Location of toxicity within the Mediterranean sponge Crambe crambe (Demospongiae: Poecilosclerida)

Within-specimen location of toxicity in Crambe crambe (Schmidt) has been addressed by complementary procedures on specimens collected in north-east Spain (Western Mediterranean) in winter of 1993. The toxicity of the distal (ectosome) and basal (choanosome) sponge parts have been analysed and the main cellular types present in these two layers have been studied by light and electron microscopy. The toxicity of the three main cell types, separated by the gradient-density method, has also been analysed. Three main fractions, each of them enriched in a different cellular type, were obtained: Fraction 1 (interface between 2 and 5% Ficoll) contained 90+/-0.9% (mean +/- SE) of spherulous cells and 10% of different cell types consisting of choanocytes (5 +/- 0.54%), and unidentified cells or cell debris (5 +/- 0.84%); Fraction 2 (interface between 5 and 8% Ficoll) was enriched in choanocytes (70 +/- 0.95%), and also contained spherulous cells (11.8 +/- 0.73%), archeocytes (6.2 +/- 0.74%) and unidentified sponge cells (12 +/- 0.74%); Fraction 3 (interface between 8 and 11% Ficoll) mainly consisted of archeocytes and archeocyte-like cells (75 +/- 0.66%), together with spherulous cells (7 +/- 0.74%) and other unidentified sponge cells and cell aggregates mainly formed by choanocytes (18 +/- 0.41%). Toxicity [measured in toxicity units, TU, using the Microtox(R) procedure] was significantly higher in the sponge ectosome (12.45 +/- 1.4 TU) than in the choanosome (2.58 +/- 0.92 UT). Only the abundance of spherulous cells in the sponge tissues correlated well with the pattern of toxicity observed, and this was corroborated by the toxic behaviour of the three cellular fractions obtained: the one enriched in spherulous cells was highly toxic (9.08 UT), whereas those enriched in choanocytes and in archeocytes were almost inactive (0.48 UT) or totally innocuous, respectively. All these results point to the spherulous cells being responsible for the storage (and possibly production) of the toxic compounds in C. crambe. Toxicity is concentrated in the sponge periphery. Spherulous cells are also concentrated in this area and can also be observed outside the sponge exopinacoderm. These results correlate well with the assumption of a defensive role of toxicity, since encounters with potential epibionts, predators and competitive neighbours take through this peripheral zone. However, we two types of spherulous cells (orange and colourless, respectively) coexisting in the same sponge zones as well as in Cell Fraction I. Thus, we cannot at present determine whether one or both types are responsible for the toxicity encountered, although it is likely that the two correspond to different states of the same cell type.