Corals, like other cnidarians, are venomous animals that rely on stinging cells (nematocytes) and their toxins to catch prey and defend themselves against predators. However, little is known about the chemical arsenal employed by stony corals, despite their ecological importance. Here, we show large differences in the density of nematocysts and whole-body hemolytic activity between different species of reef-building corals. In the branched coral Stylophora pistillata, the tips of the branches exhibited a greater hemolytic activity than the bases. Hemolytic activity and nematocyst density were significantly lower in Stylophora that were maintained for close to a year in captivity compared to corals collected from the wild. A cysteine-containing actinoporin was identified in Stylophora following partial purification and tandem mass spectrometry. This toxin, named Δ-Pocilopotoxin-Spi1 (Δ-PCTX-Spi1) is the first hemolytic toxin to be partially isolated and characterized in true reef-building corals. Loss of hemolytic activity during chromatography suggests that this actinoporin is only one of potentially several hemolytic molecules. These results suggest that the capacity to employ offensive and defensive chemicals by corals is a dynamic trait within and between coral species, and provide a first step towards identifying the molecular components of the coral chemical armament. Sci Rep Sci Rep 1579 scirep Scientific Reports 2045-2322 Nature Publishing Group PMC5762905 PMC5762905.1 5762905 5762905 29321526 10.1038/s41598-017-18355-1 18355 1 Article The chemical armament of reef-building corals: inter- and intra-specific variation and the identification of an unusual actinoporin in Stylophora pistilata Ben-Ari Hanit 1 2 Paz Moran 1 Sher Daniel dsher@univ.haifa.ac.il 1 1 0000 0004 1937 0562 grid.18098.38 Department of Marine Biology, Leon H. As their name attests, the physical structure of most coral reefs is composed of hermatypic (reef-building) stony corals. As a result of anthropogenic stress and global climate change, hermatypic corals are increasingly facing challenges such as loss of zooxanthellae due to bleaching 1 , 2 , increased microbial loads 3 – 8 , competition with macroalgae 9 , and the potential for nutrient limitation and loss of exoskeleton due to ocean acidification 10 . Under each of these scenarios, reef-building corals may need to rely even more on offensive or defensive chemicals to survive in a changing world 2 , 11 . After identifying consistent differences between hermatypic corals, we focused further studies on Stylophora pistillata , one of the most abundant corals in the Indo-Pacific and a commonly used model for studying various aspects of coral physiology, genetics and ecology (e.g. 47 – 53 ). In particular, we assessed to what extent the number of nematocysts and hemolytic activity differ within colonies of this branching coral. Finally, using bioassay-guided column chromatography and mass spectrometry, we identify a new group of coral-derived hemolysins belonging to the actinoporin family. Results Coral species differ widely in nematocyst density and tissue hemolytic activity To assess the extent to which reef-building corals produce and deliver toxins, we collected fragments from seven abundant reef-building organisms (six anthozoan corals and the hydrozoan “fire coral” Millepora sp ) from the Gulf of Aquaba (N 29° 30.211′ E 34° 55.068), at the northern tip of the Red Sea. For clarity, we refer to all these organisms, including Millepora sp, as “corals”, using the genus name throughput the manuscript. As shown in Fig. Nevertheless, the most abundant protein in each of these pools (comprising 48-90% of the total ion current) was identified as an actinoporin with high identity (60%) to the hemolytic toxin Sticholysin-2 from the sea anemone Stichodactyla helianthus 58 (see below). There was no other protein shared among all three pools. These results strongly suggest that the actinoporin is the hemolysin responsible for the majority of the hemolytic activity in these pools. We thus propose the rational nomenclature of Δ-Pocilopotoxin-Spi1 (Δ-PCTX-Spi1) for this hemolysin, the first hemolytic toxin to be partially isolated and characterized in true reef-building corals 59 . are known to cause painful stings when touched, leading to their common name, “Fire Corals”. Previous studies have shown that fire corals produce phospholipase A2 toxins (PLA2s) 37 , 38 , and one might expect that these membrane-disrupting enzymes would cause hemolysis. However, while several publications describe the purification or partial purification of PLA2s from cnidarians, none of these proteins have been shown to be hemolytic 87 , 88 . Our own experiments with commercially-available bee venom PLA2 revealed high phospholipase activity but no hemolysis (not shown). We therefore propose that the Millepora PLA2s are not hemolytic, and that, similar to jellyfish and some sea anemones 16 , Millepora have lost the ability We propose that these can be considered as quantitative functional traits related to prey capture or defense from predators. Our results reveal large and consistent differences in these two traits, both between stony coral species and within colonies of Stylophora . Furthermore, these traits are dynamic over time, as seen in the reduction in both traits in Stylophora colonies maintained in water tables compared to colonies from the wild, and we propose that these changes are related to predation pressure. Material and Methods Coral collection and maintenance All coral colonies and coral fragments were collected from an in situ coral nursery near the Interuniversity Institute for Marine Sciences (IUI N 29° 30.211′ E 34° 55.068) at a 5–12 m depth located at the Gulf of Eilat, a northeastern extension of the Red Sea (Fig. 1 ). The use of corals for this research was approved by the Israel Nature and Parks Authority (authorizations 2012/38770, 2014/40464 and 2015/40869).