11th ICRS Abstract book - Nova Southeastern University

8-13
Assessment Of Coral Microbiota: The Impact Of Untested Assumptions,
Unintended Biases, And Undefined Variables
Shawn POLSON 1,2 , Sara POLSON 1 , Julie HIGGINS 1 , Cheryl WOODLEY* 1
1 Center for Coastal Environmental Health and Biomolecular Research, NOAA National
Ocean Service, Charleston, SC, 2 Marine Biomedicine and Environmental Sciences
Center, Medical University of South Carolina, Charleston, SC
The study of microbiology has emerged as an important component of coral biology, with
numerous studies examining coral-associated microbial ecology in hopes of
understanding the dynamic nature of these communities. Such studies have provided
numerous insights into the composition of these communities, but a significant level of
project-to-project variability is apparent when reviewing these results. The study
presented here utilized culture-dependent (~650 isolates) and independent (~25,000
sequences from 88 16S rDNA libraries) analyses to assess the microbial community
structure associated with acroporid corals of the Florida reef tract (USA). The study was
designed to allow for the impact of multiple sample variables including sample
collection/processing methodology, disease state, acroporid species, regional geography,
and limited temporal variation to be simultaneously assessed. Sample processing
methodology was found to introduce significant bias into the resulting microbial
community composition detected. Lumping coral health states into defined “diseases”,
based solely upon field observations of similarity in disease signs, was also demonstrated
to be a potential source of variation found in the past literature. The existence of marked
temporal variation within a population of corals was also detected. The results of this
study indicate that future studies of coral-associated microbial communities must take
great care to control for numerous variables (many of which are currently poorly
understood). Key among these factors is an in depth assessment of the physiological
state of the coral during sample collection. Without increased efforts to address the role
of such secondary variables on microbial community composition (both actual and
detected) it is likely that increased fragmentation in the body of literature will occur,
hampering efforts to understand the nature of this important component of the coral
holobiont.
8-14
Changes in Coral Associated Microbial Communities During A Bleaching Event
David BOURNE* 1 , Yuki IIDA 1 , Sven UTHICKE 1 , Carolyn SMITH-KEUNE 1,2
1 Australian Institute of Marine Science, Townsville, Australia, 2 Centre of Marine Studies,
University of Queensland, Brisbane, Australia
Environmental stressors such as increased sea surface temperatures are well known for
contributing to coral bleaching, however the effect of increased temperatures and
subsequent bleaching on coral associated microbial communities is poorly understood.
Colonies of the hard coral Acropora millepora were tagged on a reef flat off Magnetic
Island (Great Barrier Reef) and surveyed over 2.5 years, which included a severe
bleaching event in January/February 2002. Daily average water temperatures exceeded
the previous 10-year average by more than 1ºC for extended periods with field based
visual surveys recording all tagged colonies displaying signs of bleaching. During the
bleaching period, direct counts of coral zooxanthellae densities decreased by ~64%,
before recovery to pre-bleaching levels after the thermal stress event. A subset of three
tagged coral colonies were sampled through the bleaching event and changes in the
microbial community elucidated. DGGE analysis demonstrated conserved bacterial
banding profiles between the three coral colonies confirming previous studies
highlighting specific microbial associations. As coral colonies bleached, the microbial
community shifted and redundancy analysis (RDA) of DGGE-banding patterns revealed
a correlation of increasing temperature with the appearance of Vibrio-affiliated
sequences. Clone libraries hybridised with Vibrio-specific oligonucleotide probes
confirmed an increase in the fraction of Vibrio-affiliated clones during the bleaching
period. Post-bleaching, the coral microbial associations again shifted, returning to a
profile similar or identical to the fingerprints prior to bleaching. This provided further
evidence for corals selecting and shaping their microbial partners. For non-bleached
samples, a close association with Spongiobacter related sequences were revealed by both
clone libraries and DGGE profiling. Despite Vibrio species being previously implicated
in bleaching of specific coral species, it is unsure if the relative increase in retrieved
Vibrio sequences is due to bacterial infection or an opportunistic response to
compromised health and changing environmental parameters of the coral host.
