NEWS FEATURE

Prehistoric animals, in living colorPaleontologists are looking beyond bones to reveal the hues of prehistoric animals that

vanished millions of years ago. But the young field has its share of disagreements.

Amber Dance, Science Writer

Michael Benton used to tell his paleontology students thatthey would never know the true color of a dinosaur. Afterall, even fossils that sport light or dark patches may notindicate the creature’s original hue. But in recent years, thevertebrate paleontologist at the University of Bristol in theUnited Kingdom has had to revise those lectures.

In 2010, Benton and colleagues found evidencethat the feathered dinosaur Sinosauropteryx primahad reddish-brown stripes on its tail (1). The same year,another group claimed that birdlike Anchiornis huxleyibore a red crest on its head (2). Since then, others havediscovered that a marine ichthyosaur was dark-colored,and that the early bird Confuciusornis sanctus possesseddark feathers with light wing tips (3, 4) (Table 1). Sud-denly, the color of prehistoric animals has become anactive topic for research rather than speculation, turningBenton into an optimist: “If you’re a scientist, never sayanything is impossible,” he now tells students.

Most of these colorful revelations have emerged fromfossils that contain evidence of melanin pigments—responsible for earth tones, such as red, black, brown,and buff—or the tiny cellular bags, called melano-somes, which produce and store melanins. But somescientists are already identifying the brighter hues ofancient snakes and insects. “Probably all of the colorscan eventually be identified,” predicts Benton.

The work is not only helping to repaint the colorfulpictures of dinosaurs that charm schoolchildren andmuseum-goers. Color can provide clues about howanimals lived, mated, or died. Bright feathers suggestthat animals used those hues to select a desirablemate, for example, whereas green coloration wouldoffer camouflage amid foliage, indicating there musthave been predators about. “Color has roles in lots ofdifferent parts of animal behavior and social interactions,”

Researchers reconstructed the plumage color of a Jurassic birdlike dinosaur called A. huxleyi using color-imparting melanosomes, the morphology ofwhich had been preserved in fossil feathers. Image courtesy of Michael DiGiorgio (artist).

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says Nick Edwards, a paleontologist at the Universityof Manchester in the United Kingdom.

But the fledgling field of paleo-color has also gen-erated controversy. Some scientists dispute whethermelanosomes could be preserved in fossils, and arguethat researchers are being fooled by what are in factfossilized bacteria. Others question the use of certainmetals as proxies for pigments. “I think what’s happeningis what often happens when people come up with newideas,” says Luis Chiappe, Vice President for Research andCollections at theNatural HistoryMuseumof Los Angeles.“We got super excited because we had figured out waysof potentially devising the colors of these extinct ani-mals. . . now we’re starting to realize it’s not as simple.”

The Race to Color a DinosaurIn retrospect, it’s not so surprising that pigment tracescan survive for so long. Melanins, derived from the aminoacid tyrosine, are remarkably tough polymers, and theytightly cross-link with proteins, such as the keratin infeathers. This strengthens tissues, and could even

be responsible for helping to preserve fossil feathers,says Benton.

Nineteenth century fossil hunters certainly knew thatmelanins could stand the test of time. Some even wroteletters with the melanin pigment ink that they foundpreserved in the ink sacs of fossilized squids. And it wasduring his own studies of squid fossils in 2006 that JakobVinther, then a graduate student at Yale University inNew Haven, Connecticut, began to wonder whethermelanins could persist in other fossils. A handful of sci-entists, including his supervisor Derek Briggs, had pre-viously seen minuscule, sausage-shaped formations in avariety of fossils, and labeled them fossil bacteria (5, 6).Vinther suggested these formations could be mela-nosomes instead. Melanosome shape, he reasoned,could even give clues to color: sausage-shaped mela-nosomes contain the black pigment eumelanin, whereasround melanosomes hold reddish-brown pheomelanin.

