Multiple introgression events from ghost Rüppell’s fox mitochondrial lineages into red fox

Mitochondrial introgression has been reported between red fox (Vulpes vulpes) and Rüppell’s fox (Vulpes rueppellii). While an evolutionary scenario of old divergence followed by recent mitochondrial introgression has been proposed, the directionality, prevalence, and timing of this event remain unclear. To further investigate this scenario, we analysed mitogenomes (n=85), including four newly generated red fox mitogenomes from Türkiye and a new Rüppell’s fox mitogenome from the United Arab Emirates, and partial mitochondrial DNA sequences (n=320) from both species across their ranges, including a newly comprehensive sampling in the Anatolian Peninsula (n=80). Our results are consistent with unidirectional mitochondrial introgression from the desert-adapted Rüppell’s fox into the generalist red fox, likely driven by climatic shifts promoting secondary contact and asymmetrical reproductive behaviour. Phylogenetic analysis unveiled two deeply divergent mitochondrial lineages of Rüppell’s fox sampled among red fox individuals: one distributed across Türkiye, Iran and Tunisia, and another restricted to Iran, suggesting that likely now-extinct (ghost) mitochondrial lineages may have introgressed into red foxes at least twice. Estimates of the time to the most recent common ancestor indicate that introgressed and contemporary Rüppell’s fox mitochondrial lineages diverged approximately 230 kya, predating the current intraspecific mitochondrial diversification of Rüppell’s fox lineages (~72 kya). This study highlights how comprehensive mitogenomic data and exhaustive regional surveys are critical for elucidating the complexity of introgression patterns in closely related canids.