The story of the peppered moth, Biston betularia, is one of the most famous evolutionary tales known to the public, and is a staple of both popular literature and biology texts. It’s appealing because it’s an example of “evolution in action”: a case in which we could see evolution happening over only one or two human generations, and we now understand the forces of natural selection that caused the evolutionary change.

The change was a simple one: the replacement of an originally light-colored form of the moth (called the “typica” form, with a “peppered” appearance; see below) by a dark-colored form (“carbonaria“) in Britain, all occurring between about 1850 and 1900.  The difference between the forms was due to variants of a single gene, with the carbonaria form being dominant over the typica form, so that having one copy of the “dark” allele makes you fully dark. (There’s also an intermediate “half dark” form called “insularia“.)

Why the evolutionary change? (Remember, evolution can be defined as “genetic change in populations”). The Wikipedia article on “Peppered moth evolution” gives a good summary of the situation.

Early experiments demonstrated that the likely cause was bird predation. Birds are visual predators and pick moths off the trees. Before Britain’s Industrial Revolution, the trees were light in color, often festooned with lichens, and so light-colored moths were camouflaged, less likely to be eaten than were the carbonarias. The first dark-colored moth, a rarity, was described in 1848 in Manchester, and then the frequency of this “morph” (color variant) rose rapidly as soot from factories darkened the trees and killed off the lichens.

This change in tree color, most pronounced in industrial areas like Manchester, now reversed the selective advantage of the carbonaria versus typica forms: the dark moths now were more camouflaged and less likely to be eaten. The frequency of that form then rose strongly in many areas: to nearly 100% in  the grim town of Manchester (apologies to Matthew). All of this change took place over only about 50 years, so natural selection was very strong.

Here are two photos showing the typica and carbonaria forms on darkened versus “normal” trees: you can see how the wrong-colored moths stick out, and so would be obvious to birds looking for a meal:

A soot-darkened tree:

A tree not exposed to pollution. Notice the speckled color of the typica form, which blends in well with the tree bark:

After the passage of clean air legislation in Britain in the 1950s, pollution abated and tree color began to return to its normal light appearance. As one might expect, the frequency of the light typica form began to increase, and it’s now well above 70% in most locations.

Experiments by Bernard Kettlewell in the 1950s, involving placing moths on trees and then later recapturing them, suggested the bird predation theory, as dark moths released in light woods were recaptured less often than were light moths, suggesting that the wrong-colored moths had a higher mortality. The opposite was the case for releases in polluted woods. Other research, involving placing moths on trees, suggested the same scenario.

There were, however, problems with the design and execution of these experiments, and I acquired a bit of notoriety in 1998 by pointing these issues out, adding that the demonstration of bird predation as the cause of the evolution was suggestive but not conclusive. I was somewhat demonized by British evolutionists who were wedded to the whole story, and my objections were, of course, picked up by creationists, who misused them to cast doubt on all of evolution. If the textbook case of natural selection was flawed, they claimed, then the whole Darwinian enterprise was wrong.

However, Michael Majerus, a researcher in Cambridge, repeated the release-recapture experiments properly beginning in 2001 (he used only light-colored woods, for the dark, polluted woods were gone), and found the same result as did Kettlewell. In Majerus’s light colored woods, the dark moths that were released along with light ones were recaptured less frequently, and Majerus actually observed birds and bats eating these moths. (Sadly, he died before the work was published, but his colleagues wrote it up and got it published.) Majerus’s findings, along with the observation that the same species in North America experienced the same changes in morph frequency coincident with both the rise and fall of pollution in the U.S., satisfied me that the story of the moths is now pretty solid. There is now very little doubt that we have a case of evolution by natural selection in action, and we pretty much know what kind of selection was operating.

For decades we’ve known that the difference between the carbonaria and typica forms was due to change at a single gene, for genetic crosses showed a nice Mendelian segregation, with carbonaria behaving as a dominant allele. But we didn’t know the exact gene involved, although in recent years it was narrowed down to a 400,000-base section of the moth’s genome. Although the fact that this is a case of evolution by natural selection doesn’t depend on knowing the exact gene involved, to get the complete story from gene to color to the ecological forces involved (bird predation), it would be nice to know what that gene is and what it does.

A big group of researchers has now reported in Nature that they found the gene. The paper is by Arjen E. van’t Hof et al. (reference below, sadly, no free download), and identifies the gene causing the moth color difference as cortex, a well-known gene that’s been studied in fruit flies (Drosophila). A companion paper in the same issue by Nicola Nadeau et al. (reference below, also behind a paywall) shows that mutations at cortex are involved in patterning and mimicry in many species of Lepidoptera. I won’t describe that paper in detail, as it’s only tangentially relevant here.

