11th ICRS Abstract book - Nova Southeastern University

7-25
Cytotoxcity in The Innate Immune Responses Of The Sea Fan Coral To Infection
By A Fungal Pathogen
Laura D. MYDLARZ* 1 , Laura R. HUNT 1 , C. Drew HARVELL 2
1 Department of Biology, University of Texas at Arlington, Arlington, TX, 2 Ecology and
Evolutionary Biology, Cornell University, Ithaca, NY
The sea fan Gorgonia ventalina possesses effective innate immune mechanisms to fight
infection by the opportunistic fungus Aspergillus sydowii. Some of the innate immune
responses, while efficient at halting pathogen growth and invasion, can lead to
cytotoxicity and self harm in the coral host. Of specific interest are immune mechanisms
which produce reactive oxygen and concomitant production of anti-oxidants to mitigate
the toxic effects. We have recently characterized an oxidative burst in the sea fan in
reaction to fungal signals and have elucidated several roles for the released reactive
oxygen which include halting growth of the fungus and signaling apoptosis. In addition
using corals naturally infected with fungus, we have described cellular immune
mechanisms such as amoebocyte aggregation, activation of the prophenoloxidase
cascade, apoptotic events and increased production of anti-oxidants to reduce the
potential for self harm. The successful use of the sea fan as a model for uncovering
resistance and immune mechanisms and relationships to other coral diseases will also be
summarized.
7-26
Production of Cyanotoxins by Black Band Disease Cyanobacteria
Laurie RICHARDSON* 1 , Miroslav GANTAR 1 , Jamie MYERS 1 , Aaron MILLER 1
1 Biological Sciences, Florida International University, Miami, FL
Black band disease (BBD) of corals is a cyanobacterial dominated microbial mat that
migrates across coral colonies, killing and lysing coral tissues. While it is known that
sulfide, produced by sulfate reducing BBD bacteria, is lethal to coral, inhibition of sulfate
reduction in active BBD infections does not stop tissue lysis or disease progression.
Therefore, there must be additional toxin(s) produced. Using HPLC/MS we detected the
cyanotoxin microcystin in 22 field samples of BBD collected from five coral species on
nine reefs of the Florida Keys and Bahamas. Two cyanobacterial cultures isolated from
BBD, Geitlerinema and Leptolyngbya spp., contained microcystin based on HPLC/MS,
with toxic activity confirmed using the protein phosphatase inhibition assay. DGGE
analysis revealed that the cultured Geitlerinema and Leptolyngbya strains were present in
BBD field samples. HPLC analysis identified the microcystin variants -LY, -LR, -LF, -
LW and –RR in the field and laboratory samples. Exposure of coral fragments to purified
microcystin resulted in tissue stress, which was exacerbated by co-exposure with sulfide.
Cyanobacterial toxicity was further tested on a comparative basis using 10 strains that
were isolated from BBD and 10 strains from non-BBD sources on coral reefs, including
benthic mat forming cyanobacteria and non-pathogenic cyanobacterial patches on healthy
corals. Toxicity was measured using cell extracts and included ELISA, the protein
phosphatase inhibition assay, and HPLC. We found toxicity in both BBD and non-BBD
strains, with levels that varied from non-detectable to 3.5 ug/g of dry biomass within each
group (BBD and non-BBD). The ELISA assay was microcystin and nodularin congenerindependent,
and discrimination between these two toxins is underway. The variant and
relative toxicity of microcystins and other cyanotoxins can be dependent on
cyanobacterial physiology and environmental factors, which may play a role in the
pathogenicity of BBD.
Oral Mini-Symposium 7: Diseases on Coral Reefs
7-27
Significance Of Immunological Responses in The Black-Spined Sea Urchin, diadema
Antillarum, To Caribbean-Wide Mass Mortality
Gregory BECK* 1 , Robert MILLER 2 , Aaron ADAMS 3 , John EBERSOLE 1
1 Biology, University of Massachusetts - Boston, Boston, MA, 2 Marine Science Institute,
University of California - Santa Barbara, Santa Barbara, CA, 3 Mote Marine Laboratory,
Pineland, FL
Mass mortality of the grazer Diadema antillarum, probably caused by a water-borne pathogen,
was a major factor leading to a phase shift from coral to algal domination of Caribbean reefs.
Recovery or restoration of Diadema may be critical to reef restoration. Recent initiation of
recovery of Diadema in St. Croix allows research that addresses basic questions on how
recovery/restoration may be influenced by immunological processes. What are the basic
features of immunological responses in Diadema? How does strength of immune responses in
Diadema compare to other Caribbean urchins that did not die off (e.g., Tripneustes ventricosa,
Echinometra lucunter, or Echinometra viridis)? How does strength of immune response vary
among reefs with strong versus weak recovery, large versus small individuals, healthy versus
sick/dying individuals?
We used coelomic fluid and coelomocytes (blood cells) extracted from Diadema and other
urchins from several locations on St. Croix to test humoral immune responses. Coelomocyte
concentrations did not differ consistently between Diadema and the other urchins. Protein
levels in coelomic fluids were higher in Diadema than the other urchin species. All urchins
released O2-, antimicrobial peptides, and phenoloxidase when stimulated by various agents -
with one conspicuous and statistically significant exception: Diadema from all locations did not
respond as efficiently as other urchins to lipopolysaccharide (a typical component of bacterial
cell walls). These results suggest a defect in immune response that is specific to D. antillarum
and independent of stressors associated with particular environments.
These studies will provide the information required to understand whether a weakened immune
system was responsible for the mass mortality, and how much strengthening of immune systems
has occurred since. Effective management of a recovery may depend on knowing whether
diseases continue to impact Diadema population growth due to weak immune responses, and
whether an abundant recovered/restored population could experience another die-off.
7-28
Cleaner Shrimp periclemenes Pedersoni Reduce Ectoparasite Loads On A Caribbean
Reef Fish
Amber MCCAMMON 1 , Donna NEMETH* 2 , Paul SIKKEL 3
1 Department of Biology, Florida Atlantic University, Boca Raton, FL, 2 Science and Math,
University of the Virgin Islands, St. Thomas, Virgin Islands (U.S.), 3 Department of Biology,
Centre College, Danville, KY
Reef fish are known to make regular visits to ‘cleaning stations’, yet the benefit to the fish is not
well studied in Caribbean species. Periclemenes pedersoni cleaner shrimp associate with the
anemone Bartholomea annulata, and have been hypothesized to participate in cleaning
symbioses. Experiments were conducted in a seminatural environment to determine the
effectiveness of these cleaner shrimp at removing parasitic monogeneans trematodes from host
reef fish, Acanthurus coeruleus. Wild-caught fish were exposed to seawater flow from an open
system aquarium exhibit containing high densities of monogenean eggs and larvae
(Neobenedenia). Fish were assigned to one of two treatments: one giving fish access to cleaner
shrimps that also included the anemone Bartholomea annulata for cover, and one that included
anemones with no shrimps. After 15 days, fish were fresh-water dipped to remove parasites,
which were preserved in ethanol and counted under a microscope. Average monogenean loads
were 4 times higher on fish without access to cleaners, compared to fish with access to cleaners
(monogeneans load per fish averaged 101 vs. 26, respectively, ANOVA p