Three-dimensional visualization of fossil flowers, fruits, seeds, and other plant remains using synchrotron radiation X-ray tomographic microscopy (SRXTM): new insights into Cretaceous plant diversity
Summary (3 min read)
INTRODUCTION
- THE PAST thirty to forty years have seen significant advancesin understanding patterns of structural diversification during the early phases of angiosperm evolution.
- Over time technical advances in SEM also improved resolution in the routine microscopy of surface features.
- Two-dimensional sections can be constructed in multiple orientations, 3-D reconstructions can be created and manipulated, and complex specimens, such as flowers, can be dissected digitally rather than physically.
- In this paper the authors provide an overview of the SRXTM techniques applied so far to understand and visualize the detailed structure of Cretaceous fossil flowers and other plant mesofossils.
FOSSIL MATERIAL
- The first substantial discovery of a Cretaceous mesofossil assemblage containing well-preserved flowers was from fluviatile-lacustrine sediments of late Santonian to early Campanian age at the Åsen locality, Southern Sweden (Friis and Skarby, 1981).
- Intermediate in size between the larger fossils that have typically been the focus of Cretaceous paleobotanical research, and fossil pollen and spores , the flowers, fruits, seeds and other fossils that comprise mesofossil assemblages rarely exceed more than a few millimeters in length.
- Comparable assemblages of Cenozoic age, prepared using the same techniques, typically contain fossils with a much wider range of sizes (Tiffney, 1984; Eriksson et al., 2000a), are usually preserved as lignite rather than charcoal, and rarely contain fossil flowers (Friis et al., 2011).
- In the process of charcoalification, the incomplete combustion of plant material under conditions of reduced oxygen resulted in excellent preservation of the 3-D form and cellular detail of diverse plant parts.
- The application of the new technique to charcoalified material has allowed mesofossils to be examined with an unprecedented level of detail and is advancing their understanding of Cretaceous plant diversity in substantial ways.
SRXTM TECHNIQUES
- Synchrotron radiation hard X-ray tomographic microscopy represents a great advance over the application of conventional X-ray approaches in paleontology and has proved a powerful technique for the non-destructive investigation of internal structure in a variety of optically opaque samples.
- Unlike SEM no coating is required, and unlike with conventional sectioning of living material dehydrating, fixing or staining of the specimen is not needed.
- Data from 2-D radiographs provide useful but cumulative Downloaded From: https://bioone.org/journals/Journal-of-Paleontology on 30 May 2022 Terms of Use: https://bioone.org/terms-of-use information on the internal structure of the specimen along the beam path.
- When the analyzed sample is made of light elements, or elements with a similar atomic number Z, contrast is instead obtained by exploiting the refraction of the X-ray beam at material boundaries in the study object and the resulting interference phenomena.
- This permits clear visualization of internal boundaries, although the actual contrast between regions with different composition is not improved compared to standard absorption contrast tomography.
RECENT DEVELOPMENTS IN SRXTM TECHNIQUES
- —For parallel beam geometry, the available field of view is determined by the optical configuration chosen and the resolution that is required.
- When expansion of the available field of view in the direction parallel to the rotation axis is desirable, but high resolution is still required, a stack of several independent tomographic scans can be acquired.
- New developments in detector technology (e.g., sCMOS), as well as improvements in scintillating materials, have significantly improved the signal-to-noise ratio in tomographic datasets and resulted in improved density resolution.
- Downloaded From: https://bioone.org/journals/Journal-of-Paleontology on 30 May 2022 Terms of Use: https://bioone.org/terms-of-use shortcomings, but techniques for fusing and integrating the information provided by the different contrasts are also being developed.
- High quality scintillators and digital cameras it is difficult to completely avoid rings, one of the most common class of artifacts in reconstructed tomographic slices.
SRXTM COMPARED TO LABORATORY X-RAY SOURCES
- The very significant photon density reached by third generation synchrotrons brings huge advantages compared to traditional X-ray laboratory sources, especially when exceptional spatial, temporal and density resolution is required.
- The high brilliance of synchrotron light allows increased spatial and temporal resolution, including the routine and rapid detection of details as small as 1 lm in millimeter-sized samples.
- Throughput times of only a few minutes also permit a large amount of material to be scrutinized in a short time enabling the selection of the most significant and best-preserved specimens for more detailed analysis.
- The monochromaticity of the X-ray beam also enables quantitative measurements of material properties and easier identification of different phases, if the radiation energy is properly tuned, since beam hardening artifacts, which are often characteristic of laboratory X-ray sources, can be avoided.
- Unlike laboratory based X-ray CT, the coherence of synchrotron light allows SRXTM to exploit both absorption and phase contrast imaging, enabling optimal investigation of both low- and high-absorbing samples.
SEM COMPARED TO X-RAY APPROACHES
- In SEM the signal is the result of the interaction of the electron beam with atoms at or near the surface of the sample.
- In the most common or standard detection mode, secondary electrons are measured.
- Chemical information can be obtained, if the back-scattered secondary electrons (BSE) are detected, since their signal is strongly related to the atomic Downloaded From: https://bioone.org/journals/Journal-of-Paleontology on 30 May 2022 Terms of Use: https://bioone.org/terms-of-use number, but in all cases, the information retrieved from SEM studies is limited to data from the surface of the specimen.
- Their depth of penetration has been widely exploited in paleontology.
- Initial efforts using X-rays were limited in their resolution, but new approaches using SRXTM provide both high penetration and high resolution to reveal fine details of internal structure.
