Many of the small, cysteine-rich ion-channel modulatory peptides found in Cnidaria are distantly related to vertebrate defensins (of the trans-defensin superfamily). Transcriptomic and proteomic studies of the endemic Australian speckled sea anemone (Oulactis sp.) yielded homologous peptides to known defensin sequences. We extended these data using existing and custom-built hidden Markov models to extract defensin-like families from the transcriptomes of seven endemic Australian cnidarian species. Newly sequenced transcriptomes include three species of Actiniaria (true sea anemones); the speckled anemone (Oulactis sp.), Oulactis muscosa, Dofleinia cf. armata and a species of Corallimorpharia, Rhodactis sp. We analyzed these novel defensin-like sequences along with published homologues to study the evolution of their physico-chemical properties in vertebrate and invertebrate fauna. The cnidarian trans-defensins form a distinct cluster within the chemical space of the superfamily, with a unique set of motifs and biophysical properties. This cluster contains identifiable subgroups, whose distribution in chemical space also correlates with the divergent evolution of their structures. These sequences, currently restricted to cnidarians, form an evolutionarily distinct clade within the trans-defensin superfamily.