Measurements of photosynthesis, respiration, electron transport activity and ATP were used to locate the site(s) of copper toxicity in the fouling diatom, Amphora coffeaeformis. Results of this and previous research suggest that, except at very high concentrations ( 5 × 10 -6 M), ionic copper does not target these processes, but rather one which occurs at or near the plasmalemma. This may reflect a reduced ability of ionic copper to cross the membranes of the chloroplasts and mitochondria, and possibly the plasmalemma as well. In contrast, copper bound to HQO (8-hydroxyquinoline) or HQA (8-hydroxyquinaldine) was highly toxic to electron transport and ATP production, although the ligands alone had no effect on either process. This supports a membrane-carrier role for HQO and HQA, which give copper access to internal mitochondrial and possibly chloroplast membranes.In a second set of experiments, A. coffeaeformis cells were treated with enzymes in order to elucidate external copper tolerance mechanisms. Cells treated with the polysaccharidase, p-glucuronidase, became more sensitive to copper, whereas those treated with the protease, trypsin, showed improved growth and no increase in sensitivity to copper. This supports a polysaccharide-based tolerance mechanism.
When cultured on different types of solid media, the marine‐fouling diatom Amphora coffeaeformis (Ag.) Kütz. consistently formed two distinct colonial morphotypes named tight and fuzzy. Tight colonies were comprised mainly of small, morphologically distorted, nonmotile cells, whereas morphologically normal and highly motile cells formed the fuzzy colonies. Cells from tight colonies were less adherent to glass, grew more slowly in liquid media, and had a slightly decreased viability on plates with copper than cells from fuzzy colonies. Whereas the protein profiles of the two types of cells were nearly identical in polyacrylamide gels stained with Coomassie blue, cells from tight colonies produced a significantly lower amount of a protease‐resistant, low Mr polysaccharide or glycoconjugate as detected in silver‐stained gels. The frequency of appearance of the fuzzy and tight morphotypes was not influenced by the mode of nutrition or the type of substratum to which the algal cells adhered. However, certain formulations of solid medium and the presence of growth‐inhibitory concentrations of copper in agar plates favored the formation of tight colonies. Due to their frequencies and patterns of appearance, it was clear that the two naturally formed morphotypes were not the consequence of spontaneous mutations, genetic rearrangement, or selection of stable natural variants, and we have hypothesized that they were linked to a normal physiological behavior. The tight colonial morphotype was used as a valuable marker to screen for true motility/adhesion mutants within an ultraviolet‐mutagenized population of A. coffeaeformis. Seven mutants were isolated that were non‐motile on agar plates, poorly adherent to glass, and distinguished from naturally formed cells from tight colonies by their inability to form fuzzy colonies upon subculture on solid media.
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