*3.4. Antibacterial Activity and Chemical Ecology of the Egg Masses*

As part of a screening study on the antimicrobial properties of molluskan egg masses, *Dicathais orbita* was identified as a species of particular interest, with the lipophylic extracts showing strong activity against a range of human and marine bacterial pathogens [66,84]. Bio-guided fractionation identified the brominated indole precursors of Tyrian purple as being responsible for this activity [12]. Based on this activity, Benkendorff *et al*. [12] proposed that defense of the developing embryos against ubiquitous marine pathogens could be the naturally selected role for these brominated indoles in Muricidae evolution. Consistent with this, the surface of the egg capsules of *D. orbita* were found to have very low levels of bacterial biofilm formation, with a high proportion of dead bacteria indicated by live/dead bacterial staining [85]. Using the MTS cell proliferation and broth dilution assay, extracts containing the Tyrian purple precursors from the surface of *D. orbita* egg capsules were effective at inhibiting the growth of the marine biofilm forming bacteria *Pseudoalteromonas* sp. S91, as well as the molluskan pathogen *Vibrio harveyi* [28]. The egg capsules of *D. orbita* were also found to have no protists on the surface and were relatively free of algal fouling compared to other gastropod egg masses [86]. The low surface fouling on these egg masses is likely to be due to a combination

of chemical, physical and mechanical defense mechanisms preventing bacterial attachment and persistence on the surface [85].

To investigate whether fatty acids could contribute to the observed antibacterial activity in liphophylic extracts from mollusks [66,76], Benkendorff *et al*. [70] tested a series of lipid mixtures modeled on those found in the egg masses. The lipid mixture modeled on the fatty acid and sterol composition of *D. orbita* and similar Neogastropoda had very limited antibacterial activity against marine pathogens, especially when compared to species with gelatinous egg masses and a high content of polyunsaturated fatty acids [70]. It is possible that the bioactive indoles in *D. orbita* egg masses [12,60] negate the requirement for antimicrobial polyunsaturated fatty acids, or perhaps the transfer of bioactive indoles for defense of the egg masses was selected for due to the absence of alternative secondary metabolites with antibacterial activity in these egg masses.
