Potential of CLSM in studying some modern and fossil palynological objects.

JOURNAL OF MICROSCOPY(2018)

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摘要
We have tested possibilities and limitations of confocal laser scanning microscopy to study the morphology of pollen and spores and inner structure of sporoderms. As test objects, we used pollen grains of the modern angiosperm Ribes niveum (Grossulariaceae) and Datura metel (Solanaceae), fossil angiosperm pollen grains of Pseudointegricorpus clarireticulatum and Wodehouseia spinata dated to the Late Cretaceous, fossil gymnosperm pollen grains of Cycadopites-type dated to the Middle Jurassic, and fossil megaspores Maexisporites rugulaeferus, M. grosstriletus, and Trileites sp. dated to the Early Triassic. For comparative purpose, we studied the same objects with application of conventional light, scanning electron (to entire pollen grains and spores or to semithin sections of their walls), or transmission electron microscopy. The resolution of confocal microscope is much lower than that of electron microscopes, as are its abilities to reconstruct the surface patterns and inner structure. On the other hand, it can provide information that is unreachable by other microscopical methods. Thus, the structure of endoapertures in angiosperm pollen grains can be directly observed. It is also helpful in studies of asymmetrical pollen and pollen grains bearing various appendages and having complicated exine structure, because rotation of 3-D reconstructions allows one to examine all sides and structures of the pollen grain. The exact location of all visible and concealed structures in the sporoderm can be detected; this information helps to describe the morphology and inner structure of pollen grains and to choose necessary directions of further ultrathin sectioning for a transmission electron microscopical study. In studies of fossil pollen grains that are preserved in clumps and stuck to cuticles, confocal microscope is useful in determining the number of apertures in individual pollen grains. This can be done by means of virtual sections through 3-D reconstructions of pollen grains. Fossil megaspores are too large and too thick-walled objects for a confocal study; however, confocal microscope was able to reveal a degree of compression of fossil megaspores, the presence of a cavity between the outer and inner sporoderm layers, and to get some information about sporoderm inner structure. Lay description Pollen grains and spores of terrestrial plants are microscopical objects (), studied within the scope of palynology - a discipline akin to both biology and geology. Pollen grains and spores are very diverse and those of different plants differ from each other, allowing palynologists to be aware which plant produced a given pollen grain or spore. On this basis, palynological data are used for the systematics and phylogeny of different groups of extinct and living plants, dating and stratigraphy of geological deposits (), studies of honeys, allergies, as well as for some other purposes. Light microscope, a workhorse of palynology, is conventionally used to observe the general morphology of pollen grains and spores, whereas scanning and transmission electron microscopes provide detailed information about their surface pattern and inner structure. Light microscope gives a lower resolution; electron-microscopical methods are laborious, expensive and destructive. We have tested pros and cons of confocal microscope for studies of fossil and modern pollen and spores. The resolution of confocal laser scanning microscope is higher than that of traditional light microscope and much lower than that of electron microscopes, as are its abilities to reconstruct the surface patterns and inner structure. On the other hand, it can provide information that is unreachable by other microscopical methods. It also allows one to study palynological objects without destroying them, that is particularly pertinent for fossil objects. For this study, we used as test objects pollen grains and spores of different morphologies, botanical affinities, and preservation. We have studied pollen grains of a living angiosperm plants and those of extinct angiosperms, which are more than 66 million years (Ma) in age, pollen grains of extinct gymnosperms (more than 170 Ma), and spores of fossil lycopsids (about 252 Ma). The structure of endoapertures in modern angiosperm pollen grains was directly observed as well as appendages and complicated wall structure in fossil angiosperm pollen grains. The ability of confocal microscope to rotate 3-D reconstructions of studied objects was very helpful. Virtual sections under arbitrary angles through 3-D reconstructions of pollen grains of gymnosperms helped us to understand their morphology in spite of the unusual preservation of these fossils. Fossil megaspores turned to be too large and too thick-walled objects for a confocal study, but some useful information also was amassed.
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关键词
Confocal laser scanning microscopy (CLSM),light microscopy (LM),megaspores,pollen grains,scanning electron microscopy (SEM),transmission electron microscopy (TEM)
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