Short Communication Post-anoxic Viability and Developmental Rate of Artemia Franciscana Encysted Embryos

Free-living animals differ widely in their tolerance to the absence of molecular oxygen (anoxia): most die within hours or days under these conditions, and even well-adapted forms such as intertidal invertebrates do not survive more than a month or so of continuous anoxia (Hochachka, 1980; Hochachka and Guppy, 1987; Bryant, 1991). A striking exception is the brine shrimp, Artemia franciscana, whose encysted embryos (cysts) have previously been reported to survive 4 months of continuous anoxia with no decrease in viability (Dutrieu and ChrestiaBlanchine, 1966) and 7 months with only a 25 % reduction (Stocco etal. WIT). Both studies were terminated at the times given so that anoxic survival limits of this remarkable organism remain to be determined. In view of these observations, and the interest in cyst metabolism during anoxia (see Hand and Gnaiger, 1988; Clegg and Jackson, 1989a; Hand, 1990; Hofmann and Hand, 1990), further study of anoxic survival seemed worthwhile. Activated cysts in 'vacuum-packed' cans (San Francisco Bay Brand, Hayward, California) were processed and stored at -20°C (Clegg, 1986). These cysts are 'activated' in the sense that their hydration under permissive conditions (aerobic sea water at 25°C) results in the resumption of metabolism and development. Activated cysts contrast with the diapause cysts released from maternal females: diapause is a state of obligatory dormancy that is terminated by desiccation, among other treatments, resulting in activated cysts (Drinkwater and Clegg, 1991). Both diapause and activated cysts experience hypoxia/anoxia in nature (see Drinkwater and Clegg, 1991). Dried cysts (about 100 mg) were hydrated under anoxic conditions (Clegg and Jackson, 1989a) by the addition of 7 ml of anoxic sterile sea water (deoxygenated with 100% N2 for 4h) containing penicillin ( l J m g r a r 1 ) and streptomycin ( l ^ m g m T 1 ) in an 8ml screw-cap glass vial (N2 in the lml gas phase). The caps were wrapped with several layers of Parafilm to prevent loosening, and the vials were stored on their sides under ambient conditions of light and temperature (21-24°C). After anoxia, the cysts from each vial were collected by nitration and some were immediately assayed for emergence of embryos from their surrounding