Briggs, a paleontologist, was skeptical, but willing totest the idea. He suggested that Vinther take a closer lookat a fossil feather that haddark and light stripes. If theblobs

Illustrations courtesy of Lucy Reading (graphic artist).

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were melanosomes, they should be found only in the darksections. If theywere bacteria, they ought to be all over thefeather. When Vinther examined the light sections with ascanning electron microscope, he saw nothing more thanthe imprint of the feather itself. But in the dark sections hefound the oblong bodies, along with their impressions inthe rock (7). His hypothesis appeared to be correct.

Once Vinther’s idea went public in 2008, the racewas on to color in a dinosaur. It ended in a dead heat:Benton’s and Vinther’s teams unveiled the colors ofstripe-tailed S. prima and red-headed A. huxleyiwithindays of each other in early 2010 (1, 2).

Other discoveries followed fast, and Benton saysthat the majority of paleontologists have come to acceptthat the preserved blobs are melanosomes. EvenMichaelWuttke, the paleontologist recently retired from theGeneral Department of Cultural Heritage Rhineland-Palatinate in Germany, who originally thought they werebacteria (5), says he is now “absolutely convinced” thatmelanosomes are preserved in hairs and feathers.

However, evolutionary biologist Mary Higby Schweit-zer, of North Carolina State University in Raleigh, callsthe conclusions “overstated,” a position she admits hasmade her unpopular among most paleo-color aficio-nados. “I’mnot against melanosomes preserving,” saysSchweitzer. But, she adds, it’s “more parsimonious toassume a microbial origin until disproven with dataother than pictures.”

Rainbow of PossibilitiesResearchers have started to collect such data. JohanLindgren, a paleontologist at Lund University in Swe-den, uses time-of-flight secondary ion mass spec-trometry (ToF-SIMS) and other methods to analyzethe chemical composition of fossils. By exciting thefossil surface with an ion beam, Lindgren can mea-sure the masses of the atoms and molecules thatfly off, and he has discovered black eumelanin pig-ment itself in fossils, such as a fish eye and an

A. huxleyi specimen (8, 9). Lindgren concludes thatat least some of the microbodies in fossils are indeedmelanosomes.

Vinther, now a paleontologist at the University ofBristol, also uses ToF-SIMS; he and other scientists saythis direct evidence of the chemical signature ofmelanin pigments clinches their argument. “As far asI’m concerned, case closed,” says Briggs.

However, some critics point out that these analysismethodsmay offer amisleading picture of pigmentation.Essentially, they can only look at a limited number ofspots from a given specimen. Scientists have to chip off asmall bit of the fossil to subject it to mass spectrometry;to minimize damage they typically only look at a handfulof chips. And Vinther and colleagues studied onlyone isolated feather to decide that Archaeopteryx lith-ographica was probably black (10). This kind of extrapo-lation could bewrong, say Schweitzer and others. Imaginetrying to determine the coloration of a modern-day pea-cock from pigments at just a few dozen spots, she cau-tions: you might not come up with the right pattern.

To avoid this problem, Edwards and colleagues useX-ray analysis (which causes no damage) to probe pig-ments across an entire fossil. Along with collaboratorsat the Stanford Synchrotron Radiation Lightsource (SSRL)in Menlo Park, California, these researchers use a tech-nique called synchrotron rapid-scanning X-ray fluo-rescence that causes different elements to emit acharacteristic burst of light. In fossil organisms such asC. sanctus, the scientists see copper, which is one ofseveral metal ions that can bind to melanins, and is alsofound in tyrosinase, an enzyme that controls the produc-tion of melanins (4). Another technique, X-ray absorptionspectroscopy, revealed that the copper was bound to or-ganic molecules, which were “most probably derived fromprecursor pigment molecules,” says Roy Wogelius, a Uni-versity of Manchester geologist on the team. Based onthese techniques, the researchers concluded that the tips

Elucidating Structural ColorPigments are not the only source of animal colors. The bright hues of a butterfly’s wing and the metallicsheen of a beetle are structural colors, produced when light bounces off multilayer, nanoscale structures thatscatter the rays. Maria McNamara, of the University College Cork in Ireland, has shown that these nano-structures can occasionally survive the fossilization process (15), allowing researchers to decipher the color ofinsects that lived long ago.

