Isotope geolocation and population genomics in Vanessa cardui: Short- and long-distance migrants are genetically undifferentiated

The painted lady butterfly Vanessa cardui is renowned for its virtually cosmopolitan distribution and the remarkable long-distance migrations that are part of its annual, multi-generational migratory cycle. Recently, V. cardui individuals were found north and south of the Sahara in the autumn, suggesting distinct migratory behaviours within the species. However, the evolutionary and ecological factors shaping these differences in migratory behaviour remain largely unexplored. Here, we performed whole-genome resequencing and analysed the hydrogen and strontium isotopes of 40 V. cardui individuals simultaneously collected in the autumn from regions both north and south of the Sahara. Our investigation revealed two main migratory groups: (i) short-distance migrants, journeying from temperate Europe to the circum-Mediterranean region and (ii) long-distance migrants, originating from Europe, crossing the Mediterranean Sea and Sahara, and reaching West Africa, covering up to over 4,000 km. Despite these stark differences in migration distance, a genome-wide analysis revealed that both short- and long-distance migrants belong to a single intercontinental panmictic population extending from northern Europe to sub-Saharan Africa. Contrary to common biogeographic patterns, the Sahara is not a catalyst for population structuring in this species. No significant genetic differentiation or signs of adaptation and selection were observed between the two migratory phenotypes (pairwise FST = 0.001 ± 0.006). Nonetheless, two individuals, which were early arrivals to West Africa and covered longer migration distances, exhibited some genetic differentiation. The lack of genetic structure between short- and long-distance migrants suggests that migration distance in V. cardui is a plastic response to environmental conditions. Significance statement Although migratory insects dominate living biomass fluxes and impact agriculture, ecosystems, and human communities, little is known about the controls of their migratory behavior. Our study develops an interdisciplinary framework, applied to the migratory butterfly V.cardui , to explore the genetic basis of variation in insect migration behavior. We leverage new generation isotope geolocation techniques to uncover striking differences in butterfly behaviour, with some individuals migrating short distances within the circum-Mediterranean region and others migrating thousands of kilometers across the Mediterranean Sea and Sahara. This major difference does not coincide with genetic differentiation or population structure and is likely a plastic response to environmental cues. This study provides a ground-breaking framework to study migration in insects. ### Competing Interest Statement The authors have declared no competing interest.