APPLICATION OF SRXTM TECHNIQUES TO CRETACEOUS FOSSIL PLANTS
- To fully understand the structure and organization of fossil flowers and other mesofossils it is necessary to study both external and internal features.
- In the case of flowers that are open external morphology and the general position of floral parts, and their number, can often be studied using SEM alone, especially when multiple specimens are available.
- A further problem with conventional sectioning is that many critical fossils are unique, for instance with a type specimen (e.g., holotype of Silvianthemum suecicum; Fig. 2) or where there is only one specimen of a taxon (e.g., holotype and only specimen of Monetianthus mirus Friis et al., 2009; Fig. 5).
- SRXTM allows investigation of external and internal features at high resolution, provides 3-D reconstructions, allows 2-D sections of the same specimen to be prepared in all directions and creates the possibility of virtual dissections.
CONCLUSIONS
- SRXTM has already been informative where it has been deployed in paleobotany, but the full possibilities of these techniques are still relatively underexplored.
- New methods of pre-preparation will need to be devised if such specimens are to yield useful information with SRXTM.
- Nevertheless, the capability of SRXTM studies to provide details of internal structure in a new way from a great range of fossil material significantly extends the structural and anatomical information that can potentially be obtained from fossil plants.
- In particular, it largely shifts the efforts of the investigator from preparation work in the laboratory to analysis and visualization on the computer.
- The coupling of higher magnification microscope objectives with thinner scintillators could push the spatial resolution to the submicron regime.
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Citations
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...Volumes can be assessed using advanced methods such as x-ray computed tomography (CT) on fruits (Stuppy et al., 2003) or synchrotron radiation x-ray tomographic microscopy applied in paleobiological studies (e.g. on fruits and seed; Friis et al., 2014)....
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...Direct measurement of a single seed volume is not easy to achieve, and published data are based mainly on sophisticated methods such as x-ray CT (Stuppy et al., 2003; Friis et al., 2014)....
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...SRXTM has been used to image minute charcoalified seeds and flowers from the Cretaceous Period [77,78], which resemble Cooksonia in their preservation, size fraction and in their rarity....
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References
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...Reconstruction approaches that are being developed in the medical field to overcome scan deterioration when a metal implant cannot be avoided (Prell et al., 2010; Boas and Fleischmann, 2011) may be helpful in these cases....
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271 citations
"Three-dimensional visualization of ..." refers methods in this paper
...Comparable assemblages of Cenozoic age, prepared using the same techniques, typically contain fossils with a much wider range of sizes (Tiffney, 1984; Eriksson et al., 2000a), are usually preserved as lignite rather than charcoal, and rarely contain fossil flowers (Friis et al., 2011)....
[...]
...Comparable assemblages of Cenozoic age, prepared using the same techniques, typically contain fossils with a much wider range of sizes (Tiffney, 1984; Eriksson et al., 2000a), are usually preserved as lignite rather than charcoal, and rarely contain fossil flowers (Friis et al....
[...]
261 citations
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...Extensive datasets on fossil plants have been accumulated from SRXTM analyses at the Tomcat beamline (Stampanoni et al., 2006) at the Swiss Light Source (SLS) covering material from the Carboniferous (Scott et al....
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...Extensive datasets on fossil plants have been accumulated from SRXTM analyses at the Tomcat beamline (Stampanoni et al., 2006) at the Swiss Light Source (SLS) covering material from the Carboniferous (Scott et al., 2009), Permian (Slater et al., 2011), Cretaceous (Friis et al., 2007; von Balthazar…...
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247 citations
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...A suite of algorithms for the mitigation of rings and other artifacts are continuously being developed and several software routines based on different approaches (Sijbers and Postnovz, 2004; Boin and Haibel, 2006; Titarenko et al., 2010) are available for clearing such artifacts from reconstructed slices....
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207 citations
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...Although most features indicate affinity with Nymphaeales the systematic position of Carpestella lacunata is not fully resolved (von Balthazar et al., 2008)....
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...The early presence of the Nymphaeales in angiosperm diversification is supported by macrofossil occurrences such as the compression fossils of Pluricarpellatia peltata B. Mohr, Bernardes-de-Oliveira and David W. Taylor (2008) from the Early Cretaceous of Brazil....
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...These characters confirmed the earlier placement of the fossil flower in the Nymphaeales and Monetianthus mirus remains the oldest well-documented floral record of this basal lineage of angiosperms....
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...Also, in Illicium the single ovule fills out the whole ovary cavity while in Nymphaeales this is not the case....
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...Monetianthus mirus: the recognition of Nymphaeales in the Early Cretaceous....
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Frequently Asked Questions (2)
Q2. What future works have the authors mentioned in the paper "Three-dimensional visualization of fossil flowers, fruits, seeds, and other plant remains using synchrotron radiation x-ray tomographic microscopy (srxtm): new insights into cretaceous plant diversity" ?
SRXTM has already been informative where it has been deployed in paleobotany, but the full possibilities of these techniques are still relatively underexplored. The combination of SRXTM with the availability of a large numbers of diverse and well-preserved specimens offers the possibility of a new phase of rapid progress in their understanding of Cretaceous and other fossil plants. In many cases, the critical details revealed by the application of SRXTM have created opportunities to compare the fine structure of fossils with those of extant taxa, raising the possibility of also using SRXTM to study complex 3-D structures in living plants. This kind of material requires higher energies, and particularly for larger specimens where lateral merging and vertical stacking is required the acquisition time may be extensive.