McNamara and her team examined the greenish-blue fossils of 47-million-year-old forester moths byusing electron microscopy and spectrophotometry to analyze how they scatter light (16). The researchersconcluded that the moths’ wings contained multilayer structures that would have produced color in life, butthe wings probably weren’t originally greenish-blue. That’s because the fossilization process can change thearchitecture of the nanostructures, and therefore the color they produce. To simulate this process, McNa-mara exposed modern-day beetles to high temperatures and pressure in an autoclave. She found that thehighest temperatures turned their wings black, whereas lower temperatures shifted the hue toward the blueend of the spectrum (17).

Based on their measurements, McNamara and colleagues decided that the forester moth would haveactually been a matte yellowish-green, likely matching leaves in its environment. This implies that theseprehistoric moths had already evolved camouflage mechanisms to deter predators.

Meanwhile, other researchers have identified structural color in a 50-million-year-old beetle, andMcNamara and others have found it in fossil feathers (18–21). “It is an important area,” says Nick Edwards, apaleontologist at the University of Manchester in the United Kingdom. “The structural aspects to coloration . . .

can have a profound effect on how coloration can be perceived by another organism.”

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and one side of A. lithographica feathers were darklypigmented, whereas the other side was lighter (11).

Vinther and Ryan Carney, a paleontologist at theUniversity of South Florida in Tampa who worked withhim on the A. lithographica feather, are not convinced.The researchers point out that copper binds to all kindsof organic compounds, including the humic acids indecomposing biomaterials. Briggs and Schweitzer addthat copper compounds could have moved around be-tween fossilizing organisms and the surrounding rock.Nevertheless, Wogelius and Edwards argue that be-cause they map the entire fossil, they can correlate or-ganic copper complexes with the places theyexpect to see pigmentation, such as in feathers andfossil melanosomes (4, 11). “That’s why the wholeimage is so important,” says Edwards. “It’s anotherway to check whether we’re being fooled by somesort of geochemical process.”

From Color to BehaviorArmed with these discoveries, paleontologists arenow considering how color fits in with their suspicionsabout prehistoric animal behavior. For example, Vinthersuggests that the findings of colorful fossil feathers couldhelp scientists understand why feathers evolved. Thevery first proto-feather filaments were probably no goodfor flight, and paleontologists think they helped keepdinosaurs warm. But it’s a big leap from single filamentsto the branched, flat-feathers needed to soar into theprehistoric skies. Pigments seem to appear around thesame time as flat feathers, and Vinther suggests theyserved as a “billboard” for displays—to attract a mate orindicate one’s species—before they ever took creaturesairborne. Scientists already knew that birds and dino-saurs had ornamental feathers or even bony headgear,notes Chiappe, but the color findings certainly supportthe idea that those structures were for show.

Pigmentation isn’t always about flash, though. Inone study, Lindgren found dark eumelanin pigment inthree fossil reptiles: a leatherback turtle, a lizard-likeaquatic mosasaur, and an ichthyosaur (3). Lindgrensuspects the ichthyosaur was uniformly dark, like mod-ern deep-diving sperm whales, which are camouflagedby their coloration in the ocean’s depths. This theoryis supported by other evidence that the icthyosaurmight have been a deep diver, such as its large eyes

(12). Back at the surface, the dark pigment might alsohave helped the reptiles absorb heat while basking inthe sun, as it does for modern leatherbacks.