These are the salient results of the van’t Hoff et al paper:

• The gene where the change resides that made the light-colored moths dark is called cortex, and is known in Drosophila to be involved in cell division of the egg in females. It wasn’t known, at least in flies, to be involved in color.

• The association between cortex and color was found by “association mapping” in B. betularia. The researchers took a bunch of moths of both the carbonaria and typica forms (and insularia as well), and sequenced DNA in the region where the mutation was known to reside, looking for a consistent change in the DNA that would distinguish the various forms with near-perfect ability.

• The change was found not to be a single-base mutation in the DNA, but an insertion of a “transposable element” (or “transposon”)—a bit of DNA that can move around in the genome—into the carbonaria form. The whole inserted region comprises 21,925 nucleotides, and involves a single element present in 2.3 copies that has moved as a unit into the cortex gene.

• The transposable element actually activated (rather than silenced) the cortex gene, increasing the amount of its product in a manner we don’t understand. Curiously, the increased gene activity in the dark form was far more pronounced in larvae (caterpillars, which the paper calls “crawlers”) rather than pupae or adults, presumably because the precursors for scale development and their color are being formed at this stage of development.

• The association between the transposon and color was nearly perfect, but not 100% so. Every one of the typica and insularia moths lacked the element, while 105 of 110 black carbonaria forms had the element. This means that other genes besides cortex may influence color—or developmental/environmental effects that aren’t genetic—but that the insertion in cortex is likely the most important one: the one that produced the fuel for natural selection on color.

• By looking at the DNA around the transposon, the researchers could estimate the age of the mutation. If it arose recently in a single individual, most carriers of the mutation would have similar sequences nearby, as recombination wouldn’t have time to put the transposon next to varied DNA from other individuals. If the mutation was old, the surrounding regions of the carbonaria form would be more diverse, as recombination would have combined the inserted gene with other nearby genes from different individuals. Using simulations, the researchers gave the most likely date of the mutation as 1819, shortly before it was first seen in the wild. The “interquartile range” for the dates, which I take to be the range of dates between the 25% and 75% likelihood that it originated, is 1681-1806. Below is the graph showing the probability density of when the mutation to carbonaria originated (i.e., when the transposon moved). You can see that all the dates are fairly recent, so this mutation did not occur thousands of years ago. The highest probability is at 1819, not far from when it was first seen in the wild (1848; dotted line). A new variant butterfly would be found pretty quickly by the Brits, who are avid butterfly and moth collectors!

• Finally, the paper by Nadeau et al. immediately following this one in the journal showed that in the butterfly genus Heliconius, which is full of mimics as well as “warningly colored” species, cortex is involved in scale development, and thus probably scale color. The gene has been recruited to a new function in Heliconius, although it may still play some role in cell division during the formation of the butterfly egg.

The upshot.  As I said, we can regard this paper as lagniappe: we already knew the main points of the story—that mutation in a single gene was the basis for an adaptive change in moth color based on bird predation. Now we know that the single gene is cortex.

We also know that the change in cortex, causing a change in moth color from light to dark, was due to the insertion of a mobile transposable element, not the “conventional” mutation we think of: a change in a single base of a DNA sequence. This is still a “mutation”, but a large change in the gene that altered how much product it produced. There may be other adaptations in other species known to rest on insertions or removals of transposons, but I don’t know of any. (Readers with that knowledge should weigh in.)

The mutation occurred only shortly before the environmental change—pollution—that caused the evolution of the color difference. That’s interesting, but it wasn’t necessary, for mutations like this occur continuously, and can hang around permanently. That’s because, although natural selection weeds such genes out of populations (dark moths would be at a disadvantage before the Industrial Revolution), mutation keeps putting them back in, so there is a reservoir of low-frequency mutations hanging around that could be the basis for a new adaptation should the environment change. (This is called a “mutation/selection equilibrium.”) Remember, though, that those mutations aren’t hanging around for the purpose of providing future adaptive evolution.  Errors in DNA happen randomly, cannot arise to anticipate the organisms’s future needs, and sometimes, but not usually, turn out to be useful.

Finally, this paper, and the adjoining one on Heliconius, show that while fruit flies are a good “model organism”—a species that has taught us a lot about development and genetics (after all, most of what we know about Mendelian genetics was worked out in Drosophila)—it didn’t tell us much about evolution in Biston betularia. This is for a simple reason: flies don’t have scales, and so scale color couldn’t be studied using Drosophila. What happened, as we see so often in evolution, is that a gene that does something in one species can be co-opted for a different function in another species.  This twist on evolution was only realized after we developed new genetic and developmental tools over the last 30 years.

h/t: Matthew Cobb, Jonathan

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van’t Hof, A. et al. 2016. The industrial melanism mutation in British peppered moths is a transposable element. Nature 534:102-105.

Nadeau, N. J. et al. 2016.  The gene cortex controls mimicry and crypsis in butterflies and moths. Nature 534:106-110.