Addition of germanic acid into the culture medium of the diatom Synedra acus subsp. radians (Kutz.) Skabitsch. had nearly no influence on the culture growth at the Ge/Si molar ratio 0.01, but stopped it at ratios 0.05 and higher. It was shown by mass-spectrometry that at the Ge/Si ratio 0.01 germanium was incorporated in both the cytoplasm and siliceous valves, whereas at Ge/Si 0.05 it was incorporated into the cytoplasm but almost failed to accumulate in the valves. At Ge/Si 0.1 germanium was accumulated in the cytoplasm, but its incorporation into the valves terminated. Studies on the cell morphology by light, epifluorescence, and transmission electron microscopy showed that high concentrations of germanic acid induced disorders in morphogenesis of the siliceous frustule and accumulation of large rhodamine-stainable electron-dense inclusions. Model chemical experiments with over-saturated solutions of silicic acid in the presence of polyallylamine revealed that addition of 5% germanic acid considerably accelerated coagulation of silica. Hence, the toxic effect of germanic acid on diatoms could be caused by changes in coagulation of silica.
The planktonic diatoms, Asterionella formosu Kutz. and Synedru acus Hass., were cultivated in 96-well flat-bottom plastic plates with 100 pL of medium per well. At regular time intervals, cells were counted using an inverted microscope. Typically, the number of cells per well increased over 10 days from 50-200 to >1000. The first growth inhibition experiments were successfully run at room temperature using natural illumination. However, for better control of the growth conditions, a special micro-incubator was manufactured. This device hosted two incubation plates and was equipped with Peltier elements for cooling below room temperature, and a small luminescent lamp. For automatic counting, two digital micro-photographs of each well were taken, first at a 0.2 msec exposure and the second at a 5 msec exposure. The images were treated by means of custom software based on ImagePro to identify and count the diatom cells. Cultivation of diatoms on the micro-scale proved to be a convenient technique. Using this technique, we were able to study the effects of mercaptoethanol, Cu2+ on A. formosa and Cu2+, Hg2+, Cd2* and phenanthroline upon the growth of S. ucus.
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