Hybrid zones can be abrupt and leaky
Above: From the cover: Fire‐bellied toad (Bombina bombina) from southern Poland, where they form a famous hybrid zone with their sister species the yellow‐bellied toad (Bombina variegata). Photo credit: Christophe Dufresnes.
I was not even born when Jacek Szymura and Nick Barton initiated their pioneering research on the hybridizing fire-bellied toads Bombina bombina and Bombina variegata in the late 1970s / early 1980s. These tiny amphibians are quite distinctive in looks, sounds and habitats, but still they utterly hybridize wherever they get the chance along an extensive area of contact in Central Europe. This area soon became a playground to study hybrid zones and understand how different species are maintained apart despite interbreeding and gene flow, a major question in evolutionary biology. Dozens of papers on this iconic system, some highly influential, were published in subsequent decades.
It thus felt natural to include the B. bombina/variegata pair in my post-doc project a couple of years ago: a comparative framework of amphibian hybrid zones analyzed with high-throughput sequencing data to characterize the genetic and biogeographic features of speciation. For Bombina, we specifically aimed at testing whether the two species kept their genetic integrity everywhere they hybridize, or whether there was noticeable variation linked to their dynamic biogeographic history. And simply to revisit this famous speciation model for the first time with genome-wide markers.
Cover article: (Free to read online for a year.)
Dufresnes C, Suchan T, Smirnov NA, Denoël M, Rosanov JM, Litvinchuk SN. 2021. Revisiting a speciation classic: Comparative analyses support sharp but leaky transitions between Bombina toads. Journal of Biogeography 48(3): 548-560. https://onlinelibrary.wiley.com/doi/full/10.1111/jbi.14018
To get fresh samples, Dr. Tomasz Suchan and I initially went into the field near Krakow in Poland, home of the first Bombina hybrid zone ever documented. But decades after Jacek Szymura cycled around these once abundant populations, most of them had since disappeared following landscape changes. Exploration of suitable habitats along the Carpathian foothills led us to discover a new contact, although now fragmented by … the E40 highway, which was precisely built along the foothills!
In parallel, we were able to resurrect a forgotten collection of specimens sampled in the 1990s by Drs. Spartak Litvinchuk and Juriy Rosanov across a wide transition zone in the Ukraine Transcarpathians, and Dr. Nazar Smirnov recently documented another area of hybridization in the Ukraine Ciscarpathians. Combined with previous literature – thanks to the support of Prof. Alexey Yanchukov – new and published data could be gathered across no less than 11 contact zones!
Fine-scale distributions of the two species in Central Europe, and admixture analyses of toads collected across two parapatric ranges in Poland (left) and Ukraine (right), based on genome-wide markers. In these examples, most individuals (= horizontal bars) are either pure B. bombina (blue) or B. variegata (green), but with slight admixture by the other species. Credit: Prof. Christophe Dufresnes.
Finally, to properly reconstruct the history of hybrid zone formation, we had to map the numerous lineages of each species. Dr. Mathieu Denoël spent days buried under amphibian atlases to accurately delimit range extents, which were then exploited to predict past distributions and routes of expansions from the various glacial refugia.
We were quite amazed that in all contact zones, the exact same pattern stood out: sharp but leaky transitions (hence the title of our paper), in the sense that the genomes of the two species always locally admix, but only a few alleles made it far into the foreign ranges. This means that species boundaries between B. bombina and B. variegata remain robust despite the ongoing gene exchange, probably because the hybrids are not that fit. Based on hundred-times more loci than the classic Bombina research, our study thus provides decisive empirical evidence that hybridization should not be viewed as a force of “despeciation”, as long as reproductive barriers that prevent hybrids to spread are strong enough, in the form of developmental problems or maladaptation.
Dr. Nazar Smirnov taking notes on a B. variegata site in Western Ukraine. Credit: Dr. Ihor Skilsky.
Moreover, our results also implied that the biogeographic history of these numerous hybrid zones did not really influence how the toads mixed in the end. This was surprising since in other amphibians, we had shown that hybrid zones could have quite different genetic structures depending on the lineages involved, the time since contacts, etc. Here, such replicability indicates that when two species are sufficiently differentiated, hybrid swarms remain geographically restricted, no matter how and when the contact is established.
What next? In parallel to this publication, we have pushed analyses further to look at the genomic architecture of reproductive isolation, i.e. the number of barrier loci that cause incompatibilities between species, by comparing multiple pairs of European frogs and toads. It turns out that with their sharp transitions, the Bombina pair lies at the upper edge of the speciation continuum, where many parts of the genome ceased to admix. Another team has also been working on a reference genome, from which they have already identified the sex chromosomes. These developments mark the advent of a new exciting era for evolutionary research in the timeless Bombina model!
A precious finding a few hundred meters south of the E40 highway in southern Poland: a B. variegata individual (slightly admixed by B. bombina) in a forest track puddle. Credit: Prof. Christophe Dufresnes.
LASER, College of Biology & the Environment, Nanjing Forestry University, Nanjing, China.