Vladimir A. Lukhtanov saw Agrodiaetus butterflies of several species flying together, and even though they all looked much the same in most respects — all an inch to an inch and a half wide, with the same sort of spots, and, upon examination, same-shaped genitalia — the males of different species had wings of different color. Why?

Lukhtanov is an entomologist at St. Petersburg State University in Russia, where one of his students was Nikolai P. Kandul. Kandul came to Harvard and completed a doctorate in organismic and evolutionary biology this year, working in the laboratory of Hessel professor of biology Naomi E. Pierce. With Kandul as link, collaborating teams — consisting of Lukhtanov and one other from St. Petersburg, and Kandul, Pierce, and two others from Harvard — analyzed the Agrodiaetus butterflies that Lukhtanov had seen.

Agrodiaetus damon male	Agrodiaetus damon female
Photographs by Nikolai Kandul

The researchers built a phylogeny, or family tree, of 15 species. They were relatively young species and genetically very closely related. Kandul and his colleagues believe that the species formed in a process called “reinforcement.” Reinforcement occurs when natural selection strengthens behavioral discrimination to avoid costly interspecies matings that tend to produce weedy, sterile hybrids. The evolutionary process of reinforcement can eventuate in speciation, providing a direct link between Darwin’s theory of natural selection and the origin of new species.

Male butterflies of various Agrodiaetus species display a range of wing color to the human eye. Tests by researchers using ultraviolet light confirm that butterflies see similar differences.

Nikolai Kandul and Vladimir Lukhtanov

“The phenomenon of reinforcement,” says Kandul, “is one of the very few mechanisms that has natural selection playing a role in speciation. It might be widespread, but it is hard to find good evidence of it.” The joint study of Agrodiaetus butterflies, reported in Nature, details a probable instance, rarely demonstrable in nature, of the reinforcement model of evolution at work.

For one species of butterfly to become two, two branches of the species must stop breeding with each other, as though Cantabrigians decided to stop breeding with other Americans. Two branches of the species may be separated geographically — by a mountain range, for instance. (The genus Agrodiaetus lives from Spain to Central Asia, sometimes amid mountain ranges.) After hundreds of generations, the butterflies may find that they are no longer able to interbreed and have become clearly distinct species. But what if a nascent species that has developed some random genetic difference while living in isolation comes round the mountain and finds itself in seductive proximity to one or more other developing species? They must all take vows of reproductive isolation if they are to successfully branch into different species. Biologists can get excited by these Agrodiaetus butterflies because they appear to show in their colorful wings how reproductive isolation can be aided.

These photographs show pairs of chromosomes in four Agrodiaetus species. "Usually butterfly species from the same genus have very similar numbers of chromosomes," says Nikolai Kandul. "This group is fascinating because every species has a different number, and the range in number in the genus is huge." These differences in chromosome number probably drive speciation, says Kandul. The more a breeding pair differs in their number of chromosomes, the less successful their union will be.

Nikolai Kandul

The collaborators discovered that separated species of Agrodiaetus butterfly living in isolation tend to look alike. In species living together, on the other hand, the males of each species look strikingly different: their wings may be silver, brown, or blue. (All females are brown.) The males’ divergent wing colors allow females, which breed only once, to identify a same-species mate and avoid producing hybrids through interbreeding. Natural selection acts through reinforcement against maladaptive hybrids and favors colorful wings as ways of distinguishing species. It promotes the evolution of differences between nascent species and thus speciation. Thus do those colorful wings drive a wedge between species.                                                               

Nikolai Kandul e-mail address: kandulfas [dot] harvard [dot] edu (kandulfas [dot] harvard [dot] edu

)nature website: www.nature.com/nature/journal/v436/n7049/abs/nature03704.htm