Selection of taxonomic criteria in the marine Pyrenomycetes, especially for the characterization of genera, is an unsettled problem. Kohlmeyer separates species primarily by ascospore ornamentation patterns (1961), and genera by certain perithecial characteristics (1960). Johnson contends, on the other hand, that species differing in the texture and origin of ascospore appendages (syn.: ornamentations, processes) may belong in different genera (Johnson, 1963 a—d; Johnson and Cavaliere, 1963). Cavaliere (Cavaliere, 1966 a—c; Cavaliianic and Johnson, 1966 a—b), finding little support for Kohlmeyer’s (1960) position, suggests more weight be given ornamentation patterns, should these prove constant.
Clarifying the taxonomic and biological significance of marine pyrenomycete-ascospore appendages is the primary purpose of this investigation. Athorough morphological and microscopic cytochemica] study is made to determine the origin, variability, and composition of the basic textural types of appendages, that is, the membranous, flexuous, rigid, exposed mucilaginous, and enclosed mucilaginous types. These are presumed to arise through epispore rupture, wall growth, and secretion occurring within the spores or within the ascus cytoplasm (Johnson, 1963a—d; Johnson and Cavaliere, 1963; Wilson, 1965). Attention is given mainly to ascospores of Ceriosporopsis halima Linder, Corollospora maritima Werdermann, Halosphaeria medioSetigera Cribb and Cribb, Lulworthia grandispora Meyers, and Lulworthia medusa (Ellis and Everhart) Cribb and Cribb cultured under various conditions of salinity and temperature. These species exhibit the entire range of diversity in appendage texture and origin, and they fruit readily in culture. Lulworthia fucicola Sutherland, Halosphaeria tubulifera Kohlmeyer, and Remispora ornata Johnson and Cavaliere are considered also.
A second objective is to describe in detail the organization and composition of the walls, protoplasts, and inclusions of the ornamented spores throughout their development and during early stages of germination. Considerable attention is given to these structures in their relation to the origin and nature of appendages. However, this is carried a step further, and many spore features are examined carefully by means of supravital and morphological stains, digestive enzymes, and controlled microscopic histochemical tests. Electrophoresis is used also, as well as several histochemical techniques, to determine whether the proteins associated with polyphosphates in volutin are basic. Unlike spores of the more familiar terrestrial fungi, those of the marine Pyrenomycetes studied here are large and hyaline, and permit the resolution of considerable cytological detail.
Another purpose is testing the applicability of modern histochemical methods, including fluorescence microscopy, to fungus spore cytology. Microscopic cytochemical studies of fungi are relatively few, and with certain exceptions (e.g., Breslau, 1955; Fuller, 1960; Roth and Winkelmann, 1960; Pomeranz, 1962), seldom mention controls. More tests and controls in my study are used than are needed to characterize chemically various structures, so that their sensitivity and specificity on fungi as test organisms might be evaluated. Methods for detecting enzymes and minerals other than phosphates are excluded, as are quantitative procedures.
Devising media that support normal ascosporic development through several transfers is the fourth objective. Such media were needed for my study, and also in many other areas of investigation (Johnson and Sparrow, 1961). Nutritional factors affecting mycelial growth, generally emphasized in cultural studies of lignicolous marine fungi (Johnson and Sparrow, 1961; Sguros and Simms, 1963), are of no concern here.