Oral Mini-Symposium 8: Coral Microbial Interactions
8-15
Metagenomic Analysis Of The Microbial Community Associated With The Coral Porites
Astreoides
Linda WEGLEY 1 , Robert EDWARDS* 2 , Forest ROHWER 1
1 Biology, San Diego State University, San Diego, CA, 2 San Diego State University, San Diego,
CA
The coral holobiont is a dynamic assemblage of the coral animal, zooxanthellae, endolithic
algae and fungi, Bacteria, Archaea and viruses. Zooxanthellae and some Bacteria form
relatively stable and species-specific associations with corals. The symbiontic algae, or
zooxanthellae, that reside within the coral tissues have been studied extensively, however very
little is understood about the other microbes within the coral holobiont. To better understand the
roles of the microbes associated with corals, a fractionation procedure was used to separate the
microbes, mitochondria, and viruses from the coral animal cells and zooxanthellae. The
resulting metagenomic DNA was sequenced using pyrosequencing. Fungi, Bacteria, and phage
were the most commonly identified organisms in the metagenome. Three of the four fungal
phyla were represented, including a wide diversity of fungal genes involved in carbon and
nitrogen metabolism, suggesting that the endolithic community is more important than
previously appreciated. In particular, the data suggested that endolithic fungi could be
converting nitrate and nitrite to ammonia, which would enable fixed-nitrogen to cycle within
the coral holobiont. The most prominent bacterial groups were Proteobacteria (68%),
Firmicutes (10%), Cyanobacteria (7%), and Actinobacteria (6%). Functionally, the bacterial
community was primarily heterotrophic and included a number of pathways for the degradation
of aromatic compounds, the most abundant being the homogentisate pathway. The most
abundant phage family was the ssDNA Microphage and most of the eukaryotic viruses were
most closely related to those known to infect aquatic organisms. This study provides a
metabolic and taxonomic snapshot of microbes associated with the reef-building coral Porites
astreoides and presents a basis for understanding how coral-microbial interactions structure the
holobiont and coral reefs.
8-16
Comparison of bacterial communities on corals containing different Symbiodinium
(Zooxanthellae) clades
Raechel LITTMAN* 1 , David BOURNE 2 , Bette WILLIS 1
1 Marine and Tropical Biology, James Cook University, Townsville, Australia, 2 Australian
Institute of Marine Science, Townsville, Australia
Reef-building corals host a variety of micro-organisms, including symbiotic dinoflagellates,
Symbiodinium (zooxanthellae) and an array of bacteria. Previous studies have suggested that
bacteria can provide many benefits to corals through fixation and passage of nitrogen and
carbon to the coral host and production of secondary metabolites such as antibiotics. Many of
the coral’s nutritional needs are supplied by Symbiodinium, and it has been shown that different
genetic types of this algal symbiont differentially affect the physiology of the coral host. For
instance, corals containing Symbiodinium clade D tend to be more tolerant to heat stress.
However, little is known about how Symbiodinium affects the entire holobiont. Arguably,
changes in the algal symbionts may lead to differences in the bacterial populations on corals.
For instance, Symbiodinium contributes to the mucus layer, providing nutrients to potentially
important bacterial inhabitants. To aid in our understanding of possible interactions between
bacterial and Symbiodinium communities, this study compares bacteria community profiles of
juvenile Acropora millepora and A. tenuis experimentally infected with two different clades of
Symbiodinium, C1 and D. Moreover, bacterial communities are compared on corals containing
C1 and D that have been experimentally bleached to determine whether coral-associated
bacterial communities are more resilient to change on corals containing heat tolerant algal
symbionts. This will aid in understanding the relationships between microbial inhabitants and
how these communities can be destabilized if Symbiodinium is lost from the host.
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