We applied quantitative histochemical techniques and digital image analysis to study seasonal cycles of use of lipid and protein reserves during vitellogenesis in the pearl oyster Pinctada mazatlanica. Female gonad samples were collected seasonally during an annual cycle and processed histologically to characterize the gametogenic cycle, analyze variations in the frequency and size of vitellogenic and postvitellogenic oocytes and calculate the ooplasm:nucleoplasm ratio for both types of oocytes. Lipid and protein inclusions in each type of oocyte were identified using Sudan Black B and Schiff's ninhydrin stains. In both cases, quantification of lipid and protein components was performed through measuring variations in the color coverage area of the oocyte with a digital image analysis system. With this procedure, we calculated a lipid index and a protein index to refer oocyte quality. The lipid index was higher in winter, suggesting a strategy towards storage in the gonad. The protein index was highest during spring in vitellogenic oocytes and during winter in postvitellogenic oocytes, indicating that proteins are actively used during oocyte growth. These results, together with data of the ooplasm:nucleoplasm ratio, suggest differential accumulation of lipid and protein components within the ooplasm during oocyte development. Quantitative histochemistry and digital image analysis represent a combination of reliable techniques for evaluating reproductive processes and oocyte growth and quality.
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Vol. 24 • No. 4