The ultrastructure of ascosporogenesis in freeze-substituted Thelebolus crustaceus: enveloping membrane system and ascospore initial development

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Abstract
High-pressure and propane-jet freezing were essential for determining the origin and development of the enveloping membrane system during ascosporogenesis in Thelebolus crustaceus. Prior to the completion of mitotic divisions within the ascus, invaginations of the plasma membrane initiated formation of the enveloping membrane system. Fusion of individual units of the closely spaced double membranes resulted in the formation of a cylinder around most of the ascus protoplasm. This double-membraned cylinder enveloped individual nuclei with accompanying cytoplasm and organelles to form ascospore initials. Envelopment of asci nuclei to form ascospore initials appeared to be facilitated by a nuclear-associated organelle and closely associated microtubule organizing center. Initially, wall materials and (or) precursors were deposited between the closely spaced double membranes from within the ascospore initials. Secondary wall formation appeared to be deposited, in part, from the epiplasm. Microtubules located adjacent to the inside wall layer of the ascospore initials appeared to contribute to an elliptical shape. Subsequent to epispore wall formation, numerous microtubules were found associated with the outer membranes of the ascospores and appeared to interconnect the spores into a single mass before discharge. Key words: Thelebolus crustaceus, ascosporogenesis, high-pressure freezing, propane-jet freezing, ultrastructure, laser scanning confocal microscopy.

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