Marine mollusc shell growth has been widely measured using fluorochrome marking. In order to test the efficiency and reliability of calcein staining on Pinctada margaritifera shells and pearls, the present study examined two administration methods, different concentrations and several immersion times. Immersion in a 150 mg L−1 calcein solution for 12 h to 24 h appeared to be the best method for marking P. margaritifera shells. For pearl marking, injection of a 200 mg L−1 calcein solution into the pearl pouch was the optimal method. Calcein marking was then used to measure the influence of food resource levels on the shell growth. Groups of 23-month-old P. margaritifera were fed at three trophic levels for two months. The two highest food levels tested (6000 cell mL−1 and 15 000 cell mL−1) induced uniform growth between the dorsal and ventral sides of shell, whereas the lowest food level (800 cell mL−1) induced greater growth on the dorsal side. Shell deposits from the ventral side were observed using a scanning electron microscope, revealing that the difference of the trophic level over two months had modified the thickness of the aragonite tablets formed. These results showed that the trophic level is a major factor conditioning P. margaritifera development.

-Cryopreservation is a valuable tool for genetic improvement programs. Several bivalve mollusc species have already been the subject of such programs and the Tahitian black pearl oyster industry is now planning the development of selective breeding for desirable traits in Pinctada margaritifera. The ability to cryopreserve spermatozoa would, therefore, offer significant benefits to the cultured black pearl industry. Spermatozoa were cryopreserved with cryoprotectant agent (CPA) 0.7 M trehalose in 0.8 M Me 2 SO and a two-step freezing process was used: straws were first maintained in nitrogen vapour for 10 minutes, then directly plunged into liquid nitrogen and stored for one week before use. The viability of thawed sperm was 23% lower than that of fresh sperm. When using thawed sperm, therefore, a higher sperm/egg ratio of 100 000:1 was required to reach 80% oocyte fertilization, compared with 100:1 for fresh sperm. Nevertheless, this first demonstration of cryopreserved sperm fertility in black pearl oyster confirms the hatchery applicability of the cryopreservation technique defined here. Monitoring for larval viability during the first 23 days of life revealed no significant differences between the progeny produced with cryopreserved sperm and that produced using fresh sperm.