Study on butterfly breeding sheds light on the evolution of iridescent colors

At her Florida-based butterfly farm, Edith Smith bred a shinier and bluer Common Buckeye but it was Rachel Thayer, a graduate student at the University of California, Berkeley (UC Berkeley) who described the genetic and physical variations that are fundamental to the newly acquired iridescence of the butterfly.

The butterflies that Edith Smith selectively bred are much bluer and more iridescent than the wild Common Buckeye, which is mostly brown (see below). The breeding, UC Berkeley researchers discovered, changed the structure of the wing scales to produce a blue rather than golden structural color. Image Credit: Edith Smith.

During the process, Thayer observed how butterflies are easily able to alter their wing colors over merely a few generations and eventually identified the first gene that has been demonstrated to impact the so-called “structural color.” This structural color is fundamental to the iridescent golden, green, blue, and purple colors of a majority of the butterflies.

Thayer’s findings serve as a preliminary point for novel genetic techniques to find out how butterflies are able to create complex nanostructures with optical characteristics. This could eventually assist engineers to devise innovative methods to create photonic nanostructures for iridescent colors or solar panels for cosmetics, clothing, and paints.

Structural color is not the same as pigment color, like the one on a canvas or in the human skin that reflects or absorbs varying colors of light. Rather, structural color originates from the interaction of light with a solid material just like how a transparent bubble turns into a colorful luster. When the light enters it, it re-bounces out and interferes with the light reflected from the surface in a way that annuls all except for one color.

The Common Buckeye—Junonia coenia—is a predominantly brown butterfly with colorful and spectacular spots. It is found across the United States and usually bred by butterfly farmers intended for wedding ceremonies or for butterfly gardens.

At the Shady Oak Butterfly Farm located in Brooker, Florida, Smith’s breeding experiments with this kind of butterfly were perfect for Thayer’s research on structural color.

Edith noticed that sometimes these butterflies have just a few blue scales on the very front part of the forewing and started breeding the blue animals together. So, effectively, she was doing an artificial selection experiment, guided by her own curiosity and intuition about what would be interesting.”

Rachel Thayer, Graduate Student, Department of Integrative Biology, University of California, Berkeley

In a study that was recently published online in the eLife journal, Thayer along with Nipam Patel, a professor of molecular and cell biology at UC Berkeley and is currently on leave as director of the Marine Biological Laboratory in Woods Hole, Massachusetts, has elucidated the physical variations in wing scales—linked with Smith’s experiment carried out on the Common Buckeye—and reported a single genetic regulator of blue iridescence.

“I especially loved the clear evolutionary context: being able to directly compare the ‘before’ and ‘after’ and piece together the whole story,” added Thayer. “We know that blueness in J. coenia is a recent change, we know explicitly what the force of selection was, we know the time frame of the change. That doesn’t happen every day for evolutionary biologists.”

Structural color produces showy butterflies

Thayer believes that scores of butterflies have been analyzed due to the spectacular structural color in their wing scales. The blue morpho is the most spectacular of all, with 5″ wings of iridescent blue lined with black.

But Thayer’s research focused on a less spectacular genus, Junonia, and noted that the radiant color is common across the 10 species, and even among the dull ones. Under a microscope, the pansy J. atlites, which is an ordinary light gray butterfly, was observed to have radiant rainbow-colored scales whose colors merge together into gray when seen with the naked eye.

According to Thayer, one key lesson derived from the study is that “most butterfly patterns probably have a mix of pigment color and structural color, and which one has the strongest impact on wing color depends on how much pigment is there.”

Thayer raised the crossbred, bluer variety acquired from Smith and also the wild, brownish Common Buckeye.

With the help of an advanced helium ion microscope, Thayer imaged the wings’ scales to observe which structures of the scale account for the color and to find out whether the change in color was caused by the change in structural color, or if this is merely a loss of brown pigment that makes the blue color to stand out.

Thayer did not notice any variation in the amount of brown pigment present on the scales, but she did observe a major variation in the width of chitin, the powerful polymer from which the scale is developed and that is also responsible for the production of the structural color.

The thickness of the chitin layer in the wild buckeye measured approximately 100 nm, producing a golden color that merged with the brown pigment. The bluer buckeye butterfly had chitin that had a thickness of around 190 nm—roughly the thickness of a soap bubble—that yielded a blue iridescence that surpassed the brown pigment.

They are actually creating the color the same way a soap bubble iridescence works; it’s the same phenomenon physically.”

Rachel Thayer, Graduate Student, Department of Integrative Biology, University of California, Berkeley

In addition, Thayer observed that, while the scales from the Junonia butterflies have an extensive tiny structure, structural color emerges from the scale’s base or bottom.

That is not intuitive, because the top part of the scale has all of these curves and grooves and details that really catch your eye, and the most famous structural colors are elaborate structures, often in the top part of the scale. But the simple, flat layer at the bottom of the scale controls structural coloration in each species we checked.”

Rachel Thayer, Graduate Student, Department of Integrative Biology, University of California, Berkeley

“The color comes down to a relatively simple change in the scale: the thickness of the lamina,” stated Patel. “We believe that this will be a genetically tractable system that can allow us to identify the genes and developmental mechanisms that can control structural coloration.”

Furthermore, Thayer examines the scales of mutant buckeyes produced by scientists from Cornell University, but these scales did not have a crucial gene, known as optix, that regulates color.

The micrograph images revealed that the absence of this gene also raised the width of the thin film of chitin present in the scales, resulting in blue color. Optix can be described as a regulatory gene that mediates several other butterfly genes, which Thayer will subsequently be studying.

“One thing that I thought was cool about our findings was seeing that the same mechanism that has recurred over millions of years of butterfly evolution could be reproduced really rapidly in (Smith’s) artificial section experiment. That says that color evolving by changes in lamina thickness is a repeatable, important phenomenon,” Thayer concluded.

Frances Allen, a research scientist at the Department of Materials Science and Engineering of UC Berkeley is also the study’s co-author. The study was financially supported by the National Science Foundation (DEB-1601815, DGE-1106400).

This video produced by the Marine Biological Laboratory explains how a selective mating experiment by a curious butterfly farmer-led scientists to a deeper understanding of how butterfly wing color is created and evolves. Video Credit: Emily Greenhalgh, MBL.