Three‐dimensional virtual histology of silurian osteostracan scales revealed by synchrotron radiation microtomography

We used propagation phase contrast X‐ray synchrotron microtomography to study the three‐dimensional (3D) histology of scales of two osteostracans, Tremataspis and Oeselaspis, members of a jawless vertebrate group often cited as the sister group of jawed vertebrates. 3D‐models of the canal systems and other internal structures are assembled based on the virtual thin section datasets and compared with previous models based on real thin sections. The primary homology framework of the canal systems in the two taxa is revised and new histological details are revealed based on the results of this work. There is no separation of vascular canals and lower mesh canals in the Tremataspis scale, contrary to previous results. The secondary upper mesh canals have a limited distribution to the anterior region of the Tremataspis scale. The upper and lower mesh canal systems of Tremataspis have different geometries, inferred to reflect different developmental origins: we interpret the upper system as a probable epithelial invagination, the lower system as entirely vascular. Oeselaspis has no equivalent of the upper mesh canal system. The upper mesh canal system of Tremataspis may have been sensory in function. In Oeselaspis, numerous polyp‐shaped structures opening from the canal system onto the surface of the scale resemble the innervation tracts for neuromast organs. The growth of the Oeselaspis scale proceeds by addition of small odontodes containing unmineralized lacunae, which may further mineralize and become more compact. Our results highlight that 3D‐histological investigation on scales and other dermal skeletons of osteostracans is necessary to fully appreciate the diversity of skeletal histologies in the group. Traditional 3D‐models based on thin sections alone are not reliable and should no longer be used as the basis for homology assessments or functional hypotheses. J. Morphol. 276:873–888, 2015. © 2015 Wiley Periodicals, Inc.     

Linking evolution and development: Synchrotron Radiation X‐ray tomographic microscopy of planktic foraminifers

Abstract: Making the link between evolutionary processes and development in extinct organisms is usually hampered by the lack of preservation of ontogenetic stages in the fossil record. Planktic foraminifers, which grow by adding chambers, are an ideal target organism for such studies as their test incorporates all prior developmental stages. Previously, studies of development in these organisms were limited by the small size of their early chambers. Here, we describe the application of synchrotron radiation X‐ray tomographic microscopy (SRXTM) to document the ontogenetic history of the foraminifers Globigerinoides sacculifer and Globorotalia menardii. Our SRXTM scans permit resolution at submicrometre scale, thereby displaying additional internal structures such as pores, dissolution patterns and complexity of the wall growth. Our methods provide a powerful tool to pick apart the developmental history of these microfossils and subsequently assist in inferring phylogenetic relationships and evolutionary processes.     

Different speciation for bromine in brown and red algae,revealed by in vivo X‐ray absorption spectroscopic studies

Members of various algal lineages are known to be strong producers of atmospherically relevant halogen emissions, that is a consequence of their capability to store and metabolize halogens. This study uses a noninvasive, synchrotron‐based technique, X‐ray absorption spectroscopy, for addressing in vivo bromine speciation in the brown algae Ectocarpus siliculosus, Ascophyllum nodosum, and Fucus serratus, the red algae Gracilaria dura, G. gracilis, Chondrus crispus, Osmundea pinnatifida, Asparagopsis armata, Polysiphonia elongata, and Corallina officinalis, the diatom Thalassiosira rotula, the dinoflagellate Lingulodinium polyedrum and a natural phytoplankton sample. The results highlight a diversity of fundamentally different bromine storage modes: while most of the stramenopile representatives and the dinoflagellate store mostly bromide, there is evidence for Br incorporated in nonaromatic hydrocarbons in Thalassiosira. Red algae operate various organic bromine stores – including a possible precursor (by the haloform reaction) for bromoform in Asparagopsis and aromatically bound Br in Polysiphonia and Corallina. Large fractions of the bromine in the red algae G. dura and C. crispus and the brown alga F. serratus are present as Br^? defects in solid KCl, similar to what was reported earlier for Laminaria parts. These results are discussed according to different defensive strategies that are used within algal taxa to cope with biotic or abiotic stresses.     

Support and challenges to the melanosomal casing model based on nanoscale distribution of metals within iris melanosomes detected by X‐ray fluorescence analysis

Melanin within melanosomes exists as eumelanin or pheomelanin. Distributions of these melanins have been studied extensively within tissues, but less often within individual melanosomes. Here, we apply X‐ray fluorescence analysis with synchrotron radiation to survey the nanoscale distribution of metals within purified melanosomes of mice. The study allows a discovery‐based characterization of melanosomal metals, and, because Cu is specifically associated with eumelanin, a hypothesis‐based test of the ‘casing model’ predicting that melanosomes contain a pheomelanin core surrounded by a eumelanin shell. Analysis of Cu, Ca, and Zn shows variable concentrations and distributions, with Ca/Zn highly correlated, and at least three discrete patterns for the distribution of Cu vs. Ca/Zn in different melanosomes – including one with a Cu‐rich shell surrounding a Ca/Zn‐rich core. Thus, the results support predictions of the casing model, but also suggest that in at least some tissues and genetic contexts, other arrangements of melanin may co‐exist.     

