Figure 1. EDS spectrum of framboid. EDS spectrum of framboid showing an iron-oxygen signature. Pt is from coating for SEM. Area in red box was scanned for elements. [larger view].
Some of you might remember a paper published in Science that rocked the paleontological world by revealing that a broken thigh bone from Tyrannosaurus rex contained soft tissue. When this soft tissue was analyzed, it was identified as collagen from the blood vessels and, because it was found inside a dinosaur fossil, it was assumed to be ancient dinosaur tissue. Analysis of the protein sequence of this collagen indicated that chickens are the closest living relatives of this iconic dinosaur. However, a research team from my alma mater, the University of Washington in Seattle, just published a paper that questions whether this soft tissue really is preserved dinosaur tissue. After meticulous analysis, they hypothesize that this biological material might actually be a bacterial “biofilm” instead.
According to the 2007 research teams led by Mary Schweitzer from North Carolina State University in Raleigh, and John Asara of Harvard Medical School and Beth Israel Deaconess Medical Center in Boston, reconstructed amino acid sequences of a proteinaceaus substance found in a fractured Tyrannosaurus rex thigh bone bore striking similarities to collagen found in chickens, while several other protein strands were similar to amphibian collagen.
However, paleontologist Thomas Kaye of the University of Washington, and his research team contend that what was really inside the T. rex bone was not collagen from the dinosaur’s blood vessels, but instead, it was a slimy biofilm produced by bacteria that lived within the spaces once occupied by blood vessels and cells.
A bacterial biofilm is a micro-ecosystem consisting mainly of a community of diverse types of bacteria that produce a polymeric matrix that sticks them to a living or inert surface. One familiar example of a biofilm is that layer of slime that grows on a person’s teeth overnight.
Kaye compares this to what happens if you leave a pail of rainwater sitting in your backyard. Basically, after sitting for several weeks, the inner walls of the bucket feel slippery from the bacterial film that has grown on its inner walls. After time, this biofilm hardens through mineralization and takes on the shape of the surface it grew on. This is the same process that causes the formation of hardened deposits known as calculus on a person’s teeth.
“We are not experts in the field,” cautions Kaye. “We are not disagreeing with the fact that their instruments detected protein. We are offering an alternative explanation” for the source of this material.
When he began this work, Kaye’s goal was to be the second person to find preserved dinosaur tissue, although he discovered something far different. In short, Kaye was doing what scientists often do: he was trying to duplicate Schweitzer’s and Asara’s findings. To do this, he went to the same formation where Schweitzer’s sample came from, dug up a 65-million-year-old fossilized bone from a turtle (figure 2), cracked it open, and looked at what was inside using a Scanning Electron Microscope (SEM) (figure 3);
Figure 2. Well preserved complete bone used in initial investigation. Exceptionally well preserved small phalange from the Lance formation used for initial survey. No cracks or deformities present. Specimen was pressure fractured and directly examined under the SEM. UWBM 89327 Scale bar, 10 mm. [larger view].
Figure 3. Iron oxide framboids. An iron oxide framboid cluster in dinosaur trabecular bone found commonly throughout time and taxa. At approximately 10 microns in diameter they are closely matched in size to red blood cells and typical pyrite framboids. UWBM 89327 Scale bar, 3 μm. [larger view].
Kaye’s team was surprised when SEM revealed that these supposedly rare iron-containing structures were very common. What previously had been identified as fragments of blood cells due to the presence of iron were actually microscopic spheres containing iron, known as framboids.
The team found similar spheres in a variety of other fossils from different time periods. Interestingly, they also found iron-containing spheres in an ammonite fossil, a marine mollusc whose closest living relatives include the squid and octopus. These spheres were observed in a cavity within the ammonite fossil where blood had never been, so the residual iron could not have had any relationship to the presence of blood (figure 4A);
Figure 4. Tubular branching structures. Branching, transparent tube-like structures that match the porosity of the trabecular bone. Note small red grains that were found to be iron oxide framboids. These structures remain in acid baths after demineralization. Some are pliable, others frangible. Scale bars, 100 μm. Photos Z stacked, 7 images, unsharp mask, gamma adjusted.
After subjecting several fossils to demineralizing chemical baths as described in the original work, the team found a sediment consisting of a variety of branching structures that resemble capillaries (figure 4A-C). Some of these structures were soft and pliable while others were brittle and easily fractured, as described in Schweitzer’s and Asara’s research. These structures were then analyzed using energy dispersive spectroscopy (EDS).
Kaye’s team then examined a variety of other bones, including a turtle’s, and found similar structures. They also analyzed collagen and a modern biofilm that grew on microscope slides that they had placed into a nearby pond with a high iron content, and found that the fossil proteinaceous material more closely resembled slime from pond water (figure 9);
Figure 9. Infrared spectral comparison. Infrared spectra showing similarity of modern biofilms and modern collagen compared to fossil coatings. Cross correlation shows that the fossil material more closely resembles the modern biofilm than the modern collagen. [larger view].
The team also used SEM to examine some of the demineralized structures they recovered from the acid baths and found that they resemble decomposing bacteria
Rhodococcus sp. This species of bacteria has a variety of microscopic shapes, ranging from small, round cocci to long filaments (figure 10);
Figure 10. Osteocytes and lacunae. (A and B) Osteocytes found floating free in acid baths with fillapodia. (C,D,E) Fractured lacunae examined with SEM show filaments and spheres consistent with bacterial forms. UWBM 89325, UWBM 89322 Scale bars, A,B 10 μm, C-E 1 μm.
“We determined that these structures were too common to be exceptionally preserved tissue,” observed Kaye. “We realized it couldn’t be a one-time exceptional preservation,” contrary to Schweitzer’s and Asara’s argument. This led Kaye’s team to investigate the possibility that bacteria might be the source for this biological material.
As the result of their meticulous investigation, the team concluded that the voids in dinosaur bone provide the micro-environmental equivalent of a natural cave where bacteria can colonize and form biofilms. Further, when the cavity that biofilms have grown on is subsequently removed, as with an acid bath, the remaining biofilm retains much of the original morphology of the surface it originally grew on. This is the likely explanation for the quantity and similarity of structures found in fossil bone. This also indicates that these structures are unlikely to be preserved dinosaurian tissues but instead, are the product of common bacterial growth.
“From this evidence, we could determine that what had previously been reported as dinosaurian soft tissues were in fact biofilms, or slime,” Kaye concluded.
Kaye, T.G., Gaugler, G., Sawlowicz, Z., Stepanova, A. (2008). Dinosaurian Soft Tissues Interpreted as Bacterial Biofilms. PLoS ONE, 3(7), e2808. DOI: 10.1371/journal.pone.0002808.