Eclipse 80i lm

Morphology, cell sizes (n = 36), and life cycle were observed by light microscopy (LM) with an Nikon Eclipse 80i LM, Plan Apo VC 100 × 1.40 objective and a DS-5 M camera (Nikon Instruments, Netherlands), either with differential interference contrast or at chlorophyll autofluorescence mode (filter emission = 600 LP, excitation = 480/40). For TEM, 3-week-old cultures were fixed with a standard chemical fixation protocol modified for green algae (2.5% glutaraldehyde, 1% OsO₄ in 20 mM caccodylate buffer, pH = 7.0) according to Holzinger et al. (2009) (link). Specimens were then dehydrated in increasing ethanol concentrations, transferred to modified Spurr’s resin, and polymerized for 8 h at 80 °C. For observation at the TEM, ultrathin sections were prepared with an ultramicrotome (Reichert Ultracut, Leica Microsystems, Austria), counterstained with uranyl acetate and Reynold’s lead citrate, and examined using a Zeiss LIBRA 120 transmission electron microscope (Carl Zeiss AG, Germany) at 80 kV. Images were captured with a TRS 2 k SSCCD camera (Albert Tröndle Restlichtverstarker Systeme, Moorenweis, Germany) and further processed using Adobe Photoshop software (Adobe Systems Inc., USA).