Lipids status and copper in a single astrocyte of the rat model for amyotrophic lateral sclerosis: Correlative synchrotron‐based X‐ray and infrared imaging

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease, causing death of motor neurons controlling voluntary muscles. The pathological mechanisms of the disease are only partially understood. The hSOD1‐G93A ALS rat model is characterized by an overexpression of human mutated SOD1, causing increased vulnerability by forming intracellular protein aggregates, inducing excitotoxicity, affecting oxidative balance and disturbing axonal transport. In this study we followed the bio‐macromolecular organic composition and compartmentalization together with trace metal distribution in situ in single astrocytes from the ALS rat model and compared them to the control astrocytes from nontransgenic littermates by simultaneous use of two synchrotron radiation‐based methods: Fourier transform infrared microspectroscopy (SR‐FTIR) and hard X‐ray fluorescence microscopy (XRF). We show that ALS cells contained more Cu, which colocalized with total lipids, increased carbonyl groups and oxidized lipids, thus implying direct involvement of Cu in oxidative stress of lipidic components without direct connection to protein aggregation in situ.     

Lithiation‐Induced Dilation Mapping in a Lithium‐Ion Battery Electrode by 3D X‐Ray Microscopy and Digital Volume Correlation

Recent advances in high‐resolution 3D X‐ray computed tomography (CT) allow detailed, non‐destructive 3D structural mapping of a complete lithium‐ion battery. By repeated 3D image acquisition (time lapse CT imaging) these investigations of material microstructure are extended into the fourth dimension (time) to study structural changes of the device in operando. By digital volume correlation (DVC) of successive 3D images the dimensional changes taking place during charge cycling are quantified at the electrode level and at the Mn₂O₄ particle scale. After battery discharging, the extent of lithiation of the manganese (III/IV) oxide grains in the electrode is found to be a function of the distance from the battery terminal with grains closest to the electrode/current collector interface having the greatest expansion (≈30%) and grains furthest from the current collector and closest to the counter electrode showing negligible dilation. This implies that the discharge is limited by electrical conductivity. This new CT+DVC technique is widely applicable to the 3D exploration of the microstructural degradation processes for a range of energy materials including fuel cells, capacitors, catalysts, and ceramics.     

Vibrational Circular Dichroism (VCD), VCD Exciton Coupling,and X‐ray Determination of the Absolute Configuration of an α,β‐Unsaturated Germacranolide

The absolute configuration of 1 was deduced by vibrational circular dichroism together with the evaluation of the Flack and Hooft X‐ray parameters. Vibrational circular dichroism exciton coupling, using the carbonyl group signals, confirmed the absolute configuration of 2 . In addition, sodium borohydride reduction of the 11,13‐double bond of 6‐epi‐desacetyllaurenobiolide ( 1 ) yields an almost equimolecular mixture of C11 epimers, while reduction of the same double bond of 6‐epi‐laurenobiolide ( 2 ) provided almost exclusively the (11S) diastereoisomer 4 . Chirality 27:247–252, 2015. © 2015 Wiley Periodicals, Inc.     

Elemental composition of strawberry plants inoculated with the plant growth‐promoting bacterium Azospirillum brasilense REC3, assessed with scanning electron microscopy and energy dispersive X‐ray analysis

The elemental composition of strawberry plants (Fragaria ananassa cv. Macarena) inoculated with the plant growth‐promoting bacterium Azospirillum brasilense REC3, and non‐inoculated controls, was studied using scanning electron microscopy (SEM) and energy dispersive X‐ray (EDS) analysis. This allowed simultaneous semi‐quantification of different elements in a small, solid sample. Plants were inoculated and grown hydroponically in 50% or 100% Hoagland solution, corresponding to limited or optimum nutrient medium, respectively. Bacteria‐inoculated plants increased the growth index 45% and 80% compared to controls when grown in 100% and 50% Hoagland solution, respectively. Thus, inoculation with A. brasilense REC3 in a nutrient‐limited medium had the strongest effect in terms of increasing both shoot and root biomass and growth index, as already described for Azospirillum inoculated into nutrient‐poor soils. SEM‐EDS spectra and maps showed the elemental composition and relative distribution of nutrients in strawberry tissues. Leaves contained C, O, N, Na, P, K, Ca and Cu, while roots also had Si and Cl. The organic fraction (C, O and N) accounted for over 96.3% of the total chemical composition; of the mineral fraction, Na had higher accumulation in both leaves and roots. Azospirillum‐inoculated and control plants had similar elemental quantities; however, in bacteria‐inoculated roots, P was significantly increased (34.33%), which constitutes a major benefit for plant nutrition, while Cu content decreased (35.16%).     

3‐D visualisation,printing, and volume determination of the tracheal respiratory system in the adult desert locust,Schistocerca gregaria

Here, we describe a single micro‐CT scan with a spatial resolution of 10 μm of a 10‐day‐old adult male Schistocerca gregaria (Forskål) (Orthoptera: Acrididae) and we compare our tracheal volume (V_T) determination with published work on the subject. We also illustrate the feasibility of performing non‐invasive ‘virtual dissection’ on insects after performing micro‐CT. These post‐processing steps can be performed using free downloadable 3‐D software. Finally, the values of producing stereo‐lithography (STL) files that can be viewed or used to print out 3‐D models as teaching aids are discussed.