards and their malarial parasites were used to reconstruct the evolutionary relationships and estimate the ages of the populations for both host and parasite. The findings of these analyses indicate that the sky island populations of S. jarrovii have been isolated for hundreds of thousands of years and did not experience gene flow during the last woodland expansion. In contrast, the results indicate that the malarial infection occurred more recently, possibly during the Holocene woodland expansion. In addition, the prevalence of the malarial infection was compared to multiple attributes of the lizards. This analysis revealed a negative relationship between the genetic diversity of the lizard populations and the prevalence of infection. Furthermore, lizard populations with lower prevalence of infection have a lower frequency of multiclonal infections.