Recently, Vinther and colleagues used melanosomesto discern the identity of the mysterious Tullimonstrumgregarium fossils, first found in Illinois in the 1950s byamateur paleontologist Francis Tully. The creature’ssquishy finned body, elephantine trunk ending in eightsharp teeth, and bizarre, barbell-like midbody organearned it the nickname “Tully monster,” and paleon-tologists have long debated whether it might havebeen a mollusk, a worm, or a vertebrate. Using electronmicroscopy and ToF-SIMS, the team identified mela-nosomes in the creature’s eyes, arranged in layers typ-ical of vertebrates. The team concluded that the Tullymonster was a vertebrate, albeit an unusual one (13).

These kinds of studies demonstrate the impact thatpaleo-color is having on the wider field, says Carney:“It allows us to gain more information than just whatthey looked like; we can infer things about their func-tion and behavior.”

Beyond MelaninsMelanins are only part of the natural color palette.Other pigments, such as the plant-derived orangecarotenoids, are commonly found in modern species,from flamingoes to goldfish. (And pigments aren’talways the color source; see Elucidating StructuralColor.) But these molecules are much less stable thanmelanin pigments, and lack the heavy elements neededfor X-ray analysis at SSRL, making it challenging to findthem in fossils.

Still, it might be possible to deduce their presence.Maria McNamara, a paleobiologist at University CollegeCork in Ireland, has found a way to infer these othercolors in a 10-million-year-old, exquisitely-preservedsnake skin from Spain (14). The snakeskin is mineral-ized, its biological structures replaced by calcium phos-phate. Nonetheless, the snakeskin preserves thecharacteristic microscopic structures of individualcells. In reptiles, three distinct cell types are largely re-sponsible for producing color: melanophores makeblack; xanthophores hold red, yellow, and orange ca-rotenoids; and iridophores contain crystals that scatterlight. A combination of melanophores with iridophoresor xanthophores produces green coloration.

Based on samples taken from a leatherback turtle fossil (A) (scale bar, 10 cm), this ion image (B) (scale bar, 3 μm) showsthe spatial distribution of peaks characteristic of eumelanin (green), silicon oxide (blue), and sulfate (red) superimposedonto a scanning electron microscopy image of the “skin.” An enlargement (C) (scale bar, 300 nm) of the demarcatedarea in B (white box) shows a melanosome-like microbody. Reproduced from ref. 3, with permission from MacmillanPublishers Ltd.: Nature, copyright 2014.

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Each pigment cell type has a different shape, size,and location, soMcNamara and her team used electronmicroscopy to identify them in the fossil snakeskin, andconcluded that the creature once sported blotches ofgreen, black, and yellow-green, with a pale underbelly.“These types of blotches are really good at breaking upthe body outline for camouflage,”McNamara explains.She believes that the snake probably evaded predatorsduring the day by hiding among leaves, as manymodern snakes do.

“It’s an amazing specimen, and it really opens thedoor to expanding our palette of color reconstruction,”says Carney. McNamara suspects other fossils mightyield to her technique; fish and amphibians also havediverse types of pigment cells.

But Matthew Shawkey, an evolutionary biologist atthe University of Ghent in Belgium, is skeptical of thefindings because, he says, the fossil cells that McNamaraidentified do not look much like the modern versions,meaning they may not be pigment cells at all. Briggs,who advised McNamara during her postdoctoratestudies, suspects that the cells changed shape as theskin mineralized (as, for example, muscle cells havebeen shown to do in some fossils), and so he believesthe results are valid. Briggs doubts that the method willfind wide use, though, because it requires fossils withfinely preserved details.

Although some scientists remain to be convincedabout paleo-color studies, the public has embraced thediscoveries, a side benefit of thework that’s not lost on theresearchers. Both Edwards’ and McNamara’s team havetaken part in the United Kingdom’s Royal Society SummerScience Exhibition in London, a prestigious outreachevent, and McNamara also showcased her research atthe World Science and Technology Fair in Bangkok in2012. “Science can often be intimidating to studentsand kids,” says Carney. “Dinosaurs, especially coloredflying dinosaurs, are a great way to get them excited.”

