11th ICRS Abstract book - Nova Southeastern University

8.214
Bacterial Diversity Associated With Tropical Azooxanthellate Hexacorals And
Octocorals
Lory Z. SANTIAGO-VÁZQUEZ* 1 , Wolfram M. BRÜCK 2 , Thomas B. BRÜCK 3 ,
Angela P. DUQUE-ALARCÓN 4 , Peter J. MCCARTHY 5 , Russell G. KERR 6
1 Biotechnology, University of Houston, Clear Lake, Houston, TX, 2 Centre of Applied
Marine Biotechnology, Letterkenny Institute of Technology, CAMBio, Letterkenny,
Ireland, 3 Corporate Research and Development, Süd-Chemie AG, München, Germany,
4 Center of Excellence in Biomedical and Marine Biotechnology, Department of Chem.
and Biochem., Florida Atlantic University, Boca Raton, FL, 5 Center for Ocean
Exploration, Harbor Branch Oceanographic Institution, Ft. Pierce, FL, 6 Department of
Chemistry, University of Prince Edward Island, Charlottetown, PE, Canada
There is limited information on the microbial communities associated with
azooxanthellate soft corals. Most of these organisms inhabit inaccessible deep or cold
water environments. However, some tropical azooxanthellate hexacorals and octocorals
grow at moderate depths (40-100m) reached by SCUBA. This study examined the
bacterial symbionts of some of these corals, the hexacoral Cirrhipathes lutkeni and the
octocorals Leptogorgia minimata, Swiftia exertia, and Iciligorgia schrammi. For this
purpose, we used fluorescence in situ hybridization (FISH) and traditional plate culture.
FISH counts for the hexacoral C. lutkeni showed a predominance of γ- and α-
Proteobacteria, and Actinobacteria. FISH counts for the three octocorals showed a
concentration of α-, β-, and γ-Proteobacteria, however, Actinobacteria were present in
low amounts. When examining the bacterial diversity using plate culture, it was found
that these cultures were highly selective for γ-Proteobacteria. The next two most
prominent groups present were α-Proteobacteria and Firmicutes. The predominance of γ-
Proteobacteria observed correlates well with the bacterial populations of other soft corals.
Some γ-Proteobacteria are symbiotic in nature and thus may be important for coral
health. This study provides further insight into the microbial ecology of these unique
organisms.
8.215
Bacterial Community Associated With Tissue And Skeleton Of Three Scleractinian
Species: galaxea Fascicularis, pavona Cactus And turbinaria Reniformis
Pascale TREMBLAY* 1,2 , Markus G. WEINBAUER 3 , Christian NOZAIS 1 , Cécile
ROTTIER 2 , Christine FERRIER-PAGÈS 2
1 Département de Biologie, Chimie et Géographie, Université du Québec à Rimouski,
Rimouski, QC, Canada, 2 Centre Scientifique de Monaco, Monaco, Monaco, 3 Microbial
Ecology and Biogeochemistry Group, Laboratoire d'Océanographie de Villefranche,
CNRS-UPMC, UMR 7093, Villefranche-sur-Mer, France
A study was performed, to assess the bacterial community composition and richness of
the tissue and skeleton of three scleractinian coral species Galaxea fascicularis, Pavona
cactus and Turbinaria reniformis. The main question was to assess if this bacterial
diversity was species specific. For this purpose, corals were incubated all together in the
same environment and bacterial diversity analyzed using denaturing gradient gel
electrophoresis (DGGE) on nested PCR amplified fragments of the 16S rRNA gene. The
multidimensional scaling (MDS) and cluster analyses showed that the bacterial
community in the tissue was species-specific, conversely to the community associated to
the skeleton, which was more homogenous between species. Results concerning tissue
indeed showed that P. cactus was different from G. fascicularis (richness and Shannon p
< 0.0001) and from T. reniformis (Shannon Index p = 0.0335) and G. fascicularis was
different of T. reniformis (richness p = 0.0190). The Shannon Index and richness
parameter were not significantly different between tissue and skeleton, except for P.
cactus (p = 0.0023 and 0.0038 respectively). Mucus excretion was significantly different
(p < 0.0001) for the three species, with a higher excretion in G. fascicularis (178.2 ± 17.4
nmol C mg protein-1 h-1) than in P. cactus (48.4 ± 3.3) and T. reniformis (18.4 ± 1.4).
The mucus of G. fascicuaris and P. cactus also mainly contains galactose and glucose
compared to the mucus of T. reniformis that contains glucose and xylose. This different
excretion and composition can partly explain the observed differences in bacterial
diversity.
