Identification of Natural Hybrids between Ahlbergia frivaldszkyi (Lederer, 1853) and Callophrys rubi (Linnaeus, 1758) (Lepidoptera, Lycaenidae) Using Mitochondrial and Nuclear Markers

SIMPLE SUMMARY: Butterfly specimens with unusual morphological characters (e.g., unusual wing coloration) have contradictory interpretations in the literature and have been considered by different authors either as previously undescribed taxa, putative hybrids, or aberrations of well-known species. Such individuals clearly represent a taxonomic problem that needs to be addressed by scientists. The application of molecular techniques could shed light on the origin of morphological uncertainty. Here we use a combination of mitochondrial and nuclear DNA markers to analyze three lycaenid butterflies with unusual wing pattern, which are thought to represent naturally occurring hybrids due to their intermediate phenotype. We confirm their hybrid origin and indicate that the specimens are wild-caught hybrids between females of Callophrys rubi and males of Ahlbergia frivaldszkyi. Our data indicate that gene flow across species boundaries in these butterflies can occur long after speciation. ABSTRACT: Natural hybridization is rather widespread and common in animals and can have important evolutionary consequences. In terms of taxonomy, exploring hybridization and introgression is crucial in defining species boundaries and testing taxonomic hypotheses. In the present paper, we report on natural hybrid specimens between Ahlbergia frivaldszkyi (Lederer, 1853) and Callophrys rubi (Linnaeus, 1758). To test the hypothesis of their hybrid origin, we employed the molecular mitochondrial (COI gene) and nuclear (wingless, RPS5, and Ca-ATPase genes) markers commonly used in phylogenetic studies and explored the morphology of the specimens. Our analysis revealed that hybrids bear mitochondrial haplotypes of C. rubi, while nuclear fragments are heterozygous, sharing a combination of A. frivaldszkyi and C. rubi lineages. The hybrid specimens combine morphological characters of both genera. Our results for the first time empirically demonstrate the possibility of genetic introgression between these species and between the genera Callophrys and Ahlbergia on the whole.