Soft tissue preservation

Much of my current work involves understanding how soft tissues preserve in exceptional fossils and what biomolecules persist deep into the geologic record. Doing so will lead to better interpretations of fossil data and a better understanding of the biology of extinct organisms. My work involves experimental taphonomy utilizing decay and maturation experiments as well as modern analytical techniques on fossils themselves. These projects are highly collaborative in nature given their span of paleontology, geology, biology, and chemistry.

My first foray into this subject was during my masters thesis. This thesis is available through my Researchgate page. The thesis was also presented at the 2016 Society of Vertebrate Paleontology annual meeting. The slides from the talk are also available through Researchgate. I also presented the thesis in a poster at the 2015 annual meeting of the Palaeontological Association (again available through Researchgate). The first paper to come from my masters thesis, showing through decay and maturation experiments common to experimental taphonomy that keratin is not likely to preserve in fossils of appreciable age or burial depth, was published in Palaeontology:

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Pyrolysis GC-MS results of thermally matured feathers

Saitta, E. T., Rogers, C., Brooker, R. A., Abbott, G. D., Kumar, S., O’Reilly, S. S., Donohoe, P., Dutta, S., Summons, R.E., & Vinther, J. (2017). Low fossilization potential of keratin protein revealed by experimental taphonomy. Palaeontology60(4), 547-556.

Abstract

Recent studies have suggested the presence of keratin in fossils dating back to the Mesozoic. However, ultrastructural studies revealing exposed melanosomes in many fossil keratinous tissues suggest that keratin should rarely, if ever, be preserved. In this study, keratin’s stability through diagenesis was tested using microbial decay and maturation experiments on various keratinous structures. The residues were analysed using pyrolysis-gas chromatography-mass spectrometry and compared to unpublished feather and hair fossils and published fresh and fossil melanin from squid ink. Results show that highly matured feathers (200–250°C/250 bars/24 h) become a volatile-rich, thick fluid with semi-distinct pyrolysis compounds from those observed in less degraded keratins (i.e. fresh, decayed, moderately matured, and decayed and moderately matured) suggesting hydrolysis of peptide bonds and potential degradation of free amino acids. Neither melanization nor keratin (secondary) structure (e.g. ⍺- vs β-keratin) produced different pyrograms; melanin pyrolysates are largely a subset of those from proteins, and proteins have characteristic pyrolysates. Analyses of fossil fur and feather found a lack of amides, succinimide and piperazines (present even in highly matured keratin) and showed pyrolysis compounds more similar to fossil and fresh melanin than to non-matured or matured keratin. Although the highly matured fluid was not water soluble at room temperature, it readily dissolved at elevated temperatures easily attained during diagenesis, meaning it could leach away from the fossil. Future interpretations of fossils must consider that calcium phosphate and pigments are the only components of keratinous structures known to survive fossilization in mature sediments.

A second paper deriving from my masters thesis discusses the early taphonomic changes that might occur in keratinous structures and whether the resulting textures would ever be expected in the fossil record. It also challenges structures identified as dinosaur erythrocytes, suggesting that they are taphonomic artifacts of degraded organic material. This paper was published in Palaios and reported by Phys.org and the University of Bristol. It also formed the basis of an article I wrote for The Conversation.

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Structures in moderately matured dermis (above) match those interpreted to be dinosaur blood cells (below) in shape and in their range of shapes

Saitta, E. T., Rogers, C. S., Brooker, R. A., & Vinther, J. (2017). Experimental taphonomy of keratin: a structural analysis of early taphonomic changes. Palaios32(10), 647-657.

Abstract

The evolution of integumentary structures, particularly in relation to feathers in dinosaurs, has become an area of intense research. Our understanding of the molecular evolution of keratin protein is greatly restricted by the fact that such information is lost during diagenesis and cannot be derived from fossils. In this study, decay and maturation experiments are used to determine if different keratin types or integumentary structures show different patterns of degradation early in their taphonomic histories and if such simulations might advance our understanding of different fossilization pathways. Although different distortion patterns were observed in different filamentous structures during moderate maturation and ultrastructural textures unique to certain types of scales persisted in moderate maturation, neither of these have been observed in fossils. It remains uncertain whether these degradation patterns would ever occur in natural sediment matrix, where microbial and chemical decay happens well before significant diagenesis. It takes some time for remains to be buried, meaning that keratin may not be left for moderate maturation to produce such patterns. Higher, more realistic maturation conditions produce a thick, and water soluble fluid that lacks all morphological and ultrastructural details of the original keratin, suggesting that such textural or distortion patterns are unlikely to be preserved in fossils. Although different degradation patterns among keratinous structures are intriguing, it is unlikely that such information could be recorded in the fossil record. Calcium phosphates and pigments are the surviving components of integumentary structures. Thus, the results here are likely of more relation to the archaeological record than fossil record. Other noteworthy results of these experiments are that melanin may not be the leading factor in determining the rate of microbial decay in feathers but may reduce the rate of degradation from maturation, that the existence of rachis filamentous subunits similar to plumulaceous barbules is supported, and that previously reported dinosaur ‘erythrocytes’ may be taphonomic artifacts of degraded organic material.