The discoveries are certainly enticing, agrees PhilipDonoghue, a paleobiologist at the University of Bristol.But then, it’s not too surprising to find that ichthyosaurswere dark or that early birds flashed ornamental feathers.He wants more. To achieve truly novel insights, scientistsmay need to identify colors in many more fossils. Thatcould allow them to investigate the geographical distri-bution of different color patterns, how they vary within aspecies, or how colors evolved over time.

Following the initial thrill of the first paleo-color stud-ies, Donoghue says that researchers now need to posesome key questions: “What work canweput that data to?”he asks. “What hypotheses can we now test?” With anyluck, as the paleo-color field matures, such inquiries willoffer new insights into the lives of prehistoric animals,even as it brightensmuseumexhibits and children’s bookswith true-to-life colors from the distant past.

1 Zhang F, et al. (2010) Fossilized melanosomes and the colour of Cretaceous dinosaurs and birds. Nature 463(7284):1075–1078.2 Li Q, et al. (2010) Plumage color patterns of an extinct dinosaur. Science 327(5971):1369–1372.3 Lindgren J, et al. (2014) Skin pigmentation provides evidence of convergent melanism in extinct marine reptiles.Nature 506(7489):484–488.4 Wogelius RA, et al. (2011) Trace metals as biomarkers for eumelanin pigment in the fossil record. Science 333(6049):1622–1626.5 Wuttke M (1983) “Weichteil-Erhaltung” durch lithifizierte Mikroorganismen bei mittel-eozänen Vertebraten aus den Ölschiefern der“Grube Messel” bei Darmstadt. Senck Leth 64(5/6):509–527.

6 Davis PG, Briggs DEG (1995) Fossilization of feathers. Geology 23(9):783–786.7 Vinther J, Briggs DEG, Prum RO, Saranathan V (2008) The colour of fossil feathers. Biol Lett 4(5):522–525.8 Lindgren J, et al. (2012) Molecular preservation of the pigment melanin in fossil melanosomes. Nat Commun 3(5):824.9 Lindgren J, et al. (2015) Molecular composition and ultrastructure of Jurassic paravian feathers. Sci Rep 5:13520.

10 Carney RM, Vinther J, Shawkey MD, D’Alba L, Ackermann J (2012) New evidence on the colour and nature of the isolatedArchaeopteryx feather. Nat Commun 3(1):637.

11 Manning PL, et al. (2013) Synchrotron-based chemical imaging reveals plumage patterns in a 150 million year old early bird. J Anal AtSpectrom 28(7):1024–1030.

12 Motani R, Rothschild BM, Wahl W, Jr (1999) Large eyeballs in diving ichthyosaurs. Nature 402(6763):747.13 Clements T, et al. (2016) The eyes of Tullimonstrum reveal a vertebrate affinity. Nature 532(7600):500–503.14 McNamara ME, et al. (2016) Reconstructing carotenoid-based and structural coloration in fossil skin. Curr Biol 26(8):1075–1082.15 McNamara ME, Briggs DEG, Orr PJ, Noh H, Cao H (2012) The original colours of fossil beetles. Proc Biol Sci 279(1731):1114–1121.16 McNamara ME, et al. (2011) Fossilized biophotonic nanostructures reveal the original colors of 47-million-year-old moths. PLoS Biol

9(11):e1001200.17 McNamara ME, et al. (2013) The fossil record of insect color illuminated by maturation experiments. Geology 41(4):487–490.18 Parker AR, McKenzie DR (2003) The cause of 50 million-year-old colour. Proc Biol Sci 270(Suppl 2):S151–S153.19 McNamara ME, Briggs DEG, Orr PJ, Field DJ, Wang Z (2013) Experimental maturation of feathers: Implications for reconstructions of

fossil feather colour. Biol Lett 9(3):20130184.20 Vinther J, Briggs DEG, Clarke J, Mayr G, Prum RO (2010) Structural coloration in a fossil feather. Biol Lett 6(1):128–131.21 Li Q, et al. (2012) Reconstruction of Microraptor and the evolution of iridescent plumage. Science 335(6073):1215–1219.

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