Sequestration and in vivo effect of lead on DE2009 microalga, using high-resolution microscopic techniques

Algae are primary producers in a wide variety of natural ecosystems, and these microorganisms have been used in bioremediation studies. Nevertheless, very little is known about the in vivo effect of heavy metals on individual living cells. In this paper, we have applied a method based on confocal laser scanning microscopy and lambda scan function (CLSM-λscan) to determine the effect of lead (Pb), at different concentrations, on the DE2009 microalga. At the same time, we have optimized a method based on CLSM and image-analysis software (CLSM-IA) to determine in vivo biomass of this microorganism. The results obtained by lambda scan function indicated that the pigment peak decreases while the concentration of metal increases at pH 7. On the other hand at pH 4 there is no good correlation between the concentration of metal and the intensity of the emission of fluorescence of the pigment. Also, in some cases a displacement of the Chl a peak towards 680 nm is produced. Total and individual biomass determined by CLSM-IA shows statistically significant differences between unpolluted and 10 mM polluted cultures. Complementary studies using electron microscopy techniques coupled to energy dispersive X-ray microanalysis (EDX) demonstrate that the microalga can sequestrate Pb extra- and intracellularly.