I was coauthor of a paper providing an overview of the various processes involved in fossilization, highlighting how decay is far from the only factor to consider when studying fossil taphonomy or designing experiments. The paper was published in BioEssays.

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Exceptionally preserved fossils. A) Sinosauropteryx prima (NIGP 127586), a feathered dinosaur from the Cretaceous Jehol Biota preserving melanized tissues (feathers, eyes, and abdominal organs). B) Aquilonifer spinosus (OUMNH C.29695), a Silurian arthropod preserved in three dimensions in volcanic ash-hosted carbonate concretions from Herefordshire. Image at left shows a surface captured during serial grinding, image at right shows a three dimensional reconstruction from serial photographs.[112] C) Belemnotheutis antiquus(NHMUK 25966), a Jurassic stem group decabrachian (belemnoid) from Christian Malford, Wiltshire, UK, preserving creamy colored musculature replaced by calcium phosphate and organic arm hooks. D) Fossil Anolis lizard preserved in Miocene Dominican amber.[113] Image at left is a photograph of specimen, image at right shows 3D reconstruction using micro CT. E) Haootia quadriformis, a possible medusozoan from the Ediacaran of Newfoundland. F) Pyritised specimens of the trilobite Triarthrus eatoni (ROM 62891), with preserved limbs from the Late Ordovician Beecher’s Trilobite Bed, New York, State. Image credits to the authors, except C (Jonathan Jackson, NHM) D (Russell Garwood and Emma Sherratt) E (Alex Liu), F (David Rudkin).

Parry, L. A., Smithwick, F., Norden, K., Saitta, E. T., Lozano-Fernandez, J., Tanner, A., Bernard Caron, J., Edgecombe, G. D., Briggs, D. E. G., & Vinther, J. (2017). Soft-bodied fossils are not simply rotten carcasses—towards a holistic understanding of exceptional fossil preservation. BioEssays. doi: 10.1002/bies.201700167

Abstract

Exceptionally preserved fossils are the product of complex interplays of biological and geological processes including burial, autolysis and microbial decay, authigenic mineralization, diagenesis, metamorphism, and finally weathering and exhumation. Determining which tissues are preserved and how biases affect their preservation pathways is important for interpreting fossils in phylogenetic, ecological, and evolutionary frameworks. Although laboratory decay experiments reveal important aspects of fossilization, applying the results directly to the interpretation of exceptionally preserved fossils may overlook the impact of other key processes that remove or preserve morphological information. Investigations of fossils preserving non-biomineralized tissues suggest that certain structures that are decay resistant (e.g., the notochord) are rarely preserved (even where carbonaceous components survive), and decay-prone structures (e.g., nervous systems) can fossilize, albeit rarely. As we review here, decay resistance is an imperfect indicator of fossilization potential, and a suite of biological and geological processes account for the features preserved in exceptional fossils.

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‘Synthetically fossilized’ lizard head

One project involves improved equipment and methods for maturation experiments using a large-scale, sediment-based approach with the goal of modeling fossilization processes to create ‘synthetic fossils’ in the lab, in an analogous manner to the production of synthetic diamonds. Such a technique holds great potential for better understanding fossilization and testing a wide range of hypotheses for many years to come. The initial results were presented in a pre-conference, informal series of talks prior to the 2016 annual meeting of the Palaeontological Association. I also presented this work during the Romer session of the 2017 annual meeting of the Society of Vertebrate Paleontology. The method was mentioned by my collaborator, Tom Kaye, in the early part of a podcast interview with Palaeocast. The first paper from this project is under review.

The paper itself was published in Palaeontology. The work was covered in a variety of media outlets including Science.