Poster Mini-Symposium 8: Coral Microbial Interactions
8.216
Bacterial Quorum Sensing Signals And Settlement Of Coral Larvae
Cory KREDIET* 1 , Kim RITCHIE 2 , Mikhail MATZ 3 , Max TEPLITSKI 4
1 School of Natural Resources and Environment, University of Florida, Gainesville, FL, 2 Marine
Microbiology, Mote Marine Laboratory, Sarasota, FL, 3 Section of Integrative Biology,
University of Texas, Austin, TX, 4 Soil and Water Science, University of Florida, Gainesville,
FL
The settlement cue perceived by coral larvae is currently unknown. Several research groups
demonstrated that coral larvae prefer to settle on substrates that are colonized by coralline algae
or by mats (biofilms) formed by coralline algae and associated microbes. Formation and
function of microbial biofilms involves quorum sensing (QS) signals (acyl homoserine lactones,
AHLs). Both bacteria and eukaryotes produce vitamin signals with newly discovered functions
in QS and host-microbial interactions. While the chemical characterization of the settlement
cue is ongoing elsewhere, we tested a hypothesis that known signals commonly associated with
microbial biofilms may function as settlement cues for larvae of stony corals. These settlement
experiments involved C4-homoserine lactone, 3-oxo-C12-homoserine lactone, lumichrome and
riboflavin, each compound is known to function in bacterial cell-to-cell communication. Acyl
homoserine lactones (AHL) and a riboflavin derivative lumichrome are also involved in
interactions between bacteria and their eukaryotic hosts. These molecules have also been
shown to contribute to settlement or metamorphosis of marine organisms. Presence of AHLs,
lumichrome and riboflavin in coral-associated microbes and in coralline algae was investigated.
Their role in settlement was investigated using two complementary approaches. First,
transgenic microbial biofilms expressing AHL-lactonase were constructed to test the
consequences of AHL hydrolysis in larval settlement. Chemicals were also impregnated onto
C18-bonded silica resin to simulate biologically-relevant release rates of the compounds into
the medium during the settlement experiment. Two settlement experiments were carried out to
date with larvae of Acropora palmata and Montastraea faveolata. A strong correlation between
the treatments and settlement rates has yet to be elucidated. Although optimization of
techniques and larval rearing is ongoing, sub-optimal larval and settlement conditions may have
affected the outcome of the experiments.
8.217
Efficient Isolation Of Bacteria That Induce Settlement And Metamorphosis Of acropora
Larvae
Kanae MATSUSHIMA* 1 , Masayuki HATTA 2
1 Graduate School of Humanities and Sciences, Ochanomizu University, Tokyo, Japan, 2 Marine
and Coastal Biology Center, Ochanomizu University, Tokyo, Japan
Biofilms on submarine substrata can act as efficient inducers for settlement and metamorphosis
of Acropora larvae. Indeed, bacteria that induce metamorphosis have been identified from
biofilms; one strain of Pseudoalteromonas and three of Alteromonas. Since micro-environments
on substrata are very diverse, there could be more bacteria species capable of inducing
metamorphosis. Then we tried screening of such bacteria by a new method. Each bacteria
source was swabbed from a 1cm2 surface of tiles submerged in a reef for 3 months, and
suspended in sterilized seawater (SSW). Membrane filters were soaked in each suspension and
incubated on agar media. For passage, the bacteria-grown filter (mixed culture filter; MCF) was
broken into pieces in SSW. We tested each MCF for the activity to induce metamorphosis. Out
of 32 MCF lineages from 16 sources, metamorphosis-inducing batches were obtained in 11
from 8. Although individual batches varied in their activities, every lineage contained one or
more batches giving 100% metamorphosis. Some lineages retained their activities through
passages. Next, a total of 230 independent strains were isolated from 12 active MCFs, and 19
isolates turned out to induce metamorphosis, including 17 giving 100% metamorphosis.
According to 16SrDNA sequencing analyses, 17 isolates, except 2 that failed to be identified,
fell into 7 species in 4 genera; Pseudoalteromonas, Alteromonas, Vibrio and one in alphaproteobacteria.
Our original culture method using MCFs enabled us to get metamorphosisinducing
bacteria very efficiently, compared with the conventional isolation method (1/80 and
3/500 isolates). This study reveals that inducer bacteria spread over in wide taxa. In addition,
we found two Pseudoalteromonas species that inhibit metamorphosis. These findings may
suggest very high diversity of bacteria concerned with the metamorphosis decision of Acropora
larvae.
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