Diane Lazinsky

EFFECT OF LIGHT CYCLE ON DIATOM FATTY ACID COMPOSITION AND QUANTITATIVE MORPHOLOGY1

Quantitative cytological and fatty acid composition was determined for the diatom Cyclotella meneghiniana Kütz, Data from four separate experiments were examined to elucidate changes that may occur with respect to daily photoperiod. Overall, fatty acid composition is similar to that reported for other diatoms with the exception that the C16 fatty acids constitute approximately 70% of all fatty acids. The major fatty acids are C14:0, 16:1, 16:0, 18:0, and 20:5. Fatty acids that are present in minor amounts are iso‐14:0, iso‐15:0, 15:0, 17:0, 18:4, 18:2, 18:1, 19:0, 20:0, 22:0, and 23:0.

Effects of chlorinated benzenes on diatom fatty acid composition and quantitative morphology. II: 1,3,5-trichlorobenzene

Cells of the diatomCyclotella meneghiniana were exposed in a closed system to 0.245 ppm 1,2,4-trichlorobenzene. Response to this chlorinated benzene was monitored over a period of 5 days by quantitative ultrastructure and fatty acid percent composition. Over the time period examined, 41 significant morphological changes and 12 significant changes in fatty acid composition were observed. Autophagic-like vacuoles were frequently observed, indicating lysis of cellular constituents. In general, there was an increase in the C20:5 fatty acid, which may be indicative of photosynthetic impairment, since this fatty acid probably substitutes for linolenic acid in diatoms. The most significant numbers of changes were observed after 8 hr of exposure to this lipophilic toxicant, and these changes occurred in membranous organelles. It is suggested that the daily photoperiodic variation in lipid content of phytoplankton may be an important consideration in evaluating effects of lipohilic toxicants.

Quantitative ultrastructural changes associated with lead-coupled luxury phosphate uptake and polyphosphate utilization

Quantitative electron microscopy (stereology) was used to assess the ultrastructural response of three algae representative of the classes Chlorophyceae, Cyanophyceae, and Bacillariophyceae to lead-coupled polyphosphate degradation. The organisms were exposed to a culture medium concentration of 20 ppb Pb for 3 hr at the time of luxury phosphate uptake and subsequently transferred to phosphorus and lead-free medium. A differential sensitivity was observed as follows:Plectonema > Scenedesmus > Cyclotella. InPlectonema andScenedesmus, detrimental cytological changes were observed when the polyphosphate relative volume dropped below 0.5%, which was approximately the P-starvation level of polyphosphate. Few significant ultrastructural changes were observed inCyclotella after one week in P-deficient medium. At this time, the relative volume of polyphosphate was still 1.5%. Although a few significant ultrastructural changes occurred with phosphate deprivation, the greatest numbers of changes occurred in cells that had been exposed to a short-term (3 hr) low level of Pb. Changes in the relative volume of polyphosphate in all three organisms suggest thatPlectonema andScenedesmus have higher phosphate nutrient requirements thanCyclotella. The ecological implications of metal sequestering by polyphosphate are discussed.

Effects of chlorinated benzenes on diatom fatty acid composition and quantitative morphology. IV: Pentachlorobenzene and comparison with trichlorobenzene isomers

Cells of the diatomCyclotella meneghiniana were exposed in a closed system to 0.095 ppm pentachlorobenzene over a period of 5 days. Changes in fatty acid and morphological percent composition were monitored to determine the effect of the toxicant. The greatest morphological change observed was an increase in lipid volume. Most morphological changes occurred in the 1 hour and 5 day sampling periods. Few changes in morphological characteristics or fatty acid percent composition were observed at eight hours, when the cells were in the dark. The C18∶1 and C20∶5 fatty acids were most variable with exposure to pentachlorobenzene. Results suggest that at sublethal doses, lipophilic toxicants exert effects that are biphasic. That is, immediately measurable effects are observed in the cells that include increases in storage products and changes in membranous organelles. Long-term effects are postulated to be the result of mobilization of lipophilic toxicants that have partitioned into lipid stores and are more available when lipids are metabolized. Although pentachlorobenzene has a higher octanol/water partition coefficient, it appears to exert fewer cellular changes than any trichlorobenzene isomer.