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Saitta, E. T., Kaye, T. G., & Vinther, J. (2018). Sediment-encased maturation: a novel method for simulating diagenesis in organic fossil preservation. Palaeontology. doi: 10.1111/pala.12386

Abstract

Exceptional fossils can preserve diagenetically‐altered biomolecules. Understanding the pathways that lead to such preservation is vital to utilizing fossil information in evolutionary and palaeoecological studies. Experimental taphonomy explores the stability of tissues during microbial/autolytic decay or their molecular stability through maturation under high pressure and temperature. Maturation experiments often take place inside sealed containers, preventing the loss of labile, mobile or volatile molecules. However, wrapping tissues inside aluminium foil, for example, can create too open a system, leading to loss of both labile and recalcitrant materials. We present a novel experimental procedure for maturing tissues under elevated pressure/temperature inside compacted sediment. In this procedure, porous sediment allows maturation breakdown products to escape into the sediment and maturation chamber, while recalcitrant, immobile components are contained, more closely mimicking the natural conditions of fossilization. To test the efficacy of this procedure in simulating fossil diagenesis, we investigate the differential survival of melanosomes relative to proteinaceous tissues through maturation of fresh lizard body parts and feathers. Macro‐ and ultrastructures are then compared to fossils. Similar to many carbonaceous exceptional fossils, the resulting organic components are thin, dark films composed mainly of exposed melanosomes resting on the sediment in association with darkened bones. Keratinous, muscle, collagenous and adipose tissues appear to be lost. Such results are consistent with predictions derived from non‐sediment‐encased maturation experiments and our understanding of biomolecular stability. These experiments also suggest that organic preservation is largely driven by the original molecular composition of the tissue and the diagenetic stability of those molecules, rather than the tissue’s decay resistance alone; this should be experimentally explored in the future.

I am currently working on several ongoing projects. I am performing a series of chemical analyses on various fossils of keratinous structures to determine what components of these structures survive into the fossil record.

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Dinosaur Provincial Park, Alberta, Canada – Aseptic collection of dinosaur bones

Similarly, I am using a variety of chemical analyses to understand the organics present in fossil dinosaur (e.g., Mesozoic) bone. This work involves samples that I aseptically excavated from Dinosaur Provincial Park in the summer of 2016. I wish to understand whether or not these organics are ancient, endogenous, and/or well-preserved. I presented a poster on this topic entitled “Life inside a dinosaur bone: a thriving microbiome” at the 2017 annual meeting of the Palaeontological Society. We have published a preprint of the manuscript in bioRxiv. The preprint was covered in a piece by The Atlantic.

Saitta, E. T., Liang, R. Lau, C. Y., Brown, C. M., Longrich, N. R., Kaye, T. G., Novak, B. J., Salzberg, S., Donohoe, P., Dickinson, M., Vinther, J., Bull, I. D., Brooker, R. A., Martin, P., Abbott, G. D., Knowles, T. D. J., Penkman, K., & Onstott, T. C. (2018). Life Inside A Dinosaur Bone: A Thriving Microbiome. bioRxiv, 400176. doi: 10.1101/400176

Abstract

Fossils were long thought to lack original organic material, but the discovery of organic molecules in fossils and sub-fossils, thousands to millions of years old, has demonstrated the potential of fossil organics to provide radical new insights into the fossil record. How long different organics can persist remains unclear, however. Non-avian dinosaur bone has been hypothesised to preserve endogenous organics including collagen, osteocytes, and blood vessels, but proteins and labile lipids are unstable during diagenesis or over long periods of time. Furthermore, bone is porous and an open system, allowing microbial and organic flux. Some of these organics within fossil bone have therefore been identified as either contamination or microbial biofilm, rather than original organics. Here, we use biological and chemical analyses of Late Cretaceous dinosaur bones and sediment matrix to show that dinosaur bone hosts a diverse microbiome. Fossils and matrix were freshly-excavated, aseptically-acquired, and then analysed using microscopy, spectroscopy, chromatography, spectrometry, DNA extraction, and 16S rRNA amplicon sequencing. The fossil organics differ from modern bone collagen chemically and structurally. A key finding is that 16S rRNA amplicon sequencing reveals that the subterranean fossil bones host a unique, living microbiome distinct from that of the surrounding sediment. Even in the subsurface, dinosaur bone is biologically active and behaves as an open system, attracting microbes that might alter original organics or complicate the identification of original organics. These results suggest caution regarding claims of dinosaur bone soft tissue preservation and illustrate a potential role for microbial communities in post-burial taphonomy.

In the process of running these analyses, I have also started to investigate the organics of fossil eggshells. Preliminary results were presented at the 2017 International Workshop on Konservat-Lagerstätten in Cork, Ireland.

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Tom Kaye’s lab, Sierra Vista, Arizona – Examining ‘synthetic fossils’ under laser fluorescence