Increased Zooxanthellae Nitric Oxide Synthase Activity Is Associated With Coral Bleaching HENRY TRAPIDO-ROSENTHAL 1, *, SANDRA ZIELKE 1 , RICHARD OWEN 1,2 , LUCY BUXTON 1,3 , BRIAN BOEING 1,4 , RANJEET BHAGOOLI 1,5 , AND JESSICA ARCHER 1,6 1 Bermuda Biological Station for Research, Inc., Ferry Reach GE-01, Bermuda; 2 Environment Agency, Westbury-on-Trym, Bristol BS106BF, UK; 3 Department of Environmental Sciences, University of Technology, Sydney, New South Wales 2065, Australia; 4 Department of Oceanography, University of Hawaii, Honolulu, Hawaii 96822; 5 Department of Chemistry, Biology and Marine Sciences, University of the Ryukyus, Nishihara, Okinawa, 903-0213 Japan; and 6 Department of Biology, Evergreen State University, Olympia, Washington 98505 Coral bleaching, the breakdown of the symbiotic rela- tionship between host corals and their photosynthetic dinoflagellate endosymbionts, is a phenomenon of major ecological significance. The cellular and molecular mech- anisms underlying bleaching are poorly understood. Here we report substantial increases in nitric oxide synthase (NOS) activity in symbionts associated with bleaching cor- als. Nitric oxide (NO), a product of NOS activity, is a highly reactive and membrane-permeable molecule that has the potential to exert cytotoxic effects in host cells either di- rectly or in combination with molecular oxygen or super- oxide radicals. This potential allows us to suggest that coral bleaching may be a stress response initiated by the symbi- ont, rather than by the host. Coral reefs are biodiverse ecosystems of enormous eco- logical and economic importance (1, 2). The corals that constitute the foundations of these ecosystems are symbiotic assemblages consisting of the host coral and symbiotic dinoflagellate algae of the genus Symbiodinium, known as zooxanthellae, which provide the host organism with pho- tosynthetically derived nutrients and promote calcification. Environmental stressors can induce the phenomenon of bleaching, which refers to the loss of symbiotic algae or, less commonly, the degradation of algal pigments (3). Se- verely bleached corals can suffer depressed growth rates, increased susceptibility to disease and, at the extreme, mor- tality (4, 5). The cellular and molecular mechanisms that underlie the breakdown of the symbiotic partnership are still poorly understood. The study of these mechanisms is an active research area, in which a consensus is developing that a primary step in the bleaching process occurs in the zoo- xanthellae (3, 6). Many investigations into the molecular mechanisms of bleaching have used experimentally bleached symbioses— that is, assemblages exposed to acute temperature stress 2–5 °C above or below ambient for 12–72 h (4). However, bleaching in the field is not usually associated with such abrupt, acute changes in temperature and usually occurs after a more gradual onset of temperature stress that may persist for several weeks (7). The extent to which the mechanisms of experimental bleaching relate to the ob- served declines in symbiont densities in the field is uncer- tain, unless these mechanisms are investigated under both experimental and field conditions. Results from our labora- tory indicate that nitric oxide (NO), generated by algal nitric oxide synthase (NOS), is closely associated with coral bleaching both in artificially induced bleaching in the lab- oratory setting and during naturally occurring bleaching episodes in the field. Nitric oxide synthases convert arginine to citrulline and NO (8). Zooxanthellae that have been freshly iso- lated from a non-stressed symbiotic relationship with a host cnidarian have a free amino acid pool that is dom- inated by arginine (9). In addition, these symbionts pos- sess a low basal level of NOS activity (10). We examined the effects of elevated or depressed water temperatures that induce the breakdown of cnidarian-dinoflagellate symbioses on symbiont NOS activity, which was mea- Received 6 November 2003; accepted 30 November 2004. * To whom correspondence should be addressed, at School of Ocean and Earth Science and Technology, University of Hawaii, Honolulu 96822. E-mail: hank@bbsr.edu Reference: Biol. Bull. 208: 3– 6. (February 2005) © 2005 Marine Biological Laboratory 3