Synergistic effects of nutrients and lead on the quantitative ultrastructure ofCyclotella (Bacillariophyceae)

A four-way completely crossed factorial enrichment experiment (2T×2Si×2Pb×2P) was used to analyze the short (2 hr) and long (48 hr) term effects of nutrients and lead on the quantitative ultrastructure ofCyclotella, a diatom. Some of the cytological modifications that have been reported to occur in other algae as a result of heavy metal exposure occurred inCyclotella, but were mediated in some cases by the addition of low levels of nutrients. While it is extremely difficult to simulate the natural environment where a variety of parameters may be changing simultaneously, it is suggested that factorial design experiments approximate it more closely. Furthermore, environmental factors that may change the availability of metals may also change the physiological state of the cells, which may also either amplify or mitigate the metal effects.

Morphometric analysis of phosphate and chromium interactions in Cyclotella meneghiniana

Quantitative electron microscopy was used to evaluate interactive effects of chromium additions and phosphate nutrient status on the diatom Cyclotella meneghiniana. Cells of differing phosphate status were exposed to hexavalent chromium for 7 days and morphological changes in both phosphate alone and phosphate and chromium treatments were observed. Changes in chloroplast relative volume and number/ volume were observed in all treatments. Changes in mitochondrial and vacuolar volume and an increase in presumed autophagic activity were apparent in all chromium treatments. An increase in polyphosphate was also found in most chromium treated cells. These morphological modifications are discussed in relation to changes observed with exposure to other heavy metals.

PARAFORMALDEHYDE -GLUTARALDEHYDE AS A ROUTINE PHYTOPLANKTON FIXATIVE

Several fixatives such as Lugols iodine solution and osmium tetroxide traditionally have been employed in phytoflagellate studies. Lugols iodine (LI) stains cytoplasmic components, making the algae easier to study with a light microscope while at the same time it preserves their flagella. However, this fixative is not suitable for electron microscopy. Osmium tetroxide (Os) is also an excellent fixative for flagella (Rosowski, 1977). But due to its extreme toxicity, it is not useful for shipboard sampling and sample transport. Since we have successfully employed paraformaldehyde-glutaraldehyde (PFG) modified after Karnovsky (1965) in our laboratory for ultrastructural and cytochemical studies of cultured diatoms, we attempted to use this fixative for routine light and electron microscopic study of phytoflagellates to avoid toxicity problems and provide samples that would be useful for light microscopy, SEM, and TEM. Unialgal cultures of Pandorina and Euylena were employed in the present study and fixed by one of the following methods:

Ultrastructural modification of three blue-green algae following heavy metal exposure

Algal responses to heavy metal exposure may vary with the species and similarly, various metals may cause a differing response within a species. In blue-green algae metals can be accumulated, and reduction in growth rates and inhibition of phosphate uptake are co~on effects that have been attributed to heavy metal treatment. More specifically, heavy metals can induce drastic effects at the ultrastructural level such as changes in the volume and number of cell-sustaining inclusions. The present study was conducted to determine the effects of various heavy metals on the ultrastructure of three blue-green algae and thus to evaluate their susceptibility to a particular heavy metal.

Accumulation and cellular effects of heavy metals in benthic and planktonic algae

Concentration levels of heavy metals which are found in the Laurentian Great Lakes (ppb range) can be accumulated by algae and can exert toxicological effects on algal species in natural assemblages. One of the most common mechanism for sequestering metals in algae is the incorporation of these metals into polyphosphate bodies. Presumably, the movement of species that have sequestered metals in this manner by water mass movements and circulation patterns could lead to the dispersal of metals to areas that are removed from the point source of pol lut ion. Evidence was also obtained that indicated that tub i f i c id worms in sediments from Saginaw Bay had lead accumulations in the cells l in ing the gut. The gut contents, as examined by x-ray energy dispersive analysis, included blue-green algae that had polyphosphate bodies and lead associated with them, leading us to believe that there is a transfer of lead from the algae in the gut into the tub i f i c id body.

Synergistic effects of phosphorus nutrient status and lead exposure in three algae

There is some indication in the literature that nutrient status may mitigate toxicity effects of heavy metals in algae. Monahan (1973) demonstrated that cells of the green alga Ho~otila which were phosphate sufficient were less susceptible to lead toxicity than cells which had no phosphate. Similarly, our laboratory (Sicko-Goad and Stoermer, 1979) reported that lead may be incorporated into polyphosphate bodies thereby reducing the amount of lead available to intracellular sites. We designed a series of experiments to determine (i) the effects of lead exposure on certain algae and (2) the effects of phosphorus nutrient status on lead toxicity.